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Motawi, T. K., A. E. Mady, M. Elhelbawy, R. M. Talaat, and N. N. Shahin, "Association of long noncoding RNA H19 rs2839698 C/T with hepatitis B virus infection and hepatocellular carcinoma risk.", Journal of biochemical and molecular toxicology, vol. 38, issue 4, pp. e23673, 2024. Abstract

The intricate pathogenesis of the hepatitis B virus (HBV) and its progression to hepatocellular carcinoma (HCC) have not yet been fully elucidated. H19 is one of the earliest imprinted long noncoding RNAs (lncRNAs) associated with liver pathobiology. This study investigated the association of H19 single nucleotide polymorphisms (SNPs) rs2839698 C/T and rs217727 C/T with HBV and HBV-related HCC and their correlation with H19 expression level. A total of 230 subjects were enrolled in this study including 100 HBV-infected patients, 30 HBV-related HCC patients, and 100 apparently healthy controls. TaqMan genotyping human assays were utilized to assess allelic discrimination for H19 SNPs. H19 expression was assessed using quantitative real-time polymerase chain reaction (qRT-PCR). Our findings showed that H19 rs2839698 was linked to a higher incidence of HBV infection and HBV-related HCC. Individuals who bear the CT genotype of rs2839698 were more susceptible to HBV infection (OR = 3.05; 95% CI 1.714-5.457; p < 0.001). Those harboring the TT genotype were more prone to develop HCC (OR = 2.625; 95% CI 1.037-6.64; p = 0.038). Our data revealed that rs2839698 could function as a promising predictor of HCC risk. Furthermore, H19 was significantly downregulated in HBV (p < 0.01) and HCC (p < 0.01) patients versus the control group. Significant upregulation of H19 in HCC patients with cirrhosis (p < 0.001) was detected. Altogether, this is considered the first prospective case-control study to address the implication of the genetic variations of H19 SNPs in HBV and HBV-related HCC in Egyptian patients.

Shahin, N. N., O. G. Shaker, and M. O. Mahmoud, "GOAT rs10096097 and CREB1 rs6740584 single nucleotide polymorphisms are associated with type 2 diabetes mellitus in Egyptians.", Archiv der Pharmazie, pp. e2400011, 2024. Abstract

Diabetes mellitus (DM) is a chronic disorder that affects nearly half a billion people around the world and causes millions of deaths annually. Treatment of diabetes or related complications represents an economic burden not only for developing countries but also for the developed ones. Hence, new efficient therapeutic and preventive strategies and screening tools are necessary. The current work aimed to assess the potential association of single nucleotide polymorphisms (SNPs) in ghrelin O-acyltransferase (GOAT) rs10096097, cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) rs6740584, and v-maf musculoaponeurotic fibrosarcoma oncogene homolog A (MafA) rs62521874 genes with type 2 DM susceptibility in Egyptians. A total of 96 patients with type 2 DM along with 72 healthy individuals participated in this study. Genotyping was executed via real-time polymerase chain reaction (PCR), and the serum protein levels of GOAT, CREB, and MafA were measured by enzyme-linked immunosorbent assay (ELISA). Genotyping revealed a significant association of GOAT rs10096097 and CREB1 rs6740584 SNPs with type 2 diabetes risk, with significantly higher GOAT rs10096097 G allele and CREB1 rs6740584 T allele frequencies in diabetic patients than in controls. However, insignificant association was identified between the MafA rs62521874 SNP and diabetes in the examined sample of the Egyptian residents. Serum GOAT, CREB1, and MafA protein levels did not vary significantly between diabetic and control individuals. Yet, significant variation in serum GOAT and CREB1 levels was detected between CREB1 rs6740584 genotypes within the diabetic group, with CT and TT genotype carriers showing higher levels than AA genotype patients. GOAT rs10096097 and CREB1 rs6740584, but not MafA rs62521874, SNPs are associated with type 2 diabetes risk in the studied Egyptians.

Haridy, S. F. A., N. N. Shahin, M. I. Shabayek, M. M. Selim, M. A. Abdelhafez, and T. K. Motawi, "Diagnostic and prognostic value of the RUNXOR/RUNX1 axis in multiple sclerosis.", Neurobiology of disease, vol. 178, pp. 106032, 2023. Abstract

The runt-related transcription factor-1 (RUNX1) gene with its lncRNA RUNXOR are recently becoming a research focus in various diseases, specifically immune-related diseases as they are implicated in multiple pathways. Interestingly, their role in multiple sclerosis (MS) remains unstudied. The present study explored the role of RUNXOR/RUNX1 in the development and progression of MS and investigated their possible mechanism of action. We measured the serum expression levels of lncRNA RUNXOR, as well as RUNX1, microtubule associated protein 2 (MAP2), nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) mRNAs in 30 healthy controls and 120 MS patients subdivided into 4 groups: 30 clinically isolated syndrome patients, 30 relapsing-remitting MS (RRMS) patients in relapse, 30 RRMS patients in remission and 30 secondary progressive MS patients. Additionally, we measured the serum protein levels of RUNX1, MAP2, NGF, BDNF and interleukin-10 (IL-10). All measured RNA expression levels were markedly downregulated and, consequently, the protein levels of RUNX1, MAP2, NGF, BDNF and IL-10 were significantly decreased in MS patients compared to healthy controls. Moreover, the levels of the measured parameters varied significantly within the MS groups. According to receiver-operating-characteristic (ROC) curve and logistic regression analyses, lncRNA RUNXOR, RUNX1 mRNA and its protein levels were predictors of disease progression, in addition to RUNX1 mRNA exhibiting a diagnostic potential. Altogether, this study suggests the implication of the RUNXOR-RUNX1 axis in MS development, progression, and increased MS-related disability, and highlights the potential utility of the studied parameters as promising diagnostic/prognostic biomarkers for MS.

