Loss of end-differentiated ß-cell phenotype following pancreatic islet transplantation.

Replacement of pancreatic ß-cells through deceased donor islet transplantation is a proven therapy for preventing recurrent life-threatening hypoglycemia in type 1 diabetes. Although near-normal glucose levels and insulin independence can be maintained for many years following successful islet transplantation, restoration of normal functional ß-cell mass has remained elusive. It has recently been proposed that dedifferentiation/plasticity towards other endocrine phenotypes may play an important role in stress-induced ß-cell dysfunction in type 2 diabetes. Here we report loss of end-differentiated ß-cell phenotype in 2 intraportal islet allotransplant recipients. Despite excellent graft function and sustained insulin independence, all examined insulin-positive cells had lost expression of the end-differentiation marker, urocortin-3, or appeared to co-express the α-cell marker, glucagon. In contrast, no insulin+ /urocortin-3- cells were seen in nondiabetic deceased donor control pancreatic islets. Loss of end-differentiated phenotype may facilitate ß-cell survival during the stresses associated with islet isolation and culture, in addition to sustained hypoxia following engraftment. As further refinements in islet isolation and culture are made in parallel with exploration of alternative ß-cell sources, graft sites, and ultimately fully vascularized bioengineered insulin-secreting microtissues, differentiation status immunostaining provides a novel tool to assess whether fully mature ß-cell phenotype has been maintained.

Associations between dietary intake and the presence of the metabolic syndrome in patients with non-alcoholic fatty liver disease.

BACKGROUND: Although dietary habits have been associated with the likelihood of the metabolic syndrome (MetS) in the general population, similar associations in non-alcoholic fatty liver disease (NAFLD) patients have not been explored. The aim of this cross-sectional study was to assess the presence of the MetS and to explore its potential association with dietary habits in a sample of NAFLD patients. METHODS: Seventy-three adult patients with recent NAFLD diagnosis based on elevated liver enzyme levels and evidence of hepatic steatosis on ultrasound were enrolled. Participants' habitual food consumption was retrospectively assessed through a food frequency questionnaire and adherence to the Mediterranean diet (MD) was assessed via the Mediterranean Diet Score (MedDietScore). The presence of the MetS was defined as the concomitant presence of at least three of its individual components, according to the criteria proposed by a recent joint statement of several major organisations. RESULTS: The MetS was present in 46.5% of the sample, with increased waist circumference values and decreased high-density lipoprotein cholesterol levels being the most prevalent disorders (63% and 88.7%, respectively). Consumption of refined grains [odds ratio (OR) = 1.02, 95% confidence interval (CI) = 1.00-1.05] and red meat and products (OR = 1.10, 95% CI = 1.01-1.21) were positively associated with the presence of the MetS, whereas the consumption of whole grains (OR = 0.92, 95% CI = 0.84-0.99) and MedDietScore (OR = 0.87, 95% CI = 0.76-0.99) were negatively associated, after adjusting for participants' age, sex, daily energy intake and time spent in sedentary activities. CONCLUSIONS: Low refined grain and red meat intake, high whole grain intake and high adherence to the MD were associated with lower odds of the MetS in NAFLD patients.

Keratin 7 expression in hepatic cholestatic diseases.

