High-depth African genomes inform human migration and health.

The African continent is regarded as the cradle of modern humans and African genomes contain more genetic variation than those from any other continent, yet only a fraction of the genetic diversity among African individuals has been surveyed1. Here we performed whole-genome sequencing analyses of 426 individuals-comprising 50 ethnolinguistic groups, including previously unsampled populations-to explore the breadth of genomic diversity across Africa. We uncovered more than 3 million previously undescribed variants, most of which were found among individuals from newly sampled ethnolinguistic groups, as well as 62 previously unreported loci that are under strong selection, which were predominantly found in genes that are involved in viral immunity, DNA repair and metabolism. We observed complex patterns of ancestral admixture and putative-damaging and novel variation, both within and between populations, alongside evidence that Zambia was a likely intermediate site along the routes of expansion of Bantu-speaking populations. Pathogenic variants in genes that are currently characterized as medically relevant were uncommon-but in other genes, variants denoted as 'likely pathogenic' in the ClinVar database were commonly observed. Collectively, these findings refine our current understanding of continental migration, identify gene flow and the response to human disease as strong drivers of genome-level population variation, and underscore the scientific imperative for a broader characterization of the genomic diversity of African individuals to understand human ancestry and improve health.

Behavioral effects of toll-like receptor-4 antagonist 'eritoran' in an experimental model of depression: role of prefrontal and hippocampal neurogenesis and γ-aminobutyric acid/glutamate balance.

Depression is the disease of the modern era. The lack of response to the available antidepressants, which were developed on the basis of the monoaminergic deficit hypothesis of depression, has encouraged scientists to think about new mechanisms explaining the pathogenesis of depression. In this context, the inflammatory theory has emerged to clarify many aspects of depression that the previous theories have failed to explain. Toll-like receptor-4 (TLR-4) has a regulatory role in the brain's immune response to stress, and its activation is suggested to play a pivotal role in the pathophysiology of depression. In this study, we tested eritoran (ERI), a TLR-4 receptor-4 antagonist, as a potential antidepressant. We investigated the effect of long-term administration of ERI in three different doses on behavioral changes, hippocampal and prefrontal cortex (PFC) neurogenesis, and γ-aminobutyric acid (GABA)/glutamate balance in male Wistar rats exposed to chronic restraint stress (CRS). Long-term administration of ERI ameliorated CRS-induced depressive-like symptoms and hypothalamic-pituitary-adrenal axis hyperactivity alongside reducing levels of hippocampal and PFC inflammatory cytokines, restoring GABA and glutamate balance, and enhancing PFC and hippocampal neurogenesis, by increasing BDNF gene and protein expression in a dose-dependent manner. The results demonstrate an antidepressant-like activity of ERI in Wistar rats exposed to CRS, which may be largely mediated by its ability to reduce neuroinflammation, increase BDNF, and restore GABA/glutamate balance in prefrontal cortex and hippocampus. Nonetheless, further studies are needed to characterize the mechanism of the antidepressant effect of ERI.

Laboratory validation of formal concept analysis of the methylation status of microarray-detected genes in primary breast cancer.

Breast cancer is the leading cause of cancer-related mortality. DNA methylations play important roles in cancer development and progression. Formal concept analysis was previously utilized for data mining hypermethylated and hypomethylated genes in breast cancer molecular subtypes in illumina methylation-based microarray database, to laboratory validate their outputs; HS3ST2 (heparan sulfate d-glucosaminyl 3-O-sulfonyl transferase-2) and MUC1 (mucin-1) were retrieved. Both play important roles in progression and invasion of breast cancer. The methylation status of both genes was laboratory validated using methylation-based polymerase chain reaction in breast cancer subtypes luminal A (early stages) and luminal B (late stages) in comparison with benign conditions and normal breast to conclude their roles in tumor invasion and to validate the newly developed algorithm (formal concept analysis). Significant cancer-specific hypermethylation of HS3ST2 was detected in luminal B (chi square = 30.6, p = 0.000), while significant cancer-specific hypomethylation of MUC1 was detected in luminal B (chi square = 30.5, p = 0.001) breast cancer. The median levels of the percentage of methylated allele of both genes were significantly discriminative between luminal A and luminal B subtypes and benign and healthy control groups. Detection of MUC1 and HS3ST2 promoter methylation status appears to be useful molecular markers for assessing the progressive state of the disease and could be helpful in discriminating breast cancer molecular subtypes. These results validate the methylation-based microarray analysis, thus trust their output in the future.

