ABSTRACT
Supplements containing pharmacological concentrations of biotin are commercially available. The mechanisms by which biotin at pharmacological concentrations exerts its action have been the subject of multiple investigations, particularly for biotin's medicinal potential and wide use for cosmetic purposes. Several studies have reported that biotin supplementation increases cell proliferation; however, the mechanisms involved in this effect have not yet been characterized. In a previous study, we found that a biotin-supplemented diet increased spermatogonia proliferation. The present study was focused on investigating the molecular mechanisms involved in biotin-induced testis cell proliferation. Male BALB/cAnNHsd mice were fed a control or a biotin-supplemented diet (1.76 or 97.7 mg biotin/kg diet) for eight weeks. Compared with the control group, the biotin-supplemented mice presented augmented protein abundance of the c-kit-receptor and pERK1/2Tyr204 and pAKTSer473, the active forms of ERK/AKT proliferation signaling pathways. No changes were observed in the testis expression of the stem cell factor and in the serum levels of the follicle-stimulating hormone. Analysis of mRNA abundance found an increase in cyclins Ccnd3, Ccne1, Ccna2; Kinases Cdk4, Cdk2; and E2F; and Sp1 & Sp3 transcription factors. Decreased expression of cyclin-dependent kinase inhibitor 1a (p21) was observed but not of Cdkn2a inhibitor (p16). The results of the present study identifies, for the first time, the mechanisms associated with biotin supplementation-induced cell proliferation, which raises concerns about the effects of biotin on male reproductive health because of its capacity to cause hyperplasia, especially because this vitamin is available in large amounts without regulation.
Subject(s)
Biotin/toxicity , Cell Proliferation/drug effects , Dietary Supplements/toxicity , Follicle Stimulating Hormone/blood , Spermatogonia/drug effects , Stem Cell Factor/metabolism , Testis/drug effects , Animals , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Male , Mice, Inbred BALB C , Mitogen-Activated Protein Kinases/metabolism , Phosphorylation , Proto-Oncogene Proteins c-akt/metabolism , Proto-Oncogene Proteins c-kit/genetics , Proto-Oncogene Proteins c-kit/metabolism , Signal Transduction , Sp1 Transcription Factor/genetics , Sp1 Transcription Factor/metabolism , Sp3 Transcription Factor/genetics , Sp3 Transcription Factor/metabolism , Spermatogonia/metabolism , Spermatogonia/pathology , Testis/metabolism , Testis/pathologyABSTRACT
Toxicological studies so far suggest that excessive use of malathion, an organophosphate insecticide, causes serious ill-effects in mammalian reproductive physiology. The present study aims at assessing malathion-induced toxicity in a dose- and time-dependent manner with mitigating effects of N-acetyl-l-cysteine. The testicular germ cell viability was monitored using MTT assay, where NAC, being an antioxidant significantly reduced malathion-induced toxicity by enhancing the frequency of cell viability. The histomorphological analysis showed that NAC successfully diminished several apoptotic features in testicular cells, induced by malathion. The differential EB/AO staining revealed a significant decline in the percentage of apoptosis after NAC supplementation. NAC also diminished the malathion-induced DNA fragmentation along with significantly reduction in oxidative stress parameters causing decrease in lipid peroxidation and enhancement of ferric reducing antioxidant power within testicular germ cells. Thus, NAC mitigated the malathion-induced toxicity, proving its potential in infertility treatment.
Subject(s)
Acetylcysteine/pharmacology , Apoptosis/drug effects , Malathion/toxicity , Spermatogonia/metabolism , Testis/metabolism , Animals , Goats , Male , Spermatogonia/pathology , Testis/pathologyABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Moringa oleifera Lam. is widely cultivated in Asian and African countries for its medicinal and dietary significance. The leaves are highly nutritious and are known to possess various biological activities. MATERIALS AND METHODS: Pre-pubertal Swiss albino male mice were injected with single dose of cyclophosphamide (CP, 200mg/kg body weight) or ethanolic extract of Moringa oleifera leaves (MOE, 100mg/kg body weight) intraperitoneally. In combination group, MOE was administered 24h prior to CP injection. RESULTS: CP induced a significant decrease in testicular weight (p<0.01) and depletion of germ cells (p<0.001) and higher level of DNA damage (p<0.001) compared to control. The expression of P53, Bax, Cytochrome C (Cyt C) was increased while there was a decrease in the expression of Bcl2, c-Kit and Oct4. Administration of MOE 24h prior to CP treatment ameliorated the depletion (p<0.001), DNA damage (p<0.001) and apoptosis (p<0.01) of germ cells induced by CP. The mitigating effect of MOE appears to be mediated by up-regulating the expression of c-Kit and Oct4 transcripts in P53-independent manner. CONCLUSION: MOE protects the spermatogonial cells from CP-induced damage by modulating the apoptotic response elicited by CP and therefore can be considered as an efficient method of male fertility preservation.
