Gouda K. Helal

Influence of p-coumaric acid on doxorubicin-induced oxidative stress in rat’s heart

The therapeutic value of doxorubicin (DOX) as anticancer antibiotic is limited by its cardiotoxicity. The implication of natural phenolic acids in the prevention of many pathologic diseases has been reported. Herein, the ability of p-coumaric (PC) acid, a member of phenolic acids, to protect rats heart against DOX-induced oxidative stress was investigated. Three main groups of albino rats were used; DOX, PC, and PC plus DOX-receiving animals. Corresponding control animals were also used. DOX was administered i.p. in a single dose of 15mgkg(-1). PC alone, in a dose of 100mgkg(-1), was orally administered for five consecutive days. In PC/DOX group, rats received PC 5 days prior to DOX. DOX-induced high serum levels of lactic dehydrogenase (LDH) and creatine phosphokinase (CPK), were reduced significantly by PC administration, compared to DOX-receiving rats. Pretreatment with PC ameliorated the cardiac content of glutathione (GSH), and superoxide dismutase (SOD) & catalase (CAT) activities, compared to DOX-receiving rats. On the other hand, accumulation of cardiac content of MDA significantly decreased following PC pretreatment, compared to DOX-treated rats. The data presented here indicate that PC protects rats hearts against DOX-induced oxidative stress in the heart. It may be worthy to consider the usefulness of PC as adjuvant therapy in cancer management.

Exosomes: From garbage bins to promising therapeutic targets

Intercellular communication via cell-released vesicles is a very important process for both normal and tumor cells. Cell communication may involve exosomes, small vesicles of endocytic origin that are released by all types of cells and are found in abundance in body fluids, including blood, saliva, urine, and breast milk. Exosomes have been shown to carry lipids, proteins, mRNAs, non-coding RNAs, and even DNA out of cells. They are more than simply molecular garbage bins, however, in that the molecules they carry can be taken up by other cells. Thus, exosomes transfer biological information to neighboring cells and through this cell-to-cell communication are involved not only in physiological functions such as cell-to-cell communication, but also in the pathogenesis of some diseases, including tumors and neurodegenerative conditions. Our increasing understanding of why cells release exosomes and their role in intercellular communication has revealed the very complex and sophisticated contribution of exosomes to health and disease. The aim of this review is to reveal the emerging roles of exosomes in normal and pathological conditions and describe the controversial biological role of exosomes, as it is now understood, in carcinogenesis. We also summarize what is known about exosome biogenesis, composition, functions, and pathways and discuss the potential clinical applications of exosomes, especially as biomarkers and novel therapeutic agents.

Exosomal miR-940 maintains SRC-mediated oncogenic activity in cancer cells: A possible role for exosomal disposal of tumor suppressor miRNAs

Exosomes have emerged as important mediators of diverse biological functions including tumor suppression, tumor progression, invasion, immune escape and cell-to-cell communication, through the release of molecules such as mRNAs, miRNAs, and proteins. Here, we identified differentially expressed exosomal miRNAs between normal epithelial ovarian cell line and both resistant and sensitive ovarian cancer (OC) cell lines. We found miR-940 as abundant in exosomes from SKOV3-IP1, HeyA8, and HeyA8-MDR cells. The high expression of miR-940 is associated with better survival in patients with ovarian serous cystadenocarcinoma. Ectopic expression of miR-940 inhibited proliferation, colony formation, invasion, and migration and triggered G0/G1 cell cycle arrest and apoptosis in OC cells. Overexpression of miR-940 also inhibited tumor cell growth in vivo. We showed that proto-oncogene tyrosine-protein kinase (SRC) is directly targeted by miR-940 and that miR-940 inhibited SRC expression at mRNA and protein levels. Following this inhibition, the expression of proteins downstream of SRC, such as FAK, paxillin and Akt was also reduced. Collectively, our results suggest that OC cells secrete the tumor-suppressive miR-940 into the extracellular environment via exosomes, to maintain their invasiveness and tumorigenic phenotype.

