Short- and long-term effects of antiretroviral therapy on peripheral regulatory CD4+/CD25hi/CD127low T lymphocytes in people living with HIV/AIDS

ABSTRACT The effect of antiretroviral therapy (ART) on CD4+/CD25hi/CD127low T lymphocyte changes in people living with HIV/AIDS (PLWHA) is still a matter of debate. From October 2015 to December 2019, peripheral blood from 70 cases of PLWHA were collected for the detection of CD4+/CD25hi/CD127low T lymphocytes by flow cytometry. Statistical analysis was performed to detect changes of CD4+/CD25hi/CD127low T lymphocytes in patients with different duration of ART and different treatment effects. We found that the number of CD4+/CD25hi/CD127low T lymphocytes in ART-naive PLWHA were lower than those in healthy volunteers (10.3±٦.٠ cells/uL vs 31.7±8.0 cells/uL, P < 0.05). CD4+/CD25hi/CD127low T lymphocyte counts increased to 17.8±٤.٠ cells/uL 6 months post-ART and 25.0±١١.٩ cells/uL 9 months post-ART, respectively (P < 0.05). There was no significant difference in CD4+/CD25hi/CD127low T lymphocyte counts between PLWHA who reached a complete immune reconstruction after ART and healthy volunteers. The growth of CD4+/CD25hi/CD127low T lymphocyte counts in patients who had baseline CD4 > 200 cells/uL was greater than those who had baseline CD4 ≤ 200 cells/uL (12.6±٤.٦ cells/uL vs 5.6±٥.٠ cells/uL, P = 0.027). CD4+/CD25hi/CD127low T lymphocyte counts were positively correlated with CD4+ T lymphocyte counts (r = 0.923, P < 0.001) and CD4+/CD8+ ratio (r = 0.741, P < 0.001), but were negatively correlated with HIV-VL (r = −0.648, P = 0.000). In conclusion, the results of the present study showed that changes in CD4+/CD25hi/CD127low T lymphocyte counts can be used to assess the effect of ART in PLWHA.

Regulatory T cells (CD4+CD25+FOXP3+) in lupus nephritis

Background: Systemic lupus erythromatosus (SLE) is an autoimmune disease with 20–65% of patients developing lupus nephritis (LN). Studies have reported 10% of LN patients will end up with end stage renal disease and their mortality rate is higher compared to patients without LN. Abnormality of regulatory T cells (Tregs) level is thought to be a potential factor for this LN development. The aim of study was to evaluate the percentage of Tregs in LN patients.Methods: This was a comparative cross sectional study involving LN patients and age and gender matched controls with a 2:1 ratio. The patients were grouped into active and inactive LN based on their lupus activity index; complement levels, ANA, dsDNA antibodies, ESR, SLE Disease Activity Index (SLEDAI2K) score and also urine PCI (uPCI>0.05 for active group). Disease history, demographic data, routine blood test, peripheral blood for differentials count were taken and recorded. Peripheral blood mononuclear cells were stained with CD4, CD25 and Foxp3 antibodies and percentage of Tregs was analysed using BD fluorescence-activated cell sorting (FACS) cytometer. We compared demographic and laboratory parameters between healthy controls and LN patients as well as active and inactive LN patients.Results: A total of 34 LN patients (32 females, 2 males) were recruited. Their mean age and disease duration were 37.97±11.14 years and 110.95±65.07 months respectively.  Thirteen matched controls with mean age 35.23±7.89 years were enrolled. There was no demographic difference between 2 groups of LN patients. Tregs were significantly lower in active LN compared to inactive LN and healthy control (0.44±0.37% vs. 1.89±0.46% vs. 3.12±0.56% of the CD4+, P<0.001). C3 and C4 complement fragments were significantly reduced in patients with active disease (C3; 50.92±28.43 vs. 76.31±25.63, P=0.011) and (C4; 11.17±8.41 vs. 16.70±6.50 P=0.044). Proteinuria was significantly higher while serum albumin levels were significantly lower in active patients compared to inactive patients and healthy control (urine PCI; 0.25(0.15-0.3) vs. 0.03(0.01-0.05) vs. 0.01, P<0.001) and (albumin; 29.89±6.87 vs. 36.87±3.58 vs. 40.62±1.89mmol/L, P<0.001). We found positive inversely correlation between Tregs with SLEDAI2K (r = -0.572, P=0.011) and proteinuria (r = -0.451, P=0.007).Conclusions: Tregs, C3 and C4 complements, and albumin were significantly lower while proteinuria was significantly higher in active LN. There was positive inversely correlation between the percentage of Tregs with SLEDAI2K score and proteinuria.

T cell receptor rearrangement excision circles ( TRECs) and CD31+ regulatory T cells for assessing recent thymic output in patients with chronic hepatitis B / 中华微生物学和免疫学杂志

