The effect of calpain inhibitor-I on copper oxide nanoparticle-induced damage and cerebral ischemia-reperfusion in a rat model.

This study aimed to investigate the effects of the calpain inhibitor N-Acetyl-Leu-Leu-norleucinal (ALLN) on neuroapoptotic cell damage caused by Copper Oxide Nanoparticles (CuO-NP) and exacerbation of damage through brain ischemia/reperfusion (I/R) in a rat model. Male Wistar Albino rats (n=80) were divided into eight groups: Control, I/R, CuO-NP, CuO-NP+I/R, I/R+ALLN, CuO-NP+ALLN, CuO-NP+I/R+ALLN, and DMSO. Biochemical markers (MBP, S100B, NEFL, NSE, BCL-2, Cyt-C, Calpain, TNF-α, Caspase-3, MDA, and CAT) were measured in serum and brain tissue samples. Histological examinations (H&E staining), DNA fragmentation analysis (TUNEL) were performed, along with Caspase-3 assessment. The ALLN-treated groups exhibited significant improvements in biochemical markers and a remarkable reduction in apoptosis compared to the damaged groups (CuO-NP and I/R). H&E and Caspase-3 staining revealed damage-related morphological changes and reduced apoptosis in the ALLN-treated group. However, no differences were observed among the groups with TUNEL staining. The findings suggest that ALLN, as a calpain inhibitor, has potential implications for anti-apoptotic treatment, specifically in mitigating neuroapoptotic cell damage caused by CuO-NP and I/R.

Effects of Varicocele Repair on Sperm DNA Fragmentation and Seminal Malondialdehyde Levels in Infertile Men with Clinical Varicocele: A Systematic Review and Meta-Analysis.

PURPOSE: Varicoceles can be a source of elevated seminal oxidative stress (OS) and sperm DNA fragmentation (SDF). However, it remains unclear whether varicocele repair (VR) could reduce these parameters. This systematic review and meta-analysis (SRMA) aims to investigate the impact of VR on SDF and seminal malondialdehyde (MDA). MATERIALS AND METHODS: A literature search was performed in Scopus, PubMed, Ovid, Embase, and Cochrane databases. This SRMA included randomized controlled trials and observational studies reporting the pre- and postoperative levels of SDF and seminal OS in infertile men with clinical varicocele that underwent VR. Subgroup analyses included techniques of VR and SDF testing. The effect size was expressed as standardized mean difference (SMD). RESULTS: Out of 1,632 abstracts assessed for eligibility, 29 studies with 1,491 infertile men were included. The analysis showed a significant reduction in SDF after VR, compared to preoperative values (SMD -1.125, 95% confidence interval [CI] -1.410, -0.840; p<0.0001) with high inter-study heterogeneity (I²=90.965%). Reduction in SDF was evident with microsurgical technique and non-microsurgical inguinal approaches (SMD -1.014, 95% CI -1.263, -0.765; p<0.0001, and SMD -1.495, 95% CI -2.116, -0.873; p<0.0001), respectively. Reduction in SDF was significant irrespective of testing was done by sperm chromatin dispersion (SMD -2.197, 95% CI -3.187, -1.207; p<0.0001), sperm chromatin structure assay (SMD -0.857, 95% CI -1.156, -0.559; p<0.0001) or TUNEL (SMD -1.599, 95% CI -2.478, -0.719; p<0.0001). A significant decrease in seminal MDA levels was observed following VR (SMD -2.450, 95% CI -3.903 to -0.997, p=0.001) with high inter-study heterogeneity (I²=93.7%). CONCLUSIONS: Using pre- and post-intervention data, this SRMA indicates a significant reduction in SDF and seminal MDA levels in infertile men with clinical varicocele treated with VR. These findings may have important implications for the future management of this selected group of infertile patients.

In 2D and 3D Cell Culture Models, Effects of Endothelial Cells on E-cadherin / ß-catenin Expression Levels and Spheroid Sizes in Ishikawa Cells.

