Assessment of platelet growth factors in supernatants from rehydrated freeze-dried equine platelets and their effects on fibroblasts in vitro.
Abstract: To determine whether platelet growth factors are preserved in supernatants obtained from rehydrated trehalose-stabilized, freeze-dried (lyophilized) equine platelets and whether those growth factors stimulate fibroblast proliferation and migration and enhance fibroblast-associated contraction in a collagen gel assay. Methods: 6 clinically normal adult horses. Methods: Blood samples were obtained from 6 horses, and washed platelets were prepared via differential centrifugation. Washed platelets were freeze-dried in a physiologic buffer with a mixture of trehalose and polyethylene glycol 4000. Rehydrated platelet supernatants and releasates prepared from fresh washed platelets stimulated with thrombin or platelet-activating factor were evaluated for transforming growth factor beta1 and platelet-derived growth factor-BB by use of ELISAs. Effects of rehydrated freeze-dried platelet supernatants on fibroblast proliferation, migration, and collagen gel contraction were compared with effects of 1%, 2.5%, or 10% fetal bovine serum (FBS). Results: Supernatants from freeze-dried platelets contained similar amounts of growth factors as thrombin- and platelet-activating factor-stimulated platelet releasates. The supernatants significantly enhanced fibroblast proliferation and migration in a scratch assay, compared with FBS-free control or low (1%) FBS conditions. Additionally, supernatants from freeze-dried platelets enhanced contraction of fibroblast-seeded collagen gels, compared with the effect of 1% FBS. Conclusions: The preparation technique preserved platelet growth factors, enhanced fibroblast proliferation and migration, and improved fibroblastseeded collagen gel contraction under conditions of low FBS concentration; these platelet supernatant preparations may prove useful as an aid to conventional wound management.
Publication Date: 2008-11-05 PubMed ID: 18980435DOI: 10.2460/ajvr.69.11.1512Google Scholar: Lookup
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- Comparative Study
- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research determines that growth factors in platelets from horse blood, even after undergoing freeze-drying, can stimulate growth and movement in fibroblasts, a type of cell linked to repairing tissue damage. The method used may prove useful for improving conventional wound treatment methods.
Research Methods
The procedure details:
- The extraction of blood samples from six healthy adult horses.
- The preparation of washed platelets from these samples through differential centrifugation— a method for separating different components of the blood.
- The freeze-drying of these washed platelets in a physiologic buffer mixed with trehalose and polyethylene glycol 4000, commonly used as stability agents in freeze-drying processes.
- The rehydration of the freeze-dried platelets.
- The analysis of the supernatants and releasates of these rehydrated platelets, and from fresh washed platelets stimulated with thrombin or platelet-activating factor.
Observations and Results
- The supernatants of the freeze-dried platelets contained similar amounts of growth factors—specifically transforming growth factor beta1 and platelet-derived growth factor-BB—as the stimulated platelet releasates.
- These supernatants notably amplified fibroblast proliferation and migration in a scratch assay, compared to FBS-free control or low (1%) FBS conditions.
- Moreover, these supernatants from freeze-dried platelets stimulated contraction of fibroblast-seeded collagen gels more than 1% FBS.
Implications and Conclusions
The findings of this research have significant implications. They show that:
- The procedure used to prepare these freeze-dried platelets effectively maintained the platelets’ growth factors.
- The growth factors held in the supernatants effectively enhanced fibroblast proliferation and movement—factors which play a key role in tissue repair and wound healing—as well as improving the contraction of fibroblast-seeded collagen gels under low FBS concentrations.
- These platelet supernatant preparations could therefore potentially be utilized as a supplement to traditional wound management techniques.
Cite This Article
APA
Tablin F, Walker NJ, Hogle SE, Pratt SM, Norris JW.
(2008).
Assessment of platelet growth factors in supernatants from rehydrated freeze-dried equine platelets and their effects on fibroblasts in vitro.
Am J Vet Res, 69(11), 1512-1519.
https://doi.org/10.2460/ajvr.69.11.1512 Publication
Researcher Affiliations
- Department of Anatomy, Physiology and Cell Biology, School of Veterinary Medicine, University of California, Davis, CA 95616, USA.
MeSH Terms
- Animals
- Blood Platelets / chemistry
- Cell Movement / drug effects
- Cell Proliferation / drug effects
- Centrifugation / veterinary
- Enzyme-Linked Immunosorbent Assay
- Fibroblasts / drug effects
- Freeze Drying / veterinary
- Horses
- Platelet-Derived Growth Factor / analysis
- Platelet-Derived Growth Factor / pharmacology
- Transforming Growth Factor beta1 / analysis
- Transforming Growth Factor beta1 / pharmacology
Citations
This article has been cited 8 times.- Seidel SRT, Fülber J, Barbosa ÂP, Penatti NMA, Demasi M, Baccarin RYA. Medium-term storage of platelet-derived orthobiologics: a feasible alternative for equine practice. Front Vet Sci 2025;12:1720164.
- Warin R, Vongchan P, Suriyasathaporn W, Boripun R, Suriyasathaporn W. In Vitro Assessment of Lyophilized Advanced Platelet-Rich Fibrin from Dogs in Promotion of Growth Factor Release and Wound Healing. Vet Sci 2022 Oct 15;9(10).
- Garbin LC, Contino EK, Olver CS, Frisbie DD. A safety evaluation of allogeneic freeze-dried platelet-rich plasma or conditioned serum compared to autologous frozen products equivalents in equine healthy joints. BMC Vet Res 2022 Apr 18;18(1):141.
- Seidel SRT, Vendruscolo CP, Moreira JJ, Fülber J, Ottaiano TF, Oliva MLV, Michelacci YM, Baccarin RYA. Does Double Centrifugation Lead to Premature Platelet Aggregation and Decreased TGF-β1 Concentrations in Equine Platelet-Rich Plasma?. Vet Sci 2019 Aug 21;6(3).
- Liu Z, Jin H, Xie Q, Jiang Z, Guo S, Li Y, Zhang B. Controlled Release Strategies for the Combination of Fresh and Lyophilized Platelet-Rich Fibrin on Bone Tissue Regeneration. Biomed Res Int 2019;2019:4923767.
- Seabaugh KA, Thoresen M, Giguère S. Extracorporeal Shockwave Therapy Increases Growth Factor Release from Equine Platelet-Rich Plasma In Vitro. Front Vet Sci 2017;4:205.
- Shiga Y, Kubota G, Orita S, Inage K, Kamoda H, Yamashita M, Iseki T, Ito M, Yamauchi K, Eguchi Y, Sainoh T, Sato J, Fujimoto K, Abe K, Kanamoto H, Inoue M, Kinoshita H, Furuya T, Koda M, Aoki Y, Toyone T, Takahashi K, Ohtori S. Freeze-Dried Human Platelet-Rich Plasma Retains Activation and Growth Factor Expression after an Eight-Week Preservation Period. Asian Spine J 2017 Jun;11(3):329-336.
- Li Q, Reed DA, Min L, Gopinathan G, Li S, Dangaria SJ, Li L, Geng Y, Galang MT, Gajendrareddy P, Zhou Y, Luan X, Diekwisch TG. Lyophilized platelet-rich fibrin (PRF) promotes craniofacial bone regeneration through Runx2. Int J Mol Sci 2014 May 14;15(5):8509-25.
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