Adipose‐derived mesenchymal stem cells (ASC) hold great promise in the treatment of many disorders including musculoskeletal system, cardiovascular and/or endocrine diseases. However, the cytophysiological condition of cells, used for engraftment seems to be fundamental factor that might determine the effectiveness of clinical therapy. In this study we investigated growth kinetics, senescence, accumulation of oxidative stress factors, mitochondrial biogenesis, autophagy and osteogenic differentiation potential of ASC isolated from horses suffered from equine metabolic syndrome (EMS). We demonstrated that EMS condition impairs multipotency/pluripotency in ASCs causes accumulation of reactive oxygen species and mitochondria deterioration. We found that, cytochrome c is released from mitochondria to the cytoplasm suggesting activation of intrinsic apoptotic pathway in those cells. Moreover, we observed up‐regulation of p21 and decreased ratio of Bcl‐2/BAX. Deteriorations in mitochondria structure caused alternations in osteogenic differentiation of ASCEMS resulting in their decreased proliferation rate and reduced expression of osteogenic markers BMP‐2 and collagen type I. During osteogenic differentiation of ASCEMS, we observed autophagic turnover as probably, an alternative way to generate adenosine triphosphate and amino acids required to increased protein synthesis during differentiation. Downregulation of PGC1α, PARKIN and PDK4 in differentiated ASCEMS confirmed impairments in mitochondrial biogenesis and function. Hence, application of ASCEMS into endocrinological or ortophedical practice requires further investigation and analysis in the context of safeness of their application.

A horse suffering from an undetected hoof bone fracture was diagnosed three weeks after injury. The formation of callus tissue was detected at the fracture site. Standard orthopaedic screw application was augmented by a novel method, a combination of stem cells and plasma components. For experimental therapy, fat tissue and blood samples were collected from the patient to isolate stem cells and plasma proteins. The obtained and characterised mesenchymal stem cell population was applied to the wound area, together with an implant prepared from plasma, wrapped over the orthopaedic screw. Additionally, cells with implant were differentiated in vitro into bone tissue, to evaluate if cells could successfully produce extracellular matrix in such material. Three weeks after application, the hoof was significantly regenerated, and after three months -the bone was completely rebuilt. The in vitro experiment also gave positive results, with completely differentiated cells after three weeks. Our data show that enriching the standard orthopaedic material with mesenchymal stem cells adds therapeutic value to the treatment of refractory bone fractures.

One of the most common reasons for horse lameness is subchondral bone cysts (SBCs), which are especially evident in young horse athletes. It is believed that SBC development is strongly associated with an individual's bone growth and/or bone microstructure impairment. Current methods of SBC treatment include pharmacological treatment or surgical procedures which may allow the bone within the cyst to rebuild and be restored to properly developed bone tissue. Thus, we propose filling the SBCs with a 3D complex of alginate hydrogel and autologous adipose derived mesenchymal stem cells (ASCs). We have observed at the in vitro level, that this hydrogel complex induces osteogenic and chondrogenic differentiation potential through the upregulation of bone morphogenetic protein, osteopontin, collagen type I and aggrecan mRNA levels. Moreover, we detected the creation of a 3D extracellular matrix (EM). To investigate the complex in vivo, we chose 8 horses of varying age suffering from SBC, which resulted in lameness, to undergo experimental surgery. We documented the horses' clinical appearance, lameness and radiographic appearance, to determine that there was clinical improvement in 87.75% of the patients (n=7, out of 8 horses) 6 months postoperatively and 100% (n=8, out of 8 horses) a year after surgery. These results are promising for the potential of this procedure to become the standard in SBC treatment.

Stem cell based therapy are now commonly applied in human and veterinary medical practice especially in orthopaedics. Mesenchymal stromal stem cells isolated from adipose tissue (ASC) are first choice option due to relatively non-invasive and safe procedure of tissue harvesting. However, ASC therapeutic potential strongly rely on patients general health condition, age and life-style. For that reason, to enhance therapeutic potential of cells, they are modified in vitro using different approaches. Previous studies have shown, that ASC treated with resveratrol, herein called SIRT+, are characterised by decreased senescence, increased proliferation rate and improved clinical outcome in autologous therapies. Herein, SIRT + cells in alginate hydrogel were applied to 5 years old warm breed mare was clinically evaluated due to the left hind lameness due to subchondral bone cyst. The therapeutic effect was assessed by the analysis of lameness score and radiological evaluation. This case report demonstrates the therapeutic potential of SIRT + cells in the treatment of orthopaedics disorders in horses as complete bone remodelling occurred after therapy and horse came back to training.