Protective effect of N-acetylcysteine against carmustine-induced myelotoxicity in rats.

Carmustine (BCNU) is used to treat a variety of tumors, in particular gliomas. However, the success of such treatment is limited by severe myelosuppression. The role of N-acetylcysteine (NAC) in protection against BCNU-induced myelotoxicity is still needed to be explored. The aim of this work is to study the possible protective role of NAC against BCNU-induced myelotoxicity through evaluation of apoptosis, lipid peroxidation, antioxidant enzymes (superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase(CAT)) as well as glutathione (GSH) content in bone marrow cells of rats. Administration of BCNU in a single dose (30 mg/kg, i.p.) significantly decreased RBCs, WBCs and platelets counts as well as hemoglobin level. In addition, BCNU produced a significant apoptotic effect as well as a significant lipid peroxidation in bone marrow cells. Pretreatment of animals with NAC (150 mg/kg, i.p.) daily for 5 days significantly ameliorated the changes in oxidant and antioxidant parameters as well as apoptosis induced by BCNU. In addition the pattern of blood parameters was shifted markedly to normal values in animals pretreated with NAC when compared to BCNU-treated group. Conclusively, NAC could have a potential protective effect against BCNU-induced myelotoxicity; an effect that is mainly attributed to the antioxidant property.

Systemic administration of Zn2+ during the reperfusion phase of transient cerebral ischaemia protects rat hippocampus against iron-catalysed postischaemic injury.

1 The aim of the present study was to test the protective role of intravenous Zn2+ against iron‐catalysed reperfusion injury in the hippocampus of ischaemic rats. 2 One hundred adult male Wistar albino rats were randomly divided into five groups. Rats in the first group were subjected to surgery (sham operation) without induction of cerebral ischaemia and injected with normal saline (i.v.). The second group of sham‐operated rats were injected with 6 mg/kg, i.v., ZnCl2. In the third group, rats were subjected to cerebral ischaemia for 60 min. Animals in the fourth group were subjected to cerebral ischaemia for 60 min followed by 8 h reperfusion. In the fifth group, rats were subjected to cerebral ischaemia for 60 min, followed by 8 h reperfusion with injection of a single dose of ZnCl2 (6 mg/kg, i.v.) during the first 5 min of the reperfusion period. After reperfusion, animals were killed, their brains were dissected out on ice and the two hippocampi from each animal were isolated and analysed. 3 Cerebral ischaemia induced an increase in the iron content, lipidic peroxidation, apoptosis and metallothionein (MT) in the hippocampus. These effects were significantly increased in the hippocampus of ischaemic rats subjected to 8 h reperfusion compared with ischaemic non‐reperfused rats. Intravenous administration of ZnCl2 decreased the accumulation of iron, lipidic peroxidation and apoptosis produced by reperfusion, but increased the level of MT. 4 Data from the present study suggest that, after 1 h ischaemia, there is an increase in the permeability of the blood–brain barrier and this allows penetration of i.v. injected ZnCl2, which can induce expression of brain MT, increase the anti‐oxidant capacity and diminish iron‐catalysed lipid peroxidation and apoptosis. This may give new insights as to how to improve the outcome for stroke patients.

Carbon tetrachloride-induced liver injury in mice is tissue factor dependent

Tissue factor (TF) is a membranous glycoprotein that activates the coagulation system when blood vessels or tissues are damaged. TF was up-regulated in monocrotaline (MCT)/lipopolysaccharide (LPS) hepatotoxicity model. The present study aimed to test the hypothesis that TF-dependent fibrin deposition occurs in liver toxicity induced by CCl4 in mice. Pericentral deposition of TF and fibrin is induced after CCl4-induced liver toxicity. The toxicity was evaluated by determination of serum activities of ALT, AST and ALP as well as GSH content and histopathological changes. The results showed that injection of mice with TF-antisense deoxyoligonucleotide (TF-AS) prevented the accumulation of TF and fibrin in the hepatic tissues. Furthermore, it significantly restored blood biochemical parameters, GSH content and distorted histopathological features caused by CCl4. The current study demonstrates that TF activation is associated with CCl4-induced liver injury. Furthermore, administration of TF-AS successfully prevented this type of liver injury.

