Pathology_Robbins

Pathology_Robbins. Fetal hydrops refers to the accumulation of edema fluid in the fetus during intrauterine growth. The causes of fetal hydrops are manifold; the most important are listed in Table 7.8 . In the past, hemolytic anemia caused by Rh blood group incompatibility between mother and fetus (immune hydrops) was the most common cause, but with the successful prophylaxis of this disorder during pregnancy, other causes of nonimmune hydrops have emerged as the principal culprits. The fluid accumulation can be quite variable, ranging in degree from progressive, generalized edema of the fetus (hydrops fetalis), a usually lethal condition, to more localized and less marked edematous processes, such as isolated pleural and peritoneal effusions or postnuchal fluid collections (cystic hygroma), that often are compatible with life ( Fig. 7.27 ). The mechanism of immune hydrops is discussed first, followed by other important causes of fetal hydrops. Table 7.8 Major Causes of Fetal Hydrops


Pathology_Robbins

Pathology_Robbins. Fig. 7.27 ). The mechanism of immune hydrops is discussed first, followed by other important causes of fetal hydrops. Table 7.8 Major Causes of Fetal Hydrops TurnersyndromeTrisomy21,trisomy18 *Thecauseoffetalhydropsmaybeundetermined(“idiopathic”)inupto20%ofcases.DatafromMachinGA:Hydrops,cystichygroma,hydrothorax,pericardialeffusions,andfetalascites.InGilbert-BarnessE,etal,editors:Potter’s pathology of the fetus, infant, and child, St.Louis,2007,Mosby,pp33. Fig. 7.27 Hydropsfetalis.(A)Generalizedaccumulationoffluidinthefetus.(B)Fluidaccumulationparticularlyprominentinthesofttissuesoftheneck.Thisconditionhasbeentermedcystic hygroma. Cystichygromasarecharacteristicallyseenwith,butnotlimitedto,constitutionalchromosomalanomaliessuchas45,Xkaryotypes.(Courtesy of Dr. Beverly Rogers,Department of Pathology,University of Texas Southwestern Medical Center,Dallas,Texas.) http://ebooksmedicine.net


Pathology_Robbins

Pathology_Robbins. Immune hydrops results from an antibody-induced hemolytic disease in the newborn that is caused by blood group incompatibility between mother and fetus. Such an incompatibility occurs when the fetus inherits red cell antigenic determinants from the father that are foreign to the mother. The most common antigens to result in clinically significant hemolysis are the Rh and ABO blood group antigens. Of the numerous antigens included in the Rh system, only the D antigen is a major cause of Rh incompatibility. Fetal red cells may reach the maternal circulation during the last trimester of pregnancy, when the cytotrophoblast is no longer present as a barrier, or during childbirth itself (fetomaternal bleed). The mother then becomes sensitized to the foreign antigen and produces antibodies that, in future pregnancies, can freely traverse the placenta to the fetus, in which they cause red cell destruction. With initiation of immune hemolysis, progressive anemia in the fetus leads to tissue


Pathology_Robbins

Pathology_Robbins. in future pregnancies, can freely traverse the placenta to the fetus, in which they cause red cell destruction. With initiation of immune hemolysis, progressive anemia in the fetus leads to tissue ischemia, intrauterine cardiac failure, and peripheral pooling of fluid (edema). As discussed later, cardiac failure may be the final pathway by which edema occurs in many cases of nonimmune hydrops as well.


Pathology_Robbins

Pathology_Robbins. Several factors influence the immune response to Rh-positive fetal red cells that reach the maternal circulation: Concurrent ABO incompatibility protects the mother against Rh immunization, because the fetal red cells are promptly coated by isohemagglutinins (preformed anti-A or anti-B antibodies) and removed from the maternal circulation. The antibody response depends on the dose of immunizing antigen, so hemolytic disease develops only when the mother has experienced a significant transplacental bleed (more than 1 mL of Rh-positive red cells). The isotype of the antibody is important, because immunoglobulin G (IgG) (but not IgM) antibodies can cross the placenta. The initial exposure to Rh antigen evokes the formation of IgM antibodies, so Rh disease is very uncommon with the first pregnancy. Subsequent exposure during the second or third pregnancy generally leads to a brisk IgG antibody response.


Pathology_Robbins

Pathology_Robbins. Appreciation of the role of previous sensitization in the pathogenesis of Rh-hemolytic disease of the newborn has led to its therapeutic control. Currently, Rh-negative mothers are given Rh immune globulin (RhIg) at 28 weeks and within 72 hours after delivery of an Rh-positive baby. The RhIg masks the antigenic sites on the fetal red cells that may have leaked into the maternal circulation during childbirth, thus preventing long-lasting sensitization to Rh antigens.


Pathology_Robbins

Pathology_Robbins. As a result of the remarkable success achieved in prevention of Rh hemolysis, fetomaternal ABO incompatibility currently is the most common cause of immune hemolytic disease of the newborn. Although ABO incompatibility occurs in approximately 20% to 25% of pregnancies, hemolysis develops in only a small fraction of infants born subsequently, and in general the disease is much milder than Rh incompatibility. In part the less severe course of disease can be attributed to the expression of A and B antigens on many cells other than red cells which act like sponge for the transferred antibody. ABO hemolytic disease occurs almost exclusively in infants of blood group A or B who are born to mothers of blood group O. The normal anti-A and anti-B isohemagglutinins in group O mothers usually are of the IgM type and therefore do not cross the placenta. However, for reasons not well understood, certain group O women possess IgG antibodies directed against group A or B antigens (or both) even


Pathology_Robbins

Pathology_Robbins. of the IgM type and therefore do not cross the placenta. However, for reasons not well understood, certain group O women possess IgG antibodies directed against group A or B antigens (or both) even without previous sensitization. Therefore, the firstborn may be affected. There is no effective method of preventing hemolytic disease resulting from ABO incompatibility.


Pathology_Robbins

Pathology_Robbins. The major causes of nonimmune hydrops include those disorders associated with cardiovascular defects, chromosomal anomalies, and fetal anemia. Structural cardiovascular defects and functional abnormalities (i.e., arrhythmias) may result in intrauterine cardiac failure and hydrops. Among the chromosomal anomalies, 45,X karyotype (Turner syndrome) and trisomies 21 and 18 are associated with fetal hydrops; the basis for this disorder usually is the presence of underlying structural cardiac anomalies, although in Turner syndrome there may be an abnormality of lymphatic drainage from the neck leading to postnuchal fluid accumulation (resulting in cystic hygromas). Fetal anemias resulting from causes other than Rh or ABO incompatibility also may result in hydrops. In fact, in some parts of the world (e.g., Southeast Asia), severe fetal anemia caused by homozygous α-thalassemia probably is the most common cause of fetal hydrops.


Pathology_Robbins

Pathology_Robbins. Transplacental infection by parvovirus B19 is increasingly recognized as an important cause of fetal hydrops. The virus gains entry into erythroid precursors (normoblasts), where it replicates. The ensuing cellular injury leads to the death of the normoblasts and aplastic anemia. Parvoviral intranuclear inclusions can be seen within circulating and marrow erythroid precursors ( Fig. 7.28 Fig. 7.28 BonemarrowfromaninfantinfectedwithparvovirusB19.Thearrows pointtotwoerythroidprecursorswithlargehomogeneousintra-nuclearinclusionsandasurroundingperipheralrimofresidualchromatin. http://ebooksmedicine.net The basis for hydrops in fetal anemia of immune and nonimmune causes is tissue ischemia with secondary myocardial dysfunction and circulatory failure. Additionally, secondary liver failure may occur, with loss of synthetic function contributing to hypoalbuminemia, reduced plasma osmotic pressure, and edema.


Pathology_Robbins


Pathology_Robbins

Pathology_Robbins. Early recognition of fetal hydrops is imperative, because even severe cases can sometimes be salvaged with timely therapy. Immune hydrops that results from Rh incompatibility can be predicted with reasonable certainty, because its severity correlates well with rapidly rising Rh antibody titers in the mother during pregnancy. Antenatal identification and management of the at-risk fetus have been facilitated by amniocentesis and the advent of chorionic villus and fetal blood sampling. The direct anti-globulin test (direct Coombs test) (Chapter 12) using fetal cord blood yields a positive result if the red cells have been coated by maternal antibody. In addition, cloning of the RHD gene has resulted in efforts to determine fetal Rh status by sequencing cell free DNA in maternal blood. When identified, cases of severe intrauterine hemolysis may be treated by fetal intravascular transfusions via the umbilical cord and early delivery. Postnatally, phototherapy is helpful, because visible


Pathology_Robbins

Pathology_Robbins. identified, cases of severe intrauterine hemolysis may be treated by fetal intravascular transfusions via the umbilical cord and early delivery. Postnatally, phototherapy is helpful, because visible light converts bilirubin to readily excreted dipyrroles. As already discussed, in an overwhelming majority of cases, administration of RhIg to the mother can prevent the occurrence of immune hydrops in subsequent pregnancies. Group ABO hemolytic disease is more difficult to predict but is readily anticipated by awareness of the blood incompatibility between mother and father and by hemoglobin and bilirubin determinations in the vulnerable newborn. In fatal cases of fetal hydrops, a thorough postmortem examination is imperative to determine the cause and to exclude a potentially recurring cause such as a chromosomal abnormality.


Pathology_Robbins

Pathology_Robbins. Themostcommoncausesoffetalhydropsarenonimmune(chromosomalabnormalities,cardiovasculardefects,andfetalanemia),whereasimmunehydropshasbecomelessfrequentasaresultofRhantibodyprophylaxis. Erythroblastosisfetalis(circulatingimmatureerythroidprecursors)isacharacteristicfindingoffetalanemia-associatedhydrops. Hemolysis-inducedhyperbilirubinemiacanresultinbilirubintoxicity(kernicterus)inthebasalgangliaandbrainstem,particularlyinprematureinfants. Malignant neoplasms constitute the second most common cause of death in children between the ages of 4 and 14 years; only accidents exact a higher toll. Benign tumors are even more common than cancers. It is sometimes difficult to segregate, on morphologic grounds, true tumors from tumorlike lesions in the infant and child. In this context, two special categories of tumor-like lesions should be recognized:


Pathology_Robbins

Pathology_Robbins. Heterotopia or choristoma refers to microscopically normal cells or tissues that are present in abnormal locations. Examples are a pancreatic tissue “rest” found in the wall of the stomach or small intestine and a small mass of adrenal cells found in the kidney, lungs, ovaries, or elsewhere. Heterotopic rests usually are of little clinical significance, but on the basis of their appearance they can be confused with neoplasms.


Pathology_Robbins

Pathology_Robbins. Hamartoma refers to an excessive but focal overgrowth of cells and tissues native to the organ in which it occurs. Although the cellular elements are mature and identical to those found in the remainder of the organ, they do not reproduce the normal architecture of the surrounding tissue. The line of demarcation between a hamartoma and a benign neoplasm frequently is tenuous and is variously interpreted. Hemangiomas, lymphangiomas, rhabdomyomas of the heart, and adenomas of the liver are considered by some researchers to be hamartomas and by others to be true neoplasms. Virtually any neoplasm may be encountered in the pediatric age group, but three—hemangiomas, lymphangiomas, and teratomas—deserve special mention here.


