Hanan Shehata

Expression of HYAL1 and Survivin RNA as Diagnostic Molecular Markers for Bladder Cancer

PURPOSE Urinary tumor markers that help in the early detection of bladder cancer promise a significant improvement in sensitivity, specificity and convenience over conventional, invasive diagnostic tests. We assessed the diagnostic efficacy of hyaluronidase (HYAL1) and survivin for early bladder cancer detection. MATERIALS AND METHODS The study included 166 patients diagnosed with bladder carcinoma, 112 with benign bladder lesions and 100 healthy volunteers who served as controls. All underwent serological assessment of schistosomiasis antibody, urine cytology, and hyaluronidase (HYAL1) and survivin RNA estimation by qualitative and semiquantitative reverse transcriptase-polymerase chain reaction in urothelial cells from voided urine. RESULTS Positivity rates of HYAL1 RNA and survivin RNA on qualitative reverse transcriptase-polymerase chain reaction were significantly different among the 3 groups. Mean rank using semiquantitative method was increased in the malignant vs the other groups. The best cutoff for HYAL1 and survivin RNA was 0.25 each. Using these cutoffs HYAL1 and survivin RNA sensitivity was 91% and 75%, respectively, with absolute specificity. HYAL1 RNA detected all patients with stages 0 and I bladder cancer (p <0.037). Urine cytology sensitivity improved when combined with hyaluronidase or survivin RNA on semiquantitative reverse transcriptase-polymerase chain reaction. CONCLUSIONS The detection of urinary HYAL1 and survivin RNA is a promising noninvasive test for bladder cancer early detection. HYAL1 RNA was more sensitive and specific than urine cytology. Semiquantitative reverse transcriptase-polymerase chain reaction is favored for its high sensitivity and specificity.

Histone Deacetylases Enzyme, Copper, and IL-8 Levels in Patients With Alzheimer’s Disease :

Background: Alzheimers disease (AD) is a neurodegenerative disorder characterized by progressive loss of cognitive abilities. Epigenetic modification, oxidative stress, and inflammation play an important role in the pathogenesis of the disease. We aimed to detect noninvasive peripheral biomarkers with a high degree of sensitivity and specificity in diagnosis and progression of AD. Methods: A total of 25 elderly patients with AD and 25 healthy control participants were selected and subjected to cognitive assessment and laboratory measures including histone deacetylases (HDACs), copper, and interleukin 8 (IL-8) levels. Results: The levels of HDACs, copper, and IL-8 were significantly higher in patients with AD (P < .001) and had a significant negative effect on all cognitive assessment tests. Receiver–operating curve (ROC) analysis revealed that HDACs and copper levels had higher sensitivity and specificity. Conclusions: Plasma levels of HDACs and copper may be used as peripheral biomarkers in diagnosis of AD, while IL-8 level could be a useful biomarker in following AD progression.

Antineoplastic Effects of Bee Honey and Nigella sativa on Hepatocellular Carcinoma Cells

Objectives: To evaluate in vitro antitumor effects of bee honey (BH) and Nigella sativa (NS) on HepG2 through their antioxidant and apoptotic activities. Methods: HepG2 cell line was treated with different concentrations of diluted unfractionated BH and different concentrations of alcohol extract of NS. Exposure lasted for different time durations (6-72 hours), both dose–response and time course–response were conducted. Cell viability was tested by trypan blue exclusion test. Total antioxidant status and caspase-3 activity were estimated in the cell lysate. Nitric oxide levels were measured in culture supernatants of both treated and untreated HepG2 at all indicated times. Results: Treatment of HepG2 cells with BH and NS leads to a significant decrease in both the number of viable HepG2 cells and the levels of nitric oxide on one hand, but improvement of the total antioxidant status and caspase-3 activity on the other, especially in HepG2 cells treated with higher doses of BH and NS (20% and 5000 μg/mL, respectively) and for longer duration (72 hours). Conclusions: BH and NS are effective in reducing the viability of HepG2 cells, improving their antioxidant status and inducing their apoptotic death.

Detection of survivin protein in aqueous humor and serum of retinoblastoma patients and its clinical significance

OBJECTIVES There is growing biomedical interest in survivin as a diagnostic and prognostic factor in human neoplasms. The present work assessed survivin expression in retinoblastoma. DESIGN AND METHODS Survivin was quantitatively measured in 82 aqueous humor and serum samples from 41 children with retinoblastoma and nonmalignant ophthalmic diseases. After treatment, 12 serum samples from retinoblastoma patients were included. RESULTS Survivin levels were higher in aqueous humor and serum of retinoblastoma patients than the controls (P<0.05). In aqueous, it was significantly correlated with the tumor stage and optic nerve affection. The best cutoff values for survivin in aqueous and serum that differentiate between retinoblastoma and nonmalignant ophthalmic diseases were 25.2 pg/mg protein and 12.9 pg/mL, respectively. The sensitivity and specificity were 100% and 90% in serum and 94% and 48% in aqueous, respectively. Serum survivin has decreased significantly after treatment. CONCLUSION Measurement of survivin protein might have a great diagnostic and prognostic potential in retinoblastoma.

