SDF-1和VEGF在糖尿病大鼠视网膜病变中的表达及AMD3100干预作用
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温州市科技局对外科技合作交流项目(No.H20100012)


The molecular biological expression of SDF-1 and VEGF in rat diabetic retinopathy and the intervention effect of AMD3100
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Scientific and Technological Cooperation and Exchange Item of Wenzhou Science and Technology Bureau(No.H20100012)

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    摘要:

    目的:探讨不同时期糖尿病大鼠视网膜中基质细胞衍生因子-1(stromal cell derived factor-1,SDF-1)和血管内皮生长因子(vascular endothelial growth factor,VEGF)mRNA的表达,以及不同剂量SDF-1拮抗剂3100对SDF-1表达的抑制作用。

    方法:将实验大鼠分为正常组、拮抗剂组、糖尿病组。腹腔注射链脲佐菌素建立大鼠糖尿病模型。拮抗剂组、糖尿病组分别于成模后1wk玻璃体腔注射AMD3100和PBS。随后各组球后多次球后注射相应药物。实验方法采用RT-PCR和Western-Blot。RT-PCR实验共饲养三组大鼠60只,各组均于1,3,5mo处死部分大鼠,各组提取标本,RT-PCR检测并HE染色观察视网膜血管增生情况。Western-Blot实验将18只大鼠分为正常对照组、4组不同剂量AMD3100的拮抗剂组及糖尿病组共6组,饲养时间均为3mo。

    结果:RT-PCR检测1,3,5mo各同龄组中三组间大鼠视网膜SDF-1和VEGF mRNA的表达均有显著统计学差异(P<0.01),且均以糖尿病组表达最高,拮抗剂组的表达低于糖尿病组。随着糖尿病病程的延长,SDF-1和VEGF mRNA的表达进一步增加,视网膜HE染色切片上突破内界膜的内皮细胞核数明显增加。Western-Blot检测表明,在一定范围内随着拮抗剂AMD3100浓度的增大,SDF-1和VEGF的蛋白表达量逐渐减少,差异具有显著统计学差异(P<0.01); 当继续增加浓度(玻璃体腔注射>10μg/μL),SDF-1和VEGF的蛋白表达量无明显改变,差异无统计学意义(4和5组相比,P>0.05)。

    结论:随着糖尿病视网膜病变病程的进展,糖尿病大鼠视网膜组织里VEGF和SDF-1 mRNA的表达量增加。SDF-1受体拮抗剂 AMD3100可降低SDF-1和VEGF的蛋白表达。这种拮抗作用在一定的浓度范围内,其抑制作用有剂量依从性,剂量越大,抑制作用越强。

    Abstract:

    AIM: To measure the expression of stromal cell derived factor-1(SDF-1)and vascular endothelial growth factor(VEGF)in retina of diabetic rats model at the different stage and explore the inhibitory effect of AMD3100 on the expression of SDF-1 and VEGF mRNA by RT-PCR and Western-Blot test.

    METHODS: RT-PCR and Western-Blot tests were carried out. In RT-PCR test, 60 adult SD rats were divided into normal group, antagonist group, and diabetes group. After diabetic rat model was induced using streptozotocin and antagonist group and diabetic group were injected intravitreously and postocularly with AMD3100 and PBS respectively. All rats were killed and the retina was extracted. after 1,3,5 months and the HE stain of paraffin sections was used and the expression of SDF-1 and VEGF mRNA were measured with RT-PCR. In Western-Blot test, 18 rats were divided into normal group, diabetes group and four antagonist groups which were using different concentration of AMD3100, and killed after 3 months.

    RESULTS: SDF-1 and VEGF mRNA were expressed in normal group, antagonist group and diabetes group. At the same age group(1, 3 and 5 months)and among the normal group, antagonist group and diabetes group, the difference of expression of SDF-1 and VEGF mRNA were significant. The expressions in diabetic group were always highest and antagonist group lower than diabetic group. The expression of SDF-1 and VEGF mRNA was increased significantly with the extension of disease. The HE Stain of paraffin sections showed DM group had more cell nucleus which protruded internal limited membranes than normal control group and antagonist group. The Western-Blot test showed in 4 antagonist groups the SDF -1 and VEGF protein expression levels gradually decreased with the increases of SDF-1 antagonist AMD3100 concentration, the difference was significant. When intravitreous injected concentration of AMD3100 increased over 10μg/μL, the expression of SDF-1 and VEGF protein did not change, the difference was not statistically significant

    CONCLUSION: With the progression of diabetic retinopathy, the expression of VEGF and SDF-1 mRNA in the retinal tissue of diabetic rats increased. The antagonist AMD3100 could reduce the expression of SDF-l, VEGF and inhibit the development of new blood vessels. In a certain concentration range, this inhibitory effect of AMD3100 was dose-dependent.

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游逸安,赖中燕,周倩倩. SDF-1和VEGF在糖尿病大鼠视网膜病变中的表达及AMD3100干预作用.国际眼科杂志, 2013,13(10):1960-1964.

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  • 收稿日期:2013-05-30
  • 最后修改日期:2013-09-16
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  • 在线发布日期: 2013-09-23
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