Shehata, N. I., M. A. Abdelsamad, H. A. A. Amin, N. A. H. Sadik, and A. A. Shaheen, "Ameliorating effect of ketogenic diet on acute status epilepticus: Insights into biochemical and histological changes in rat hippocampus.", Journal of food biochemistry, vol. 46, issue 9, pp. e14217, 2022. Abstractjournal_of_food_biochemistry_-_2022_-_shehata_-_ameliorating_effect_of_ketogenic_diet_on_acute_status_epilepticus__insights.pdf

This study aimed to evaluate the potential neuroprotective effects of ketogenic diet (KD) against the neuronal disruptions induced by SE in lithium-pilocarpine rat model of status epilepticus (SE). Four groups of female rats include; groups I and III received standard diet and groups II and IV received KD for 3 weeks. Groups I and II were left untreated, while groups III and IV were injected with LiCl (127 mg/kg, i.p.) followed by pilocarpine HCl (10 mg/kg, i.p.) 18-24 h later, repeatedly, till induction of SE. 72 h post-SE, KD effectively ameliorated the balance between excitatory (glutamate) and inhibitory (GABA) neurotransmitters and the oxidative stress indices, increased adenine nucleotides and decreased immunoreactivity of iNOS, TNFα, glial fibrillary acidic protein, and synaptophysin. Thiswas in association with improvement in inflammatory response and neuronal tissue characteristics in hippocampus of SE rats. Histological changes showed preservation of neuronal integrity. These findings highlight the protective effects of KD in the acute phase post-SE via ameliorating biochemical and histological changes involved. PRACTICAL APPLICATIONS: Epilepsy is the fourth most common neurological disorder that requires lifelong treatment. It stigmatizes patients and their families. The use of the ketogenic diet (KD) as a therapy for epilepsy developed from observations that fasting could reduce seizures. From 1920s, the KD was a common epilepsy treatment until it was gradually superseded by anticonvulsant drugs so that by the 1980s it was rarely used. However, there has been a resurgence of interest and usage of the KD for epilepsy since the turn of the century. Despite its long history, the mechanisms by which KD exhibits its anti-seizure action are not fully understood. Our study aims to identify the mechanism of KD which may help further studies to achieve the same benefits with a drug or supplement to overcome its unpalatability and gastrointestinal side effects.

Shehata, N. I., D. M. Abd El-Salam, R. M. Hussein, and S. M. Rizk, "Effect of safranal or candesartan on 3-nitropropionicacid-induced biochemical, behavioral and histological alterations in a rat model of Huntington's disease.", PloS one, vol. 18, issue 11, pp. e0293660, 2023. Abstractjournal.pone_.0293660.pdf

3-nitropropionic acid (3-NP) is a potent mitochondrial inhibitor mycotoxin. Systemic administration of 3-NP can induce Huntington's disease (HD)-like symptoms in experimental animals. Safranal (Safr) that is found in saffron essential oil has antioxidant, anti-inflammatory and anti-apoptotic actions. Candesartan (Cands) is an angiotensin receptor blocker that has the potential to prevent cognitive deficits. The present study aims to investigate the potential neuroprotective efficacy of Safr or Cands in 3-NP-induced rat model of HD. The experiments continued for nine consecutive days. Rats were randomly assigned into seven groups. The first group (Safr-control) was daily intraperitoneally injected with paraffin oil. The second group (Cands- and 3-NP-control) daily received an oral dose of 0.5% carboxymethylcellulose followed by an intraperitoneal injection of 0.9% saline. The third and fourth groups received a single daily dose of 50 mg/kg Safr (intraperitoneal) and 1 mg/kg Cands (oral), respectively. The sixth group was daily treated with 50 mg Safr kg/day (intraperitoneal) and was intraperitoneally injected with 20 mg 3-NP/ kg, from the 3rd till the 9th day. The seventh group was daily treated with 1 mg Cands /kg/day (oral) and was intraperitoneally injected with 20 mg 3-NP/ kg, from the 3rd till the 9th day. The present results revealed that 3-NP injection induced a considerable body weight loss, impaired memory and locomotor activity, reduced striatal monoamine levels. Furthermore, 3-NP administration remarkably increased striatal malondialdehyde and nitric oxide levels, whereas markedly decreased the total antioxidant capacity. Moreover, 3-NP significantly upregulated the activities of inducible nitric oxide synthase and caspase-3 as well as the Fas ligand, in striatum. On the contrary, Safr and Cands remarkably alleviated the above-mentioned 3-NP-induced alterations. In conclusion, Safr and Cands may prevent or delay the progression of HD and its associated impairments through their antioxidant, anti-inflammatory, anti-apoptotic and neuromodulator effects.