OBJECTIVE: Increasing evidence shows that three dimensional cell culture models better reflect the in vivo tumor microenvironment than two dimensional cell culture models. Co-culture models are ideal cell culture models for understanding the communication between cells and the in vivo microenvironment. Changes in expression levels of E-cadherin are closely related to cancer metastasis and progression. ß-catenin mediates cell adhesion of E-cadherin. Endothelial cells are stromal cell components in the tumor microenvironment. It is known that there is little or no expression of E-cadherin in endothelial cells. METHODS: In our study, both two-dimensional and three dimensional mono-culture and co-culture models were created using Huvec and Ishikawa cells (endometrial cancer cell lines) to better reflect cell interactions. Spheroids were followed for three days in the three-dimensional cell culture model. E-cadherin and ß-catenin expression levels of two-dimensional and three dimensional mono-culture and co-culture models were measured by immunofluorescence staining. Spheroid images were recorded using a Z-stack. Intensity measurements in both two-dimensional and three-dimensional mono-culture and co-culture models were made using the Image J software. Study groups were evaluated by one-way analysis of variance (One-Way ANOVA). Values of p <0.05 were considered statistically significant. RESULTS: The size of the co-culture spheroids was recorded significantly larger than the mono-culture spheroids (p <0.0001). In two (p = 0.0175) and three dimensional models (p <0.0001), expression levels of E-cadherin in the mono-culture of Ishikawa cells were recorded significantly higher than in Huvec and co-culture cells. Likewise, while the expression levels of ß-catenin were higher in the mono-culture of Ishikawa cells in two-dimensional models (p <0.05), no significant difference was observed in three dimensional models (p> 0.05). CONCLUSION: In summary, it has been noted that the expression levels of E-cadherin are significantly reduced in co-cultures of Ishikawa cells with Huvec cells in both two and three dimensions . These results support the idea that endothelial cells may cause changes in endometrial cancer progression by suppressing E-cadherin expression in Ishikawa cells.

Effects of Vincamine on Testicular Dysfunction in Alloxan-induced Diabetic Male Rats.

Background: Diabetes mellitus (DM) is frequently linked with problems of several organ systems, including retinopathy, neuropathy, and nephropathy. Additionally, patients have changes in sexual functioning, such as decreased libido and fertility. Vincamine, a monoterpenoid indole alkaloid, has hypoglycemic and antioxidant effects. Objectives: This research assessed the impact of vincamine on testicular dysfunction in alloxan-induced male rats by measuring fasting blood glucose, oxidative stress, seminal analysis, and histological examination of the testis. Methods: Wister-albino male rats were randomized into the following groups at random: Untreated-healthy, untreated-DM, vincamine-treated (20 mg/kg) DM, vincamine-treated (40 mg/kg) DM, and clomiphene-treated DM (5 mg/kg). On day 14, rats were sacrificed, and semen/blood samples were collected. Sperm count, motility, and morphological abnormalities were noted by microscopic examination. The testis was examined histopathologically and assessed using Johnsen's score. Results: Compared with the untreated diabetic group, a dosage of 40 mg/kg vincamine generate a significant reduction in fasting blood sugar (FBG). Compared with the untreated diabetic group, the vincamine-treated rats produced greater plasma testosterone levels and Johnsen scores. In the vincamine 20 mg/kg group, sperm concentration was higher than in the vincamine 40 mg/kg group. Conclusions: It is possible that vincamine has a potential preventive effect against diabetes-related reproductive problems attributable to its antioxidant activity and capacity to restore testicular steroidogenesis.

Chronic unpredictable stress disturbs the blood-testis barrier affecting sperm parameters in mice.

