Impacts of Acrylamide on testis and spermatozoa.

Acrylamide (ACR) is an industrial chemical used to produce polyacrylamide, a synthetic polymer with a wide range of applications. Depending on the dosage, its presence in occupational and environmental sources poses potential health risks to humans and animals. ACR can be formed in starchy foods cooked at high temperatures. Its effects on human sperm are not well understood. Animal studies indicate that ACR induces toxicity in the male reproductive system through oxidative stress mechanisms. Exposure to ACR alters the normal structure of testicular tubules, leading to congestion, interstitial edema, degeneration of spermatogenic cells, formation of abnormal spermatid giant cells, and necrosis and apoptosis. It also disrupts the balance of important biomarkers such as malondialdehyde, nitric oxide, superoxide dismutase, catalase, and glutathione. ACR has a negative impact on mitochondrial function, antioxidant enzymes, ATP production, and sperm membrane integrity, resulting in decreased sperm quality. Furthermore, it interferes with the expression of steroidogenic genes associated with testosterone biosynthesis. This review explores the detrimental effects of ACR on sperm and testicular function and discusses the potential role of antioxidants in mitigating the adverse effects of ACR on male reproduction.

Co-treatment with bone marrow-derived mesenchymal stem cells and curcumin improved angiogenesis in myocardium in a rat model of MI.

BACKGROUND: The cardioprotective properties of mesenchymal stem cells and the therapeutic potential of curcumin (CUR) have been explored. Combining these approaches may enhance stem cell effectiveness and expedite healing. This study aimed to investigate the synergistic effects of co-treating bone marrow mesenchymal stem cells (BMSCs) with curcumin on vascular endothelial growth factor (VEGF) levels, in a rat model of myocardial ischemia (MI). METHODS AND RESULTS: Sixty-five male rats were divided into four groups: G1 (healthy control), G2 (MI induced by isoproterenol hydrochloride), G3 (treated with BMSCs), and G4 (co-treated with curcumin and BMSCs). Blood and tissue samples were collected at specific time points (day 1, 7, 15 and 21) after MI induction. Serum levels of lactate dehydrogenase (LDH), creatine kinase (CK), cardiac troponin I (cTnI), aspartate aminotransferase (AST), CK-MB and VEGF were measured. VEGF mRNA and protein expression were evaluated using RT-qPCR and Western blot techniques. Histopathological assessments were performed using H&E staining and CD31 immunofluorescence staining. VEGF expression significantly increased on days 7 and 15 in the CUR-BMSCs group, peaking on day 7. Western blot analysis confirmed elevated VEGF protein expression on days 7 and 15 post-MI. ELISA results demonstrated increased serum VEGF levels on days 7 and 15, reaching the highest level on day 7 in CUR-BMSCs-treated animals. Treated groups showed lower levels of LDH, AST, CK, CK-MB and cTnI compared to the untreated MI group. H&E staining revealed improved myocardial structure, increased formation of new capillaries, in both treatment groups compared to the MI group. CONCLUSION: Combining curcumin with BMSCs promotes angiogenesis in the infarcted myocardium after 15 days of MI induction. These findings suggest the potential of this combined therapy approach for enhancing cardiac healing and recovery.

Corrigendum to "Protective effects of chlorogenic acid against ionizing radiation-induced testicular toxicity" [Heliyon 8 (10) (October 2022) Article e10798].

[This corrects the article DOI: 10.1016/j.heliyon.2022.e10798.].

Detrimental effects of vitrification on integrin genes (α9 and ß1) and in vitro fertilization in mouse oocytes.

OBJECTIVE: Integrins are known as key molecules that importantly involve in fertilization. This study aimed to evaluate effects of vitrification on fertilization rate and expression of integrin genes, α9 and ß1, on mice oocytes in GV and MІІ stages. MATERIALS AND METHODS: From the ovarian tissue and fallopian tube of NMRI mice, germinal vesicle (GV, n = 200) and metaphase II (MII, n = 200) oocytes were obtained. Then, oocytes were distributed into 4 groups including non-vitrified GV, non-vitrified MII, vitrified GV, and vitrified MII. Cryotop method was used for vitrification and oocytes (for 4 weeks) were kept in liquid nitrogen. After that, by using an inverted microscope, the rate of survived oocytes was assessed. Also, in vitro fertilization (IVF) for oocytes, obtained from in vitro maturated MII and mice ovaries (ovulated MII), was done to assess embryos at differenced stages (2-cells, morula, and hatched). Finally, RT-qPCR was performed to investigate the mRNA expression of integrin genes (α9 and ß1). RESULTS: After vitrification, the rate of survived oocytes, 68.65%for GV and 65.07% % for MII, did not show a remarkable difference related to non-vitrified groups, while the fertilization rate in vitrified groups remarkably decrease compared to non-vitrified groups (p < 0.05). Also, the expression of α9 and ß1 genes was significantly altered in vitrified groups when compared to non-vitrified groups (p < 0.05). There was no significant difference in embryo developmental rates for non-vitrified and vitrified groups. CONCLUSION: Cryotop method for vitrification caused an alternation in oocyte quality by reducing fertilization rate and integrin gene expression.