Shama, S., H. Jang, X. Wang, Y. Zhang, N. N. Shahin, T. K. Motawi, S. Kim, S. Gawrieh, and W. Liu, "Phosphatidylethanolamines Are Associated with Nonalcoholic Fatty Liver Disease (NAFLD) in Obese Adults and Induce Liver Cell Metabolic Perturbations and Hepatic Stellate Cell Activation.", International journal of molecular sciences, vol. 24, issue 2, 2023. Abstract

Pathogenesis roles of phospholipids (PLs) in nonalcoholic fatty liver disease (NAFLD) remain incompletely understood. This study investigated the role of PLs in the progression of NAFLD among obese individuals via studying the alterations in serum PL composition throughout the spectrum of disease progression and evaluating the effects of specific phosphatidylethanolamines (PEs) on FLD development in vitro. A total of 203 obese subjects, who were undergoing bariatric surgery, were included in this study. They were histologically classified into 80 controls (C) with normal liver histology, 93 patients with simple hepatic steatosis (SS), 16 with borderline nonalcoholic steatohepatitis (B-NASH) and 14 with progressive NASH (NASH). Serum PLs were profiled by automated electrospray ionization tandem mass spectrometry (ESI-MS/MS). HepG2 (hepatoma cells) and LX2 (immortalized hepatic stellate cells or HSCs) were used to explore the roles of PL in NAFLD/NASH development. Several PLs and their relative ratios were significantly associated with NAFLD progression, especially those involving PE. Incubation of HepG2 cells with two phosphatidylethanolamines (PEs), PE (34:1) and PE (36:2), resulted in significant inhibition of cell proliferation, reduction of mitochondrial mass and membrane potential, induction of lipid accumulation and mitochondrial ROS production. Meanwhile, treatment of LX2 cells with both PEs markedly increased cell activation and migration. These effects were associated with a significant change in the expression levels of genes involved in lipogenesis, lipid oxidation, autophagy, apoptosis, inflammation, and fibrosis. Thus, our study demonstrated that elevated level of PEs increases susceptibility to the disease progression of obesity associated NAFLD, likely through a causal cascade of impacts on the function of different liver cells.

Motawi, T. M. K., N. A. H. Sadik, D. Sabry, S. A. Fahim, and N. N. Shahin, "rs62139665 Polymorphism in the Promoter Region of EpCAM Is Associated With Hepatitis C Virus-Related Hepatocellular Carcinoma Risk in Egyptians", Frontiers in Oncology, vol. 11, pp. 754104, 2022. AbstractWebsite

Hepatocellular carcinoma (HCC) is a universal health problem that is particularly alarming in Egypt. The major risk factor for HCC is hepatitis C virus (HCV) infection which is a main burden in Egypt. The epithelial cell adhesion molecule (EpCAM) is a stem cell marker involved in the tumorigenesis and progression of many malignancies, including HCC. We investigated the association of -935 C/G single nucleotide polymorphism in EpCAM promoter region (rs62139665) with HCC risk, EpCAM expression and overall survival in Egyptians. A total of 266 patients (128 HCV and 138 HCC cases) and 117 age- and sex-matched controls participated in this study. Genotyping, performed using allelic discrimination and confirmed by sequencing, revealed a significant association between EpCAM rs62139665 and HCC susceptibility, with higher GG genotype and G allele distribution in HCC patients than in non-HCC subjects. Such association was not detected in HCV patients compared to controls. EpCAM gene expression levels, determined in blood by RT-qPCR, and its serum protein expression levels, determined by ELISA, were significantly higher in GG relative to GC+CC genotype carriers in HCV and HCC patients in a recessive model. ROC analysis of EpCAM protein levels revealed significant discriminatory power between HCC patients and non-HCC subjects, with improved diagnostic accuracy when combining α-fetoprotein and EpCAM compared to that of α-fetoprotein alone. Altogether, EpCAM rs62139665 polymorphism is significantly associated with HCC and with EpCAM gene and protein expression levels in the Egyptian population. Moreover, serum EpCAM levels may hold promise for HCC diagnosis and for improving the diagnostic accuracy of α-fetoprotein.

Shahin, N. N., R. N. Shamma, and I. S. Ahmed, "A Nano-Liposomal Formulation of Caffeic Acid Phenethyl Ester Modulates Nrf2 and NF-κβ Signaling and Alleviates Experimentally Induced Acute Pancreatitis in a Rat Model.", Antioxidants (Basel, Switzerland), vol. 11, issue 8, pp. 1536, 2022. Abstract

The currently available management strategies for acute pancreatitis are inadequately effective which calls for exploration of new approaches to treat this condition. Caffeic acid phenethyl ester (CAPE) is a major bioactive constituent of honeybee propolis with promising therapeutic and preventive applications. However, its pharmaceutical potential and clinical use are hindered by its poor water solubility and limited plasma stability. In this study, we aimed to prepare, characterize and evaluate a CAPE-loaded nanoliposomal formulation to improve the efficacy of CAPE for the management of acute pancreatitis. The CAPE-loaded nanoliposomes (CAPE-loaded-NL) were prepared by a thin layer evaporation technique and were optimized using three edge activators. CAPE-loaded-NL were characterized for their vesicle size (VS), zeta potential (ZP), encapsulation efficiency (EE), polydispersity index (PDI), crystalline state and morphology. The protective effect of the optimal CAPE-loaded-NL was evaluated in a rat model of acute pancreatitis induced by administering a single intraperitoneal injection of L-ornithine. Oral pretreatment with CAPE-loaded-NL significantly counteracted ornithine-induced elevation in serum activities of pancreatic digestive enzymes and pancreatic levels of malondialdehyde, nuclear factor kappa B (NF-κB) p65, tumor necrosis factor-alpha, nitrite/nitrate, cleaved caspase-3 and myeloperoxidase activity. Moreover, pretreatment with CAPE-loaded-NL significantly reinstated the ornithine-lowered glutathione reductase activity, glutathione, nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 levels and ATP/ADP ratio, and potentiated the Bcl-2/Bax ratio in pancreatic tissue. CAPE-loaded-NL displayed superior antioxidant, anti-inflammatory and anti-apoptotic effects compared to free CAPE oral suspension and achieved a more potent correction of the derangements in serum amylase and pancreatic myeloperoxidase activities. The histological observations were in line with the biochemical findings. Our results suggest that CAPE-loaded-NL provide a promising interventional approach for acute pancreatitis mainly through the enhancement of the exerted antioxidant, anti-inflammatory and anti-apoptotic effects which may be mediated, at least in part, through modulation of Nrf2 and NF-κβ signaling.

