补肾活血方对骨髓间充质干细胞移植治疗薄型子宫内膜大鼠子宫内膜CD34、整合素ανβ3、LIF表达的影响

Effect of Bushen Huoxue Recipe on the expression of CD34,integrin ανβ3 and leukocyte inhibitory factor( LIF) in the endometrium of thin endometrial rats treated with bone marrow mesenchymal stem cell transplantation

目的探讨补肾活血方对骨髓间充质干细胞(bone marrow mesenchymalstem cells,BMSCs)移植治疗薄型子宫内膜模型大鼠子宫内膜血管内皮细胞标志蛋白CD34、子宫内膜容受性标志物整合素ανβ3、白细胞抑制因子(LIF)表达的影响。方法从6~8周龄雄性SD大鼠股骨骨髓中分离提取BMSCs,采用全骨髓贴壁法将其纯化,实验选用纯度较高的第3代细胞,分别采用尾静脉(IV)和宫腔注射(IU)的方法移植到雌性SD大鼠体内;给予雌性SD大鼠宫腔灌注95%无水乙醇及灌胃羟基脲加尾静脉注射高分子右旋糖苷,建立肾虚血瘀薄型子宫内膜模型;将实验大鼠分为正常组、模型组、阳性对照组、BMSCs+IV组、BMSCs+IU组、BMSCs+IV+中药组、BMSCs+IU+中药组;各干预组分别给予BMSCs静脉或宫腔移植并联合中药干预后经过大鼠3个动情周期,处死取材。免疫组化、Western blot检测子宫内膜中CD34、整合素ανβ3及LIF的表达情况。结果 BMSCs+IV组、BMSCs+IU组、BMSCs+IV+中药组、BMSCs+IU+中药组均能一定程度地增加子宫内膜CD34、整合素ανβ3、LIF的表达(P <0.05);在免疫组化检测整合素ανβ3表达方面,BMSCs+IV组与BMSCs+IU组差异有统计学意义(P <0.01);BMSCs+IV+中药组与BMSCs+IV组,BMSCs+IU+中药组与BMSCs+IU组在免疫组化检测整合素ανβ3表达及Western blot检测整合素ανβ3、LIF表达方面差异有统计学意义(P <0.05)。结论尾静脉注射BMSCs增加整合素ανβ3表达优于宫腔注射BMSCs,加用补肾活血方后能更好地增加整合素ανβ3和LIF的表达。

Objective To investigate the effect of Bushen Huoxue Recipe on the expression of CD34,integrin ανβ3 and leukocyte inhibitory factor( LIF) in endometrium of rats with thin endometrium treated by bone marrow mesenchymal stem cells( BMSCs)transplantation. Methods BMSCs were isolated from femur bone marrow of male SD rats aged 6 ~ 8 weeks. BMSCs were purified by whole bone marrow adherent method. The cells of passage 3 with high purity were transplanted into the experimental rats by tail vein( IV) and intrauterine( IU) injection respectively. Female SD rats were perfused with 95% absolute ethanol and intrauterine injection of hydroxyurea plus high molecular dextran via tail vein to establish the model of kidney deficiency and blood stasis. The experimental rats were divided into control group,model group,positive control group,BMSCs + IV group,BMSCs + IU group,BMSCs + IV +traditional Chinese medicine group and BMSCs + IU + traditional Chinese medicine group;each group was intervened by corresponding Methods,and then the rats were sacrificed after three estrous cycles. The expression of vascular endothelial cell marker protein CD34,endometrial receptive marker integrin ανβ3 and LIF were detected by immunohistochemistry and Western blot. Results BMSCs + IV group,BMSCs + IU group,BMSCs + IV + traditional Chinese medicine group and BMSCs + IU + traditional Chinese medicine group could all increase the expression of CD34,integrin ανβ3 and LIF in endometrium to a certain extent( P < 0. 05);the expression of integrin ανβ3 in the tail vein injection group was significantly higher than that in the intrauterine injection group( P < 0. 01). There were significant differences in the expression of CD34,integrin ανβ3 by immunohistochemistry and the expression of integrin ανβ3 and LIF detected by Western blot among BMSCs + IV group,BMSCs + IU group,BMSCs + IV + traditional Chinese medicine groupand BMSCs + IU + traditional Chinese medicine group( P < 0. 05). Conclusion The expression of integrin ανβ3 increased by tail vein injection was higher than that by intrauterine injection;combined with Bushen Huoxue Decoction,the expression of integrin ανβ3 and LIF showed a better efficacy.