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.

The potential DNA toxic changes among workers exposed to antimony trioxide.

Occupational exposure to antimony has gained much interest when specific toxic effects were noticed among workers processing antimony. Thus, the aim of the present work was to investigate the potential DNA oxidative damage occurring among Egyptian workers occupationally exposed to antimony trioxide. The study was conducted on 25 subjects exposed to antimony trioxide while working in the polymerization process of polyester in Misrayon and Polyester Fiber Company, KafrEldawwar, Beheira, Egypt. Urinary antimony levels were assessed using inductive coupled plasma-optical emission spectrometry (ICP-OES) and considered as a biological exposure index. DNA damage and total oxidant capacity (TOC) were assessed using ELISA. DNA damage was detected in the form of increased apurinic/apyrimidinic (AP) sites among antimony trioxide-exposed workers compared to control subjects, but it could not be explained by oxidative mechanisms due to lack of significant correlation between DNA damage and measured TOC. Antimony trioxide might have a genotoxic impact on occupationally exposed workers which could not be attributed to oxidative stress in the studied cases.

Study of the role of tumor necrosis factor-α (-308 G/A) and interleukin-10 (-1082 G/A) polymorphisms as potential risk factors to acute kidney injury in patients with severe sepsis using high-resolution melting curve analysis.

RATIONAL: Septic acute kidney injury (AKI) is a prevalent complication in intensive care units with an increased incidence of complications. OBJECTIVE: The aim of the present study was to assess the use of high-resolution melting curve (HRM) analysis in investigating whether the genetic polymorphisms; -308 G/A of tumor necrosis factor-α (TNF-α), and -1082 G /A of Interleukin-10 (IL-10) genes may predispose patients diagnosed with severe sepsis to the development of AKI. METHODS: One hundred and fifty patients with severe sepsis participated in the present study; only sixty-six developed AKI. Both polymorphisms were studied using HRM analysis. MAIN FINDINGS: The low producer genotype of both studied polymorphism of TNF-α and IL-10 genes was associated with AKI. Using logistic regression analysis, the low producer genotypes remained an independent risk factor for AKI. A statistically significant difference was detected between both studied groups as regards the low producer genotype in both TNF-α (-308 G/A) and interleukin-10 (IL-10) (-1082 G/A) polymorphisms being prevalent in patients developing AKI. Principle conclusions: The low producer genotypes of both TNF-α (-308 G/A) and IL-10 (-1082 G/A) polymorphisms could be considered a risk factor for the development of AKI in critically ill patients with severe sepsis, thus management technique implemented for this category should be modulated rescuing this sector of patients from the grave deterioration to acute kidney injury. Using HRM for genotyping proved to be a highly efficient, simple, cost-effective genotyping technique that is most appropriate for the routine study of large-scale samples.

Mitochondrial DNA Copy Number in Egyptian Patients with Hepatitis C Virus-Related Hepatocellular Carcinoma.

AIM: To assess the use of mitochondrial DNA (mtDNA) content as a noninvasive molecular biomarker in hepatitis C virus-related hepatocellular carcinoma (HCV-HCC). MATERIALS AND METHODS: A total of 135 participants were enrolled in the study. Equal numbers of subjects were enrolled in each of three clinically defined groups: those with HCV-related cirrhosis (HCV-cirrhosis), those with HCV-HCC, and a control group of age- and sex-matched healthy volunteers with no evidence of liver disease. mtDNA concentrations were determined using a quantitative real-time polymerase chain reaction (PCR) technique. RESULTS: mtDNA content was lowest among the HCV-HCC cases. No statistically significant difference was observed between the group of HCV-cirrhosis and the control group as regards mtDNA level. HCC patients with multicentric hepatic lesions had significantly lower mtDNA content than HCC patients with less advanced disease. When a receiver operating characteristic curve analysis was used, a cutoff of 34 was assigned for mtDNA content to distinguish between HCV-HCC and HCV-cirrhosis patients who are not yet complicated by malignancy. Lower mtDNA content was associated with HCC risk when using either or both healthy controls and HCV-cirrhosis groups for reference. CONCLUSIONS: mtDNA content analysis could serve as a noninvasive molecular biomarker that reflects tumor burden in HCV-HCC cases and could be used as a predictor of HCC risk in patients of HCV-cirrhosis. In addition, the nonsignificant difference of mtDNA level between HCV-cirrhosis patients and healthy controls could eliminate the gray zone created by the use of alpha-fetoprotein in some cirrhotic patients.

Correlation between waist:height ratio and serum nitric oxide level in children with atopic dermatitis.

Atopic dermatitis (AD) is the most common chronic inflammatory disease of early childhood. This study shows that nitric oxide levels positively correlate with the clinical severity of AD, waist:height ratio, and weight.

Occurrence of the polyomavirus among kidney transplant recipients: a single-center study.