Kotb, R. M., S. S. Ibrahim, O. M. Mostafa, and N. N. Shahin, "Potential role of CXCR4 in trastuzumab resistance in breast cancer patients.", Biochimica et biophysica acta. Molecular basis of disease, vol. 1868, issue 11, pp. 166520, 2022. Abstract

Despite the efficacy of trastuzumab in treating HER2-positive breast cancer patients, a significant proportion of patients relapse after treatment. The role of C-X-C chemokine receptor type 4 (CXCR4) in trastuzumab resistance was studied only in cell lines and the underlying mechanisms remain largely unclear. This study investigated the role of CXCR4 in trastuzumab resistance in breast cancer patients and explored the possible underlying mechanisms. The study was performed retrospectively on tissue samples from 62 breast cancer patients including 42 who were treated with trastuzumab and chemotherapy and 20 who received chemotherapy alone in adjuvant setting. Expression levels of CXCR4 and its regulators hypoxia-inducible factor 1-alpha (HIF-1α), tristetraprolin (TTP), human antigen R (HuR), itchy E3 ubiquitin protein ligase (ITCH), miR-302a and miR-494 were determined and their associations with tumor recurrence and disease-free survival were analyzed. In trastuzumab-treated patients, high CXCR4 expression was associated with recurrence and was an independent predictor of progression risk after therapy. CXCR4 correlated positively with its transcriptional regulator, HIF-1α, and negatively with its post-translational regulator, ITCH. HIF-1α, HuR and ITCH were significantly associated with clinical outcome. In chemotherapy-treated patients, neither CXCR4 nor any of its regulators were associated with recurrence or predicted disease progression risk after chemotherapy. In conclusion, this study suggests a potential role for CXCR4 in recurrence after trastuzumab-based therapy in human breast cancer that could be mediated, at least in part, by hypoxia and/or decreased ubiquitination. These findings highlight the potential utility of CXCR4 as a promising target for enhancing trastuzumab therapeutic outcome.

Motawi, T. M. K., N. A. H. Sadik, D. Sabry, S. A. Fahim, and N. N. Shahin, "rs62139665 Polymorphism in the Promoter Region of EpCAM Is Associated With Hepatitis C Virus-Related Hepatocellular Carcinoma Risk in Egyptians", Women in Hepato Pancreatic Biliary (HPB) Tumors: 2021, Volume I: Frontiers, 2022.
El-Fattah, A. A. A., N. A. H. Sadik, O. G. Shaker, A. M. Kamal, and N. N. Shahin, "Serum Long Non-Coding RNAs PVT1, HOTAIR, and NEAT1 as Potential Biomarkers in Egyptian Women with Breast Cancer.", Biomolecules, vol. 11, issue 2, 2021. Abstract

Long non-coding RNAs play an important role in tumor growth, angiogenesis, and metastasis in several types of cancer. However, the clinical significance of using lncRNAs as biomarkers for breast cancer diagnosis and prognosis is still poorly investigated. In this study, we analyzed the serum expression levels of lncRNAs PVT1, HOTAIR, NEAT1, and MALAT1, and their associated proteins, PAI-1, and OPN, in breast cancer patients compared to fibroadenoma patients and healthy subjects. Using quantitative real-time PCR (qRT-PCR), we compared the serum expression levels of the four circulating lncRNAs in patients with breast cancer ( = 50), fibroadenoma ( = 25), and healthy controls ( = 25). The serum levels of PAI-1 and OPN were measured using ELISA. Receiveroperating-characteristic (ROC) analysis and multivariate logistic regression were used to evaluate the diagnostic value of the selected parameters. The serum levels of HOTAIR, PAI-1, and OPN were significantly higher in breast cancer patients compared to controls and fibroadenoma patients. The serum level of PVT1 was significantly higher in breast cancer patients than in the controls, while that of NEAT1 was significantly lower in breast cancer patients compared to controls and fibroadenoma patients. Both ROC and multivariate logistic regression analyses revealed that PAI-1 has the greatest power in discriminating breast cancer from the control, whereas HOTAIR, PAI-1, and OPN have the greatest power in discriminating breast cancer from fibroadenoma patients. In conclusion, our data suggest that the serum levels of PVT1, HOTAIR, NEAT1, PAI-1, and OPN could serve as promising diagnostic biomarkers for breast cancer.

Arab, H. H., M. M. Safar, and N. N. Shahin, "Targeting ROS-Dependent AKT/GSK-3β/NF-κB and DJ-1/Nrf2 Pathways by Dapagliflozin Attenuates Neuronal Injury and Motor Dysfunction in Rotenone-Induced Parkinson's Disease Rat Model.", ACS chemical neuroscience, vol. 12, issue 4, pp. 689-703, 2021. Abstract