Objective To evaluate the clinical value of combined detection of T cell receptor rear-rangement excision circles ( TRECs) and CD31+ regulatory T ( Treg) cells for accessing the recent thymic output in patients with chronic hepatitis B. Methods Four groups involving 135 subjects were set up in this study as follows: mild chronic hepatitis B ( Mild CHB, n=35 ) , moderate chronic hepatitis B ( Moderate CHB, n=35 ) , severe chronic hepatitis B ( Severe CHB, n=35 ) and healthy control ( HCs, n=30 ) groups. CD4+CD25+Treg cells in these subjects were sorted out using magnetic cell separation. The ratio of peripheral CD31+Treg cells to Treg cells in each group was analyzed by flow cytometry. Real-time PCR was performed to detect TRECs in CD4+CD25+Treg cells. The percentages of CD3+, CD4+ and CD8+T cell sub-sets were also measured. Results The ratios of CD31+Treg/Treg cells and the numbers of TRECs in pe-ripheral blood of the Moderate CHB and Severe CHB groups were significantly lower than those of the Mild CHB and HCs groups (P<0. 05), while no statistical difference was found between the mild CHB and HC groups (P>0. 05). No significant difference in the percentages of CD3+, CD4+ or CD8+ T cell subsets was observed between the four groups (P>0. 05). CD31+ Treg/Treg cell ratio had a positive correlation with the number of TRECs (r=0. 551, P=0. 014). Conclusions Both CD31+Treg/Treg cell ratio and the number of TRECs were reduced in the peripheral blood of patients with moderate or severe CHB. CD31+Treg/Treg cell ratio and the number of TRECs were positively correlated and could be used as new indices to evaluate recent thymus output.

Inhibition of Toxoplasma gondii excretory - secretory antigens on growth of murine Lewis lung carcinoma / 中国血吸虫病防治杂志

Objective To investigate the effect of Toxoplasma gondii excretory-secretory antigens (ESA) on CD4+ CD25+ Foxp3+ T (Treg) cells in mice carrying Lewis lung carcinoma, and examine the inhibitory effect of T. gondii ESA on tumor growth. Methods C57BL/6 mice were randomly assigned into the PBS group (n = 14) and the Lewis group (n = 34). Mice in the Lewis group were subcutaneously injected with 2 × 105 Lewis lung carcinoma cells in the right axilla, while animals in the PBS group were injected with the same volume of sterile PBS. On day 7 post-injection (D7), mice in the PBS group were further divided into the PBS2 group and the PBS2 + ESA group, of 7 mice in each group, and mice in the Lewis group were further divided into the Lewis2 group and the Lewis2 + ESA group, of 17 mice in each group. Then, mice in the PBS2 + ESA group and the Lewis2 + ESA group were intraperitoneally injected with 100 μL of ESA. The mouse spleen coefficient was calculated in each group 7 days post-injection with ESA, and the changes of Treg cell counts and the long-term tumor growth were measured in tumor-bearing mice. Results The spleen coefficient was significantly greater in the PBS2 + ESA group and the Lewis2 + ESA group than in the PBS2 (0.66% ± 0.09% vs. 0.30% ± 0.02%, P < 0.05) and Lewis2 groups (0.69% ± 0.07% vs. 0.33% ± 0.03%, P < 0.05) 7 days post-treatment with ESA, respectively, and the percentage of splenic Treg cells in splenocytes was significantly lower in the PBS2 + ESA group and the Lewis2 + ESA group than in the PBS2 (1.28% ± 0.14% vs. 2.06% ± 0.07%, P < 0.05) and Lewis2 groups (1.58% ± 0.14% vs. 2.44% ± 0.23%, P < 0.05), respectively. T. gondii ESA treatment caused a delay in tumor growth, and the tumor size was significantly smaller in the Lewis2 + ESA group than in the Lewis2 group (P < 0.05). Conclusion T. gondii ESA may reduce the proportion of splenic Treg cells in splenocytes and inhibit tumor growth in mice carrying Lewis lung carcinoma.

Inhibition of Toxoplasma gondii excretory - secretory antigens on growth of murine Lewis lung carcinoma / 中国血吸虫病防治杂志

Objective To investigate the effect of Toxoplasma gondii excretory-secretory antigens (ESA) on CD4+ CD25+ Foxp3+ T (Treg) cells in mice carrying Lewis lung carcinoma, and examine the inhibitory effect of T. gondii ESA on tumor growth. Methods C57BL/6 mice were randomly assigned into the PBS group (n = 14) and the Lewis group (n = 34). Mice in the Lewis group were subcutaneously injected with 2 × 105 Lewis lung carcinoma cells in the right axilla, while animals in the PBS group were injected with the same volume of sterile PBS. On day 7 post-injection (D7), mice in the PBS group were further divided into the PBS2 group and the PBS2 + ESA group, of 7 mice in each group, and mice in the Lewis group were further divided into the Lewis2 group and the Lewis2 + ESA group, of 17 mice in each group. Then, mice in the PBS2 + ESA group and the Lewis2 + ESA group were intraperitoneally injected with 100 μL of ESA. The mouse spleen coefficient was calculated in each group 7 days post-injection with ESA, and the changes of Treg cell counts and the long-term tumor growth were measured in tumor-bearing mice. Results The spleen coefficient was significantly greater in the PBS2 + ESA group and the Lewis2 + ESA group than in the PBS2 (0.66% ± 0.09% vs. 0.30% ± 0.02%, P < 0.05) and Lewis2 groups (0.69% ± 0.07% vs. 0.33% ± 0.03%, P < 0.05) 7 days post-treatment with ESA, respectively, and the percentage of splenic Treg cells in splenocytes was significantly lower in the PBS2 + ESA group and the Lewis2 + ESA group than in the PBS2 (1.28% ± 0.14% vs. 2.06% ± 0.07%, P < 0.05) and Lewis2 groups (1.58% ± 0.14% vs. 2.44% ± 0.23%, P < 0.05), respectively. T. gondii ESA treatment caused a delay in tumor growth, and the tumor size was significantly smaller in the Lewis2 + ESA group than in the Lewis2 group (P < 0.05). Conclusion T. gondii ESA may reduce the proportion of splenic Treg cells in splenocytes and inhibit tumor growth in mice carrying Lewis lung carcinoma.

