Elaheh Amini

Polycystic ovarian syndrome (PCOS) is a common endocrine-metabolic condition characterized by persistent anovulation, and hormonal imbalance leading to disrupted follicular maturation, and overall ovarian dysfunction. This study aimed to investigate the regenerative effects of adipose-derived mesenchymal stem cell-derived exosomes (AD-MSCs-Exo) on ovarian restoration, and granulosa cells (GCs) functionality in a PCOS murine model. PCOS was induced via a single subcutaneous injection of estradiol valerate. Exosomes were isolated from AD-MSCs using the ExoCIB kit and characterized by DLS, TEM and flowcytometery. NMRI mice received intraperitoneal injection of AD-MSCs-Exo (25, 50, 75, and 100 µg/kg body weight) twice over a two-week period. Following treatment, the mice were anaesthesia and sacrificed by cervical dislocation. The harvested ovarian tissues were subjected to histological assessment; FSH, LH, and estradiol levels were measured from serum. Further, GCs were isolated, and identified through FSH expression. Subsequently, the levels of GCs key regulator genes including FOXO3, Map1lc3b, and SF-1 were assessed by qRT-PCR. AD-MSCs-Exosome-like nanovesicles isolated in range of 30-80 nm with bi-membrane spherical shape and 54.8% expression of CD9. The hormonal analysis revealed improved estradiol levels and a lowered LH/FSH ratio, indicating endocrine stabilization under exposure with AD-MSCs-Exo. In addition, treatment with AD-MSCs-Exo significantly improved ovarian histoarchitecture, and the optimum response was observed following exposure to 75 µg/kg of AD-MSCs-Exo. However, administration of the highest dose (100 µg/kg) was associated with inflammatory responses and partial tissue degeneration. Gene expression analysis also showed a marked up-regulation of FOXO3, along with increased Map1lc3b and SF-1 levels. Accordingly, AD-MSCs-Exosome like nanovesicles at optimum concentration exert regenerative effects on PCOS-induced ovarian dysfunction, which emphasizes the necessity for precise dose selection to achieve maximal therapeutic efficacy as a promising cell-free strategy for PCOS treatment.

Background: Male infertility affects a substantial portion of the population, and extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) have been proposed as a cell‐free therapeutic approach. This study evaluated the effects of adipose tissue‐derived MSC EVs (ADSC‐EVs) on spermatogonial stem cell (SSC) function in vitro and on spermatogenesis in vivo. Methods and Results: Rat ADSCs were cultured to passage 3, and EVs were isolated from serum‐free conditioned medium by ultracentrifugation (100,000 × g , 70 min). In vitro, SSCs co‐cultured with sertoli cells were treated with 10, 50, and 100 µg/mL ADSC‐EVs; cell viability was measured by MTT after 4 days and gene expression (SYCP3, DAZL) assessed by qRT‐PCR after 10 days. In vivo, immature male Wistar rats (21 days old, n = 6 per group) were randomly assigned to control or intratesticular injection of ADSC‐EVs at 0.25, 1.25, or 2.5 mg/kg; testes were collected 10 days postinjection for histological (hematoxylin and eosin [H&E]) assessment and qRT‐PCR analysis. In vitro, 10 µg/mL ADSC‐EVs produced the highest SSC viability (78.06%) and significantly upregulated SYCP3 (~5.3‐fold, p < 0.001) while DAZL showed a nonsignificant change. In vivo, low‐dose ADSC‐EVs (0.25 mg/kg) increased the presence of spermatozoa within the lumen of seminiferous tubules and moderately upregulated SYCP3 (fold change ~1.95), whereas ADSC‐EVs at 2.5 mg/kg induced seminiferous epithelium degeneration and absence of mature spermatozoa. DAZL expression was downregulated in vivo at the tested doses. Conclusions: ADSC‐EVs exert dose‐dependent, dual effects on spermatogenesis: at low concentrations, they promote SSC differentiation (SYCP3‐associated), while higher concentrations can damage seminiferous architecture and impair sperm release. These findings emphasize the importance of dose optimization for potential therapeutic applications.

Citalopram (CT) has indeed been demonstrated to induce sexual dysfunction, although its capacity to be harmful to the male reproductive system has not been fully explored. The current investigation analyzed the mechanism underlying the protective impact of date palm pollen (DPP) on CT‐induced Sertoli cell (TM4 cells) apoptosis and clarified the harmful effects of CT on the mouse males’ reproductive system. Using flow cytometry, the apoptotic rate in each group was determined. The impact of DPP on the expression of Bcl-2 , Nrf2 , and SOD1 molecular targets was identified. Also, male NMRI mice were orally treated simultaneously with CT and DPP for 30 days, and the testes underwent histological analysis. Our findings demonstrated that adding different doses of DPP to Sertoli cells raised the mRNA expression levels of Nrf2 , SOD1 , and Bcl-2 to varying degrees ( p < 0.01). Our findings also demonstrated that DPP decreased the rate of TM4 cell death brought on by CT ( p < 0.01). In addition, many abnormalities were observed in testicular tissue in mice treated with CT. Our results showed that DPP can reduce the harmful effects of CT on testicular tissue and restore spermatogenesis. These results lend credence to the idea that CT can result in testicular injury and infertility. In contrast, treatment with DPP can lessen the negative effects of CT, most likely through the regeneration of testicular tissue and antioxidant systems.

An effective approach to enrich the in vitro maturation (IVM) of oocyte medium as one of the main assisted reproduction technologies is the use of antioxidants to minimize oxidative stress. This study examined the effects of copper nanoparticles (CuNPs) synthesized by chemical (Ch-NPs) and green (G-CuNPs) methods on the IVM process of mouse oocytes and the development of the embryo in comparison to control oocytes (without nanoparticles treatment). Hydroalcoholic (G-H-CuNPs) and aqueous (G-A-CuNPs) Rosmarinus officinalis extracts were used for green synthesis. Here, Ch-NPs showed much less nuclear maturation and survival rate (44.92 ± 4.52; 66.21 ± 6.22) than the control (73.36 ± 7.40; 89.33 ± 4.40), respectively (P < 0.001). In contrast, G-H-CuNPs treated oocytes exhibited a significant increase (72.28 ± 5.51; 79.37 ± 6.29) compared to the Ch-NPs (P < 0.05). The level of ROS in Ch-NPs exposed oocytes was significantly higher than in the control (P < 0.001). The fertilization rate exhibited a significant elevation in the G-H-CuNPs (96.00 ± 2.45) compared to the control (71.14 ± 5.20) and Ch-NPs (50.00 ± 0.01) (P < 0.05). The 8-cell and blastocyst (BL) rates in the G-H-CuNPs (70.32 ± 3.78) revealed notably higher than those in the control (64.29 ± 3.69) and Ch-NPs (36.67 ± 10.22) (P < 0.05). In summary, results exhibited that G-CuNPs promote mouse oocyte maturation, fertilization, and embryo development more than Ch-NPs. The follow-up studies propose looking into the safety and applicability of green-synthesized CuNPs in human-assisted reproductive technologies. Graphical Abstract