In vitro culture of equine preantral follicles obtained via the Biopsy Pick-Up method

The objective was to conduct a preliminary evaluation of the efficacy of two media for in vitro culture of equine preantral follicles. Ovarian cortical strips were obtained from mares (N = 10) via the Biopsy Pick-Up method during the breeding season. Ovarian tissue was immediately submitted to histological analysis (noncultured control; D0) or cultured in situ for 1 day (D1) or 7 days (D7) in either alpha-MEM or TCM-199 and submitted to histological analysis, generating five treatment groups: noncultured control, alpha-MEM:D1, TCM-199:D1, alpha-MEM:D7, and TCM-199:D7. Preantral follicles were evaluated for follicle class (primordial, transitional, primary, and secondary) and morphology (normal vs. abnormal). A total of 142 preantral follicles were analyzed in five replicates. No follicles were observed in the TCM-199:D7 treatment group. The proportion of primordial follicles was higher (P < 0.03) in the control compared to the alpha-MEM:D7 treatment group. The proportion of primary follicles was higher (P < 0.04) in the alpha-MEM:D7 treatment group compared to the control. The proportion of developing follicles (transitional, primary, and secondary) was higher (P < 0.03) in the alpha-MEM:D7 treatment group compared to the control group. There was a greater (P < 0.004) percentage of morphologically normal developing follicles in the alpha-MEM:D1 treatment group compared to the TCM-199:D1 treatment group. Overall, the percentage of morphologically normal follicles was higher in the control group (72%; P < 0.02) and alpha-MEM:D1 group (84%; P < 0.0001) compared to the alpha-MEM:D7 (27%) treatment group. Mean follicle diameter was greater (P < 0.04) in the alpha-MEM:D7 treatment group (40.6 +/- 1.1 mu m) compared to the control group (37.3 +/- 0.7 mu m). Mean oocyte diameter was greater in the alpha-MEM:D1 (31.0 +/- 0.7 mu m; P < 0.006), TCM-199:D1 (30.7 +/- 1.8 mu m; P < 0.006), and alpha-MEM:D7 (33.2 +/- 1.8 mu m; P < 0.006) treatment groups compared to the control group (27.4 +/- 0.9 mu m). In conclusion, based on these preliminary data, in vitro culture of equine ovarian fragments obtained in vivo via the Biopsy Pick-Up method promoted preantral follicle development and follicle and oocyte growth in alpha-MEM for 7 days, with some follicles remaining morphologically normal throughout the culture period. (C) 2013 Elsevier Inc. All rights reserved.