RESEARCH QUESTION: Does chronic stress affect the key proteins and sperm parameters of the blood-testis barrier (BTB)? DESIGN: C57Bl/6 mice were divided into two groups: a non-treated control group and a chronic unpredictable stress (CUS) applied group. The stress status of the animals was confirmed with behavioural tests. Histopathologic evaluation was conducted by haematoxylin and eosin staining and electron microscope. Malondialdehyde, corticosterone and testosterone levels were evaluated in peripheral blood. Expression levels of BTB proteins, namely zonula occludens-1 (ZO-1), claudin-11 (CLDN11) and clathrin in Sertoli cells, were assessed by Western blotting and immunofluorescence techniques. Sperm samples were collected from cauda epididymis, and sperm parameters analysed. RESULTS: The stress model was confirmed by behavioural tests. Histopathological evaluation of the testes demonstrated a mild degeneration in seminiferous tubules. Malondialdehyde (P = 0.008) and corticosterone levels increased (P = 0.004) and testosterone levels decreased (P = 0.005) in the CUS group. Electron microscopic evaluation confirmed the damage in BTB integrity in the CUS group. Western blot analysis showed that ZO-1 and CLDN11 levels were significantly decreased, although clathrin levels were unchanged. Although sperm concentration and total motility rate were not significantly different between the groups, progressive motility (P = 0.03), normal sperm morphology (P = 0.04), chromatin integrity (toluidine blue) (P = 0.002) and the acrosomal reaction rate (P = 0.002) were significantly decreased, and acrosomal abnormality rate was dramatically increased (P = 0.04) in the CUS group. CONCLUSIONS: In mice, CUS disrupted BTB integrity and impaired sperm parameters. A decrease in ZO-1 and CLDN11 expression levels may be proposed as the causative factor.

Leptin in sperm analysis can be a new indicator.

Purpose Radiotherapy, chemotherapy, various tumors and invasive surgery can result in ejaculatory dysfunction and testicular insufficiency. Sperm cryopreservation is the only method which can provide a baby for couples. Cryopreservation freezes tissues and cells, allowing them to be stored for future use by stopping all biological activities. Cryopreservation can cause some harmful changes in the structure and function of the sperm. Leptin molecule plays many roles in most biological processes including the satiety and cell renewal, proliferation, angiogenesis, modulation of energy expenditure and regulation of the neuroendocrine system. Leptin was also reported to be associated with spermatogenesis in several studies. Methods This study aims to use leptin molecule as a parameter for sperm motility and DNA fragmentation before and after the cryopreservation. In this study, semen samples were taken from 30 normospermic male volunteers. Each semen sample was examined for the same parameters before and after the cryopreservation. Samples were analyzed before and after cryopreservation in terms of sperm motility by morphological sperm analysis with spermac stain dye, DNA fragmentation by TUNEL assay, ultrastructural analysis with transmission electron microscopy (TEM), seminal leptin levels by ELISA method and reactive oxygen species (ROS) levels by colorimetric method. Results Decreased sperm motility, distribution of sperm morphology and increased DNA fragmentation were determined after cryopreservation. Similarly, seminal ROS and leptin levels were also increased significantly. There was a negative correlation between seminal leptin and sperm motility. Additionally, there was a positive correlation between seminal leptin and DNA fragmentation. Conclusion According to these results, leptin molecule can be used as a marker for sperm motility and DNA fragmentation before and after cryopreservation. We think that the results of this study can contribute to further studies in the clinical aspect.

A comparative study of fixatives for axolotl blastema tissue / Estudio comparativo de fijadores para el tejido blastema del axolotl

Regeneration is defined as tissue renewal and functional restoration process of the damaged parts of the body after an injury. Ambystoma mexicanum, commonly named the Axolotl, is one of the unique vertebrates, which has a remarkable ability to regenerate their extremities following the amputation. Although the process of regeneration includes several periods, it can be divided into two main phases; blastema formation and dedifferentiation. In the couple of hours following the amputation, wound closure occurs by migration of epithelial cells around the amputation site followed by macrophage infiltration and dedifferentiation of cells to turn into stem cells. Accumulated stem cells form a very authentic tissue type called blastema, which is crucial for successful regeneration. In order to evaluate this exceptional tissue and acquire high quality images, it is crucial to employ specific procedures to prepare the tissue for imaging. Here, in this study, we aimed to investigate success of various fixative solutions (Carnoy's, Bouin's, % 10 NBF, Clarke's, Alcoholic Formaline and AFA) to monitor the fixed blastema. Our data reveals that integrity of the blastema tissue differs among used fixatives and a significant difference is observed between the samples in terms of staining quality.

