Objective To investigate whether and how regulatory T cells(Tr) affect macrophages foam-cell formation, and thereby investigate the mechanism of Tr in the development of atherosclerosis. Methods Tr were isolated from lymphocyte suspensions by magnetic cell sorting-column and analyzed by flow cytometry. Macrophages were cultured alone, with CD4~+ CD25~+ T, or CD4~+ CD25~- T cells in the pres-ence of oxLDL for 48 h to transform macrophages into foam cells. Oil red O staining and cellular lipid meas-urement were used to identify macrophage foam cell formation. Semi-quantitative PCR, quantitative real-time PCR and Western blot analysis were carried out to explore the mechanism of Tr-mediated suppression on macrophages foam cell formation. Results Foam cell formation, as identified by oil red O staining, was readily apparent in cells treated with CD4~+ CD25~- T cells and without T cells. After treatment with Tr, a marked decrease(13.9% ± 5.6% ) in foam cell count was observed, compared with untreated cells(13.9% ±5.6% vs 52.9% ± 10.4%, P<0.001 ) or CD4~+ CD25~- T-treated cells(13.9% ± 5.6% vs 53. 1% ± 17.2%, P<0.001 ), 52.9% ± 10.4% and 53.1% ± 17.2%, respectively (P<0.001). The similar effect of Tr was obtained when extracted oil red O and measured by a spectrophotometer. Cholesteryl ester ac-cumulation also used to quantify foam cell formation. Compared with untreated (no T) and CD4~+ CD25~- Tr-treated macrophage cells (CD25~-), the lipids accumulation in CD4~+ CD25~+ Tr-treated macrophage foam cells(CD25~+) was significantly reduced. Total cellular cholesterol and cellular cholesteryl ester was siginifi-cantly reduced in CD25~+ cultures relative to no T[TC(total cholesterol): 57.46 ± 17.92 vs 159.48 ± 16.38, P<0.01 ; CE(esterified cholesterol): 26.68 ± 8.88 vs 102.54 ± 16.67, P<0.001] or CD25~- (TC: 58.50±7.00 vs 150.55±25.11, P<0.01; CE: 26.68±8.88 vs96.90 ± 11.95, P<0.001) cul-tures. Moreover, PCR and Western blot analysis showed that the expression of both CD36 and SRA in Tr-treated macrophage foam cells was significantly down-regulated. Conclusion Results collectively suggest that CD4~+ CD25~+ Tr cells may inhibit macrophage foam-cell formation, which is largely attributed to a down-regulated expression in scavenger receptor in Tr-treated macrophage foam cells.

The plasma levels of atrial natriuretic factor (ANF), angiotension II (AII, vaso-pressin (VP) and endogenous digitalis-like factor (EDLF) were determined in 23 patients with mi-tral stenosis before and after percutaneous balloon mitral valvuloplasty (PBMV). Before PBMV, the plasma levels of ANF and EDLF were elevated in all patients with lower level of All than nor-mal. After the procedure, with a concomitant decrease of left atrial pressure, the plasma ANF de-creased at 15 minutes (from 317. 4?96. 2 to 164. 9?56. 8ng/L , P

Objective:To evaluate whether oral administration of heat shock protein60(HSP60) could induce immune tolerance and its effect on the progression of aortic atherosclerotic plaque in hypercholesterolemic apolipoprotein(apo) E-/-mice.Methods:8 week-old mice were divided into two groups that were orally administrated PBS plus HSP60 and only PBS separately for 5 days,and then a high-cholesterol diet was started 2 days after the last treatment for 12 weeks.Pathological analysis of plaque was performed,and percentage of CD4+CD25+regulatory T cells in the splenocytes was analyzed,proliferation response of the splenocytes to HSP60 was detected and cytokines in the superanant were measured.Results:Compared with control animals,oral tolerance to HSP60 resulted in a significant decrease of atherosclerotic plaques in size.Tolerance to HSP60 induced a significant increase of CD4+CD25+ cells in the spleen.Specific proliferation response of splenocytes to HSP60 was significantly suppressed and TGF-? in the superanant increased while IFN-? decreased significantly.Conclusion:HSP60 specific tolerance can be induced via oral administration of HSP60 which in turn attenuates progression of plaque.This provides a new immunologic approach for treatment of atherosclerosis.