Subject(s)
Cyclophosphamide/toxicity , Moringa oleifera , Plant Extracts/pharmacology , Protective Agents/pharmacology , Spermatogonia/drug effects , Animals , Cytochromes c/genetics , DNA Damage , Ethanol/chemistry , Male , Mice , Octamer Transcription Factor-3/genetics , Plant Leaves/chemistry , Proto-Oncogene Proteins c-bcl-2/genetics , Proto-Oncogene Proteins c-kit/genetics , RNA, Messenger/metabolism , Sexual Development , Solvents/chemistry , Sperm Count , Spermatogonia/metabolism , Spermatogonia/pathology , bcl-2-Associated X Protein/geneticsABSTRACT
Epidemiological reports have indicated a correlation between the increasing bisphenol A (BPA) levels in the environment and the incidence of male infertility. In this study, the protective effects of melatonin on BPA-induced oxidative stress and apoptosis were investigated in the rat testes and epididymal sperm. Melatonin (10 mg/kg body weight (bw)) was injected concurrently with BPA (50 mg/kg bw) for 3 and 6 weeks. The administration of BPA significantly increased oxidative stress in the testes and epididymal sperm. This was associated with a decrease in the serum testosterone level as well as sperm quality, chromatin condensation/de-condensation level, and the percentage of haploid germ cells in the semen. BPA administration caused a significant increase in apoptosis accompanied by a decrease in the expression of the antiapoptotic proteins Bcl-2 in the testes and epididymal sperm. The concurrent administration of melatonin decreased oxidative stress by modulating the levels of glutathione, superoxide dismutase, and catalase as well as the malondialdehyde and hydrogen peroxide concentrations in the testes and sperm. Melatonin sustained Bcl-2 expression and controlled apoptosis. Furthermore, melatonin maintained the testosterone levels, ameliorated histopathological changes, increased the percentages of seminal haploid germ cells, and protected sperm chromatin condensation process, indicating appropriate spermatogenesis with production of functional sperm. In conclusion, melatonin protected against BPA-induced apoptosis by controlling Bcl-2 expression and ameliorating oxidative stress in the testes and sperm. Thus, melatonin is a promising pharmacological agent for preventing the potential reproductive toxicity of BPA following occupational or environmental exposures.