Pathology_Robbins

Pathology_Robbins. Virtually any neoplasm may be encountered in the pediatric age group, but three—hemangiomas, lymphangiomas, and teratomas—deserve special mention here. Hemangiomas are the most common neoplasms of infancy. Both cavernous and capillary hemangiomas may be encountered (Chapter 10), although the latter often are more cellular than in adults and thus may appear deceptively worrisome. In children, most hemangiomas are located in the skin, particularly on the face and scalp, where they produce flat to elevated, irregular, red-blue masses; the flat, larger lesions are referred to as port-wine stains. Hemangiomas may enlarge as the child ages, but in many instances they spontaneously regress (


Pathology_Robbins

Pathology_Robbins. Fig. 7.31 ). The vast majority of superficial hemangiomas have no more than a cosmetic significance; rarely, they may be the manifestation of a hereditary disorder associated with disease within internal organs, such as the von Hippel-Lindau syndrome (Chapter 10). A subset of CNS cavernous hemangiomas can occur in the familial setting; affected families harbor mutations in one of three cerebral cavernous malformation (CCM) genes. Lymphangiomas represent the lymphatic counterpart of hemangiomas. Microscopic examination shows cystic and cavernous spaces lined by endothelial cells and surrounded by lymphoid aggregates; the spaces usually contain pale fluid. They may occur on the skin but, more importantly, they also are encountered in the deeper regions of the neck, axilla, mediastinum, and retroperitoneum. Although histologically benign, they tend to increase in size after birth and may encroach on mediastinal structures or nerve trunks in axilla.


Pathology_Robbins

Pathology_Robbins. Teratomas may occur as benign, well-differentiated cystic lesions (mature teratomas), as lesions of indeterminate potential (immature teratomas), or as unequivocally Fig. 7.31 Congenitalcapillaryhemangioma(A)atbirthand(B)at2yearsofageafterthelesionhadundergonespontaneousregression.(Courtesy of Dr. Eduardo Yunis, Children’s Hospital of Pittsburgh, Pittsburgh, Pennsylvania.) http://ebooksmedicine.net malignant teratomas (usually admixed with another germ cell tumor component such as an endodermal sinus tumor). Sacrococcygeal teratomas are the most common teratomas of childhood, accounting for 40% or more of cases (


Pathology_Robbins

Pathology_Robbins. Fig. 7.32 ). In view of the overlap in the mechanisms underlying congenital malformations and oncogenesis, it is interesting that approximately 10% of sacrococcygeal teratomas are associated with congenital anomalies, primarily defects of the hindgut and cloacal region and other midline defects (e.g., meningocele, spina bifida) not believed to result from local effects of the tumor. Approximately 75% of these tumors are mature teratomas with a benign course, and about 12% are unmistakably malignant and lethal (Chapter 18). The remainder are designated immature teratomas, and their malignant potential correlates with the amount of immature tissue elements present. Most of the benign teratomas are encountered in younger infants (4 months of age or younger), whereas children with malignant lesions tend to be somewhat older. The organ systems involved most commonly by malignant neoplasms in infancy and childhood are the hematopoietic system, neural tissue, and soft tissues (


Pathology_Robbins

Pathology_Robbins. The organ systems involved most commonly by malignant neoplasms in infancy and childhood are the hematopoietic system, neural tissue, and soft tissues ( Table 7.9 ). This distribution is in sharp contrast with that in adults, in whom epithelial tumors of the lung, heart, prostate, and colon are the most common forms. Malignant neoplasms of infancy and childhood differ biologically and histologically from those in adults. The main differences are as follows: Relatively frequent demonstration of a close relationship between abnormal development (teratogenesis) and tumor induction (oncogenesis), suggesting a common stem cell defect. Prevalence of genetic abnormalities or familial syndromes that predispose to cancer Tendency of fetal and neonatal malignancies to regress spontaneously or to undergo “differentiation” into mature elements Fig. 7.32 Sacrococcygealteratoma.Notethesizeofthelesioncomparedwiththatoftheinfant.


Pathology_Robbins

Pathology_Robbins. Fig. 7.32 Sacrococcygealteratoma.Notethesizeofthelesioncomparedwiththatoftheinfant. Table 7.9 Common Malignant Neoplasms of Infancy and Childhood 10–14 Years of 0–4 Years of Age 5–9 Years of Age Age • Better survival or cure of many childhood tumors, so that much attention is now paid to minimizing the adverse delayed effects of chemotherapy and radiotherapy in survivors, including the development of second malignancies


Pathology_Robbins

Pathology_Robbins. Many malignant pediatric neoplasms are histologically unique. In general, they tend to exhibit a primitive (embryonal) rather than pleomorphic-anaplastic microscopic appearance, and frequently they exhibit features of organogenesis specific to the site of tumor origin. Because of their primitive histologic appearance, many childhood tumors have been collectively referred to as small, round, blue-cell tumors. Sheets of cells with small, round nuclei characterize these tumors, which include neuroblastoma, lymphoma, rhabdomyosarcoma, Ewing sarcoma (peripheral neuroectodermal tumor), and some cases of Wilms tumor. Sufficient distinctive features usually are present to permit definitive diagnosis on the basis of histologic examination alone, but molecular studies are becoming increasingly useful, both for diagnosis and prognosis of childhood cancers. Three common tumors—neuroblastoma, retinoblastoma, and Wilms tumor—are described here to highlight the differences between pediatric tumors


Pathology_Robbins

Pathology_Robbins. both for diagnosis and prognosis of childhood cancers. Three common tumors—neuroblastoma, retinoblastoma, and Wilms tumor—are described here to highlight the differences between pediatric tumors and those in adults.


Pathology_Robbins

Pathology_Robbins. The term neuroblastic includes tumors of the sympathetic ganglia and adrenal medulla that are derived from primordial neural crest cells populating these sites; neuroblastoma is the most important member of this family.


Pathology_Robbins

Pathology_Robbins. It is the second most common solid malignancy of childhood after brain tumors, accounting for 7% to 10% of all pediatric neoplasms, and as many as 50% of malignancies diagnosed in infancy. Neuroblastomas demonstrate several unique features in their natural history, including spontaneous regression and spontaneous or therapy-induced maturation. Most occur sporadically, but 1% to 2% are familial, with autosomal dominant transmission, and in such cases the neoplasms may involve both of the adrenals or multiple primary autonomic sites. Germline mutations in the anaplastic lymphoma kinase (ALK) gene have been linked to the familial predisposition to neuroblastoma. Somatic gain-of-function ALK mutations are also observed in 8% to 10% of sporadic neuroblastomas, and are markers of adverse prognosis. Clinical trials using inhibitors that target the mutated ALK tyrosine kinase are underway. http://ebooksmedicine.net


Pathology_Robbins

Pathology_Robbins. Someneoplasmsshowsignsofmaturation,eitherspontaneousortherapy-induced.Largercellshavingmoreabundantcytoplasmwithlargevesicularnucleiandaprominentnucleolus,representingganglion cells invariousstagesofmaturation,maybefoundintumorsadmixedwithprimitiveneuroblasts(ganglioneuroblastoma). Lesionsthatareevenbetterdifferentiatedcontainmanymorelargecellsresemblingmatureganglioncellsintheabsenceofresidualneuroblasts;suchneoplasmsmeritthedesignationganglioneuroma ( Fig.7.33B ).MaturationofneuroblastsintoganglioncellsusuallyisaccompaniedbytheappearanceofSchwanncells. Many factors influence prognosis, but the most important are the stage of the tumor and the age of the patient. • Staging of neuroblastomas ( Table 7.10 ) assumes great importance in establishing a prognosis. Special note should be taken of stage 4S (S means special), because the outlook for these patients is excellent, despite the spread of disease. As noted in


Pathology_Robbins

Pathology_Robbins. Table 7.10 , the primary tumor would be classified as stage 1 or 2 but for the presence of metastases, which are limited to liver, skin, and bone marrow, without bone involvement. Infants with 4S tumors have an excellent prognosis with minimal therapy, and it is not uncommon for the primary or metastatic tumors to undergo spontaneous regression. The biologic basis for this welcome behavior is not clear. Unfortunately, most (60% to 80%) children Fig. 7.33 (A)Neuroblastoma.Thistumoriscomposedofsmallcellsembeddedinafinelyfibrillarmatrix(neuropil).AHomer-Wrightpseudorosette(tumorcellsarrangedconcentricallyaroundacentralcoreofneuropil)isseenintheupperrightcorner.(B)Ganglioneuromas,arisingfromspontaneousortherapy-inducedmaturationofneuroblastomas,arecharacterizedbyclustersoflargeganglioncellswithvesicularnucleiandabundanteosinophiliccytoplasm(arrow).Spindle-shapedSchwanncellsarepresentinthebackgroundstroma.


Pathology_Robbins

Pathology_Robbins. present with stage 3 or 4 tumors, and only 20% to 40% present with stage 1, 2A, 2B, or 4S neuroblastomas. Age is the other important determinant of outcome. The outlook for children younger than 18 months is much more favorable than for older children at a comparable stage of disease. Most neoplasms diagnosed in children during the first 18 months of life are stage 1 or 2, or stage 4S (“low” risk category in Table 7.10), whereas neoplasms in older children fall into the “intermediate” or “high” category of risk. Histology is an independent prognostic variable in neuroblastic tumors; evidence of schwannian stroma and gangliocytic differentiation is indicative of a favorable prognosis.


Pathology_Robbins

Pathology_Robbins. Histology is an independent prognostic variable in neuroblastic tumors; evidence of schwannian stroma and gangliocytic differentiation is indicative of a favorable prognosis. Amplification of the MYCN oncogene has profound impact on prognosis. MYCN is located on the distal short arm of chromosome 2. MYCN amplification is present in about 25% to 30% of primary tumors, most in advanced-stage disease; the greater the number of copies, the worse the prognosis. Amplification of MYCN does not karyotypically manifest at the resident 2p23p24 site, but rather as extrachromosomal double minute chromosomes or homogeneously staining regions on http://ebooksmedicine.net Stage4Anyprimarytumorwithdisseminationtodistantlymphnodes,bone,bonemarrow,liver,skin,and/orother0.9 organs(exceptasdefinedforstage4S)0.8 Stage4SLocalizedprimarytumor(asdefinedforstage1,2A,or 2B)withdisseminationlimitedtoskin,liver,and/orbonemarrow(<10%ofnucleatedcellsareconstitutedbyneoplasticcells;>10%involvementofbonemarrowis


Pathology_Robbins

Pathology_Robbins. Stage4SLocalizedprimarytumor(asdefinedforstage1,2A,or 2B)withdisseminationlimitedtoskin,liver,and/orbonemarrow(<10%ofnucleatedcellsareconstitutedbyneoplasticcells;>10%involvementofbonemarrowis S, Special.AdaptedfromBrodeurGM,PritchardJ,BertholdF,etal:Revisionsofthe internationalneuroblastomadiagnosis,staging,andresponsetotreatment,J Clin Oncol 11:1466,1993. 0.7 0.6 0.5 0.4 0.3 0.2 0.1 Fig. 7.34 ). MYCN amplification is currently the most important genetic abnormality used in risk stratification of neuroblastic tumors and automatically renders a tumor as “high” risk, irrespective of stage or age.