Dapagliflozin, a selective sodium-glucose co-transporter 2 (SGLT2) inhibitor, has emerged as a promising neuroprotective agent in murine models of epilepsy and obesity-induced cognitive impairment through its marked antioxidant/antiapoptotic features. However, the impact of dapagliflozin on the pathogenesis of Parkinson's disease (PD) is lacking. Hence, the present study aimed at exploring the potential neuroprotective effects of dapagliflozin against PD-associated neurodegenerative aberrations/motor dysfunction in rotenone-induced PD rat model. Rotenone (1.5 mg/kg) was subcutaneously administered every other day for 3 weeks. The expression of target signals was investigated using qPCR, Western blotting, ELISA, and immunohistochemistry. Dapagliflozin (1 (mg/kg)/day, by gavage for 3 weeks) attenuated PD motor dysfunction and improved motor coordination in the open-field and rotarod tests without triggering hypoglycemia. It also diminished the histopathologic alterations and α-synuclein expression and augmented tyrosine hydroxylase and dopamine levels. Dapagliflozin markedly alleviated neuronal oxidative stress via lowering lipid peroxides with consequent restoration of the disturbed DJ-1/Nrf2 pathway. Moreover, dapagliflozin counteracted ROS-dependent neuronal apoptosis and upregulated GDNF and its downstream PI3K/AKT/GSK-3β (Ser9) pathway. Meanwhile, it suppressed neuroinflammation via curbing the activation of NF-κB pathway and TNF-α levels. Together, these pleiotropic neuroprotective effects highlight the promising role of dapagliflozin in the management of PD.

Hanafy, M. M., J. Z. Lindeque, S. A. EL-Maraghy, A. - H. Z. Abdel-Hamid, and N. N. Shahin, "Time-based investigation of urinary metabolic markers for Type 2 diabetes: Metabolomics approach for diabetes management.", BioFactors (Oxford, England), 2021. Abstract

Diabetes is considered one of the most important health emergencies worldwide and Egypt has 8.2 million diabetic patients according to the International Diabetes Federation report in 2017. The objective of this study was to monitor the time-course variation in the metabolic profile of diabetic rats to detect urinary metabolic biomarkers using the metabolomics approach. Type 2 diabetes was induced in male Wistar albino rats using a single intraperitoneal injection of 40 mg/kg of streptozotocin following oral administration of 10% fructose in drinking water for 3 weeks. Then, urine was collected for 24 h from rats at three time points (0, 2, and 4 weeks after confirmation of diabetes), and were analyzed by nuclear magnetic resonance (H -NMR), followed by multivariate data analysis. The results from H -NMR pointed out that d-glucose, taurine, l-carnitine, l-fucose, 1,5-anhydrosorbitol, and d-galactose levels showed consistent significant variation (p < 0.05) between the positive (diabetic) and negative (normal) controls during the whole experimental period. Also, with the disease progression, myoinositol, and l-phenylalanine levels were significantly altered (p < 0.05) after 2 weeks and this alteration was maintained till the end of the 4-week experimental period in the positive control group. From the results of the present study, it could be concluded that we cannot depend only on glucose levels for prognostic purposes since there are other metabolic disturbances in diabetes which need to be tracked for better disease prognosis.

Motawi, T. M. K., Z. M. Abdel-Nasser, and N. N. Shahin, "Ameliorative Effect of Necrosulfonamide in a Rat Model of Alzheimer's Disease: Targeting Mixed Lineage Kinase Domain-like Protein-Mediated Necroptosis.", ACS chemical neuroscience, vol. 11, issue 20, pp. 3386-3397, 2020. Abstract

Alzheimer's disease (AD) is a progressively debilitating neurodegenerative disorder that has no effective remedy, so far, with available therapeutic modalities being only symptomatic and of modest efficacy. Necroptosis is a form of controlled cell death with a recently emerging link to the pathogenesis of several neurodegenerative diseases. This study investigated the role of necroptosis in the pathogenesis of AD and evaluated the potential beneficial effect of the necroptosis inhibitor, necrosulfonamide (NSA), in a rat model of AD. AD was induced by oral administration of AlCl (17 mg/kg/day) for 6 consecutive weeks. Administration of NSA (1.65 mg/kg/day) intraperitoneally for 6 weeks significantly amended AlCl-induced spatial learning and memory deficits, as demonstrated by enhanced rat performance in Morris water and Y-mazes. NSA alleviated the abnormally high hippocampal expression of tumor necrosis factor-alpha (TNF-α), β-site amyloid precursor protein cleaving enzyme 1 (BACE1), β-amyloid, glycogen synthase kinase-3β (GSK-3β), phosphorylated tau protein, and acetylcholinesterase with concordant replenishment of acetylcholine. The amendments of AD perturbations achieved by NSA correlated with its inhibitory effect on the phosphorylation of the key necroptotic executioner, mixed lineage kinase domain-like protein (MLKL). Histopathological alterations supported the biochemical findings. In conclusion, NSA treatment represents a promising anti-Alzheimer's approach, mitigating AD neuropathologies via targeting MLKL-dependent necroptosis.

Motawi, T. K., N. N. Shahin, K. Awad, A. S. Maghraby, D. N. Abd-Elshafy, and M. M. Bahgat, "Glycolytic and immunological alterations in human U937 monocytes in response to H1N1 infection.", IUBMB life, vol. 72, issue 11, pp. 2481-2498, 2020. Abstract

We monitored changes that took place in glycolytic enzymes, the pyruvate end product of glycolysis, tumor necrosis factor α (TNFα), and toll-like receptors (TLRs) both at the transcriptional and translational levels upon direct interaction between PR8-H1N1 and the human monocytes U937 in vitro system. U937 were first treated with H1N1 infectious viral particles or phorbol-12-myristate-13-acetate (PMA) or left untreated and later infected with the H1N1 virus. Levels of phosphofructokinase 1 (PFK1) and pyruvate were biochemically quantified. In addition, levels of TNFα, TLR3, and TLR7 were measured by ELISA. The transcriptional profiles of PFKs, inflammatory cytokines, TLR3 and TLR7 were relatively quantified by qRT-PCR. The results generally revealed significant changes in both the transcriptional and translational profiles of the studied biochemical and immunological parameters upon influenza infection in a time-dependent manner. In conclusion, H1N1 infection triggers transcriptional and translational changes in immortalized human monocytes, which might serve as markers for infection subject for further validation for their specificities.

