LncRNAs NEAT1, HOTAIR, and GAS5 expression in hypertensive and non-hypertensive associated cerebrovascular stroke patients, and its link to clinical characteristics and severity score of the disease.

Background: Cerebrovascular stroke (CVS) is a potentially fatal disease. The most common risk factor for CVS is hypertension. Aim: While most studies in the field have focused on the functional roles of long noncoding RNAs (lncRNAs) NEAT1, GAS5, and HOTAIR in CVS, less attention has been paid to their clinical relevance to stroke incidence and prognosis. Also, a link has not yet been made between these lncRNAs and hypertension, our study aim was to investigate whether the expression of these lncRNAs differed between CVS with and without hypertension, as well as to compare each group to controls. Method: In total, 181 CVS patients were enrolled, including 91 chronic hypertensive patients with stroke, 90 stroke patients without hypertension, and 51 control subjects. blood samples were collected on the day of recruitment from patients with CVS and controls. Real-time qRT-PCR was used to detect the expression of target lncRNAs in serum. Results: When compared to controls, there was a statistically higher level of lncNEAT1 in each case group (median (IQR) = 3.68 (1.35-7.35) and 3.05 (0.95-6.45) for the hypertensive and non-hypertensive groups, respectively, with a significantly higher level in the hypertensive group (P = 0.04). When compared to controls, lncHOTAIR was significantly downregulated in all case groups (medians in hypertensive and non-hypertensive patients were 0.13, and 0.34, respectively), with a significantly lower level in the hypertensive group (P = 0.05). LncGAS5 levels in patients were significantly lower (median (IQR) = 0.16 (0.02-0.55) and 0.25 (0.03-0.99) for the hypertensive and non-hypertensive groups, respectively) compared to controls, with a significantly lower level in the hypertensive group (P = 0.02). There was a significant positive correlation between NEAT1 and GAS5, but a significant negative correlation between each with HOTAIR in both patients' groups. We also detected a significant negative correlation between each NEAT1 or GAS5 and NIHSS score while a significant positive correlation between HOTAIR and NIHSS. ROC curve analysis for GAS5 was able to differentiate patients with CVS hypertensive from patients with CVS non-hypertensive. Conclusion: Patients in each case group had statistically higher levels of NEAT1 and lower levels of HOTAIR and GAS5 compared to control levels, with higher significant NEAT1 but lower significant HOTAIR and GAS5 in the hypertensive group. Therefore, lncRNAs NEAT1, HOTAIR, and GAS5 could be used as diagnostic and prognostic biomarkers of CVS that correlate with NIHSS score and could produce a novel target for CVS therapy.

Study of the role of microRNAs 16 and 135a in patients with lifelong premature ejaculation receiving fluoxetine daily for 3 months: A prospective case control study.

We aimed to determine the level of miRNAs 16 and 135a in lifelong premature ejaculation (LPE) patients versus controls. Moreover, we evaluated the potential interplay between the studied miRNAs and fluoxetine in these patients after utilizing fluoxetine daily for 3 months. The study involved 60 consecutive LPE patients and 20 healthy age matched individuals as controls. The median miRNA16 was significantly higher in the controls (1.02) compared to the patients (0.31) (p < 0.001). Moreover, the median miRNA-135a was significantly higher in the controls compared to the patients 1.02 and 0.35, p < 0.001, respectively. In addition, the median pre-treatment miRNA16 in the responders was 0.29 that significantly increased to 0.66 (p < 0.001). The median pre-treatment miRNA-135a in the responders was 0.27 that significantly increased to 0.65 (p < 0.001). Furthermore, considering EXP(ß) for the odds ratio evaluation, with a 95% degree of confidence, a 1 fold increase in pre-treatment miRNA 135a fold change decreases the odds for being responsive to SSRI by 0.028. Meanwhile, there was non-significant association between fluoxetine responsiveness and age, pre-treatment miRNA 16, pre-treatment PEDT and pre-treatment IELT. The current study had shown that a lower pre-treatment miRNA 135a was significantly associated with response to fluoxetine.

Genetic signature of CTLA-4, BTLA, TIM-3 and LAG-3 molecular expression in colorectal cancer patients: Implications in diagnosis and survival outcomes.

