Faculty of Veterinary Medicine, Laboratory of Carnivore Reproduction (LRC), State University of Ceara, Fortaleza, CE, Brazil.
Theriogenology. 2010 Sep 15;74(5):749-55. doi: 10.1016/j.theriogenology.2010.03.028. Epub 2010 May 26.
The objective was to evaluate the effects of various concentrations of exogenous FSH during in vitro culture of isolated canine preantral follicles. Preantral secondary follicles (>200 microm) were isolated by microdissection and cultured for 18 d in supplemented alpha-Minimum Essential Medium (alpha-MEM). There were three treatment groups: 1) absence of FSH (control medium); 2) FSH100 (fixed concentration of 100 ng/mL throughout the entire culture period); and 3) sequential FSH (FSHSeq - 100, 500, and 1,000 ng/mL were added sequentially). Following culture, all follicles from all treatments were still viable (marked green by calcein-AM). The initial (D0) average follicle diameter for the control, FSH100, and FSHSeq was (mean +/- SEM) 298.96 +/- 7.02, 286.00 +/- 5.87, and 275.39 +/- 174 6.55 microm, respectively (P > 0.05). Mean diameter of follicles treated with FSHSeq on Day 18 (D18-439.80 +/- 14.08 microm) was greater than those of the other treatments (P < 0.05). Daily follicular growth rate (microm/d) of follicles in the FSHSeq treatment (6.47 +/- 0.55) was significantly faster than for both the control (3.67 +/- 0.32) and FSH100 (4.47 +/- 0.38) treatments. Furthermore, FSH100 and FSHSeq treatments had a significantly higher rate of antrum formation than the control group on D12 of culture, whereas after D12, FSH100 had a significantly higher rate of extrusion compared to the control (P < 0.05). In conclusion, the sequential addition of FSH to the culture medium maintained the survival of isolated canine preantral follicles and promoted an increased rate of follicular growth and antrum formation.
目的在于评估在犬类初级腔前卵泡的体外培养过程中，添加不同浓度外源性 FSH 的效果。通过显微解剖分离出大于 200μm 的次级腔前卵泡，并在添加了α-最低必需培养基（α-MEM）的条件下培养 18 天。有三个实验组：1）无 FSH（对照培养基）；2）FSH100（整个培养周期内始终保持 100ng/ml 的固定浓度）；3）序贯 FSH（FSHSeq-100、500 和 1000ng/ml 依次添加）。培养结束后，所有实验组的卵泡仍然存活（用 calcein-AM 标记为绿色）。对照组、FSH100 组和 FSHSeq 组的初始（D0）卵泡平均直径分别为（平均值 ± SEM）298.96 ± 7.02μm、286.00 ± 5.87μm 和 275.39 ± 174.65μm（P > 0.05）。FSHSeq 组在第 18 天（D18-439.80 ± 14.08μm）的卵泡平均直径大于其他两组（P < 0.05）。FSHSeq 组卵泡的每日卵泡生长率（μm/d）为 6.47 ± 0.55，显著快于对照组（3.67 ± 0.32）和 FSH100 组（4.47 ± 0.38）（P < 0.05）。此外，在培养的第 12 天，FSH100 组和 FSHSeq 组的窦腔形成率显著高于对照组，而在第 12 天之后，FSH100 组的卵泡排出率显著高于对照组（P < 0.05）。总之，在培养基中序贯添加 FSH 可维持犬类初级腔前卵泡的存活，并促进卵泡生长和窦腔形成速度的提高。
