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Pathology_Robbins. Primarily, TGF-β drives scar formation by stimulating matrix synthesis through decreased matrix metalloproteinase (MMP) activity and increased activity of tissue inhibitors of proteinases (TIMPs). TGF-β also applies brakes to the inflammation that accompanies wound healing by inhibiting lymphocyte proliferation and the activity of other leukocytes.


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Pathology_Robbins. The ECM is a network of interstitial proteins that constitutes a significant proportion of any tissue. Cell interactions with ECM are critical for development and healing, as well as for maintaining normal tissue architecture ( Fig. 1.13 ). Much more than a simple “space filler” around cells, ECM serves several key functions: Mechanical support for cell anchorage and cell migration, and maintenance of cell polarity. Control of cell proliferation, by binding and displaying growth factors and by signaling through cellular receptors of the integrin family. The ECM provides a depot PROLIFERATION, DIFFERENTIATION, PROTEIN SYNTHESIS, ATTACHMENT, MIGRATION, SHAPE CHANGE Fig. 1.13 Interactions of ECM and growth factor–mediated cell signaling. Cell surface integrins interact with the cytoskeleton at focal adhesion complexes (protein aggregates that include vinculin, α-actinin, and talin; see


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Pathology_Robbins. Fig. 1.16C ). This can initiate the production of intracellular messengers or can directly trans-duce signals to the nucleus. Cell surface receptors for growth factors can activate signal transduction pathways that overlap with those mediated through integrins. Signals from ECM components and growth factors can be integrated by the cells to produce a given response, including changes in proliferation, locomotion, and/or differentiation. for a variety of latent growth factors that can be activated within a focus of injury or inflammation. Scaffolding for tissue renewal. Because maintenance of normal tissue structure requires a basement membrane or stromal scaffold, the integrity of the basement membrane or the stroma of parenchymal cells is critical for the organized regeneration of tissues. Thus, ECM disruption results in defective tissue regeneration and repair, for example, cirrhosis of the liver resulting from the collapse of the hepatic stroma in various forms of hepatitis.


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Pathology_Robbins. Establishment of tissue microenvironments. The basement membrane acts as a boundary between the epithelium and underlying connective tissue; it does not just provide support to the epithelium but is also functional, for example, in the kidney, forming part of the filtration apparatus. The ECM is constantly being remodeled; its synthesis and degradation accompany morphogenesis, tissue regeneration and repair, chronic fibrosis, and tumor invasion and metastasis. ECM occurs in two basic forms: interstitial matrix and basement membrane ( Fig. 1.14


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Pathology_Robbins. Fig. 1.14 Interstitial matrix is present in the spaces between cells in connective tissue, and between the parenchymal epithelium and the underlying supportive vascular and smooth muscle structures. The interstitial matrix is synthesized by mesenchymal cells (e.g., fibroblasts), forming an amorphous three-dimensional gel. Its major constituents are fibrillar and nonfibrillar collagens, as well as fibronectin, elastin, proteoglycans, hyaluronate, and other constituents (see later). Basement membrane. The seemingly random array of interstitial matrix in connective tissues becomes highly organized around epithelial cells, endothelial cells, and smooth muscle cells, forming the specialized basement membrane. This is synthesized conjointly by the overlying epithelium and the underlying mesenchymal cells, forming a flat lamellar “chicken wire” mesh (although labeled as a membrane, it is quite porous). The major constituents are amorphous nonfibrillar type IV collagen and laminin.)


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Pathology_Robbins. The components of the ECM fall into three groups of proteins ( Fig. 1.15 Fibrous structural proteins such as collagens and elastins that confer tensile strength and recoil Water-hydrated gels such as proteoglycans and hyaluronan that permit compressive resistance and lubrication Adhesive glycoproteins that connect ECM elements to one another and to cells Collagens. Collagens are composed of three separate polypeptide chains braided into a ropelike triple helix. About 30 collagen types have been identified, some of which are unique to specific cells and tissues.


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Pathology_Robbins. Some collagen types (e.g., types I, II, III, and V collagens) form linear fibrils stabilized by interchain hydrogen bonding; such fibrillar collagens form a major proportion of the connective tissue in structures such as bone, tendon, cartilage, blood vessels, and skin, as well as in healing wounds and scars. The tensile strength of the fibrillar collagens derives from lateral crosslinking of the triple helices by covalent bonds, an unusual A Fibrillar collagen and elastin B Proteoglycan C bFGF regulation by association with the extracellular matrix


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Fig.

Pathology_Robbins. Fig.


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Pathology_Robbins. 1.15 ECM components. (A) Fibrillar collagen, and elastic tissue structures. Because of rodlike fibril stacking and extensive lateral crosslinking, collagen fibers have marked tensile strength but little elasticity. Elastin also is crosslinked but differs in having large hydrophobic segments that form a dense globular configuration at rest. As stretch is exerted, the hydrophobic domains are pulled open, but the crosslinks keep the molecules intact; release of the tension allows the hydrophobic domains of the proteins to refold. (B) Proteoglycan structure. The highly negatively charged sulfated sugars on the proteoglycan “bristles” attract sodium and water to generate a viscous compressible matrix. (C) Regulation of basic FGF (bFGF, FGF-2) activity by ECM and cellular proteoglycans. Heparan sulfate binds bFGF secreted in the ECM. Syndecan is a cell surface proteoglycan with a transmembrane core protein and extracellular glycosaminoglycan side chains that bind bFGF, and a cytoplasmic tai


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Pathology_Robbins. sulfate binds bFGF secreted in the ECM. Syndecan is a cell surface proteoglycan with a transmembrane core protein and extracellular glycosaminoglycan side chains that bind bFGF, and a cytoplasmic tail that interacts with the intracellular actin cytoskeleton. Syndecan side chains bind bFGF released from damaged ECM, thus facilitating bFGF interaction with cell surface receptors.


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Pathology_Robbins. post-translational modification that requires hydroxylation of lysine residues in collagen by the enzyme lysyl oxidase. Because lysyl oxidase is a vitamin C-dependent enzyme, children with ascorbate deficiency have skeletal deformities, and people of any age with vitamin C deficiency heal poorly and bleed easily because of “weak” collagen. Genetic defects in collagens cause diseases such as osteogenesis imperfecta and certain forms of Ehlers-Danlos syndrome (Chapter 7). • Nonfibrillar collagens variously contribute to the structures of planar basement membranes (type IV collagen); help regulate collagen fibril diameters or collagen-collagen interactions via so-called “fibril-associated collagen with interrupted triple helices” (FACITs, such as type IX collagen in cartilage); and provide anchoring fibrils within basement membrane beneath stratified squamous epithelium (type VII collagen).


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Pathology_Robbins. Elastin. The ability of tissues to recoil and recover their shape after physical deformation is conferred by elastin ( Fig. 1.15 ). Elasticity is especially important in cardiac valves and large blood vessels, which must accommodate recurrent pulsatile flow, as well as in the uterus, skin, and ligaments. Morphologically, elastic fibers consist of a central core of elastin with an associated meshlike network composed of fibrillin. The latter relationship partially explains why fibrillin defects lead to skeletal abnormalities and weakened aortic walls, as in individuals with Marfan syndrome. Fibrillin also controls the availability of TGF-β (Chapter 7).


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Pathology_Robbins. Fig. 1.15 ). Proteoglycans form highly hydrated gels that confer resistance to compressive forces; in joint cartilage, proteoglycans also provide a layer of lubrication between adjacent bony surfaces. Proteoglycans consist of long polysaccharides called glycosaminoglycans (examples are keratan sulfate and chondroitin sulfate) attached to a core protein; these are then linked to a long hyaluronic acid polymer called hyaluronan in a manner reminiscent of the bristles on a test-tube brush. The highly negatively charged, densely packed sulfated sugars attract cations (mostly sodium) and abundant water molecules, producing a viscous, gelatin-like matrix. Besides providing compressibility to tissues, proteoglycans also serve as reservoirs for secreted growth factors (e.g., FGF and HGF). Some proteoglycans are integral cell membrane proteins that have roles in cell proliferation, migration, and adhesion, for example, by binding and concentrating growth factors and chemokines ( Fig. 1.15


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Pathology_Robbins. Fig. 1.15 Adhesive glycoproteins and adhesion receptors. These are structurally diverse molecules variously involved in cell-cell, cell-ECM, and ECM-ECM interactions ( Fig. 1.16 ). Prototypical adhesive glycoproteins include fibronectin (a Fig. 1.16 CellandECMinteractions:adhesiveglycoproteinsandintegrinsignaling.(A)Fibronectin consistsofadisulfide-linkeddimer,withseveraldistinctdomainsthatallowbindingtoECMandintegrins,thelatterthrougharginine-glycine-asparticacid(RGD)motifs.(B)Thecross-shapedlaminin moleculeisoneofthemajorcomponentsofbasementmembranes;itsmultidomainstructureallowsinteractionsbetweentypeIVcollagen,otherECMcomponents,andcell-surfacereceptors.(C)Integrinsandintegrin-mediatedsignalingeventsatfocaladhesioncomplexes.Eachα–β heterodimericintegrinreceptorisatrans-membranedimerthatlinksECMandtheintracellularcytoskeleton.Itisalsoassociatedwithacomplexoflinkingmolecules(e.g.,vinculinandtalin)thatcanrecruitandactivatekinasesthatultimatelytriggerdownstreamsignalingcascades.


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Pathology_Robbins. major component of the interstitial ECM) and laminin (a major constituent of basement membrane). Integrins are representative of the adhesion receptors, also known as cell adhesion molecules (CAMs); the CAMs also include immunoglobulins family members, cadherins, and selectins. Fibronectin is a large (450 kD) disulfide-linked heterodimer that exists in tissue and plasma forms; it is synthesized by a variety of cells, including fibroblasts, monocytes, and endothelium. Fibronectin has specific domains that bind to distinct ECM components (e.g., collagen, fibrin, heparin, and proteoglycans), as well as integrins ( Fig. 1.16 ). In healing wounds, tissue and plasma fibronectin provide a scaffold for subsequent ECM deposition, angiogenesis, and reepithelialization. Laminin is the most abundant glycoprotein in the basement membrane. It is an 820-kD cross-shaped heterotrimer that connects cells to underlying ECM components such as type IV collagen and heparan sulfate (


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Pathology_Robbins. Fig. 1.16 ). Besides mediating the attachment to the basement membrane, laminin can also modulate cell proliferation, differentiation, and motility.


