| BackgroundA plenty of Epidemic and clinical investigation showed that incidence of atherosclerosis(AS) was increased among the patients with chronic inflammatory or autoimmune disease. The clinical picture implied that a intimate relationship existed between AS and immunoinflammatory respond.Above these,a hypothesis was introduced that AS was an immunoinflammatory disease.But dose the presence of inflammation result in atherosclerosis, or just result from it? Cytokines is the representative products of immunoinflammatory respond. It would contribute to the investigation of the immunoinflammatory target to identify its regular patterns. NF-κB and AP-1 are the common molecules in the upstream of cytokines and mediators of inflammation. JNK and p38 have a close relationships with immunoinflammatory respond. So It is important to identify the rules of those molecules during AS for revealing the immunoinflammatory mechanism of AS. Saponins of Panax Notoginseng(PNS) played an important role in anti-atherosclerosis and suppressing active chronic inflammation.So it canbe presumed there are some relationships beteween the two actions.Then it is very important to obtain the dynamic informations of those molecules in AS region for reveal the rule of its signal transduction and prevention and cure the immunoinflammatory AS with PNS.Methodes1. The inflammatory effect by Zym was observated while intraperitoneal injection the suspl(20mg/kg) of Zymosan A(Zym) made with liquid paraffin to rats every six days.AS was induced with the combination of Zym and ovalbumin (OVA) base on cholesterol diet.2. RayBio? Rat Cytokine Antibody Array 1.1 was applied to detect the dynamic informations of cytokines during the immunoinflammatory AS and the effects of PNS.3. Immunohistochemistry was used to detect the dynamic informations of NFkB/P65 and c-Jun/AP-1 in the region of AS.The foam cells was induced by oxLDL, oxLDL+Zym, oxLDL+Zym +PNS50μg, oxLDL+Zym +PNS100μg with peritoneal macrophage of mouse. The levels of Phospho-p38 MAP Kinase and Phospho-SAPK/JNK was detectd with Western blot. Results1. Compared with pro-injection the white blood cells counts of rats were increased from 24 hours to 120 hours, and it was the highest at 24 hours, then it was decreased to normal level at 144 hours after the second injection of Zym. None of animals was dead during the three weeks of the whole period of experiment. It showed that the controllable inflammation can be induced in this way. Rats were stimulated with the combination of Zym and OVA based on cholesterol diet for ten weeks. The serum total cholesterol (TC) and circulating immune complex (CIC) were higher than control groups in model groups.The foam cells origin from monocytes were found in the aorta walls.2. It was showed that IL-1α,MCP-1,TNF-αwere higher than control in the model groups in rat cytokine antibody array and the high levels of IL-1α,MCP-1can be decreased by PNS but not TNF-αin third week of the period of experiment. The expressed levels of more chemotatic factors were detected at the sixth week, for example CINC-2,CNTF,FKN,LIX while the expression of IL-6 begin to increase and TNF-αwas still in the high level. The inhibitory effects of PNS were significantly. GM-CSF,IFN-γ,Leptin were increased obviously excepted some chemotatic factors and immunoinflammatory factors in the tenth week. They could be decreased by PNS. CINC-2,LIX,TNF-α,VEGF were in a persistent heighten tendency in the model groups during the whole period of exeriment while CINC-3,GM-CSF,IFN-γ,IL-1β,Leptin heightened gradually and Fractalkine,IL-6,IL-1αfluctuated. MCP-1 increased significantly at third week but surgeless after that. MIP-3a,TIMP-1 changed little during the whole period. Exceped IL-6 increased in sixth week, the two anti-inflammtory factors IL-4,IL-10 did not change obviously.3. NF-κB/P65 expressed in the nuclei of endothelial cell and inflammatory cell in sub-endothelia mainly in the aorta walls of rats. It implied that NF-κB was activited in these cells. NF-κB increased gradually from third week accompany with the proceeding of AS in model group but not in control. c-Jun/AP-1 expressed in the nuclei of all type of cells in the aorta walls concluding endothelial cells, inflammatory cells in sub-endothelia and smooth muscle cells. The high expression was persistent from third week during the whole period of experiment. The changes of the two molecules were inhibited by PNS significantly. The form of foam cells from macrophage was contributed by Zym and The sum of deposite lipid in the foam cells increased obviously by Zym. The effect could be inhibited by PNS. Phospho-p38 MAP Kinase and Phospho-SAPK/JNK expressed little in group of ox-LDL but higher in group of ox-LDL+Zmy. The levels decreased when PNS presented in a dose dependently. Only Phospho-JNK2 were detected in the western blot system but not Phospho-JNK1.Conclusions1. The foam cells can be induced by combination of Zym with OVA in the aorta wall in rats basing on the cholesterol diet. It is a practicable model of immunoinflammatory As in rodents (rat). Simultaneously, immunoinflammatory response is the important reason of pathogenesis of AS except of hyperlipemia.2. It was shown that the main effect came from different cytokines in the different stages of immunoinflammatory As by the information from rat cytokines antibody array. The broken balance between anti-inflammatory factors and pro-inflammatory factors was the criticality in pathogenesis of AS. This must be the reason of those clinical picture that the incidence of AS remained significiant increase in all world although the drug of anti-hyperlipermia have been applied widely and was increased among the patients with chronic inflammatory or autoimmune disease.3. The excess expression of NF-κB/p65 and c-Jun/AP-1 is one of the mechanism of cytokines disproportion. Intervention the two common upstream molecules can regulate the levels of those disorder cytokines.4. The abnormal activity of JNK and p38 were involved in the proceeding of foam cell induced by combination of Zym and oxLDL.JNK2 was the main subset of JNK in it. The expression of cytokines were regulated by Phospho-p38 MAP Kinase and Phospho-SAPK/JNK through many transcription factors including AP-1and NF-κB.5. It would not present the desired effect by intervention only one of the cytokines because of the cross talking among the enormous cytokines.The integerate levels of cytokins in AS could be regulated by PNS. This is why PNS can prevent and cure AS. The inhibition of the abnormal activity of NF-κB,AP-1,p38,JNK is the mechanism of regulation the cytokines and mediators of inflammation then prevention and cure AS by PNS. |
