Characterization of equine mesenchymal stromal cells from non-invasive sources

INTRODUCTION: Mesenchymal stromal cells (MSC) have been extensively studied for their promising capabilities in regenerative medicine. In general, most cell-based therapies consist of the use of autologous MSC. Nevertheless, the low immunogenicity of MSC – indicated by the absence of the expression of major histocompatibility complex (MHC) class II and co-stimulatory molecules CD40, CD80 and CD86 – and their immunosuppressive capacities, enable their use in clinical allogeneic applications [1]. An allogeneic source would provide an off-the-shelf, more standardized and readily available product without the inherent lag period associated with isolation and expansion of autologous MSC [2,3]. Although bone marrow is the best known source for isolating equine MSC, alternative sources such as umbilical cord blood (UCB), umbilical cord matrix (UCM) and peripheral blood (PB) have been reported also. METHODS: Equine MSC were isolated from 6 mares (PB) and their foals (UCB & UCM) at parturition. Following parameters were analyzed: (i) success rate of isolation, (ii) proliferation capacity, (iii) tri-lineage differentiation ability, (iv) immunophenotypical protein and (v) immunomodulatory mRNA profiles. Linear regression models were fit to determine the association between the source of MSC (UCB, UCM, PB) and (i) the moment of first observation, (ii) the moment of first passage, (iii) cell proliferation data, (iv) the expression of markers related to cell immunogenicity, and (v) the mRNA profile of immunomodulatory factors, except for hepatocyte growth factor (HGF) as no normal distribution could be obtained for the latter variable. To evaluate the association between the source of MSC and the mRNA expression of HGF, the non-parametric Kruskal-Wallis test was performed instead. RESULTS: While equine MSC could be isolated from all the UCB and PB samples, isolation from UCM was only successful in 2 samples. Proliferation data showed that equine MSC from all three sources could be easily expanded, although UCB-derived MSC appeared significantly faster in culture than PB- or UCM-derived MSC. Equine MSC from both UCB and PB could be differentiated towards the osteo-, chondro- and adipogenic lineage, in contrast to UCM-derived MSC where only chondro- and adipogenic differentiation could be confirmed. Regardless of the source, equine MSC expressed the immunomodulatory genes CD40, CD80, HGF and TGF-β. In contrast, no mRNA expression was found for CD86, IDO and TNFα. DISCUSSION & CONCLUSIONS: Our data strengthen recent findings that inherent differences exist between MSC from different tissues, as suggested for human MSC. Combining all the observations in this present study, we propose UCB as the most promising non-invasive alternative source for MSC and UCM as the least feasible source due to high contamination risks. Moreover, our data indicate that UCB-derived MSC could be suited for allogeneic use, although their immunogenicity potential needs to be addressed in more detail in future studies.