Expression of PD-L1 on Peripheral Blood T Cells of Patietients with Coronary Heart Disease and Regulation of Simvastatin and Captorpril
|School||Southern Medical University,|
|Keywords||Simvastatin PD-L1 Patients with coronary heart disease Captopril Lymphocytes ACS Statins ACEI Regulatory role Costimulatory signal Unstable angina Acute coronary syndrome Normal control group Vascular endothelial function Thrombotic complications Damage response doctrine Atherosclerosis Inhibitors Peripheral blood Apoptosis|
Coronary heart disease (Coronary Heart Disease, CHD) is one of the contemporary threat to human healthy aging disease, the leading cause of death of the middle-aged. Therefore, the prevention and treatment of coronary heart disease, reduce the incidence rate has been the concern of the world. China's CHD prevalence and mortality rates showed an increasing trend, according to the World Health Organization (WHO) estimates: around the year 2020, China will usher in the \Atherosclerosis (atherosclerosis, AS) is a major pathological basis of coronary heart disease. The atherosclerosis pathogenesis studies have experienced a century and a half, a large number of AS pathogenesis doctrine, which consist primarily of endothelial injury doctrine, the doctrine of lipid infiltration, thrombosis borne doctrine arterial smooth muscle cell proliferation doctrine. 1999, Ross R Professor new arguments on the basis of the doctrine of his damage response clearly stated that \The study confirmed that the inflammation involved in various stages of of AS formation and development until the final thrombotic complications. In recent years, the role of the immune factors in the occurrence and development of AS being awareness and attention, more scholars AS is an autoimmune disease. In the AS, the variety involved in the immune cells and inflammatory factors constitute the intricate mutual coordination interaction network, inflammation, and immune to influence the process of the AS. T lymphocytes (peripheral blood T cells, PBTCs) is a major immune cells in the atherosclerotic plaque. AS pathological features of smooth muscle cell proliferation, lipid deposition, macrophage and lymphocyte infiltration and foam cell formation. By T-cell-specific CD3 antibody staining in AS, different periods of development have different degrees of T lymphocytes, the majority of these T cells is active, the expression of adhesion molecules and other surface molecules, secretion of cytokines and proliferation. The active plaque unstable plaque, T cells increased significantly. Many studies on humans and rodents confirmed, T lymphocytes promote atherosclerotic plaque inflammation reaction, and affect the progress of the damage and remodeling. T cell activation is to play a pathophysiological role in the premise, the activation requires two signals, the first signal is the TCR-CD3 complex of the surface of T lymphocytes with antigen presenting cells (APC) on the MHC-Ⅱ class antigen peptide complex combination, given the specificity of the immune response, but does not induce T cell proliferation and cytokine secretion; second signal is antigen-nonspecific costimulatory signal, stimulated by T cells co ligand binding molecules and the surface of APC induction start to maintain and regulate the activation cascade reaction to determine the T-cell activation and proliferation, or unresponsive state even apoptosis. PD-L1 (programmed death-ligand-1) molecule is B-7 family members, which are widely expressed in lymphocytes and other peripheral tissues and cells. Studies have confirmed that the PD-L1 having two receptors: the receptors have been identified the PD-1 (Programmed death-1) and another unknown receptor. PD-L1 interaction with different receptors play a positive and negative dual role. PD-L1 costimulatory signal whether to participate in the development and progression of the AS, what is its mechanism of action, it is not yet conclusive. Pi-reductase inhibitors, statins (Statins) drugs are methyl hydroxyl coenzyme A (HMG-COA) can lower serum cholesterol levels, is the most commonly used class of drugs for the treatment of hyperlipidemia and AS. Studies have shown that statins reduce cardiovascular disease morbidity and mortality, not only in its lipid-lowering effects, what is also associated with a variety of other pharmacological effects, statins also include improving endothelial function, increasing NO bioavailability, anti oxidation, plaque stabilization, regulation of progenitor cells, anti-inflammatory, immunomodulatory multiple vascular protective effect. In recent years, statins anti-inflammatory effects of the increasing number of immunomodulatory function, but still not perfect. Belonging to the angiotensin-converting enzyme (ACE) inhibitors captopril, plasma and local tissue renin - angiotensin system (RAS) activity can be suppressed to reduce afterload, increased cardiac output, inhibition of the cardiac and vascular remodeling inhibition of sympathetic activity, in addition, research has shown, the ACEI class of drugs also have anti-inflammatory, restoration and maintenance of the vascular endothelial function, anti-platelet