We evaluated keratin 7 (K7) hepatocellular expression in 92 patients with common types of acute and chronic cholestatic diseases caused by bile duct obstruction/destruction or parenchymal lesions [acute hepatitis (n=20), mixed/pure cholestasis (n=16), primary biliary cholangitis-PBC (n=35), primary sclerosing cholangitis-PSC (n=10), vanishing bile duct syndrome (n=3), complete large bile duct obstruction due to space-occupying lesions (n=8)]. K7 immunohistochemical hepatocellular expression and ductular reaction (DR) were semi-quantitatively assessed. Results were correlated with liver enzyme serum levels, cholestasis type, histological features, hepatocellular Ki67 labelling index (LI) and HepPar1 expression. Hepatocellular K7 expression was detected in 87% (81/92) cases and in all cholestatic disease types with lowest incidence in pure/mixed cholestasis and highest in incomplete bile duct obstruction (iBDO), reaching 100% in PSC. K7-positive hepatocytes had low Ki67 LI (0-5%) retaining HepPar1 expression, irrespective of disease type. PSC cases had high K7 hepatocellular expression even with intact bile ducts, a feature that may aid differential diagnosis of cholestatic syndromes. K7 hepatocellular expression significantly correlated with cholestasis type, bile duct loss and fibrosis stage. It was higher in milder acute cholestatic hepatitis showing inverse correlation with hepatocyte proliferation and serum transaminase levels. In iBDO, younger age independently correlated with high K7 expression, while serum GGT levels showed a nearly significant correlation. Correlation with DR findings implied that K7-positive hepatocytes may result through metaplasia. In conclusion, K7 hepatocellular expression is a sensitive though non-specific marker of cholestasis. It may represent a cytoprotective reaction of resting hepatocytes in cholestasis of longer duration especially in younger patients.

Right sited renal cell carcinoma metastasizing to the contralateral ovary: case report and review of the literature.

Ovarian metastases from renal cell carcinoma are rare, with only 22 cases reported in the literature. We report a case of a 45-year-old woman, who developed left ovarian and right adrenal metastases 3 months after diagnosis of clear cell renal cell carcinoma and review the literature. This is the fourth reported case of right renal cell carcinoma metastasizing to the left ovary. The patient is alive 4 years after resection of the ovarian tumor, treated with sunitinib. We conclude that, although rare, metastatic renal cell carcinoma should be included in the differential diagnosis of ovarian tumors with clear cell histology.

Liver biopsy in alcoholic and non-alcoholic steatohepatitis patients.

Alcoholic liver disease and non-alcoholic liver disease share a similar histological spectrum that starts with 'simple' steatosis, and may be accompanied by inflammation. Alcoholic steatohepatitis and non-alcoholic steatohepatitis (NASH) are progressive forms of alcoholic liver disease and non-alcoholic liver disease, respectively, and can evolve into cirrhosis. The currently accepted minimum diagnostic criteria for steatohepatitis include steatosis, lobular inflammation and hepatocellular injury, but not fibrosis. Steatosis involving more than 5% of hepatocytes is required for the diagnosis of non-alcoholic fatty liver disease, but is not necessary for the diagnosis of alcoholic liver disease. Lobular inflammation is usually mild and frequently consists of a mixed, acute and chronic, inflammatory cell infiltrate composed of neutrophils and mononuclear cells. The presence of large numbers of neutrophils favors an alcoholic etiology. Hepatocellular injury in fatty liver disease usually occurs in the form of ballooning, but it can also present as apoptotic (acidophilic) bodies and lytic necrosis. The characteristic pattern of fibrosis in non-cirrhotic steatohepatitis is pericellular/perisinusoidal and is the result of deposition of collagen in the space of Disse. In both alcoholic steatohepatitis and NASH, sinusoidal collagen formation is the result of hepatic stellate cell activation that, in NASH, has been correlated with the grade of steatosis and fibrosis.

Serum apoptotic caspase activity as a marker of severity in HBeAg-negative chronic hepatitis B virus infection.