Expression of transcription factor Bach2 in CD4+CD25+CD45RA- T cells in patients with systemic lupus erythematosus and its effect / 上海交通大学学报（医学版）

Objective: To study the expression of transcription factor BTB and CNC homology 2 (Bach2) in CD4+CD25+CD45RA- T cells from patients with systemic lupus erythematosus (SLE) and its effect on cell function. Methods: The CD4+CD25+CD45RA- T cells from active SLE patients and healthy volunteers were sorted by flow cytometry. The expression of Bach2 in CD4+CD25+CD45RA- T cells was detected by fluorescence quantitative PCR and Western blotting. The correlation between the median flourscence indensity (MFI) of Bach2 in CD4+CD25+CD45RA- T cells and the disease activity index of SLE (SLEDAI) was analyzed. The MFI of Bach2 in IL-17+CD4+CD25+CD45RA- T cells was compared with that in IL-17-CD4+CD25+CD45RA- T cells by flow cytometry. In Bach2 overexpression system, the expression of IL-17 in CD4+CD25+CD45RA- T cells was detected by flow cytometry and the concentration of IL-17 in the culture supernants was detected by ELISA. Results: The mRNA and protein expressions of Bach2 in CD4+CD25+CD45RA- T cells from SLE patients were significantly lower than those in healthy controls (P<0.01). There was a significant negative correlation between the MFI of Bach2 and SLEDAI (R2=0.433, P=0.001) in patients with SLE. The expression of Bach2 in IL-17+CD4+CD25+CD45RA- T cells was significantly lower than that in IL-17-CD4+CD25+CD45RA- T cells (P=0.013). When Bach2 was overexpressed, the percentage of CD4+CD25+CD45RA- T cells from SLE patients expressing inflammatory factor IL-17 decreased significantly (P=0.032) and the IL-17 concentration in cell culture supernatants markedly decreased (P=0.008). Conclusion: The expression of Bach2 in CD4+CD25+CD45RA- T cells from SLE patients decreases, and overexpression of Bach2 in the cells leads to the falling expression of IL-17.

Schistosoma japonicum heat shock protein 60 enhances regulatory T cell im-munosuppressive function by promoting the expressions of IL-10 and TGF-β / 中国血吸虫病防治杂志

Objective To explore the possible mechanisms by which Schistosoma japonicum heat shock protein 60(SjH-SP60)enhances CD4+CD25+regulatory T cell(Treg)immunosuppressive function.Methods An in vitro method was used to investigate the effect of SjHSP60 on Treg immunosuppressive activity.Co-cultures in transwells and in vitro suppression assay were performed to investigate how SjHSP60 enhanced the immunosuppressive function of Tregs.Intracellular cytokine staining combined with flow cytometry was used to detect Treg-expressing IL-10 and TGF-β,and flow cytometry was also used to analyze the expressions of Foxp3 and CTLA-4 in Tregs.Results SjHSP60 enhanced the immunosuppressive function of Tregs.Soluble cytokines IL-10 and TGF-β mediated inhibitory activity of SjHSP60-triggered Tregs.SjHSP60 induced significant increases in both IL-10 and TGF-β expressions of Tregs.Further investigation showed significant increased Foxp3 and CTLA-4 in SjHSP60-trggered Tregs.Conclusion SjHSP60 enhances Treg immunosuppressive function by promoting the expressions of IL-10 and TGF-β,possibly due to SjHSP60-mediated induction of Foxp3 and CTLA-4 in Tregs.

Progress on exosome and effect on regulatory T cells / 中国免疫学杂志

Regulatory T (Treg) cells is an indispensable subset of T lymphocyte with the ability of immunosuppression in the periphery.Thymus-derived Treg cells(CD4+CD25+FOXP3+Treg cells) play a fundamental role in maintaining immune homeostasis in vivo.Treg cells have been actively involved in the onset and development of major human diseases including malignant tumors, autoimmune diseases,infectious diseases,allergic diseases and graft versus host disease(GVHD).Exosomes are membranous vesicles of endosomal origin that released from multiple cells into the extracellular space.They are currently considered to be vehicles containing protein,RNA and MicroRNA which can been transferred to recipient cells to modulate their activity by cell-contact-independent mecha-nism.Exosomes have a great impact on the induction and proliferation of Treg cells,understanding the relation of them will lead to novel therapeutic approaches for cancer immunotherapy,treating autoimmunity,infection,allergy and organ transplantation.