Objective To investigate whether and how CD4 + CD25 + regulatory T cells(Tr) affect oxidized low density lipoprotein(oxLDL) induced proinflammatory response in macrophages.Methods Tr were isolated from lymphocyte suspensions by magnetic cell sorting-column and analyzed by flow cytometry.Macrophages were cultured alone,with CD4 + CD25 + Tr or CD4 + CD25 - Tr in the presence of oxLDL for 48 h.The phenotype of macrophages was determined by flow cytometry.NO production was assessed by Griess reaction an iNOS mRNA was isolated by RT-PCR.ELISA were used to measure the production of cytokine/chemokine like MCP-1,MMP-9,TNF-ct,TGF-β and IL-10 in macrophages response to oxLDL.Results Our data showed that with oxLDL challenge,the Tr modulated macrophages have decreased NO production and iNOS expression,decreased HLA-DR and CD86 expression,and down-regulated proinflammatory cytokine/chemokine production.Conclusion Tr can inhibit the proinflammatory properties of macrophages and steer macrophage differentiation toward an anti-inflammatory cytokine producing phenotype.

AIM: To study the role of NOS2 in the devel op ment of cardiac dysfunction after rat myocardial infarction by observing effects of S-methylisothiourea (SMT) on left ventricular morphology and haemodynamics. METHODS: An selective NOS2 inhibitor, was used. Administration o f SMT by gavage began 30 min before coronary ligation. Six weeks later, left ventricular morphologic and haemodynamic parameters were o bserved,and NOS2 expression, plasma NO 2-/NO 3- level and myocardial fibr osis were studied. RESULTS: Six weeks after myocardial infarction, NOS2 level in ra t non-infarct cardiac muscle, plasma NO 2-/NO 3- level, CVP and LVEDP were higher than that in controls. Long term administration of SMT decreased plasma NO 2-/NO 3- level [(26.6?6.1) ?mol/L vs (50.1?10.4) ?mol/L, P

AIM: To evaluate whether tolerogenic dendritic cells (DC) loaded with heat shock protein 60 (HSP60) inhibit the progression of aortic atherosclerotic plaque in hypercholesterolemic apolipoprotein E (Apo-E) -null mice. METHODS: Bone marrow derived DC of the mice were loaded with HSP60 and co-cultured with rapamycin to generate tolerogenic DC. The tolerogenic DC, DC loaded only HSP60 and PBS were injected into the ApoE-null mice at 8 weeks of age for three times at a one-week interval. 8 weeks after the last injection, aorta were harvested for HE staining and anti-CD4~+T cell immunostaining. Responses of pleenic cells to HSP60 were also evaluated. RESULTS: Compared with DC, DC_ HSP60 expressed higher levels of CD86, and stimulated T lymphocytes to proliferation significantly, while the tolerogenic DC expressed lower levels of CD86, and inhibited T lymphocytes to proliferation. After immunization with different injection, the numbers of CD4~+ T cells in plaque were increased significantly in DC_ HSP60 group vs in PBS group (P

Objective:To investigate the expression of transcription factor BACH2 on PBMC in patients with coronary artery disease( CAD) and discuss its significance and role.Methods:The circulating levels of BACH2 and TLR4 in PBMC were evaluated by RT-PCR and Western blot separately.Serum levels of signature cytokines of T-helper cells,cytokines and ox-LDL were evaluated by ELISA.The data among groups were analyzed by one-way ANOVA.Correlation between variables were subjected to linear regression analysis.Results:The expression level of BACH2 down-regulated and TLR4 up-regulated significantly in PBMCs of ACS patients had significantly higher concentrations of serum ox-LDL,cytokines of TH1 cells,TH2 cells and TH17 cells and low level of cytokine of Treg cells.Negative correlation exists significantly between BACH2 and TLR4 in patients with CAD and between BACH2 and IFN-γ,IL-4 and IL-17,and between BACH2 and ox-LDL in serum.However, positive correlation exists significantly between BACH2 and TGF-β1.Conclusion:Increased serum concentrations of ox-LDL probably down-regulated the expression of BACH2 in ACS patients by activating TLR4 on PBMC, resulting in promoting production of pro-inflammatory cytokines and suppressing production of anti-inflammatory cytokine and then might cause ACS.