Motawi, T. K., N. N. Shahin, A. S. Maghraby, M. Kirschfink, D. N. Abd-Elshafy, K. Awad, and M. M. Bahgat, "H1N1 Infection Reduces Glucose Level in Human U937 Monocytes Culture.", Viral immunology, vol. 33, issue 5, pp. 384-390, 2020. Abstract

Infection with influenza A (H1N1) virus contributes significantly to the global burden of acute respiratory diseases. Glucose uptake and metabolic changes are reported in different cell types after infections with different virus types, including influenza A virus. Alteration of glucose metabolism specifically in immune cells has major health consequences. The aim of this study was to monitor glucose concentration in unstimulated and stimulated U937 human monocytes with infectious or heat inactivated H1N1 or or in nonpathogenically stimulated monocytes with phorbol-12-myristate-13-acetate. Stimulated or unstimulated U937 human monocytes were subjected to H1N1 infection for different time points and the glucose profile in the growth medium was measured post infection. Results showed that regardless to whether the initial stimuli on U937 cells were of pathogen or nonpathogen origins, challenge infection by H1N1 causes a significant reduction of glucose levels 36 h post infection. In conclusion, H1N1 infection has a direct effect on the glucose uptake of U937 cells . This effect can be related to either H1N1 infection or cell differentiation status that might occur due to the exerted stimuli.

Shahin, N. N., G. T. Abd-Elwahab, A. A. Tawfiq, and H. M. Abdelgawad, "Potential role of aryl hydrocarbon receptor signaling in childhood obesity.", Biochimica et biophysica acta. Molecular and cell biology of lipids, vol. 1865, issue 8, pp. 158714, 2020. Abstract

BACKGROUND: There is a growing concern that junk food has contributed to the childhood obesity epidemic. Recently, experimental studies suggested that the aryl hydrocarbon receptor (AHR) gene is strongly linked to western diet-induced obesity.

AIM: This study investigated the potential role of AHR signaling in childhood obesity and the possible associations of the AHR-aryl hydrocarbon receptor repressor (AHRR)-cytochrome P450 1B1 (CYP1B1) axis with fatty acid homeostasis and the appetite-related hormones, leptin and ghrelin.

SUBJECTS AND METHODS: The study included 80 children; 54 obese and 26 non-obese of matched age and sex. Demographic data, anthropometric measurements, and lipid profile were assessed. Expression of AHR signaling genes was analyzed in blood cells by qRT-PCR. Serum insulin, leptin and ghrelin levels were measured using ELISA.

RESULTS: The statistical power of this study, calculated using G*Power version, was 90% (α = 0.05). AHR and CYP1B1 gene expression levels were upregulated in the obese group compared to controls, whereas AHRR, stearoyl-CoA desaturase 1 (SCD1), and peroxisome proliferator-activated receptor-γ (PPARγ) were downregulated. Serum leptin correlated positively, while serum ghrelin correlated negatively with both AHR and CYP1B1. Stratification of obese children by age revealed more activated AHR signaling in younger than in older children. Receiver-operating-characteristic (ROC) analysis revealed that AHR, AHRR and CYP1B1 could discriminate between obese and normal weight children. Multivariate analysis showed that AHRR, CYP1B1 and ghrelin could be significant independent predictors of obesity.

CONCLUSION: This study provides new insights into the molecular mechanisms contributing to childhood obesity by revealing alterations in the AHR-AHRR-CYP1B1 axis, which could serve as a promising therapeutic target for childhood obesity.

Abo-Elfadl, M. T., A. M. Gamal-Eldeen, M. F. Ismail, and N. N. Shahin, "Silencing of the cytokine receptor TNFRSF13B: A new therapeutic target for triple-negative breast cancer.", Cytokine, vol. 125, pp. 154790, 2020. Abstract

BACKGROUND: TNFRSF13B, TACI, is a member of the TNF receptor superfamily; it plays a key role in cancer cell proliferation and progression.

METHOD: Influence of silencing of human cytokine receptors on cell viability was screened by Luminescent Cell Viability Assay, after transfection of the siRNA library to find the maximum cell death superhits in both triple-negative MDA-MB-231 and double-positive MCF7 breast cells. The mode of cell death was investigated by dual DNA fluorescence staining. The expression of mRNAs of TACI, BAFF, BAFF-R, and APRIL was explored by qPCR. Immunocytofluorescence analysis was used to evaluate changes in TACI, Bcl-2, TNFR2, cyclin-D2, and PCNA. NF-kB p65, cell cycle, and necrosis/apoptosis (late and early) were analyzed by flow cytometry.

RESULTS: TACI is the most potent cytotoxic superhit resulted from high-throughput screening of the siRNA library, in both types of cells. Our findings indicated that silencing receptor TACI in both types of breast cancer cells led to significant cell death, after different intervals from siRNA transfection. Cell death mediators (TNFR2, Bcl-2, and NF-κB) were significantly decreased after TACI silencing. The key factors for cell division (Cyclin-D2 and PCNA) were significantly increased in silenced cells of both types but the cell cycle was arrested before the completion of mitosis. Expression of BAFF, BAFF-R and APRIL mRNA in TACI-silenced cells showed significant upregulation in MDA-MB-231 cells, while only BAFF-R and APRIL showed significant downregulation in MCF7 cells.

CONCLUSION: TACI silencing can be a new and promising therapeutic target for mesenchymal-stem like triple-negative breast cancer subtype.