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Pathology_Robbins. Integrins are a large family of transmembrane heterodimeric glycoproteins composed of α-and β-subunits that allow cells to attach to ECM constituents such as laminin and fibronectin, thus functionally and structurally linking the intracellular cytoskeleton with the outside world. Integrins also mediate cell-cell adhesive interactions. For instance, integrins on the surface of leukocytes are essential in mediating firm adhesion to and transmigration across the endothelium at sites of inflammation (Chapter 3), and they play a critical role in platelet aggregation (Chapter 4). Integrins attach to ECM components via a tripeptide arginine-glycine-aspartic acid motif (abbreviated RGD). In addition to providing focal attachment to underlying substrates, binding through the integrin receptors can also trigger signaling cascades that influence cell locomotion, proliferation, shape, and differentiation ( Fig. 1.16


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Pathology_Robbins. Fig. 1.16 Cell proliferation is fundamental to development, maintenance of steady-state tissue homeostasis, and replacement of dead or damaged cells. The key elements of cellular proliferation are accurate DNA replication accompanied by the coordinated synthesis of all other cellular constituents, followed by equal apportionment of DNA and other cellular constituents (e.g., organelles) to daughter cells through mitosis and cytokinesis. The sequence of events that results in cell division is called the cell cycle. The cell cycle consists of G1 (pre-synthetic growth), S (DNA synthesis), G2 (premitotic growth), and M (mitotic) phases; quiescent cells that are not actively cycling are in the G0 state. (


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Pathology_Robbins. Fig. 1.17 ). Cells can enter G1 either from the G0 quiescent cell pool or after completing a round of mitosis. Each stage requires completion of the previous step, as well as activation of necessary factors (see later); nonfidelity of DNA replication or cofactor deficiency results in arrest at the various transition points. Fig. 1.17 Cell-cyclelandmarks.Thefigureshowsthecell-cyclephases(G0,G1,G2,S,andM),thelocationoftheG1restrictionpoint,andtheG1/SandG2/Mcell-cyclecheckpoints.G1restrictionpointreferstothephaseinG1atwhichthecellgetscommittedtothecellcyclewithoutfurtherneedofthegrowthfactorthatinitiatedcelldivision.Cellsfromlabiletissuessuchastheepidermisandthegastrointestinaltractmaycyclecontinuously;stablecellssuchashepatocytesarequiescentbutcanenterthecellcycle;permanentcellssuchasneuronsandcardiacmyocyteshavelostthecapacitytoproliferate.(Modified from Pollard TD, Earnshaw WC: Cellbiology,Philadelphia, 2002, Saunders.)


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Pathology_Robbins. The cell cycle is regulated by numerous activators and inhibitors. Cell-cycle progression is driven by proteins called cyclins—named for the cyclic nature of their production and degradation—and cyclin-associated enzymes called cyclin-dependent kinases (CDKs) ( Fig. 1.18 ). CDKs acquire the ability to phosphorylate protein substrates (i.e., kinase activity) by forming complexes with the relevant cyclins. Transiently increased synthesis of a particular cyclin leads to increased kinase activity of the appropriate CDK binding partner; as the CDK completes its round of phosphorylation, the associated cyclin is degraded and the CDK activity abates. Thus, as cyclin levels rise and fall, the activity of associated CDKs likewise waxes and wanes.


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Pathology_Robbins. More than 15 cyclins have been identified; cyclins D, E, A, and B appear sequentially during the cell cycle and bind to one or more CDKs. The cell cycle thus resembles a relay race in which each leg is regulated by a distinct set of cyclins: as one collection of cyclins leaves the track, the next set takes over.


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Pathology_Robbins. Embedded in the cell cycle are surveillance mechanisms primed to sense DNA or chromosomal damage. These quality-control checkpoints ensure that cells with genetic imperfections do not complete replication. Thus, the G1-S checkpoint monitors the integrity of DNA before irreversibly committing cellular resources to DNA replication. Later in the cell cycle, the G2-M check point ensures that there has been accurate DNA replication before the cell actually divides. When cells do detect DNA irregularities, checkpoint activation delays cell-cycle progression and triggers DNA repair mechanisms. If the genetic derangement is too severe to be repaired, the cells either undergo apoptosis or enter a nonreplicative state called senescence— primarily through p53-dependent mechanisms (see later). Enforcing the cell-cycle checkpoints is the job of CDK inhibitors (CDKIs); they accomplish this by modulating CDK-cyclin complex activity. There are several different CDKIs:


Pathology_Robbins

Pathology_Robbins. Enforcing the cell-cycle checkpoints is the job of CDK inhibitors (CDKIs); they accomplish this by modulating CDK-cyclin complex activity. There are several different CDKIs: One family of CDKIs—composed of three proteins called p21 (CDKN1A), p27 (CDKN1B), and p57 (CDKN1C)— broadly inhibits multiple CDKs Another family of CDKIs has selective effects on cyclin CDK4 and cyclin CDK6; these proteins are called p15 (CDKN2B), p16 (CDKN2A), p18 (CDKN2C), and p19 (CDKN2D) Defective CDKI checkpoint proteins allow cells with damaged DNA to divide, resulting in mutated daughter cells at risk for malignant transformation


Pathology_Robbins

Pathology_Robbins. Defective CDKI checkpoint proteins allow cells with damaged DNA to divide, resulting in mutated daughter cells at risk for malignant transformation An equally important aspect of cell growth and division is the biosynthesis of other cellular components needed to make two daughter cells, such as membranes and organelles. Thus when growth factor receptor signaling stimulates cell-cycle progression, it also activates events that promote changes in cellular metabolism that support growth. Chief among these is the Warburg effect, mentioned earlier, marked by increased cellular uptake of glucose and glutamine, increased glycolysis, and (counterintuitively) decreased oxidative phosphorylation. These changes are major elements of cancer-cell growth and are discussed in greater detail in Chapter 6. Not all stem cells are created equal. During development, totipotent stem cells can give rise to all types of http://ebooksmedicine.net


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Pathology_Robbins. Not all stem cells are created equal. During development, totipotent stem cells can give rise to all types of http://ebooksmedicine.net Fig. 1.18 Roleofcyclins,CDKs,andCDKinhibitorsinregulatingthecellcycle.Shaded arrows representthephasesofthecellcycleduringwhichspecificcyclin-CDKcomplexesareactive.Asillustrated,cyclinD-CDK4,cyclinD-CDK6,andcyclinE-CDK2regulatetheG1-to-StransitionbyphosphorylatingtheRbprotein(pRb).CyclinA-CDK2andcyclinA-CDK1areactiveintheSphase.CyclinB-CDK1isessentialfortheG2-to-Mtransition.TwofamiliesofCDKinhibitorscanblockactivityofCDKsandprogressionthroughthecellcycle.Theso-called“INK4inhibitors,”composedofp16,p15,p18,andp19,actoncyclinD-CDK4andcyclinD-CDK6.Theotherfamilyofthreeinhibitors,p21,p27,andp57,caninhibitallCDKs.


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Pathology_Robbins. differentiated tissues; in the mature organism, adult stem cells in various tissues only have the capacity to replace damaged cells and maintain cell populations within the tissues where they reside. There also are populations of stem cells between these extremes with varying capacities to differentiate into multiple cell lineages. Thus, depending on the source and stage of development, there may be limits on the cell types that a stem cell population can generate. In normal tissues (without neoplasia, degeneration, or healing), there is a homeostatic equilibrium between the replication, self-renewal, and differentiation of stem cells and the death of the mature, fully differentiated cells (


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Pathology_Robbins. Fig. 1.19 ). The dynamic relationship between stem cells and terminally differentiated parenchyma is nicely exemplified by the continuously dividing epithelium of the skin. Thus, stem cells at the basal layer of the epithelium progressively differentiate as they migrate to the upper layers of the epithelium before dying and being shed. Under conditions of homeostasis, stem cells are characterized by two important properties: Self-renewal, which permits stem cells to maintain their numbers. Self-renewal may follow asymmetric or symmetric division.


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Pathology_Robbins. Self-renewal, which permits stem cells to maintain their numbers. Self-renewal may follow asymmetric or symmetric division. Asymmetric division refers to cell replication in which one daughter cell enters a differentiation pathway and gives rise to mature cells, whereas the other remains undifferentiated and retains its self-renewal capacity. By contrast, in symmetric division, both daughter cells retain self renewal capacity. Such divisions are seen early in embryogenesis (when stem cell populations are expanding) and under conditions of stress, such as in the bone marrow following chemotherapy. Although there is a tendency in the scientific literature to partition stem cells into several different subsets, fundamentally there are only two varieties:


Pathology_Robbins

Pathology_Robbins. Although there is a tendency in the scientific literature to partition stem cells into several different subsets, fundamentally there are only two varieties: Embryonic stem cells (ES cells) are the most undifferentiated. They are present in the inner cell mass of the http://ebooksmedicine.net blastocyst, have virtually limitless cell renewal capacity, and can give rise to every cell in the body; they are thus said to be totipotent ( Fig. 1.20 ). ES cells can be maintained for extended periods without differentiating; thereafter, appropriate culture conditions allow them to form specialized cells of all three germ cell layers, Fig. 1.19 Mechanismsregulatingcellpopulations.Cellnumberscanbealteredbyincreasedordecreasedratesofstemcellinput,celldeathresultingfromapoptosis,orchangesintheratesofproliferationordifferentiation.


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Pathology_Robbins. Fig. 1.19 Mechanismsregulatingcellpopulations.Cellnumberscanbealteredbyincreasedordecreasedratesofstemcellinput,celldeathresultingfromapoptosis,orchangesintheratesofproliferationordifferentiation. (Modified from McCarthy NJ, et al: Apoptosis in the development of the immune system: growth factors, clonal selection and bcl-2, CancerMetastasisRev11:157, 1992.) including neurons, cardiac muscle, liver cells, and pancreatic islet cells.


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Pathology_Robbins. • Tissue stem cells (also called adult stem cells) are found in intimate association with the differentiated cells of a given tissue. They are normally protected within specialized tissue microenvironments called stem cell niches. Such niches have been demonstrated in many organs, most notably the bone marrow, where hematopoietic stem cells congregate in a perivascular niche. Other niches for stem cells include the bulge region of hair follicles; the limbus of the cornea; the crypts of the gut; the canals of Hering in the liver; and the subventricular zone in the brain. Soluble factors and other cells within the niches keep the stem cells quiescent until there is a need for expansion and differentiation of the precursor pool ( Fig. 1.21


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Pathology_Robbins. Fig. 1.21 Adult stem cells have a limited repertoire of differentiated cells that they can generate. Thus, although adult stem cells can maintain tissues with high (e.g., skin and gastrointestinal tract) or low (e.g., endothelium) cell turnover, the adult stem cells in any given tissue can usually only produce cells that are normal constituents of that tissue.


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Pathology_Robbins. Hematopoietic stem cells are the most extensively studied; they continuously replenish all the cellular elements of the blood as they are consumed. They can be isolated directly from bone marrow, as well as from the peripheral blood after administration of certain colony stimulating factors (CSF) that induce their release from bone marrow niches. Although rare, hematopoietic stem cells can be purified to virtual homogeneity based on cell surface markers. Clinically, these stem cells can be used to repopulate marrows depleted after chemotherapy (e.g., for leukemia), or to provide normal precursors to correct various blood cell defects (e.g., sickle cell disease; see Chapter 12).