Subject(s)
Antioxidants/therapeutic use , Apoptosis/drug effects , Benzhydryl Compounds/toxicity , Dietary Supplements , Endocrine Disruptors/toxicity , Melatonin/therapeutic use , Phenols/toxicity , Testis/drug effects , Animals , Benzhydryl Compounds/antagonists & inhibitors , Chromatin Assembly and Disassembly/drug effects , Endocrine Disruptors/chemistry , Environmental Pollutants/antagonists & inhibitors , Environmental Pollutants/toxicity , Epididymis/drug effects , Epididymis/metabolism , Epididymis/pathology , Infertility, Male/blood , Infertility, Male/chemically induced , Infertility, Male/metabolism , Infertility, Male/prevention & control , Male , Oxidative Stress/drug effects , Phenols/antagonists & inhibitors , Proto-Oncogene Proteins c-bcl-2/agonists , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , Proto-Oncogene Proteins c-bcl-2/metabolism , Random Allocation , Rats, Sprague-Dawley , Semen Analysis , Spermatogenesis/drug effects , Spermatogonia/drug effects , Spermatogonia/metabolism , Spermatogonia/pathology , Testis/metabolism , Testis/pathology , Testosterone/blood , Testosterone/metabolism , Vacuoles/drug effects , Vacuoles/pathologyABSTRACT
Refsum disease is caused by a deficiency of phytanoyl-CoA hydroxylase (PHYH), the first enzyme of the peroxisomal alpha-oxidation system, resulting in the accumulation of the branched-chain fatty acid phytanic acid. The main clinical symptoms are polyneuropathy, cerebellar ataxia, and retinitis pigmentosa. To study the pathogenesis of Refsum disease, we generated and characterized a Phyh knockout mouse. We studied the pathological effects of phytanic acid accumulation in Phyh(-/-) mice fed a diet supplemented with phytol, the precursor of phytanic acid. Phytanic acid accumulation caused a reduction in body weight, hepatic steatosis, and testicular atrophy with loss of spermatogonia. Phenotype assessment using the SHIRPA protocol and subsequent automated gait analysis using the CatWalk system revealed unsteady gait with strongly reduced paw print area for both fore- and hindpaws and reduced base of support for the hindpaws. Histochemical analyses in the CNS showed astrocytosis and up-regulation of calcium-binding proteins. In addition, a loss of Purkinje cells in the cerebellum was observed. No demyelination was present in the CNS. Motor nerve conduction velocity measurements revealed a peripheral neuropathy. Our results show that, in the mouse, high phytanic acid levels cause a peripheral neuropathy and ataxia with loss of Purkinje cells. These findings provide important insights in the pathophysiology of Refsum disease.
Subject(s)
Ataxia/pathology , Purkinje Cells/pathology , Refsum Disease/pathology , Animals , Ataxia/enzymology , Ataxia/physiopathology , Automation , Behavior, Animal/drug effects , Central Nervous System/abnormalities , Central Nervous System/drug effects , Central Nervous System/enzymology , Central Nervous System/pathology , Dietary Supplements , Disease Models, Animal , Gait/drug effects , Gene Targeting , Genetic Vectors , Lipidoses/enzymology , Lipidoses/pathology , Male , Mice , Mixed Function Oxygenases/deficiency , Mixed Function Oxygenases/genetics , Peripheral Nervous System Diseases/enzymology , Peripheral Nervous System Diseases/pathology , Phenotype , Phytanic Acid/blood , Phytol/administration & dosage , Phytol/pharmacology , Purkinje Cells/drug effects , Purkinje Cells/enzymology , Refsum Disease/enzymology , Refsum Disease/physiopathology , Spermatogonia/drug effects , Spermatogonia/enzymology , Spermatogonia/pathologyABSTRACT
Quercetin, a dietary-derived falvonol-type flavonoid, is ubiquitous in fruits and vegetables and plays important roles in human health by virtue of its antioxidant function. The present study was performed to investigate effects of quercetin on oxidative damage that was induced by an environmental endocrine disrupter, Aroclor 1254 (A1254), in cultured spermatogonial cells of embryonic chickens. Spermatogonial cells were dispersed from 18-day-old embryo and exposed to A1254 alone or in combination with quercetin. The oxidative damage was estimated by measuring contents of thiobarbituric acid-reactive substances (TBARS, an indicator of lipid peroxidation), activity of superoxide dismutase (SOD, a scavenger of superoxide), and activity of glutathione (GSH, an intracellular antioxidant). Results showed that quercetin had no deleterious effect on spermatogonial cells at 0.01 approximately 1 microg/ml. Exposure to A1254 (10 microg/ml) induced an increase of spermatogonial cell number, and membrane integrity was damaged by elevation of lactate dehydrogenase (LDH) leakage. Exposure to A1254 also induced an elevation in TBARS but a decrease in SOD activity and GSH content. However, compared with A1254 treatment alone, simultaneous supplementation with quercetin decreased LDH leakage to maintain the cell integrity, decreased the levels of TBARS to quench the free radicals, increased SOD activity and GSH content to restore the endogenous antioxidant defense system. Thus, quercetin displayed protective effects on spermatogonial cells from A1254-induced oxidative damage through increasing intracellular antioxidant levels and decreasing lipid peroxidation. Consequently, the antioxidant, such as quercetin, from food or feed consumed by human and animals may attenuate the negative effects of environmental endocrine disrupters.