Bovine adipose tissue-derived mesenchymal stem cells self-assemble with testicular cells and integrates and modifies the structure of a testicular organoids.

Mesenchymal stem cells (MSC) display self-renewal and mesodermal differentiation potentials. These characteristics make them potentially useful for in vitro derivation of gametes, which may constitute experimental therapies for human and animal reproduction. Organoids provide a spatial support and may simulate a cellular niche for in vitro studies. In this study, we aimed at evaluating the potential integration of fetal bovine MSCs derived from adipose tissue (AT-MSCs) in testicular organoids (TOs), their spatial distribution with testicular cells during TO formation and their potential for germ cell differentiation. TOs were developed using Leydig, Sertoli, and peritubular myoid cells that were previously isolated from bovine testes (n = 6). Thereafter, TOs were characterized using immunofluorescence and Q-PCR to detect testicular cell-specific markers. AT-MSCs were labeled with PKH26 and then cultured with testicular cells at a concentration of 1 × 106 cells per well in Ultra Low Attachment U-shape bottom (ULA) plates. TOs formed by testicular cells and AT-MSCs (TOs + AT-MSCs) maintained a rounded structure throughout the 28-day culture period and did not show significant differences in their diameters. Conversely, control TOs exhibited a compact structure until day 7 of culture, while on day 28 they displayed cellular extensions around their structure. Control TOs had greater (P < 0.05) diameters compared to TOs + AT-MSCs. AT-MSCs induced an increase in proportion of Leydig and peritubular myoid cells in TOs + AT-MSCs; however, did not induce changes in the overall gene expression of testicular cell-specific markers. STAR immunolabelling detected Leydig cells that migrated from the central area to the periphery and formed brunches in control TOs. However, in TOs + AT-MSCs, Leydig cells formed a compact peripheral layer. Sertoli cells immunodetected using WT1 marker were observed within the central area forming clusters of cells in TOs + AT-MSCs. The expression of COL1A associated to peritubular myoids cells was restricted to the central region in TOs + AT-MSCs. Thus, during a 28-day culture period, fetal bovine AT-MSCs integrated and modified the structure of the TOs, by restricting formation of branches, limiting the overall increase in diameters and increasing the proportions of Leydig and peritubular myoid cells. AT-MSCs also induced a reorganization of testicular cells, changing their distribution and particularly the location of Leydig cells.

Bovine fetal mesenchymal stem cells exert antiproliferative effect against mastitis causing pathogen Staphylococcus aureus.

Staphylococcus aureus is the most commonly isolated pathogen from clinical bovine mastitis samples and a difficult pathogen to combat. Mesenchymal stem cells (MSC) are multipotent progenitor cells equipped with a variety of factors that inhibit bacterial growth. The aim of the present study was to evaluate the in vitro antibacterial potential against S. aureus of conditioned medium (CM) from MSC derived from fetal bovine bone marrow (BM-MSC) and adipose tissue (AT-MSC). BM-MSC, AT-MSC and fetal fibroblasts (FB) cultures were activated by infection with S. aureus. Bacterial growth was evaluated in presence of CM, concentrated CM (CCM), activated CM (ACM) and concentrated ACM (CACM) from BM-MSC, AT-MSC and FB. Gene expression of ß-defensin 4A (bBD-4A), NK-lysine 1 (NK1), cathelicidin 2 (CATHL2), hepcidin (HEP) and indoleamine 2,3 dioxygenase (IDO) and protein expression of bBD-4A were determined in activated and non-activated cells. The majority of BM-MSC and AT-MSC expressed CD73, Oct4 and Nanog, and were negative for CD34. Growth of S. aureus decreased when it was exposed to CM from BM-MSC, AT-MSC and FB. Moreover, growth of S. aureus in CCM, ACM and CACM was lower compared to controls of CM from BM-MSC and AT-MSC. Activated AT-MSC increased mRNA levels of bBD4A and NK1, and protein levels of bBD4A in CM. Thus, CM from fetal bovine BM-MSC and AT-MSC has the capacity to reduce in average ~30% of S. aureus relative growth under in vitro conditions. The in vitro antibacterial effect of fetal bovine MSC may be mediated by bBD4A and NK1 activity.