Impact of genetic polymorphisms of four cytokine [G] genes on treatment induced viral clearance in HCV infected Egyptian patients

Background: Many factors contribute for viral clearance and response to antiviral therapy. Genetic polymorphisms of cytokines, chemokines, and their receptors can alter the immune response against Hepatitis C virus [HCV]

Aim of the study: The aim of the current study is to assess single nucleotide polymorphism [SNP] in the promoter region of IL-10, TNF-alpha, IFN-y and TGF-p as predictors of response to combined Pegylated interferon ot/ribavirin [PEG-IFN/RBV] therapy in chronic HCV infected Egyptian patients

Patients and methods: The study was conducted on 150 HCV infected patients and 100 apparently healthy control subjects. All patients were treated with PEG-IFN/RBV. They were classified according to their icsponse to treatment

Genotyping of IL-10, TNF-alpha, IFN-y and TGF-p were performed on peripheral blood DNA using polymerase chain reaction-restriction fragment-length polymorphism [PCR-RFLP] and primer specific assays

Results: Overall, 83/150 [55.3%] patients achieved sustained virological response [SVR], whereas 67 [44.7%] did not. Age and BMI were significantly lower in patients who achieved SVR [P < 0.05]. IL-10 at site [-1082] GG genotype was associated with SVR where odds ratio was 1.98 with 95% confidence interval [1.34-3.65]

None of the other genes showed a significant association with SVR

Conclusion: Analysis of IL-10 SNP at promoter site [-1082] could be used as a pretreatment predictor of response to combined PEG-IFN/RBV treatment

Circulating MiRNA-21 and programed cell death [PDCD] 4 gene expression in hepatocellular carcinoma [HCC] in Egyptian patients

Background: Circulating microRNAs [miRNAs] are endogenous, small [17-25 nucleo-tides] non-coding RNAs that are overexpressed in many human cancers including hepatocellular carcinoma [HCC]. Moreover, circulating miRNAs can reflect the level of tissue miRNAs, so could be potential tumor markers. miRNA-21 regulates post-transcriptional expression of tumor suppressor gene; programed cell death 4 [PDCD4] gene which implies that miRNA-21 might be a novel diagnostic and/or prognostic marker for cancer

Objective: To evaluate the diagnostic and prognostic potential of circulating miRNA-21 and study the expression of PDCD4 gene as a target of miRNA-21 in HCC in Egyptian patients

Subjects and methods: This study was conducted on 30 HCC patients, 20 chronic liver disease [CLD] patients due to HCV infection and 20 healthy subjects. Serum alpha fetoprotein [AFP] was measured for all participants. The relative plasma expression of each of miRNA-21 and PDCD4 gene was determined in whole blood samples using real-time polymerase chain reaction

Results: The results revealed over expression of miRNA-21 and under expression of PDCD4 gene in HCC group [p < 0.05] compared to both CLD and healthy subjects, while no significant change was detected between CLD and healthy subjects. miRNA-21 expression was negatively correlated with PDCD4 gene expression. miRNA-21 expression increased significantly with presence of cirrhosis, increased number of focal lesions, larger size of tumor, advanced tumor stage and presence of vascular invasion. Receiver Operator of Characteristics [ROC] curve analysis of plasma miRNA-21 revealed that, at a cut-off value of 3.93 [fold expression], the sensitivity and specificity for differentiation of HCC cases were 93% and 90%, respectively

Conclusion: Circulating miRNA-21 could be a novel early diagnostic and prognostic biomarker for detection of HCC. Approaches interfering with the miRNA-2 l/PDCD4-axis, or releasing PDCD4 expression, may have a strong basis for therapeutic uses in cancer in the future

Impact of IL28B polymorphism on treatment induced viral clearance in HCV infected Egyptian patients