BACKGROUND AND AIMS: In chronic hepatitis C and non-alcoholic fatty liver disease, apoptotic caspases are activated in liver, and serum caspase activity has been suggested as a sensitive marker of early liver injury. An investigation was carried out into whether the serum levels of caspase-generated fragments of cytokeratin-18 (CK-18) are associated with the severity of liver lesions in hepatitis B e antigen (HBeAg)-negative chronic hepatitis B virus (HBV) infection. Patients/ METHODS: CK-18 fragment serum levels were determined in 115 treatment-naive, consecutive HBV patients and 30 healthy controls. Hepatic-expression of CK-18 fragments was evaluated by immunocytochemistry in chronic hepatitis B patients. RESULTS: CK-18 fragment levels (U/l) were significantly lower in healthy controls (mean (SD), 154 (31)) than in 53 inactive carriers (172 (24), p = 0.003) and in 62 chronic hepatitis B patients (474 (488), p<0.001). The receiver operating characteristic curve showed excellent diagnostic accuracy (c-statistic: 0.87) for differentiating inactive carriers from chronic hepatitis B patients. A CK-18 fragment cut-off level of 240 U/l gave a sensitivity of 60%, and a specificity and positive predictive value of 100% for chronic hepatitis B diagnosis. CK-18 fragment levels were also lower in inactive carriers than in 16 chronic hepatitis B patients with transiently normal alanine aminotransferase (ALT; 327 (256), p = 0.001), offering good accuracy for such a differentiation (c-statistic: 0.78). In chronic hepatitis B patients, serum CK-18 fragments correlated positively with ALT/aspartate aminotransferase (AST), viraemia, grading score and their immunohistochemical hepatic expression, and negatively with platelet counts, but not with fibrosis or steatosis severity. CONCLUSIONS: Serum apoptotic caspase activity is strongly associated with the presence of liver injury in patients with HBeAg-negative chronic HBV infection. CK-18 fragment levels seem to be a very useful marker for differentiation between the inactive HBV carrier state and HBeAg-negative chronic hepatitis B, but not for estimation of the severity of liver histological lesions among HBeAg-negative chronic hepatitis B patients.

KRAS mutation testing for predicting response to anti-EGFR therapy for colorectal carcinoma: proposal for an European quality assurance program.

Novel therapeutic agents targeting the epidermal growth factor receptor (EGFR) have improved outcomes for patients with colorectal carcinoma. However, these therapies are effective only in a subset of patients. Activating mutations in the KRAS gene are found in 30-40% of colorectal tumors and are associated with poor response to anti-EGFR therapies. Thus, KRAS mutation status can predict which patient may or may not benefit from anti-EGFR therapy. Although many diagnostic tools have been developed for KRAS mutation analysis, validated methods and standardized testing procedures are lacking. This poses a challenge for the optimal use of anti-EGFR therapies in the management of colorectal carcinoma. Here we review the molecular basis of EGFR-targeted therapies and the resistance to treatment conferred by KRAS mutations. We also present guideline recommendations and a proposal for a European quality assurance program to help ensure accuracy and proficiency in KRAS mutation testing across the European Union.

Evaluation of combined h-TERT, bcl-2, and caspases 3 and 8 expression in cutaneous malignant melanoma based on tissue microarrays and computerized image analysis.

PURPOSE: Deregulation of apoptotic pathways in cutaneous malignant melanoma appears to be correlated with chemoresistance and poor prognosis. Furthermore, telomerase (especially h-TERT) expression induces proliferation and also represents a potential target for vaccination regarding some types of malignancies. MATERIALS AND METHODS: Using tissue microarrays (TMA) technology, 25 paraffin-embedded tissue samples of histologically confirmed malignant melanomas were cored at a diameter of 2 mm and re-embedded into one recipient block (final TMA density 24/25-96%). Immunohistochemistry (IHC) was performed by the use of anti-bcl-2, anti-caspase 3, anti-caspase 8 and anti- h-TERT antibodies. Protein expression levels were evaluated using a computerized image analysis system (CIA). SPSS (chi square test and inter-rater kappa) was used for statistical analysis. RESULTS: Strong protein expression was observed in 1/24 (4.1%), 1/24 (4.1%), 2/24 (8.2%), and 4/24 (16.4%) cases regarding h-TERT, caspase 3, caspase 8 and bcl-2, respectively. Moderate was observed in 7/24 (29.1%), 8/24 (32.2%), 5/24 (20.2%), and 8/24(32.2%) cases, whereas reduced or absent expression demonstrated 16/24 (65%), 15/24 (60.2%), 17/24 (68.5%), and 12/24 (50 %) cases. Statistical significance was assessed correlating age to caspase 3 (p=0.05), Breslow's thickness to telomerase (p=0.013) and to bcl-2 (p=0.053), Clark's level to telomerase (p=0.008) and to bcl-2 (p=0.022), and finally ulceration to telomerase expression (p=0.007). CONCLUSION: bcl-2 and telomerase expression are correlated to critical parameters of malignant melanoma, affecting its biological behavior. Furthermore, downregulation of proteins such as caspases 3/8, which normally induce apoptosis, is perhaps associated with resistance of the applied chemotherapeutic strategies in this type of malignancy.