Effects of pentoxifylline, 7-nitroindazole, and imipramine on tumor necrosis factor-α and indoleamine 2,3-dioxygenase enzyme activity in the hippocampus and frontal cortex of chronic mild-stress-exposed rats

Objectives: This study aimed to investigate the role of tumor necrosis factor (TNF)-α and the neuronal nitric oxide synthase enzyme in dysregulation of indoleamine 2,3-dioxygenase (IDO) enzyme, and hence serotonin availability in chronic mild stress (CMS), an animal model of depression. Methods: Rats were divided into five groups: two control and CMS-exposed for 6 weeks, and another three groups exposed to CMS and administered pentoxifylline 50 mg/kg/day intraperitoneally, 7-nitroindazole 40 mg/kg/day subcutaneously, or imipramine 20 mg/kg/day intraperitoneally for the previous 3 CMS weeks. Rats were assessed for neurochemical and immunohistochemical abnormalities. Results: Pentoxifylline-, 7-nitroindazole-, and imipramine-treated rats showed amelioration of CMS-induced behavioral deficits that was accompanied by significant reduction in kynurenine/serotonin molar ratio and nitrates/nitrites in frontal cortex and hippocampus. In the pentoxifylline and 7-nitroindazole groups, serum TNF-α was reduced relative to the CMS group (18.54 ± 0.85 and 19.16 ± 1.54 vs 26.20 ± 1.83 pg/mL, respectively; P < 0.05). Exposure to CMS increased TNF-α and IDO immunohistochemical staining scores in both hippocampus and midbrain raphe nuclei. 7-Nitroindazole and pentoxifylline significantly (P < 0.05) reduced TNF-α immunostaining in hippocampus and raphe nuclei, with significant (P < 0.01) reduction of IDO immunostaining in raphe nuclei. Likewise, imipramine reduced TNF-α immunostaining (P < 0.05) in hippocampus. Conclusion: Neuronal nitric oxide synthase and TNF-α may play a concerted role in modulating IDO enzyme activity in CMS-exposed rats and provide additional evidence for possible alternative approaches to switch the neurobiological processes in depression.

Evaluation of Circulatory RNA-Based Biomarker Panel in Hepatocellular Carcinoma

BackgroundThe circulating transcriptome (coding and non-coding) plays a critical role in cancer. Novel accurate strategies for early detection of hepatocellular carcinoma (HCC) are strongly needed.Patients and MethodsWe chose an HCC-specific RNA-based biomarker panel based on the integration of differential lysosomal-associated membrane protein 2 (LAMP2) gene expression with its selected epigenetic regulators using bioinformatic methods. This was followed by RT-qPCR validation in serum of 78 patients with HCC, 36 patients with chronic hepatitis C (CHC) infection and 44 healthy volunteers. We used risk-score analysis to evaluate the diagnostic efficacy of the serum profiling system. Moreover, in twenty of the 78 HCC cases involved in the study we examined the expression of RNA-based biomarker panel in both HCC and adjacent non-tumor tissues and assessed their correlation with the serum level of this panel.ResultsThe four ribonucleic acid (RNA)-based biomarker panel [long non-coding RNA-C terminal binding protein, androgen responsive (lncRNA–CTBP), microRNA-16-2 (miR-16-2), microRNA-21-5-P (miR-21-5p) and LAMP2], had high sensitivity and specificity for discriminating HCC from healthy controls and also from CHC patients. Among these four RNAs, serum miR-16-2 and miR-21-5p were independent prognostic factors.ConclusionThe circulatory RNA-based biomarker panel can serve as a potential biomarker for HCC diagnosis and prognosis.

HER1 R497K and HER2 I655V polymorphisms are linked to development of breast cancer.