Safar, M. M., N. N. Shahin, A. F. Mohamed, and N. F. Abdelkader, "Suppression of BACE1 and amyloidogenic/RAGE axis by sitagliptin ameliorates PTZ kindling-induced cognitive deficits in rats.", Chemico-biological interactions, vol. 328, pp. 109144, 2020. Abstract

The debilitating nature of cognitive impairment in epilepsy and the potential of some traditional antiepileptics to further deteriorate cognitive function are areas of growing concern. Glucagon-like peptide-1 (GLP-1) deficiency has been linked to reduced seizure threshold as well as cognitive dysfunction. Here, we tested whether sitagliptin (SITA), by virtue of its neuroprotective properties, could alleviate both epilepsy and associated cognitive dysfunction in a rat model of kindling epilepsy. Chemical kindling was induced by subconvulsive doses of pentylenetetrazol (PTZ) (30 mg/kg; i.p). SITA (50 mg/kg; p.o) was administered 1 h before PTZ injections. SITA conceivably attenuated PTZ hippocampal histological insult, preserved neuronal integrity and amended neurotransmitter perturbations in rat hippocampi paralleled with enhanced hippocampal GLP-1 levels as well as the downstream cAMP content and protein kinase A (PKA) activity. Moreover, SITA improved cognitive functioning of rats in the Morris water maze which was coupled with hampered hippocampal p(Ser)-tau and β-amyloid proteins. SITA replenished p(Ser)-glycogen synthase kinase-3β (GSK-3β). It also opposed the boosted matrix metalloproteinase-9 (MMP-9), brain-derived neurotrophic factor (BDNF), and insulin-like growth factor-1 (IGF-1) levels associated with PTZ administration along with mitigation of both β-secretase-1 (BACE1) immunoreactivity and receptor for advanced glycation end products (RAGE) protein level in rat hippocampi. In conclusion, SITA subdues epileptic and cognitive upshots of PTZ kindling in rats, which might correspond to the modulation of BACE1, amyloidogenic/RAGE axis as well as GSK-3β/MMP-9/BDNF signaling cascade. SITA effects are probably mediated via boosting GLP-1 and subsequently enhancing GLP-1/GLP-1R signaling.

Abdelmonem, M., N. N. Shahin, L. A. Rashed, H. A. A. Amin, A. A. Shamaa, and A. A. Shaheen, "Hydrogen sulfide enhances the effectiveness of mesenchymal stem cell therapy in rats with heart failure: In vitro preconditioning versus in vivo co-delivery.", Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie, vol. 112, pp. 108584, 2019 Feb 18. Abstract

Stem cell therapy represents a promising therapeutic avenue for cardiac disorders, including heart failure. Although stem cell transplantation showed encouraging preliminary results, the outcomes of clinical studies are still unsatisfactory. This study aimed to compare the outcomes of two therapeutic approaches, in vivo co-delivery of sodium hydrogen sulfide (NaHS) concomitant with bone marrow-derived mesenchymal stem cell (BMSC) transplantation and in vitro preconditioning of BMSCs with NaHS, both of which are intended to promote the success of stem cell therapy in rats with isoprenaline-induced heart failure. Heart failure developed 4 weeks after the subcutaneous injection of isoprenaline (170 mg/kg) for 4 consecutive days. The in vivo approach involved the co-delivery of intraperitoneally administered NaHS concomitant with BMSC transplantation for a period of 14 days. The in vitro approach involved preconditioning BMSCs with NaHS for 30 min before transplantation. Compared to treatment with BMSCs alone, in vitro preconditioning of BMSCs with NaHS improved left ventricular function as measured by echocardiography and electrocardiography and enhanced stem cell homing, proliferation and differentiation as manifested by higher cardiac expression of GATA-4 and myocyte enhancer factor 2. Moreover, the measurement of cardiac transforming growth factor beta 1 levels and histopathological investigation revealed mitigated fibrosis and myocardial injury scores. Compared with BMSC therapy alone, the in vivo approach enhanced stem cell homing and differentiation, alleviated fibrosis and augmented vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS) expression. In conclusion, NaHS can potentiate the efficiency of BMSC therapy for heart failure by in vitro preconditioning or in vivo co-delivery. The in vitro approach is superior with regard to improving cardiac function in addition to enhancing stem cell proliferation, while the in vivo approach is superior with regard to increasing cardiac VEGF and eNOS expression.

Shahin, N. N., N. A. El-Nabarawy, A. S. Gouda, and B. Mégarbane, "The protective role of spermine against male reproductive aberrations induced by exposure to electromagnetic field - An experimental investigation in the rat.", Toxicology and applied pharmacology, vol. 370, pp. 117-130, 2019. Abstract

The exponentially increasing use of electromagnetic field (EMF)-emitting devices imposes substantial health burden on modern societies with particular concerns of male infertility. Limited studies have addressed the modulation of this risk by protective agents. We investigated the hazardous effects of rat exposure to EMF (900 MHz, 2 h/day for 8 weeks) on male fertility and evaluated the possible protective effect of the polyamine, spermine, against EMF-induced alterations. Exposure to EMF significantly decreased sperm count, viability and motility, and increased sperm deformities. EMF-exposed rats exhibited significant reductions in serum inhibin B and testosterone along with elevated activin A, follicle-stimulating hormone, luteinizing hormone and estradiol concentrations. Testicular steroidogenic acute regulatory protein (StAR), c-kit mRNA expression and testicular activities of the key androgenic enzymes 3β- and 17β-hydroxysteroid dehydrogenases were significantly attenuated following exposure to EMF. Exposure led to testicular lipid peroxidation, decreased catalase and glutathione peroxidase activities and triggered nuclear factor-kappa B p65, inducible nitric oxide synthase, cyclooxygenase-2 and caspase-3 overexpression. EMF-exposed rats showed testicular DNA damage as indicated by elevated comet parameters. Spermine administration (2.5 mg/Kg/day intraperitoneally for 8 weeks) prevented EMF-induced alterations in the sperm and hormone profiles, StAR and c-kit expression and androgenic enzyme activities. Spermine hampered EMF-induced oxidative, inflammatory, apoptotic and DNA perturbations. Histological and histomorphometric analysis of the testes supported all biochemical findings. In conclusion, rat exposure to EMF disrupts sperm and hormone profiles with underlying impairment of steroidogenesis and spermatogenesis. Spermine confers protection against EMF-associated testicular and reproductive aberrations, at least in part, via antioxidant, anti-inflammatory and anti-apoptotic mechanisms.