OBJECTIVE: Accumulating evidences suggest that immune checkpoints (ICs) inhibit immune response against cancerous cells and promote tumor cell survival. Up-regulation of ICs in tumor microenvironment is reported in patients with colorectal cancer (CRC). Thus, evaluating the peripheral blood expression of ICs may be used as non-invasive biomarkers for diagnosis and prognosis of CRC. METHODS: This study included 60 primary and treatment naïve CRC patients along with 15 age and sex matched healthy volunteers as a control group. Total RNA was extracted from peripheral blood samples and gene expression of cytotoxic T lymphocyte antigen-4 (CTLA-4), B and T lymphocyte attenuator (BTLA), T-cell immunoglobulin and mucin domain-containing protein 3 (TIM-3), and Lymphocyte activation gene-3 (LAG-3) was measured by quantitative real time polymerase chain reaction (qRT-PCR). All patients were followed for 12 months to correlate the measured ICs to patients' survival. RESULTS: The gene expression of CTLA-4, BTLA, TIM-3 and LAG-3 was significantly up-regulated in CRC patients compared to the control group (p < 0.001). Individually, CTLA-4 and BTLA showed 85% sensitivity in discriminating CRC patients from control group (p < 0.001). On the other hand, TIM-3 and LAG-3 expression showed higher sensitivity (93%) for diagnosis of CRC (p < 0.001). Conversely, CTLA-4 or BTLA strongly predicted CRC patients' survival (p < 0.001) compared to TIM-3 (p = 0.018) or LAG-3 (p = 0.035). CTLA-4, BTLA, TIM-3 and LAG-3 were independent prognostic factors of survival after adjustment for age and gender. CONCLUSION: The current study provided evidence that blood gene expression of ICs was up-regulated in CRC patients and associated with cancer stage and patients' survival, which highlights the diagnostic and prognostic values of ICs expression in CRC. Further investigations and validations in larger cohorts are required.

Association between TNF-α, Interleukin-18 Polymorphisms and Risk of Hepatocellular Carcinoma in Egyptian patients.

OBJECTIVE: To evaluate the association of gene polymorphisms of the SNP of TNF-α gene -238G>A and IL-18 gene-607C>A with the development of hepatocellular carcinoma among Egyptian patients. METHODS: One hundred and fifty patients were allocated to this study; eighty patients with hepatocellular carcinoma (Group A), seventy cancer-free HCV age, and sex-matched patients (Group B). We analyzed two Single nucleotide polymorphisms (SNPs) (TNF-α-238G>A and IL-18-607C>A) by real-time polymerase chain reaction using sequence-specific primers (PCR-SSP). RESULTS: Significant higher risk of HCC was associated with genotype IL-18-607AA (p <0.001), OR: 5(2.188-11.47), allele IL-18 -607/A (P=0.001), OR: 2.1(1.32-3.3). A significant association was found between the size of HFL in the HCC group and different genotypes of IL18 genes (P=0.013) where 62.5% of patients with tumor size >5 cm carried the risky (AA) genotype on the other hand the SNP of TNF-α gene -238G>A showed no statistically significant association between the two groups. CONCLUSION: The SNP -607C>A in the IL18 gene was associated with increased HCC risk in Egyptian patients suggesting its use as a potential diagnostic non-invasive tool that allows to identify a new group of HCC patients at an earlier stage.
.

Urinary pigment epithelium-derived factor as a marker of diabetic nephropathy in Egyptian patients with type 2 diabetes mellitus.

Diabetic nephropathy (DN) is the most common cause of end-stage renal disease. Urinary pigment epithelium-derived factor (PEDF) has also been shown to suppress the expression of fibrogenic, pro-inflammatory, and angiogenic factors, thus contributing to pathological changes in early DN. We aimed to study the role of urinary PEDF as a biomarker for the detection of chronic kidney disease progression in patients with type 2 diabetes mellitus (T2DM). Sixty patients with T2DM were recruited in addition to 20 nondiabetic healthy volunteers. Urinary PEDF using enzyme-linked immunoassay technique was performed to all subjects, and correlations between it and different clinical parameters were examined. Our study showed a statistically significant correlation between urinary PEDF level and duration of DM (P <0.001), glycosylated hemoglobin (P <0.001), serum creatinine (P <0.001), urinary albumin-to-creatinine ratio (P <0.001), and stage of diabetic retinopathy by fundus examination (P <0.001). Urinary PEDF is a good indicator of progression of DN and microvascular damage (as a complication of diabetes) in general. It was also increased in case of poor diabetic control.

Expression of miRNAs-122, -192 and -499 in end stage renal disease associated with acute myocardial infarction.

INTRODUCTION: New diagnostic tools are needed to accurately detect acute myocardial infarction (AMI) in patients with end stage renal disease (ESRD) presenting with ischemic chest pain. We aimed in this study to investigate circulating miR-122, -192 and -499 expression levels in patients with AMI on top of ESRD and evaluate the potential of these miRNAs as blood-based biomarkers for AMI in patients with ESRD. MATERIAL AND METHODS: The study included 80 ESRD patients without AMI, 80 patients with ESRD associated with AMI and 60 healthy subjects. Assessment of microRNAs was done using SYBR Green based real-time PCR. RESULTS: Levels of miR-122 were 28-fold and 20-fold higher in controls than in ESRD patients with or without AMI respectively (p < 0.001), while no differences were detected between the two patient groups (p = 0.9). Levels of miR-192 showed a marked increase in ESRD patients with and without AMI compared to the control group (> 500-fold, > 8000-fold respectively, p ≤ 0.001). Patients who developed AMI had lower expression than ESRD patients without AMI (p < 0.001). Non-significant miR-499 elevation was found in ESRD patients without cardiac disease compared to the control group, while highly significant elevation of miR- 499 was demonstrated in ESRD patients who developed AMI compared to other ESRD patients and the control group (> 100-fold, > 350-fold respectively, p = 0.001). CONCLUSIONS: Altered expression of miR-122 and -192 may contribute in pathogenesis of ESRD. MiR-192 and -499 may serve as potential biomarkers for AMI in ESRD. Further studies are needed to correlate these miRNAs with disease progression and outcome.