A RelA(p65) Thr505 phospho-site mutation reveals an important mechanism regulating NF-κB-dependent liver regeneration and cancer.

Post-translational modifications of nuclear factor (NF)-κB subunits provide a mechanism to differentially regulate their activity in response to the many stimuli that induce this pathway. However, the physiological significance of these modifications is largely unknown, and it remains unclear if these have a critical role in the normal and pathological functions of NF-κB in vivo. Among these, phosphorylation of the RelA(p65) Thr505 residue has been described as an important regulator of NF-κB activity in cell lines, but its physiological significance was not known. Therefore, to learn more about the role of this pathway in vivo, we generated a knockin mouse with a RelA T505A mutation. Unlike RelA knockout mice, the RelA T505A mice develop normally but exhibit aberrant hepatocyte proliferation following liver partial hepatectomy or damage resulting from carbon tetrachloride (CCl4) treatment. Consistent with these effects, RelA T505A mice exhibit earlier onset of cancer in the N-nitrosodiethylamine model of hepatocellular carcinoma. These data reveal a critical pathway controlling NF-κB function in the liver that acts to suppress the tumour-promoting activities of RelA.

Diagnostic performance of FibroTest, SteatoTest and ActiTest in patients with NAFLD using the SAF score as histological reference.

BACKGROUND: Blood tests of liver injury are less well validated in non-alcoholic fatty liver disease (NAFLD) than in patients with chronic viral hepatitis. AIMS: To improve the validation of three blood tests used in NAFLD patients, FibroTest for fibrosis staging, SteatoTest for steatosis grading and ActiTest for inflammation activity grading. METHODS: We pre-included new NAFLD patients with biopsy and blood tests from a single-centre cohort (FibroFrance) and from the multicentre FLIP consortium. Contemporaneous biopsies were blindly assessed using the new steatosis, activity and fibrosis (SAF) score, which provides a reliable and reproducible diagnosis and grading/staging of the three elementary features of NAFLD (steatosis, inflammatory activity) and fibrosis with reduced interobserver variability. We used nonbinary-ROC (NonBinAUROC) as the main endpoint to prevent spectrum effect and multiple testing. RESULTS: A total of 600 patients with reliable tests and biopsies were included. The mean NonBinAUROCs (95% CI) of tests were all significant (P < 0.0001): 0.878 (0.864-0.892) for FibroTest and fibrosis stages, 0.846 (0.830-0.862) for ActiTest and activity grades, and 0.822 (0.804-0.840) for SteatoTest and steatosis grades. FibroTest had a higher NonBinAUROC than BARD (0.836; 0.820-0.852; P = 0.0001), FIB4 (0.845; 0.829-0.861; P = 0.007) but not significantly different than the NAFLD score (0.866; 0.850-0.882; P = 0.26). FibroTest had a significant difference in median values between adjacent stage F2 and stage F1 contrarily to BARD, FIB4 and NAFLD scores (Bonferroni test P < 0.05). CONCLUSIONS: In patients with NAFLD, SteatoTest, ActiTest and FibroTest are non-invasive tests that offer an alternative to biopsy, and they correlate with the simple grading/staging of the SAF scoring system across the three elementary features of NAFLD: steatosis, inflammatory activity and fibrosis.