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Pathology_Robbins. Fig. 1.21 Stem cell niches in various tissues. (A) Skin stem cells are located in the bulge area of the hair follicle, in sebaceous glands, and in the lower layer of the epidermis. (B) Small intestine stem cells are located near the base of the crypt, above Paneth cells. (C) Liver stem cells (oval cells) are located in the canals of Hering (thick arrow), structures that connect bile ductules (thin arrow) to parenchymal hepatocytes. Bile duct cells and canals of Hering are stained here with an immunohistochemical stain for cytokeratin 7. (C, Courtesy Tania Roskams, MD, University of Leuven, Belgium.)


Pathology_Robbins

Pathology_Robbins. Besides hematopoietic stem cells, the bone marrow (and notably, other tissues such as fat) also contains a population of mesenchymal stem cells. These are multipotent cells that can differentiate into a variety of stromal cells including chondrocytes (cartilage), osteocytes (bone), adipocytes (fat), and myocytes (muscle). Because these cells can be expanded to large numbers, they represent a potential means of manufacturing the stromal scaffolding needed for tissue regeneration.


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Pathology_Robbins. The ability to identify, isolate, expand, and transplant stem cells has given birth to the new field of regenerative medicine. Theoretically, the differentiated progeny of ES or adult stem cells can be used to repopulate damaged tissues or to construct entire organs for replacement. In particular, there is considerable excitement about the therapeutic opportunities for restoring damaged tissues that have low intrinsic regenerative capacity, such as myocardium after a myocardial infarct or neurons after a stroke. Unfortunately, despite an improving ability to purify and expand stem cell populations, much of the initial enthusiasm has been tempered by difficulties encountered in introducing and functionally integrating the replacement cells into sites of damage.


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Pathology_Robbins. More recently it has been possible to generate pluripotential cells, resembling ES cells, that are derived from the patient into whom they will be implanted. To accomplish this, a handful of genes have been identified whose products can—remarkably—reprogram somatic cells to achieve the “stem-ness” of ES cells. When such genes are introduced into fully differentiated cells (e.g., fibroblasts), induced pluripotent stem cells (iPS cells) are generated ( Fig. 1.22 ), albeit at low frequency. Because these cells are derived from the patient, their differentiated progeny (e.g., insulin-secreting β-cells in a patient with diabetes) can be engrafted without eliciting an immunologically mediated rejection reaction that would occur if the differentiated cells were derived from ES cells obtained from another donor.


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Pathology_Robbins. Fig. 1.22 The production of induced pluripotent stem cells (iPS cells). Genes that confer stem cell properties are introduced into a patient’s differentiated cells, giving rise to stem cells that can be induced to differentiate into various lineages. (Modified from Hochedlinger K, Jaenisch R: Nuclear transplantation, embryonic stem cells, and the potential for cell therapy, N Engl J Med 349:275–286, 2003.) This survey of selected topics in cell biology serves as a basis for our later discussions of pathology, and we will refer back to it throughout the book. Students should, however, remember that this summary is intentionally brief, and more information about some of the fascinating topics reviewed here can be readily found in textbooks devoted to cell and molecular biology. Batista PJ, Chang HY: Long noncoding RNAs: cellular address codes in development and disease, Cell 152:1298, 2013. [Good review regarding lncRNA biology.]


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Pathology_Robbins. Batista PJ, Chang HY: Long noncoding RNAs: cellular address codes in development and disease, Cell 152:1298, 2013. [Good review regarding lncRNA biology.] Cech TR, Steitz JA: The noncoding RNA revolution trashing old rules to forge new ones, Cell 157:77, 2014. [An excellent review of the roles played by noncoding RNAs.] Hübner MR, Eckersley-Maslin MA, Spector DL: Chromatin organization and transcriptional regulation, Curr Opin Genet Dev 23:89, 2013. [A nice discussion of genome organization and chromatin structure– function relationships that regulate cell type–specific nuclear transcription.] Jarovcevski M, Akbarian S: Epigenetic mechanisms in neurologic disease, Nat Med 18:1194, 2012. [A well-written overview of genomic organization and transcriptional regulation, with a specific focus on neurologic disease.]


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Pathology_Robbins. Meller VH, Joshi SS, Deshpande N: Modulation of chromatin by non-coding RNA, Annu Rev Genet 49:673, 2015. [An excellent overview of the roles played by noncoding RNAs in nuclear organization.] Minarovits J, Banati F, Szenthe K, et al: Epigenetic regulation, Adv Exp Med Biol 879:1, 2016. [A brief primer on the pathways that regulate chromatin structure and accessibility.] Rowley MJ, Corces VG: The three-dimensional genome: principles and roles of long-distance interactions, Curr Opin Cell Biol 40:8, 2016. [An interesting discussion regarding the mechanisms by which three-dimensional conformations can influence nuclear transcription.] Teperino R, Lempradl A, Pospisilik JA: Bridging epigenomics and complex disease: the basics, Cell Mol Life Sci 70:1609, 2013. [An introductory review of the epigenetic basis for human disease.]


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Pathology_Robbins. Teperino R, Lempradl A, Pospisilik JA: Bridging epigenomics and complex disease: the basics, Cell Mol Life Sci 70:1609, 2013. [An introductory review of the epigenetic basis for human disease.] Andersson ER: The role of endocytosis in activating and regulating signal transduction, Cell Mol Life Sci 69:1755, 2011. [Overview of endocytosis with specific emphasis on its role in modulating intracellular signaling.] Choi AM, Ryter SW, Levine B: Autophagy in human health and disease, N Engl J Med 368:651, 2013. [A superb review concerning the physiologic and pathophysiologic aspects of autophagy.] English AR, Zurek N, Voeltz GK: Peripheral ER structure and function, Curr Opin Cell Biol 21:596, 2009. [An overview of the structural and functional organization of the endoplasmic reticulum and its relationship to other cellular organelles.]


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Pathology_Robbins. Guillot C, Lecuit T: Mechanics of epithelial tissue homeostasis and morphogenesis, Science 340:1185, 2013. [A topical discussion about cellular interactions and the mechanical basis of tissue maintenance.] Hetz C, Chevet E, Oakes SA: Proteostasis control by the unfolded protein response, Nat Cell Biol 17:829, 2015. [Mechanisms underlying endoplasmic reticulum editing and cellular homeostasis.] Kaur J, Debnath J: Autophagy at the crossroads of catabolism and anabolism, Nat Rev Mol Cell Biol 16:461, 2015. [An excellent review of the mechanisms and consequences of cellular autophagy.] Simons K, Sampaio JL: Membrane organization and lipid rafts, Cold Spring Harb Perspect Biol 3:1, 2013. [A nice review of the general principles of membrane architecture and emphasizing domain organization.]


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Pathology_Robbins. Wong E, Cuervo AM: Integration of clearance mechanisms: the proteasome and autophagy, Cold Spring Harb Perspect Biol 2:1, 2010. [An overview of intracellular degradation pathways, specifically focusing on the elimination of aberrant or abnormal constituents.] Andersen JL, Kornbluth S: The tangled circuitry of metabolism and apoptosis, Mol Cell 49:399, 2013. [A solid review of the interplay between cell metabolism, cell proliferation, and cell death.] Dang CV: Links between metabolism and cancer, Genes Dev 26:877, 2012. [An excellent review on metabolic functions of mitochondria.] Friedman JR, Nunnari J: Mitochondrial form and function, Nature 505:335, 2014. [A good overview of mitochondrial replication, and response to cellular injury.] Tait SW, Green DR: Mitochondria and cell death: outer membrane permeabilization and beyond, Nat Rev Mol Cell Biol 11:621, 2010. [A review of the role of mitochondria in cell death pathways.]


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Pathology_Robbins. Tait SW, Green DR: Mitochondria and cell death: outer membrane permeabilization and beyond, Nat Rev Mol Cell Biol 11:621, 2010. [A review of the role of mitochondria in cell death pathways.] Duronio RJ, Xiong Y: Signaling pathways that control cell proliferation, Cold Spring Harb Perspect Biol 5:1, 2013. [An excellent overall review of cell signaling and proliferation.] Morrison DK: MAP kinase pathways, Cold Spring Harb Perspect Biol 4:1, 2012. [A review of mitogen-activated kinase signaling pathways.] Perona R: Cell signalling: growth factors and tyrosine kinase receptors, Clin Transl Oncol 8:77, 2011. [An overview of signaling pathways with an emphasis on how these become dysregulated in malignancy.] Alvarado AS, Yamanaka S: Rethinking differentiation: stem cells, regeneration, and plasticity, Cell 157:110, 2014.


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Pathology_Robbins. Alvarado AS, Yamanaka S: Rethinking differentiation: stem cells, regeneration, and plasticity, Cell 157:110, 2014. Fuchs E, Chen T: A matter of life and death: self-renewal in stem cells, EMBO Rep 14:39, 2013. [A scholarly review on the conceptual framework and experimental underpinnings of our understanding regarding stem cell renewal, using cutaneous stem cells as a paradigm.] Li M, Liu GH, Izpisua-Belmonte JC: Navigating the epigenetic landscape of pluripotent stem cells, Nat Rev Mol Cell Biol 13:524, 2012. [A good discussion of the epigenetic regulation of stem cell proliferation and subsequent differentiation.] Martello G, Smith A: The nature of embryonic stem cells, Annu Rev Cell Dev Biol 30:647, 2014. [A good, comprehensive overview of cell plasticity and stemness.] http://ebooksmedicine.net studentconsult.com Cell Injury, Cell Death, and Adaptations Other Pathways of Cell Death


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Pathology_Robbins. Cell Injury, Cell Death, and Adaptations Other Pathways of Cell Death The field of pathology is devoted to understanding the causes of disease and the changes in cells, tissues, and organs that are associated with disease and give rise to the presenting signs and symptoms in patients. There are two important terms that students will encounter throughout their study of pathology and medicine: Etiology refers to the underlying causes and modifying factors that are responsible for the initiation and progression of disease. It is now clear that many common diseases, such as hypertension, diabetes, and cancer, are caused by a combination of inherited genetic susceptibility and various environmental triggers. Elucidating the genetic and environmental factors underlying diseases is a major theme of modern medicine.


Pathology_Robbins

Pathology_Robbins. Pathogenesis refers to the mechanisms of development and progression of disease, which account for the cellular and molecular changes that give rise to the specific functional and structural abnormalities that characterize any particular disease. Thus, etiology refers to why a disease arises and pathogenesis describes how a disease develops ( Fig. 2.1 Defining the etiology and pathogenesis of disease not only is essential for understanding disease but also is the basis for developing rational treatments and effective preventive measures. Thus, pathology provides the scientific foundation for the practice of medicine.


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Pathology_Robbins. To render diagnoses and guide therapy in clinical practice, pathologists identify changes in the gross or microscopic appearance (morphology) of cells and tissues, and biochemical alterations in body fluids (such as blood and urine). Pathologists also use a variety of morphologic, molecular, and other techniques to define the biochemical, structural, and functional changes that occur in cells, tissues, and organs in response to injury. We begin, in this chapter, with a discussion of cellular abnormalities induced by a variety of internal (e.g., genetic) and external (e.g., environmental) abnormalities and stresses.