Serum Soluble Interleukin 2 Receptor Level as a Marker of Acute Rejection in Pediatric Kidney Transplant Recipients.

BACKGROUND: Despite advances in immunosuppression, acute allograft rejection remains one of the key factors affecting patient and graft survival in pediatric kidney transplantation. The aim of the study is to evaluate the role of serum soluble IL2 receptor (sIL2R) level as a noninvasive assessment parameter of acute rejection (AR). METHOD: Serum sIL2R level was measured (using enzyme-linked immune-sorbent assay technique) in 60 pediatric kidney transplant recipients (30 recipients with AR and 30 transplant recipients with stable graft function). RESULTS: The mean values of sIL2R level in patients experiencing AR (14.8 ± 6.54) ng/mL were significantly higher than that in patients with stable graft functions (6.44 ±1.95) ng/mL (p = 0.0001). In addition; patients with AR proved by graft biopsy had their mean values of sIL2R level (16.19 ± 7.48) ng/mL significantly higher than that of other recipients in the study population (p = 0.032). CONCLUSION: Serum sIL2R level may serve as a noninvasive diagnostic indicator in pediatric kidney transplant recipients experiencing AR.

Genotypic Identification of Cystoisospora in Immunocompromised Patients Using Tm-Variation Analysis.

Cystoisospora is responsible for morbidity in immunocompromised patients. PCR is sensitive for diagnosing Cystoisospora; however, it needs reevaluation for differential molecular diagnosis of cystoisosporiasis. We aimed at evaluating melting curve analysis (MCA) after real-time PCR (qPCR) in diagnosis and genotyping of Cystoisospora as an alternative to conventional PCR. We included 293 diarrheic stool samples of patients attending the Department of Clinical Oncology and Nuclear Medicine of Cairo University Hospitals, Egypt. Samples were subjected to microscopy, nested PCR (nPCR), and qPCR targeting the internal transcribed spacer 2 region (ITS2) of the ribosomal RNA (r RNA) gene followed by melting temperatures (Tms) analysis and comparing the results to PCR-RFLP banding patterns. Using microscopy and ITS2-nPCR, 3.1% and 5.8% of cases were Cystoisospora positive, respectively, while 10.9% were positive using qPCR. Genotyping of Cystoisospora by qPCR-MCA revealed 2 genotypes. These genotypes matched with 2 distinct melting peaks with specified Tms at 85.8°C and 88.6°C, which indicated genetic variation among Cystoisospora isolates in Egypt. Genotype II proved to be more prevalent (65.6%). HIV-related Kaposi sarcoma and leukemic patients harbored both genotypes with a tendency to genotype II. Genotype I was more prevalent in lymphomas and mammary gland tumors while colorectal and hepatocellular tumors harbored genotype II suggesting that this genotype might be responsible for the development of cystoisosporiasis in immunocompromised patients. Direct reliable identification and differentiation of Cystoisospora species could be established using qPCR-Tms analysis which is useful for rapid detection and screening of Cystoisospora genotypes principally in high risk groups.

Aberrant p16INK4A methylation: Relation to viral related chronic liver disease and hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is currently the fifth most common solid tumor worldwide and the third leading cause of cancer related deaths. Several studies have shown that the tumor suppressor gene p16INK4A is frequently downregulated by aberrant methylation of the 5'-cytosine-phosphoguanine island within the promoter region. AIM: To find out the frequency of methylated p16INK4A in the peripheral blood of HCC and cirrhotic patients and to evaluate its role in hepatocarcinogenesis. PATIENTS AND METHODS: This study was performed on 58 subjects: 30 HCC patients, 20 cirrhotic patients, and eight healthy volunteers. Methylation of p16INK4A was examined using methylation specific polymerase chain reaction (PCR) (MSP). Comparison of quantitative variables between the study groups was done using Mann-Whitney U test for independent samples when not normally distributed. For comparing categorical data, Chi-square (χ(2)) test was performed. Exact test was used instead when the expected frequency was less than 5. RESULTS: Methylation of p16INK4A was found in 6.7% of HCC patients, 5% of liver cirrhosis (LC) patients, and none of the healthy volunteers; 66.67% of the p16INK4A-methylated cases (2/3) were positive for anti-hepatitis C virus (HCV) antibodies (one of them had HCC). All HCC cases with aberrant p16INK4A methylation show very high serum alpha fetoprotein (AFP) level (9,080; 30,000 µg/mL). There were no significant associations between the status of p16INK4A methylation and tumor size. CONCLUSION: Hypermethylation of p16INK4A was found to be infrequent among Egyptian patients with HCC.