| Atherosclerosis(AS)is the main pathological process of coronary heart disease(CHD),cerebrovascular accident and peripheral vascular disease.In this century,it is presently one of the most harmful human diseases with higheat morbidity.In order to conquer this disease,many medical groups have paid attention to a series of research on the aspects of pathology,epidemiology and etiology,but the main mechanism of AS is still unknown.Take a wide view on the study process of AS,it has experienced three stages from vascular repair to nonspecific and specific immunity.In the early of 20 century, AS was regarded as excess lipidoses on the arterial canal.To the 70's,Russell Ross found out the reparative process and chronic inflammation might be the main point. The damage of oxLDL on the endothelium could induce the activity of nonspecific immunity system.And a lot of leucocytes and macrophages were stimulated to migrate through the endarterium,swallowed oxLDL and overloaded to die.The toxic lipid was released to stimulate more cells.It has been generally proved that AS is the process of chronic inflammation.The artery is similar to the battlefield,toxic substances,such as oxLDL,act as attacker,and the defense is nonspecific immunity system.In the 90's,Erling Falk,Aarhus and Micheal David almost proposed that the rapid progress of AS was highly correlative with the unstability of plaque in the same time,they also indicated that unstable plaque may be comprised of more lipoids,inflammation and dead cells.Inflammatory cells gathered in the plaque,and released a great quantity of inflammatory factors and active substances.This would lead to cell necrosis and matrix degradation,and then the plaque became more unstable.Thrombosis would form and block up the blood supply,if the plaque was ruptured.And this was regarded as acute coronary syndrome,a clinic syndrome defined with the characteristic of unstable plaque and clinical manifestation of unstable angina and myocardial infarction.It has been found that neutrophilic leukocytes in the peripheral blood are increased and the serum level of TNF-α,IL-6 and CRP are also elevated in the ACS patients.Furthermore,K.Hansson found that activated T cells existed in the plaque,even in the peripheral blood were also detected.More interest was paid on the relationship between the specific immunity system and AS.The specific immunity is more specific and exact than the nonspecific immunity. The nonspecific immunity assaults with all of the material out of our own,but monoclonal T cells just recognize one specific antigen.Hundreds of millions kinds of T cells are flowing with blood and dwelling in the lymphatic organ.Specific antigen could be recognize and extracted by antigen presenting cells(APC),such as macrophagocytes and dendritic cells et al.When specific T cell clone accepts the signal from antigen presenting cells,it will be activated and differentiate.The specific T cell clone could differentiate into Th cell which can induce immune responses and Ts cell which lead to immunodepressive.The exist of activated T cells in the plaque and peripheral blood suggests that immunologic mechanism may play the important role in the genesis and development of AS.The relationship between the immune system and AS is father-child relation or brother relation.And the exist of activated specific T cells clone become the core element of the question whether AS finally belong to specific immune or nonspecific immune.T cells in the peripheral blood could be divided into different kinds of subgroups according to their phenotype and function.Usually two subgroups of CD4+ T cell and CD8+ T cell is firstly be separated on their phenotype of CD expression.And T cells can also be divided into three subgroups of help T cell(Th), cytotoxic T cell(Tc)and suppressor T cell(Ts)on their different functions.CD4+ Th cells can be classified into three subgroups of Th0,Th1 and Th2 according to the cytokine secreted.The native T cells,I.e.Th0 cell,can produce many kinds of cytokine soon after stimulated by antigen.And then Th0 cell will differentiate to Th1 and Th2 cell according to the different