Effects of high risk human papillomavirus loading on the expression of locally regulatory T cells in differ-ent cervical lesions / 实用医学杂志

Objective To compare the synchronous changes of high risk human papillomavirus load(HPV-DNA)and CD4+CD25+Foxp3+regulatory T cells(Treg)in local microenvironment of cervix,and investigate the ef-fects of HPV virus replication and progression of cervical lesions on Treg cells. Methods 304 cases of HR-HPV infection with cervical lesions were divided into 5 groups,cervical intraepithelial neoplasia(CIN)I,CINII,CINIII, cervical cancer and chronic cervicitis.The HPV-DNA of cervical secretion was detected by PCR fluorescence,and the relative Treg cells numbers from cervical brush samples were determined by flow cytometry with CD4+CD25+Foxp3+gating,and the data were statistically analyzed. Results(1)There was significant difference of cervical Treg cells in different degrees of cervical lesion and different copy numbers by variance comparison(F = 24.93, 109.86,P < 0.05),and a further pairwise comparison showed that there was no significant difference of Treg cell between chronic cervicitis and CINI and low load(HPV DNA 104~105copies/mL)and medium load(HPV DNA 105~106copies/mL)(P>0.05).There was a significant difference between the other groups(P<0.05);(2)Treg cells as variable,interaction effect of cervical lesions and viral load factors were significant different(F=3.39,P<0.05). The effect of different cervical lesions and HR-HPV viral load on the expression of Treg cells was differen-tial. An overall showing with the degree of cervical lesions increased,HR-HPV virus copy number increased, Treg cells expression increased gradually;(3)CD4+CD25+Foxp3+Treg cells were highly expressed in cervical cancer patients,but the expression level fluctuated widely and the numerical distribution was the most dispersedly. Conclusion The immune suppression function of local CD4+CD25+Foxp3+Treg cells with different cervical lesions and different HR-HPV DNA may bilaterally regulate the prognosis of cervical lesions,as a whole,between Treg cells and HR-HPV load and cervical lesions were the consistent progress trend.

Expression of transcription factor Bach2 in CD4+CD25+CD45RA-T cells in patients with systemic lupus erythematosus and its effect / 上海交通大学学报（医学版）

Objective·To study the expression of transcription factor BTB and CNC homology 2 (Bach2) in CD4+CD25+CD45RA-T ceils from patients with systemic lupus erythematosus (SLE) and its effect on cell function.Methods·The CD4+CD25+CD45RA T cells from active SLE patients and healthy volunteers were sorted by flow cytometry.The expression of Bach2 in CD4+CD25+CD45RA T cells was detected by fluorescence quantitative PCR and Western blotting.The correlation between the median flourscence indensity (MFI) of Bach2 in CD4+CD25+CD45RA-T cells and the disease activity index of SLE (SLEDAI) was analyzed.The MFI of Bach2 in IL-17+CD4+CD25+CD45RA-T ceils was compared with that in IL-17-CD4+CD25+CD45RA-T cells by flow cytometry.In Bach2 overexpression system,the expression of IL-17 in CD4+CD25+CD45RA T ceils was detected by flow cytometry and the concentration of IL-17 in the culture supernants was detected by ELISA.Results·The mRNA and protein expressions of Bach2 in CD4+CD25+CD45RA-T cells from SLE patients were significantly lower than those in healthy controls (P＜0.01).There was a significant negative correlation between the MFI of Bach2 and SLEDAI (R2=0.433,P=-0.001) in patients with SLE.The expression of Bach2 in IL-17+CD4+CD25+CD45RA-T cells was significantly lower than that in IL-17-CD4+CD25+CD45RA-T cells (P=-0.013).When Bach2 was overexpressed,the percentage of CD4+CD25+CD45RA-T cells from SLE patients expressing inflammatory factor IL-17 decreased significantly (P=0.032) and the IL-17 concentration in cell culture supernatants markedly decreased (P=0.008).Conclusion·The expression of Bach2 in CD4+CD25+CD45RA-T cells from SLE patients decreases,and overexpression of Bach2 in the cells leads to the falling expression of IL-17.

Unbalanced expression of aryl hydrocarbon receptor in peripheral blood CCR6+CD4+ and CD4+CD25+T cells of rheumatoid arthritis / Expressão não equilibrada do receptor de hidrocarboneto arílico nos linfócitos T CCR6+ CD4+ e CD4+ CD25+ do sangue periférico na artrite reumatoide

ABSTRACT Objective: The goal of this study was to analyze the role of aryl hydrocarbon receptor in peripheral blood CCR6+CD4+ and CD4+CD25+T cells of patients with rheumatoid arthritis. Methods: Flow cytometry was applied to determine the proportion of AhR positive cells in CCR6+CD4+T, CD4+CD25+T and peripheral blood peripheral mononuclear cells from each subject. AhR mRNA and CYP1A1 mRNA relative expression levels were tested by real-time PCR. Results: The percentage of AhR positive cells in peripheral blood mononuclear cells was higher in RA group than that in healthy cases [(35.23 ± 10.71)% vs. (18.83 ± 7.32)%, p < 0.01]. The expression levels of AhR and CYP1A1 were both increased in patients with RA while compared to controls [(3.71 ± 1.63) vs. (2.00 ± 1.27), p = 0.002; (2.62 ± 2.08) vs. (0.62 ± 0.29), p < 0.01, respectively]. In RA patients, the percentage of AhR positive cells in CD4+CD25+T cells was significantly lower than that from controls [17.90 (6.10 ± 80.10)% vs. (52.49 ± 19.18)%, p < 0.01]; In healthy controls, the percentage of AhR positive cells in CD4+CD25+T cells was significantly higher than that in CCR6+CD4+T cells, and was also significantly higher than that in PBMCs [(52.49 ± 19.18)% vs. (23.18 ± 5.62)% vs. (18.06 ± 7.80)%, X 2 = 24.03, p < 0.01]; in RA patients, the percentage of AhR positive cells in CCR6+CD4+T cells was significantly increased than that in CD4+CD25+T cells and PBMCs [(46.02 ± 14.68)% vs. 17.90 (6.10 ± 80.10)% vs. (34.22 ± 10.33)%, X 2 = 38.29, p < 0.01]; Nevertheless, no statistically significant relationship was found between clinical data and AhR positive cells in CCR6+CD4+T and CD4+CD25+T cells. Conclusion: AhR may participate in the pathological progress of RA by controlling the differentiation of Th17 and Treg cells in peripheral blood.

