title: Preparation and application of an innovative thrombocyte/leukocyte-enriched plasma to promote tissue repair in chelonians.
abstract: Platelet concentrates are widely used in mammalian regenerative medicine to improve tissue healing. Chelonians (Testudines) would benefit from the application of thrombocyte preparations to regenerate damaged tissues, since traumatic injuries are leading causes of morbidity and mortality for both wild-living and domesticated animals. The aim of this study was to establish a protocol that optimized the recovery of the thrombocytes from blood samples and to show the efficacy of thrombocyte-enriched plasma in chelonians. Peripheral blood samples were obtained from Testudo spp. (n = 12) and Trachemys scripta elegans (n = 10). Blood cells were fractionated by sodium diatrizoate-sodium polysucrose density gradient using a two-step centrifugation protocol. Thrombocytes and leukocytes were isolated and resuspended to obtain thrombocyte-leucocyte rich plasma (TLRP). The mean recovery of leukocytes and thrombocytes was 48.9% (±4.0 SEM, n = 22) of the whole blood cell content. No statistically significant difference was observed between blood samples collected from different turtle species. The ability of TLRP to form a gel was evaluated by adding variable concentrations of calcium gluconate at room temperature and at 37°C. A reliable and consistent clotting of the TLRP was obtained in glass tubes and dishes by adding 5-20% v/v of a 100 mg/ml solution of calcium gluconate. Furthermore, in order to test the clinical efficacy of TLRP, a preliminary evaluation was performed on four turtles (Testudo spp.) with traumatic injuries. In all the four animals, a successful clinical outcome was observed. The results demonstrated that a thrombocyte-enriched plasma, comparable to mammalian platelet rich plasma, can be prepared from chelonian blood samples. Furthermore, although the low number of cases presented does not allow definitive conclusions from a clinical point of view, their outcome suggests that TLRP application could be further investigated to improve the healing process of both soft and hard tissue injuries in chelonians.
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Di Ianni
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Merli
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Burtini
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Virna
authors.initials: V
authors.lastname: Conti
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Igor
authors.initials: I
authors.lastname: Pelizzone
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Rosanna
authors.initials: R
authors.lastname: Di Lecce
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Enrico
authors.initials: E
authors.lastname: Parmigiani
authors.affiliation: Clinica Veterinaria Ambulatorio Squassino, Asti, Italy.
authors.forename: Gian Paolo
authors.initials: GP
authors.lastname: Squassino
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Maurizio
authors.initials: M
authors.lastname: Del Bue
authors.affiliation: Osteoarticular Regeneration Laboratory, Rizzoli Orthopaedic Institute, 40136, Bologna, Italy.
authors.forename: Enrico
authors.initials: E
authors.lastname: Lucarelli
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Roberto
authors.initials: R
authors.lastname: Ramoni
authors.affiliation: Department of Veterinary Science, Università degli Studi di Parma, Via Del Taglio 10, 43126, Parma, Italy.
authors.forename: Stefano
authors.initials: S
authors.lastname: Grolli
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000166:cytology*; D002498:Centrifugation; D015925:Cryopreservation; D007962:Leukocytes / Q000166:cytology*; D010949:Plasma / Q000166:cytology*; D044968:Regenerative Medicine / Q000379:methods*; D014426:Turtles / Q000097:blood* / Q000293:injuries* / Q000502:physiology