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Pathology_Robbins. Cells actively interact with their environment, constantly adjusting their structure and function to accommodate changing demands and extracellular stresses. The intracellular milieu of cells is normally tightly regulated such that it remains fairly constant, a state referred to as homeostasis. As cells encounter physiologic stresses (such as increased workload in the heart) or potentially injurious conditions (such as nutrient deprivation), they can undergo adaptation, achieving a new steady state and preserving viability and function. If the adaptive capability is exceeded or if the external stress is inherently harmful or excessive, cell injury develops ( Fig. 2.2 ). Within certain limits, injury is reversible, and cells return to their stable baseline; however, if the stress is severe, persistent, or rapid in onset, it results in irreversible injury and death of the affected cells. Cell death is one of the most crucial events in the evolution http://ebooksmedicine.net


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Pathology_Robbins. MOLECULAR, FUNCTIONAL, AND MORPHOLOGIC ABNORMALITIES IN CELLS AND TISSUES CLINICAL MANIFESTATIONS: SIGNS AND SYMPTOMS OF DISEASE Fig. 2.1 Stepsintheevolutionofdisease.Onlyselectedmajorcauses(etiologies)areshown. of disease in any tissue or organ. It results from diverse causes, including ischemia (lack of blood flow), infections, toxins, and immune reactions. Cell death also is a normal and essential process in embryogenesis, the development of organs, and the maintenance of tissue homeostasis. Because damage to cells is the basis of all disease, in this chapter we discuss first the causes, mechanisms, and consequences of the various forms of acute cell injury, including reversible injury and cell death. We then consider cellular adaptations to stress and conclude with two other processes that affect cells and tissues: the deposition of abnormal substances and cell aging. NORMAL CELL (homeostasis) REVERSIBLE INJURY Injurious stimulus Severe, progressive IRREVERSIBLE INJURY


Pathology_Robbins

Pathology_Robbins. NORMAL CELL (homeostasis) REVERSIBLE INJURY Injurious stimulus Severe, progressive IRREVERSIBLE INJURY Fig. 2.2 Sequenceofreversiblecellinjuryandcelldeath.Necrosisandapoptosisarethetwomajorpathwaysofcelldeathandarediscussedindetaillater. The causes of cell injury span a range from gross physical trauma, such as after a motor vehicle accident, to a single gene defect that results in a nonfunctional enzyme in a specific metabolic disease. Most injurious stimuli can be grouped into the following categories.


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Pathology_Robbins. Hypoxia and ischemia. Hypoxia, which refers to oxygen deficiency, and ischemia, which means reduced blood supply, are among the most common causes of cell injury. Both deprive tissues of oxygen, and ischemia, in addition, results in a deficiency of essential nutrients and a build up of toxic metabolites. The most common cause of hypoxia is ischemia resulting from an arterial obstruction, but oxygen deficiency also can result from inadequate oxygenation of the blood, as in a variety of diseases affecting the lung, or from reduction in the oxygen-carrying capacity of the blood, as with anemia of any cause, and carbon monoxide (CO) poisoning.


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Pathology_Robbins. Toxins. Potentially toxic agents are encountered daily in the environment; these include air pollutants, insecticides, CO, asbestos, cigarette smoke, ethanol, and drugs. Many drugs in therapeutic doses can cause cell or tissue injury in a susceptible patient or in many individuals if used excessively or inappropriately (Chapter 7). Even innocuous substances, such as glucose, salt, water and oxygen, can be toxic. Infectious agents. All types of disease-causing pathogens, including viruses, bacteria, fungi, and protozoans, injure cells. The mechanisms of cell injury caused by these diverse agents are discussed in Chapter 9.


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Pathology_Robbins. Immunologic reactions. Although the immune system defends the body against pathogenic microbes, immune reactions also can result in cell and tissue injury. Examples are autoimmune reactions against one’s own tissues, allergic reactions against environmental substances, and excessive or chronic immune responses to microbes (Chapter 5). In all of these situations, immune responses elicit inflammatory reactions, which are often the cause of damage to cells and tissues. Genetic abnormalities. Genetic aberrations can result in pathologic changes as conspicuous as the congenital malformations associated with Down syndrome or as subtle as the single amino acid substitution in hemoglobin giving rise to sickle cell anemia (Chapter 7). Genetic defects may cause cell injury as a consequence of deficiency of functional proteins, such as enzymes in inborn errors of metabolism, or accumulation of damaged DNA or misfolded proteins, both of which trigger cell death when they are beyond repair.


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Pathology_Robbins. Nutritional imbalances. Protein–calorie insufficiency among impoverished populations remains a major cause of cell injury, and specific vitamin deficiencies are not uncommon even in developed countries with high standards of living (Chapter 8). Ironically, excessive dietary intake may result in obesity and also is an important underlying factor in many diseases, such as type 2 diabetes mellitus and atherosclerosis. Physical agents. Trauma, extremes of temperature, radiation, electric shock, and sudden changes in atmospheric pressure all have wide-ranging effects on cells (Chapter 8). http://ebooksmedicine.net Aging. Cellular senescence results in a diminished ability of cells to respond to stress and, eventually, the death of cells and of the organism. The mechanisms underlying cellular aging are discussed at the end of this chapter.


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Pathology_Robbins. With this introduction, we proceed to a discussion of the progression and morphologic manifestations of cell injury, and then to the biochemical mechanisms in injury caused by different noxious stimuli. Although various injurious stimuli damage cells through diverse biochemical mechanisms, all tend to induce a stereotypic sequence of morphologic and structural alterations in most types of cells. Reversible injury is the stage of cell injury at which the deranged function and morphology of the injured cells can return to normal if the damaging stimulus is removed Fig. 2.3 ). In reversible injury, cells and intracellular organelles typically become swollen because they take in water as a result of the failure of energy-dependent ion pumps in the plasma membrane, leading to an inability to maintain ionic and fluid homeostasis. In some forms of injury, degenerated organelles and lipids may accumulate inside the injured cells.


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Pathology_Robbins. Thecytoplasmofinjuredcellsalsomaybecomeredder(eosinophilic),achangethatbecomesmuchmorepronouncedwithprogressiontonecrosis(describedlater).Otherintracellularchangesassociatedwithcellinjury( Fig.2.3 )include(1)plasmamembranealterationssuchasblebbing,blunting,ordistortionofmicrovilli,andlooseningofintercellularattachments; Swelling of endoplasmic reticulum and mitochondria Breakdown of plasma membrane, organelles, and nucleus; leakage of contents Fig. 2.3 Reversiblecellinjuryandnecrosis.Theprincipalcellularalterationsthatcharacterizereversiblecellinjuryandnecrosisareillustrated.Byconvention,reversibleinjuryisconsideredtoculminateinnecrosisiftheinjuriousstimulusisnotremoved. withdetachmentofribosomesanddissociationofpolysomes;and(4)nuclearalterations,suchasclumpingofchromatin.Thecytoplasmmaycontainso-called“myelinfigures,”whicharecollectionsofphospholipidsresemblingmyelinsheathsthatarederivedfromdamagedcellularmembranes.


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Pathology_Robbins. In some situations, potentially injurious insults induce specific alterations in cellular organelles, such as the ER. The smooth ER is involved in the metabolism of various chemicals, and cells exposed to these chemicals show hypertrophy of the ER as an adaptive response that may have important functional consequences. For instance, many drugs, including barbiturates, which were commonly used as sedatives in the past and are still used as a treatment for some forms of epilepsy, are metabolized in the liver by the cytochrome P-450 mixed-function oxidase system found in the smooth ER. Protracted use of http://ebooksmedicine.net


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Pathology_Robbins. Fig. 2.4 Morphologicchangesinreversibleandirreversiblecellinjury(necrosis).(A)Normalkidneytubuleswithviableepithelialcells.(B)Early(reversible)ischemicinjuryshowingsurfaceblebs,increasedeosinophiliaofcytoplasm,andswellingofoccasionalcells.(C)Necrotic(irreversible)injuryofepithelialcells,withlossofnucleiandfragmentationofcellsandleakageofcontents.(Courtesy of Drs. Neal Pinckard and M.A.Venkatachalam, University of Texas Health Sciences


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Pathology_Robbins. Center, San Antonio,Texas.) barbiturates leads to a state of tolerance, marked by the need to use increasing doses of the drug to achieve the same effect. This adaptation stems from hypertrophy (an increase in volume) of the smooth ER of hepatocytes and a consequent increase in P-450 enzymatic activity. P-450– mediated modification of compounds sometimes leads to their detoxification, but in other instances converts them into a dangerous toxin; one such example involves carbon tetrachloride (CCl4), discussed later. Cells adapted to one drug demonstrate an increased capacity to metabolize other compounds handled by the same system. Thus, if patients taking phenobarbital for epilepsy increase their alcohol intake, they may experience a drop in blood concentration of the anti-seizure medication to subtherapeutic levels because of smooth ER hypertrophy in response to the alcohol.


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Pathology_Robbins. With persistent or excessive noxious exposures, injured cells pass a nebulous “point of no return” and undergo cell death. The clinical relevance of defining this transition point is obvious—if the biochemical and molecular changes that predict cell death can be identified, it may be possible to devise strategies for preventing the transition from reversible to irreversible cell injury. Although there are no definitive morphologic or biochemical correlates of irreversibility, it is consistently characterized by three phenomena: the inability to restore mitochondrial function (oxidative phosphorylation and adenosine triphosphate [ATP] generation) even after resolution of the original injury; the loss of structure and functions of the plasma membrane and intracellular membranes; and the loss of DNA and chromatin structural integrity. As discussed in more detail later, injury to lysosomal membranes results in the enzymatic dissolution of the injured cell, which is the culmination of


Pathology_Robbins

Pathology_Robbins. loss of DNA and chromatin structural integrity. As discussed in more detail later, injury to lysosomal membranes results in the enzymatic dissolution of the injured cell, which is the culmination of necrosis.


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Pathology_Robbins. When cells are injured they die by different mechanisms, depending on the nature and severity of the insult. • Severe disturbances, such as loss of oxygen and nutrient supply and the actions of toxins, cause a rapid and uncontrollable form of death that has been called “accidental” cell death. The morphological manifestation of accidental cell death is necrosis (Greek, necros = death) ( Table 2.1 ). Necrosis is the major pathway of cell death in many commonly encountered injuries, such as those resulting from ischemia, exposure to toxins, various infections, and trauma. Necrosis is traditionally considered the inevitable end result of severe damage that is beyond salvage and is not thought to be regulated by specific signals or biochemical mechanisms; in other words, necrosis happens accidentally because the injury is too severe to be repaired and many cellular constituents simply fail or fall apart.