Protective effects of chlorogenic acid against ionizing radiation-induced testicular toxicity.

Background: Testicular tissues could damage by ionizing radiation (IR) during the treatment of pelvic cancers. The aim of this study was to investigate both the protective and therapeutic effects of chlorogenic acid (CGA) on IR-induced mouse testis tissue damage. Methods: In this experimental study, 70 mice were divided into 3 groups, including group 1 (normal saline), group 2 (IR + normal saline), and group 3 (IR + 5, 10, 20, 40, and 80 mg/kg) CGA via I.P injection. Animals in groups 2 and 3 received a dose of 2.0 Gy total-body irradiation in a single fraction. At two determined time points (16 h and 35 days after exposure), the testis and caudal part of both epididymis were isolated and underwent subsequent analyses. Results: The results showed that irradiation of mice caused massive damage to spermatogenesis, seminiferous tubules, basal lamina, Leydig cells, and sperm parameters. Further biochemical assessment of the data demonstrated that 40 mg/kg CGA almost restored MDA to a normal level. In addition, the level of SOD, TAC, and GSH were significantly increased in the 40 mg/kg CGA treated group. Molecular evidence confirmed the protective effects of CGA and also revealed that the ratio of Bax/Bcl-2 in the presence of 40 mg/kg CGA was significantly decreased compared to IR and some treated groups. Conclusion: The protective and therapeutic effects of CGA on testis were found to be positively correlated with the dose level.

Chlorogenic acid improves functional potential of follicles in mouse whole ovarian tissues in vitro.

BACKGROUND: Chlorogenic acid (CGA) is one of the well-known polyphenol compounds possessing several important biological and therapeutic functions. In order to optimize a culture system to achieve complete development of follicles, we focused on the effects of CGA supplementation during in vitro culture (IVC) on follicular development, oxidative stress, antioxidant capacity, developmental gene expression, and functional potential in cultured mouse ovarian tissue. METHODS AND RESULTS: The collected whole murine ovaries were randomly divided into four groups: (1) non-cultured group (control 1) with 7-day-old mouse ovaries, (2) non-cultured group (control 2) with 14-day-old mouse ovaries, (3) cultured group (experimental 1) with the culture plates containing only the basic culture medium, (4) cultured group (experimental 2) with the culture plates containing basic culture medium + CGA (50, 100 and 200 µmol/L CGA). Afterward, histological evaluation, biochemical analyses, the expression assessment of genes related to follicular development and apoptosis as well as the analysis of 17-ß-estradiol were performed. The results showed that supplementation of ovarian tissue with the basic culture media using CGA (100 µmol/l) significantly increased the survival, developmental and functional potential of follicles in whole mouse ovarian tissues after 7 days of culture. Furthermore, CGA (100 µmol/L) attenuated oxidative damage and enhanced the concentration of antioxidant capacity along with developmental gene expression. CONCLUSION: It seems that supplementation of ovarian tissue with culture media using CGA could optimize follicular growth and development in the culture system.

Lysophosphatidic acid supports the development of vitrified ovarian follicles by decreasing the incidence of cell death: An experimental study.