influence of cytokines,antigen characteristic and other hormones.Th1 cells could synthesize some cytokines such as IL-2,IFN-γand TNF-α,and promote the cellular immune.Although Th2 cells could enhance the antibody mediated humoral immunity by the secretion of cytokines of IL-4 and IL-10.In the normal circumstance,Th1/Th2 cells keep the voluntary balance to maintain proper functions.But in the pathological conditions,the balance may be broken down,the disequilibrium of subgroups possibly lead to immune disorder.In order to illuminate specific antigen activated T cells clone expands in AS,we should firstly detect whether the unbalance of T cells subgroups exist or not in the peripheral blood.T cell receptor(TCR)is the receptor of the specific antigen on the surface of T cell.The main function of TCR is antigen recognition.The beginning of immune response was based on the TCR on the CD4+ T cell to recognize the MHC-antigen compounds on the APC surface.TCR has always been proven to be the core point of the research on the stimulation of T cells.TCR is a dimeride structure consisted ofα/βorγ/δsubunit.Two peptide chains connected with disulfide bond form transmembrance polypeptides.In our peripheral bloodα/βpeptide chains is expressed by most of T cells(95%),soα/βTCR is the main antigen recognized receptor on the mature T cells,α/βTCR can distinguish the specific antigen-MHC compounds,combine with noncovalent interaction,and the stimulated signal conducts into cellular in the last.TCR consists with three complementary determining region(CDR).Among them,CDR3 is just the binding site with the antigen determinant.And CDR3 is the hypervariable region of TCR adapted with all kinds of antigen outside.That to say,different kinds of CDR3 sequences maybe represent different T cell clone.The study on the sequence of CDR3 has already proven to be the well tools reflecting the function of T cells.In order to find the expands of specific T cell clone corresponded with unknown disease,we should detect the sequence of CDR3 in the PBTC from ACS patients.Although until now the mechanism of AS is still unknown,it is too difficult in the fight with CHD.Scientists have tried their best to find the methods and drug to prevent and control the process of AS for the past century.Three main progresses were experienced in the treatment of acute myocardial infarction.The establishment of institution of cardiac intensive care and the treatment of intravenous thrombolytic therapy change the CHD treatment from conservative therapy to active therapy to pursue the best result for more survival myocardial muscle.It has been believed that the active therapy not only turns down the mortality of AMI,but also decreases the incidence of ischemia heart disease and refractory heart failure.In the past 20 years,the main noticeable improvements are coronary intervention treatment and intensive lipid lowing therapy.The intervention treatment eliminates the blockage of plaque,change the narrow situation and recover the blood perfusion. And intensive lipid lowing therapy could stabilize the plaque and control the process of AS unless lowing the hyperlipemia.But the mechanism of pleiotropic effect of statins is still unknown.Some study found that statins may have the potential capability of immunological regulation.The inflammatory reaction is the main cause of unstable plaque,and then statins could stabilize the plaque.We presume that statins maybe control the process the unstable plaque by the function of immunoregulation.So we present a follow up research on the ACS patients after six month lowing lipid therapy to observe the effect on the immune system of statins.It is expected to prove the relationships between the specific immune and the process of AS.In this study,we recruited 81 hospital patients from Jan 2006 to July 2006 in zhujiang hospital.All the patients were divided into five group:control group(Control),chest pain syndrome group with normal coronary angiograph(CPS), stable angina group(SAP),unstable angina group(UAP)and acute myocardial infarction group(AMI).In the first part,the level of Th1/Th2 cytokine in the peripheral