"Bath Salts" intoxication with multiorgan failure and left-sided ischemic colitis: a case report.

BACKGROUND: In the recent years, a new group of designer drugs, under the brand name of bath salts has emerged as a new trend. They are mainly b-ketone amphetamine analogs and are derivatives of cathinone, a monoamine alkaloid. They are abused for psychostimulant effects. Their primary ingredient 3,4-methylenedioxypyrovalerone (MDPV), has alerted authorities worldwide due to its severe physiological and behavioral toxicities. Description of Case:We present the case of a 47-year-old man with coma, seizures, multi-organ failure and ischemic colitis after intoxication with bath salts containing MDPV. After supportive care, he had a successful outcome. To our knowledge, this report is the first to describe ischemic colitis after MDPV intoxication. Clinicians need to be especially alert since MDPV is not detected by routine screens, and its overdose can be life-threatening. CONCLUSION: Ischemic colitis should be recognized as a potential complication of bath salts ingestion in order to prevent unnecessary interventions, such as diagnostic laparotomy, which could worsen patient's condition. Hippokratia 2015; 19 (4): 363-365.

NFκB1 is a suppressor of neutrophil-driven hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) develops on the background of chronic hepatitis. Leukocytes found within the HCC microenvironment are implicated as regulators of tumour growth. We show that diethylnitrosamine (DEN)-induced murine HCC is attenuated by antibody-mediated depletion of hepatic neutrophils, the latter stimulating hepatocellular ROS and telomere DNA damage. We additionally report a previously unappreciated tumour suppressor function for hepatocellular nfkb1 operating via p50:p50 dimers and the co-repressor HDAC1. These anti-inflammatory proteins combine to transcriptionally repress hepatic expression of a S100A8/9, CXCL1 and CXCL2 neutrophil chemokine network. Loss of nfkb1 promotes ageing-associated chronic liver disease (CLD), characterized by steatosis, neutrophillia, fibrosis, hepatocyte telomere damage and HCC. Nfkb1(S340A/S340A)mice carrying a mutation designed to selectively disrupt p50:p50:HDAC1 complexes are more susceptible to HCC; by contrast, mice lacking S100A9 express reduced neutrophil chemokines and are protected from HCC. Inhibiting neutrophil accumulation in CLD or targeting their tumour-promoting activities may offer therapeutic opportunities in HCC.

Detection of hepatitis-B virus-DNA and mutations in k-ras and p53 genes in human hepatocellular carcinomas.

Hepatitis B virus (HBV) infection is considered as one of the major risk factors in the development of human hepatocellular carcinoma (HCC). Recent studies have also suggested the implication of oncogene and onco-suppressor genes in liver carcinogenesis. We studied 41 cases of HCC for the presence of HBV DNA and point mutations in codon 12 of K-ras and codon 249 of p53. We used 'nested' PCR for the amplification of HBV because of the expected low incidence of the virus DNA in the samples. PCR was also used for the amplification of K-ras and p53 regions that contain the codons of interest, followed by RFLP analysis for the detection of point mutations. HBV DNA was amplified in 22 cases (53.7%), while 5 cases (12.2%) appeared to carry mutations in codon 12 of K-ras and 7 cases (17.1%) had mutations in codon 249 of the p53 gene. These results further support the correlation between HBV infection and HCC and also indicate an implication of K-ras and p53 genes in hepatocarcinogenesis.

Osteopontin expression in ovarian carcinomas and tumors of low malignant potential (LMP).

Expression of osteopontin (OPN) by ovarian tumors is not known. Neoplasms arising from the ovarian epithelium are distinguished in adenocarcinomas and borderline tumors (LMPs), which overall have a favorable prognosis even with omental implants. Tissues from primary ovarian tumors and their metastases from 30 patients (16 LMPs and 14 adenocarcinomas) were evaluated for OPN expression by immunohistochemistry, Western blotting, and in situ hybridization. OPN was weak or absent in 93% of ovarian adenocarcinomas or their metastases. In contrast, 81.5% of the LMPs and 50% of omental and lymph node implants were OPN positive (P < .028). Histological type, grade, or clinical stage did not correlate with OPN expression. Expression of OPN primarily by ovarian neoplasms with favorable prognosis is an intriguing new finding of potential importance in the pathogenesis of ovarian LMPs.

