Hepatitis B infection and the pathogenesis of hepatocellular carcinoma

June l990

SELECTED SUMMARIES

of the bypassed segment and is reversible with antibiotics (Gastroenterology 1982;82:535-5481 or bypass takedown, it is reasonable to assume that liver damage can be reversed if takedown is performed befora cirrhosis has developed. Furthermore, serum hepatic enzymes; and bilirubin levels are insensitive markers of ongoing hepatic damage. Therefore, reversal of the JI bypass should be performed before biochemical or clinical evidence of hepatic dysfunction occurs. One discussant of the present paper warned that asymptomatic patients after JI bypasses are “time bombs waiting to explode.” In my opinion, the skeletal abnormalities caused by JI bypass have received too little attention. Arthritis, presumably caused by immune complexes containing antigens from bacteria in the bypassed segment [South Med J 1987;80:768-7721, can be disabling. The present study shows that the arthritis resolves if treated early with bypass reversal. Not surprisingly, crippling arthritis did not improve substantially after bypass reversal; two patients had no relief. Another factor to consider is osteomalacia resulting from hypovitaminosis D. In one study, 75% of patients after JI bypass had osteomalacia as determined by bone biopsy (Ann Surg 1979;189:785790). Importantly, none of these patients had symptoms, and noninvasive diagnostic tests were normal. Therefore, jejunoileal bypass causes both arthritis and osteomalacia. If left untreated, the arthritis may become crippling and irreversable. Osteomalacia is common, assymptomatic and detectable only by bone biopsy. For all these reasons, I believe the presence of a JI bypass is indication for reversal. Should gastric restrictive surgery be added to the JI bypass reversal? For two reasons, the answer is “yes” for nearly all patients. First, there is ample evidence that weight gain is inevitable and rapid after reversal. Appetite and well-being return. In one study, patients gained an average of 3 kg/ma after reversal, increasing 45 kg over 19 mo [Arch Surg 1981;116:320-324). In the present study [mean follow-up, 88 mo]. average weight gain after reversal was 38 kg in the patients who did not undergo gastroplasty. The second reason to add gastric restrictive surgery to the JI bypass reversal is the fact that weight loss [a] lessens morbidity and (b] prolongs life in morbidly obese patients. For example, weight loss has been shown to affect favorably cardiac function in morbidly obese men (N Engl J Med 3986;314:334-339) Weight loss has also been shown to improve blood lipid profiles and diabetes mellitus in obese patients (N Engl J Med 1988;319:1173-1179; Ann Surg 1987;206:316-323). Mortality rates in patients who have undergone successful gastric restrictive operations are similar to those in nonobese patients [Am J Surg 1989;157:150-155). For these reasons, gastric restrictive procedures should be added to reversal operations in most patients. The present study performed vertical-banded gastroplasties as the gastric restrictive procedure. In my opinion, the literature supports the claim that the Roux-en-Y gastric bypass is a better operation than the vertical-banded gastroplasty. Randomized trials have established the longevity and superiority of the gastric bypass compared with gastroplasty (Ann Surg 1982;196:389-399; Ann Surg 1987;205:613-624; Am J Surg 1989;157:93-102). During a reversal procedure, the “Roux limb” is already constructed, facilitating gastric: bypass. Regardless of the type of gastric procedure, the reconstruction should be performed at a center familiar with gastric restrictive procedures. In summary, the present study nicely reviews the JI bypass syndrome and then accurately describes the clinical course in patients after reversal with or without gastric restrictive operations. The authors have provided a significant contribution to the treatment of an iatrogenic life-threatening disease which was inflicted upon nearly 25,000 patients in this country between 1970 and 1980. R. C. THIRLEIY.M.D.

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HEPATITIS B INFECTION AND THE PATHOGENESIS OF HEPATOCELLULAR CARCINOMA C&sari FV, Klopchin K, Moriyama T, et al. (Department of Molecular and Experimental Medicine, Research Institute of Scripps Clinic, LaJolla, California; Departments of Pathology, University of Texas Medical Schools at Houston and Galveston, Texas: Laboratory of Reproductive Physiology, University of Pennsylvania School of Veterinary Medicine, Philadelphia, Pennsylvania; and Department of Biochemistry, Howard Hughes Medical Institute, University of Washington, Seattle, Washington). Molecular pathogenesis of hepatocellular carcinoma in hepatitis B virus transgenic mice. Cell 1989;59:1145-1156 [December).

