Influence of culture conditions on the secretome of mesenchymal stem cells derived from feline adipose tissue: Proteomics approach.

This study aimed to describe the secretome of mesenchymal stem cells derived from feline adipose tissue (AD-MSCs) and compare the effects of different culture conditions on AD-MSC proteomics using a shotgun approach. Adipose tissue was collected from 5 female cats and prepared to culture. Conditioned media was collected at third passage, in which the cells were cultured under 4 conditions, normoxia with fetal bovine serum (N + FBS), hypoxia with FBS (H + FBS), normoxia without FBS (N - FBS), and hypoxia without FBS (H - FBS). Then, the secretome was concentrated and prepared for proteomic approaches. Secretomes cultured with FBS-free medium had more than twice identified proteins in comparison with the secretomes cultured with FBS. In contrast, hypoxic conditions did not increase protein amount and affected only a small proteome fraction. Relevant proteins were related to the extracellular matrix promoting environmental modulation, influencing cell signaling pathways, and providing a suitable environment for cell proliferation and maintenance. Moreover, other proteins were also related to cell adhesion, migration and morphogenesis. Culture conditions can influence protein abundance in AD-MSC secretome, and can give also more specificity to cell and cell-free treatments for different diseases.

Allogenic mesenchymal stem cell-conditioned medium does not affect sperm parameters and mitigates early endometrial inflammatory responses in mares.

This study aimed to evaluate the effects of mesenchymal stem cell-conditioned medium (MSC-CM) on sperm parameters, intrauterine polymorphonuclear neutrophils (PMN), intrauterine fluid accumulation (IUF), and fertility in mares. In experiment 1, two ejaculates from ten stallions were extended to 50 million sperm/mL using a milk-based extender. Thereafter, 20 mL of extended semen was added of MSC-CM as follows: 0, 5, 10, 15, and 20 mL. Sperm kinetics and plasma membrane integrity were evaluated immediately after dilution (T0) and 2 h post-incubation at 37 °C (T2). In experiment 2, mares characterized as resistant (n = 13) or susceptible (n = 7) to endometritis were inseminated with fresh semen 24 h post-induction of ovulation in two (Control and CM-1) and three (Control, CM-1, and CM-2) cycles in a crossover, as follows: control, no pharmacological interference; CM-1, supplementation of semen insemination dose at 3:4 (v:v, MSC-CM:semen); CM-2, 30 mL of MSC-CM was infused into the uterus 24 h before insemination. Endometrial cytology and uterine fluid were collected 6 and 24 h after insemination to evaluate the number of PMNs and concentrations of interleukins IL6, IL10, and TNFα. IUF was determined by ultrasonography 24 and 48 h after insemination. Pregnancy status was diagnosed 14 days after ovulation. The addition of MSC-CM to semen did not influence sperm parameters at T0 and T2 (P > 0.05) and reduced (CM-1; P < 0.05) the number of PMNs at 6 h post-insemination in resistant mares. In susceptible mares, PMNs at 6 and 24 h post-insemination, as well as IUF were reduced (P < 0.05) in both treated cycles (CM-1 and CM-2). In addition, MSC-CM downregulated IL6 and upregulated IL10 concentrations in the uterus of susceptible mares after insemination. There were no differences in fertility rates among groups both in resistant (Control, 77%, 10/13; CM-1, 62%, 8/13) and susceptible mares (Control, 42.8%, 3/7; CM-1, 57.1%, 4/7; CM-2, 85.7%. 6/7). In conclusion, MSC-CM did not affect sperm parameters when mixed with diluted semen, and reduced post-insemination inflammatory responses in mares.

Allogeneic mesenchymal stem cell transplantation in healthy equine superficial digital flexor tendon: A study of the local inflammatory response.

The superficial digital flexor tendon (SDFT) is a structure frequently affected by injuries in high-performance athletic horses, and there are limited therapeutic options. Regenerative medicine has evolved significantly in treating different illnesses. However, understanding the cellular behaviour during mesenchymal stem cell (MSC) transplantation in healthy tissues is not fully known yet. To address the inflammatory response induced by allogeneic MSC transplantation, this study evaluated the local inflammatory response after the application of allogeneic adipose tissue-derived mesenchymal stem cells (AT-MSCs) in the equine tendon compared to an autologous transplant and the control group. Eighteen thoracic limbs (TL) in nine animals were divided into three groups and subjected to the application of AT-MSCs in the healthy tendon. In the allogeneic group (Gallog), the animals received an allogeneic AT-MSC application in the TL. The autologous group (Gauto) received an application of autologous cells in the TL, and in the control group (Gcont), phosphate-buffered saline (PBS) was applied. There were no significant differences among the evaluated groups in the physical, morphological, thermography, and ultrasonography analyses. A higher number of CD3-positive lymphocytes was observed in the Gauto group compared to the control (P < 0.05). Additionally, we did not observe different expressions of CD172 and microvascular density among the groups. The allogeneic transplantation of AT-MSCs did not result in an adverse or inflammatory reaction that compromised the use of these cells in this experiment. Their behaviour was similar to that of autologous transplantation.