Co-expression of trypsin and tumour-associated trypsin inhibitor (TATI) in colorectal adenocarcinomas.

Trypsin and its specific inhibitor, TATI (tumour-associated trypsin inhibitor), are expressed in normal human pancreas and in a variety of tumours. The aim of the present study was to assess the parallel expression of trypsin and TATI in colorectal cancer, in comparison with their expression in normal epithelial tissue, since proteases and their inhibitors are thought to be co-expressed in malignant neoplasms. We also assessed the possible significance of their expression as a means of differentiation between normal and malignant tissue. We examined qualitatively and semi-quantitatively the immunohistochemical expression of trypsin and TATI on paraffin-embedded serial tissue sections from 91 colorectal adenocarcinomas. The reverse-transcriptase-polymerase-chain reaction (RT-PCR) was also performed on fresh malignant tissue from 55 of the above adenocarcinomas. Normal and non-malignant tissues adjacent to the tumours were also evaluated. Cytoplasmic expression of trypsin (more than 25% of the cancer cells positive) was found in 67 (73.6%) adenocarcinomas, whereas TATI was expressed in the cytoplasm of 59 (64.8%) cases studied. Statistical analysis using Spearman's test has demonstrated a significant correlation between trypsin and TATI immunohistochemical expression (p<0.01). RT-PCR showed co-expression of trypsin and TATI mRNA in all carcinomas studied. Distinct patterns of trypsin and TATI immunohistochemical expression were observed in adjacent, non-malignant tissues, where both trypsin and TATI mRNA were also detected. Normal tissues were negative by immunohistochemistry. Our results indicate co-expression of trypsin and TATI in colorectal tumours both at the mRNA and protein level. We conclude that in colorectal neoplasms, high levels of trypsin and TATI may be important for malignant tumour formation and/or metastatic process.

Donor lung pretreatment with surfactant in experimental transplantation preserves graft hemodynamics and alveolar morphology.

In experimental lung transplantation, the reduction of endogenous surfactant properties occurs after graft preservation and transplant reperfusion. The aim of this study was to evaluate the efficacy of donor lung pretreatment with exogenous surfactant on graft damage after ischemia and reperfusion. Fourteen (control group A, n = 8; study group B, n= 6) young female white pigs (mean weight 27 +/- 3.5 kg) were used in a newly developed autotransplantation model within situcold ischemia. In study group B, before thoracotomy, 1.5 ml/kg surfactant apoprotein-A-free surfactant was administrated into the left main bronchus via flexible bronchoscopy. Belzer UW solution was used for lung preservation. Cold ischemia was achieved for 3 hr with interlobar lung parenchyma temperature at 8 +/- 1.3 degrees C, and central temperature maintained at 37.20 +/- 0.5 degrees C. Animals were sacrificed after 3 hr of graft reperfusion. At the end of reperfusion, pulmonary vascular resistance index (was 447.80 dyn/sec.cm(5).m(2)(+/-66.8) in group A vs 249.51 in group B (P< 0.001) and serum nitric oxide was adequately preserved. The mean alveolar surface area estimated by computerized morphometry was 5280.84 (4991.1) microm(2)(group A) vs 3997.89 (3284.70) microm(2)(group B;P< 0.005). Histology revealed milder macrophage and lymphocyte infiltration in group B at the end of reperfusion. Pretreatment of donor lung with an surfactant apoprotein-A -free surfactant agent appears to be beneficial in terms of maintaining serum NO and reducing hemodynamic disturbances. Furthermore, alveolar histology and stereomorphology are better preserved.

