Secretoma liofilizzato (lio-secretoma) derivante da cellule stromali mesenchimali per il trattamento di patologie muscoloscheletriche in medicina veterinaria

In the last decade, regenerative medicine has gained interest in the veterinary clinical practice, since it offers a promising therapeutic alternative, especially for the treatment of musculoskeletal disorders (MSDs), characterized by multimodal management and complex healing processes. Among MSDs, osteoarthritis affects a high percentage of canine, equine, and even human populations, thus representing a one-health challenge. Furthermore, there is still a lack of effective long-term treatment for osteoarthritis. In this scenario, Mesenchymal stromal cells (MSCs) have been shown to modulate inflammation and stimulate the repair of damaged tissue, and they can have a positive long-term impact on the animal’s quality of life. Despite promising results obtained in experimental and clinical studies, MSCs therapeutic applications are seriously influenced by the in-vivo implantation of an active and proliferating cell population. Recently, the new trend has been represented by MSCs-derived secretome, a cell-free product composed of soluble factors and extracellular vesicles, with similar biological activity but greater practical advantages when compared to the cells themselves. The present thesis work focused on a freeze-dried form of secretome, called Lyo-secretome, prepared from adipose-derived MSCs from canine and equine species. The first aim was the validation of the technological process for veterinary clinical-use medicinal production of different canine and equine batches of Lyo-secretome, and its characterization in terms of quantitative and qualitative analysis. Furthermore, in vitro metabolic tests were performed on target cells, such as tenocytes, chondrocytes, and synovial fluid derived-MSCs, demonstrating positive proliferative effects. Additionally, proteomic analysis demonstrated the presence of proteins involved in regulating the inflammatory response. The next step was a double-blinded randomized controlled clinical trial, enrolling dogs and horses suffering from naturally occurring osteoarthritis to assess the in vivo safety and efficacy of Lyo-secretome. The protocol included a double intra-articular injection of Lyo-secretome resuspended in hyaluronic acid, at 40 days interval. Clinical and orthopedic examinations, as well as owner questionnaires, were used to evaluate the patients during the 180-day follow-up. Results indicated that the double intra-articular injection of allogeneic Lyosecretome is safe and does not induce systemic adverse response for both species. A mild to moderate joint flare was observed in some patients after the injection, although they were self-limiting and not correlated to the treatment, since no differences were found with the control group (hyaluronic acid). Regarding the efficacy in canine patients, the study revealed mild clinical improvement following the two injections in both treatment and control groups, without highlighting one treatment to be significantly superior, even if a positive tendency in terms of lameness and pain score, as well as an improvement in the Helsinki Chronic Pain Index score was observed in the Lyo-secretome group.