Treatment of Equine Tarsus Long Medial Collateral Ligament Desmitis with Allogenic Synovial Membrane Mesenchymal Stem/Stromal Cells Enhanced by Umbilical Cord Mesenchymal Stem/Stromal Cell-Derived Conditioned Medium: Proof of Concept.

Horses are high-performance athletes prone to sportive injuries such as tendonitis and desmitis. The formation of fibrous tissue in tendon repair remains a challenge to overcome. This impels regenerative medicine to develop innovative therapies that enhance regeneration, retrieving original tissue properties. Multipotent Mesenchymal Stem/Stromal Cells (MSCs) have been successfully used to develop therapeutic products, as they secrete a variety of bioactive molecules that play a pivotal role in tissue regeneration. These factors are released in culture media for producing a conditioned medium (CM). The aforementioned assumptions led to the formulation of equine synovial membrane MSCs (eSM-MSCs)-the cellular pool that naturally regenerates joint tissue-combined with a medium enriched in immunomodulatory factors (among other bioactive factors) produced by umbilical cord stroma-derived MSCs (eUC-MSCs) that naturally contribute to suppressing the immune rejection in the maternal-fetal barrier. A description of an equine sport horse diagnosed with acute tarsocrural desmitis and treated with this formulation is presented. Ultrasonographic ligament recovery occurred in a reduced time frame, reducing stoppage time and allowing for the horse's return to unrestricted competition after the completion of a physical rehabilitation program. This study focused on the description of the therapeutic formulation and potential in an equine desmitis treatment using the cells themselves and their secretomes.

Allogenic Synovia-Derived Mesenchymal Stem Cells for Treatment of Equine Tendinopathies and Desmopathies-Proof of Concept.

Tendon and ligament injuries are frequent in sport horses and humans, and such injuries represent a significant therapeutic challenge. Tissue regeneration and function recovery are the paramount goals of tendon and ligament lesion management. Nowadays, several regenerative treatments are being developed, based on the use of stem cell and stem cell-based therapies. In the present study, the preparation of equine synovial membrane mesenchymal stem cells (eSM-MSCs) is described for clinical use, collection, transport, isolation, differentiation, characterization, and application. These cells are fibroblast-like and grow in clusters. They retain osteogenic, chondrogenic, and adipogenic differentiation potential. We present 16 clinical cases of tendonitis and desmitis, treated with allogenic eSM-MSCs and autologous serum, and we also include their evaluation, treatment, and follow-up. The concerns associated with the use of autologous serum as a vehicle are related to a reduced immunogenic response after the administration of this therapeutic combination, as well as the pro-regenerative effects from the growth factors and immunoglobulins that are part of its constitution. Most of the cases (14/16) healed in 30 days and presented good outcomes. Treatment of tendon and ligament lesions with a mixture of eSM-MSCs and autologous serum appears to be a promising clinical option for this category of lesions in equine patients.

Human umbilical cord tissue-derived mesenchymal stromal cells attenuate remodeling after myocardial infarction by proangiogenic, antiapoptotic, and endogenous cell-activation mechanisms.

INTRODUCTION: Among the plethora of cells under investigation to restore a functional myocardium, mesenchymal stromal cells (MSCs) have been granted considerable interest. However, whereas the beneficial effects of bone marrow MSCs (BM-MSCs) in the context of the diseased heart are widely reported, data are still scarce on MSCs from the umbilical cord matrix (UCM-MSCs). Herein we report on the effect of UCM-MSC transplantation to the infarcted murine heart, seconded by the dissection of the molecular mechanisms at play. METHODS: Human umbilical cord tissue-derived MSCs (UCX®), obtained by using a proprietary technology developed by ECBio, were delivered via intramyocardial injection to C57BL/6 females subjected to permanent ligation of the left descending coronary artery. Moreover, medium produced by cultured UCX® preconditioned under normoxia (CM) or hypoxia (CMH) was collected for subsequent in vitro assays. RESULTS: Evaluation of the effects upon intramyocardial transplantation shows that UCX® preserved cardiac function and attenuated cardiac remodeling subsequent to myocardial infarction (MI). UCX® further led to increased capillary density and decreased apoptosis in the injured tissue. In vitro, UCX®-conditioned medium displayed (a) proangiogenic activity by promoting the formation of capillary-like structures by human umbilical vein endothelial cells (HUVECs), and (b) antiapoptotic activity in HL-1 cardiomyocytes subjected to hypoxia. Moreover, in adult murine cardiac Sca-1+ progenitor cells (CPCs), conditioned medium enhanced mitogenic activity while activating a gene program characteristic of cardiomyogenic differentiation. CONCLUSIONS: UCX® preserve cardiac function after intramyocardial transplantation in a MI murine model. The cardioprotective effects of UCX® were attributed to paracrine mechanisms that appear to enhance angiogenesis, limit the extent of the apoptosis, augment proliferation, and activate a pool of resident CPCs. Overall, these results suggest that UCX® should be considered an alternative cell source when designing new therapeutic approaches to treat MI.

Towards an advanced therapy medicinal product based on mesenchymal stromal cells isolated from the umbilical cord tissue: quality and safety data.

INTRODUCTION: Standardization of mesenchymal stromal cells (MSCs) manufacturing is urgently needed to enable translational activities and ultimately facilitate comparison of clinical trial results. In this work we describe the adaptation of a proprietary method for isolation of a specific umbilical cord tissue-derived population of MSCs, herein designated by its registered trademark as UCX®, towards the production of an advanced therapy medicinal product (ATMP). METHODS: The adaptation focused on different stages of production, from cell isolation steps to cell culturing and cryopreservation. The origin and quality of materials and reagents were considered and steps for avoiding microbiological and endotoxin contamination of the final cell product were implemented. Cell isolation efficiency, MSCs surface markers and genetic profiles, originating from the use of different medium supplements, were compared. The ATMP-compliant UCX® product was also cryopreserved avoiding the use of dimethyl sulfoxide, an added benefit for the use of these cells as an ATMP. Cells were analyzed for expansion capacity and longevity. The final cell product was further characterized by flow cytometry, differentiation potential, and tested for contaminants at various passages. Finally, genetic stability and immune properties were also analyzed. RESULTS: The isolation efficiency of UCX® was not affected by the introduction of clinical grade enzymes. Furthermore, isolation efficiencies and phenotype analyses revealed advantages in the use of human serum in cell culture as opposed to human platelet lysate. Initial decontamination of the tissue followed by the use of mycoplasma- and endotoxin-free materials and reagents in cell isolation and subsequent culture, enabled the removal of antibiotics during cell expansion. UCX®-ATMP maintained a significant expansion potential of 2.5 population doublings per week up to passage 15 (P15). They were also efficiently cryopreserved in a DMSO-free cryoprotectant medium with approximately 100% recovery and 98% viability post-thaw. Additionally, UCX®-ATMP were genetically stable upon expansion (up to P15) and maintained their immunomodulatory properties. CONCLUSIONS: We have successfully adapted a method to consistently isolate, expand and cryopreserve a well-characterized population of human umbilical cord tissue-derived MSCs (UCX®), in order to obtain a cell product that is compliant with cell therapy. Here, we present quality and safety data that support the use of the UCX® as an ATMP, according to existing international guidelines.