Akmal A. El-Ghor

β-Glucan inhibits the genotoxicity of cyclophosphamide, adriamycin and cisplatin

The inhibitory effects of beta-glucan (betaG), one of the biological response modifiers, on the induction of chromosomal aberrations in the bone marrow and spermatogonial cells of mice treated with various anti-neoplastic drugs were investigated. beta-Glucan (100 mg/kg bw, i.p.) pre-treatment reduced the total number of cells with structural chromosomal aberrations scored after the treatment with cyclophosphamide (CP) (2.5 mg/kg bw, i.p.) adriamycin (ADR) (12 mg/kg bw, i.p.) and cis-diamminedichloroplatinum-II (cisplatin) (5 mg/kg bw, i.p.) by about 41.1, 26.9 and 57.7% in bone marrow and 44.4, 55 and 57.1% in spermatogonial cells, respectively. This protective effect of beta-glucan could be attributed to its scavenging ability to trap free-radicals produced during the biotransformation of these anti-neoplastic drugs. Beta-glucan also markedly restored the mitotic activity of bone marrow cells that had been suppressed by the anti-neoplastic drugs. These results indicate that in addition to the known immunopotentiating activity of beta-glucan, it plays a role in reducing genotoxicity induced by anti-neoplastic drugs during cancer chemotherapy.

Microsatellite instability at three microsatellite loci (D6mit3, D9mit2 and D15Mgh1) located in different common fragile sites of rats exposed to cadmium

Cadmium (Cd) is a non-essential element and is a widespread environmental pollutant. Exposure to cadmium can result in cytotoxic, carcinogenic and mutagenic effects. Mutagenesis is an indicative of genetic instability and can be assayed using microsatellites instability. The aim of the present study is to investigate; based on the rat model, the effects of oral acute (single 8.8mg/kg BW, 1/10 LD(50)) and sub-chronic (2.93mg/kg BW, 1/30 LD(50), for 4 weeks) doses of cadmium chloride on microsatellite instability at D6mit3, D9mit2 and D15Mgh1 loci, which are located in three different common fragile sites (6q21, 9q32-q33 and 15p14, respectively), within rat genome. In the acute study, no microsatellite instability (MSI) was observed in all the three tested loci (D6mit3, D9mit2 and D15Mgh1). In the sub-chronic study, the MSI were observed in the three studied loci and was in the form of deletion of 2-3bp or addition of 3-6bp. These finding may indicate the sensitivity of microsatellite sequences located at the fragile sites and the sensitivity of the simple sequence repeats (SSRs) assay for the detection of small variations in DNA sequence. However, additional chronic toxicological trials are needed in order to assess genotoxic effects of chronic exposure to Cd.

Normalization of Nano-Sized TiO2-Induced Clastogenicity, Genotoxicity and Mutagenicity by Chlorophyllin Administration in Mice Brain, Liver, and Bone Marrow Cells

The intensive uses of titanium dioxide (TiO2) nanoparticles in sunscreens, toothpaste, sweats, medications, etc. making humans exposed to it daily by not little amounts and also increased its risks including genotoxicity. Thus, the present study was designed as one way to reduce nano-titanium-induced clastogenicity, genotoxicity, and mutagenicity in mice by co-administration of the free radical scavenger chlorophyllin (CHL). In addition, markers of oxidative stress were detected to shed more light on mechanism(s) underlying nano-sized TiO2 genotoxicity. Male mice were exposed to multiple injection into the abdominal cavity for five consecutive days with either CHL (40 mg/kg bw/day), or each of three dose levels of nano-sized TiO2 (500, 1000, or 2000 mg/kg bw/day) alone, or both simultaneously and sacrificed by cervical dislocation 24 h after the last treatment. After CHL co-administration, the observed dose-dependent genotoxicity of TiO2 nanoparticles indicated by the significant elevations in frequencies of both micronuclei and DNA damage induction was significantly decreased and returned to the negative control level. The observed induced mutations in p53 exons 5, 7, & 8 and 5 & 8 in the liver and brain, respectively, were declined in most cases. Moreover, CHL significantly decreased hepatic malondialdehyde level and significantly increased glutathione level and superoxide dismutase, catalase, and glutathione peroxidase activities that were significantly disrupted in animal groups treated with nano-TiO2 alone. In conclusion, the evidenced in vivo genotoxicity of nano-TiO2 in the present study was normalized after CHL co-administration which supports the previously suggested oxidative stress as the possible mechanism for titanium toxicity.