BACKGROUND: Polymorphism of the genes of Human Epidermal growth factor receptor1 (HER1) and receptor2 (HER2) have been reported to be linked to pathogenesis of several malignant tumors but still there is contradiction regarding their association with breast cancer. OBJECTIVE: In this case control study we aimed to analyze the frequency of HER1 R497K (rs 11543848) and HER2 I655V (rs 1136201) Polymorphisms in breast cancer. SUBJECT AND METHOD: The frequency of HER1 Arg(R) 497Lys (K) and HER2 Ile (I) 655Val (V) polymorphisms were tested in 64 breast cancer patients and 86 normal control by polymerase chain reaction followed by restriction fragment polymorphism detection. Immunohistochemical analysis was done for HER2 protein on the available 18 malignant tissue samples. RESULTS: HER1 497K and HER2 655V variant had significantly increased breast cancer risk (OR=2.6, 95% CI 1.6–4.2, OR=2.2, 95% CI 1.2–4.1, p< 0.05) respectively. Moreover, combined HER1 K497 and HER2 V655 variant was detected in 26.6% malignant in comparison to 8.14% of control group (OR=4.1, 95% CI 1.58–10.57), but, no significant association was noticed between both Polymorphisms and clinicopathological features of the disease. As regard HER2 immunohistochemical expression no significant correlation was revealed with HER2 655V polymorphism. CONCLUSIONS: Our findings suggest that HER1 497K and HER2 655V polymorphisms are potential risk factor for development of breast cancer.

Breast tissue–based microRNA panel highlights microRNA-23a and selected target genes as putative biomarkers for breast cancer

We explored the differential expression of breast tissue-based panel of microRNAs (miRNAs) and their potential application as prognostic markers of breast cancer (BC). This study was divided into the following phases: (1) A panel of 6 BC characteristic miRNAs, which were retrieved based on the microarray signature profiling (released by miRWalk), was explored using SYBR Green-based polymerase chain reaction (PCR) array in 16 cancerous and 16 noncancerous breast tissue; (2) pathway enrichment analysis of the key miRNA target genes; (3) marker choice and validation by real-time PCR in a larger set of 76 patients with BC, 36 benign breast conditions, and 36 healthy volunteers; (4) validation of miRNA (miR)-23a target genes (forkhead box m [FOXM1] and histidine-rich glycoprotein [HRG]) by conventional reverse transcriptase (RT)-PCR; and (5) the prognostic significance of the investigated parameters in the BC validation group was explored. In PCR array-based miRNA expression analysis, 4 miRNAs were found to be altered more than twice (miR-96, miR-29c, miR-221, and miR-23a). Bioinformatic analysis of the target genes revealed enrichment for special biological process categories, that is, cell cycle, angiogenesis, apoptosis, cell proliferation, and cell adhesion. miR-23a, HRG messenger RNA, and FOX messenger RNA were positive in BC by 82.9%, 72.4%, and 71.1%, respectively. The overall concordance rates between miR-23a with HRG and FOXM1 tissue RNAs were 91% and 79%, respectively. The median follow-up period was 49 months. mi-23a and HRG RNA were significant independent prognostic markers in relapse-free survival. miR-23a may have an oncogenic function and enhance BC progression by directly activating FOXM1 and HRG at RNA level.

Behavioural, metabolic, and endothelial effects of the TNF-α suppressor thalidomide on rats subjected to chronic mild stress and fed an atherogenic diet

There is accumulating evidence suggesting that depression is a risk factor for cardiovascular diseases. This study aimed to examine the hypothesis that the proinflammatory cytokine TNF-α would partially explain the link between depression and atherosclerotic endothelial changes. Rats were distributed among 6 groups: (i) control group; (ii) group subjected to chronic mild stress (CMS); (iii) group fed a cholesterol-cholic acid-thiouracil (CCT diet); and (iv) CMS group fed the CCT diet and treated with the vehicle for 8 weeks. The last 2 groups were subjected to CMS-CCT and received thalidomide (THAL) or imipramine (IMIP). Rats were assessed behaviorally (sucrose preference, open field, and forced-swimming tests). TNF-α protein was assessed from the serum, aorta, and liver. Aortic TNF-α gene expression (assessed using RT-PCR), serum lipid profile, and insulin levels were measured. Endothelial function was assessed in isolated aortic rings. The THAL and IMIP groups showed ameliorated CMS-CCT-related behavioral changes. CMS-CCT-induced metabolic and endothelial dysfunctions were improved in the THAL group but were worsened in the IMIP group. RT-PCR showed a significant reduction of aortic TNF-α mRNA expression in the THAL and IMIP treatment groups. These data paralleled the findings for aortic immunohistochemistry. The THAL group, but not the IMIP group, showed improved CMS-CCT-induced changes in the vascular reactivity of the aortic rings. Thus, TNF-α provides a target link between depression, metabolic syndrome, and endothelial dysfunction. This could open a new therapeutic approach to address the comorbidities of depression.

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.