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Pathology_Robbins. • In contrast, when the injury is less severe, or cells need to be eliminated during normal processes, they activate a precise set of molecular pathways that culminate in death. Because this kind of cell death can be manipulated Table 2.1 Features of Necrosis and Apoptosis http://ebooksmedicine.net Fig. 2.5 Therelationshipamongcellularfunction,celldeath,andthemorphologicchangesofcellinjury.Notethatcellsmayrapidlybecomenonfunctionalaftertheonsetofinjury,althoughtheyarestillviable,withpotentiallyreversibledamage;withalongerdurationofinjury,irreversibleinjuryandcelldeathmayresult.Notealsothatcelldeathtypicallyprecedesultrastructural,lightmicroscopic,andgrosslyvisiblemorphologicchanges. by therapeutic agents or genetic mutations, it is said to be “regulated” cell death. The morphologic appearance of most types of regulated cell death is apoptosis (see


Pathology_Robbins

Pathology_Robbins. Table 2.1 ). In some instances, regulated cell death shows features of both necrosis and apoptosis, and has been called necroptosis. The discovery of these previously unrecognized forms of cell death that were regulated by identifiable genes and signaling pathways showed that cell death can be a controlled process. The idea of regulated cell death also raises the possibility that specific molecular pathways can be targeted therapeutically to prevent the loss of cells in pathologic conditions. Apoptosis is a process that eliminates cells with a variety of intrinsic abnormalities and promotes clearance of the fragments of the dead cells without eliciting an inflammatory reaction. This “clean” form of cell suicide occurs in pathologic situations when a cell’s DNA or proteins are damaged beyond repair or the cell is deprived of necessary survival signals. But unlike necrosis, which is always an indication of a pathologic process, apoptosis also occurs in healthy tissues. It serves to


Pathology_Robbins

Pathology_Robbins. beyond repair or the cell is deprived of necessary survival signals. But unlike necrosis, which is always an indication of a pathologic process, apoptosis also occurs in healthy tissues. It serves to eliminate unwanted cells during normal development and to maintain constant cell numbers, so it is not necessarily associated with pathologic cell injury. These types of physiologic cell death are also called programmed cell death.


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Pathology_Robbins. It is important to point out that cellular function may be lost long before cell death occurs, and that the morphologic changes of cell injury (or death) lag far behind loss of function and viability ( Fig. 2.5 ). For example, myocardial cells become noncontractile after 1 to 2 minutes of ischemia, but may not die until 20 to 30 minutes of ischemia have elapsed. Morphologic features indicative of the death of ischemic myocytes appear by electron microscopy within 2 to 3 hours after the death of the cells, but are not evident by light microscopy until 6 to 12 hours later. Necrosis is a form of cell death in which cellular membranes fall apart, and cellular enzymes leak out and ultimately digest the cell (


Pathology_Robbins

Pathology_Robbins. Necrosis is a form of cell death in which cellular membranes fall apart, and cellular enzymes leak out and ultimately digest the cell ( Fig. 2.3 ). Necrosis elicits a local host reaction, called inflammation, that is induced by substances released from dead cells and which serves to eliminate the debris and start the subsequent repair process (Chapter 3). The enzymes responsible for digestion of the cell are derived from lysosomes and may come from the dying cells themselves or from leukocytes recruited as part of the inflammatory reaction. Necrosis often is the culmination of reversible cell injury that cannot be corrected.


Pathology_Robbins

Pathology_Robbins. The biochemical mechanisms of necrosis vary with different injurious stimuli. These mechanisms include: failure of energy generation in the form of ATP because of reduced oxygen supply or mitochondrial damage; damage to cellular membranes, including the plasma membrane and lysosomal membranes, which results in leakage of cellular contents including enzymes; irreversible damage to cellular lipids, proteins, and nucleic acids, which may be caused by reactive oxygen species (ROS); and others. These biochemical mechanisms are discussed later when we consider the individual causes of cell necrosis. Figs.2.3 2.4C


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Pathology_Robbins. http://ebooksmedicine.net Fig. 2.6 Coagulativenecrosis.(A)Awedge-shapedkidneyinfarct(yellow) withpreservationoftheoutlines.(B)Microscopicviewoftheedgeoftheinfarct,withnormalkidney(N) andnecroticcellsintheinfarct(I).Thenecroticcellsshowpreservedoutlineswithlossofnuclei,andaninflammatoryinfiltrateis present(difficulttodiscernatthismagnification). Morphologic Patterns of Tissue Necrosis In severe pathologic conditions, large areas of a tissue or even entire orgrans may undergo necrosis. This may happen in association with marked ischemia, infections, and certain inflammatory reactions. There are several morphologically distinct patterns of tissue necrosis that may provide etiologic clues. Although the terms that describe these patterns do not reflect underlying mechanisms, such terms are commonly used and their implications are understood by pathologists and clinicians. grossappearances;theexceptionisfibrinoidnecrosis,whichis detectedonlybyhistologicexamination.


Pathology_Robbins

Pathology_Robbins. Fig.2.10 ).Theimmunologicallymediateddiseases(e.g.,polyarteritisnodosa)inwhichthistypeofnecrosisisseenaredescribedinChapter5. Leakage of intracellular proteins through the damaged cell membrane and ultimately into the circulation provides a means of detecting tissue-specific necrosis using blood or serum samples. Cardiac muscle, for example, contains a unique isoform of the enzyme creatine kinase and of the contractile protein troponin, whereas hepatic bile duct epithelium contains the enzyme alkaline phosphatase, and hepatocytes contain transaminases. Irreversible injury and cell death in these tissues elevate the serum levels of these proteins, which makes them clinically useful markers of tissue damage. Apoptosis is a pathway of cell death in which cells activate enzymes that degrade the cells’ own nuclear DNA and nuclear and cytoplasmic proteins (


Pathology_Robbins

Pathology_Robbins. Apoptosis is a pathway of cell death in which cells activate enzymes that degrade the cells’ own nuclear DNA and nuclear and cytoplasmic proteins ( Fig. 2.11 ). Fragments of the apoptotic cells then break off, giving the appearance that is responsible for the name (apoptosis, “falling off”). The plasma membrane of the apoptotic cell remains intact, but the membrane is altered in such a way that the fragments, called apoptotic bodies, become highly “edible,” leading to their rapid consumption by phagocytes. The dead cell and its fragments are cleared with little leakage of cellular contents, so apoptotic cell death does not elicit an inflammatory reaction. Thus, apoptosis differs in many respects from necrosis ( Table 2.1 Causes of Apoptosis Apoptosis occurs in many normal situations and serves to eliminate potentially harmful cells and cells that have http://ebooksmedicine.net


Pathology_Robbins

Pathology_Robbins. Table 2.1 Causes of Apoptosis Apoptosis occurs in many normal situations and serves to eliminate potentially harmful cells and cells that have http://ebooksmedicine.net Fig. 2.11 Apoptosis.Thecellularalterationsinapoptosisareillustrated.Contrastthesewiththechangesthatcharacterizenecroticcelldeath,shownin Fig.2.3 .Apoptotic body Phagocyte Phagocytosis of apoptotic cells and fragments NORMALCELLAPOPTOSIS Table 2.2 Physiologic and Pathologic Conditions Associated With Apoptosis Condensation of chromatin Table 2.2 ). It also occurs as a pathologic event when cells are damaged, especially when the damage affects the cell’s DNA or proteins; thus, the irreparably damaged cell is eliminated.


Pathology_Robbins

Pathology_Robbins. Table 2.2 ). It also occurs as a pathologic event when cells are damaged, especially when the damage affects the cell’s DNA or proteins; thus, the irreparably damaged cell is eliminated. • Physiologic apoptosis. During normal development of an organism, some cells die and are replaced by new ones. In mature organisms, highly proliferative and hormone-responsive tissues undergo cycles of proliferation and cell loss that are often determined by the levels of growth factors. In these situations, the cell death is always by apoptosis, ensuring that unwanted cells are eliminated without eliciting potentially harmful inflammation. In the immune system, apoptosis eliminates excess leukocytes left at the end of immune responses as well as lymphocytes that recognize self-antigens and could cause autoimmune diseases if they were not purged.


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Pathology_Robbins. TurnoverofproliferativetissuesLossofgrowthfactorsignaling(e.g.,intestinalepithelium,(presumedmechanism)lymphocytesinbonemarrow,andthymus) • Apoptosis in pathologic conditions. Apoptosis eliminates cells that are damaged beyond repair. This is seen when there is severe DNA damage, for example, after exposure to radiation and cytotoxic drugs. The accumulation of misfolded proteins also triggers apoptotic death; the underlying mechanisms of this cause of cell death and its significance in disease are discussed later, in the context of ER stress. Certain infectious agents, particularly some viruses, induce apoptotic death of infected cells. Mechanisms of Apoptosis


Pathology_Robbins

Pathology_Robbins. Mechanisms of Apoptosis Apoptosis is regulated by biochemical pathways that control the balance of death-and survival-inducing signals and ultimately the activation of enzymes called caspases. Caspases were so named because they are cysteine proteases that cleave proteins after aspartic acid residues. Two distinct pathways converge on caspase activation: the mitochondrial pathway and the death receptor pathway ( Fig. 2.12 ). Although these pathways can intersect, they are generally induced under different conditions, involve different molecules, and serve distinct roles in physiology and disease. The end result of apoptotic cell death is the clearance of apoptotic bodies by phagocytes. • The mitochondrial (intrinsic) pathway seems to be responsible for apoptosis in most physiologic and pathologic situations. Mitochondria contain several proteins that are capable of inducing apoptosis, including cytochrome c. When mitochondrial membranes http://ebooksmedicine.net


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Pathology_Robbins. PhagocyteMitochondria Receptor-ligand interactions • Fas • TNF receptor Cell injury • Growth factor withdrawal • DNA damage (by radiation, toxins, free radicals) • Protein misfolding (ER stress) Cytochrome c and other pro-apoptotic proteins Regulators (Bcl-2, Bcl-xL) Caspase activation Adaptor proteins Breakdown of cytoskeleton Endonuclease activation Ligands for phagocytic cell receptors Apoptotic body Nuclear fragmentation BH3 sensors Bcl-2 family effectors (Bax, Bak)


Pathology_Robbins


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Pathology_Robbins. become permeable, cytochrome c leaks out into the cytoplasm, triggering caspase activation and apoptotic death. A family of more than 20 proteins, the prototype of which is Bcl-2, controls the permeability of mitochondria. In healthy cells, Bcl-2 and the related protein Bcl-xL, which are produced in response to growth factors and other stimuli, maintain the integrity of mitochondrial membranes, in large part by holding two proapoptotic members of the family, Bax and Bak, in check. When cells are deprived of growth factors and survival signals, or are exposed to agents that damage DNA, or accumulate unacceptable amounts of misfolded proteins, a number of sensors are activated. These sensors are called BH3 proteins because they contan the third domain seen in Bcl-family proteins. They in turn shift this delicate, life-sustaining balance in favor of pro-apoptotic Bak and Bax. As a result, Bak and Bax dimerize, insert into the mitochondrial membrane, and form channels through which


Pathology_Robbins

Pathology_Robbins. in turn shift this delicate, life-sustaining balance in favor of pro-apoptotic Bak and Bax. As a result, Bak and Bax dimerize, insert into the mitochondrial membrane, and form channels through which cytochrome c and other mitochondrial proteins escape into the cytosol. After cytochrome c enters the cytosol, it, together with certain cofactors, activates caspase-9. The net result is the activation of a caspase cascade, ultimately leading to nuclear fragmentation and formation of apoptotic bodies.