Host and viral determinants of the outcome of exposure to HCV infection genotype 4: a large longitudinal study.

OBJECTIVES: The objective of this study was to characterize the factors that influence the outcome of exposure to hepatitis C virus (HCV) genotype 4 (HCV-G4) and the course of recent infection. METHODS: In this longitudinal study, we prospectively assessed the clinical, genetic, virological, and immunological parameters and retrospectively determined single-nucleotide polymorphisms at interleukin-28B (IL-28B) rs12979860 in a well-characterized large cohort recently exposed to HCV-G4. RESULTS: A total of 136 subjects with acute HCV (new viremia, seroconversion, and HCV-specific T-cell responses) were identified. Forty-eight subjects (35%) had spontaneous viral clearance and 88 subjects developed chronic HCV of which 42 subjects were treated with pegylated interferon monotherapy, with a sustained virologic response (SVR) rate of 88%. Twenty-six subjects developed HCV-specific T-cell immune responses without detectable viremia or seroconversion. IL-28B-CC (odds ratio (OR) 14.22; P<0.0001), multispecific T-cell responses (OR=11.66; P<0.0001), >300 IU/l alanine aminotransferase (ALT) decline within 4 weeks (OR=6.83; P<0.0001), jaundice (OR=3.54; P=0.001), female gender (OR=2.39; P=0.007), and >2.5 log10 HCV-RNA drop within 8 weeks (OR=2.48; P=0.016) were independently associated with spontaneous clearance. ALT normalization and undetectable HCV-RNA predicted SVR. Exposed apparently uninfected participants had a higher frequency of IL-28B-CC than patients with unresolved acute HCV (P<0.001). IL-28B-CC was associated with multispecific T-cell response (r(2)=0.0.835; P<0.001). CONCLUSIONS: IL-28B-CC genotype, multispecific HCV T-cell responses, rapid decline in ALT, and viral load predict spontaneous clearance and response to acute HCV-G 4 therapy. IL-28B-CC genotype correlates with developing early multispecific T-cell responses. These findings have important implications for predicting the outcome of HCV exposure and acute infection and identifying patients likely to benefit from therapy.

Antiproliferative effect of hepatitis C virus on mitogen-stimulated peripheral blood mononuclear cells: potential role in viral persistence in Egyptian patients.

OBJECTIVES: We aimed to study the effect of hepatitis C virus (HCV) and sera of chronic HCV patients on phytohemagglutinin (PHA)-stimulated normal donor PBMCs and to study the effect of chronic HCV infection on some cytokine profile. SUBJECTS AND METHODS: (3)H-Thymidine uptake was utilized to study effect of pelleted virus and patients sera on PBMCs proliferation in vitro. The study included 337 Egyptian chronic liver patients from Ain Shams University Hospitals and 90 healthy control subjects. The patients' group included chronic hepatitis C (250 subjects), and other chronic liver diseases (87 subjects). All subjects' sera were subjected to RT-PCR for HCV RNA detection, IL-4, IL-1beta, and TNF-alpha measurement by EIA, and biochemical measurement of ALT and albumin. RESULTS: Treatment of PHA-stimulated normal donor PBMCs with pelleted virus led to decrease (dose response) in their rate of proliferation. This was partially reversed after addition of HCV RNA positive patients' sera. HCV RNA positive patients were significantly higher in IL-4 and ALT, and lower in IL-1beta and albumin than HCV RNA negative patients. CONCLUSION: HCV infection suppresses early immune response. This leads to increased IL-4 Th2 cytokine.

The role of interleukin-12, and tissue antioxidants in chronic sinusitis.

BACKGROUND: The role of cytokines and reactive oxygen species formation in the pathogenesis of chronic sinusitis is still not fully understood. This work was designed to determine if patients with chronic sinusitis demonstrate altered levels of serum IL-12 and/or tissue antioxidants. SUBJECTS AND METHODS: Mucosal biopsy specimens from the uncinate process of patients with chronic sinusitis were obtained from 52 patients using functional endoscopic sinus surgery. Normal mucosa samples were collected from 20 healthy controls. Patients' group was further classified according to computerized tomography findings into mild and severe subgroups. Serum IL-12 was estimated using enzyme immunoassay (EIA). The levels of tissue uric acid, and reduced glutathione were determined biochemically, alpha-tocopherol was measured by HPLC. Superoxide dismutase (SOD) activity was determined by spectrophotometry. RESULTS: A significant decrease in serum IL-12, tissue alpha-tocopherol and SOD in patient group was demonstrated (p < 0.05). Tissue uric acid and reduced glutathione showed primary increase in mild subgroup followed by significant drop in severe subgroup (p < 0.05). Negative significant correlation was observed between glutathione, uric acid, and SOD, and the severity of the disease (p < 0.05) independent of the cellularity of the biopsy. CONCLUSION: The presented data suggests a possible role of IL-12, and tissue antioxidants in development and progression of chronic sinusitis. Adjuvant antioxidant treatment may have role in achieving better prognosis of the disease.