Shehata, N. I., S. M. Abo Zeid, S. A. Abd El Aziz, and H. M. Abdelgawad, "Mitigation of streptozotocin-induced alterations by natural agents via upregulation of PDX1 and Ins1 genes in male rats.", Journal of food biochemistry, vol. 46, issue 5, pp. e14086, 2022. Abstractjournal_of_food_biochemistry_-_2022_-_shehata_-_mitigation_of_streptozotocin-induced_alterations_by_natural_agents_via.pdf

Diabetes is the most common endocrine disorder contributing to high morbidity and mortality globally. Due to the side effects reported for anti-diabetic drugs, there is a rising interest in herbal medicine for diabetes mitigation. However, rare studies were performed to analyze the molecular effects of natural agents on diabetes. Therefore, this study is the first to assess the possible ameliorating effects of avocado and cinnamon extracts on streptozotocin (STZ)-induced disturbances in the gene expression of PDX1 and Ins1 in type-2 diabetic rats, in comparison to metformin. A total number of 50 male Albino rats were randomly divided into five groups; normal control, STZ-induced diabetic group (65 mg/kg b.w divided into three doses 5 days apart), three other STZ-diabetic groups were treated orally for 6 weeks with metformin (500 mg/kg b.w/day), an avocado fruit ethanolic extract (300 mg/kg b.w/day), or a cinnamon aqueous extract (200 mg/kg b.w/day). All rats were fasted overnight then euthanized, blood was collected, and serum was separated for biochemical analyses. The pancreas was processed for the evaluation of PDX1 and Ins1 gene expression by qRT-PCR. Oral administration of the avocado/cinnamon extract significantly improved the altered levels of blood glucose, insulin, lipid profile, and total antioxidant capacity (TAC) and upregulated the pancreatic expression of PDX1 and Ins1 genes. In conclusion, this study added another mechanism of anti-diabetic action for the plant extracts via upregulating the gene expression of PDX1 and Ins1 in STZ-diabetic rats. Metformin has a more profound effect than the plant extracts; however, cinnamon has a comparable effect. PRACTICAL APPLICATIONS: Diabetes is a worldwide chronic serious problem; therefore, different efficacious drug treatments are currently available. However, several side effects of these drugs also negatively affect patient welfare. Therefore, the present study provides an additive/adjunct biochemical rationale for further natural intervention strategy for type-2 diabetes through the introduction of plant-derived products that revealed a wide range of therapeutic effects without causing untoward actions. This study revealed that cinnamon and avocado extracts exhibited marked anti-diabetic and antioxidant effects in rats, as compared to the standard drug, metformin. These results augmented the previous ones and added a new molecular mechanism of action through upregulation of Ins1 and PDX1 in type-2 diabetic rats.

Nossier, A. I., N. I. Shehata, S. M. Morsy, D. F. Saeed, N. M. Elsayed, M. F. Ismail, and S. Eissa, "Determination of certain urinary microRNAs as promising biomarkers in diabetic nephropathy patients using gold nanoparticles.", Analytical biochemistry, vol. 609, pp. 113967, 2020. Abstract

Diabetic nephropathy (DN) is a major leading cause of kidney failure. So, early detection of DN by assessing urinary microRNAs (miRNAs) expression may be of clinical value. In this study, the diagnostic value of two urinary miRNAs (miR-210 & miR-34a) as biomarkers for diagnosis of DN was assessed using a simple colorimetric gold nanoparticle (AuNP) assay and real-time PCR. MiR-(210 & 34a) were markedly up-regulated in DN groups (micro-albuminuric and macro-albuminuric groups) compared to the non-albuminuric group and healthy controls. The sensitivity and specificity for the qualitative detection of urinary miR-(210 & 34a) using the AuNP assay were (78% and 72%) & (81% and 69%), respectively, which were consistent with the results of real-time PCR. There was a highly significant correlation between urinary miR-(210 & 34a) detected by either qRT-PCR or qualitative AuNP assay. Accordingly, this simple AuNP assay may be considered a valid test for the detection of these two urinary miRNAs as potential biomarkers that can aid in the noninvasive diagnosis of DN.

