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Summary

Equine neonatal chondrocytes were cultured in three-dimensional fibrin matrices under conditions of immediate implantation or implantation following monolayer culture for 6 days, and 3 cell concentrations (1 × 105, 1 × 106, and 5 × 106 chondrocytes/ cm3). Equine fibrinogen was collected by cryoprecipitation and polymerized by use of activated bovine thrombin. The fibrin implants were harvested and analyzed histologically and biochemically at 3, 7, and 14 days after the chondrocytes were implanted in fibrin. The differentiation ratio (ratio of rounded, chondrocyte-like cells to stellate, fibroblast-like cells) was statistically higher for implants that received 5 × 106 precultured cells at all time periods than for implants that received 1 × 105 or 1 × 106 precultured cells. The differentiation ratio was statistically higher for implants that received 5 × 106 immediately implanted cells than for other implants at 7 days after implantation. At 14 days, implants that received 5 × 106 precultured chondrocytes had a higher differentiation ratio than did implants that received 5 × 106 chondrocytes that had not been precultured. Among implants that received precultured chondrocytes, total glycosaminoglycan and chondroitin sulfate content was lowest for implants that received only 1 × 105 cells. Among implants that received chondrocytes that had not been precultured, glycosaminoglycan content was not significantly different among the 3 cell concentrations, and chondroitin sulfate content was different only between implants that received 5 × 106 vs 1 × 106 cells. Only after the longest incubation period and at the highest cell concentration studied did preculturing of chondrocytes improve maintenance of phenotype. Preculturing did not appear to influence proteoglycan synthesis.

Free access
in American Journal of Veterinary Research

Summary

Equine articular chondrocytes were isolated from explant cartilage cultures by digestion in a 0.075% collagenase solution for 15 to 19 hours. Cartilage from late-term fetal and neonatal foals resulted in mean chondrocyte yield of 51.99 × 106 cells/g of cartilage (wet weight), compared with a yield of 17.83 × 106 cells/g for foals 3 to 12 months old. Propagation of chondrocytes in monolayer and 3-dimensional culture was accomplished, using Ham’s F-12 as the basal medium, with supplements of fetal bovine serum (10%), ascorbic acid, α-ketoglutarate, and l-glutamine. The medium was buffered with hepes, and penicillin and streptomycin were added for microorganism control. In primary monolayer cultures of freshly isolated chondrocytes, the population doubling time was approximately 6 days. Dedifferentiation of chondrocytes toward a more fibroblastic-appearing cell was observed after the fifth passage (subculture), but was hastened by lower cell-plating density. Chondrocytes were frozen for periods of up to 9 months, using 10% dimethyl sulfoxide as the cryoprotectant. Cell viability of late-term fetal and neonatal foal chondrocytes after storage at −196 C decreased from 86% at 3 weeks to 31% at 12 weeks. Viability of cells derived from older foals and young adult horses was considerably better than that of cells from neonatal foals. Frozen chondrocytes can be stored for extended periods and thawed for immediate implantation or can be sustained in vitro in monolayer or 3-dimensional culture. Such cultures would be suitable for cartilage resurfacing experiments or in vitro assessment of various pharmaceuticals.

Free access
in American Journal of Veterinary Research

Abstract

Objective

To determine the relation between epidural injectate volume (ml/kg of body weight) and its craniad migration in calves and pigs.

Animals

23 neonatal calves and 26 feeder pigs.

Procedure

Animals were randomly assigned to receive different volumes of new methylene blue (NMB, 1.2 mg/ml in 0.9% saline solution). Injections were made into the sacrococcygeal intervertebral space in calves and the lumbosacral intervertebral space in pigs, immediately after euthanasia. Sagittal sections of the spine were made at necropsy, and craniad migration of NMB was determined and rounded to the nearest intervertebral space.

Results

In calves treated with 0.05, 0.1, or 0.15 ml of NMB/kg, mean ± SEM number of stained spinal segments was 5 ± 0.3, 8 ± 0.6, and 8 ± 0.6, respectively. Craniad migration of NMB was significantly greater for 0.15 and 0.1 ml/kg volumes versus 0.05 ml/kg. In pigs treated with 0.05, 0.1, 0.2, or 0.3 ml of NMB/kg, mean number of stained spinal segments was 8 ± 1.1, 8 ± 0.9, 10 ± 1.2, and 18 ± 2.0. Craniad dye migration was significantly greater in the 0.3 ml/kg group versus the 3 lower volume groups. Linear regression performed on both sets of data after logarithmic transformation of spaces migrated to correct for non-normality was significant (P < 0.05), and R 2 values of 0.49 and 0.55 were obtained for calves and pigs, respectively.

Conclusions

There is a significant correlation between volume (ml/kg) of NMB injected in the epidural space and its craniad migration in calves and pigs.

Clinical Relevance

Results provide a basis for determination of volume of injectate to be given to reach a minimal desired level and should be a useful baseline for future investigations of epidural drug administration. (Am J Vet Res 1997;58:786–790)

Free access
in American Journal of Veterinary Research

progressive increase in survival rates during the early neonatal period. Although an early, complete repair of TOF is preferred by some, 3 in many cases, a 2-stage approach has been advocated as the key to successfully managing these patients. 4 In the 2

Full access
in American Journal of Veterinary Research

, Kerr CL , Pearce SG , et al . Comparison of two laparoscopic suture patterns for repair of experimentally ruptured urinary bladders in normal neonatal calves . Vet Surg 2005 ; 34 : 47 – 54 . 10.1111/j.1532-950X.2005

Full access
in American Journal of Veterinary Research