RESUMO Objetivo: Analisar o papel do receptor de hidrocarboneto arílico (AhR) nos linfócitos T CCR6+ CD4+ e CD4+ CD25+ no sangue periférico de pacientes com artrite reumatoide (AR). Métodos: Foi aplicada citometria de fluxo para determinar a proporção de células AhR positivas em linfócitos CCR6+ CD4+ e CD4+ CD25+ do sangue periférico e células mononucleares periféricas de cada indivíduo. Os níveis de expressão relativa de ácido ribonucleico mensageiro (do inglês ribonucleic acid, RNAm,) de AhR e RNAm de enzima de primeiro estágio essencial para o AhR (CYP1A1) foram testados por reação em cadeia de polimerase (do inglês polymerase chain reaction, PCR,) em tempo real. Resultados: A percentagem de células AhR positivas nas células mononucleares do sangue periférico foi maior no grupo com AR do que nos indivíduos saudáveis [(35,23 ± 10,71)% vs. (18,83 ± 7,32)%, (p < 0,01)]. Os níveis de expressão de AhR e CYP1A1 estavam aumentados em pacientes com AR quando comparados com os controles [(3,71 ± 1,63) vs. (2,00 ± 1,27), p = 0,002; (2,62 ± 2,08) vs. (0,62 ± 0,29), p < 0,01, respectivamente]. Em pacientes com AR, a percentagem de células AhR positivas nos linfócitos T CD4+ CD25+ foi significativamente inferior à dos controles [17,90 (6,10 ± 80,10)]% vs. (52,49 ± 19,18)%, p < 0,01]; em controles saudáveis, a percentagem de células AhR positivas nos linfócitos T CD4+ CD25+ foi significativamente mais elevada do que nos linfócitos T CCR6+ CD4+ e também foi significativamente maior do que nas células mononucleares do sangue periférico (do inglês peripheral blood mononuclear cells, PBMC,) [(52,49 ± 19,18)% vs. (23,18 ± 5,62)% vs. (18,06 ± 7,80)%, X 2 = 24,03, p < 0,01]; em pacientes com AR, a percentagem de células AHR positivas nos linfócitos T CCR6+ CD4+ era significativamente maior em comparação com os linfócitos T CD4+ CD25+ e PBMC (46,02 ± 14,68)% vs. [17,90 (6,10 ± 80.10)]% vs. (34,22 ± 10,33)%, X2 = 38,29, p < 0,01]; no entanto, não foi encontrada correlação estatisticamente significativa entre os dados clínicos e células AhR positivas em linfócitos T CCR6+ CD4+ e CD4+ CD25+. Conclusão: O Ahr pode participar do progresso patológico da AR ao controlar a diferenciação de linfócitos Th17 e Treg no sangue periférico.

Relationship between HBeAg from HBsAg positive mothers and regulatory T cells in neonates and its influence on HBV intrauterine transmission / 中华流行病学杂志

Objective To explore the relationship between HBeAg in HBsAg positive mothers and CD4 + CD25 + Foxp3 + regulatory T cells (Treg) in newborns,as well as how they would influence the increasing risk on HBV intrauterine transmission.Methods We collected information on general demographic characteristics and delivery on 270 HBsAg positive mothers and their newborns from the Third People's Hospital of Taiyuan.Fluorescence quantitative polymerase chain reaction (FQ-PCR) and chemiluminescence immunoassay (CLIA) were used to detect HBV DNA and HBV serological markers in peripheral blood from both mothers and neonates.The expression of Treg and other immune cells in peripheral blood of neonates were detected with flow cytometry (FCM).Results Maternal HBeAg positive rates were associated with an increased risk of intrauterine transmission (0R=4.08,95％ CI:1.89-8.82).Rates of T.reg in newborns born to HBsAg-positive mothers were higher than that of the negative group (Z=2.29,P=0.022).Each pair of the subjects was assigned to five different groups according to the HBeAg titers of mothers.Frequencies of both Treg and HBeAg in newboms and HBV DNA in mothers between the above said 5 groups showed similar trends of changing patterns and the differences between groups were statistically significant (x2=18.73,P＜0.001;x2=181.60,P＜0.001;x2=183.09,P＜0.001).Results from partial correlation analysis showed that after adjusting for neonatal HBeAg and maternal HBV DNA,mother's HBeAg titers were positively related to the percentage of Treg in their newboms (rs=0.19,P=0.039).In addition,the frequencies of Treg were negatively correlated with pDC and CD4 + T cell in their newborns (rs=-0.21,P=0.017;r,=-0.23,P=0.009).Conclusion HBeAg from HBsAg positive mothers might have inhibited the function of neonatal DC cells and T cells to reduce the immune response to HBV by up-regulating the proportion of Treg and finally increased the risk of HBV intrauterine transmission.