Pathology_Robbins

Pathology_Robbins. DNA ploidy is another prognostic factor, with tumors that are hyperdiploid (with whole chromosome gains) having more favorable prognosis than tumors that are near-diploid. The latter subset, although closer to diploidy on absolute number, tends to have multiple structural rearrangements between, and within, chromosomes that can result in adverse molecular events, such as MYCN amplification.


Pathology_Robbins

Pathology_Robbins. Although age, stage, MYCN status, and ploidy are used clinically for assigning prognostication, many other “experimental” molecular aberrations have been identified that might also have prognostic bearings, or be potential candidates for targeted therapy. For example, expression of TrkA, a high-affinity receptor for nerve growth factor that is indicative of differentiation toward sympathetic ganglia lineage, is associated with favorable prognosis. Overall, besides MYCN amplification and ALK mutations (latter in ~10%), de novo neuroblastomas have few recurrent “hotspot” mutations. However, a very high frequency of relapsed neuroblastomas (>75%) have mutations in the RAS-MAP kinase signaling pathway, suggesting that relapsed tumors might be targeted with therapies against these oncogenic pathways. Children younger than 2 years with neuroblastomas generally present with a protuberant abdomen resulting


Pathology_Robbins

Pathology_Robbins. Fig. 7.34 (A)FISHusingafluorescein-labeledcosmidprobeforN-myc onatissuesectioncontainingneuroblastomaattachedtothekidney.Notetheneuroblastomacellsontheupperhalfofthephotowithlargeareasofstaining(yellow-green);thiscorrespondstoamplifiedN-MYC intheformofhomogeneouslystainingregions.Renaltubularepithelialcellsinthelowerhalfofthephotographshownonuclearstainingandbackground(green) cytoplasmicstaining.(B)AKaplan-Meiersurvivalcurveofinfantsyoungerthan1yearofagewithmetastaticneuroblastoma.The3-yearevent-freesurvivalofinfantswhosetumorslackedMYCN amplificationwas93%,whereasthosewithtumorsthathadMYCN amplificationhadonlya10%event-freesurvival.(A, Courtesy Dr. Timothy Triche, Children’s Hospital, Los Angeles, California. B, Reproduced with permission from Brodeur GM: Neuroblastoma: biological insights into a clinical enigma, NatRevCancer3:203–216, 2003.) from an abdominal mass, fever, and weight loss. In older children the neuroblastomas may remain unnoticed until metastases cause hepatomegaly,


Pathology_Robbins

Pathology_Robbins. into a clinical enigma, NatRevCancer3:203–216, 2003.) from an abdominal mass, fever, and weight loss. In older children the neuroblastomas may remain unnoticed until metastases cause hepatomegaly, ascites, and bone pain. Neuroblastomas may metastasize widely through the hematogenous and lymphatic systems, particularly to liver, lungs, bones, and the bone marrow. In neonates, disseminated neuroblastomas may manifest with multiple cutaneous metastases associated with deep blue discoloration to the skin (earning the rather unfortunate moniker of “blueberry muffin baby”). About 90% of neuroblastomas, regardless of location, produce catecholamines (similar to the catecholamines associated with pheochromocytomas), which constitutes an important diagnostic feature (i.e., elevated blood levels of catecholamines and elevated urine levels of catecholamine metabolites such as vanillylmandelic acid [VMA] and homovanillic acid [HVA]). Despite the elaboration of catecholamines, hypertension is much


Pathology_Robbins

Pathology_Robbins. catecholamines and elevated urine levels of catecholamine metabolites such as vanillylmandelic acid [VMA] and homovanillic acid [HVA]). Despite the elaboration of catecholamines, hypertension is much less frequent with these neoplasms than with pheochromocytomas (Chapter 20).


Pathology_Robbins


Pathology_Robbins

Pathology_Robbins. Neuroblastomassecretecatecholamines,whosemetabolites(VMA/HVA)canbeusedforscreeningpatients. Retinoblastoma is the most common primary intraocular malignancy of children. The molecular genetics of retinoblastoma has been discussed previously (Chapter 6). Approximately 40% of the tumors are associated with a germline mutation in the RB gene and are therefore heritable. The remaining 60% of the tumors develop sporadically, and these have somatic RB gene mutations. Familial cases typically are associated with development of multiple tumors that are bilateral, although they may be unifocal and unilateral. All of the sporadic, nonheritable tumors are unilateral and unifocal. Patients with familial retinoblastoma also are at increased risk for the development of osteosarcoma and other soft tissue tumors. MORPHOLOGYRetinoblastomastendtobenodularmasses,usuallyintheposteriorretina,oftenwithsatelliteseedings(


Pathology_Robbins

Pathology_Robbins. Tumorcellsmaydisseminatebeyondtheeyethroughtheopticnerveorsubarachnoidspace.ThemostcommonsitesofdistantmetastasesaretheCNS,skull,distalbones,andlymphnodes. The presenting findings include poor vision, strabismus, a whitish hue to the pupil (“cat’s eye reflex”), and pain and tenderness in the eye. The median age at presentation is 2 years, although the tumor may be present at birth. Untreated, the tumors usually are fatal, but when treated with chemotherapy, radiotherapy, and (in locally advanced tumors) enucleation, survival is the rule. As noted earlier, some tumors spontaneously regress, and patients with familial retinoblastoma are at increased risk for the development of osteosarcoma and other soft tissue tumors.


Pathology_Robbins

Pathology_Robbins. Wilms tumor, or nephroblastoma, is the most common primary tumor of the kidney in children, with most cases occurring in children between 2 and 5 years of age. This tumor illustrates several important concepts of childhood tumors: the relationship between congenital malformation and increased risk of tumors; the histologic similarity between tumor and developing organ; and finally, the remarkable success in the treatment of childhood tumors. Each of these concepts is presented in the following discussion.


Pathology_Robbins

Pathology_Robbins. Three groups of congenital malformations are associated with an increased risk for Wilms tumor. These are WAGR syndrome (i.e., Wilms tumor, aniridia, genital abnormalities, and mental retardation); Denys-Drash syndrome (DDS), and Beckwith-Wiedemann syndrome (BWS). Of patients with WAGR syndrome approximately one in three will go on to develop this tumor. Another group of patients, those with so-called “Denys-Drash syndrome” (DDS), have an even higher risk (approximately 90%) of Wilms tumor. Patients with the WAGR syndrome, as the name indicates, have Wilms tumor, aniridia, genital abnormalities and mental retardation. DD syndrome is characterized by gonadal dysgenesis and early onset nephropathy leading to renal failure. Both of these conditions are associated with abnormalities of the Wilms tumor 1 (WT1) gene, located on 11p13. The nature of the genetic aberration differs, however: Patients with WAGR syndrome demonstrate loss of genetic material (i.e., deletions) of WT1, while


Pathology_Robbins

Pathology_Robbins. the Wilms tumor 1 (WT1) gene, located on 11p13. The nature of the genetic aberration differs, however: Patients with WAGR syndrome demonstrate loss of genetic material (i.e., deletions) of WT1, while persons with DDS harbor a dominant negative inactivating mutation in WT1 that interferes with the function of normal WT1 protein made from the other WT1 allele. WT1 is critical to normal renal and gonadal development; it is not surprising, therefore, that constitutional inactivation of one copy of this gene results in genitourinary abnormalities in humans.


Pathology_Robbins

Pathology_Robbins. A third group of patients, those with Beckwith-Wiedemann syndrome (BWS), also are at increased risk for the development of Wilms tumor. These patients exhibit enlargement of individual body organs (e.g., tongue, kidneys, or liver) or entire body segments (hemihypertrophy); enlargement of adrenal cortical cells (adrenal cytomegaly) is a characteristic microscopic feature. BWS is an example of a disorder of genomic imprinting (see earlier). The genetic locus that is involved in these patients is in band p15.5 of chromosome 11 distal to the WT1 locus. Although this locus is called “WT2” for the second Wilms tumor locus, the gene involved has not been identified. This region contains at least 10 genes that normally are expressed from only one of the two parental alleles, with transcriptional silencing of the other parental homologue by methylation of the promoter region located upstream of the transcription start site. Of all candidate “WT2” genes, imprinting abnormalities of insulinlike


Pathology_Robbins

Pathology_Robbins. silencing of the other parental homologue by methylation of the promoter region located upstream of the transcription start site. Of all candidate “WT2” genes, imprinting abnormalities of insulinlike growth factor-2 (IGF2) have the strongest relationship to tumor predisposition in persons with BWS. IGF2 normally is expressed solely from the paternal allele, whereas the maternal allele is imprinted. In some Wilms tumors, loss of imprinting (i.e., reexpression of IGF2 by the maternal allele) can be demonstrated, leading to overexpression of the IGF2 protein, which is postulated to result in both organ enlargement and tumorigenesis. Thus, these associations suggest that in some cases, congenital malformations and tumors represent related manifestations of genetic lesions affecting a single gene or closely linked genes. In addition to Wilms tumors, patients with BWS also are at increased risk for the development of hepatoblastoma, adrenocortical tumors, rhabdomyosarcomas, and pancreatic


Pathology_Robbins

Pathology_Robbins. gene or closely linked genes. In addition to Wilms tumors, patients with BWS also are at increased risk for the development of hepatoblastoma, adrenocortical tumors, rhabdomyosarcomas, and pancreatic tumors.


Pathology_Robbins

Pathology_Robbins. http://ebooksmedicine.net Fig. 7.35 Retinoblastoma.(A)Poorlycohesivetumorinretinaisseenabuttingtheopticnerve.(B)Higher-powerviewshowingFlexner-Wintersteinerrosettes(arrow) andnumerousmitoticfigures.


Pathology_Robbins

Pathology_Robbins. In contrast to syndromic Wilms tumors, the molecular abnormalities underlying sporadic (i.e., nonsyndromic) tumors, which account for 90% of cases overall in children, are only recently being elucidated. Some of them are associated with specific histologic features described later. For example, gain-of-function mutations of the gene encoding β-catenin (Chapter 6) have been demonstrated in approximately 10% of sporadic Wilms tumors. Other recurrent mutations occur in genes encoding proteins involved in microRNA processing (DROSHA, DGCR8, and DICER1); these are seen in 15% to 20% of Wilms tumors with predominantly blastemal histology (see later). It is postulated that aberrations in microRNA processing lead to reduced levels of many mature microRNAs, in particular in the miR-200 family, which is involved in “mesenchymal to epithelial transformation” during renal morphogenesis. The lack of mesenchymal to epithelial transformation likely leads to persistent blastemal “rests” in the kidney


Pathology_Robbins

Pathology_Robbins. is involved in “mesenchymal to epithelial transformation” during renal morphogenesis. The lack of mesenchymal to epithelial transformation likely leads to persistent blastemal “rests” in the kidney (see the following), which evolve into Wilms tumors. Finally, tumors with TP53 mutations are associated with an especially poor prognosis and often have a distinctive anaplastic histologic appearance, described later.