Interleukin [IL] 28B single nucleotide polymorphisms [SNP] was recently recognized as predictor of SVR in HCV infected patients treated by combination therapy of pegylatedinterferon [Peg-IFN] and ribavirin [RBV]. The aim of the current study was to assess IL 28B polymorphism SNP [rs12979860] as a predictor of response to combined Peg-INF/RBV therapy in Egyptian chronic HCV infected patients. The study was conducted on 247 HCV infected patients and 100 apparently healthy control subjects. All patients were treated with PEG-IFN-alpha/ribavirin; and they were classified according to their response to treatment. Genotyping of IL28B rs12979860 was performed on peripheral blood DNA using polymerase chain reaction restriction fragment length polymorphism [PCR-RFLP] assay. The overall frequency of IL28B genotypes was 24.7%, 50.2% and 25.1% for genotypes CC, CT and TT respectively, while the response rate was 82%, 38.7% and 43.8% for genotypes CC, CT and TT respectively, moreover, genotype CC had increased probability to HCV clearance than both genotypes CT and TT with OR 7.71 [95%CI: 3.71-15.79]. Genotyping of IL28B at SNP rs12979860 could be used as a guide to tailor treatment in Egyptian patients infected with HCV for better outcome

Value of serum lentil affinity of alpha-fetoprotein in the diagnosis of hepatocellular carcinoma on top of liver cirrhosis

Hepatocellular carcinoma [HCC] is a common complication in patients with liver cirrhosis [LC]. Detection of HCC at an early stage is critical for a favorable clinical outcome. AFP-L3% is an isoform of AFP which is very specific for HCC. The AFP-L3% is the percentage of AFP-L3 over the total AFP level. The study aimed to evaluate the utility of AFPL-3% in detection of HCC developing on top of liver cirrhosis, to compare the levels of both alpha- fetoprotein [AFP] and AFP-L3% in HCC versus LC patients without HCC and to define the cut-off level of each tumor marker with the best sensitivity and specificity for HCC detection. The study was conducted on 25 cases of HCC that developed on top of LC and 25 LC cases with no evidence of HCC, as well as 25 apparently healthy controls. The levels of AFP and AFP-L3% were measured for all cases. Biochemical parameters and viral markers were also tested. Imaging and histopathological evidence of HCC were a prerequisite for inclusion in HCC group. Patients included in the HCC group had total AFP value < 200 ng/ml, which is not a diagnostic level for HCC. Levels of AFP and AFP L-3% were significantly higher in patients with HCC compared to those without HCC [P < 0.01]. Receiver-operating characteristic [ROC] curve analysis indicated that the best cut-off value was 15.4% for AFP-L3% to detect HCC as the sensitivity, specificity, positive predictive value [PPV], negative predictive value [NPV] and accuracy were 79.2%, 100%, 100%, 83.3% and 96.2% respectively. For AFP, the best cut-off in the non diagnostic range was 77.8 ng/ ml as the sensitivity, specificity, PPV, NPV, and accuracy were 75%, 68%, 69.2%, 73.9% and 70.4% respectively, The mean serum level of AFP showed no significant difference [P>0.05] regarding Child Pugh classification, numbers of tumor foci nor rumor size, however, it showed a significant difference [P<0.05] regarding lymph nodes invasion and TNM classification in HCC patients. Meanwhile, AFP-L3% showed no significant difference [P>0.05] regarding all these parameters. There was a positive significant correlation [P<0.05] between AFP and both AST and ALT, while AFP-L3% showed inverse significant correlation [P<0.05] with PC%. No significant correlation [P>0.05] was observed between serum AFP and serum AFP-L3% in HCC patients. In patients with total AFP values <200 ng/ml who present a diagnostic dilemma, AFP-L3% had higher sensitivity, specificity, PPV, NPV and accuracy for HCC detection, and was not elevated in any of the patients without HCC with specificity of 100%

