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目的:探讨β1整合素反义寡核苷酸(ASODN)对精氨酸加压素(AVP)诱导大鼠心脏成纤维细胞(CFs)DNA合成功能的抑制作用。方法:以胰蛋白酶消化法分离、培养SD大鼠的CFs,采用MTT吸光度法及3H-胸腺嘧啶核苷(3H-TdR)掺入法,原位酶联免疫吸附测定(ELISA)等技术,观察基础状态下β1整合素ASODN对CFs细胞数目及DNA合成功能的影响;β1整合素ASODN对AVP诱导的β1整合素表达,CFs增殖及DNA合成功能的影响;基础状态下设空白对照组、反义链组和正义链组;AVP诱导下设空白对照组、1×10-7mol/L AVP组、反义链+1×10-7mol/L AVP组和正义链+1×10-7mol/L AVP组,每组均为8个复孔。结果:①反义链组的CFs细胞数目及3H-TdR掺入率明显低于空白对照组及正义链组,并且均有统计学意义(P<0.01);正义链组与对照组比较无统计学意义。②1×10-7mol/LAVP组和正义链+1×10-7mol/LAVP组的β1整合素表达水平,CFs细胞数目均高于空白对照组(P<0.05),β1整合素ASODN与AVP共同作用组的β1整合素表达水平,CFs细胞数目明显低于1×10-7mol/LAVP组和正义链+1×10-7mol/L AVP组,并且均有非常显著性意义(P<0.01)。③1×10-7mol/L AVP组和正义链+1×10-7mol/L AVP组的CFs3H-TdR掺入率均高于空白对照组(P<0.05),β1整合素ASODN与AVP共同作用组的CFs3H-TdR掺入率明显低于空白对照组、1×10-7mol/L AVP组和正义链+1×10-7mol/L AVP组,并且均有非常显著性意义(P<0.01)。结论:β1整合素ASODN不但抑制基础状态下CFs生长及DNA合成代谢,而且抑制AVP刺激β1整合素表达、CFs增殖、DNA合成的作用,说明针对特异性靶基因片段合成的ASODN可能抑制β1整合素遗传信息传递的某个环节,干扰细胞内外信息的传递,影响β1整合素介导的CFs与细胞外基质的黏附,从而产生拮抗AVP刺激心脏间质重构形成的作用。进一步提示应用反义药物防治高血压心脏间质重构的可能性。
AIM: To investigate the inhibitory effect of β1 integrin antisense oligonucleotide (ASODN) on arginine vasopressin (AVP) -induced DNA synthesis in rat cardiac fibroblasts (CFs). Methods: CFs of Sprague-Dawley rats were isolated and cultured by trypsin digestion. MTT assay and 3H-thymidine incorporation (3H-TdR) The effect of β1 integrin ASODN on the expression of β1 integrin, CFs proliferation and DNA synthesis induced by AVP was observed under the basal condition. The blank control group, antisense 1 × 10-7mol / L AVP group, 1 × 10-7mol / L AVP group and 1 × 10-7mol / L AVP group Groups, each with 8 replicate wells. Results: ① The numbers of CFs and 3H-TdR incorporation in the antisense group were significantly lower than those in the blank control group and the sense chain group (both P <0.01); there was no statistical difference between the sense strand group and the control group Significance of learning. The expression of β1 integrin and the number of CFs in 1 × 10-7mol / LAVP group and the sense strand + 1 × 10-7mol / LAVP group were significantly higher than those in the blank control group (P <0.05), and β1 integrin ASODN interacted with AVP The expression of β1 integrin and the number of CFs in CFs were significantly lower than that in 1 × 10-7mol / L AVP group and the sense strand + 1 × 10-7mol / L AVP group (P <0.01). The incorporation rate of CFs3H-TdR in the 1 × 10-7mol / L AVP group and the sense strand + 1 × 10-7mol / L AVP group was higher than that in the blank control group (P <0.05). The combined effect of β1 integrin ASODN and AVP CFs3H-TdR incorporation rate was significantly lower than the blank control group, 1 × 10-7mol / L AVP group and the sense chain + 1 × 10-7mol / L AVP group, and have very significant significance (P <0.01). CONCLUSIONS: β1 integrin ASODN not only inhibits the growth of CFs and DNA synthesis and metabolism in the basal state, but also inhibits the expression of β1 integrin, CFs proliferation and DNA synthesis induced by AVP, indicating that ASODN synthesized against specific target gene fragments may inhibit β1 integrin A certain part of the transmission of genetic information interferes with the transmission of intracellular and extracellular information and affects the adhesion between β1 integrin-mediated CFs and the extracellular matrix, thereby resulting in the antagonism of the formation of cardiac interstitial remodeling by AVP. Further prompted the use of antisense drugs to prevent hypertensive cardiac interstitial remodeling possibilities.