Pathology_Robbins

Pathology_Robbins. http://ebooksmedicine.net Fig. 7.36 Wilmstumorinthelowerpoleofthekidneywiththecharacteristictantograycolorandwell-circumscribedmargins. Patients typically present with a palpable abdominal mass, which may extend across the midline and down into the pelvis. Less often, the presenting features are fever and abdominal pain, hematuria or, occasionally, intestinal obstruction as a result of pressure from the tumor. The prognosis for Wilms tumor generally is very good, and excellent results are obtained with a combination of nephrectomy and chemotherapy. Anaplasia is a harbinger of adverse prognosis, but only if it is diffuse. If the anaplasia is focal and confined within the resected nephrectomy Wilmstumoristhemostcommonrenalneoplasmofchildhood. PatientswiththreesyndromesareatincreasedriskforWilmstumors:Denys-Drash,Beckwith-Wiedemann,andWilmstumor,aniridia,genitalabnormalities,andmentalretardation(WGAR)syndrome.


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Pathology_Robbins. PatientswiththreesyndromesareatincreasedriskforWilmstumors:Denys-Drash,Beckwith-Wiedemann,andWilmstumor,aniridia,genitalabnormalities,andmentalretardation(WGAR)syndrome. WAGRsyndromeandDDSareassociatedwithWT1inactivation,whereasBeckwith-WiedemannarisesthroughimprintingabnormalitiesattheWT2locus,principallyinvolvingtheIGF2 gene. ThemorphologiccomponentsofWilmstumorincludeblastema(small,roundbluecells)andepithelialandstromalelements. NephrogenicrestsareprecursorlesionsofWilmstumors. specimen, the outcome is no different from that for tumors without evidence of anaplasia.


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Pathology_Robbins. NephrogenicrestsareprecursorlesionsofWilmstumors. specimen, the outcome is no different from that for tumors without evidence of anaplasia. In the era predating the ready availability of molecular diagnostic assays, rendering the diagnosis of a genetic disorder depended on the identification of abnormal gene products (e.g., mutant hemoglobin or abnormal metabolites) or their clinical effects, such as mental retardation (e.g., in PKU). Several factors have since enabled the rapid expansion of molecular diagnostics from the realm of research to an almost ubiquitous presence in both academic and commercial pathology laboratories. These include (1) the sequencing of the human genome and the deposition of these data in publicly available databases; (2) the availability of numerous “off-the-shelf” polymerase chain


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Pathology_Robbins. http://ebooksmedicine.net reaction (PCR) kits tailor-made for the identification of specific genetic disorders; (3) the availability of high-resolution microarrays (“gene chips”) that can interrogate both DNA and RNA on a genomewide scale using a single platform; and, finally, (4) the emergence of automated, high-throughput, next-generation (“NextGen”) sequencing technologies. The last two advances have been especially useful in the context of new research to elucidate the genetic basis for both mendelian and complex disorders. Although a detailed discussion of molecular diagnostics is beyond the scope of this book, some of the better-known approaches are highlighted in the ensuing paragraphs. Irrespective of the technique used, the genetic aberration being queried can be either in the germline (i.e., present in each and every cell of the affected person, as with a CFTR mutation in a patient with CF) or somatic (i.e., restricted to specific tissue types or lesions, as with MYCN


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Pathology_Robbins. the germline (i.e., present in each and every cell of the affected person, as with a CFTR mutation in a patient with CF) or somatic (i.e., restricted to specific tissue types or lesions, as with MYCN amplification in neuroblastoma cells). This consideration determines the nature of the sample (e.g., peripheral blood lymphocytes [PBLs], saliva, tumor tissue) used for the assay.


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Pathology_Robbins. In general, indications for genetic analysis can be divided into inherited conditions and acquired conditions. Within inherited conditions, genetic testing can be offered at either the prenatal or postnatal stages. It may involve conventional cytogenetics, FISH, molecular diagnostics, or a combination of these techniques. Prenatal genetic analysis should be offered to all patients who are at risk of having cytogenetically abnormal progeny. It can be performed on cells obtained by amniocentesis, on chorionic villus biopsy material, or increasingly in “liquid biopsies” on maternal blood paired with next-generation sequencing. Some important indications are the following: Advanced maternal age (beyond 34 years), which is associated with greater risk of trisomies Confirmed carrier status for a balanced reciprocal trans-location, robertsonian translocation, or inversion (in such cases, the gametes may be unbalanced, so the progeny would be at risk for chromosomal disorders)


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Pathology_Robbins. Fetal abnormalities observed on ultrasound, or an abnormal result on routine maternal blood screening Determination of fetal sex when the patient or partner is a confirmed carrier of an X-linked genetic disorder Postnatal genetic analysis usually is performed on peripheral blood lymphocytes. Indications are as follows: Suspicion of a metabolic syndrome Suspected aneuploidy (e.g., features of Down syndrome) or other syndromic chromosomal abnormality (e.g., deletions, inversions) Suspected monogenic disease, whether previously described or unknown


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Pathology_Robbins. Acquired genetic alterations, such as somatic mutations in cancer, are increasingly becoming a large focus area in molecular diagnostics laboratories, especially with the advent of targeted therapies. Although single gene tests (mutations of EGFR or BRAF, amplification of HER2) have been used for a while to inform treatment decisions, the advent of cost-effective next-generation sequencing approaches has now allowed interrogations of large numbers of coding genes (often in the 100s), as well as cancer-relevant translocations, in a single assay. The clinical team typically receives a “genomic report” on the patient’s cancer, including potential molecularly targeted treatment recommendations. Another major focus of molecular diagnostics has been the rapid identification of infectious diseases, such as suspected tuberculosis or virulent pathogens such as Ebola, using DNA-based approaches. In general, these approaches have cut down the time required for diagnosis from weeks to a matter of


Pathology_Robbins

Pathology_Robbins. such as suspected tuberculosis or virulent pathogens such as Ebola, using DNA-based approaches. In general, these approaches have cut down the time required for diagnosis from weeks to a matter of days. Besides de novo identification of pathogens, molecular diagnostics laboratories can also contribute to the identification of treatment resistance (e.g., acquired mutations in influenza viruses that render them resistant to anti-virals), and to the monitoring of treatment efficacy using assays for “viral load” in the blood. Similar parameters (measuring efficacy of therapy and emergence of resistance) are also widely used in cancer patients.


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Pathology_Robbins. Because of the rapid advances in molecular diagnostics, terms such as “personalized therapy” and “precision medicine” are being increasingly used to indicate therapy tailored to the needs of the individual patient. It is hoped that such expectations will be met in the foreseeable future. We will now discuss some of the common approaches used in molecular diagnostics laboratories for assessment of inherited and acquired indications.


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Pathology_Robbins. As already discussed in this chapter, various diseases may occur as a result of copy number abnormalities, at the level of the entire chromosome (trisomy 21), chromosomal segments (22q11 deletion syndrome), or submicroscopic intragenic deletions (WAGR syndrome). Karyotype analysis of chromosomes by G banding remains the classic approach for identifying changes at the chromosomal level; however, as might be expected, the resolution with this technique is fairly low. To identify subchromosomal alterations, both focused analysis of chromosomal regions by FISH and global genomic approaches such as comparative genomic hybridization (CGH) are commonly used.


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Pathology_Robbins. FISH uses DNA probes that recognize sequences specific to chromosomal regions of greater than 100 kilobases in size, which defines the limit of resolution with this technique for identifying chromosomal changes. Such probes are labeled with fluorescent dyes and are applied to meta-phase spreads or interphase nuclei. The probe hybridizes to its complementary sequence on the chromosome and thus labels the specific chromosomal region that can then be visualized under a fluorescence microscope. The ability of FISH to circumvent the need for dividing cells is invaluable when a rapid diagnosis is warranted (e.g., in a critically ill infant suspected of having an underlying genetic disorder). Such analysis can be performed on prenatal samples (e.g., cells obtained by amniocentesis, chorionic villus biopsy, or umbilical cord blood), peripheral blood lymphocytes, and even archival tissue sections. FISH is used for the detection of numeric abnormalities of chromosomes (aneuploidy) (


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Pathology_Robbins. Fig. 7.38A ), subtle microdeletions ( Fig. 7.38B ) and complex translocations not indentifiable by routine karyotyping, and gene amplification (e.g., MYCN amplification in neuroblastomas). http://ebooksmedicine.net FISH requires previous knowledge of the one or few specific chromosomal regions suspected of being altered in the test sample. However, genomic abnormalities can also be detected without previous knowledge by using micro-array technology to perform a global genomic survey. First-generation platforms were designed for comparative genomic hybridization (CGH), whereas newer platforms incorporate single nucleotide polymorphism (SNP) genotyping approaches, offering multiple benefits.


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Pathology_Robbins. In array comparative genomic hybridization (aCGH), the test DNA and a reference (normal) DNA are labeled with two different fluorescent dyes that fluoresce red and green, respectively. The differentially labeled samples are then cohybridized to an array spotted with DNA probes that span the human genome at regularly spaced intervals, and usually cover all 22 autosomes and the sex chromosomes. At each chromosomal probe location, the binding of the labeled DNA from the two samples is compared. If the two samples are equal (i.e., the test sample is diploid), then all spots on the array will fluoresce yellow (the result of an equal admixture of green and red dyes). In contrast, if there is even a focal deletion or duplication, the probe spots corresponding to it will show skewing toward red or green (depending on gain or loss of material), allowing highly accurate determinations of copy number variants across the genome.


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Pathology_Robbins. Newer types of “SNP chips” are based on a similar concept, but the probes are designed to identify SNP sites genomewide, which has a number of advantages over CGH. As discussed in Chapter 1, SNPs are the most common type of DNA polymorphism, occurring approximately every 1000 nucleotides throughout the genome (e.g., in exons, introns, and regulatory sequences). Thus, SNPs serve both as a physical landmark within the genome as well as a genetic marker whose transmission can be followed from parent to child. There are several testing platforms that allow SNPs to be analyzed genomewide on arrays. Like CGH probes, methods involving SNPs can be used to determine copy number variations. In addition, by discriminating between SNP alleles at each particular location, they also provide zygosity data (


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Pathology_Robbins. Fig. 7.39 ). This means, for example, that SNP chips can identify uniparental disomy, in which loss of heterozygosity is seen within an affected region having diploid DNA content. The current generation of SNP arrays is quite comprehensive, with the Fig. 7.38 FISH.(A)Interphasenucleusfromamalepatientwithsuspectedtrisomy18.Threedifferentfluorescentprobeshavebeenusedina“FISHcocktail”;thegreenprobehybridizestotheXchromosomecentromere(one copy),theredprobetotheYchromosomecentromere(one copy),andtheaquaprobetothechromosome18centromere(three copies).(B)Ametaphasespreadinwhichtwofluorescentprobeshavebeenused,onehybridizingtochromosomeregion22q13(green) andtheotherhybridizingtochromosomeregion22q11.2(red).Therearetwo22q13signals.Oneofthetwochromosomesdoesnotstainwiththeprobefor22q11.2,indicatingamicrodeletioninthisregion.Thisabnormalitygivesrisetothe22q11.2deletionsyndrome(DiGeorgesyndrome).


