Ras Association Domain Family 1A Gene Promoter Methylation as a Biomarker for Chronic Viral Hepatitis C-related Hepatocellular Carcinoma.

Background One of the most prevalent aberrant epigenetic modifications found in hepatocellular carcinoma (HCC) is abnormal DNA methylation. Our study aimed to evaluate serum Ras association domain family 1A (RASSF1A) gene promoter methylation in patients with chronic viral hepatitis C (HCV)-associated liver cirrhosis with and without HCC as a potential new marker for the early detection of HCC. Methodology The 60 participants who participated in the trial were divided into the following three groups: 20 patients with newly diagnosed primary HCC on top of HCV-related liver cirrhosis, 20 patients with HCV-related liver cirrhosis, and 20 age- and sex-matched healthy individuals as a control group. All participants underwent methylation-specific polymerase chain reaction testing to detect the blood level of the RASSF1A gene's methylated promoter. Results Methylated RASSF1A was found in 30% of patients with liver cirrhosis caused by HCV and in 65% of patients with HCC, but not in any of the controls. It was discovered that the serum methylation RASSF1A had an accuracy of 82.50% and an area under the curve (AUC) of 0.825 for separating HCC patients from healthy controls. With an AUC of 0.675 and an accuracy of 67.50%, it was able to differentiate patients with HCC from those with HCV-related liver cirrhosis. Additionally, there was no statistically significant association between RASSF1A methylation status and HCC mass size (p = 0.449). Conclusions Serum RASSF1A promoter methylation status detection could be useful for detecting HCC early, especially in high-risk individuals such as those with HCV.

HIRA directly targets the enhancers of selected cardiac transcription factors during in vitro differentiation of mouse embryonic stem cells.

HIRA is a histone chaperone known to modulate gene expression through the deposition of H3.3. Conditional knockout of Hira in embryonic mouse hearts leads to cardiac septal defects. Loss of function mutation in HIRA, together with other chromatin modifiers, was found in patients with congenital heart diseases. However, the effects of HIRA on gene expression at earlier stages of cardiogenic mesoderm differentiation have not yet been studied. Differentiation of mouse embryonic stem cells (mESCs) towards cardiomyocytes mimics some of these early events and is an accepted model of these early stages. We performed RNA-Seq and H3.3-HA ChIP-seq on both WT and Hira-null mESCs and early cardiomyocyte progenitors of both genotypes. Analysis of RNA-seq data showed differential down regulation of cardiovascular development-related genes in Hira-null cardiomyocytes compared to WT cardiomyocytes. We found HIRA-dependent H3.3 deposition at these genes. In particular, we observed that HIRA influenced directly the expression of the transcription factors Gata6, Meis1 and Tbx2, essential for cardiac septation, through H3.3 deposition. We therefore identified new direct targets of HIRA during cardiac differentiation.

Relation of middle molecules levels and oxidative stress to erythropoietin requirements in high-flux versus low-flux hemodialysis.

The objective of this study is to investigate the serum beta-2-microglobulin (B2MG) and advanced oxidation protein products (AOPP) as middle molecule uremic toxins and protein carbonyl (PCO) as oxidative stress marker in uremic patients undergoing high-flux versus low-flux hemodialysis (HD) and to correlate their levels to the erythropoietin requirements for those patients. Twenty patients on chronic low-flux HD were recruited in the study. At the start of the study, all patients underwent high-flux HD for eight weeks, followed by low-flux HD for two weeks as a washout period. The patients were then subjected to another eight weeks of low-flux HD. Blood samples were obtained at the beginning and at the end of the high-flux period and the low-flux period. The mean erythropoietin dose for patients using high-flux HD was significantly lower than that for low-flux HD (P = 0.0062). Post-high flux, the B2MG and PCO levels were significantly lower than the pre-high-flux levels (P = 0.026 and 0.0005, respectively), but no significant change was observed in AOPP (P = 0.68). Post-low flux, the B2MG, AOPP and PCO were significantly higher than the pre-low-flux levels (P = 0.0002, 0.021 and <0.0001, respectively). Post-low flux, the B2MG and PCO were significantly higher than the post-high-flux levels (P <0.0001), but no significant difference was observed in AOPP (P = 0.11). High-flux HD results in reduction of some of the middle molecule toxins and PCO levels better than low-flux HD, and is associated with a better response to erythropoietin.

Methylation status of the gene promoter of cyclin-dependent kinase inhibitor 2A (CDKN2A) in ovarian cancer.

OBJECTIVE: This study investigated the methylation status of the promoter of the tumor suppressor gene cyclin-dependent kinase inhibitor 2A (CDKN2A) in ovarian cancer. DESIGN AND METHODS: CDKN2A methylation was quantified by real-time PCR in ovarian biopsies of 52 patients with ovarian cancer, 43 patients with benign ovarian tumors and 40 patients with benign uterine pathology and healthy ovaries. RESULTS: CDKN2A methylation was detected in the three groups. The methylation level was higher in the cancer patients than in the other two groups (p = 0.0003) but was not different between benign tumors and healthy ovarian tissue (p > 0.05). Using a cutoff threshold based on receiver-operating characteristic analysis, 21 patients with ovarian cancer and three patients with benign tumors were considered positive for CDKN2A methylation while all patients with healthy ovaries were considered negative. At the chosen cutoff, the diagnostic sensitivity was 40.4% and specificity 96.4%. CDKN2A methylation level and frequency were associated with high grade tumors (p = 0.0001 and p = 0.0005) but were not associated with disease stage or serum CA125 levels. However, it should be noted that most patients (92.3%) presented with advanced stage 3 or 4 disease. CONCLUSION: CDKN2A promoter methylation is common in ovarian cancer. Quantification of CDKN2A methylation may be useful in distinguishing malignant from benign ovarian tumors or healthy ovarian tissue.