Sana-Eldine, A. O., H. M. Abdelgawad, N. A. H. L. A. S. KOTB, and N. I. Shehata, "The potential effect of Schisandrin-B combination with panitumumab in wild-type and mutant colorectal cancer cell lines: Role of apoptosis and autophagy.", Journal of biochemical and molecular toxicology, vol. 37, issue 5, pp. e23324, 2023. Abstract

Panitumumab is an approved monoclonal antibody for the treatment of colorectal cancer (CRC); however, mutations in EGFR signaling pathway resulted in poor response. Schisandrin-B (Sch-B) is a phytochemical that was suggested to protect against inflammation, oxidative stress, and cell proliferation. The present study aimed to investigate the potential effect of Sch-B on panitumumab-induced cytotoxicity in wild-type Caco-2, and mutant HCT-116 and HT-29 CRC cell lines, and the possible underlying mechanisms. CRC cell lines were treated with panitumumab, Sch-B, and their combination. The cytotoxic effect of drugs was determined by MTT assay. The apoptotic potential was assessed in-vitro by DNA fragmentation and caspase-3 activity. Additionally, autophagy was investigated via microscopic detection of autophagosomes and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) measurement of Beclin-1, Rubicon, LC3-II, and Bcl-2 expression. The drug pair enhanced panitumumab cytotoxicity in all CRC cell lines where IC50 of panitumumab was decreased in Caco-2 cell line. Apoptosis was induced through caspase-3 activation, DNA fragmentation, and Bcl-2 downregulation. Caco-2 cell line treated with panitumumab showed stained acidic vesicular organelles, contrariwise, all cell lines treated with Sch-B or the drug pair displayed green fluorescence indicating the lack of autophagosomes. qRT-PCR revealed the downregulation of LC3-II in all CRC cell lines, Rubicon in mutant cell lines, and Beclin-1 in HT-29 cell line only. Sch-B at 6.5 µM promoted panitumumab-induced apoptotic cell death, in-vitro, via caspase-3 activation and Bcl-2 downregulation, rather than autophagic cell death. This novel combination therapy against CRC, allows the reduction of panitumumab dose to guard against its adverse effects.

Motawi, T. K., N. I. Shehata, M. M. ElNokeety, and Y. F. El-Emady, "Potential serum biomarkers for early detection of diabetic nephropathy.", Diabetes research and clinical practice, vol. 136, pp. 150-158, 2018. Abstract

AIM: Diabetic nephropathy (DN) is considered as one of the diabetic complications affecting up to 40% of patients with type 1 or type 2 diabetes. In clinical practice, the frequently used markers of renal disease and progression are serum creatinine, estimated glomerular filtration rate (eGFR) and albuminuria. The aim of this study is to determine new biomarkers in human serum which are promising for early detection of DN.

METHODS: This study included 50 patients with type 2 diabetes mellitus (T2DM) and 25 clinically healthy individuals. The patients were divided into two groups; group I included 25 T2DM patients with normoalbuminuria, and group II consisted of 25 T2DM patients with microalbuminuria. In all groups, neutrophil gelatinase-associated lipocalin (NGAL), β-trace protein (βTP) and microRNA- 130b (miR-130b) were estimated.

RESULTS: The serum levels of NGAL and βTP were significantly elevated in T2DM patients with microalbuminuria (group II) compared with T2DM patients with normoalbuminuria (group I) and control subjects but there was no significant difference between group I and control subjects. Serum miR-130b level was significantly decreased in patients with T2DM (groups I and II) compared with healthy control subjects, with a higher decrease in their levels in group II compared with group I.