Three lineages of transgenic mice that contained a hepatitis B virus (HBV) DNA fragment under the transcriptional control of the mouse albumin promoter were studied. At both the ribonucleic acid [RNA] and protein levels these three lineages expressed differing levels of hepatitis B surface antigen (HBsAg), presumably as a function of the integration site of the transgene within the mouse genome. However, the molecular forms of the HBV transcripts expressed by the three lineages were identical. Overproduction of the HBV large envelope polypeptide was associated with formation of branching filamentous HBsAg particles that became entrapped in the endoplasmic reticulum and caused the formation of ground glass hepatocytes. In the high and very high HBsAg-producing lineages, but not in the moderate HBsAg-producing lineage, accumulation of intracellular HBsAg eventually led to cell death and the development of an associated inflammatory response that was presumably non-antigen-specific because these mice were HBsAg tolerant (Proc Nat1 Acad Sci USA 1987;84:69096913). The time of onset of hepatic transaminase elevations and the percentage of animals with enzyme abnormalities were found to be directly proportional to the intrahepatic HBsAg concentration, When observed over a period of 2 yr, mice were observed to sequentially develop hepatic injury, regenerative hyperplasia, nuclear aneuploidy, benign hepatic adenomas, and finally multiple foci of hepatocellular carcinoma (HCC) and elevated serum a-fetoprotein (AFP) levels. The incidence of HCC corresponded with the frequency, severity, and age at onset of hepatocyte injury, which itself correlated with intracellular levels of HBsAg. Genetic background and sex also influenced the rate of HCC development, and tumor formation was favored by male sex or the presence of the transgene on heterozygous C57BL/ 6xSJL rather than mice backcrossed with the parental C57BL/6 strain. Thus, overexpression of a single structural viral gene was found to be sufficient to cause malignant transformation. Because transformation was observed in two independent lineages without evidence of transgene rearrangement or instability, direct insertional activation of a cellular oncogene was considered unlikely. Therefore, the authors suggest that in this model, severe, prolonged cellular injury induces a preneoplastic proliferative response that fosters

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secondary genetic events that program the cell for unrestrained growth and neoplastic transformation. Comment. The link between chronic hepatitis B infection and hepatocellular carcinoma has been recognized for 2 decades (Lancet 1970;1:1243-1247). Following the cloning and sequencing of the HBV genome, it became apparent that during prolonged infection, viral deoxyribonucleic acid (DNA] sequences integrate in the host

cell genome, where they and the flanking cellular sequences are commonly rearranged (N Engl J Med 1981;305:1067-1073; J Cell Biochem 1982;20:293-301). These observations immediately suggested the possibility that such integration and rearrangement events might directly activate cellular protooncogenes and hence account for the increased incidence of hepatic malignancy seen in chronic carriers of hepatitis B. Such mechanisms involving active transcription of oncogenes indeed appear to occur in selected human hepatitis B-related hepatocellular carcinomas (Nature 1986; 322:70-72; Am J Gastroenterol 1987;82:512-518). However, such events appear to be rare and thus do not explain the majority of HBV-associated hepatic malignancies. In contrast to such hypotheses, the present manuscript suggests alternative mechanisms for induction of malignant transformation of hepatocytes containing integrated HBV sequences. Hepatitis B virus-transgenic mice have previously been shown to be tolerant to HBV gene products (Proc Nat1 Acad Sci USA 1987;84:6909-6913), yet in lineages with extensive overproduction and intracellular accumulation of HBsAg particles, cell death and secondary inflammatory responses occur. The present findings suggest that the proliferative response to chronic cell injury in such animals contributes to the indirect accumulation of multiple chromosomal abnormalities or other genetic changes sufficient for acquisition of a malignant phenotype. The authors suggest that the propensity to liver cell injury and HCC formation is a direct function of quantity of HBsAg synthesized. However, complete data are provided for only three lineages. The authors do remark, however, that of the many HBV-transgenic mice previously studied and described in the literature, only the two “high-producing” lineages reported in the