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Pathology_Robbins. • The death receptor (extrinsic) pathway of apoptosis. Many cells express surface molecules, called death receptors, that trigger apoptosis. Most of these are members of the tumor necrosis factor (TNF) receptor family, which contain in their cytoplasmic regions a conserved “death domain,” so named because it mediates interaction with other proteins involved in cell death. The prototypic death receptors are the type I TNF receptor and Fas (CD95). Fas ligand (FasL) is a membrane protein expressed mainly on activated T lymphocytes. When these T cells recognize Fas-expressing targets, Fas molecules are crosslinked by FasL and bind adaptor proteins via the death domain. These then recruit and activate caspase-8, which, in turn, activates downstream caspases. The death receptor pathway is involved in the elimination of self-reactive lymphocytes and in the killing of target cells by some cytotoxic T lymphocytes (CTLs) that express FasL.


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Pathology_Robbins. In either pathway, after caspase-9 or caspase-8 is activated, it cleaves and thereby activates additional caspases that cleave numerous targets and ultimately activate enzymes that degrade the cells’ proteins and nucleus. The end result is the characteristic cellular fragmentation of apoptosis.


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Pathology_Robbins. • Clearance of apoptotic cells. Apoptotic cells and their fragments entice phagocytes by producing a number of “eat-me” signals. For instance, in normal cells, phosphatidylserine is present on the inner leaflet of the plasma membrane, but in apoptotic cells this phospholipid “flips” to the outer leaflet, where it is recognized by tissue macrophages, leading to phagocytosis of the apoptotic cells. Cells that are dying by apoptosis also secrete soluble factors that recruit phagocytes. The plasma membrane alterations and secreted proteins facilitate prompt clearance of the dead cells before the cells undergo membrane damage and release their contents (which can induce inflammation). Numerous macrophage receptors have been shown to be involved in the binding and engulfment of apoptotic cells. The phagocytosis of apoptotic cells is so efficient that dead cells disappear without leaving a trace, and inflammation is virtually absent. http://ebooksmedicine.net


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Pathology_Robbins. http://ebooksmedicine.net MORPHOLOGYInH&E-stainedtissuesections,thenucleiofapoptoticcellsshowvariousstagesofchromatincondensationandaggregationand,ultimately,karyorrhexis( Fig.2.13 );atthemolecularlevel,thisisreflectedinthefragmentationofDNAintonucleosome-sizedpieces.Thecellsrapidlyshrink,formcytoplasmicbuds,andfragmentintoapoptoticbodiesthatarecomposedofmembrane- boundpiecesofcytosolandorganelles(Fig.2.11 ).Becausethesefragmentsarequicklyextrudedandphagocytosedwithoutelicitinganinflammatoryresponse,evensubstantialapoptosismaybehistologicallyundetectable. In addition to necrosis and apoptosis, two other patterns of cell death have been described that have unusual features. Although the importance of these pathways in disease remains to be established, they are the subjects of considerable current research, and it is useful to be aware of the basic concepts.


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Pathology_Robbins. Necroptosis. This form of cell death is initiated by engagement of TNF receptors as well as other, poorly defined triggers. Unlike the extrinsic pathway of apoptosis, which also is downstream of TNF receptors, in necroptosis, kinases called receptor-interacting protein (RIP) kinases are activated, initiating a series of events that result in the dissolution of the cell, much like necrosis. The name necroptosis implies that there are features of both necrosis and apoptosis. Some infections are believed to kill cells by this pathway, and it has been hypothesized to play a role in ischemic injury and other pathologic situations, especially those associated with inflammatory reactions in which the cytokine TNF is produced. However, when and why it occurs and how significant it is in human diseases is not well understood.


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Pathology_Robbins. Pyroptosis. This form of cell death is associated with activation of a cytosolic danger-sensing protein complex called the inflammasome (Chapter 5). The net result of inflammasome activation is the activation of caspases, some of which induce the production of cytokines that induce inflammation, often manifested by fever, and others trigger apoptosis. Thus, apoptosis and inflammation coexist. The name pyroptosis stems from the association of apoptosis with fever (Greek, pyro = fire). It is thought to be one mechanism by which some infectious


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Pathology_Robbins. Fig. 2.13 Morphologicappearanceofapoptoticcells.Apoptoticcells(someindicatedbyarrows)incolonicepitheliumareshown.(Somepreparativeregimensforcolonoscopymayinduceapoptosisinepithelialcells,whichexplainsthepresenceofdeadcellsinthisbiopsy.)Notethefragmentednucleiwithcondensedchromatinandtheshrunkencellbodies,somewithpiecesfallingoff.(Courtesy of Dr. Sanjay Kakar, Department of Pathology, University of California San Francisco, San Francisco, California.) microbes cause the death of infected cells. Its role in other pathologic situations is unknown.


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Pathology_Robbins. Autophagy (“self-eating”) refers to lysosomal digestion of the cell’s own components. It is a survival mechanism in times of nutrient deprivation, so that the starved cell can live by eating its own contents and recycling these contents to provide nutrients and energy. In this process, intracellular organelles and portions of cytosol are first sequestered within an ER-derived autophagic vacuole, whose formation is initiated by cytosolic proteins that sense nutrient deprivation ( Fig. 2.14 ). The vacuole fuses with lysosomes to form an autophagolysosome, in which lysosomal enzymes digest the cellular components. In some circumstances, autophagy may be associated with atrophy of tissues (discussed later) and may represent an adaptation that helps cells survive lean times. If, however, the starved cell can no longer cope by devouring its contents, autophagy may eventually lead to apoptotic cell death.


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Pathology_Robbins. Extensive autophagy is seen in ischemic injury and some types of myopathies. Polymorphisms in a gene involved in autophagy have been associated with inflammatory bowel disease, but the mechanistic link between autophagy and intestinal inflammation is not known. The role of autophagy in cancer is discussed in Chapter 6. Thus, a once little-appreciated survival pathway in cells may prove to have wide-ranging roles in human disease. http://ebooksmedicine.net Fig. 2.14 Autophagy. Cellular stresses, such as nutrient deprivation, activate autophagy genes, which initiate the formation of membrane-bound vesicles in which cellular organelles are sequestered. These vesicles fuse with lysosomes, in which the organelles are digested, and the products are used to provide nutrients for the cell. The same process can trigger apoptosis by mechanisms that are not well defined.


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Pathology_Robbins. Reversible cell injury: cell swelling, fatty change, plasma membrane blebbing and loss of microvilli, mitochondrial swelling, dilation of the ER, eosinophilia (resulting from decreased cytoplasmic RNA) Necrosis: Accidental cell death manifested by increased cytoplasmic eosinophilia; nuclear shrinkage, fragmentation, and dissolution; breakdown of plasma membrane and organellarmembranes; abundant myelin figures; leakage and enzymaticdigestion of cellular contents Morphologic types of tissue necrosis: under different conditions, necrosis in tissues may assume specific patterns: coagulative, liquefactive, gangrenous, caseous, fat, and fibrinoid. Apoptosis: regulated mechanism of cell death that serves toeliminate unwanted and irreparably damaged cells, with theleast possible host reaction, characterized by enzymatic degradation of proteins and DNA, initiated by caspases; and byrapid recognition and removal of dead cells by phagocytes Apoptosis is initiated by two major pathways:


Pathology_Robbins

Pathology_Robbins. Apoptosis is initiated by two major pathways: Mitochondrial (intrinsic) pathway is triggered by loss ofsurvival signals, DNA damage, and accumulation ofmisfolded proteins (ER stress); associated with leakage ofproapoptotic proteins from mitochondrial membrane intothe cytoplasm, where they trigger caspase activation; inhibited by anti-apoptotic members of the Bcl family, which are induced by survival signals including growthfactors. Death receptor (extrinsic) pathway is responsible forelimination of self-reactive lymphocytes and damage byCTLs; is initiated by engagement of death receptors(members of the TNF receptor family) by ligands onadjacent cells. Two other unusual pathways of cell death are necroptosis(which, as the name implies, includes features of both necrosisand apoptosis and is regulated by particular signaling pathways)and pyroptosis, which can lead to the release of proinflammatory cytokines and may initiate apoptosis.


Pathology_Robbins

Pathology_Robbins. Autophagy is an adaptation to nutrient deprivation in whichcells digest their own organelles and recycle them to provideenergy and substrates. If the stress is too severe for theprocess to cope with it, it results in cell death by apoptosis. Before discussing individual mechanisms of cell injury and death, some general principles should be emphasized. The cellular response to injurious stimuli depends on the type of injury, its duration, and its severity. Thus, low doses of toxins or a brief period of ischemia may lead to reversible cell injury, whereas larger toxin doses or longer ischemic times may result in irreversible injury and cell death.


Pathology_Robbins

Pathology_Robbins. The consequences of an injurious stimulus also depend on the type, status, adaptability, and genetic makeup of the injured cell. The same injury has vastly different outcomes depending on the cell type. For instance, striated skeletal muscle in the leg tolerates complete ischemia for 2 to 3 hours without irreversible injury, whereas cardiac muscle dies after only 20 to 30 minutes of ischemia. The nutritional (or hormonal) status also can be important; understandably, a glycogen-replete hepatocyte will survive ischemia better than one that has just burned its last glucose molecule. Genetically determined diversity in metabolic pathways can contribute to differences in responses to injurious stimuli. For instance, when exposed to the same dose of a toxin, individuals who inherit variants in genes encoding cytochrome P-450 may catabolize the toxin at different rates, leading to different outcomes. Much effort is now directed toward understanding the role of genetic polymorphisms in


Pathology_Robbins

Pathology_Robbins. in genes encoding cytochrome P-450 may catabolize the toxin at different rates, leading to different outcomes. Much effort is now directed toward understanding the role of genetic polymorphisms in responses to drugs and toxins, a field of study called pharmacogenomics. In fact, genetic variations influence susceptibility to many complex diseases as well as responsiveness to various therapeutic agents. Using the genetic makeup of the individual patient to guide therapy is one example of “precision medicine.”


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Pathology_Robbins. Cell injury usually results from functional and biochemical abnormalities in one or more of a limited number of essential cellular components ( Fig. 2.15 ). As we discuss in more detail later, different external insults and endogenous perturbations typically affect different cellular organelles and biochemical pathways. For instance, deprivation of oxygen and nutrients (as 42CHAPTER 2 Cell Injury, Cell Death, and Adaptations Fig. 2.15 The principal biochemical mechanisms and sites of damage in cell injury. Note that causes and mechanisms of cell death by necrosis and apoptosis are shown as being independent but there may be overlap; for instance, both may contribute to cell death caused by ischemia, oxidative stress, or radiation. ATP, Adenosine triphosphate; ROS, reactive oxygen species.


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Pathology_Robbins. in hypoxia and ischemia) primarily impairs energy-dependent cellular functions, culminating in necrosis, whereas damage to proteins and DNA triggers apoptosis. However, it should be emphasized that the very same injurious agent may trigger multiple and overlapping biochemical pathways. Not surprisingly, therefore, it has proved difficult to prevent cell injury by targeting an individual pathway.