AIM: To examine whether tolerogenic dendritic cells(DC) loaded with heat shock protein 60(HSP60) could restore endothelial function in hypercholesterolemic apolipoprotein E(apoE)-null mice.METHODS: Bone marrow derived DC of the mice was loaded with HSP60 and co-cultured with rapamycin to generate tolerogenic DC.The tolerogenic DC,DC loaded only with HSP60(DChsp) and saline were injected into the apoE-null mice at 6 weeks of age for two times at a one-week interval.C57BL/6 mice at the same age were taken as normal control two weeks after the last injection.Aorta was harvested for ex vivo vascular ring tension test.Immune parameters were also analyzed in vitro and in vivo.RESULTS: Compared with the non loaded DC,HSP60 pulsed DC expressed higher levels of CD86,and stimulated T lymphocytes to proliferation significantly,while the tolerogenic DC expressed lower levels of CD86,and inhibited T lymphocytes to proliferation.After immunization with different injection,Ach-induced relaxation was reduced significantly in(DChsp) group compared with saline group(P

To investigate the function of dendritic cells (DC) in patients with unstable angina, 10 mL of blood was drawn from 30 subjects. 15 patients diagnosed as having unstable angina and 15 healthy subjects were included in an observation and a control groups respectively. The mononuclear cells were separated from the peripheral blood and cultured in RPMI1640 supplemented with recombinant human granulocyte/macrophage-colony stimulating factor (rh GM-CSF) and recombinant human interleukin-4 (rh IL-4) to induce dendritic cells. The shape and ultrastructure of DC was examined with electronic microscope. The phenotype of DC was analyzed with FACS and the alloantigen presenting capacity of DC was evaluated by mixed lymphocyte reaction (MLR). The expression rate of CD86 of DC in patients with unstable angina was (40.7±3.6) %, which was obviously higher than that of normal DC (29.6±2.5 %) (P<0.001). The capacity of the DCs in unstable angina patients to induce allogenic T cells (OD 2.73±1.10), was significantly higher than that of the normal DC (OD:0.9±0.21) (P<0.005). It is suggested that the function of DC in patients with unstable angina is increased, which may play an important role in the initiation of immune reaction in the plaque.

The function of CD4+CD25+regulatory T lymphocytes (Treg) in patients with acute coronary syndrome (ACS) and the effects of atorvastatin were investigated. Forty-eight patients with ACS were randomly divided into two groups: group C receiving conventional therapy (n=24), and group C+A receiving conventional therapy+atorvastatin (10 mg/day, n=24). T lymphocytes from ACS patients (before and 2 weeks after the treatment) or 18 healthy subjects were separated and the flow cytometry was used to measure the percentage of Treg. The inhibitory ability of Treg on effector T cells was determined by mixed lymphocyte reaction (MLR). ELISA was used to measure the serum levels of cytokines (IL-10, TGF-β1 and IFN-γ) before and after treatment. The results showed that as compared with normal control group, Treg percentage was decreased significantly (P<0.01), the in- hibitory ability of Treg on the T lymphocytes proliferation was reduced (P<0.01), IFN-γ, levels were increased and IL-10 and TGF-β1 levels were lowered in ACS patients. After treatment with atorvas- tatin, Treg percentage and the inhibitory ability of Treg on T lymphocytes proliferation were signifi- cantly increased in ACS patients. Serum IFN-γ, was decreased significantly, while IL-10 and TGF-β1 were elevated significantly as compared with the non-atorvastatin group. The number of Treg was positively correlated with serum TGF-β1, but negatively with serum IFN-γ and CRP. It was concluded that ACS was associated with decreased number and defected function of Treg, which may play an important role in initiating immune-inflammatory response in ACS. The inhibitory ef- fects of atorvastatin on inflammation in ACS may be due to its beneficial effects on Treg and restora- tion of immune homeostasis.