CONCLUSION: Our results suggest that serum NGAL and βTP as tubular and glomerular markers respectively, together with serum miR-130b may be independent and reliable biomarkers for early detection of DN in patients with T2DM.

Motawi, T. M. K., D. Sabry, N. I. Shehata, M. M. William, and A. T. Fahim, "Impact of FOXP1 rs2687201 genetic variant on the susceptibility to HCV-related hepatocellular carcinoma in Egyptians.", Journal of biochemical and molecular toxicology, vol. 36, issue 3, pp. e22965, 2022. Abstract

Hepatocellular carcinoma (HCC) constitutes a challenging health problem in Egypt due to the high incidence of hepatitis C virus (HCV) infection. Improved understanding of genetic mechanisms underlying the individual predisposition to HCC will lead to enhancements in the early diagnosis, treatment, and prevention of this disease. Transcription factor forkhead box P1 (FOXP1) is involved in the cellular processes of proliferation, differentiation, metabolism, and longevity. In addition, it has been implicated in hepatic tumorigenesis. The present study explored the association of C/A single-nucleotide polymorphism in the FOXP1 gene (rs2687201) with HCC susceptibility in HCV Egyptian patients. The study included 108 patients with HCV-dependant HCC, 86 HCV patients, and 80- age and gender-matched healthy controls. rs2687201 genotyping was performed by allelic discrimination method using TaqMan real-time PCR assays while FOXP1 gene expression and protein level were determined using qRT-PCR and enzyme-linked immunoassay, respectively. Our results revealed a significant association between FOXP1 rs2687201 and HCC risk where (A) allele was significantly more frequent in patients with HCC compared to controls (odds ratio [OR]: 1.88, 95% confidence interval [CI]: 1.17-3.04, p = 0.01) and to HCV patients (OR: 1.85, 95% CI: 1.62-2.94, p = 0.012). Furthermore, FOXP1 gene and protein expression levels were remarkably higher in (CA + AA) than in CC genotype carriers in a dominant model. The (CA + AA) genotype displayed a significantly shorter overall survival than the CC genotype in HCC patients. In conclusion, FOXP1 gene polymorphism rs2687201 is significantly associated with HCC, but not with HCV infection, in Egyptian patients.

Gibriel, A. A., M. F. Ismail, H. Sleem, N. Zayed, A. Yosry, S. M. El-Nahaas, and N. I. Shehata, "Diagnosis and staging of HCV associated fibrosis, cirrhosis and hepatocellular carcinoma with target identification for miR-650, 552-3p, 676-3p, 512-5p and 147b.", Cancer biomarkers : section A of Disease markers, vol. 34, issue 3, pp. 413-430, 2022. Abstract

BACKGROUND: Chronic HCV infection progresses to fibrosis, cirrhosis and hepatocellular carcinoma (HCC). The latter represents the third most common cause for cancer mortality. Currently, there is no reliable non-invasive biomarker for diagnosis of HCV mediated disorders.

OBJECTIVE: Profiling expression signature for circulatory miRNAs in the plasma of 167 Egyptian patients (40 healthy, 48 HCV fibrotic, 39 HCV cirrhotic and 40 HCV-HCC cases).

METHODS: QRTPCR was used to quantify expression signature for circulatory miRNAs.

RESULTS: MiR-676 and miR-650 were powerful in discriminating cirrhotic and late fibrosis from HCC. MiR-650 could distinguish mild (f0-f1) and advanced (f2-f3) fibrosis from HCC cases. MiR-650 and miR-147b could distinguish early fibrosis from healthy controls meanwhile miR-676 and miR-147b could effectively distinguish between mild chronic and (f1-f3) cases from healthy individuals. All studied miRNAs, except miR-512, can differentiate between (f0-f3) cases and healthy controls. Multivariate logistic regression revealed three potential miRNA panels for effective differentiation of HCC, cirrhotic and chronic liver cases. MiR-676-3p and miR-512-5p were significantly correlated in (f1-f3) fibrosis meanwhile miR-676 and miR-512 could differentiate between cirrhosis and (f0-f3) cases. Both miR-650 and miR-512-5p were positively correlated in the cirrhotic group and in (f0-f4) group. Putative targets for investigated miRNAs were also determined.