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Pathology_Robbins. (Courtesy of Dr. Nancy R.Schneider and Jeff Doolittle,Cytogenetics Laboratory,University of Texas Southwestern Medical Center,Dallas,Texas.) http://ebooksmedicine.net 2.00 1.00 0.00 number –1.00 –2.00 1.0 AA 0.8 p13.31 p12.3 p12.1


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Pathology_Robbins. Fig. 7.39 AnalysisofcopynumbervariationviaSNPcytogenomicarray.GenomicDNAislabeledandhybridizedtoanarraycontainingpotentiallymillionsofprobespots.Copynumberisdeterminedbyoverallintensityandgenotypeisdeterminedbyallelicratio.Theexampleshownistheparmofchromosome12inapediatricleukemia.Here,thenormalareas(green) showneutral(diploid)DNAcontentandthezygosityplotshowstheexpectedratioofAA,AB,andBBSNPgenotypes.Theanomalousarea(red) showsdecreasedoverallintensity,andthezygosityplotshowslossofheterozygosity(theABgenotype);together,thesefindingsindicatethepresenceofadeletionononecopyofchromosome12p.(Modified from Paulsson K, et al: Genetic landscape of high hyper-diploid childhood acute lymphoblastic leukemia, PNAS107[50]:21719–21724, 2010.) largest containing greater than 4 million SNP probes. As a of one of the terminator nucleotides. After size separaresult, this technology is the mainstay of genomewide tion by capillary electrophoresis, the exact sequence can association studies (GWASs,


Pathology_Robbins

Pathology_Robbins. As a of one of the terminator nucleotides. After size separaresult, this technology is the mainstay of genomewide tion by capillary electrophoresis, the exact sequence can association studies (GWASs, described later). be “read” and compared with the normal sequence to detect the presence of mutations. Many applications of


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Pathology_Robbins. Sanger sequencing (and other PCR-based approaches) DirectDetectionofDNAMutationsbyare starting to give way to next-generation sequencing (discussed later), particularly when the analysis of large genes or multiple genes is required. Nonetheless, even PCR analysis, which involves exponential amplification of in 2016, Sanger sequencing remains the “gold standard” DNA, is now widely used in molecular diagnosis. If RNA for many genotyping assays. is used as the substrate, it is first reverse-transcribed to • Another approach for identifying mutations at a specific obtain cDNA and then amplified by PCR. This method nucleotide position (say, a codon 12 mutation in the involving reverse transcription (RT) often is abbreviated as KRAS oncogene that converts glycine [GGT] to aspartic RT-PCR. To amplify a DNA segment of interest, two acid [GAT]) would be to add fluorescently labeled primers that bind to the 3′ and 5′ ends of the normal nucleotides C and T to the PCR mixture, which are sequence


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Pathology_Robbins. a DNA segment of interest, two acid [GAT]) would be to add fluorescently labeled primers that bind to the 3′ and 5′ ends of the normal nucleotides C and T to the PCR mixture, which are sequence are designed. By using appropriate DNA poly-complementary to either the wild-type (G) or mutant merases and thermal cycling, the target DNA is greatly (A) sequence, respectively. Because these two nucleoamplified, producing millions of copies of the DNA tides are labeled with different fluorophores, the fluosequence between the two primer sites. The DNA sequence rescence emitted by the resulting PCR product can be of of the PCR product can then be analyzed in several ways: one or another color, depending on whether a C or a T • Sanger sequencing, named after its inventor, Nobel-becomes incorporated in the process of primer extenlaureate Frederick Sanger, has for many years been the sion (


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Pathology_Robbins. Fig. 7.40 ). The advantage of this allele-specific “workhorse” of genome sequencing, including the orig-extension strategy is that it is more sensitive than Sanger inal Human Genome Project. Here, the amplified DNA sequencing, allowing for the identification mutated is mixed with a DNA polymerase, a DNA primer, nucle-DNA sequences even if they make up a small fraction otides, and four dead-end (di-deoxy terminator) nucleo-(~1%) of the total DNA (as may be the case in clinical tides (A, T, G, and C) labeled with different fluorescent specimens obtained from patients suspected of harbor-tags. The ensuing reaction produces a series of DNA ing a malignancy). An ultrasensitive variant of this molecules of all possible lengths up to a kilobase or so, assay, known as droplet digital PCR, literally separates each labeled with a tag that corresponds to the base at the template DNA (or cDNA) into thousands to millions which the reaction stopped because of the incorporation of microscopic oil


Pathology_Robbins

Pathology_Robbins. separates each labeled with a tag that corresponds to the base at the template DNA (or cDNA) into thousands to millions which the reaction stopped because of the incorporation of microscopic oil “droplets,” such that any given http://ebooksmedicine.net


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Pathology_Robbins. Fig. 7.40 Allele-specificpolymerasechainreaction(PCR)analysisformutationdetectioninaheterogeneoussamplecontaininganadmixtureofnormalandmutantDNA.Nucleotidescomplementarytothemutantandwild-typenucleotidesatthequeriedbasepositionarelabeledwithdifferentfluorophoressuchthatincorporationintotheresultingPCRproductyieldsfluorescentsignalsofvariableintensityinaccordancewiththeratioofmutanttowild-typeDNApresent. droplet contains no more than one strand of template DNA, along with requisite PCR reagents. Each such “minireactor” then generates its own PCR product, which can be digitally integrated and analyzed for the presence of mutant alleles. By assessing each template individually in parallel, rather than in a single PCR reaction, droplet digital PCR has exquisite sensitivity, being able to detect as little as 1 mutant DNA strand in 10,000 or more wild-type molecules.


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Pathology_Robbins. Increasingly, nearly all of molecular diagnostics applications requiring assessment of DNA (or RNA) are being supplanted by so-called “next-generation sequencing” (NGS) technologies, so named because the Sanger sequencing mentioned earlier is now considered “first generation.” The availability of NGS technology has the potential to alter molecular diagnostics radically, by the sheer volume of sequencing data (more than 1 gigabase pairs or 1,000,000,000 base pairs of DNA per day!) at relatively low costs. The entire human genome has a little more than 3 gigabases, so true “whole genome sequencing” can be performed several times in a matter of days. In contrast with Sanger sequencing, NGS technologies use platforms in which sequencing of multiple fragments of the human genome (DNA or cDNA) can occur in parallel (“massively parallel sequencing”), significantly enhancing its speed (


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Pathology_Robbins. Fig. 7.41 ). Fluorescently labeled nucleotides are incorporated that are complementary to the template DNA strands, which are immobilized on a solid phase, with one nucleotide added per template per cycle. An image is captured at the end of each cycle, and the cycles are repeated until a “read” of sufficient length is generated to map each read back to the human genome using sophisticated bioinformatics. In addition to differences of time and scale, another key difference between NGS and traditional Sanger sequencing is the ability of the former to detect variant alleles that occur at low frequency in samples that contain a heterogeneous mixture of cells. With Sanger sequencing, the lower limit of detection for variant allele frequency is 20% or more, thus requiring either homogeneous samples (such as germline DNA) or some form of enrichment, such as microdissection of tumor cells from a biopsy that also contains normal tissue. The ability of NGS to “read” the same region of the


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Pathology_Robbins. samples (such as germline DNA) or some form of enrichment, such as microdissection of tumor cells from a biopsy that also contains normal tissue. The ability of NGS to “read” the same region of the genome multiple times (from 100to 1000-fold redundancy) allows detection of allele frequencies as low as 1%, making NGS particularly useful when searching for mutations in tissue biopsies that contain more nonneoplastic stroma than the tumor cells.


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Pathology_Robbins. Clinical and research applications of NGS (which can be performed on either germline or somatic DNA, or both) include targeted sequencing (typically a panel of a few to several hundred genes); whole exome sequencing (WES), which interrogates all coding regions of the human genome; and whole genome sequencing (WGS), which includes the remaining 99% of the human genome, including regions that express noncoding RNAs. In addition to DNA, NGS can also be used for measuring the transcriptome (RNA-Seq) and genomewide binding sites for transcription factors or histones (chromatin immunoprecipitation and sequencing, or ChIP-Seq).


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Pathology_Robbins. Direct diagnosis of mutations is possible only if the gene responsible for a genetic disorder is known and its sequence has been identified. In several diseases that have a genetic basis, including some common disorders, direct genetic diagnosis is not possible, either because the causal gene has not been identified or because the disease is multifactorial (polygenic) and no single gene is involved. In such cases, two types of analyses can be performed for unbiased identification of disease-associated gene(s): linkage analysis and GWASs. In both instances, surrogate markers in the genome, also known as marker loci, must be used to localize the chromosomal regions of interest, based on their linkage to one or more putative disease-causing genes. The marker loci used are naturally occurring variations in DNA sequences known as polymorphisms, typically SNPs, as described previously.


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Pathology_Robbins. Two technologic breakthroughs have enabled the application of SNPs to high-throughput “gene hunting.” First is the completion of several large genomic sequencing projects (the HapMap project and the 1000 genome project) that have provided linkage disequilibrium patterns in multiple ethnoracial groups, based on genomewide SNP mapping. The entire human genome can now be divided into blocks known as “haplotypes,” which contain varying numbers of contiguous SNPs on the same chromosome that are in linkage disequilibrium and hence inherited together as a cluster. As a result, rather than querying every single SNP in the human genome, comparable information about shared DNA can be obtained simply by http://ebooksmedicine.net A AC C C G T GG A B T A AC C GG T Add all four nucleotides, labeled with different dyes C G T Wash, four-color imaging C G T C G T C G T G A C G A C Cleave dye and terminating Repeat C A T G groups, wash Top: CATCGT Bottom: CCCCCC


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Fig. 7.41

Pathology_Robbins. Fig. 7.41


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Pathology_Robbins. with permission from Metzker M: Sequencing technologies—the next generation, NatRevGenet11:31–46, 2010, © Nature Publishing Group.) looking for shared haplotypes, using single or a small number of SNPs that “tag” or identify a specific haplotype. Second, it is now possible to genotype millions of SNPs simultaneously, in a cost-effective way, using high-density SNP arrays, as described earlier.


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Pathology_Robbins. Linkage analysis deals with assessing shared marker loci (i.e., SNPs) in family members exhibiting the disease or trait of interest, with the assumption that SNPs in linkage disequilibrium with the disease allele are transmitted through pedigrees. With time it becomes possible to define a “disease haplotype” based on a panel of SNPs, all of which cosegregate with the putative disease allele. Eventually, linkage analysis facilitates localization and cloning of the disease allele. Linkage analysis is most useful in mendelian disorders that are related to one gene with profound effects and high penetrance.


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Pathology_Robbins. It is now established that some of the most common human diseases, such as hypertension, diabetes, mental disorders, and asthma, have a polygenic basis, with multiple genetic loci contributing small independent effects, resulting in a disease phenotype. Conventional linkage analyses lack the statistical power to detect such genetic variants. In GWASs, large cohorts of patients with and without a disease (rather than families) are examined across the entire genome for variant SNPs that are overrepresented in persons with the disease. This identifies regions of the genome that contain a variant gene or genes that confer disease susceptibility. The causal variant within the region is then provisionally identified using a “candidate gene” approach, in which genes are selected based on how tightly they are associated with the disease and whether their biologic function seems likely to be involved in the disease under study. In addition to polygenic diseases, GWASs also have led to the


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Pathology_Robbins. tightly they are associated with the disease and whether their biologic function seems likely to be involved in the disease under study. In addition to polygenic diseases, GWASs also have led to the identification of genetic loci that modulate common quantitative traits in humans, such as height, body mass, hair and eye color, and bone density.