present manuscript have been noted to manifest hepatocyte injury and inflammation and to develop HCC. The transgenic model of HBV integration used by Chisari et al. has a number of additional analogies to human HBV-related disease. Thus, male mice exhibited higher levels of HBsAg expression and higher rates of malignant transformation than female mice. Not only does this model recapitulate the male predominance among human HCC patients, but the strain difference in susceptibility to malignant transformation noted in this study also may be analogous to racial imbalances reported in human hepatic carcinoma rates. Finally, some histological features of this animal model, such as the presence of ground glass hepatocytes, appear to be similar to those seen in human HBsAg carriers who indeed remain at risk for development of hepatic malignancies even in the absence of progression to cirrhosis or other manifestations of chronic liver disease. However, in contrast to the total absence of fibrosis or cirrhosis in this HBV-transgenic mouse model, human HBV-related HCC is associated with cirrhosis in 60%-90% of cases [Ann Intern Med 1988;108:390-401). Because a variety of circumstantial evidence also suggests that the majority of HBV-induced liver cell injury in the human disease is mediated by antigen-specific immune responses, it is difficult to infer the extent to which the direct cytopathic effect of intracellular HBsAg accumulation contributes to human hepatocyte injury or to assess whether the resulting patterns of inflammatory response and/or hepatocyte regeneration seen after these two forms of cell death are equally likely to predispose to malignant transformation. In addition, progression through a stage of hepatic adenoma formation before malignant transformation appears to be

GASTROENTEROLOGY

Vol. 98, No. 6

a typical feature of this animal model but has not been suggested as a common feature of human HBV-related HCC development. Despite these differences between human HBV-related liver disease and this HBV-transgenic mouse model, the observations made in this study are likely to influence our conceptual understanding of HBV-related HCC pathogenesis in humans. These findings suggest that in this disease, as in other forms of injury-induced increases in the rate of cell proliferative responses, random secondary genetic changes rather than direct activation of oncogenes by integrated viral DNA sequences may be the predominant cause of malignant transformation. D. L. THIELE, M.D.

HOW DOES THE SCHISTOSOME HOST DEFENSES?

EVADE

Chun-Pin C, Caulfield J. (Departments of Rheumatology/ Immunology and Pathology, Harvard Medical School, the Brigham and Women’s Hospital, and the Department of Tropical Public Health, Harvard School of Public Health, Boston, Massachusetts]. Human lipoprotein binding to schistosomula of schistosoma mansoni. Am J Path01 1989;135:10151024 (December). It has been unclear how schistosomes survive in an environment that is presumed to produce antibodies against foreign tissue antigens. Following infection by the freshwater stage of these parasites, called cercariae, larvae migrate through lungs, liver, and finally portal veins, where they differentiate into adults. During these tissue migrations they are exposed to a variety of antibody-producing cells, yet no effective antibody defense is mounted and the adult worms may survive for 20 or more years. In vitro experishow ments by Butterworth (J Exp Med 1977;145:136-149) that the parasites can be killed by human eosinophils in an antibody-dependent cytotoxic reaction. In vivo, however, both the larvae and the adult worms seem immune from antibody-mediated assault. The main theory to explain this immunity has emphasized the role of host or hostlike antigens on the surface of the parasite (Damian R, Am Naturalist 1964;98:129-149). There is experimental support for this hypothesis. Worms taken from infected mice will survive if injected into monkeys. If the monkeys have been immunized against mouse blood cells, however, the worms are killed, suggesting that the injected schistosomes had developed mouse antigens on their surface. However, this common antigen theory does not fully explain the schistosome.+ apparent immunity. If the adult worms from mice are washed for 4 h, there is an increase in the binding of mouse antischistosomal antibody. This increased binding can be reversed if serum is added to the preparation (Rasmussen KJ, Parasitology 1985;71:530-534). These observations suggest the possible role of a component of host serum that may actually provide protection to the parasite. The authors have previously shown that low-density lipoprotein (LDL) can inhibit the binding of antischistosomal antibody to schistosomal larvae. In the present study the role of LDL was further defined by measuring the degree of binding of labeled antibodies in the presence of LDL under a variety of conditions. The LDL was added to a test solution of worms and the percent of the worm surface area coated