CONCLUSIONS: Investigated miRNAs could assist in staging and diagnosis of HCV associated disorders.

Ahmed, L. A., N. I. Shehata, N. F. Abdelkader, and M. M. Khattab, "Tempol, a Superoxide Dismutase Mimetic Agent, Ameliorates Cisplatin-Induced Nephrotoxicity through Alleviation of Mitochondrial Dysfunction in Mice", PLOS ONE, vol. 9, issue 10 e108889, 2014. AbstractTempol, a Superoxide Dismutase Mimetic Agent, Ameliorates Cisplatin-Induced Nephrotoxicity through Alleviation of Mitochondrial Dysfunction in Mice.pdf

Background: Mitochondrial dysfunction is a crucial mechanism by which cisplatin, a potent chemotherapeutic agent,
causes nephrotoxicity where mitochondrial electron transport complexes are shifted mostly toward imbalanced reactive
oxygen species versus energy production. In the present study, the protective role of tempol, a membrane-permeable
superoxide dismutase mimetic agent, was evaluated on mitochondrial dysfunction and the subsequent damage induced by
cisplatin nephrotoxicity in mice.
Methods and Findings: Nephrotoxicity was assessed 72 h after a single i.p. injection of cisplatin (25 mg/kg) with or without
oral administration of tempol (100 mg/kg/day). Serum creatinine and urea as well as glucosuria and proteinuria were
evaluated. Both kidneys were isolated for estimation of oxidative stress markers, adenosine triphosphate (ATP) content and
caspase-3 activity. Moreover, mitochondrial oxidative phosphorylation capacity, complexes I–IV activities and mitochondrial
nitric oxide synthase (mNOS) protein expression were measured along with histological examinations of renal tubular
damage and mitochondrial ultrastructural changes. Tempol was effective against cisplatin-induced elevation of serum
creatinine and urea as well as glucosuria and proteinuria. Moreover, pretreatment with tempol notably inhibited cisplatininduced
oxidative stress and disruption of mitochondrial function by restoring mitochondrial oxidative phosphorylation,
complexes I and III activities, mNOS protein expression and ATP content. Tempol also provided significant protection
against apoptosis, tubular damage and mitochondrial ultrastructural changes. Interestingly, tempol did not interfere with
the cytotoxic effect of cisplatin against the growth of solid Ehrlich carcinoma.
Conclusion: This study highlights the potential role of tempol in inhibiting cisplatin-induced nephrotoxicity without
affecting its antitumor activity via amelioration of oxidative stress and mitochondrial dysfunction.

Eldin, A. A., A. A. Shaheen, H. M. Abd-Elgawad, and N. I. Shehata, "Protective effect of taurine and quercetin against renal dysfunction associated with the combined use of gentamycin and diclofenac.", Indian J Biochem Biophys, vol. 45, issue 5, pp. 332-340, 2008. Abstract

The potential protective effects of taurine and quercetin against gentamycin (GM)/diclofenac (DC) combined nephrotoxicity were investigated in rats. The results showed that administration of DC alone at an oral dose of 5 mg/kg b.wt/day for 28 days had no significant effect on the measured parameters, except for marked increase in urinary uronic acid excretion. Administration of GM alone at a dose of 100 mg/kg b.wt/day i.p. for 8 days resulted in obvious nephrotoxicity. Combined GM-DC treatment led to the most pronounced nephrotoxicity, as indicated by greater elevations in serum urea, creatinine and urinary N-acetyl-beta-D-glucosaminidase (NAG), together with severe depression of renal cortical Na , K+-ATPase, compared to GM-treated group. Moreover, only combined treatment resulted in significant decrease in urinary potassium and renal cortical glutathione peroxidase (GSHPx), together with an increase in renal cortical lipid peroxidation products (LPOs). Co-administration of taurine or quercetin normalized creatinine clearance and ameliorated the elevations in urinary proteins, uronic acids, NAG and renal cortical LPOs in GM/DC treated rats. The study justifies the use of taurine and quercetin as renoprotective agents against the nephrotoxicity caused by GM/DC therapy.

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