Hepatocellular carcinoma and macrophage interaction induced tumor immunosuppression via Treg requires TLR4 signaling.

AIM: To investigated the interaction between toll-like receptor 4 (TLR4)-activated hepatoma cells and macrophages in the induction of tumor-immune suppression mediated by CD4+CD25(high) family of transcription factor P3 (FOXP3) regulatory T cells (Tregs). METHODS: The proportion of FOXP3+ Tregs was identified in peripheral blood and tumor tissues of 60 hepatocellular carcinoma (HCC) patients. TLR4 expression was examined in tumor tissues and cell lines. The correlation was examined between FOXP3+ Tregs in peripheral blood and TLR4 expression of HCC tissues. Following activation of TLR4 in H22 murine hepatoma cells pre-incubated with lipopolysaccharide (LPS) and co-cultured with macrophage cell line RAW246.7, the synthesis of cytokines tumor necrosis factor-α, CCL22, and interleukin (IL)-10 by the two cell lines was detected and analyzed. RESULTS: FOXP3+ Tregs were enriched in tumor sites, and circulating FOXP3+ Tregs were increased in HCC patients in correlation with multiple tumor foci and up-regulated TLR4 expression in HCC tissues. Semi-quantitative analysis indicated that TLR4 was over-expressed in HCC compared with the matched normal tissues. Cell cultivation experiments indicated that the mRNAs of IL-10 and CCL22 were significantly up-regulated in the RAW246.7 cell line when co-cultured with LPS pre-incubated H22 cells. CONCLUSION: In hepatoma cell lines, TLR4 may indirectly facilitate the recruitment of Tregs to the tumor site and promote intrahepatic metastasis through its interaction with macrophages.

The populations of CD4+CD25highFoxp3+ T regulatory cells, Foxp3 mRNA expression related to disease activity of svstemic Iupus ervthematosus / 中华风湿病学杂志

ObjectiveTo investigate the ratios of peripheral blood CD4+CD25highFoxp3+ regulatory T cells of systemic lupus erythematosus(SLE) patients,and explore its association with disease activity and nephropathy.MethodsPBMC lymphocytes in 42 patients with SLE and PBMC in 40 normal healthy donors were evaluated for the proportion of Treg cells,as a percentage of the total CD4+ cells,by flow cytometric analysis.Levels of mRNA for Foxp3 were measured with a real-time quantitative PCR.The proportion of Treg cells and its association with SLEDAI,nephropathy,serum anti-dsDNA antibody,and C3 levels were analyzed.Statistical analysis was conducted with t-test and Spearman's correlation analysis.ResultsPatients with active disease had statistically significantly lower levels of CD4+CD25highFoxp3+ Treg than normal controls [(4±3)％ vs (7±4)％,P＜0.05],while no significant difference could be found between patients with nonactive disease and normal controls(P＞0.05).The percentage of peripheral blood CD4+CD25highFoxp3+ Treg/CD4+ in patients with active disease was significantly lower when compared to patients with non-active disease [(9±6)％ vs (10±6)％,P＜0.05],and it was related to the disease activity.SLE patients with nephropathy had significantly lower levels of CD4+CD25highFoxp3+ Treg and CD4+CD25highFoxp3+ Treg/CD4+ than patients without nephropathy(P＜0.05).Foxp3 mRNA levels were lower in PBMC from active disease patients than those in non-active disease.In addition,there was a negative correlation between the populations of CD4+CD25highFoxp3+ and SLEDAI(r=-0.5892,P＜0.05).There was a negative correlation between the percentage of CD4+CD25highFoxp3+/CD4+ and SLEDAI (r=-0.4962,P＜0.05),while there was a positive correlation between the percentage of CD4+CD25highFoxp3+/CD4+ and C3(r=0.3867,P＜0.05).There was a positive correlation between the populations of CD4+CD25highFoxp3+ and Foxp3 mRNA(r=0.6142,P＜0.01 ).ConclusionThese suggest that the decrease of CD4+CD25highFoxp3+ Treg and Foxp3 mRNA expression may play a crucial role in the pathogenesis of SLE.

Regulatory T cells are increased in tuberculosis patients and closely correlate with its immunopathology / 中华微生物学和免疫学杂志

Objective To determine whether regulatory T cells(Tr)are increased in patients with tuberculosis and whether they are associated with its immunopathology.Meantime,to investigate the possibility of tuberculosis(TB)as a model for studying Tr functions.Methods The lymphocyte subsets were isolated from peripheral blood mononuclear cells by sorting with flow cytometry.Total cellular RNA was extracted and RT-PCR was performed to detect the Foxp3 mRNA in purified CD3+CIM+T cells,CD3+CD8+T cells and non-CD3+CD4+CD8+T cells.Using FACS analysis.we further investigated the distribution of Foxp3+ population in CD4+ CD25+T cells.Finally,we compared the percentage of CD4+CD25highFoxp3+T cells present in 51 active patients with tuberculosis and 40 uninfected healthy control subjects by FACS.The detection of Tr infiltration of Foxp3+ cells were performed with immunohistochemistry(IHC)method on tuberculosis pathological sections.Results Foxp3 was specific expressed in CD3+CD4+T cells,either in tuberculosis patients or healthy control subjects.Foxp3+ T cells took about 85%fraction of CD4+ CD25highpopulation.We used CD4+CD25high Foxp3+as a detective markers for Tr in the FACS analysis.The results showed that patients with active TB had a 4.4 fold higher percentage within the CD4+T cells in peripheral blood compared to healthy control group(modian,1.01%vs 0.23%,P<0.01).Much higher frequency of Tr were found along with T cells infiltration at the tuberculosis pathological tissues.A few individuals that we can followed indicated the expanded Tr was declined after curative treatment with operation.Conclusion Tr cells are increased in tuberculosis patients and closely correlate with its immunopathology.Tuberculosis should be a valuable model for Tr functional study.