Hydroxychloroquine augments early virological response to pegylated interferon plus ribavirin in genotype-4 chronic hepatitis C patients.

The therapeutic effect of pegylated interferon (peg‐IFN) alfa‐2a combined with ribavirin (RBV) on chronic hepatitis C Egyptian patients is low and further efforts are required to optimize this therapy for achievement of higher rates of virological response. This study aimed to evaluate the safety and efficacy of hydroxychloroquine (HCQ) in combination with pegylated interferon plus ribavirin on early virological response (EVR) in chronic hepatitis C Egyptian patients. Naïve 120 Egyptian patients with chronic hepatitis C virus infection were divided into two groups. Group 1 have administered the standard of care therapy (pegylated interferon alfa‐2a plus ribavirin) for 12 weeks, (n = 60). Group 2 have administered hydroxychloroquine plus standard of care therapy for 12 weeks, (n = 60). Therapeutics included hydroxychloroquine (200 mg) oral twice daily, peginterferon alfa‐2a (160 μg) subcutaneous once weekly and oral weight‐based ribavirin (1000–1200 mg/day). Baseline characteristics were similar in the two groups. The percentage of early virological response was significantly more in patients given the triple therapy than in patients given the standard of care [54/60 (90%) vs. 43/60 (71.7%); P = 0.011; respectively]. Biochemical response at week 12 was also significantly higher in patients given the triple therapy compared with the standard of care [58/60 (96.7%) vs. 42/60 (70%); P < 0.001; respectively]. Along the study, the observed adverse events were mild and similar across treatment groups. Addition of hydroxychloroquine to pegylated interferon plus ribavirin improves the rate of early virological and biochemical responses in chronic hepatitis C Egyptian patients without an increase in adverse events. J. Med. Virol. 88:2170–2178, 2016.

Probing the putative α7 nAChR/NMDAR complex in human and murine cortex and hippocampus: Different degrees of complex formation in healthy and Alzheimer brain tissue

α7 nicotinic acetylcholine receptors (nAChRs) and N-methyl-D-aspartate receptors (NMDARs) are key mediators of central cholinergic and glutamatergic neurotransmission, respectively. In addition to numerous well-established functional interactions between α7 nAChRs and NMDARs, the two receptors have been proposed to form a multimeric complex, and in the present study we have investigated this putative α7 nAChR/NMDAR assembly in human and murine brain tissues. By α-bungarotoxin (BGT) affinity purification, α7 and NMDAR subunits were co-purified from human and murine cortical and hippocampal homogenates, substantiating the notion that the receptors are parts of a multimeric complex in the human and rodent brain. Interestingly, the ratios between GluN1 and α7 levels in BGT pull-downs from cortical homogenates from Alzheimer’s disease (AD) brains were significantly lower than those in pull-downs from non-AD controls, indicating a reduced degree of α7 nAChR/NMDAR complex formation in the diseased tissue. A similar difference in GluN1/α7 ratios was observed between pull-downs from cortical homogenates from adult 3xTg-AD and age-matched wild type (WT) mice, whereas the GluN1/α7 ratios determined in pull-downs from young 3xTg-AD and age-matched WT mice did not differ significantly. The observation that pretreatment with oligomeric amyloid-β1–42 reduced GluN1/α7 ratios in BGT pull-downs from human cortical homogenate in a concentration-dependent manner provided a plausible molecular mechanism for this observed reduction. In conclusion, while it will be important to further challenge the existence of the putative α7 nAChR/NMDAR complex in future studies applying other methodologies than biochemical assays and to investigate the functional implications of this complex for cholinergic and glutamatergic neurotransmission, this work supports the formation of the complex and presents new insights into its regulation in healthy and diseased brain tissue.

