Abstract
Simple SummaryRegenerative medicine using platelet-based blood products or adult stem cells offers the prospect of better clinical outcomes with many diseases. In veterinary medicine, most progress has been made with the development and therapeutic use of these regenerative therapeutics in horses, but the clinical need is given in dogs as well. Our aim was to transfer previous advances in the development of horse regenerative therapeutics, specifically the use of platelet lysate for feeding stem cell cultures, to the dog. Here, we describe the scalable production of canine platelet lysate, which could be used in regenerative biological therapies. We also evaluated the canine platelet lysate for its suitability in feeding canine stem cell cultures in comparison to equine platelet lysate used for equine stem cell cultures. Platelet lysate production from canine blood was successful, but the platelet lysate did not support stem cell culture in dogs in the same beneficial way observed with the equine platelet lysate and stem cells. In conclusion, canine platelet lysate can be produced in large scales as described here, but further research is needed to improve the cultivation of canine stem cells.Platelet lysate (PL) is an attractive platelet-based therapeutic tool and has shown promise as xeno-free replacement for fetal bovine serum (FBS) in human and equine mesenchymal stromal cell (MSC) culture. Here, we established a scalable buffy-coat-based protocol for canine PL (cPL) production (n = 12). The cPL was tested in canine adipose MSC (n = 5) culture compared to FBS. For further comparison, equine adipose MSC (n = 5) were cultured with analogous equine PL (ePL) or FBS. During canine blood processing, platelet and transforming growth factor-β1 concentrations increased (p < 0.05 and p < 0.001), while white blood cell concentrations decreased (p < 0.05). However, while equine MSC showed good results when cultured with 10% ePL, canine MSC cultured with 2.5% or 10% cPL changed their morphology and showed decreased metabolic activity (p < 0.05). Apoptosis and necrosis in canine MSC were increased with 2.5% cPL (p < 0.05). Surprisingly, passage 5 canine MSC showed less genetic aberrations after culture with 10% cPL than with FBS. Our data reveal that using analogous canine and equine biologicals does not entail the same results. The buffy-coat-based cPL was not adequate for canine MSC culture, but may still be useful for therapeutic applications.
Highlights
Regenerative medicine has gained tremendous attention in recent years in the veterinary field, including a growing number of studies and clinical applications in dogs.In regenerative medicine, biological materials such as cells, growth factors, and matrix substances are used to regenerate defective tissues and organs, aiming to regain their full functionality
We showed that the obtained equine PL (ePL) supported equine mesenchymal stromal cells (MSC) expansion and basic characteristics [29]
Confirming the findings described above, the metabolic activity of canine MSC was decreased in canine PL (cPL) 10% medium as compared to fetal bovine serum (FBS) medium (p < 0.05 in P5), and even more decreased in cPL 2.5% medium (p < 0.05 in passage 3 (P3), p < 0.01 in P4)
Summary
Regenerative medicine has gained tremendous attention in recent years in the veterinary field, including a growing number of studies and clinical applications in dogs.In regenerative medicine, biological materials such as cells, growth factors, and matrix substances are used to regenerate defective tissues and organs, aiming to regain their full functionality. MSC-based therapies are supported by a growing body of evidence in veterinary medicine. Canine MSC have been used successfully in several in vivo studies, especially for the treatment of osteoarthritis [8,9,10], and for various other conditions such as atopic dermatitis [11,12], diabetes mellitus [13], inflammatory bowel disease [14], or to support neuroregeneration following vertebral compression fractures [15]. One of the critical factors is the in vitro cultivation of MSC before administration to the patient. In this context, the cell culture medium represents a crucial element of the cell culture process and may strongly affect efficacy and quality of therapy [16]. The critically discussed fetal bovine serum (FBS) is still the gold standard for in vitro cultivation of MSC in animal species
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