Motawi, T. M. K., N. A. H. Sadik, D. Sabry, N. N. Shahin, and S. A. Fahim, "rs2267531, a promoter SNP within glypican-3 gene in the X chromosome, is associated with hepatocellular carcinoma in Egyptians.", Scientific reports, vol. 9, issue 1, pp. 6868, 2019. Abstract

Hepatocellular carcinoma (HCC) is a major health concern in Egypt owing to the high prevalence of hepatitis C virus (HCV) infection. HCC incidence is characterized by obvious male predominance, yet the molecular mechanisms behind this gender bias are still unidentified. Functional variations in X-linked genes have more impact on males than females. Glypican-3 (GPC3) gene, located in the Xq26 region, has lately emerged as being potentially implicated in hepatocellular carcinogenesis. The current study was designed to examine the association of -784 G/C single nucleotide polymorphism (SNP) in GPC3 promoter region (rs2267531) with HCC susceptibility in male and female Egyptian HCV patients. Our results revealed a significant association between GPC3 and HCC risk in both males and females, evidenced by higher C allele and CC/C genotype frequencies in HCC patients when compared to controls. However, no such association was found when comparing HCV patients to controls. Moreover, GPC3 gene and protein expression levels were significantly higher in CC/C than in GG/G genotype carriers in males and females. The CC/C genotype exhibited a significant shorter overall survival than GG/G genotype in HCC patients. In conclusion, GPC3 rs2267531 on the X chromosome is significantly associated with HCC, but not with HCV infection, in the Egyptian population.

Ali, S. O., N. N. Shahin, M. M. Safar, and S. M. Rizk, "Therapeutic potential of endothelial progenitor cells in a rat model of epilepsy: Role of autophagy", Journal of Advanced Research, vol. 18, pp. 101 - 112, 2019. AbstractWebsite

Epilepsy is one of the most well-known neurological conditions worldwide. One-third of adult epileptic patients do not respond to antiepileptic drugs or surgical treatment and therefore suffer from the resistant type of epilepsy. Stem cells have been given substantial consideration in the field of epilepsy therapeutics. The implication of pathologic vascular response in sustained seizures and the eminent role of endothelial progenitor cells (EPCs) in maintaining vascular integrity tempted us to investigate the potential therapeutic effects of EPCs in a pentylenetetrazole (PTZ)-induced rat model of epilepsy. Modulation of autophagy, a process that enables neurons to maintain an equilibrium of synthesis, degradation and subsequent reprocessing of cellular components, has been targeted. Intravenously administered EPCs homed into the hippocampus and amended the deficits in memory and locomotor activity. The cells mitigated neurological damage and the associated histopathological alterations and boosted the expression of brain-derived neurotrophic factor. EPCs corrected the perturbations in neurotransmitter activity and enhanced the expression of the downregulated autophagy proteins light chain protein-3 (LC-3), beclin-1, and autophagy-related gene-7 (ATG-7). Generally, these effects were comparable to those achieved by the reference antiepileptic drug, valproic acid. In conclusion, EPCs may confer therapeutic effects against epilepsy and its associated behavioural and biochemical abnormalities at least in part via the upregulation of autophagy. The study warrants further research in experimental and clinical settings to verify the prospect of using EPCs as a valid therapeutic strategy in patients with epilepsy.

Shahin, N. N., N. F. Abdelkader, and M. M. Safar, "A Novel Role of Irbesartan in Gastroprotection against Indomethacin-Induced Gastric Injury in Rats: Targeting DDAH/ADMA and EGFR/ERK Signaling.", Scientific reports, vol. 8, issue 1, pp. 4280, 2018 Mar 09. Abstract

The advent of angiotensin II type 1 receptor blockers (ARBs) as intriguing gastroprotective candidates and the superior pharmacokinetics and pharmacodynamics displayed by irbesartan compared to many other ARBs raised the interest to investigate its gastroprotective potential in a rat model of gastric injury. Irbesartan (50 mg/Kg) was orally administered to male Wistar rats once daily for 14 days; thereafter gastric injury was induced by indomethacin (60 mg/Kg, p.o). Irbesartan reduced gastric ulcer index, gastric acidity, and ameliorated indomethacin-induced gastric mucosal apoptotic and inflammatory aberrations, as demonstrated by hampering caspase-3, prostaglandin E and tumor necrosis factor-alpha levels and cyclooxygenase-2 mRNA expression. This ARB increased mucosal dimethylarginine dimethylaminohydrolase-1 (DDAH-1) gene expression and decreased elevated levels of matrix metalloproteinase-9, asymmetric dimethylarginine (ADMA), epidermal growth factor receptor (EGFR) mRNA and phosphorylated extracellular signal-regulated kinase 1 and 2 (pERK1/2). Histopathological evaluation corroborated biochemical findings. Overall efficacy of irbesartan was comparable to ranitidine, the widely used H receptor blocker. In conclusion, irbesartan exerts significant gastroprotection against indomethacin-induced mucosal damage via acid-inhibitory, anti-inflammatory, anti-apoptotic and extracellular matrix remodeling mechanisms that are probably mediated, at least partly, by down-regulating DDAH/ADMA and EGFR/ERK1/2 signaling.