Serum nitrates and vasoactive intestinal peptide in patients with gastroesophageal reflux disease.

BACKGROUND: The normal esophageal motility is a balance between excitatory cholinergic "muscarinic" innervations and noncholinergic nonadrenergic inhibitory innervations. The latter is mediated by nitric oxide (NO) and/or vasoactive intestinal polypeptide (VIP). METHODS: The study included 50 patients with gastro-esophageal reflux disease (GERD), and 10 healthy controls of matched age and sex. Patients were divided into five groups according to the degree of lower end esophagitis (Savary-Miller classification). Serum VIP was measured using enzyme immunoassay after peptide purification. Serum nitrate as an index of nitric oxide generation was determined biochemically. RESULTS: Serum nitrate and VIP levels were significantly higher in GERD patients than the control group (p < 0.001). Grade 0 serum nitrates was significantly higher than the control group (p < 0.05) with some overlap between the individual values of the two groups. Serum VIP was significantly higher in grade 0 group compared to control group (p < 0.001) with no overlap in the individual values. There was a significant positive correlation between the grade of lower end esophagitis and each of serum nitrate and VIP (p < 0.001), as well as between serum nitrate and each of serum VIP, cigarette smoking index (CSI) and BMI (p < 0.001). CONCLUSION: Abnormally high levels of serum VIP and NO may have a role in the pathogenesis of GERD. Exposure of esophageal mucosa to noxious effect of acid refluxed due to the relaxant effect of VIP on lower esophageal sphincter may cause increased NO levels. BMI and CSI are risk factors for GERD progression.

Vascular endothelial growth factor and interleukin-8 are associated with poor prognosis in epithelial ovarian cancer patients.

BACKGROUND: Ovarian cancer represents an important problem in gynecologic oncology. A growing tumor induces the host endothelial cells to proliferate and supply the requisite vascular support allowing tumor development. Vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8) have been demonstrated to induce angiogenesis in epithelial tumors in vivo. PATIENTS AND METHODS: This study included 24 tumors from patients with epithelial ovarian cancer in different stages, in addition to 20 tissue samples of benign ovarian lesions as a control group. VEGF has been measured in the cytosolic fractions using enzyme immunoassay and confirmed by Western blot analysis. Tissue IL-8 mRNA was assessed using reverse transcriptase polymerase chain reaction and immunohistochemistry for its protein. RESULTS: VEGF mean rank was significantly higher in ovarian cancer tumors compared to benign lesions (P < 0.001). Moreover, it was increased with advanced stages (P < 0.05) and in patients with poor survival (P < 0.05). Eight samples were positive for IL-8 mRNA, seven of them were in malignant group, with highest frequency in stages III and IV of the disease (6/12, 50%). IL-8 correlated with poor survival of the patients (P < 0.05). Log rank of Kaplan-Meier survival analysis was significant for FIGO stage, VEGF, and IL-8 (P < 0.05). CONCLUSION: These results indicate that VEGF and IL-8 are related to the malignant transformation process and can be considered as indicators of poor prognosis in epithelial ovarian cancer patients.

Lipopolysaccharide repeated challenge followed by chronic mild stress protocol introduces a combined model of depression in rats: reversibility by imipramine and pentoxifylline.

OBJECTIVES: The present study examined the effect of combined exposure to repeated challenge using low doses of lipopolysaccharide (LPS) and chronic mild stress (CMS) together. This combined exposure is thought to expose the animals to more realistic challenges, testable on different levels (behavioral, neurochemical, immunohistochemical and gene expression). The role of glial cells was examined, as well. Additionally, the effects of chronic administration of the tricyclic antidepressant imipramine and the anti-TNF-α pentoxyphylline were investigated. METHODS: Wistar rats were exposed to either repeated LPS (50µg/kg i.p.) over 2weeks, CMS protocol for 4weeks or LPS over 2weeks then 4weeks of CMS. Two groups of rats were exposed to LPS/CMS protocol and treated with either imipramine or pentoxifylline. Rats were examined for behavioral, neurochemical and gene expression changes. RESULTS: Animals exposed to LPS/CMS elaborated depressive-like symptoms with significant increase in both serum corticosterone and TNF-α levels compared to those in the saline, LPS or CMS groups. Hippocampal kynurenine/tryptophan ratio and TNF-α gene expression showed significant increase in the LPS/CMS model compared to those in saline, LPS or CMS groups. The immunohistochemical findings scrutinized the topography of the examined effects. Chronic treatment with imipramine or pentoxifylline significantly ameliorated the behavioral, neurochemical, immunohistochemical and TNF-α gene expression changes induced by the LPS/CMS protocol. CONCLUSION: This study gave a clue to the neurobiological processes underlying, at least, the subtypes of depressive disorders. It highlighted the possible interactions between stress and immune-inflammatory pathways in the pathogenesis of depression and suggested a new animal model of depression that addresses these pathways.