Background: Lysophosphatidic acid (LPA) contributes to follicular activation, oocyte maturation, in vitro fertilization, and embryo implantation. Objective: This study was designed to evaluate the effects of LPA to improve the development of isolated follicles derived from whole mouse cultured vitrified ovaries. Materials and Methods: In this experimental study, first, the 1-wk-old mouse ovaries in the non-vitrified and vitrified groups were cultured in the presence of 20 µM of LPA for 1 wk. Then, their isolated preantral follicles were cultured individually for 12 days in the presence or absence of 40 µM of LPA. The following evaluations were done for the cultured follicles: a viability test using Calcein AM staining, flow cytometry using annexin V/Pi, and analysis of the expression of genes by real-time reverse transcription polymerase chain reaction. The maturation rates of the oocytes were compared among groups and some of the released metaphase II oocytes were subjected to in vitro fertilization. Results: In all LPA treated groups, the rates of survival and follicular development were higher, and the incidence of cell death and expression of pro-apoptotic genes were lower, than in the non-LPA supplemented groups (p = 0.035). There was no significant difference between the vitrified and non-vitrified groups regarding follicular or oocyte development, but the expression of Bad and LPA receptors genes was significantly altered in the vitrified LPA supplemented group in comparison with the non-vitrified LPA supplemented group (p = 0.028). Conclusion: LPA improved the survival and developmental potential of the isolated follicles. Despite some alterations in the expression of apoptosis-related genes in the vitrified ovaries, LPA had positive effects on the survival and development of these follicles.

Effect of chlorogenic acid on follicular development, hormonal status and biomarkers of oxidative stress in rats with polycystic ovary syndrome.

Polycystic ovarian syndrome (PCOS) is a complex endocrine and metabolic disorder. Chlorogenic acid (CGA) bears antioxidant properties with protective effects on different tissues. This study was conducted to evaluate the effect of CGA on follicular development, hormonal status and biomarkers of oxidative stress in a rat model of PCOS. In this experimental study, 18 rats were divided into three equal groups including: control, non-treated PCOS [(estradiol valerate (EV): 40.00 mg kg-1 intramuscularly)], and PCOS-CGA (EV: 40.00 mg kg-1 intramuscularly and CGA: 100 mg kg-1 intraperitoneally once a week for eight consecutive weeks). At the end of treatment period, all rats were anesthetized. Then 5.00 mL blood samples of rats in the three groups were taken and prepared for hormonal analyses and their ovaries were isolated and dissected mechanically free of fat and mesentery. The ovaries underwent the following analyses: Morphological study with Hematoxylin and Eosin staining and biochemical study using the malondialdehyde (MDA) level and total antioxidant activity. Data were analyzed using one-way ANOVA and post hoc Tukey's test. The serum level of luteinizing hormone, estrogen, testosterone, antioxidant capacity, glutathione and the number of cystic follicles in the PCOS group treated with 100 mg kg-1 Chlorogenic acid compared to the non-treated PCOS group were significantly decreased, however, the serum level of follicle stimulating hormone, progesterone, MDA and the number of secondary, graafian follicles and corpus luteum were significantly increased. Chlorogenic acid could be effective in ameliorating follicular development as well as hormonal and biochemical disorders in rats with PCOS.

The Effects of Lysophosphatidic Acid on The Incidence of Cell Death in Cultured Vitrified and Non-Vitrified Mouse Ovarian Tissue: Separation of Necrosis and Apoptosis Border.

OBJECTIVE: The aim of the present study was to examine whether lysophosphatidic acid (LPA) could decrease cell death and improve in vitro culture (IVC) conditions in cultured vitrified mouse ovarian tissue. MATERIALS AND METHODS: In this experimental study, we collected and randomly divided 7-day-old mouse ovarian tissues into vitrified and non-vitrified groups. The ovaries were cultured in the presence and absence of LPA for one week. Morphology and follicular development were evaluated by hematoxylin and eosin (H&E) and Masson's trichrome (MTC) staining. The incidence of cell death was assessed by flow cytometry using annexin V/propidium iodide (PI) and a caspase-3/7 assay in all studied groups. RESULTS: The vitrified groups had a significantly decreased follicle developmental rate compared to the non-vitrified groups (P<0.05). Overall, qualitative and quantitative results showed prominent follicular degeneration in the vitrified groups compared with the respective non-vitrified groups. Both LPA treated groups had a significantly higher proportion of preantral follicles compared to the non-LPA treated groups (P<0.05). Flow cytometry analysis results showed significantly greater early and late apoptotic cells in all groups (17.83 ± 8.80%) compared to necrotic cells (7.97 ± 0.92%, P<0.05). The percentage of these cells significantly increased in the vitrified groups compared with non-vitrified groups. LPA treated groups had a lower percentage of these cells compared to non-LPA treated groups (P<0.05). The lower enzyme activity was observed in non-vitrified (especially in the LPA+ groups) cultured ovaries compared to the vitrified group (P<0.05). CONCLUSION: Both vitrification and IVC adversely affected cell survival and caused cell death. We postulated that LPA supplementation of culture medium could improve the developmental rate of follicles and act as an anti-cell death factor in non-vitrified and vitrified ovarian tissues. It could be used for in vitro maturation of ovarian tissue.

