The authors have no conflicts of interest to declare.
Local Nigerian men have been using
Infertility is a very common complication in diabetic men. It is a recognized global health challenge in all countries, both developed and developing, with some couples requiring assisted reproductive interventions to be able to conceive. A good number of diabetic – related outcomes such as generation of oxidative stress and lipid peroxidation will negatively affect individuals with both type 1 and type 2 diabetes Mellitus in several ways such as progressive genotoxicity, structural defect in testicular tissue and significantly lower sperm quality
The major source of oxidative stress (OS) in most diseases including diabetes mellitus and also in normal physiological conditions is the mitochondria. During oxidative metabolism in the mitochondria, high concentration of oxygen is reduced to water, while part of oxygen though little is transformed to O2_
Oxidative stress has been indirectly connected to the clinical consequences of sperm DNA damage
Mitochondrial exposure to ROS provokes apoptotic process through release of Apoptosis Inducing Factor (AIF) which also cause DNA fragmentation. High level of ROS agitates mitochondrial membrane polarization and activates release of the cytochrome-C protein as a trigger of apoptosis
The gradual shift to herbal therapy with its attendant increasing acceptance, even among the elite confirm the claim that herbal remedies can provide cure for several diseases, including infertility in men
Auricularia Polytricha (Mushroom) is one fungus found to have numerous medicinal properties. It belongs to the family Auriculariacaea and has been reported to have antioxidant property (Mau et al 2001), immunomodulatory activity
Auricularia Polytricha (AP) has been used locally by the Ejagham speaking people in Cross River State of Nigeria as an aphrodisiac agent in management and treatment of sexual dysfunction in men. This has been going on without the corresponding clinical trials and acceptable scientific experimentation to give credence to this age-long practice. This research is therefore intended to investigate the effect of ethanolic extract of Auricularia Polytricha AP (Mushroom) on testicular DNA expression and some oxidative stress markers of STZ-Induced diabetic rat model.
Auricularia polytricha was obtained from Etomi central market located in Etung Local Government Area of Cross River State and taken to the department of biological sciences, University of Nigeria for identification. The mushroom was dried at room temperature, powdered and subjected to crude extraction with ethanol. Modified method was used for extraction. 200g of
Ethical clearance was obtained from the University of Calabar, Nigeria with Clearance Number: UC/FBMS/19/001291.
Thirty (30) adult male Wistar rats with average weight of 150g were used for this research. The rats will be kept in clean cages and divided into six groups designated A, B, C, D, E, and F with five rats in each group. The rats were allowed to acclimatize for two weeks in animal house, Department of Anatomy, Faculty of Medicine, University of Nigeria, Enugu Campus and allowed unrestricted access to commercially available chow (livestock feed) and water.
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A | Normal control | Distilled water |
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B | Diabetic control | 65mg/kg.bw of STZ |
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C | STZ + AP (Low Dose) | STZ + 250mg/kg.bw |
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D | STZ + AP (Mid Dose) | STZ + 500mg/kg.bw |
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E | STZ + AP (High Dose) | STZ + 1000mg/kg.bw |
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F | STZ + Metformin | STZ + 40mg/kg.bw Metformin |
After fasting for twelve hours, hyperglycaemia was induced by administering streptozotocin (STZ) intra-peritoneally. STZ was reconstituted in 0.5M Sodium citrate and administered at a dose of 65mg/kg.bw
Diabetes was confirmed three days after administration of STZ using Accu-Check glucometer (Roche diagnostic, Germany) with blood samples obtained from tails of Wistar rats. Blood glucose levels (mmol/l) was checked before and after induction to ascertain hyperglycaemic state.
At termination, the animals were sacrificed with the testes removed and plotted with filter paper. Both right and left testes were weighed together and then suspended in Bouins fluid for fixation, preparatory to histological processing. The sperm was collected by mincing from caudal epididymis with anatomical scissors into a glass beaker. 19 drops of diluting solution was added to dilute the semen in a proportion of 1:20. A sample from this homogenate was used for semen analysis.
Fuelgen nuclear reaction was used for DNA demonstration (Fuelgen and Rossenbeck, 1924) to assess testicular DNA distribution.
Reagent based antioxidant enzyme assay was used to demonstrate for
Melondialdehyde (MDA)
Catalase
Soperoxide Dimutase (SOD)
Quantitative data from this research was recoded and tabularized. Statistical significance of the differences between the groups was determined using one way analysis of variance (ANOVA) using SPSS statistical analysis program. P<0.05 will be considered significant.
Oxidative stress markers (SOD, catalase and Melondialdehyde) activities of different experimental animals.
Results obtained from analysis of serum oxidative stress markers shows that both SOD and Melondialdehyde activities were remarkably (p<0.05) higher in diabetic control animals when compared with the normal control group. Values in Groups C, D and F that were administered with 250, 500mg/kg.bw
Catalase concentration in diabetic control animals was remarkably (p<0.05) higher when compared to the normal control but was not significantly (p<0.05) different in groups D (DM+500mg/kg.
From findings in this study, irregular and distorted arrangements of DNA in all diabetic groups (Groups B, C, D, E and F) when compared with the normal control group, may have been due to displacement of sertoli cell within the germinal epithelium of seminiferous tubules. Strands of DNA were also seen arrange in clusters in diabetic groups, showing altered and defective structure which might have resulted from base free side deletion, frame shift, cross-linking and chromosomal rearrangement. The intensity of magenta colour development in Feulgen reaction for DNA demonstration was proportional to DNA concentration. There was reduced colour intensity in all diabetic groups (Groups B, C, D, E and F) when compared with the normal control. This is in line with report from Aitken and Krausz (2001). However the degree of distortion and cross-linking of DNA strand in the group of diabetic animal models placed on high dose (1000mg/kg.bw) of
Extensive experimental evidence
The marginal reversal of DNA damage following 21 days of
Extensive experimental research show that oxidative stress and reduced level of androgen hormone are potent risk factors associated with dysfunction in testes and impaired spermatogenic process in hyperglycaemic rat. Saleh
In conclusion, ethanolic extract of