Aberrant methylation of RARbeta2 and APC genes in voided urine as molecular markers for early detection of bilharzial and nonbilharzial bladder cancer.

BACKGROUND: Bladder cancer cells illustrate major disruptions in their DNA methylation patterns as compared with normal ones. Authors aimed to identify epigenetic molecular markers in urine for early detection of bladder cancer. MATERIALS AND METHODS: We retrospectively analyzed the methylation status of RARß(2) and APC genes in urine samples from 210 bladder cancer patients, 61 patients with benign urological diseases, and 49 healthy volunteers by using methylation-specific PCR. RESULTS: Methylated RARß(2) and APC were significantly higher in bladder cancer patients (62.8%, 59.5%) than benign (16.4%, 5%) but not detected in healthy volunteers (0%) at (P < 0.0001). Both methylated genes showed no significant difference among clinicopathologic factors; however, they were detected in all grades and stages. Among the 128 patients with bilharzial bladder cancer, 94 (73.4%) showed methylated RARß(2) and 86 (67.2%) showed methylated APC. Homoplasmic methylation pattern of both genes were only detected in bilharzial bladder cancer cases. Both sensitivities and specificities of the methylated genes for bladder cancer detection were superior to urine cytology and when altogether combined, the sensitivities improved to (91.8%), (93.5%), (91.9%), and (80.9%) in detection of: bladder cancer, non-muscle invasive bladder cancer, low-grade tumors, and bilharzial associated bladder cancer, respectively. CONCLUSION: Thus, methylated RARß(2) and APC genes might be valuable urinary molecular markers for early detection of bilharzial and nonbilharzial bladder cancer.

The Cell Cycle Regulators P16INK4a, P15INK4b and Cyclin D1: Relationship to Clinicopathological Parameters and Disease-free Survival in Laryngeal Carcinoma Patients.

Laryngeal squamous cell carcinoma (LSCC) is a frequent malignancy with a complex and undefined etiology to date. The recently identified cyclin-dependent kinase inhibitor p15INK4b is frequently deleted in human tumors. Previous evidence has pointed to a related gene, p16INK4a, as another target for deletion. Both genes express cyclin D inhibitor proteins. To determine the importance of cell cycle regulators in LSCC relative to more traditional surgical and pathological prognostic factors, p15INK4b, p16INK4a and cyclin D1 analyses were performed. Forty-one malignant tumor tissues and 20 minimal pathological lesions (MPL) of the larynx were examined for deletion of the p16INK4a and p15INK4b genes using polymerase chain reaction. Cyclin D1 expression was studied by Western blotting. Deletions of p16INK4a and p15INK4b were observed in 48.8 % and 51.2% of LSCC patients, respectively. Meanwhile, no deletion was observed in MPL (p<0.001). Cyclin D1 was expressed in 43.9% of patients with LSCC versus 30% with MPL (p=0.29). Although the frequency of p16INK4a and p15INK4b deletions were higher in advanced than early tumor stages, the difference was statistically insignificant. Ninety percent of patients with deletion of p16INK4a had deletion of the p15INK4b gene. Both cyclin D1 expression and deletion of p15INK4b were found to be independent prognotic predictors of disease recurrence. p16INK4a and p15INK4b gene deletions are exclusively related to malignancy of the larynx. Cyclin D1 expression and p15INK4b gene deletion are potential prognostic indicators of recurrence of LSCC.

Detection of bladder carcinoma by combined testing of urine for hyaluronidase and cytokeratin 20 RNAs.