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Pathology_Robbins. We end this chapter to reflect on the current genomic era of medicine. Advances in understanding the molecular basis of human disease are taking place at a breathtaking pace, as are molecular diagnostic techniques. Throughout this book we will illustrate how these changes have impacted on the management of certain diseases in the clinic. By the time this book reaches the reader we hope that many more such examples will become commonplace. Bianchi DW, Parker RL, Wentworth J, et al: DNA sequencing versus standard prenatal aneuploidy screening, N Engl J Med 370:799–808, 2014. [Seminal study that demonstrated potential superiority of maternal “liquid biopsies” paired with next-generation sequencing for prenatal diagnosis of trisomies in the fetus, when compared to conventional prenatal tests.] http://ebooksmedicine.net


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Pathology_Robbins. Bresler SC, Weiser DA, Huwe PJ, et al: ALK mutations confer differential oncogenic activation and sensitivity to ALK inhibition therapy in neuroblastoma, Cancer Cell 26(5):682–694, 2014. [An important study demonstrating the potential for targeted therapy in neuroblastomas harboring ALK mutations, using small molecule ALK inhibitors.] Cutting GR: Cystic fibrosis genetics: from molecular understanding to clinical application, Nat Rev Genet 16:45–56, 2015. [An outstanding review on cystic fibrosis by one of the doyens in the field, with a particular focus on genetic and environmental modifiers of the disease.] Eleveld TF, Oldridge DA, Bernard V, et al: Relapsed neuroblastomas show frequent RAS-MAPK pathway mutations, Nat Genet 47:864– 871, 2015. [A landmark study demonstrating a high frequency of alterations in the RAS-MAPK pathway in treated neuroblastomas, providing a new opportunity for targeted therapy in this lethal tumor of childhood.]


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Pathology_Robbins. Gudbjartsson DF, Helgason H, Gudjonsson SA, et al: Large-scale whole-genome sequencing of the Icelandic population, Nat Genet 47:435–444, 2015. [A unique study that reiterates the power of modern-day next-generation sequencing, wherein the whole genomes of over 2600 Icelandic natives were sequenced for identification of genetic variants that might influence genetic diversity as well as risk for multifactorial diseases.] Hagerman PJ, Hagerman RJ: Fragile X-associated tremor/ataxia syndrome (FTAS), Ann N Y Acad Sci 1338:58–70, 2015. [An authoritative review on FMR1 protein, and the role of premutations at the fragile X loci in causing the symptomatic neurodegenerative condition, FTAS.] Jameson JL, Longo DL: Precision medicine—personalized, problematic, and promising, N Engl J Med 372:2229–2234, 2015. [An insightful review on the challenges and opportunities of personalized medicine, also known as precision medicine, in the modern era of genomics and high throughput sequencing.]


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Pathology_Robbins. Kalsner L, Chamberlain SJ: Prader-Willi, Angelman, and 15q11-q13 duplication syndromes, Pediatr Clin North Am 62:587–606, 2015. [An up-to-date review on the genetics, clinical findings, and natural history of children with these two imprinting disorders.] Page MM, Stefanutti C, Sniderman A, et al: Recent advances in the understanding and care of familial hypercholesterolemia: significance of the biology and therapeutic regulation of proprotein convertase subtilisin/kexin type 9, Clin Sci 129:63–79, 2015. [A state-of-the-art review on mechanisms of cholesterol regulation by PCSK9, and how inhibition of this protein is being leveraged in the clinic for augmenting statin effects.] Robinson JG, Farnier M, Krempf M, et al: Efficacy and safety of alirocumab in reducing lipids and cardiovascular events, N Engl J Med 372(16):1489–1499, 2015. [Seminal study establishing the benefits of PCSK9 inhibitors in hyperlipidemia.]


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Pathology_Robbins. Stoltz DA, Meyerholz DK, Welsh MJ: Origins of cystic fibrosis lung disease, N Engl J Med 372:351–362, 2015. [An exhaustive review on pathophysiological mechanisms that contribute to the panorama of pulmonary diseases in cystic fibrosis, with numerous excellent illustrations.] Trachtenberg FL, Haas EA, Kinney HC, et al: Risk factor changes for sudden infant death syndrome after initiation of Back-to-Sleep campaign, Pediatrics 129:630–638, 2012. [An important study that measures the impact of the “Back to Sleep” campaign on SIDS mortality, and what additional risk factors have emerged since then.]


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Pathology_Robbins. Wainwright CE, TRAFFIC Study Group, TRANSPORT Study Group: Lumacaftor-Ivacaftor in patients with cystic fibrosis homozygous for Phe508del CFTR, N Engl J Med 373:220–231, 2015. [Seminal study that led to FDA approval of the combination regimen of Lumacaftor and Ivacaftor, which enhance mutant Phe508del CFTR function in cystic fibrosis; Ivacaftor alone is approved for another variant of CFTR.] Walz AL, Ooms A, Gadd S, et al: Recurrent DGCR8, DROSHA, and SIX homeodomain mutations in favorable histology Wilms tumors, Cancer Cell 27(2):286–297, 2015. [Seminal sequencing study in Wilms tumors identifying mutations of genes involved in microRNA processing and consequent microRNA deregulation.]


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Pathology_Robbins. Wong LC, Behr ER: Sudden unexplained death in infants and children: the role of undiagnosed inherited cardiac conditions, Europace 16:1706–1713, 2014. [An insightful review on how unsuspected cardiac conditions, specifically “channelopathies,” leading to sudden death in infancy might be erroneously diagnosed as SIDS.] Yang Y, Muzny DM, Xia F, et al: Molecular findings among patients referred for clinical whole exome sequencing, JAMA 312:1870–1879, 2014. [An elegant study highlighting the prowess of next-generation techniques, wherein 2000 consecutive patients with suspected genetic disorders underwent germline whole exome sequencing, leading to the discovery of causal pathogenic mutations in as many as 25% of patients.] http://ebooksmedicine.net studentconsult.com Metals as Environmental Pollutants Injury by Therapeutic Drugs:Adverse Drug Reactions


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Pathology_Robbins. Metals as Environmental Pollutants Injury by Therapeutic Drugs:Adverse Drug Reactions Many diseases are caused or influenced by environmental factors. Broadly defined, the term ambient environment encompasses the various outdoor, indoor, and occupational settings in which humans live and work. In each of these settings, the air people breathe, the food and water they consume, and the toxic agents they are exposed to are major determinants of health. Other environmental factors pertain to the individual (“personal environment”) and include tobacco use, alcohol ingestion, therapeutic and “recreational” drug consumption, diet, and the like. It is generally believed that factors in the personal environment have a larger effect on human health than that of the ambient environment, but new threats related to global warming (described later) may change this equation.


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Pathology_Robbins. The term environmental disease refers to disorders caused by exposure to chemical or physical agents in the ambient, workplace, and personal environments, including diseases of nutritional origin. Environmental diseases are surprisingly common. The International Labor Organization has estimated that work-related injuries and illnesses kill more people per year globally than do road accidents and wars combined. Most of these work-related problems are caused by illnesses rather than accidents. The burden of disease in the general population created by nonoccupational exposures to toxic agents is much more difficult to estimate, mostly because of the diversity of agents and the difficulties in measuring the dose and duration of exposures. Whatever the precise numbers, environmental diseases are major causes of disability and suffering and constitute a heavy financial burden, particularly in developing countries.


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Pathology_Robbins. Environmental diseases are sometimes the consequence of major disasters, such as the methyl mercury contamination of Minamata Bay in Japan in the 1960s, the leakage of methyl isocyanate gas in Bhopal, India, in 1984, the Chernobyl nuclear accident in 1986, the Fukushima nuclear meltdown following the tsunami in 2011, and lead poisoning resulting from contaminated drinking water in the city of Flint in the United States in 2016. Fortunately, these are unusual and infrequent occurrences. Less dramatic, but much more common, are diseases and injury produced by chronic exposure to relatively low levels of contaminants. It should be noted that a host of factors, including complex interactions between pollutants producing multiplicative effects, as well as the age, genetic predisposition, and different tissue sensitivities of exposed persons, create wide variations in individual sensitivity. Disease related to malnutrition is even more pervasive. In 2010, it was estimated that 925 million


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Pathology_Robbins. different tissue sensitivities of exposed persons, create wide variations in individual sensitivity. Disease related to malnutrition is even more pervasive. In 2010, it was estimated that 925 million people were malnourished—one in every seven persons worldwide. Children are disproportionately affected by undernutrition, which accounts for more than 50% of childhood mortality worldwide.


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Pathology_Robbins. In this chapter, we first consider the emerging problem of the health effects of climate change. We then discuss the mechanisms of toxicity of chemical and physical agents, and address specific environmental disorders, including those of nutritional origin. Global temperature measurements show that the earth has warmed significantly since the early 20th century, and especially since the mid-1960s. Record-breaking global temperatures have become common, with 2005, 2010, 2014, and 2015 each setting successive high-temperature records. Of note, 15 of the 16 warmest years since 1880 have occurred during the 21st century. During 2015, the global land temperature was 0.9° C warmer than the 20th century average. Mean global ocean temperatures also set new records in 2015, with an annual average temperature 0.74° C above the 20th century average. As of this writing, 2016 is on track to approach or exceed the records just set in 2015. http://ebooksmedicine.net


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Pathology_Robbins. http://ebooksmedicine.net The rising atmospheric and ocean temperatures have led to a large number of effects that include changes in storm frequency, drought, and flood, as well as large-scale ice losses in Greenland, Antarctica, and the vast majority of the other glaciated regions on earth, as well as dramatic thinning or disappearance of Arctic ocean sea ice. The melting of land-based glacial ice and the thermal expansion of the warming oceans has produced approximately 80 mm of global average sea level rise since 1993, and the sea level currently is rising at a global average rate of 3.5 ± 0.4 mm/year. Although politicians quibble, among scientists there is a general acceptance that climate change is, at least in part, man-made. The culprit is the rising atmospheric level of greenhouse gases, particularly carbon dioxide (CO2) released through the burning of fossil fuels (


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Pathology_Robbins. Fig. 8.1A ), as well as ozone (an important air pollutant, discussed later), and methane. These gases, along with water vapor, produce the so-called “greenhouse effect” by absorbing energy radiated from Earth’s surface that otherwise would be lost into space. The annual average level of atmospheric CO2 (about 401 ppm) in 2015 was higher than at any point in approximately 650,000 years and, without changes in human behavior, is expected to increase to 500 to 1200 ppm by the end of this century—levels not experienced for tens of millions of years. This increase stems not only from increased CO2 production but also from deforestation and the attendant decrease in carbon fixation by plants. Depending on the computer model used, increased levels of greenhouse gases are projected to cause the global temperature to rise by 2°C to 5°C by the year 2100 (


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Pathology_Robbins. Fig. 8.1B ). The health consequences of climate change will depend on its extent and rapidity, the severity of the ensuing consequences, and humankind’s ability to mitigate the damaging effects. Even in the best-case scenario, however, climate change is expected to have a serious negative impact on human health by increasing the incidence of a number of diseases, including the following: Cardiovascular, cerebrovascular, and respiratory diseases, all of which will be exacerbated by heat waves and air pollution. Gastroenteritis, cholera, and other foodand waterborne infectious diseases, caused by contamination as a consequence of floods and disruption of clean water supplies and sewage treatment, after heavy rains and other environmental disasters.


