采用不同载体的玻璃化冷冻对人卵巢组织冻存效果的比较研究

摘要

目的 比较采用不同载体的玻璃化冷冻方案对人卵巢组织的卵泡形态、冻融后体外培养时卵泡生长和激素分泌能力的影响,寻找临床上简便易行且保存效果上佳的人卵巢组织玻璃化冷冻方案.方法 将16例患者的卵巢组织切割成皮质小条后随机分配到新鲜组和不同载体玻璃化组,包括冷冻管组、最小微滴组、不锈钢网筛组和固相表面玻璃化组.观察各组卵巢组织卵泡形态;冻融后的卵巢组织行体外培养,比较卵泡生长情况以及培养液中雌二醇和孕酮的浓度.结果 冻融后各组原始卵泡正常形态率均低于新鲜组,差异有统计学意义(P < 0.05).冷冻管组、最小微滴组、不锈钢网筛组和固相表面组,原始卵泡形态正常率分别为(64.57±11.84)%、(78.36±7.62)%、(77.21±6.51)%和(84.70±5.03)%.固相表面组原始卵泡形态正常率明显高于其他玻璃化组(P < 0.05).体外培养液中雌二醇和孕酮的平均水平随培养时间逐渐升高.固相表面玻璃化组两种激素平均浓度明显高于其他玻璃化组.培养后卵巢组织中原始卵泡所占比例均较新鲜组下降,生长卵泡比例升高.固相表面玻璃化组培养后生长卵泡比例明显高于其他玻璃化组.结论 固相表面玻璃化冷冻可以保存大部分原始卵泡的正常形态,并能较好地保存其体外生长和性激素分泌功能.该技术简便易行且冷冻效果好,适合人卵巢组织玻璃化冷冻的临床使用.%Objective To compare the effect of vitrification with different carriers for follicular morphology, follicular growth and hormone secretion capacity when cultured after freezing and thawing in vitro of human ovarian tissue, and to look for good simple and save effect on vitrification program of human ovarian tissue. Methods Human ovarian tissue slices from 16 patients were randomly allocated to fresh group and different vitrification groups, including TV group, MDS group, SLV group and SSV group, respectively. Follicle morphology of ovarian tissue was observed in each group; ovarian tissue after freezing and thawing was cultured in vitro, follicle growth situation and estradiol and progesterone concentration of the culture medium were compared. Results The percentages of morphologically normal primordial and primary follicles were significantly reduced (P < 0.05) in all cryopreserved tissue. The normal rate of primordial follicles morphologically was (64.57±11.84)%, (78.36±7.62)%, (77.21 ±6.51)%, (84.70±5.03)% in TV group, MDS group, SLV group, SSV group, there was a significant difference in SSV group and the other groups (P < 0.05). The levels of estradiol and progesterone in culture medium increased constantly with cultured time. SSV group showed the highest hormone levels compared with the other vitrification groups. After incubation, the proportion of primordial follicles in ovarian tissue decreased than that in the fresh group, the proportion of growing follicles increased; in the SSV group, growing follicles was significantly higher than those of the other vitrification group. Conclusion Solid phase surface vitrification can save most of the normal form of primordial follicles, and can better preserve their growth in vitro and sex hormone secretion function. The technology is simple and good for freezing effect, suitable for clinical use of human ovarian tissue vitrification.