Relationship between HBeAg from HBsAg positive mothers and regulatory T cells in neonates and its influence on HBV intrauterine transmission / 中华流行病学杂志

Objective To explore the relationship between HBeAg in HBsAg positive mothers and CD4 + CD25 + Foxp3 + regulatory T cells (Treg) in newborns,as well as how they would influence the increasing risk on HBV intrauterine transmission.Methods We collected information on general demographic characteristics and delivery on 270 HBsAg positive mothers and their newborns from the Third People's Hospital of Taiyuan.Fluorescence quantitative polymerase chain reaction (FQ-PCR) and chemiluminescence immunoassay (CLIA) were used to detect HBV DNA and HBV serological markers in peripheral blood from both mothers and neonates.The expression of Treg and other immune cells in peripheral blood of neonates were detected with flow cytometry (FCM).Results Maternal HBeAg positive rates were associated with an increased risk of intrauterine transmission (0R=4.08,95％ CI:1.89-8.82).Rates of T.reg in newborns born to HBsAg-positive mothers were higher than that of the negative group (Z=2.29,P=0.022).Each pair of the subjects was assigned to five different groups according to the HBeAg titers of mothers.Frequencies of both Treg and HBeAg in newboms and HBV DNA in mothers between the above said 5 groups showed similar trends of changing patterns and the differences between groups were statistically significant (x2=18.73,P＜0.001;x2=181.60,P＜0.001;x2=183.09,P＜0.001).Results from partial correlation analysis showed that after adjusting for neonatal HBeAg and maternal HBV DNA,mother's HBeAg titers were positively related to the percentage of Treg in their newboms (rs=0.19,P=0.039).In addition,the frequencies of Treg were negatively correlated with pDC and CD4 + T cell in their newborns (rs=-0.21,P=0.017;r,=-0.23,P=0.009).Conclusion HBeAg from HBsAg positive mothers might have inhibited the function of neonatal DC cells and T cells to reduce the immune response to HBV by up-regulating the proportion of Treg and finally increased the risk of HBV intrauterine transmission.

Effects of vasoactive intestinal peptide on the ratio of CD4+CD25+Treg/CD4+T cell and the expression of TGF-β1 in EAE rats / 西安交通大学学报(医学版)

Objective To explore the effects of vasoactive intestinal peptide (VIP)on the ratio of CD4+CD25+Treg/CD4+T cell and the expression of TGF-β1 in experimental autoimmune encephalomyelitis (EAE)rats. Methods We randomly divided 60 healthy female Wistar rats into normal control group,EAE control group,VIP low-dose group and VIP high-dose group.We used myelin basic protein (MBP)+ complete adjuvant (CFA)to establish EAE model. Since the day of model construction, the low- and high-dose VIP groups received intraperitoneal injection of 4 nmol/kg (0.2 mL)and 16 nmol/kg (0.8 mL)of VIP every other day,respectively;normal control group and EAE group received injection of saline of 0.8 mL for 10 days in a row.We recorded the peak of neurological dysfunction score (NDS)changes in the rats,observed the pathological changes and GFAP+astrocyte activation in the brain at the morbidity peak of rats with HE staining,and detected the ratio of CD4+CD25+Treg/CD4+T in the spleen with FACS and TGF-β1 cytokine level in brain tissue with ELISA.Results The peak nerve dysfunction score was decreased in each VIP dose group.In normal control group,there were decreased inflammatory cell infiltration and decreased number of active astrocytes in the brain tissue.The degree of infiltration of inflammatory cells and astrocyte activation in VIP control groups were significantly lower than those in EAE group.The CD4+CD25+Treg/CD4+T cell ratio of the spleen tissue in each dose VIP treated group rats was higher than that in EAE control group.The cytokine level of TGF-β1 in the brain tissue increased in each VIP dose group in the dose-dependent manner.Conclusion Through up-regulating the ratio of CD4+CD25+Treg/CD4+T cell in the spleen tissue,increasing TGF-β1 content in brain tissue,and inhibiting the infiltration of inflammatory cells and the astrocyte activation,VIP plays an important role in prevention and control of EAE.

Spectrum drift characteristics of CD4+ CD25+ Tregs TCR CDR3 in hepatitis B patients before and after treatment / 中国免疫学杂志

Objective:To determine the spectrum drift characteristics of CI4+CD25+Tregs TCR β chain CDR3 in patients with different phases of acute hepatitis B (AHB) and chronic hepatitis B (CHB) patients before and after the entecavir treatment.Methods:Anticoagulation venous blood was collected from 4 normal control subjects,3 AHB patients with acute phase and convalescent phase,and 4 CHB patients before and after the entecavir treatment;and peripheral blood mononuclear cells were isolated;CD4+ CD25+ Tregs were separated by using the magnetic beads,and total RNAs were extracted from CD4+ CD25+ Tregs and used for reverse transcription.The TRBV CDR3 was amplified by polymerase chain reaction (PCR) with forward primers specific for 24 TRBV families and one fluorescence-labeled common reverse primer specific for the BC region.The PCR products were sent out for Genescan,and results were analyzed for the TRBV family CDR3 spectrum characteristics by using the Peak Scanner Software vl.0.Data were analyzed with the comparative t-test to perform the statistical analysis.Results:The CDR3 spectral types of the TRBV family showed drift characteristics in 3 cases of AHB patients with acute and convalescent phases;single/oligo peak spectral type family was observed in most of patients with acute phase;multiple peak spectral type was seen in patients with convalescent phase;and the common spectrum shift of TRBV4,10,14,16,19 families seen in patients with acute phase was changed to multiple peak spectral type.The clonal expansion of TRBV family in the CD4+CD25+Tregs in PBMC from AHB patients with convalescent phase was significantly lower than AHB patients with acute phase (t =9.456,P =0.011).The clonal expansion of Tregs TRBV13.2,15,16,18,20 family seen in C HB patients before treatment may interfere the virus removal through down-regulating the body's immune response;and with the decline of viral load in serum after the antiviral treatment,the clonal expansion of Tregs TRBV1,5.2,6,12,14,24 family may help body induce immune tolerance and result in the HBV persistence.The clonal expansion of TRBV family in the CI4+CD25+Tregs in PBMC from of CHB patients after antiviral treatment was increased (t =-0.666,P =0.553).Conclusion:TRBV4,10,14,16,19 family of spectrum shift seen in AHB patients with acute phase was changed to multiple peak spectral type in patients with convalescent phase,suggesting this transition may be associated with HBsAg and HBeAg turning to negative.The clonal expansion of Tregs TRBV13.2,15,16,18,20 family seen in CHB patients before treatment may interfere the virus removal through down-regulating the body's immune response;and with the decline of viral load in serum after the antiviral treatment,the clonal expansion of Tregs TRBV1,5.2,6,12,14,24 family may help body induce immune tolerance and result in the HBV persistence.