aggregation, inhibition of the oxidative modification of low density lipoprotein, statin drugs ACEI drugs can by adjusting the costimulatory signal expression of PD-L1, delaying AS progression and thus play a role, worthy of further research and exploration. Part I: patients with coronary heart disease in peripheral blood T lymphocytes PD-L1 expression purpose: to study the expression of PD-L1 in CHD patients PBTC significance discussed in AS development. Methods: CHD patients (41 cases) for the study, including 21 patients with stable angina (SA), 20 cases of patients with acute coronary syndrome (ACS), 19 cases of non-CHD patients confirmed by coronary angiography for the normal control group, density gradient centrifugation of peripheral blood mononuclear cells (PBMC), PD-L1 protein and mRNA expression in T lymphocytes were detected by flow cytometry and RT-PCR. SPSS 13.0 software for statistical processing, all the data are used (X ± s) said. P lt; 0.05 for the difference was statistically significant results: nylon wool column method separation of T lymphocytes and flow identification CD3 T cell purity gt; 80%. 2. Flow cytometry detect the protein expression of PD-L1: PD-L1 expression on the PBTC (19.657 ± 2.588)% in the SA patient group expression rate, the ACS patient group (20.114 ± 3.265)%, normal control group (7.122 ± 2.553)%. PD-L1 expression rate differences among groups were statistically significant (P lt; 0.001), and the two groups of patients with CHD PD-L1 expression was higher than the normal control group; SA group ACS group difference was not statistically significant (P = 0.607). (See Table 1-3). 3.RT-PCR detection the PD-L1mRNA in of PBTC the expression: differences of express PD-L1mRNA results between the groups was statistically significant (P lt; 0.001). The PD-L1mRNA results show that SA group patients express 0.421 ± 0.084 ACS group is 0.406 ± 0.075, higher than the normal control group, 0.211 ± 0.064; the SA group ACS group difference was not statistically significant (P = 0.694) ( Table 1-4) Conclusion: PD-L1 expression was elevated in patients with coronary heart disease PBTCs, which may play a role in the occurrence and development of AS. The second part of simvastatin and captopril in patients with coronary heart disease in peripheral blood T lymphocytes PD-L1 expression Objective To observe the intervention effect of simvastatin and captopril regulation of the expression of PD-L1 in CHD patients PBTC explore statin drugs and ACEI drugs immunoregulatory new mechanism of AS. Experimental procedure 1, simvastatin (Simvastatin) Preparation of solution: simvastatin 4.186mg was dissolved in 2 ml of anhydrous ethanol, adding 1.8mL0.1mol/LNaOH, 80 ° C, heating 4H 0.1mol/LHCl adjusted to pH value of about 7.2, add distilled water to 10 mL of made 1mmol / L simvastatin stock solution, and then diluted in distilled water, filter sterilization made of the concentration of lumol / L simvastatin solution, 4 ℃ refrigerator save spare. 2, captopril (Captopril) Preparation of solution: captopril 2.17mg was dissolved in 2 ml of anhydrous ethanol was heated 4h, 1mmol / L stock solution in distilled water was added 10ml made, and then diluted in distilled water, was removed by filtration made of the concentration of the bacteria 1umol / L at 4 ℃ refrigerator use. 3, select the eight cases of patients with coronary heart disease are onset in patients with coronary heart disease, and confirmed by coronary angiography, blood not taking cardiovascular drugs (including statins) and other immunosuppressants, sterile extract eight patients fresh anticoagulated peripheral blood 2ml, mix well, adding a 96-well plate, each hole 100ul, the same patients were divided into blank control group, the simvastatin drug treatment group, captopril drug treatment group were added the same amount of PBS solution, simvastatin solution, captopril solution. Simvastatin solution concentration of approximately: 110.32ug / L; captopril concentration of approximately: 600.2ug / L, and incubated for 12 hours. PD-L1 protein expression on T lymphocytes, statistically detected by flow cytometry: SPSS 13.0 software for statistical processing, all the data are used (X ± s) said. P lt; O.05 was considered statistically significant. Results of the control group, simvastatin group, the the captopril group of PD-L1 protein expression (see Table 2-1 and Figure 2-1, 2-2, 2-3) were: (13.446 ± 5.535)% (11.436 ± 5.647)%, (11.744 ± 5.509)%, CHD in peripheral blood of patients with two drugs for 24 hours after the T cells, PD-L1 protein downregulation. The comparison between groups: simvastatin group and captopril, respectively, compared with the blank control group differences were statistically significant (P lt; 0.001); simvastatin group and captopril group showed no significant difference between (P = 0.475). The differences between groups were statistically significant (P lt; 0.001), the grouping compatibility effective, (see Table 2-1). Conclusion: simvastatin, captopril role in peripheral blood of patients with coronary heart disease, PD-L1 expression on the PBTC decline, which may play a role in the occurrence and development of AS.