Progress in discovery of RORγt inverse agonists for the treatment of autoimmune diseases / 中国药科大学学报

@#Abstract: As a Th17 cell-specific transcription factor, retinoic acid receptor-related orphan receptor γt (RORγt), can induce differentiation of Th17 cells and production of inflammatory factor IL-17, playing an important role in inflammation and autoimmune diseases. RORγt inverse agonists have become a research hotspot in both academia and pharmaceutical companies around the world in recent years, with great development potential. A variety of skeletal structure types have been reported, including orthosteric and allosteric inverse agonists. In this paper, the structure and functions of RORγt are introduced, and RORγt inverse agonists in clinical and preclinical studies are reviewed in order to provide reference for further research and development of RORγt inverse agonists.

Effects of sublingual immunotherapy on Treg/Th17 cell balance and related cytokines in preschool children aged 3-6 years with allergic rhinitis / 中国耳鼻咽喉头颈外科

OBJECTIVE To evaluate the impact of sublingual immunotherapy(SLIT)on the balance of Treg/Th17 cells and related cytokines in preschool children aged 3-6 years with allergic rhinitis(AR).METHODS Seventy preschool children aged 3-6 years with AR were divided into the SLIT group and the medication group,and their clinical data were retrospectively analyzed.The medication group received symptomatic treatment alone,while the SLIT group received a combined treatment of SLIT and symptomatic medication,with a 3-year follow-up period.The Treg/Th17 cell balance,serum levels of TGF-β,IL-10,IL-17,IL-21,as well as the total nasal symptom score(TNSS),total medication score(TMS),and visual analog scale(VAS)scores were compared before and after treatment between the two groups.RESULTS After 3 years of treatment,both groups showed significant improvements(P<0.05)in the percentages of CD4+CD25+Foxp3+Treg and CD4+IL-17+Th17 cells among CD4+T cells,percentages of Treg and Th17 cells,serum levels of TGF-β,IL-10,IL-17,IL-21,TNSS,TMS,and VAS scores.Moreover,the SLIT group exhibited significantly better outcomes compared to the medication group(P<0.05).CONCLUSION SLIT can modulate the balance of Treg/Th17 cells and improve serum levels of TGF-β,IL-10,IL-17,and IL-21.

Correlation analysis of serum Chemerin with disease activity and Th17/Treg in patients with rheumatoid arthritis / 天津医药

Objective To explore the correlation of serum Chemerin level with disease activity and the ratio of T helper 17/regulatory T cells(Th17/Treg)in patients with rheumatoid arthritis(RA).Methods A total of 180 patients with RA who were admitted to our hospital were regarded as the observation group.According to the DAS28 score,the observation group was divided into the high activity group(60 cases),the moderate activity group(60 cases)and the low activity group(60 cases).Another 180 healthy people who underwent physical examination in our hospital during the same period were regarded as the control group.Enzyme-linked immunosorbent assay(ELISA)was used to detect serum levels of Chemerin,interleukin-9(IL-9),interleukin-10(IL-10)and interleukin-17(IL-17).Flow cytometry was used to detect the Th17/Treg ratio.Spearman correlation analysis was applied to analyze the correlation between serum Chemerin level and DAS28 score.Pearson correlation analysis was used to analyze the correlation between serum Chemerin level and Th17,Treg cell percentage and Th17/Treg ratio.Results The results of this study showed that the serum level of Chemerin was higher in the observation group than that in the control group(P＜0.05).The serum Chemerin level was positively correlated with DAS28 score(P＜0.05).Serum Chemerin levels and DAS28 scores decreased in turn in the high,moderate and low activity groups(P＜0.05).The percentage of Th17 cells and the ratio of Th17/Treg were higher in the observation group than those in the control group,and the percentage of Treg cells was lower in the observation group than that in the control group(P＜0.05).The level of IL-10 was lower in the observation group than that in the control group,while levels of IL-17 and IL-9 were higher in the observation group than those in the control group(P＜0.05).The results of Pearson correlation analysis showed that serum Chemerin level was positively correlated with the percentage of Th17 cells and the ratio of Th17/Treg,and negatively correlated with the percentage of Treg cells(P＜0.05).Conclusion Serum Chemerin level is elevated in patients with RA,which is closely related to disease activity and Th17/Treg ratio.