Polyoma virus-associated nephropathy is an increasingly recognized cause of graft dysfunction among kidney transplant recipients and could be the result of use of potent immunosuppression following transplantation. Because there is no safe and effective anti-viral therapy available presently, screening-based prevention and pre-emptive strategy are recommended. This study, which was conducted at the Nephrology Unit, Internal Medicine Department, Alexandria University, consisted of two phases: Phase 1 was a cross-sectional study and phase 2 was a 6-month follow-up study only for polyoma virus-positive cases. Phase 1 included 75 renal allograft recipients from living donors. Urine cytology for decoy cells and quantitative real-time blood polymerase chain reaction (PCR) for the BK virus (BKV) were performed on all the study patients. Renal biopsy was performed only in patients with deteriorating renal function associated with positive urine cytology. Patients who showed positive urine cytology for decoy cells and/or positive quantitative BKV PCR assay were followed-up for six months. During follow-up, the serum creatinine level, with or without urine cytology for decoy cells, and BKV PCR viral load assay were performed. Among the 75 kidney transplant recipients studied, eight were positive for decoy cells (11%), three showed viremia by quantitative PCR for BKV (4.1%), while two others showed nephropathy (2.7%) in the form of tubulointerstitial nephritis with intra-nuclear inclusions in the tubular cells. Cases with stable renal function and positive decoy cells or viremia cleared the virus spontaneously during follow-up without any intervention. Only one case with biopsyproven nephropathy and deteriorating graft function, with undetectable BKV in blood, lost the graft while another case with viremia died during follow-up due to septicemia. Our study suggests that polyoma virus should be considered as a cause of nephropathy in renal transplant recipients. Further research is required to understand this entity better.

Evaluation of HE4 as an extrabiomarker to CA125 to improve detection of ovarian carcinoma: is it time for a step forward?

PURPOSE: To evaluate human epididymis protein 4 (HE4) as an extrabiomarker to cancer antigen 125 (CA125) to improve the detection of ovarian carcinoma. METHODS: Sixty patients with ovarian carcinoma, 50 patients with benign ovarian tumors and 30 healthy women were included in the present study. Serum concentration of HE4 was assayed using ELISA technique, while CA125 was assayed using chemiluminescent enzyme immunoassay. RESULTS: The median CA125 and HE4 serum values were significantly higher among ovarian cancer patients when compared with healthy control However, the median serum levels of CA125 but not HE4 were significantly higher among patients with benign ovarian tumors as compared to healthy women. Based on the receiver operator characteristics curve analysis, HE4 had higher sensitivities than CA125 for the detection of ovarian cancer at 90, 95 and 98 % specificities and the combination of both markers yielded a higher sensitivity than either alone. However, CA125 but not HE4 had higher sensitivities for the detection of benign ovarian tumors at the same specificities. In addition, a positive correlation was observed between HE4 and CA125 among patients with ovarian carcinoma. CONCLUSION: HE4 is a valuable marker for ovarian cancer diagnosis and when combined with CA125, they had a higher sensitivity at a set specificity, thus providing a more accurate predictor of ovarian cancer than either alone.

Toll like receptor 3 expression as a novel predictor of response to treatment in chronic hepatitis C virus patients.

PURPOSE: This work was carried out to study the level of pretreatment hepatic expression of Toll like receptor 3 (TLR3) among chronic HCV patients, aiming to determine if there are consistent differences in gene expression, between those who show complete early virological response (cEVR) at week-12 of Pegylated-Interferon α-2a plus ribavirin treatment and others who are not responding to this combination, also if this could be used to predict treatment outcomes. METHOD: A total of 61 chronic hepatitis C patients were enrolled in the study. For all of them, baseline hepatitis C virus (HCV) viral load was determined and TLR3 gene expression was examined in their hepatic percutaneous needle biopsy specimens using a real time-polymerase chain reaction technique. Hepatic TLR3 was also traced using immunohistochemistry. All patients followed a 12-week regimen of Pegylated-Interferon α-2a plus ribavirin then post-treatment viral load was assayed. RESULTS: Hepatic expression of TLR3 was significantly increased in the non-responder group as compared to those who showed complete early virological response to the used treatment regimen. Using receiver operator characteristic (ROC) curve analysis, a cutoff level of 1.5 was set for TLR3 expression to distinguish responders from non-responders to the 12-week treatment regimen used. CONCLUSION: In conclusion, measuring the hepatic expression of the interferon stimulated gene, TLR3 could provide a new molecular marker for pre-treatment prediction of complete early viral clearance, thus helping in patients' selection and optimizing treatment outcomes.

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.

Role of glutathione-S-transferase P1 hypermethylation in molecular detection of prostate cancer.

AIM: The aim of this study was to evaluate the diagnostic potential of the quantification of glutathione-S-transferase P1 (GSTP1) gene hypermethylation in molecular detection of prostate cancer in tissue biopsies. METHODS: One hundred fourteen male patients were enrolled in the study; 44 patients with histopathologically confirmed prostate adenocarcinoma, 20 patients with variable degrees of prostate intraepithelial neoplasia, and 50 with benign prostatic hyperplasia, who served as a control group. Real-time quantitative methylation-specific polymerase chain reaction was used for assessment of methylation of the promoter region of the GSTP1 gene. RESULTS: Methylation of the GSTP1 promotor was detected in 24% of patients with benign prostatic hyperplasia, 60% of patients with prostate intraepithelial neoplasia, and in 86.3% of prostate adenocarcinoma patients. A statistically significant difference in the GSTP1/MYOD1 (myogenic differentiation 1gene) methylation ratios among the three groups was observed (p=0.0001). At the cutoff value of 9, GSTP1/MYOD1 methylation ratios showed sensitivity in the detection of prostate adenocarcinoma of 71.8% and specificity of 96%. CONCLUSIONS: Methylation of the GSTP1 promotor is a common molecular alteration in prostate cancer that may be a useful adjunct to serum screening tests and digital rectal examination findings and the use of quantitative real-time methylation-specific polymerase chain reaction is a promising technique that often distinguishes malignant from nonmalignant prostate disease.

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.