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Pathology_Robbins. As we discussed in the beginning of this chapter and have alluded to throughout, there are numerous and diverse causes of cell injury and cell death. Similarly, there are many biochemical pathways that can initiate the sequence of events that lead to cell injury and culminate in cell death. Some of these pathways are recognized to play important roles in human diseases. In the following section, we organize our discussion of the mechanisms of cell injury along its major causes and pathways, and discuss the principal biochemical alterations in each. For the sake of clarity and simplicity, we emphasize the unique mechanisms in each pathway, but it is important to point out that any initiating trigger may activate one or more of these mechanisms, and several mechanisms may be active simultaneously.


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Pathology_Robbins. Deficiency of oxygen leads to failure of many energy-dependent metabolic pathways, and ultimately to death of cells by necrosis. Most cellular ATP is produced from adenosine diphosphate (ADP) by oxidative phosphorylation during reduction of oxygen in the electron transport system of mitochondria. High-energy phosphate in the form of ATP is required for membrane transport, protein synthesis, lipogenesis, and the deacylation-reacylation reactions necessary for phospholipid turnover. It is estimated that in total, the cells of a healthy human burn 50 to 75 kg of ATP every day! Not surprisingly, therefore, cells deprived of oxygen are at risk of suffering catastrophic failure of many essential functions. Oxygen deprivation is one of the most frequent causes of cell injury and necrosis in clinical medicine.


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Pathology_Robbins. Cells subjected to the stress of hypoxia that do not immediately die activate compensatory mechanisms that are induced by transcription factors of the hypoxia-inducible factor 1 (HIF-1) family. HIF-1 simulates the synthesis of several proteins that help the cell to survive in the face of low oxygen. Some of these proteins, such as vascular endothelial growth factor (VEGF), stimulate the growth of new vessels and thus attempt to increase blood flow and the supply of oxygen. Other proteins induced by HIF-1 cause adaptive changes in cellular metabolism by stimulating the uptake of glucose and glycolysis and dampening mitochondrial oxidative phosphorylation. Anaerobic glycolysis can generate ATP in the absence of oxygen using glucose derived either from the circulation or from the hydrolysis of intracellular glycogen. Understandably, normal tissues with a greater glycolytic capacity because of the presence of glycogen (e.g., the liver and striated muscle) are more likely to survive


Pathology_Robbins

Pathology_Robbins. of intracellular glycogen. Understandably, normal tissues with a greater glycolytic capacity because of the presence of glycogen (e.g., the liver and striated muscle) are more likely to survive hypoxia and decreased oxidative phosphorylation than tissues with limited glucose stores (e.g., the brain). Although it seems counterintuitive, rapidly proliferating normal cells and cancer cells rely on aerobic glycolysis to produce much of their energy, a phenomenon referred to as the Warburg effect. The reason for this is that although glycolysis yields less ATP per molecule of glucose burned than oxidative phosphorylation, metabolites generated by glycolysis and the TCA cycle serve as precursors for the synthesis of cellular constituents (e.g., proteins, lipids, and nucleic acids) that are needed for cell growth and division. Alterations in cellular metabolism are frequently seen in cancer cells, so they are discussed in more detail in Chapter 6.


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Pathology_Robbins. Persistent or severe hypoxia and ischemia ultimately lead to failure of ATP generation and depletion of ATP in cells. Loss of this critical energy store has deleterious effects on many cellular systems ( Fig. 2.16 Reduced activity of plasma membrane ATP-dependent sodium pumps, resulting in intracellular accumulation of sodium and efflux of potassium. The net gain of solute is accompanied by isoosmotic gain of water, causing cell swelling and dilation of the ER. The compensatory increase in anaerobic glycolysis leads to lactic acid accumulation, decreased intracellular pH, and decreased activity of many cellular enzymes. Prolonged or worsening depletion of ATP causes structural disruption of the protein synthetic apparatus, Fig. 2.16 Thefunctionalandmorphologicconsequencesofhypoxiaandischemia.ER, Endoplasmicreticulum. manifested as detachment of ribosomes from the rough ER (RER) and dissociation of polysomes into monosomes, with a consequent reduction in protein synthesis.


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Pathology_Robbins. manifested as detachment of ribosomes from the rough ER (RER) and dissociation of polysomes into monosomes, with a consequent reduction in protein synthesis. It also has been suggested that hypoxia per se increases the accumulation of ROS. Whether this is true is a matter of debate; however, there is ample evidence that hypoxia predisposes cells to ROS-mediated damage if blood flow (and oxygen delivery) is reestablished, a phenomenon called reperfusion injury (described later). Ultimately, there is irreversible damage to mitochondrial and lysosomal membranes, and the cell undergoes necrosis. Membrane damage is a late event in cell injury caused by diverse mechanisms, and is discussed later. Although necrosis is the principal form of cell death caused by hypoxia, apoptosis by the mitochondrial pathway is also thought to contribute.


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Pathology_Robbins. The functional consequences of hypoxia and ischemia depend on the severity and duration of the deficit. For instance, the heart muscle ceases to contract within 60 seconds of coronary occlusion. If hypoxia continues, worsening ATP depletion causes further deterioration, undergoing the sequence of changes illustrated in Fig. 2.3 and described earlier. Under certain circumstances, the restoration of blood flow to ischemic but viable tissues results, paradoxically, in increased cell injury. This is the reverse of the expected outcome of the restoration of blood flow, which normally results in the recovery of reversibly injured cells. This so-called “ischemia-reperfusion injury” is a clinically important process that may contribute significantly to tissue damage, especially after myocardial and cerebral ischemia. Several mechanisms may account for the exacerbation of cell injury resulting from reperfusion of ischemic tissues:


Pathology_Robbins

Pathology_Robbins. Several mechanisms may account for the exacerbation of cell injury resulting from reperfusion of ischemic tissues: New damage may be initiated during reoxygenation by increased generation of ROS (described in more detail below). Some of the ROS may be generated by injured cells with damaged mitochondria that cannot carry out the complete reduction of oxygen, and at the same time cellular anti-oxidant defense mechanisms may be compromised by ischemia, exacerbating the situation. ROS generated by infiltrating leukocytes also may contribute to the damage of vulnerable injured cells.


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Pathology_Robbins. The inflammation that is induced by ischemic injury may increase with reperfusion because it enhances the influx of leukocytes and plasma proteins. The products of activated leukocytes may cause additional tissue injury (Chapter 3). Activation of the complement system also may contribute to ischemia-reperfusion injury. Complement proteins may bind to the injured tissues, or to antibodies that are deposited in the tissues, and subsequent complement activation generates byproducts that exacerbate the cell injury and inflammation. Oxidative stress refers to cellular abnormalities that are induced by ROS, which belong to a group of molecules known as free radicals. Free radical-mediated cell injury is seen in many circumstances, including chemical and radiation injury, hypoxia, cellular aging, tissue injury caused by inflammatory cells, and ischemia-reperfusion injury. In all these cases, cell death may be by necrosis, apoptosis, or the mixed pattern of necroptosis.


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Pathology_Robbins. Free radicals are chemical species with a single unpaired electron in an outer orbit. Such chemical states are extremely unstable, and free radicals readily react with inorganic and organic molecules; when generated in cells, they avidly attack nucleic acids as well as a variety of cellular proteins and lipids. In addition, free radicals initiate reactions in which molecules that react with the free radicals are themselves converted into other types of free radicals, thereby propagating the chain of damage. Generation and Removal of Reactive Oxygen Species The accumulation of ROS is determined by their rates of production and removal ( Fig. 2.17 ). ROS are produced by two major pathways.


Pathology_Robbins

Pathology_Robbins. Generation and Removal of Reactive Oxygen Species The accumulation of ROS is determined by their rates of production and removal ( Fig. 2.17 ). ROS are produced by two major pathways. • ROS are produced normally in small amounts in all cells during the reduction-oxidation (redox) reactions that occur during mitochondrial respiration and energy generation. In this process, molecular oxygen is reduced in mitochondria to generate water by the sequential addition of four electrons. This reaction is imperfect, however, and small amounts of highly reactive but short-lived toxic intermediates are generated when oxygen is only partially reduced. These intermediates include superoxide (O2 • ), which is converted to hydrogen peroxide (H2O2) spontaneously and by the action of the enzyme superoxide dismutase (SOD). H2O2 is more stable than O2 • and can cross biologic membranes. In the presence of metals, such as Fe2+, H2O2 is converted to the http://ebooksmedicine.net


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Pathology_Robbins. Superoxide O2 Hydrogen peroxide Conversion Decomposition to Hydroxyl radical Lipid peroxidation Protein modifications DNA damage Membrane damage Breakdown, misfolding Mutations H2O2 OH Production of ROS: Pathologic effects Radiation Toxins Reperfusion to H2O2 H2O by glutathione by SOD peroxidase, catalase Fig. 2.17 Thegeneration,removal,androleofreactiveoxygenspecies(ROS)incellinjury.TheproductionofROSisincreasedbymanyinjuriousstimuli.Thesefreeradicalsareremovedbyspontaneousdecayandbyspecializedenzymaticsystems.Excessiveproductionorinadequateremovalleadstoaccumulationoffreeradicalsincells,whichmaydamagelipids(byperoxidation),proteins,andDNA,resultingincellinjury.SOD, Superoxidedismutase. highly reactive hydroxyl radical • OH by the Fenton reaction. The properties and pathologic effects of the major ROS are summarized in Table 2.3


Pathology_Robbins

Pathology_Robbins. highly reactive hydroxyl radical • OH by the Fenton reaction. The properties and pathologic effects of the major ROS are summarized in Table 2.3 ROS are produced in phagocytic leukocytes, mainly neutrophils and macrophages, as a weapon for destroying ingested microbes and other substances during inflammation and host defense (Chapter 3). The ROS are generated in the phagosomes and phagolysosomes of leukocytes by a process that is similar to mitochondrial respiration and is called the respiratory burst (or oxidative burst). In this process, a phagosome membrane enzyme catalyzes the generation of superoxide, which is converted to H2O2. H2O2 is in turn converted to a highly reactive compound, hypochlorite (the major component of household bleach), by the enzyme myeloperoxidase, which is present in leukocytes. The role of ROS in inflammation is described in Chapter 3.


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Pathology_Robbins. Nitric oxide (NO) is another reactive free radical produced in macrophages and other leukocytes. It can react with O2 − to form a highly reactive compound, peroxynitrite, which also participates in cell injury. The generation of free radicals is increased under several circumstances: The absorption of radiant energy (e.g., ultraviolet (UV) light, x-rays). Ionizing radiation can hydrolyze water into hydroxyl (•OH) and hydrogen (H•) free radicals. Table 2.3 Principal Free Radicals Involved in Cell Injury The enzymatic metabolism of exogenous chemicals (e.g., carbon tetrachloride—see later) Inflammation, in which free radicals are produced by leukocytes (Chapter 3) Reperfusion of ischemic tissues, as already described.