Effect of lysophosphatidic acid on the follicular development and the expression of lysophosphatidic acid receptor genes during in vitro culture of mouse ovary.

Lysophosphatidic acid (LPA) known as a serum-derived growth factor, is involved in several cell physiological functions in the female reproductive system including: oocyte maturation, in vitro fertilization and embryo implantation by its transmembrane G protein-coupled receptors. The aim of the present study was to examine the effect of LPA on in vitro follicular development of mouse ovarian tissue. Neonatal mouse ovarian tissues were cultured in five different concentrations of LPA (0, 5, 10, 20 and 40 µM). The developmental competence and the function of cultured ovarian tissue were assessed by morphological study using hematoxylin and eosin staining and hormonal analysis. The expression of LPA receptor (LPAR 1-4) genes were analyzed by real-time RT-PCR. The proportion of preantral follicles and the level of E2 hormone were significantly higher in the 20 µM LPA-treated group than those in the other treatment groups. There was a significant difference in the expression of LPAR 1-4 genes in 20 µM LPA treated group in comparison with 0 µM LPA (control group) treated and non-cultured groups. In addition, the expression of LPAR1 gene was higher than other receptor genes in all studied groups. In conclusion supplementation of the media with 20 µM LPA, could improve the survival and developmental potential of follicles and it had positive effects on cell function and stimulation of E2 synthesis in mouse whole ovarian tissues.

Vitrification by Cryotop and the Maturation, Fertilization, and Developmental Rates of Mouse Oocytes.

BACKGROUND: Oocyte cryopreservation is an important part of modern fertility treatment. The effect of vitrification on the fertilization and developmental rates of embryo is still a matter of debate. OBJECTIVES: This study aimed to investigate the effect of vitrification on the success of mouse oocyte maturation, fertilization, and preimplantation development in vitro. MATERIALS AND METHODS: In this experimental study, a total of 200 germinal vesicle (GV) and 200 metaphase II (MII) oocytes were obtained from ovaries and fallopian tubes of NMRI mice, respectively and divided into two control and experimental (vitrified) groups. Oocytes in the experimental group were vitrified by Cryotop using vitrification medium (Origio, Denmark) and kept in liquid nitrogen for one month. Then, they were cultured in maturation medium for 24 hours. In vitro maturated metaphase 2 (IVM-MII) and ovulated metaphase 2 (OV-MII) oocytes were inseminated and the fertilized embryos assessed until the hatching blastocyst stage. Outcomes were assessed for statistical significance by Chi-square test using SPSS software. RESULTS: Vitrification caused a significant reduction in the maturation rate of oocytes. Of those that matured, the fertilization rate of vitrified IVM-MII (44.1%) and OV-MII oocytes (50%) was not significantly different from each other but both were significantly lower than the control group (P < 0.05). There was no significant difference in developmental rates of both vitrified groups and the control group. CONCLUSIONS: The present study showed that vitrification using Cryotop and freezing medium can damage oocytes by reducing the maturation and fertilization rates in both developmental stages.

The effect of vitrification on mouse oocyte apoptosis by cryotop method.

BACKGROUND: Oocyte cryopreservation is one of the most important topics in the field of assisted reproductive technology to preserve women fertility, but relationship between cryopreservation and apoptosis is still a matter of debate. The present study was aimed to investigate the effects of vitrification on apoptosis in mouse oocytes by Cryotop method. METHOD: A total of 200 germinal vesicle (GV) and 200 metaphase II (MII) oocytes were obtained from ovaries and fallopian tubes of NMRI mice, respectively and divided into control and experimental groups. Oocytes in experimental group were vitrified by Cryotop using vitrification medium and were kept in liquid nitrogen for one month. The survival rate of oocytes was evaluated after 2 hour incubation time. Then, the oocyte apoptosis was evaluated by TUNEL technique and compared with those in control group. The data was compared statistically using SPSS software and chi-square test. RESULTS: The survival rates of vitrified GV (93%) and MII oocytes (88%) showed a significant decrease compared with the control group (P<0.05), but there was no significant difference in survival rate of both vitrified oocyte groups. The incidence of apoptosis in vitrified and control GV oocytes showed no significant difference (13% vs. 7%), but the rate of apoptosis in vitrified MII oocytes increased significantly not only in comparison with MII control group (25% vs. 5%) but also with vitrified GV oocytes (P<0.05). CONCLUSION: The results indicate that vitrification increases apoptosis in mouse MII oocytes and apoptosis may play a role in MII oocyte injury after vitrification.