A comparative study of the fibroblasts growth curve isolated from neck and distal limb of horse skin


1 Department of Clinical Sciences, Faculty of Veterinary Medicine, University of Zabol, Zabol- Iran

2 Department of Clinical Sciences, Faculty of Veterinary Medicine, University of Tehran, Tehran- Iran

3 Department of Food Hygiene and Quality Control, Faculty of Veterinary Medicine, University of Zabol, Zabol- Iran


BACKGROUND: Fibroblasts are one of the important cells in wound healing. These cells create a proper bed for keratinocytes migration and wound contraction.Wound healing in distal limb of horses has complications, such as formation of exuberant granulation tissue (EGT). The main factor in this problem is overgrowing of fibroblasts. ObjectiveS: The purpose of the present study was to compare fibroblast growth curve in isolated skin from horses’ neck and distal limb. Methods: 5 horses with normal hematological and clinical signs were selected. Two samples of full thickness of skin were taken from the neck and lateral metacarpal region of each horse asseptically. Then the samples were washed with PBS minced and placed in ventilated flask 25 cm2. After attaching samples to flask, 5 ml culture medium(RPMI-1640 with 10% FBS) were added and the flask was placed in an incubator at 37°c in 5% CO2. After leaving a sufficient number of cells from tissues adhered to the bottom of the flask, the cells were passaged to a new ventilated flask. After growth and proliferation of cells, they were passaged again and a suspension of cells in culture medium (10000 cells/ml) was maked. To each cell of a 24-well plate, one ml of this suspension was added. After 48 hour, cells of 3 well were detached with tripsin daily, counted and viability determinted within 8 days. Results: There was no significant difference between viable cells number but there was significant difference in viability percent of cells in neck and distal limb. The mean of population doubling time (PDT) for fibroblasts of neck is 31.73 hours and for fibroblasts of distal limb is 26.4 hours. This difference was not significant. ConclusionS: With regard to different viability percentage, it seems that the appoptosis in fibroblasts of neck skin is more regular than distal limb skin.


Azari, O., Ghamsari, S.M., Dehghan, M.M., Aghcheloo, M.R. (2007) Growth characteristics of isolated  fibroblasts from the distal limb of Thoroughberd horses and Caspian miniature horses. J Vet Res. (University of Tehran). 62: 169-176.
Bacon Miller, C., Wilson, D.A., Keegan, K.G., Kreeger, J.M., Adelstein, E.H., Ganjam, V.K. (2000) Growth Characteristics of fibroblasts isolated from the trunk and distal aspect of the limb of horses and ponies. Vet Surg. 29: 1-7.
Baxter, G.M. (1998) Wound management. In: Current Techniques in Equine Surgery and Lameness. White, N.A., Moore, J.N. (eds.). (2nd ed.) W.B. Saunders Company, Philadelphia, USA. p. 72-80.
Caron, J.P. (1999) Management of superficial wounds. In: Equine Surgery. Auer, J.A., Stick, J.A. (eds.). (2nd ed.). W.B. Saunders Company, Philadelphia, USA. p. 129-40.
Carter, C.A., Jolly, D.G., Worden, S.R., C.E., Hendren, D.G., Kane Cynthia, J.M. (2003) Platelet-rich plasma gel promotes differentiation and regeneration during equine wound healing. Exp Mol Pathol. 74: 244-255.
Cochrane, C., Rippon, M.G., Rogers, A., Walmsley, R., Knottenbelt, D., Bowler, P. (1999) Application of an in vitro model to evaluate bioadhesion of fibroblasts and epithelial cells to two different dressing. Biomaterials. 20: 1237-1244.
Deschene, K., Celeste, C., Boerboom, D., Theoret, C.L. (2012) Hypoxia regulates the expression of extracellular matrix associated proteins in equine dermal fibroblasts via HIF1. J Dermatol Sci. 65: 12-8.
Germain, L., Jean, A., Auger, F.A., Garrel, D.R. (1994) Human wound healing fibroblasts have greater contractile properties than dermal fibroblasts. J Surg Res. 57: 268-273.
Heinz, C.D., Clem, M.F. (1982) Wound healing and tissue repair. In: Text Book of Large Animal Surgery. Oehme, F.W. (ed.). (2nd ed.) Williams and Wilkins, Baltimore, USA. p. 141-153.
Hendrikson, D., Virgin, J. (2005) Factors that affect equine wound repair. Vet Clin Equine  21: 33-44.
Jacobs, K.A., Leach, D.H., Fretz, P.B., Townsend, H.G.G. (1984) Comparative aspects of the healing of excisional wounds on the leg and body of horses. Vet Surg. 13: 83-90.
Leach, D.H., Caron, J.P. (1999) The integument. In: Equine Surgery. Auer, J.A.m Stick, J.A. (eds.). (2nd ed.). WB Saunders, Philadelphia, USA. p. 121-128.
Lepault, E., Celeste, C., Dore, M., Martineau, D. Theoret, C.L. (2005) Comparative study on microvascular occlusion and apoptosis in body and limb wounds in the horse. Wound Repair Regen. 13: 520-9.
Lindsay, M.A. (1988) Wound healing in horses: guidelines for classification. J Vet Med. 83: 387-95.
McAteer, J.A., Davis, J.M. (2002) Basic cell culture technique and the maintenance of cell lines. In: Basic Cell Culture. Davis, J.M. (ed.). (2nd ed.) The Bath Press, Avon, USA. p.135-190.
Miragliotta, V., Ipina, Z., Lefebvre-Lavoie, J., Lussier, J.G., Theoret, C.L. (2008) Equine CTNNB1 and PECAM1 nucleotide structure and expression analyses in an experimental model of normal and pathological wound repair. BMC Physiol. 31: 1.
Miragliotta, V., Lefebvre-Lavoie, J., Lussier, J.G., Theoret, C.L. (2008) OB-cadherin cloning and expression in a model of wound repair in horses. Equine Vet J. 40: 643-8.
Schwartz, A.J., Wilson, D.A., Keegan, K.G., Ganiam, V.K., Sun,Y., Weber, K.T., Zhang, J. (2002) Factors regulating collagen synthesis and degradation during second intention healing of wounds in the thoracic region and distal aspect of the forelimb of horses. Am J Vet Res. 63: 1564-70.
Theoret, C.L. (2005) The pathophysiology of wound repair. Vet Clin Equine 21: 1-13.
Theoret, C.L., Barber, S.M., Moyana, T.N., Gordon, J.R. (2001) Expression of transforming growth factor β1, β2, and basic fibroblast growth factor in full-thickness skin wounds of equine limbs and thorax. J Vet Surg. 30: 269-77.
Theoret, C.L., Olutoye, O.O., Parnell, L.K., Hicks, J. (2013) Equine exuberant granulation tissue and human keloid: A comparative histopathologic study. Vet Surg. 42: 783-9.
Wilmink, J.M., Nederbragt, H., Van Weeren, P.R., Stolk, P.W.T., Barneveld, A. (2001) Differences in wound contraction between horses and ponies: the in vitro contraction capacity of fibroblasts. Equine Vet J. 33: 499-505.
Wilmink, J.M., Stolk, P.W.T., Van Weeren, P.R., Barneveld, A. (1999) Differences in second-intention wound healing between horses and ponies: macroscopical aspects. Equine Vet J. 31: 53-60.
Wilmink, J.M., Van Weeren, P.R. (2005) Second-intention repair in the horse and pony and management of exuberant granulation tissue. Vet Clin Equine 21: 15-32.