Functional characterization of α7 nicotinic acetylcholine and NMDA receptor signaling in SH-SY5Y neuroblastoma cells in an ERK phosphorylation assay

&NA; In the present study, the functional properties of &agr;7 nicotinic acetylcholine receptors (&agr;7 nAChRs) and N‐methyl‐D‐aspartate receptors (NMDARs) endogenously expressed in SH‐SY5Y human neuroblastoma cells were characterized in an extracellular‐signal regulated kinase (ERK) phosphorylation assay. Both choline and N‐methyl‐D‐aspartate (NMDA) mediated robust concentration‐dependent increases in ERK phosphorylation in the SH‐SY5Y cells, exhibiting EC50 values in good agreement with those reported for the agonists at recombinant &agr;7 nAChRs and NMDARs, respectively. Importantly, the responses evoked by choline (10 mM) and by NMDA (50 &mgr;M) were significantly inhibited by the &agr;7‐selective antagonist &agr;‐bungarotoxin (100 nM) and by the NMDAR‐selective antagonist MK‐801 (50 &mgr;M), respectively. The increased ERK phosphorylation levels observed upon co‐application of choline (1, 3, 10 mM) and NMDA (50 &mgr;M) compared to those produced by the two agonists on their own were fully reconcilable with additive effects and did not reveal substantial synergy between &agr;7 nAChR and NMDAR signaling. Interestingly, however, the responses evoked by the “choline (10 mM) – NMDA (50 &mgr;M)” combination were almost completely inhibited by &agr;‐bungarotoxin (100 nM) as well as by MK‐801 (50 &mgr;M), suggesting some sort of a link between &agr;7 nAChR‐ and NMDAR‐mediated ERK phosphorylation. Finally, oligomeric amyloid‐&bgr;1–42 peptide (1000 nM) mediated robust inhibition of the ERK phosphorylation induced by choline (10 mM), NMDA (50 &mgr;M) and the “choline (10 mM) ‐ NMDA (50 &mgr;M)” combination. In conclusion, ERK phosphorylation measurements in SH‐SY5Y cells provides a robust assay for studies of &agr;7 nAChR‐ and NMDAR‐mediating signaling and putative functional interactions between the receptors.

Coadministration of ezetimibe with pegylated interferon plus ribavirin could improve early virological response in chronic hepatitis C obese Egyptian patients.

Background Ezetimibe has been reported to inhibit viral entry and to reduce BMI and has been proposed as a novel therapeutic agent for chronic hepatitis C (CHC), potentiating the effects of pegylated interferon and ribavirin (peg-IFN/RBV). Objective The aim of the study was to assess the effects of ezetimibe coadministration with peg-IFN/RBV combination on the early virological response (EVR) rates in nonobese and obese patients with CHC genotype 4 (CHC-4). Patients and methods A total of 144 CHC-4 patients were divided into two groups; group 1 included nonobese patients (n=76) and group 2 included obese patients (n=68). Each group was further subclassified into equal control and treated groups. The control groups received peg-IFN/RBV combination for 24 weeks, and the treated groups received peg-IFN/RBV plus ezetimibe for 12 weeks and then only peg-IFN/RBV for the remaining 12 weeks. Results The study revealed that EVR significantly improved in the obese patients (85.3 vs. 64.7% in the treated and control groups, respectively, at P<0.05) without any significant improvement in the nonobese patients. Biochemical responses (defined as normalization of alanine aminotransferase at week 12) were markedly improved in the treated groups in both the nonobese and obese groups compared with their respective controls. Conclusion The addition of ezetimibe to peg-IFN/RBV combination significantly improves EVR rates in obese patients compared with nonobese patients, and remarkably improves the biochemical responses in both obese and nonobese patients with CHC-4. This may shed light on a new strategy for the treatment of CHC, particularly in obese Egyptian patients.