serum and the culture supematant of PBTC were detected by enzyme-linked immunosorbent assay(ELISA).The ratio of CD4+CD28- and CD4+CD25- T cell subgroups were checked by flow cytometry(FCM).The main purpose was to analyze the change of the balance of Th1/Th2 subgroups and other subgroups and to find out the clue that activated T cells exist in the peripheral blood of AS patients.In the second part,the sequences of TCR Vβchain were analyzed by reverse transcription polymerase chain raction(RT-PCR)and gene scan to confirm the expand of monoclonal T cells.At last,we performed a follow up of statins treatment of six months to observe the change of items above,and affirmed the immuno- regulation of statins.We got the result as follows:1.The balance of T cell subgroups were broken in the peripheral blood of ACS patients1.1 Baseline clinical characteristics in patients with CHD and healthy control subjectsAll the 81 patients recruited were separated into AMI group(n=20),UAP group(n=20),SAP group(n=20),CPS group(n=11)and Control group(n=10).There is no significant difference in the baseline clinical characteristics in all of the groups(P＜0.05),such as:age;gender;smoking;the ratio of hypertension and diabetes mellitus;the serum level of total cholesterol(TC),triglyceride(TG),HDL and LDL,et al.1.2 The serum level of Th1/Th2 cytokines in all of groupsThe serum level of IL-4 in all groups was detected as 5.31±1.4,5.91±1.02, 5.7±1.4,5.47±1.1 and 6.36±1.38pg/ml,IL-10 was 32.894±9.3,33.16±9.65,32.34±9.44, 34.5±11.15 and 35.24±10.24 pg/ml.There is no significant difference among all of the groups(P＞0.05).The serum level of IFN-γin control,CPS,SAP,UAP and AMI groups was detected as 5.664±2.17,6.354±1.68,6.564±1.50,15.714±2.68 and 14.274±3.25 pg/ml and TNF-αwas 34.24±6.98,38.4±7.81,36.64±7.56,56.54±9.58 and 55.7±10.24 pg/ml.The level of IFN-γand TNF-αwere significant elevated in UAP and AMI groups than other groups(P＜0.001).1.3 The level of Th1/Th2 cytokines in the culture supematants of PBTC of all groupsThe level of IL-4 in culture supernatants of PBTC was detected as 60.94±8.7, 63.1±10.1,62.64±9.7,63.14±9.8 and 64.2±10.0 pg/ml,IL-10 was 30.14±6.15,33.7±6.24, 32.54±7.21,32.954±7.92 and 35.2±8.1pg/ml.There is no significant difference among all of the groups(P＞0.05).The level of IFN-γin culture supernatants of PBTC in control,CPS,SAP,UAP and AMI groups was detected as 1003.954±230.5, 991.84±197.2,965.5±250.9,1756.14±306.5 and 1834.44±314.5 pg/ml and TNF-αwas 476.34±98.1,486.74±115.2,482.24±125.0,861.74±172.3 and 858.74±189.4 pg/ml.The level of IFN-γand TNF-αwere significant elevated in the culture supematants of PBTC of UAP and AMI groups than other groups(P＜0.001).1.4 The ratio of CD4+CD28- and CD4+CD25- T cell subgroups in all of the groupsThe ratio of CD4+CD28- / CD4+ T cells in control,CPS,SAP,UAP and AMI groups was 2.68±0.75%,2.79±0.49%,2.68±1.71%,10.74±5.3%and 10.94±5.97%; the ratio of CD4+CD25+/CD4+ was respectively 10.29±3.71%,10.2±3.79%, 11.4±4.27%,6.43±2.68%and 5.9±2.23%.The ratio of CD4+CD28- T cell subgroups was significant increased in the peripheral blood of ACS groups;and the ratio of CD4+CD25+ T cell subgroups was decreased in ACS groups.2.The expression of monoclonal TCR Vβsubfamilies in ACS groupsThere are more than one monoclonal TCR Vβsubfamilies which can be detected in 17 cases of ACS groups;but in only three patients of SAP groups.All of the TCR Vβsubfamilies were expressed in the polyclonal way in the patients of Normal and CPS groups.In all the 20 cases of monoclonal expression,the number of patients with the expression of Vβ3,Vβ12,Vβ14,Vβ17 and Vβ22 subfamilies were more than 25%of all the cases,and were respectively 6cases,5cases,6cases, 7cases and 5cases;the number of patients with expression of Vβ6.2,Vβ7,Vβ9,Vβ18 and Vβ21 subfamilies were less than 25%,and were repectively 3cases, 3cases,2cases,1eases and 3cases.The numbers of patients with monoclonal subfamilies in ACS group were more than in other groups significantly.3 The immunoregulation function of atorvastatin on the six month follow up3.1 Effect of therapy of atorvastatin on patients with CAD after six month follow upAfter