BACKGROUND: A new, sensitive, noninvasive method for the detection of urothelial carcinomas of the urinary bladder would open new possibilities in both the diagnosis and followup of patients. METHODS: This study included 228 patients diagnosed with bladder carcinoma, 68 patients with benign bladder lesions, and 44 healthy persons served as the control group. All were subjected to: serologic schistosomiasis antibody assay in serum, urine cytology, estimation of urine hyaluronic acid (HA) by enzyme-linked immunosorbent assay, and detection of CK-20 and hyaluronidase (HAase) by reverse transcription polymerase chain reaction (RT-PCR) in urothelial cells from voided urine. RESULTS: HA mean rank was higher in benign and malignant groups than in the healthy group (P < 0.0001) and was significantly related to tumor grade (P = 0.021). HA best-cutoff, determined using receiver operating characteristic curve to discriminate between malignant and nonmalignant groups, was 58.5 units/mg protein at 85.8% sensitivity and 60.7% specificity. HAase RNA showed superior sensitivity (90.8%) over cytology (68.9%) and CK-20 (78.1%) with specificity of 93.4%, 98.1% and 80.2%, respectively. The sensitivity reached 94.7% at a specificity of 91.5% when combined with CK-20. All 4 of the investigated markers were related to grade at P <0.05. Whereas only HAase and CK-20 were significantly related to stage (P < 0.05). As to schistosomiasis, only HAase RNA positivity was significantly associated (P = 0.038). CONCLUSIONS: HAase RNA is a promising noninvasive test with high sensitivity and specificity in bladder carcinoma detection.

Human Papillomavirus High-risk Genotypes: Relationship to Apoptosis and p53 Expression in Egyptian Patients with Laryngeal Carcinoma.

Human papillomavirus (HPV) infection is suspected of causing laryngeal carcinoma. The relationship of HPV-16 and 18 genotypes to apoptosis and p53 protein expression in Egyptian laryngeal carcinoma patients was studied. Biopsy specimens from 82 patients with laryngeal carcinoma and 28 with minimal pathological lesions serving as a control group were examined. In all specimens, HPV-16 and-18 were examined using PCR, p53 expression was studied by immunohistochemistry and DNA fragmentation to assess apoptosis was assayed using a biochemical method and gel electrophoresis. HPV-16 was detected in 51.2% of laryngeal carcinoma patients versus 14.3% of the control group (p=0.001). The surrounding areas of positive tumors were negative in 52.4% of them. HPV-16 was significantly higher in tumors with higher expression of p53 (p=0.026). An inverse significant relationship was found between HPV-16 and DNA fragmentation in the laryngeal carcinoma group (p=0.022). HPV-18 was detected in only 2.4% of laryngeal carcinoma patients. p53 protein was expressed in 76.8% of the malignant group with significant increasing positivity with increasing stage of the disease (p=0.025). Non of the control group was p53-positive. Our results suggest that highly oncogenic types of HPV may play a role in the pathogenesis of laryngeal carcinoma through inactivation of wild-type p53 with subsequent decrease in apoptosis and by induction of p53 mutation, which itself can induce malignant transformation.

Prognostic Significance of Ets-1 and the Matrix Metalloproteinase-2 to Tissue Inhibitor of Matrix Metalloproteinase-2 Ratio in Egyptian Epithelial Ovarian Cancer Patients.

The invasive potential of epithelial ovarian cancer is the main factor determining its biological behavior. Ets-1 transcription factor is involved in the activation of several proteases participating in tumor invasion and metastasis. The balance of matrix metalloproteinase-2 and its inhibitor is important in the regulation of tumor invasion. This study included 31 tumors from patients with epithelial ovarian cancer in different stages, and 20 tissue samples from benign ovarian lesions as a control group. Ets-1 was assessed using immunohistochemistry. Matrix metalloproteinase-2 (MMP-2) and the MMP-2 inhibitor (TIMP-2) were measured in the cytosolic fractions using enzyme immunoassay. MMP-2 results were confirmed by gelatin zymography. ETS-1 was expressed only in the malignant group. MMP-2 and the MMP-2:TIMP-2 ratio were significantly higher in the malignant group (p=0.045 and 0.026, respectively) with significant correlation to stage and poor survival above the specified cut-off values (p=0.006 and 0.002, respectively). Log rank of Kaplan-Meier survival analysis was significant for FIGO stage, MMP-2 and the MMP-2:TIMP-2 ratio (p<0.05). Multivariate analysis demonstrated that the MMP-2:TIMP-2 ratio is an independent prognostic parameter. Ets-1 could be a future target for therapeutic strategies. Moreover, the MMP-2:TIMP-2 ratio might serve as a new independent prognostic indicator of poor prognosis in epithelial ovarian cancer patients.