Pathology_Robbins

Pathology_Robbins. Inflammation, in which free radicals are produced by leukocytes (Chapter 3) Reperfusion of ischemic tissues, as already described. Cells have developed mechanisms to remove free radicals and thereby minimize their injurious effects. Free radicals are inherently unstable and decay spontaneously. There also are nonenzymatic and enzymatic systems, sometimes called free radical scavengers, serving to inactivate free radicals ( Fig. 2.17 The rate of decay of superoxide is significantly increased by the action of superoxide dismutase (SOD). Glutathione (GSH) peroxidases are a family of enzymes whose major function is to protect cells from oxidative damage. The most abundant member of this family, GSH peroxidase 1, is found in the cytoplasm of all cells. It catalyzes the breakdown of H2O2 by the reaction 2GSH + H2O2 → GS-SG + 2H2O. The intracellular ratio of oxidized GSH to reduced GSH is a reflection of this enzyme’s activity and thus of the cell’s ability to catabolize free radicals.


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Pathology_Robbins. Catalase, present in peroxisomes, catalyzes the decomposition of hydrogen peroxide (2H2O2 → O2 + 2H2O). It is one of the most active enzymes known, capable of degrading millions of molecules of H2O2 per second. http://ebooksmedicine.net • Endogenous or exogenous anti-oxidants (e.g., vitamins E, A, and C and β-carotene) may either block the formation of free radicals or scavenge them after they have formed. ROS causes cell injury by damaging multiple components of cells ( Fig. 2.17 Lipid peroxidation of membranes. Double bonds in membrane polyunsaturated lipids are vulnerable to attack by oxygen-derived free radicals. The lipid–radical interactions yield peroxides, which are themselves unstable and reactive, and an autocatalytic chain reaction ensues. Damage to plasma membranes as well as mitochondrial and lysosomal membranes can have devastating consequences, as discussed earlier in the context of ischemia and hypoxia.


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Pathology_Robbins. Crosslinking and other changes in proteins. Free radicals promote sulfhydryl-mediated protein crosslinking, resulting in enhanced degradation or loss of enzymatic activity. Free radical reactions also may directly cause polypeptide fragmentation. Damaged proteins may fail to fold properly, triggering the unfolded protein response, described later. DNA damage. Free radical reactions with thymine residues in nuclear and mitochondrial DNA produce single-strand breaks. Such DNA damage has been implicated in apoptotic cell death, aging, and malignant transformation of cells. In addition to the role of ROS in cell injury and the killing of microbes, low concentrations of ROS are involved in numerous signaling pathways in cells and thus in many physiologic reactions. Therefore, these molecules are produced normally but, to avoid their harmful effects, their intracellular concentrations are tightly regulated in healthy cells.


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Pathology_Robbins. Toxins, including environmental chemicals and substances produced by infectious pathogens, induce cell injury that culminates primarily in necrotic cell death. Different types of toxins induce cell injury by two general mechanism: Direct-acting toxins. Some toxins act directly by combining with a critical molecular component or cellular organelle. For example, in mercuric chloride poisoning (as may occur from ingestion of contaminated seafood) (Chapter 8), mercury binds to the sulfhydryl groups of various cell membraneproteins, causinginhibition ofATP-dependent transport and increased membrane permeability. Many anti-neoplastic chemotherapeutic agents also induce cell damage by direct cytotoxic effects. Also included in this class are toxins made by microorganisms (described in Chapter 9). These often cause damage by targeting host cell molecules that are needed for essential functions, such as protein synthesis and ion transport.


Pathology_Robbins

Pathology_Robbins. Latent toxins. Many toxic chemicals are not intrinsically active but must first be converted to reactive metabolites, which then act on target cells. Understandably, such toxins typically affect the cells in which they are activated. This is usually accomplished by cytochrome


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Pathology_Robbins. P-450 in the smooth ER of the liver and other organs. Although the metabolites might cause membrane damage and cell injury by direct covalent binding to protein and lipids, the most important mechanism of cell injury involves the formation of free radicals. Carbon tetrachloride (CCl4)—once widely used in the dry cleaning industry but now banned—and the analgesic acetaminophen belong in this category. The effect of CCl4 is still instructive as an example of chemical injury. CCl4 is converted to a toxic free radical, principally in the liver, and this free radical is the cause of cell injury, mainly by membrane phospholipid peroxidation. In less than 30 minutes after exposure to CCl4, there is sufficient damage to the ER membranes of hepatocytes to cause a decline in the synthesis of enzymes and plasma proteins; within 2 hours, swelling of the smooth ER and dissociation of ribosomes from the RER have occurred. There also is decreased synthesis of apoproteins that form complexes with


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Pathology_Robbins. and plasma proteins; within 2 hours, swelling of the smooth ER and dissociation of ribosomes from the RER have occurred. There also is decreased synthesis of apoproteins that form complexes with triglycerides and thereby facilitate triglyceride secretion; this defect results in the accumulation of lipids in hepatocytes and other cells and the “fatty liver” of CCl4 poisoning. Mitochondrial injury follows, and subsequently diminished ATP stores result in defective ion transport and progressive cell swelling; the plasma membranes are further damaged by fatty aldehydes produced by lipid peroxidation in the ER. The end result can be cell death.


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Pathology_Robbins. The accumulation of misfolded proteins in a cell can stress compensatory pathways in the ER and lead to cell death by apoptosis. During normal protein synthesis, chaperones in the ER control the proper folding of newly synthesized proteins, and misfolded polypeptides are ubiquitinated and targeted for proteolysis. If unfolded or misfolded proteins accumulate in the ER, they first induce a protective cellular response that is called the unfolded protein response ( Fig. 2.18 ). This adaptive response activates signaling pathways that increase the production of chaperones and decrease protein translation, thus reducing the levels of misfolded proteins in the cell. When a large amount of mis-folded protein accumulates and cannot be handled by the adaptive response, the signals that are generated result in activation of proapoptotic sensors of the BH3-only family as well as direct activation of caspases, leading to apoptosis by the mitochondrial (intrinsic) pathway.


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Pathology_Robbins. Intracellular accumulation of misfolded proteins may be caused by abnormalities that increase the production of misfolded proteins or reduce the ability to eliminate them. This may result from gene mutations that lead to the production of proteins that cannot fold properly; aging, which is associated with a decreased capacity to correct misfolding; infections, especially viral infections, when large amounts of microbial proteins are synthesized within cells, more than the cell can handle; increased demand for secretory proteins such as insulin in insulin-resistant states; and changes in intracellular pH and redox state. Protein misfolding is thought to be the fundamental cellular abnormality in several neurodegenerative diseases (Chapter 23). Deprivation of glucose and oxygen, as in ischemia and hypoxia, also may increase the burden of misfolded proteins. http://ebooksmedicine.net 46CHAPTER 2 Cell Injury, Cell Death, and Adaptations


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Pathology_Robbins. http://ebooksmedicine.net 46CHAPTER 2 Cell Injury, Cell Death, and Adaptations Sensor of misfolded proteins (e.g., IRE1) Signaling Signaling Cytosol Increased Reduced Increased Activation of Activation ofsynthesis of protein protein BH3 proteins caspaseschaperones synthesis degradation Reduced load of misfolded proteins ADAPTIVE TERMINAL UNFOLDED PROTEIN UNFOLDED PROTEIN RESPONSE RESPONSE: APOPTOSIS


Pathology_Robbins

Pathology_Robbins. Reduced load of misfolded proteins ADAPTIVE TERMINAL UNFOLDED PROTEIN UNFOLDED PROTEIN RESPONSE RESPONSE: APOPTOSIS Fig. 2.18 The unfolded protein response and endoplasmic reticulum (ER) stress. The presence of misfolded proteins in the ER is detected by sensors in the ER membrane, such as the kinase IRE-1, which form oligomers that are activated by phosphorylation. This triggers an adaptive unfolded protein response, which can protect the cell from the harmful consequences of the misfolded proteins. When the amount of misfolded proteins is too great to be corrected, excessive activation of ER sensors activates the mitochondrial pathway of apoptosis and the irreparably damaged cell dies; this is also called the terminal unfolded protein response.


Pathology_Robbins

Pathology_Robbins. Protein misfolding within cells may cause disease by • Cell death as a result of protein misfolding is recognized creating a deficiency of an essential protein or by induc-as a feature of a number of diseases, including the neuing apoptosis ( Table 2.4 ). rodegenerative disorders Alzheimer disease, Hunting • Misfolded proteins often lose their activity and are rapidly ton disease, and Parkinson disease, and may underlie degraded, both of which can contribute to a loss of func-type 2 diabetes as well ( Table 2.4 ). tion. If this function is essential, cellular injury ensues. One important disease in which this occurs is cystic fibro-As discussed later in the chapter, improperly folded sis, which is caused by inherited mutations in a mem-proteins can also accumulate in extracellular tissues, as in brane transport protein that prevent its normal folding. amyloidosis. Table 2.4 Diseases Caused by Misfolded Proteins


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Pathology_Robbins. Table 2.4 Diseases Caused by Misfolded Proteins Diseases Caused by Mutant Proteins That Are Degraded, Leading to Their Deficiency Retinitis pigmentosaRhodopsinAbnormal folding of rhodopsin causes photoreceptor loss and cell death, resulting in blindness Diseases Caused by Misfolded Proteins That Result From Both ER Stress-Induced Cell Loss and Functional Deficiency of the Protein aSelected illustrative examples of diseases are shown in which protein misfolding is thought to be the major mechanism of functional derangement or cell or tissue injury.CFTR, Cystic fibrosis transmembrane conductance regulator; LDL, low-density lipoprotein.


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Pathology_Robbins. Exposure of cells to radiation or chemotherapeutic agents, intracellular generation of ROS, and acquisition of mutations may all induce DNA damage, which if severe may trigger apoptotic death. Damage to DNA is sensed by intracellular sentinel proteins, which transmit signals that lead to the accumulation of p53 protein. p53 first arrests the cell cycle (at the G1 phase) to allow the DNA to be repaired before it is replicated (Chapter 6). However, if the damage is too great to be repaired successfully, p53 triggers apoptosis, mainly by stimulating BH3-only sensor proteins that ultimately activate Bax and Bak, proapoptotic members of the Bcl-2 family. When p53 is mutated or absent (as it is in certain cancers), cells with damaged DNA that would otherwise undergo apoptosis survive. In such cells, the DNA damage may result in mutations or DNA rearrangements (e.g., translocations) that lead to neoplastic transformation (Chapter 6).


Pathology_Robbins

Pathology_Robbins. A common cause of injury to cells and tissues is the inflammatory reaction that is elicited by pathogens, necrotic cells, and dysregulated immune responses, as in autoimmune diseases and allergies. In all these situations, inflammatory cells, including neutrophils, macrophages, lymphocytes, and other leukocytes, secrete products that evolved to destroy microbes but also may damage host tissues. These injurious immune reactions are classified under hypersensitivity. Their mechanisms and significance are discussed in Chapter 5. In the previous discussion, we addressed the mechanisms of cell injury according to the initiating cause, and highlighted the principal pathways of injury that are triggered in different pathophysiologic situations. Some abnormalities characterize cell injury regardless of the cause, and are thus seen in a variety of pathologic situations. Two of these changes are described next.