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Pathology_Robbins. Vector-borne infectious diseases, such as malaria and dengue fever, resulting from changes in vector number and geographic distribution related to increased temperatures, crop failures, and more extreme weather variation (e.g., more frequent and severe El Niño events). Malnutrition, caused by changes in local climate that disrupt crop production. Such changes are anticipated to be most severe in tropical locations, in which average 1958-2015 Correlation Coefficient: 0.90947 –0.4 –0.2 0 0.2 0.4 0.6 0.8 A Global temperature anomaly (° C) Fig. 8.1 Climatechange,pastandfuture.(A)CorrelationofCO2levelsmeasuredattheMaunaLoaObservatoryinHawaiiwithaverageglobaltemperaturetrendssincethemid-1960s.“Globaltemperature”inanygivenyearwasdeducedattheHadleyCenter(UnitedKingdom)frommeasurementstakenatmorethan3000weatherstationslocatedaroundtheglobe.


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Pathology_Robbins. (B)Predictedtemperatureincreasesduringthe21stcentury.Differentcomputermodelsplotanticipatedrisesinglobaltemperaturesof2°Cto5°Cbytheyear2100.(A, Courtesy of Dr. Richard Aster, Department of Geosciences, Colorado State University, Fort Collins, Colorado.) temperatures may already be near or above crop tolerance levels; it is estimated that by 2080, agricultural productivity may decline by 10% to 25% in some developing countries as a consequence of climate change.


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Pathology_Robbins. Beyond these disease-specific effects, it is estimated that the melting of glacial ice, particularly in Greenland and other parts of the Northern Hemisphere, combined with the thermal expansion of warming oceans, will raise sea levels by 2 to 6 feet by 2100. Approximately 10% of the world’s population—roughly 600 million people—live in low-lying areas that are at risk for flooding even if the rise in ocean levels is at the low end of these estimates. For example, a rise in sea level by 1.5 feet will submerge 70% of the land mass of Maldive islands by 2100 and a 3-foot rise will inundate 100% all of the islands by 2085. The resulting displacement of people will disrupt lives and commerce, creating conditions ripe for political http://ebooksmedicine.net unrest, war, and poverty, the “vectors” of malnutrition, sickness, and death. Worldwide recognition of the catastrophic effects of climate change led in late 2015 to a historic meeting of 196 countries in Paris, France, at which the


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Pathology_Robbins. of malnutrition, sickness, and death. Worldwide recognition of the catastrophic effects of climate change led in late 2015 to a historic meeting of 196 countries in Paris, France, at which the participating countries agreed to the following objective:


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Pathology_Robbins. Holding the increase in the global average temperature to well below 2°C above preindustrial levels and to pursue efforts to limit the temperature increase to 1.5°C above preindustrial levels, recognizing that this would significantly reduce the risks and impacts of climate change. Toxicology is defined as the science of poisons. It studies the distribution, effects, and mechanisms of action of toxic agents. More broadly, it also includes the study of the effects of physical agents such as radiation and heat. Approximately 4 billion pounds of toxic chemicals, including 72 million pounds of known carcinogens, are produced each year in the United States. In general, however, little is known about the potential health effects of chemicals. Of the approximately 100,000 chemicals in use in the United States, less than 1% have been tested experimentally for health effects. We now consider some basic principles regarding the toxicity of exogenous chemicals and drugs.


Pathology_Robbins

Pathology_Robbins. We now consider some basic principles regarding the toxicity of exogenous chemicals and drugs. The definition of a poison is not straightforward. It is a quantitative concept strictly dependent on dosage. The quote from Paracelsus in the 16th century that “all substances are poisons; the right dosage differentiates a poison from a remedy” is perhaps even more valid today, in view of the proliferation of therapeutic drugs with potentially harmful effects. Xenobiotics are exogenous chemicals in the environment that may be absorbed by the body through inhalation, ingestion, or skin contact ( Fig. 8.2 Chemicals may be excreted in urine or feces or eliminated in expired air, or they may accumulate in bone, fat, brain, or other tissues.


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Pathology_Robbins. Fig. 8.2 Chemicals may be excreted in urine or feces or eliminated in expired air, or they may accumulate in bone, fat, brain, or other tissues. Chemicals may act at the site of entry, or they may be transported to other sites. Some agents are not modified on entry in the body, but most solvents and drugs are metabolized to form water-soluble products (detoxification) or are activated to form toxic metabolites. Most solvents and drugs are lipophilic, which facilitates their transport in the blood by lipoproteins and penetration through lipid components of cell membranes. The reactions that metabolize xenobiotics into nontoxic products, or activate xenobiotics to generate toxic compounds ( Fig. 8.3


Pathology_Robbins

Pathology_Robbins. The reactions that metabolize xenobiotics into nontoxic products, or activate xenobiotics to generate toxic compounds ( Fig. 8.3 Fig. 8.2), occur in two phases. In phase I reactions, chemicals can undergo hydrolysis, oxidation, or reduction. Products of phase I reactions often are metabolized into water-soluble compounds through phase II reactions of glucuronidation, sulfation, methylation, and conjugation with glutathione (GSH). Water-soluble compounds are readily excreted. Toxicity Distribution to tissues Storage


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Pathology_Robbins. Toxicity Distribution to tissues Storage Fig. 8.2 Human exposure to pollutants. Pollutants contained in air, water, and soil are absorbed through the lungs, gastrointestinal (GI) tract, and skin. In the body, they may act at the site of absorption, but they generally are transported through the bloodstream to various organs, where they are stored or metabolized. Metabolism of xenobiotics may result in the formation of water-soluble compounds, which are excreted, or in activation of the agent, creating a toxic metabolite.


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Pathology_Robbins. • The most important cellular enzyme system involved in phase I reactions is the cytochrome P-450 system. The P-450 system is present in organs throughout the body, but it is most active in the endoplasmic reticulum (ER) of the liver. The system catalyzes reactions that may either detoxify xenobiotics or convert xenobiotics into active compounds that cause cellular injury. Both types of reactions may produce, as a byproduct, reactive oxygen species (ROS), which can cause cellular damage (discussed in Chapter 2). Examples of metabolic activation of chemicals through the P-450 system are the conversion of carbon tetrachloride to the toxic trichloromethyl free radical, and the generation of a DNA-binding Xenobiotic Elimination Nontoxic metabolite Reactive metabolite Effects on cellular molecules (enzymes, receptors, membranes, DNA)


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Pathology_Robbins. Xenobiotic Elimination Nontoxic metabolite Reactive metabolite Effects on cellular molecules (enzymes, receptors, membranes, DNA) Fig. 8.3 Xenobioticmetabolism.Xenobioticscanbemetabolizedtonontoxicmetabolitesandeliminatedfromthebody(detoxification).However,theirmetabolismalsomayresultinformationofareactivemetabolitethatistoxictocellularcomponents.Ifrepairisnoteffective,short-andlong-termeffectsdevelop.(Modified from Hodgson E: Atextbookofmoderntoxicology,ed 3, Fig. 1–1. Hoboken, NJ, 2004, John Wiley & Sons.) metabolite from benzo[a]pyrene, a carcinogen present in cigarette smoke. The cytochrome P-450 system also participates in the metabolism of a large number of common therapeutic drugs such as acetaminophen, barbiturates, and anti-convulsants, and in alcohol metabolism (discussed later).


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Pathology_Robbins. • P-450 enzymes vary widely in activity among different people, owing to both polymorphisms in the genes encoding the enzymes and interactions with drugs that are metabolized through the system. The activity of the enzymes also may be decreased by fasting or starvation, and increased by alcohol consumption and smoking. Air pollution is a significant cause of morbidity and mortality worldwide, particularly among at-risk individuals with preexisting pulmonary or cardiac disease. The life-giving air that we breathe is also often laden with many potential causes of disease. Airborne microorganisms have long been major causes of morbidity and death. More widespread are the chemical and particulate pollutants found in the air, both in so-called “developed” and “underdeveloped” countries. Specific hazards have been recognized for both outdoor and indoor air.


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Pathology_Robbins. The ambient air in industrialized nations is contaminated with an unsavory mixture of gaseous and particulate pollutants, more so in cities and in proximity to heavy industry. In the United States, the Environmental Protection Agency (EPA) monitors and sets allowable upper limits for six pollutants: sulfur dioxide, CO, ozone, nitrogen dioxide, lead, and particulate matter. Together, some of these agents produce the well-known smog that sometimes stifles major cities such as Cairo, Los Angeles, Houston, Mexico City, and São Paulo. It may seem that air pollution is a modern phenomenon. This is not the case; Seneca wrote in AD 61 that he felt an alteration of his disposition as soon as he left the “pestilential vapors, soot, and heavy air of Rome.” The first environmental-control law was proclaimed by Edward I in 1306 and was straightforward in its simplicity: “Whoever should be found guilty of burning coal shall suffer the loss of his head.” What has changed in modern times is the


Pathology_Robbins

Pathology_Robbins. proclaimed by Edward I in 1306 and was straightforward in its simplicity: “Whoever should be found guilty of burning coal shall suffer the loss of his head.” What has changed in modern times is the nature and sources of air pollutants, and the types of regulations that control their emission. It could be argued that modern man has lost his head to drown himself in pollution!


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Pathology_Robbins. The lungs bear the brunt of the adverse consequences of air pollution, but air pollutants affect many organ systems (as with the effects of lead poisoning and CO, discussed later). More detailed discussion of pollutant-caused lung diseases is found in Chapter 13. Here we consider the major health effects of ozone, sulfur dioxide, particulates, and CO ( Table 8.1 Table 8.1 Health Effects of Outdoor Air Pollutants DatafromHealthEffectsofOutdoorAirPollution,Part2.CommitteeoftheEnvironmentalandOccupationalHealthAssemblyoftheAmericanThoracicSociety,Am J Respir Crit Care Med 153:477,1996. http://ebooksmedicine.net


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Pathology_Robbins. http://ebooksmedicine.net Ozone is one of the most pervasive air pollutants, with levels in many cities exceeding EPA standards. It is a gas formed by sunlight-driven reactions involving nitrogen oxides, which are released mostly by automobile exhaust. Together with oxides and fine particulate matter, ozone forms the familiar smog (from the words smoke and fog). Its toxicity stems from its participation in chemical reactions that generate free radicals, which injure the lining cells of the respiratory tract and the alveoli. Low levels of ozone may be tolerated by healthy persons but are detrimental to lung function, especially in those with asthma or emphysema, and when present along with particulate pollution. Unfortunately, pollutants rarely occur singly but combine to create a veritable “witches’ brew.”