Some apoptotic signals in patients with chronic HCV infection

Hepatitis C virus [HCV] infects an estimated 3% of the world population and causes an estimated 470,000 death per year caused by complications of the end stage liver disease. Mechanisms leading to liver cell injury, are not fully understood, both immune mediated reactions and more direct cytopathic effects of HCV may be involved. Agoptosis of liver cells may play a significant role in the pathogenesis of hepatitis C. The aim of this study is to evaluate the role of apoptosis in chronic HCV infection by estimating serum levels of Fas, Fas-Ligand [FasL] and hepatocyte expression of Bcl-2 of patients with chronic hepatitis C virus infection, and find out the relationship [if any] between their levels and the degree of hepatic inflammatory activity. Sixty patients having chronic HCV infection [45 males and 15 females; mean age 36h8.6 years] were randomly selected. In addition, 20 apparently healthy subjects [12 males and 8 females; mean age 33.1 +/- 10.4 years] served as a control group. They were anti-HCV and HBsAg negative. Serum soluble Fas and Fas ligand levels were measured and HCV RNA quantitation was done to patients only. Immunohistochemical detection of Bcl-2 expression in the liver tissue was done to all patients. According to the histopathological assessment of hepatic necroinflammatory activity, patients were classified into 3 groups with minimal [n=23, GI], mild [n=23, GII] and moderate and severe [n=14, GIII] activity. Significant positive correlations were found between serum Fas and FasL and the grade of hepatic inflammatory activity. Mean serum levels of Fas in group I, II, and III were [0.7 +/- 0.3, 2.5/1.1 and 5.2 +/- 0.9 nglml] respectively with p<0.01. Mean serum levels of FasL in group GI, GII, and GIII were [0.6 +/- 0.2, 1.7+-0.8and 4 +/- 1.2 nglml] respectively with p<0.01. Bcl-2 expression was more prevalent in liver tissues of patients in GIII where 35.7% of hepatocytes, 57.1% of Kupffer cells and 50% of bile duct cells stained positively. For the three types of cells, it correlated positively with the grade of inflammatory activity [p<0.01 for each]. Percentage of Bci-2 expression in portal infiltrates in GI, GI1 and GI11 were [34.8, 78.3 and 100 respectively with p<0.01]. The results of the current work provide evidence of increased apoptosis in chronic hepatitis of moderate and severe activity and in cirrhosis due to chronic HCV infection suggesting that apoptotic cell death might be involved in the pathways of hepatocellular damage in both conditions. Serum sFas and sFasL levels positively correlated with the degree of necroinflammatory process in chronic HCV patients. Accordingly, serum sFas and sFasL could serve as non-invasive serological indicators of the hepatic inflammatory activity. Furthermore, the increase of serum sFas and sFasL [apoptotic signaling] as well as the increased expression of Bcl-2 [anti-apoptosis] at the same time, suggests that Bcl-2 might play as a protective mechanism against apoptosis. However, apoptosis caused by the activation of Fas/FasL pathway seems to occur by a mechanism that might not be blocked by Bcl-2

Relationship of tumor necrosis factor-alpha, interleukin-6 and endotoxin with the development of renal impairment in cirrhotic patients with spontaneous bacterial peritonitis

The aim of this study was to find a relation between the degree of inflammatory response spontaneous bacterial peritonitis represented by endotoxin, tumor necrosis factor-alpha and interleukin-6 levels and the development of renal impairment. Forty-two cirrhotic patients with spontaneous bacterial peritonitis were selected from the inpatients in the Liver Institute. Serum and ascitic fluid cytokines levels at the time of diagnosis were significantly higher in the patients who developed renal impairment compared to those who did not. In conclusion, the degree of inflammatory response in spontaneous bacterial peritonitis at diagnosis may be an important factor in the pathogenesis of renal impairment in such cases

Superiority of polymerase chain reaction in detection of E-coli over conventional cultural methods in cases of surgical peritonitis

The aim of this study was to determine the sensitivity of detecting microbial deoxyribonucleic acid [DNA] of E-coli using polymerase chain reaction [PCR] technique to detect it in the blood as a measure for diagnosing systemic infection and/or translocation from the gut in cases of localized peritonitis due to perforated appendicitis. Fifty patients with localized peritonitis due to perforated appendix were included in this study. The results have shown that the sensitivity of E-coli detection using the 16S rRNA and 16S rRNA+ primer set was 94% and that BG-1 and BG-4 primer set was 86%, while using either of the two sets increased the sensitivity to reach 98%. It was concluded that PCR is a highly sensitive technique which proved to be superior to the conventional blood culture methods in the detection of E-coli and can be used repeatedly to test the blood of patients for the presence of microbial DNA which could be originated from the gut