Magnetite nanoparticles inhibit tumor growth and upregulate the expression of p53/p16 in Ehrlich solid carcinoma bearing mice.

Background Magnetite nanoparticles (MNPs) have been widely used as contrast agents and have promising approaches in cancer treatment. In the present study we used Ehrlich solid carcinoma (ESC) bearing mice as a model to investigate MNPs antitumor activity, their effect on expression of p53 and p16 genes as an indicator for apoptotic induction in tumor tissues. Method MNPs coated with ascorbic acid (size: 25.0±5.0 nm) were synthesized by co-precipitation method and characterized. Ehrlich mice model were treated with MNPs using 60 mg/Kg day by day for 14 injections; intratumorally (IT) or intraperitoneally (IP). Tumor size, pathological changes and iron content in tumor and normal muscle tissues were assessed. We also assessed changes in expression levels of p53 and p16 genes in addition to p53 protein level by immunohistochemistry. Results Our results revealed that tumor growth was significantly reduced by IT and IP MNPs injection compared to untreated tumor. A significant increase in p53 and p16 mRNA expression was detected in Ehrlich solid tumors of IT and IP treated groups compared to untreated Ehrlich solid tumor. This increase was accompanied with increase in p53 protein expression. It is worth mentioning that no significant difference in expression of p53 and p16 could be detected between IT ESC and control group. Conclusion MNPs might be more effective in breast cancer treatment if injected intratumorally to be directed to the tumor tissues.

Evaluation of the role of the antioxidant silymarin in modulating the in vivo genotoxicity of the antiviral drug ribavirin in mice.

Ribavirin (1-β-d-ribofuranosyl-1,2,4-triazole-3-carboxamide) is a widely used broad-spectrum antiviral drug. Recently, several reports revealed genotoxic effects of ribavirin in vivo and in vitro, which were correlated with the production of reactive oxygen species (ROS). This study aimed to evaluate the genotoxicity of ribavirin and to investigate the role of the natural antioxidant silymarin to modulate this genotoxicity. Male albino mice (age, 8-10 weeks) were injected intraperitoneally (i.p.) with ribavirin at three dose levels (20, 75 and 130 mg/kg bw) either in a single injection (acute treatment) or in multiple injections on five consecutive days (sub-acute treatment). Other comparable groups were treated with silymarin (70 mg/kg bw) 1h before the injection with ribavirin. Mice were sacrificed at different sampling times (24, 48 and 72 h) after the last ribavirin treatment. Micronucleus (MN) and single-strand conformation polymorphism (SSCP) assays were used to assess genotoxic and cytotoxic effects of ribavirin and to evaluate the protective effect of the pre-treatment with silymarin. Our results reveal genotoxic and cytotoxic effects of ribavirin in the MN assay. Pre-treatment with silymarin reduced the toxicity of ribavirin. In the SSCP assay, ribavirin treatment did not induce any mutations in the two selected sites in the D-loop of mitochondrial DNA (mtDNA).

Genotoxic evaluation for the tricyclic antidepressant drug, amitriptyline

Tryptizol® [amitriptyline HCl (AT); El-Kahira Pharmacological and Chemical Co., Cairo, Egypt], a widely used antidepressant drug in Egypt, was evaluated for its genotoxicity. The evaluation was performed in somatic (bone marrow) and germ (spermatocytes) cells, as well as well as the sperm morphology (i.e., head and tail) and count of the resulting sperm. Three doses were tested (low, medium, and high); they were chosen according to the drug manufacturer. The low-dose group received orally 1 mg/kg body weight (b.w.) daily for a total period of 1 month; the medium-dose group received 1 mg/kg b.w. daily for 15 days and 2 mg/kg b.w. daily for another 15 days; and the high-dose group received 1 mg/kg b.w. daily for 10 days, then 2 mg/kg b.w. daily for another 10 days and, finally, 4 mg/kg b.w. daily for 10 more days. The results showed that AT treatment induced structural and numerical chromosome abnormalities in somatic cells (bone marrow) and germ cells (spermatocytes). Moreover, AT significantly reduced both the mitotic index and meiotic activity after the different treatments used. AT was found to increase significantly the incidence of sperm-cell head and tail abnormalities. The sperm-cell count was also significantly decreased with the 3 doses tested. In general, results of chromosome abnormalities in both somatic and germ cells as well as sperm morphology and count showed that the effect of AT was dose dependent. The results of the current study showed that AT is a genotoxic agent for both somatic and germ cells and should be taken under special precautions and medical supervision.