Motawi, T. K., M. R. Mohamed, N. N. Shahin, M. A. M. Ali, and M. A. Azzam, "Time-course expression profile and diagnostic potential of a miRNA panel in exosomes and total serum in acute liver injury.", The international journal of biochemistry & cell biology, vol. 100, pp. 11-21, 2018 Jul. Abstract

Circulating miRNAs have recently emerged as attractive candidates for biomarker discovery. However, they have a variant distribution in circulation, and the diagnostic significance of their compartmentalization is yet to be elucidated. This study explored the time-course expression profile and the diagnostic potential of miRNAs-122a-5p, 192-5p, 193a-3p and 194-5p in exosomal and total serum compartments in two rat models of acute liver injury (ALI). Exosomes were isolated and characterized in terms of morphology, size and CD-63 surface marker expression. Exosomal, serum and hepatic miRNAs were quantified using q-RT-PCR. An inverse expression pattern of hepatic and total serum miRNAs was observed following acetaminophen or thioacetamide-induced liver injury. Conversely, exosomal miRNAs expression pattern varied according to the type of injury. Overall, ROC analysis revealed superior discriminatory ability of exosomal miRNA-122a-5p following either acetaminophen or thioacetamide injury with earlier diagnostic potential and a wider diagnostic window compared to the corresponding total serum counterpart. Moreover, exosomal miRNAs showed higher correlation with ALT activity in both models. In conclusion, exosomal miRNA-122a-5p shows higher diagnostic performance with a broader diagnostic time window and an earlier diagnostic potential than its serum counterpart in ALI. Furthermore, exosomal miRNAs-122a-5p, 192-5p and 193a-3p exhibit an injury-specific signature in ALI and can be used not only as diagnostic tools in liver injury but also to differentiate between different etiologies of injury.

Shahin, N. N., and M. M. Mohamed, "Nano-sized titanium dioxide toxicity in rat prostate and testis: Possible ameliorative effect of morin.", Toxicology and applied pharmacology, vol. 334, pp. 129-141, 2017 11 01. Abstract

This study investigated the effect of short-term oral exposure to nano-sized titanium dioxide (nTiO2) on Wistar rat prostate and testis, and the associating reproductive-related alterations. The study also evaluated the potential ameliorative effect of the natural flavonoid, morin, on nTiO2-induced aberrations. Intragastric administration of nTiO2 (50mg/kg/day for 1, 2 and 3weeks) increased testicular gamma-glutamyltransferase (γ-GT) activity and decreased testicular steroidogenic acute regulatory protein (StAR) and c-kit gene expression, serum testosterone level and sperm count. nTiO2-treated rats also exhibited prostatic and testicular altered glutathione levels, elevated TNF-α levels, up-regulated Fas, Bax and caspase-3 gene expression, down-regulated Bcl-2 gene expression and enhanced prostatic lipid peroxidation. Sperm malformation and elevated testicular acid phosphatase (ACP) activity and malondialdehyde level, serum prostatic acid phosphatase activity, prostate specific antigen (PSA), gonadotrophin and estradiol levels occurred after the 2 and 3week regimens. Morin (30mg/kg/day administered intragastrically for 5weeks) mitigated nTiO2-induced prostatic and testicular injury as evidenced by lowering serum PSA level, testicular γ-GT and ACP activities and TNF-α level, along with hampering both intrinsic and extrinsic apoptotic pathways. Moreover, morin alleviated prostatic lipid peroxidation, raised prostatic glutathione level, and relieved testicular reductive stress. Additionally, morin increased testicular StAR and c-kit mRNA expression, raised the sperm count, reduced sperm deformities and modified the altered hormone profile. Histopathological evaluation supported the biochemical findings. In conclusion, morin could ameliorate nTiO2-induced prostatic and testicular injury and the corresponding reproductive-related aberrations via redox regulatory, anti-inflammatory and anti-apoptotic mechanisms, promoting steroidogenesis and spermatogenesis, and improving sperm count and morphology.

Motawi, T. K., O. G. Shaker, N. N. Shahin, and N. M. Ahmed, "Angiotensin-converting enzyme insertion/deletion polymorphism association with obesity and some related disorders in Egyptian females: a case-control observational study.", Nutrition & metabolism, vol. 13, pp. 68, 2016. Abstract

BACKGROUND: According to the WHO report in 2015, obesity is the fifth leading cause of death worldwide, and the prevalence of Egyptian female obesity is 37.5 %. Since obesity is highly influenced by genetics, and adipose tissue renin-angiotensin system is over-activated in obesity, the effect of angiotensin-converting enzyme (ACE) insertion/deletion (I/D) polymorphism on obesity and related disorders was studied in several populations, because of its effect on ACE activity. Our objective was to study the association of ACE I/D polymorphism with obesity and certain related disorders, namely hypertension, insulin resistance and metabolic syndrome, in Egyptian females.

METHODS: Eighty female volunteers were recruited, blood pressure and body measurements were recorded and a fasting blood sample was obtained for the quantitation of glucose, lipid profile, insulin, leptin and identification of ACE I/D polymorphs. Subjects were grouped based on hypertension and obesity states. Comparisons of continuous parameters were made with independent sample t-test between two groups. The frequencies of ACE genotypes and alleles, and the association between gene polymorphism and metabolic parameters were assessed using chi-square or Fisher's exact test.

RESULTS: Genotype frequencies were in Hardy-Weinberg equilibrium for all groups. Genotype distribution did not differ significantly between controls and cases of all the studied disorders. Although DD carriers had apparently higher parameters of blood pressure, lipid profile and insulin resistance, only diastolic blood pressure was almost significant (p = 0.057). I-carriers were significantly less susceptible to hypertension than DD carriers having normal waist/hip ratio (p = 0.007, OR = 17.29, CI = 1.81-164.96) and normal conicity index (p = 0.024, OR = 7.00, CI = 1.36-35.93). In DD genotype carriers, a significant association was found between insulin resistance and high body mass index (p = 0.004, OR = 8.89, CI = 1.94-40.71), waist circumference (p = 0.003, OR = 9.63, CI = 2.14-43.36) and waist/height ratio (p = 0.034, OR = 6.86, CI = 1.25-37.61), although the variations in percentages between DD and I-carriers were not high enough to conclude an effect of ACE I/D on such an association.

CONCLUSIONS: In this sample of Egyptian females, ACE I/D polymorphism was not significantly associated with obesity nor with any of its related disorders studied. The I allele seemed protective against hypertension in subjects with normal, not high, waist/hip ratio and conicity index compared to DD genotype carriers.