G2019S mutation of the leucine-rich repeat kinase 2 gene in a cohort of Egyptian patients with Parkinson's disease.

AIM: This work investigates the prevalence of G2019S mutation of the leucine-rich repeat kinase 2 (LRRK2) gene in a cohort of Egyptian patients with sporadic Parkinson's disease (PD) and its relation to various features of the disease. MATERIALS AND METHODS: The study included 113 patients with sporadic PD and 87 healthy individuals as a control group. Clinical assessment was done using the Unified PD Rating Scale (UPDRS) and staging of PD was done according to Hoehn-Yahr score. The G2019S mutation was detected by polymerase chain reaction (PCR) followed by restriction digestion; results were confirmed using a 5' nuclease allelic discrimination real-time PCR method. RESULTS: The G2019S mutation was detected in 11 patients (9.7%) with PD, all of whom were heterozygous, but it was not present in any of the controls. Among PD patients, carriers of the G2019S mutation had significantly higher UPDRS motor score and a higher score for resting tremor than noncarriers (p=0.019 and p=0.004, respectively). CONCLUSIONS: The G2019S mutation in the LRRK2 gene is quite common in Egyptian patients with sporadic PD. The mutation is associated with a higher degree of motor effect but does not seem to affect mentation or behavioral aspects of the disease.

Toll-like receptor 3 gene expression in Egyptian patients with glomerulonephritis and hepatitis C virus infection.

BACKGROUND: Hepatitis C virus (HCV) infection is associated with both chronic liver disorders and several extrahepatic manifestations including renal disease. Toll-like receptor 3 (TLR3), a component of the innate immune system, is a pathogen recognition receptor that recognizes viral double-stranded RNA. OBJECTIVE: This work investigated TLR3 expression in peripheral blood mononuclear cells from HCV-positive patients with glomerulonephritis. METHODS: One hundred and thirty patients with glomerulonephritis were initially enrolled in the study. After exclusion of 52 patients with secondary glomerulonephritis, 78 patients were screened for HCV infection. TLR3 expression in peripheral blood mononuclear cells was studied in 46 patients with HCV-positive glomerulonephritis and 32 patients with HCV-negative glomerulonephritis using a real-time PCR comparative quantitation approach and results were compared to a control group of 20 healthy subjects. RESULTS: TLR3 expression was significantly higher in patients with HCV-positive glomerulonephritis than in HCV negative patients and controls (p < 0.0001). TLR3 expression correlated positively with HCV viral load, interleukin-1ß, serum creatinine and inversely with creatinine clearance in patients with HCV-positive glomerulonephritis. CONCLUSION: This work shows that TLR3 expression is upregulated in HCV-positive patients with glomerulonephritis. Overexpression is associated with reduced renal function and increased interleukin 1ß level.

MEFV gene mutations in Egyptian patients with familial Mediterranean fever.

BACKGROUND: Familial Mediterranean fever (FMF) is an autosomal recessive disorder characterized by recurrent attacks of fever and serositis. The disease affects mainly Mediterranean populations and is caused by mutations in the MEFV gene. AIM: This work was carried out to identify and determine the frequencies of MEFV gene mutations in Egyptian patients in whom FMF was diagnosed. METHODS: We investigated 316 patients with a clinical diagnosis of FMF for 12 MEFV mutations including the 5 most common known mutations M694V, V726A, M694I, M680I, and E148Q by allele-specific hybridization. RESULTS: Mutations were detected in 182 (57.6%) patients: 20 were homozygous, 80 were compound heterozygous, and 82 had only one identifiable mutant allele. In patients with clinically definite FMF (n = 112), no mutations were detected in 28 patients; whereas in patients with clinically unlikely FMF (n = 48), genetic analysis established the diagnosis in 6 patients. Overall, 10 mutations were detected in our patients. The most common were M694I (34%), E148Q (22.7%), V726A (15.6%), M680I (12.1%), and M694V (7.8%). M694V was observed in severe disease and in patients with amyloidosis. CONCLUSION: We were able to identify a wide spectrum of MEFV mutations in Egyptian patients in whom FMF was diagnosed. Frequencies of individual mutations showed some differences from those in other Mediterranean populations.

Clinical usefulness of basic fibroblast growth factor and E-selectin in patients with rheumatoid arthritis.

Angiogenesis plays an important role in the pathogenesis of rheumatoid arthritis (RA). This study assesses basic fibroblast growth factor (bFGF) as an angiogenic factor and soluble E-selectin (sE-sel) as an angiogenic mediator in RA patients and correlates their levels in serum and synovial fluid (SF) with disease activity, functional status and joint derangement. Thirty RA patients and 15 osteoarthritis (OA) patients were clinically, radiologically and laboratory investigated. bFGF, sE-sel, interleukin -1 beta (IL-1beta) and IL-6 in serum of patients and 15 healthy subjects and in SF tapped from knee joints of 9 RA and 9 OA patients were measured by ELISA. Both serum bFGF and sE-sel were significantly elevated in RA compared to OA patients and controls. These levels correlated positively with functional class stages of the disease. SF levels of bFGF and sE-sel showed significant increase in RA than OA patients. Both levels showed positive correlation with each other and with disease functional stages. A positive correlation was also found between SF bFGF with grades of joint derangement assessed radiologically. No significant correlations were observed between bFGF or sE-sel and clinical parameters of disease activity or other biochemical markers. In conclusion, serum and SF b-FGF and sE-sel may be considered as makers of functional status of RA patients. SF bFGF seems to reflect progressive joint derangement.