<b>A Japanese Herbal Medicine, Shigyakusan, Induces Prolonged Survival of Fully MHC-Mismatched Murine Cardiac A</b><b>llografts</b> / 日本東洋医学雑誌

<p><b>Background </b>: Shigyakusan, a 4-component Japanese herbal medicine (Paeoniae radix, Aurantii fructus immaturus, Glycyrrhizae radix and Bupleuri radix), is used not only for cholecystitis and gastritis as an antiinflammatory agent, but also for anxiety neurosis and insomnia as an anti-anxiety agent.<br><b>Methods </b>: We investigated the effects of shigyakusan on alloimmune responses in fully MHC-mismatched murine cardiac allograft transplantation. CBA mice underwent transplantation of a C57BL/6 heart and received shigyakusan or one component of shigyakusan administered orally from the day of transplantation until 7 days afterward. Histologic studies, cytokine measurements, and flow cytometry assessments were performed.<br><b>Results </b>: Untreated CBA recipients acutely rejected C57BL/6 cardiac grafts (median survival times [MST], 7 days). On the other hand, CBA transplant recipients given shigyakusan had significantly prolonged C57BL/6 allograft survival (MST, 22.5 days). MSTs for C57BL/6 transplant recipients given Paeoniae radix, Aurantii fructus immaturus, Glycyrrhizae radix and Bupleuri radix were 11, 9.5, 18.5 and 8 days, respectively. Additionally, flow cytometry studies showed that the percentage of CD25+Foxp3+ cell populations in CD4+ cells was increased in transplant recipients given shigyakusan.<br><b>Conclusion </b>: Shigyakusan induced hyporesponsiveness to fully MHC-mismatched allogeneic cardiac allografts and may generate CD4+CD25+Foxp3+ cells in our model.</p>

Immunoregulation study of UCMSCs on UCB CD4+T lymphocytes in vitro / 中国免疫学杂志

Objective:Immunoregulation study of umbilical mesenchymal stem cell (UCMSCs) on allogeneic umbilical cord blood(UCB) CD4+T lymphocytes,which proliferation,apoptosis and the differentiation to CD4+CD25+ regulatory T cell (Treg) in vitro. Methods:Establishing on direct contact or transwell co-culture system,adopt in different proportion of UCMCs with phytohaemag-glutinin (PHA)-activated UCB CD4+T lymphocytes were co-cultured. The proliferation of lymphocyte,percent of CD4+CD25+/CD4+and Foxp3 expression, regulatory T cell marker gene were measured. Apoptosis of CD4+T lymphocytes were observed in the direct contact or transwell coculture system of UCMSCs with desamethason( DXM)-stimulated UCB CD4+T lymphocytes. Results: The UCB CD4+T lymphocytes cocultured with UCMSCs with PHA-activating for 3 days,compared with the UCMSCs free control group,the amount of cells was reduced noticeably(P<0. 05) and the percent of CD4+CD25+in CD4+T lymphocytes and Foxp3 expression significantly in-creased(P<0. 01) in a dose dependent way(P<0. 05). The UCB CD4+T lymphocytes cocultured with UCMSCs with DXM-inducing for 7 days,the apoptosis rate was significantly lower than that of the control group without UCMSCs (P<0. 01). These effects were partially attenuated in transwell coculture but could not be eliminated. Conclusion: UCMSCs are negative effect on UCB CD4+T lymphocytes-mediated immunity effects,and mainly manifested in the regulation on cell proliferate ability and differentiation rather than promoting apoptosis.

