C B Riley1, F J Archer, J V Bailey. 1. Department of Veterinary Anesthesiology, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, Canada.
Abstract
OBJECTIVE: To compare the effects of different commercial nutrient media and sera on protein synthesis and maintenance of cellular density in cultures of the equine superficial digital flexor tendon (SDFT). ANIMALS: 8 healthy 2- to 4-year-old horses. PROCEDURE: First Dulbecco's modified Eagle's medium, Ham's F12 nutrient mixture, RPMI 1640 medium, minimum essential medium with Earle's salts, minimum essential medium with Hanks' salts, and a Dulbecco's modified Eagle's medium/Ham's F12 nutrient mixture with 10% fetal bovine serum (FBS) were compared. Then FBS, fetal equine serum, and donor horse serum, each at 5, 10, and 15% in RPMI 1640 medium, were compared. Explants were cultured in roller bottles at 37 C and aerated (50% O2/45% N2/5% CO2) daily. Rates of [3H]-proline incorporation were used as a measure of the rates of total protein and collagen synthesis on days 13 and 28. Matrix cellular density of explants at days 14 and 28 was measured by computerized image analysis. RESULTS: Equine SDFT explants were cultured in all media for up to 4 weeks. Proline incorporation was greatest in Ham's F12 nutrient mixture and in RPMI 1640 medium, with the concentration of proline in medium correlating to the in vitro response. Total proline incorporation was greater in 15% FBS than in 5 or 10% FBS. Other differences among sera were not detected. Matrix cell density in 15% donor horse serum was equivalent to that in uncultured controls and higher than that in most other sera at week 2. CONCLUSION: The in vitro SDFT culture system described may be used in future studies to enhance knowledge of the biological and biochemical characteristics of intrinsic tendon healing.
OBJECTIVE: To compare the effects of different commercial nutrient media and sera on protein synthesis and maintenance of cellular density in cultures of the equine superficial digital flexor tendon (SDFT). ANIMALS: 8 healthy 2- to 4-year-old horses. PROCEDURE: First Dulbecco's modified Eagle's medium, Ham's F12 nutrient mixture, RPMI 1640 medium, minimum essential medium with Earle's salts, minimum essential medium with Hanks' salts, and a Dulbecco's modified Eagle's medium/Ham's F12 nutrient mixture with 10% fetal bovine serum (FBS) were compared. Then FBS, fetal equine serum, and donorhorse serum, each at 5, 10, and 15% in RPMI 1640 medium, were compared. Explants were cultured in roller bottles at 37 C and aerated (50% O2/45% N2/5% CO2) daily. Rates of [3H]-proline incorporation were used as a measure of the rates of total protein and collagen synthesis on days 13 and 28. Matrix cellular density of explants at days 14 and 28 was measured by computerized image analysis. RESULTS:Equine SDFT explants were cultured in all media for up to 4 weeks. Proline incorporation was greatest in Ham's F12 nutrient mixture and in RPMI 1640 medium, with the concentration of proline in medium correlating to the in vitro response. Total proline incorporation was greater in 15% FBS than in 5 or 10% FBS. Other differences among sera were not detected. Matrix cell density in 15% donorhorse serum was equivalent to that in uncultured controls and higher than that in most other sera at week 2. CONCLUSION: The in vitro SDFT culture system described may be used in future studies to enhance knowledge of the biological and biochemical characteristics of intrinsic tendon healing.