La regeneración se define como la renovación del tejido y el proceso de restauración funcional de las partes dañadas del cuerpo después de una lesión. Ambystoma mexicanum, comúnmente llamado Axolotl, es uno de los únicos vertebrados que tiene una notable capacidad para regenerar sus miembros después de una amputación. Aunque el proceso de regeneración incluye varios períodos, se puede dividir en dos fases principales: formación del blastema y desdiferenciación. En el par de horas después de la amputación, el cierre de la herida ocurre por la migración de células epiteliales alrededor del sitio de la amputación seguido por una infiltración de macrófagos y la desdiferenciación de las células para convertirse en células madre. Las células madre acumuladas forman un tipo de tejido muy diferenciado denominado blastema, que es crucial para una exitosa regeneración. Para evaluar este tejido y adquirir imágenes de alta calidad, es crucial emplear procedimientos específicos para la obtención de imágenes. En este estudio, se intentó investigar el éxito de varias soluciones fijadoras (Carnoy, Bouin, % 10 NBF, Clarke, Formalina Alcohólica y AFA) para monitorear la fijación del blastema. Nuestros datos revelan que la integridad del tejido del blastema difiere entre los fijadores utilizados y una diferencia significativa observada entre las muestras se da en términos de la calidad de tinción.

Minced nerve tissue in vein grafts used as conduits in rat tibial nerves.

INTRODUCTION: Peripheral nerve injuries are encountered frequently in clinical practice. In nerve repair, an end-to-end suture is the preferable choice of treatment. However, where primary closure is not possible, the defect is to be repaired with a nerve graft. METHODS: A total of 21 female Wistar rats weighing 230 to 290 g were used in the study. They were classified into the following 3 groups: (I) nerve graft, (II) vein graft, and (III) minced nerve graft. In group I, after exposure of the tibial nerve, a 1-cm-long nerve gap was created on the tibial nerve, and the defect was repaired epineurally by using the autogenous nerve. In group II, the 1-cm tibial nerve defect was repaired by using an autogenous vein graft. In group III, a 1-cm nerve graft was divided to 3 equal parts, with one of the nerve parts being minced with microscissors and placed in the vein graft lumen. Thereafter, a 1-cm tibial nerve defect was repaired by the vein graft filled with minced nerve tissue. The tibial function indices (TFIs) were calculated for functional assessment using the Bain-Mackinnon-Hunter formula. Light and electron microscopic evaluations were performed for morphometric assessment. In addition, the myelinated fibers were counted in all groups. RESULTS: The TFIs of group II were found to be the lowest among all the groups after the sixth week, whereas the TFI of group I was found to be better than the other groups after the sixth week. There was no difference in TFIs between group I and group III. On the basis of the number of myelinated fibers, there was no statistically significant difference between group I and group III, whereas the difference was significant (P<0.05) between groups I/III and group II. Presence of peripheral nerves in light microscopic evaluation revealed normal characteristics of myelinated fibers in all groups. The myelinated axon profile was near normal in the nerve graft group in electron microscopic evaluation. However, there were more degenerated axons with disturbed contours and vacuolizations in the vein graft group compared to the minced nerve graft group. CONCLUSIONS: We can conclude that using minced nerve tissue in vein grafts as a conduit increases the regeneration of nerves (almost like the nerve graft group) and it may not be caused by donor-site morbidity. It can be used in the repair of nerve defects instead of autogenous nerve grafts after further experimental evidence and clinical trials.