辛通畅络中药复方对血管平滑肌细胞凋亡影响的体外研究

A study on effect of sheen unobstructed network compound formula of traditional Chinese medicine on apoptosis of vascular smooth muscular cells in vitro

目的观察辛通畅络中药复方对慢性肾脏病（CKD）心血管危险因素的改善作用。方法选择sD远交系健康sPF厂vAF级雄性大鼠，体质量250—300g。以辛通畅络中药复方（柴胡12g，黄芩10g，生黄芪30g，当归10g，女贞子15g，泽兰15g，水蛭10g，细辛3g）连续灌胃大鼠10d，然后处死大鼠取动脉血，制备含10％、20％、30％含药血清培养液，体外培养血管平滑肌细胞（VSMC），以正常大鼠血清为空白对照，每组样本数为20。采用原位末端缺刻标记试验（TUNEL）和流式细胞仪分别检测各组大鼠VSMC凋亡阳性率。结果随着含中药血清浓度的增加，VSMC凋亡率逐渐增加。TUNEL法显示：含10％药物血清组VSMC凋亡率明显高于空白对照组[（9．20±0．68）％比（7．32±0．43）％，P〈0．01]，含20％药物血清组VSMC凋亡率明显高于含10％药物血清组[（10．80±0．36）％比（9．20±0．68）％，P〈0．05]，30％药物组VSMC凋亡率高于20％含药血清组，但差异无统计学意义[（11．99±0．83）％比（10．80±0．36）％，P〉0．05]。流式细胞仪检测结果显示，各含药血清组VSMC凋亡率均较空白对照组增加，且以含30％药物血清组较含10％药物血清组和含20％药物血清组增加更显著[（28．31±5．24）％比（8．30±0．66）％、（12．04±1．10）％，P〈0．05]。结论辛通畅络中药复方可以剂量依赖性地改善CKD的心血管功能，其机制可能与诱导VSMC凋亡有关。

Objective To observe the effect of symplectic unobstructed network compound prescription of traditional Chinese medicine （TCM） on cardiovascular risk factors of chronic kidney disease （CKD）. Methods Heahh male Sprague-Dawley （SD） rats in a SPF/VAF grade weighting 250 - 300 g were collected. The TCM decoction of sheen unobstructed network compound formula （ingredients： radix bupleuri 12 g, radix scutellariae 10 g, raw radix astragali 30 g, angelica sinensis 10 g, fructus ligustri lucidi 15 g, eupatorium 15 g, leech 10 g, asari 3 g） was continuously given by lavage to rats for 10 days, then they were sacrificed and their arterial blood was harvested. The serum culture solutions containing the TCM compound 10%, 20% and 30% were prepared, the vascular smooth muscle cells （VSMC） were cultivated in vitro, and the normal rat serum was used as the culture solution in blank control （n = 20）. The positive rate of rat VSMC apoptosis was detected by terminal-deoxynucleoitidyl transferase mediated nick end labeling （TUNEL） and flow cytometry in different groups, respectively. Results With the increase of serum TCM concentration in culture, the VSMC apoptosis rate was increased gradually. TUNEL method showed： The VSMC apoptosis rate in culture solution containing 10% drug serum group was obviously higher than that of blank control group [（9.20 ± 0.68）% vs. （7.32 ± 0.43）%, P 〈 0.01], in 20% drug serum group, the rate was markedly higher than that in 10% drug serum group [（10.8 ± 0.36）% vs. （9.20 ± 0.68）%, P 〈 0.05], and in 30% drug serum group, the rate was higher than 20% drug serum group, but there was no statistically significant difference [（11.92 ± 0.83）% vs. （10.80 ± 0.36）%, P 〉 0.05]. The results of flow cytometry revealed that the VSMC apoptosis rate was higher in the groups with different serum drug concentrations than that in control group, and the elevation in apoptosis rate in 30% drug group was more significant than that in 20% or 10% drug serum group [（28.31 ± 5.24）% vs. （8.30 ± 0.66）%, （12.04 ± 1.10）%, P 〈 0.05]. Conclusions The study suggests that the sheen unobstructed network of TCM compound formula induce dose-dependent improvement of cardiac vascular function of patients with chronic kidney disease, and the mechanism is possibly related to the induction of apoptosis of vascular smooth muscular ceils.