Experimental study on the expression of T cells and cytokines in the regulation of autoimmune dry eye / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To investigate the role of regulatory T cells in the pathogenesis of autoimmune dry eye and to analyze the expression of endogenous regulatory T cells and related cytokines in the lacrimal system to provide a more effective treatment for autoimmune dry eye disease treatment programs. <p>METHODS: Rabbit lacrimal gland epithelial cells isolated and cultured for a period of time were separately cultured and mixed with isolated peripheral blood lymphocytes in a ratio of 1: 1 and cultured with 5-Bromo-2-deoxyUridine(BrdU)to detect the proliferation of peripheral blood lymphocytes, and the activated autologous peripheral blood lymphocytes were transferred to donor rabbits via ear vein. The rabbits were used as the rabbit model of autoimmune dry eye and the normal rabbits which did not induce the disease as the control group. Before and after the experiment, tear film rupture time and tear secretion at 2,4,6,8wk,were observed. At 4 wk after the transfusion the rabbits were sacrificed for the collection of bilateral upper and lower lacrimal gland and conjunctiva, and pathological HE staining, and then extract the total RNA in lacrimal gland. The expression of IL-17, TNF-α, IL-6 and TGF-γ was detected. <p>RESULTS: BrdU detected peripheral lymphocyte proliferation rate of 3.72. Compared with the normal group, the time of rupture of the tear film decreased(<i>P</i><0.05), and the tear of the model group was significantly lower than that of the control group(<i>P</i><0.05). The expression of IL-17, TNF-α, TGF-γ and IL-6 in the model group were significantly higher than those in the control group(<i>P</i><0.05). The expression of IL-10 and TGF-β was significantly lower than those of the control group(<i>P</i><0.05). The ratio of CD4⁺ CD2⁺ Treg cells in lacrimal gland tissue and spleen was higher in control group. The proportion of CD4⁺CD25⁺ Treg cells in model group decreased(<i>P</i><0.05). <p>CONCLUSION: IL-17, TNF-α, IL-6 and TGF-γ play important roles in inflammatory reaction of lacrimal gland and the immunodepression active by CD4⁺CD25⁺Treg cells decreases when the rabbit model of dry eye is involved.

Role and clinical significance of imbalanced Th17/CD4+CD25+ regulatory T lymphocytes in children with Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis / 中华微生物学和免疫学杂志

Objective To investigate the changes in Th17 cells and CD4+CD25+regulatory T lym-phocytes ( Treg) as well as transcription factors and cytokines relating to them in children with Epstein-Barr virus (EBV)-associated hemophagocytic lymphohistiocytosis (HLH) and to analyze their role and clinical significance. Methods Thirty-two children with newly diagnosed EBV-associated HLH in the Hematology/Oncology Department of Zhengzhou Children′s Hospital from January 2012 to December 2016 were enrolled in this study. Thirty healthy children taking physical examination in the same hospital in the corresponding period were recruited as controls. Percentages of Th17 and Treg cells in peripheral blood T lymphocytes were detected by flow cytometry. Expression of RORγt and Foxp3 at mRNA level in peripheral blood mononuclear cells was detected by real-time PCR. Levels of IL-6, IL-17, IL-10 and TGF-β1 in serum samples were measured by ELISA. Results Compared with the control group, the EBV-associated HLH group showed in-creased percentage of Th17 cells [(1. 09±0. 43)％ vs (0. 39±0. 19)％, P<0. 05] and enhanced expres-sion of RORγt at mRNA level [(1. 41±0. 37) vs (0. 67±0. 13), P<0. 05], but decreased percentage of Treg cells [(3. 66±1. 13)％ vs (6. 80±1. 15)％, P<0. 05] and inhibited expression of Foxp3 at mRNA level [(15. 97±5. 11) vs (30. 23±4. 95), P<0. 05]. All of the above mentioned changes were reversed af-ter treatment (P<0. 05). Serum levels of IL-6 and IL-17 of EBV-associated HLH group were higher than those of control group, while serum levels of IL-10 and TGF-β1 were lower (P<0. 05). Conclusion Im-balanced Th17/Treg cells might play an important role in the pathogenesis of EBV-associated HLH. Cyto-kines relating to the maintenance of Th17/Treg cell balance could be used as indicators of disease develop-ment.

Role and clinical significance of imbalanced Th17/CD4+CD25+ regulatory T lymphocytes in children with Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis / 中华微生物学和免疫学杂志

Objective To investigate the changes in Th17 cells and CD4+CD25+regulatory T lym-phocytes ( Treg) as well as transcription factors and cytokines relating to them in children with Epstein-Barr virus (EBV)-associated hemophagocytic lymphohistiocytosis (HLH) and to analyze their role and clinical significance. Methods Thirty-two children with newly diagnosed EBV-associated HLH in the Hematology/Oncology Department of Zhengzhou Children′s Hospital from January 2012 to December 2016 were enrolled in this study. Thirty healthy children taking physical examination in the same hospital in the corresponding period were recruited as controls. Percentages of Th17 and Treg cells in peripheral blood T lymphocytes were detected by flow cytometry. Expression of RORγt and Foxp3 at mRNA level in peripheral blood mononuclear cells was detected by real-time PCR. Levels of IL-6, IL-17, IL-10 and TGF-β1 in serum samples were measured by ELISA. Results Compared with the control group, the EBV-associated HLH group showed in-creased percentage of Th17 cells [(1. 09±0. 43)％ vs (0. 39±0. 19)％, P<0. 05] and enhanced expres-sion of RORγt at mRNA level [(1. 41±0. 37) vs (0. 67±0. 13), P<0. 05], but decreased percentage of Treg cells [(3. 66±1. 13)％ vs (6. 80±1. 15)％, P<0. 05] and inhibited expression of Foxp3 at mRNA level [(15. 97±5. 11) vs (30. 23±4. 95), P<0. 05]. All of the above mentioned changes were reversed af-ter treatment (P<0. 05). Serum levels of IL-6 and IL-17 of EBV-associated HLH group were higher than those of control group, while serum levels of IL-10 and TGF-β1 were lower (P<0. 05). Conclusion Im-balanced Th17/Treg cells might play an important role in the pathogenesis of EBV-associated HLH. Cyto-kines relating to the maintenance of Th17/Treg cell balance could be used as indicators of disease develop-ment.