| Objectives1 Through the randomized, placebo-controlled comparison method, to study the effect of Tongguan capsule improve the cardiac function of patients with coronary heart disease after percutaneous coronary intervention and explore its possible mechanism.2 Based on the rat carotid artery balloon injury models, to observe the effect of Tongguan capsule mobilized the bone marrow endothelial progenitor cell repair the injury artery intimal and explore its possible mechanism.Methods1 Clinical researchTaking the randomized, placebo-controlled comparison method, sixty patients treated with percutaneous coronary intervention (PCI) were randomized to two groups:the treat group(n=30, treated with Tongguan Capsule and conventional western medicine) and the control group(n=30, treated with placebo and conventional western medicine). The drugs were administered to each groups after PCI,for taking 3 months.Observation indexes(1) The clinical characteristics were evaluated.(2) The patients before and 1 month after operation, testing the percentage of CD34 and VEGFR2 double positive cells in peripheral blood by flow cytometry.(3)Inspect the change of the LVEF (left ventricular ejection fraction) and ventricular wall motion parameters before and 3 months after PCI.(4) The incidence of the major adverse cardiac events (MACE) were observed during the 3 months after PCI. 2 Experimental research32 male Wistar rats, SPF,180-220g.10 rats serve as donor, bone marrow mononuclear cells were isolated from their femurs, then cultured and induced to endothelial progenitor cells. Fluorescence microscopy identified double-positive cells as EPCs. Then EPCs were marked by PKH26 (the red fluorescent dye of cell membrane), adjusting the density of 2X 107/ml.22 rats serve as receptor, which the left carotid arteries intima were denuded by balloon. In 24 hours after operation, transplanting the EPCs to receptor intravenous injection and then randomly divided into 2 groups:the control group (n=11, treat with lOmg/kg/d of PS), the treatment group(n=11, treat with 600mg/kg/d of Tongguan capsule), the drug and PS were taked by intragastrically for 4 weeks. After 4 weeks, packed blood from the mice hearts, placed at room temperature for 1 hour, then the serum were extracted by centrifugation, stored in-80℃refrigerator and then maked frozen sections of the injured carotid arteries, each specimen were cut 5 by randomly, each section 5μm.Observation indexes(1) A week later observed the reendothelialization area and calculated the ratio between the reendothelialization area and the total area of injury intima.(2) At 4 weeks, the injured left common carotid arteries were carefully removed for HE and elastic fibers staining. Intima area, media area and IA/MA were performed in IPP6.0, which is a automatic computerized imaging analysis system.(3)Through fluorescence microscope observed the PKH26 labeled EPCs expression at the injured arterial intima, involved in the repair of intimal injury.(4) Detect the concentration of VEGF and SDF-1 in mice serum of the two groups, explore the molecular mechanism of Tongguan Capsule repair the injured arterial intima.Research Results1 Clinical research(1) There had no difference between the treatment group and the control group before the operation(P>0.05). The level of CD34 and VEGFR2 both increased in the treatment group and control group at 30 days after the PCI, at the same time, The level of the CD34 and VEGFR2 in the treatment group, which had significant differences compared with the control group(P<0.05)。Testing that Tongguan Capsule can mobilize the bone marrow endothelial progenitor cell into the circulation, and then participating repair the injury endothelium.(2) At the 3 month time point after the PCI, the LVEF and ventricular wall motion parameters of the both groups are improved, indicating that the PCI treatment can significantly improve the cardiac functions of the CHD patients. Meanwhile, compared with the control group, the LVEF and the ventricular wall motion scores of the treatment group are further improved(P<0.05).(3)Main Adverse Cardiac Events:At 90 days follow-up, there didn't exist statistics difference in the incidence of death, non-fatal MI and TVR between two groups. The incidence of new heart failure of the treatment group was statistically lower than the control group (P<0.05), thus, the incidence of MACE in the treatment group was statistically lower than the control group (P<0.05).2 Experimental research(1) The bone marrow mononuclear cells were isolated from mice femurs, cultured by conditioned media, after 7 days can be differentiate into endothelial progenitor cells successfully. Through fluorescence microscope observed that endothelial progenitor cells swallowed Dil-acLDL show red fluorescence and swallowed FITC-UEA-1 show green fluorescence.(2) There are no differences at the total injured intima area in the two groups, but a week later the level of the reendothelialization area and the ratio between the reendothelialization area and the total area of injury intima in the treatment group, which had significant differences compared with the control group (average P<0.05)。(3) 4 weeks after artery endothelium denuded by balloon, the intimal hyperplasia was significantly in two groups. Compared to control group, the degree of treatment group was lower. The ratio of IA/MA in treatment group, which was significant differences compared with the control group (P<0.01)。(4) The PKH26 labeled EPCs were expressed at the injury carotid arteries of the treatment group and formed a complete neointimal, while only little scattered fluorescent light points at the injury carotid arteries of the control group.(5) At 2 weeks,4 weeks point after operation, the level of VEGF and SDF-1 in the serum of treatment group, which had significant differences compared to the control group(average P<0.01). Conclusion(1) Tongguan Capsule can improve left ventricular systolic function of patients after PCI. The possible mechanisms is that Tongguan Capsule could increase the number of EPC in peripheral blood, and then mobilize EPCs homing to ischemic myocardial and repair the injury vascular endothelial, established collateral circulation or differentiated into cardiomyocytes.(2) Tongguan capsule can accelerate reendothelialization of the injury arteries, inhibit rat carotid artery intimal hyperplasia and luminal stenosis, which were denuded by balloon. It has preventive effect to restenosis after PCI.(3) Tongguan capsule can mobilized the bone marrow endothelial progenitor cells into cieculation, participate repair the injury arteries intimal. The mechanism may be that Tongguan capsule can increase the degree of VEGF,SDF-llevels in peripheral blood.
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