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Pathology_Robbins. Sulfur dioxide, particles, and acid aerosols are emitted by coaland oil-fired power plants and industrial processes burning these fuels. Of these, particles appear to be the main cause of morbidity and death. Particles less than 10 µm in diameter are particularly harmful, because when inhaled they are carried by the airstream all the way to the alveoli. Here, they are phagocytosed by macrophages and neutrophils, causing the release of mediators (possibly by activating inflammasomes, Chapter 2) and inciting an inflammatory reaction. By contrast, larger particles are removed in the nose or are trapped by the mucociliary “escalator” and as a result are less dangerous.


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Pathology_Robbins. Carbon monoxide (CO) is a nonirritating, colorless, tasteless, odorless gas. It is produced by the incomplete oxidation of carbonaceous materials. Its sources include automotive engines, industries using fossil fuels, home oil burners, and cigarette smoke. The low levels often found in ambient air may contribute to impaired respiratory function but usually are not life threatening. However, persons working in confined environments with high exposure to fumes, such as tunnel and underground garage workers, may develop chronic poisoning. CO is included here as an air pollutant, but it also is an important cause of accidental and suicidal death. In a small, closed garage, exhaust from a running car engine can induce a lethal coma within 5 minutes. CO is a systemic asphyxiant that kills by binding to hemoglobin and preventing oxygen transport. Hemoglobin has a 200-fold greater affinity for CO than for O2. The carboxyhemoglobin, that is formed by binding of CO is incapable of carrying


Pathology_Robbins

Pathology_Robbins. binding to hemoglobin and preventing oxygen transport. Hemoglobin has a 200-fold greater affinity for CO than for O2. The carboxyhemoglobin, that is formed by binding of CO is incapable of carrying oxygen. Hypoxia leads to central nervous system (CNS) depression, which develops so insidiously that victims often are unaware of their plight and are unable to help themselves. Systemic hypoxia appears when the hemoglobin is 20% to 30% saturated with CO, and unconsciousness and death are probable with 60% to 70% saturation. The diagnosis of CO poisoning is based on detection of high levels of carboxyhemoglobin in the blood.


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Pathology_Robbins. Acute poisoning byCOgenerallyisaconsequenceofaccidentalexposureorsuicideattempt.Inlight-skinnedpeople,itismarkedbyacharacteristicgeneralized cherry-red color of the skin and mucous membranes, acolorimpartedbycarboxyhemoglobin.Ifdeathoccursrapidly,morphologicchangesmaynotbepresent;withlongersurvival,thebrainmaybeslightlyedematousandexhibitpunctatehemorrhagesandhypoxia-inducedneuronalchanges.Thesechangesarenotspecific;theysimplyimplysystemichypoxia(Chapter23).InvictimswhosurviveCOpoisoning,completerecoveryispossible;however,impairmentsofmemory,vision,hearing,andspeechsometimesremain. As modern homes are increasingly “buttoned up” to exclude the environment, the potential for pollution of the indoor air increases. The most common pollutant is tobacco smoke (discussed later), but additional offenders are CO, nitrogen dioxide (already mentioned as outdoor pollutants), and asbestos (discussed in Chapter 13). A few comments about some other agents are presented here.


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Pathology_Robbins. Smoke from burning of organic materials, containing various oxides of nitrogen and carbon particulates, is an irritant that predisposes exposed persons to lung infections and may contain carcinogenic polycyclic hydrocarbons. It is estimated that one-third of the world, mainly in developing areas, burn carbon-containing material such as wood, dung, or charcoal in their homes for cooking, heating, and light. Radon, a radioactive gas derived from uranium, is widely present in soil and in homes. Although radon exposure can cause lung cancer in uranium miners (particularly in those who smoke), it does not appear that low-level chronic exposures in the home increase lung cancer risk, at least for nonsmokers. Bioaerosols may contain pathogenic microbiologic agents, such as those that can cause Legionnaires’ disease, viral pneumonia, and the common cold, as well as allergens derived from pet dander, dust mites, and fungi and molds, which can cause rhinitis, eye irritation, and even asthma.


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Pathology_Robbins. Environmentaldiseasesareconditionscausedbyexposuretochemicalorphysicalagentsintheambient,workplace,andpersonalenvironments. Exogenouschemicals,knownasxenobiotics,enterthebodythroughinhalation,ingestion,andskincontact,andcaneitherbeeliminatedoraccumulateinfat,bone,brain,andothertissues. Xenobioticscanbeconvertedintonontoxicproductsortoxiccompoundsthroughatwo-phasereactionprocessthatinvolvesthecytochromeP-450system. http://ebooksmedicine.net Themostcommonairpollutantsareozone(whichincombinationwithoxidesandparticulatematterformssmog),sulfurdioxide,acidaerosols,andparticleslessthan10µmindiameter. COisanairpollutantandanimportantcauseofdeathfromaccidentsandsuicide;itbindshemoglobinwithhighaffinity,leadingtosystemicasphyxiationresultinginCNSdepression. Lead, mercury, arsenic, and cadmium, the heavy metals most commonly associated with harmful effects in human populations, are considered here.


Pathology_Robbins

Pathology_Robbins. Lead is a readily absorbed metal that binds to sulfhydryl groups in proteins and interferes with calcium metabolism, leading to hematologic, skeletal, neurologic, GI, and renal toxicities. Lead exposure occurs through contaminated air, food, and water. For most of the 20th century the major sources of lead in the environment were house paints and gasoline. Although the use of lead-based paints and leaded gas has greatly diminished, many sources of lead persist in the environment, such as mines, foundries, batteries, and spray paints, all of which constitute occupational hazards. However, flaking lead paint in older houses and soil contamination pose the major hazards for youngsters. Blood levels of lead in children living in older homes containing lead-based paint or lead-contaminated dust often exceed 5 µg/dL, the level at which the Centers for Disease Control and Prevention (CDC) recommends that measures be taken to limit further exposure. A dramatic case of lead contamination of


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Pathology_Robbins. often exceed 5 µg/dL, the level at which the Centers for Disease Control and Prevention (CDC) recommends that measures be taken to limit further exposure. A dramatic case of lead contamination of drinking water occurred in the U.S. city of Flint, Michigan, in 2014–2016. The so-called “Flint water crisis” occurred when the source of water supply to the city was changed from Lake Huron to the Flint River. Because water from the Flint River had a higher chloride concentration than the lake waters, it leached lead from century-old lead pipes. This caused an increase in lead levels in tap water above the acceptable limit of 15 parts per billion (ppb) in about 25% of the homes and in some cases as high as 13,200 ppb. As a result 6000 to 12,000 residents developed very high lead levels in their blood. Ingested lead is particularly harmful to children because they absorb more than 50% of lead from food, whereas adults absorb approximately 15%. A more permeable blood–brain barrier in children


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Pathology_Robbins. blood. Ingested lead is particularly harmful to children because they absorb more than 50% of lead from food, whereas adults absorb approximately 15%. A more permeable blood–brain barrier in children creates a high susceptibility to brain damage. The clinical features of lead poisoning are shown in


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Pathology_Robbins. Fig. 8.4 Most absorbed lead (80% to 85%) is taken up into developing teeth and into bone, where it competes with calcium, binds phosphates, and has a half-life of 20 to 30 years. About 5% to 10% of the absorbed lead remains in the blood, and the remainder is distributed throughout soft tissues. Excess lead is toxic to nervous tissues in adults and children; peripheral neuropathies predominate in adults, whereas central effects are more common in children. The effects of chronic lead exposure in children may be subtle, producing mild dysfunction, or they may be massive and lethal. In young children, sensory, motor, intellectual, and psychologic impairments have been described, including reduced IQ, learning disabilities, retarded psychomotor Fig. 8.4 Pathologicfeaturesofleadpoisoning. Adult: Headache, memory loss Child: Encephalopathy, mental deterioration BLOOD Anemia, red cell basophilic stippling PERIPHERAL NERVES Adult: Demyelination


Pathology_Robbins

Pathology_Robbins. Adult: Headache, memory loss Child: Encephalopathy, mental deterioration BLOOD Anemia, red cell basophilic stippling PERIPHERAL NERVES Adult: Demyelination BONES Child: Radiodense deposits in epiphyses development, and, in more severe cases, blindness, psychoses, seizures, and coma. Lead-induced peripheral neuropathies in adults generally remit with the elimination of exposure, but both peripheral and CNS abnormalities in children usually are irreversible. Other effects of lead exposure include the following. Excess lead interferes with the normal remodeling of calcified cartilage and primary bone trabeculae in the epiphyses in children, causing increased bone density detected as radiodense “lead lines” (


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Pathology_Robbins. Fig. 8.5 ). Lead lines of a different sort also may occur in the gums, where excess lead stimulates hyperpigmentation. Lead inhibits the healing of fractures by increasing chondrogenesis and delaying cartilage mineralization. Excretion of lead occurs by way of the kidneys, and acute exposures may cause damage to proximal tubules.


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Pathology_Robbins. Lead has a high affinity for sulfhydryl groups and interferes with two enzymes involved in heme synthesis: aminolevulinic acid dehydratase and delta ferrochelatase. Iron incorporation into heme is impaired, leading to anemia. Lead also inhibits sodium-and potassium-dependent ATPases in cell membranes, an effect that may increase the fragility of red cells, causing hemolysis. The diagnosis of lead poisoning requires constant vigilance. It may be suspected on the basis of neurologic changes in children or unexplained anemia with basophilic stippling in red cells in adults and children. Elevated blood lead and red cell free protoporphyrin levels (greater than 50 µg/dL) or, alternatively, zinc-protoporphyrin levels, are required for definitive diagnosis. In milder cases of lead exposure, anemia may be the only obvious abnormality. http://ebooksmedicine.net


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Pathology_Robbins. http://ebooksmedicine.net Fig. 8.5 Leadpoisoning.Impairedremodelingofcalcifiedcartilageintheepiphyses(arrows) ofthewristhascausedamarkedincreaseintheirradiodensity,sothattheyareasradiopaqueasthecorticalbone.(Courtesy of Dr. GW Dietz: Department of Radiology, University of Texas Southwestern Medical School, Dallas,Texas.) MORPHOLOGYThemajoranatomictargetsofleadtoxicityaretheblood,bonemarrow,nervoussystem,GItract,andkidneys( Fig.8.4 Blood changes areoneoftheearliestsignsofleadaccumulationandarecharacteristic,consistingofamicrocytic,hypo-chromicanemiaassociatedwithadistinctivepunctatebasophilic stippling ofredcells.Thesechangesinthebloodstemfromtheinhibitionofhemesynthesisinmarrowerythroidprogenitors.Anotherconsequenceofthisblockadeisthatzinc-protoporphyrinisformedinsteadofheme.Thus,elevatedbloodlevelsofzincprotoporphyrinoritsproduct,freeredcellprotoporphyrin,areimportantindicatorsofleadpoisoning.


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Pathology_Robbins. Fig.8.4 Mercury, like lead, binds to sulfhydryl groups in certain proteins with high affinity, leading to damage in the CNS and several other organs such as the GI tract and the kidneys. Humans have used mercury in many ways throughout history, including as a pigment in cave paintings, a cosmetic, a remedy for syphilis, and a component of diuretics. Poisoning from inhalation of mercury vapors has long been recognized and is associated with tremor, gingivitis, and bizarre behavior, such as that of the “Mad Hatter” in Lewis Carroll’s Alice in Wonderland (mercury formerly was used in hat-making).