FSH和IGF-1对大鼠卵巢细胞微囊化前后的影响

Effect of FSH and IGF-1 on ovarian cells before and after microencapsulation

目的:观察微囊化卵巢细胞的形态变化及卵泡刺激素(FSH)和胰岛素样生长因子-1(IGF-1)对其增殖功能的影响,探讨微囊化卵巢细胞作为内源性雌激素来源的可行性。方法:分离21 d龄雌性SD大鼠卵巢细胞,将培养至第一代的卵巢细胞微囊化包裹后接种于96孔板,加入含不同浓度的FSH和IGF-1(浓度分别为0、6.3、12.5、25、50、100 ng/mL)的无血清培养液;另将正常培养的卵巢细胞培养24 h后换无血清培养液进行相同的处理,培养44 h后,加入WST-1试剂,继续孵育4 h。全自动酶标仪测定光密度。结果:卵巢细胞在微囊内呈圆形均匀分布,48 h后形态无显著变化。FSH对培养的卵巢细胞和微囊化的卵巢细胞的增殖刺激作用均呈剂量依赖性,卵巢细胞的增殖刺激作用在FSH 50 ng/mL时,达到最高峰(P<0.05);对微囊化卵巢细胞的刺激作用在FSH 100 ng/mL时,细胞的增殖活性达到最高峰(P<0.05);IGF-1对卵巢细胞的刺激增殖作用在低剂量组不显著(6.3、12.5、25 ng/mL,P>0.05),在高剂量组(50、100ng/mL)对卵巢细胞的刺激作用与对照组相比差异有统计学意义(P<0.05,P<0.01)。IGF-1对微囊化卵巢细胞的刺激增殖作用呈剂量依赖性(P<0.05),在IGF-1 50 ng/mL和100 ng/mL组,微囊化卵巢细胞的OD值与对照组相比,差异有统计学意义(P<0.01)。结论:卵巢细胞在微囊化后,可以在微囊内存活并保持其生物学活性,并且可以对外源性的FSH和IGF-1的刺激产生反应。

Objective： To observe the effects of FSH and IGF-1 on ovarian cells before and after microencapsulation,and to evaluate the feasibility of microencapsulated ovarian cells as endogenous estrogen.Methods： The first generation of cultured ovarian cells from 21d old female SD rats was microencapsulated and seeded in 96 well plates,cultures with serum-free medium in different concentrations of FSH and IGF-1（concentrations were 0 ng/mL,6.3 ng/mL,12.5 ng/mL,25 ng/mL,50 ng/mL,100 ng/mL,respectively）were added;the other group was first generation of cultured ovarian cells cultured as the microencapsulated ovarian cells.All two groups of ovarian cells were cultured for 44 hours,then WST-1 was added.Automatic determination of optical density was performed after incubated for 4 hours.ResuLts： The effects of FSH on ovarian cells and microencapsulated ovarian cells all showed a dose-dependent manner.The proliferation of FSH on ovarian cells reached its peak at 50 ng/mL level（P 0.05）;and on microencapsulated ovarian cells at 100 ng/mL（P 0.05）.The difference of IGF-1 stimulation on proliferation of ovarian cells had statistical significance at high dose group（50 ng/mL,100 ng/mL）（P 0.05,P 0.01 respectively） compared to the control group.The same dose of IGF-1（50 ng/mL,100 ng/mL）on microcapsulated ovarian cells reached significant diffenece（P 0.01,P 0.01 respectively）.Conclusion： Ovarian cells can remain alive and maintain their biological activity after microencapsulated;also microencapsulated ovarian cells can respond to exogenous FSH and IGF-1 stimuLation.