Expression of ras and myc oncogenes in human hepatocellular carcinoma and non-neoplastic liver tissues.

An immunohistochemical assay was used to assess expression of ras p21 and myc p62 oncogene products in human hepatocellular carcinoma (HCC) and non-neoplastic liver tissues. The monoclonal antibodies Y13 259 and Myc1-9E10, specific for ras p21 and myc p62 oncoproteins, were employed on paraffin-embedded sections. Most HCCs showed enhanced ras p21 and myc p62 expression, as indicated by staining intensity. Cirrhotic livers revealed increased myc p62 and occasionally increased ras p21 expression. HBsAg+ hepatocytes showed intense immunostaining for ras p21. Fibrotic, cholestatic, fetal and normal adult liver did not present enhancement of oncoprotein production. We suggest that combined over-expression of ras and myc oncoproteins may be important for the malignant phenotypic alteration in human HCC.

Recent advances in early prostatic cancer.

Early prostatic carcinoma is a slowly progressing, localized malignant tumor which has been recently discovered with increased frequency due to the use of improved diagnostic methods. The combination of digital rectal examination, serum PSA level and transrectal ultrasound is currently the best available diagnostic tool, although other putative diagnostic markers and techniques are being investigated. Core needle biopsy may follow if there is suspicion of malignancy and in doubtful cases the most useful antibody for the immunohistochemical diagnosis of early, low grade prostatic carcinoma is clone 34 beta E12. Cytogenetic techniques and molecular biological methods are increasingly being used for further investigating localized prostate carcinomas in order to identify early molecular targets and alterations, which may lead to progression. Chromosome abnormalities, cell to cell and cell to matrix interactions, changes in the status of steroid hormone receptors, oncogenes and tumor suppressor genes, as well as other, as yet unclear, events may be of importance in prostate carcinogenesis and the progression of early malignant tumors to aggressive phenotypes. A variety of putative prognostic markers, apart from serum PSA levels, histological grade and tumor volume, such as neuroendocrine differentiation, angiogenesis, cell proliferation labeling index and ploidy analysis may prove useful in evaluating tumor progression in early prostatic carcinomas. The final and most important goal of all investigations related to early prostate cancer is to contribute to the best therapeutic management of the individual patient.

Expression of ras and c-myc oncoproteins and hepatitis B surface antigen in human liver disease.

One of the major antecedent factors preceding the development of hepatocellular carcinoma is chronic hepatitis B virus infection. Also, recent molecular studies have shown that activation of c-oncogenes might be responsible for the malignant transformation in some cases of hepatocellular carcinoma. We used immunohistochemical methods to investigate the correlation of ras and c-myc oncogene expression with the presence of HBsAg in human liver disease. Our material consisted of 23 chronic active hepatitis B needle liver biopsies and surgical specimens from 11 cases of cirrhosis, 23 hepatocellular carcinoma and 10 normal adult livers. Direct, three-step and streptavidin-biotin-complex immunoperoxidase techniques using polyclonal (anti-HBsAg) and monoclonal antibodies (anti-ras p21, anti-myc p62), were performed. Normal liver tissues were negative for all antibodies used. In HBsAg+ chronic active hepatitis B cases enhancement of c-myc, and less frequently of ras oncogene expression, was a common observation. Increased myc p62 and ras p21 expression was a finding not restricted to HBsAg+hepatocytes, which occasionally were negative for oncoprotein immunostaining. All HBsAg-chronic active hepatitis B cases were negative for ras p21 and myc p62 specific staining. Cirrhotic livers showed more frequently enhanced c-myc expression. Most of the immunostained cells were negative for HBsAg. HBsAg- cases of hepatocellular carcinoma more often showed ras p21 than myc p62 overexpression. HBsAg+ hepatocellular carcinomas presented only ras p21-positive immunostaining, which was not detected in HBsAg+ hepatocytes. Our recent data supports the view that continued expression of HBsAg in human liver disease is not necessary for the enhancement of ras and c-myc oncogene expression.