Cellequs: creating an haplo-bank of equine mesenchymal stem cells characterized by the major histocompatibility complex

Horses are important as patients and translational models, so there is an increasing demand to establish mesenchymal stem cells (MSC) biobanks for research and therapy. Allogeneic cell therapy presents advantages over autologous therapy, but immune recognition of foreign cells can compromise his safety and effectiveness. The immune recognition of MSCs can vary depending on the matching/mismatching between donor and patient for the major histocompatibility complex (MHC), and on the MSC immune profile. We aim at providing well-characterized allogeneic MSCs that can be MHC-matched with equine patients. To do this, we are creating CELLEQUS, a bank of equine MSC derived from donors homozygous for the most common MHC-haplotypes and characterised by their immune profile. To recruit MHC-matched donors, we first determined the most frequent MHC-haplotypes in the predominant equine breeds in Spain, using a validated panel of 10 intra-MHC microsatellites. The most common MHC-haplotypes were HapPRE10 (26%) and HapPRE11 (8%) in Purebred Spanish (PRE, n=326), HapPRA02 (23.8%) and COR42 (9.5%) in Purebred Arabian (PRA, n=165), HapPRE06 (9%) and HapPRE09 (9%) in Hispano-Arabian (HA, n=41), COR95 (16%) and HapPRE73 (13.6%) in Lusitano (PSL, n=132). Second, we recruited homozygous horses for the most frequent haplotypes as MSC donors (n=8). MSCs were isolated from bone marrow, characterized by their immunophenotype (CD90, CD105, CD44, CD45, CD11, MHC-I, MHC-II; flow cytometry) and gene expression analysis (MHC-I, MHC-II, IL6, COX2; RT-qPCR). MHC-I expression was lower in MSCs from HapPRE10 homozygotes, which might add advantages to their allogeneic use. MHC-II expression was variable but overall low regardless of the haplotype, while donors with the same haplotype can exhibit very different immunomodulatory profiles (COX2 and IL6), indicating that not all homozygous horses are equally suitable as donors. By offering a repository of MHC-matched MSCs, CELLEQUS Bank provides foundational resources for advancing veterinary regenerative medicine, encompassing research, future commercialization, and clinical advancement. CELLEQUS contributes paving the way for improved allogeneic MSC treatments by serving as a proof-of-concept that haplo-banks can be created in different animal species, both for research and clinical purposes. Furthermore, under the pivotal concept of One Medicine, knowledge on veterinary and human haplo-banks can be exchanged to advance their use in both species.