Lead acetate and arsenic trioxide induce instability of microsatellites at three different fragile sites (6q21, 9q32-9q33 and 15p14) within the genome of the rat.

Human exposure to metals is of increasing concern due to the well-documented toxic and carcinogenic effects of metals and metal compounds, and the rising environmental levels due to industrial processes and pollution. It has been reported that metals can be genotoxic by several modes of action including generation of reactive oxygen species and inhibition of DNA repair. The aim of this study was to evaluate microsatellite instability (MSI) in three microsatellite loci (D6mit3, D9mit2 and D15Mgh1) located within three common fragile sites in the genome of the laboratory rat (6q21, 9q32-9q33 and 15p14) exposed to acute and chronic doses of a metal salt (lead acetate trihydrate) and a metalloid oxide (arsenic trioxide). In the acute and sub-chronic studies with the two compounds, MSI was observed in the three loci as deletions or additions of microsatellite repeats. The percentages of MSI were 36.4% and 42.1% for lead acetate and arsenic trioxide, respectively. Results of the present work indicate that the microsatellites located within fragile sites provide a convenient assay system to detect changes in DNA sequences resulting from exposure to genotoxic agents.

Fibulin-2 is required for basement membrane integrity of mammary epithelium

Fibulin-2 (FBLN2) is a secreted extracellular matrix glycoprotein which has been associated with tissue development and remodelling. In the mouse mammary gland, FBLN2 can be detected during ductal morphogenesis in cap cells and myoepithelial cells at puberty and early pregnancy, respectively. In an attempt to assign its function, we knocked down Fbln2 in the mouse mammary epithelial cell line EpH4. FBLN2 reduction led to an increase in the size of spheroidal structures when compared to scrambled control shRNA-transduced cells plated on Matrigel matrix. This phenotype was associated with a disruption of the collagen IV sheath around the epithelial spheroids and downregulation of integrin β1, suggesting a role for FBLN2 in stabilizing the basement membrane (BM). In contrast to mice, in normal adult human breast tissue, FBLN2 was detected in ductal stroma, and in the interlobular stroma, but was not detectable within the lobular regions. In tissue sections of 65 breast cancers FBLN2 staining was lost around malignant cells with retained staining in the neighbouring histologically normal tissue margins. These results are consistent with a role of FBLN2 in mammary epithelial BM stability, and that its down-regulation in breast cancer is associated with loss of the BM and early invasion.

Bone marrow derived-mesenchymal stem cells downregulate IL17A dependent IL6/STAT3 signaling pathway in CCl4-induced rat liver fibrosis

Therapeutic potential of bone marrow–derived mesenchymal stem cells (BM-MSCs) has been reported in several animal models of liver fibrosis. Interleukin (IL) 17A, IL6 and Stat3 have been described to play crucial roles in chronic liver injury. However, the modulatory effect of MSCs on these markers was controversial in different diseases. BM-MSCs might activate the IL6/STAT3 signaling pathway and promote cell invasion in hepatocellular carcinoma, but the immunomodulatory role of BM-MSCs on IL17A/IL6/STAT3 was not fully elucidated in liver fibrosis. In the present study, we evaluated the capacity of the BM-MSCs in the modulation of cytokines milieu and signal transducers, based on unique inflammatory genes Il17a and Il17f and their receptors Il17rc and their effect on the IL6/STAT3 pathway in CCl4-induced liver fibrosis in rats. A single dose of BM-MSCs was administered to the group with induced liver fibrosis, and the genes and proteins of interest were evaluated along six weeks after treatment. Our results showed a significant downregulation of Il17a, Il17ra, il17f and Il17rc genes. In accordance, BM-MSCs administration declined IL17, IL2 and IL6 serum proteins and downregulated IL17A and IL17RA proteins in liver tissue. Interestingly, BM-MSCs downregulated both Stat3 mRNA expression and p-STAT3, while Stat5a gene was downregulated and p-STAT5 protein was elevated. Also P-SMAD3 and TGFβR2 proteins were downregulated in response to BM-MSCs treatment. Collectively, we suggest that BM-MSCs might play an immunomodulatory role in the treatment of liver fibrosis through downregulation of IL17A affecting IL6/STAT3 signaling pathway.