six months follow up,the serum level of TC in AMI,UAP and SAP groups was 3.89±0.26,3.98±0.35 and 3.01±0.32mmol/L,was decreased in 31.4±10.8%,22.8±7.3%and 35.3±18.8%,respectively.The serum level of LDL was 2.25±0.35,2.064±0.26 and 1.85±0.34mmol/L,and was decrease in 36.2±17.7%,37.9±11.8%and 37.6±18.9%.The level of TG and HDL were not changed remarkable. Only 1 patient in AMI groups was found with instent restenosis.There are no discrepancy in incidence rate of MACE in all groups followed up.3.2 Effect of atorvastatin on the serum level of Th1/Th2 cytokines in all of groupsThe serum level of IL-4 in SAP,UAP and AMI groups was detected as 5.15±1.72,5.88±1.07 and 6.38±1.12pg/ml,IL-10 was为35.44±6.03,32.4±5.29 and 35.4±6.0 pg/ml.There is no significant difference compared with six months before (P＞0.05).The serum level of IFN-γin SAP,UAP and AMI groups was detected as为6.07±1.04,6.75±1.02 and 6.89±1.74 pg/ml and TNF-αwas38.2±6.79,39.5±7.38 and 40.5±7.46 pg/ml.The level of IFN-γand TNF-αwere significant decrease in UAP and AMI groups compared with six months before(P＜0.001).3.3 Effect of atorvastatin on the level of Th1/Th2 cytokines in the culture supematants of PBTC of all groupsThe level of IL-4 in culture supernatants of PBTC in control,SAP,UAP and AMI groups was detected as 67.37±5.17,61.8±14.02,66.39±14.14 and 66.95±16.06 pg/ml,IL-10 was 32.5±6.81,35.3±8.2,36.3±7.94 and 39.76±8.93pg/ml.There is no significant difference among all of the groups compared with six month before (P＞0.05).The level of IFN-γin culture supematants of PBTC in control,SAP,UAP and AMI groups was detected as 1042.1±253.1,947.47±142.6,1136.4±211.7 and 1023.6±186.7pg/ml and TNF-αwas 459.7±131.6,490.88±127.3,511.74±132.8 and 519.8±159.5pg/ml.The level of IFN-γand TNF-αwere significant decreased in the culture supematants of PBTC of UAP and AMI groups compared with six months before(P＜0.001).3.4 Effect of storvastatin on the ratio of CD4+CD28- and CD4+CD25- T cell subgroups in all of the groupsThe ratio of CD4+CD28- / CD4+ T cells in control,SAP,UAP and AMI groups was 2.6±0.5%,3.7±0.94%,3.2±1.28%and 3.29±0.9%;the ratio of CD4+CD25+/CD4+ was respectively 10.05±4.95%,10.4±2.12%,10.57±2.12%and 10.24±2.03%.The ratio of CD4+CD28- T cell subgroups of ACS groups was significant decreased in the peripheral blood and the ratio of CD4+CD25+ T cell subgroups was increased compared with six months before.3.5 Effect of storvastatin on The expression of monoclonal TCR Vβsubfamilies in ACS groupsOnly 9 cases in all 17 patients of ACS groups with monoclonal expressions before were detected the same subfamilies expression after six months follow up,the remain 8 cases were in the polyclonal expression's way.In SAP group,2 cases of all the three patients with monoclonal expressions before also can detected the same subfamilies expression.The number of patients of monoclonal expression in the ACS and SAP group were comparable(P=0.3484).Through experiments of the above three parts,we can make the following conclusions:①The serum level of Th1/Th2 cytokines was changed in ACS patients,immune response may be conrespond with the pathological machenism of AS;②The level of T Th1/Th2 cytokines in the culture supematants of PBTC in ACS patients was in the unbalance way,Th1 cell was in the active condition in ACS patients;③The ratio of CD4+CD28- and CD4+CD25+ T cell subgroups were changed in ACS patients,the active T cell and the unbalance in the proliferation of T cell subgroups may play the important role in the formation of unstable plaque;④The expression of the monoclonal subfamilies of TCR Vβwere increased in ACS patients,the proliferation of monoclonal T cell increase revealed that antigen, presentation was more relatively with the unstable plaque.⑤Statins can intervene the proliferation of monoclonal T cells,the secretion of Th1 CKs,and the subgroups balance.These effects provide a theorical foundation for using the statin to act as an immune inhibitor,and have the effects of regulating immunity and anti-inflammation.Thus,the foreground of using statins to prevent and cure AS is quite vast.
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