Investigation on the incidence of IRIS in HIV patients with TB infection and its relationship with Th17/Treg, CD4 + T lymphocyte levels / 中国医师杂志

Objective:To investigate the incidence of immune reconstitution inflammatory syndrome (IRIS) in patients with HIV (HIV) and tuberculosis (TB) infection, and analyze the relationship between Th17/Treg cytokines, CD4 + T lymphocytes and IRIS. Methods:HIV patients with TB infection admitted to Public Health Clinical Center of Chengdu from June 2020 to June 2022 were divided into IRIS group (31 cases) and non IRIS group (93 cases) according to whether IRIS occurred after highly active antiretroviral therapy (HAART). The Demography data, clinical data and laboratory indicators of the two groups were compared. Multivariate logistic regression analysis was conducted to investigate the influencing factors of IRIS in HIV patients with TB infection.Results:There was no significant difference in Demography data between the two groups ( P>0.05). There was a statistically significant difference in the history of opportunistic infection between the IRIS group and the non IRIS group (χ 2=5.194, P<0.05). The levels of HIV RNA, interleukin (IL)-17, and IL-23 in the IRIS group were higher than those in the non IRIS group (all P<0.05). The levels of the γ interferon (IFN- γ), the transforming growth factor-β (TGF- β) and baseline CD4 + T lymphocyte count were lower than those in the non IRIS group (all P<0.05). The results of multivariate logistic regression analysis showed that IL-17 ( OR: 1.266, 95% CI: 1.095-1.464), IL-23( OR: 1.384, 95% CI: 1.120-1.710), and TGF- β( OR: 0.589, 95% CI: 0.436-0.797) were influencing factors for the occurrence of IRIS in HIV patients with TB infection (all P<0.05). Conclusions:For patients with high IL-17 levels, high IL-23 levels, and low TGF- β level of HIV complicated with TB infection, clinical prevention and control should be carried out as soon as possible to prevent the occurrence of IRIS.

The proportion of Th17 cells in patients with systemic sclerosis: a Meta-analysis / 中华风湿病学杂志

Objective:To clarify peripheral Th17 level in SSc patients and its correlation with disease.Methods:Chinese databases CNKI, CBM, Wanfang and VIP, and English databases PubMed, EMBASE, Web of Science, Cochrane Library and Science Direct were searched to collect a case-control study on the content of Th17 cells in peripheral blood of patients with SSc. The papers published when the database was first developed in 25 February 2021. Meta-analysis was conducted using Stata 12.0 software, and I2 and Egger tests were used to evaluate the heterogeneity and publication bias between studies. Results:A total of 26 case-controls were included in the study, including 1 160 patients with SSc and 778 healthy controls. Overall, the percentage of Th17 cells in SSc patients was higher than in healthy controls [SMD(95% CI)=1.85 (1.33, 2.38), P<0.001], which was most significant in IL-17 +Th17 concentration [SMD(95% CI)=1.88 (1.28, 2.48), P<0.001]. As for disease activity, the proportion of Th17 cells in active SSc patients was much higher than those of patients in remission [SMD(95% CI)=1.92 (1.12, 2.71), P<0.001]. SSc patients had a reduced Th17 level after receiving DMARDs treatment [SMD(95% CI)=-0.74 (-1.05, -0.42), P=0.029]. Conclusion:The number of Th17 cells increase significantly in the peripheral blood of patients with SSc, and is related to disease activity. DMARDs can be used to treat this disease by downregulating Th17 levels.

Correlation of neutrophil extracellular traps with T cells differentiation in Hashimoto′s thyroiditis mice and vitamin D′s relief effect / 中华内分泌代谢杂志

Objective:To investigate correlation between neutrophil extracellular traps(NETs) formation and T cell subsets in mice with experimental autoimmune thyroiditis(EAT) and the impact of active vitamin D intervention.Methods:Six-week-old female BALB/c mice were randomly divided into Control group, EAT group and 1, 25 dihydroxy vitamin D 3[1, 25(OH) 2D 3] treatment group(VitD group; n=6/group). HE staining was used to observe thyroid pathology. Plasma thyroglobulin antibody(TGAb), thyroid peroxidase antibody(TPOAb), and 1, 25(OH) 2D 3 were measured by ELISA. Peripheral NETs formation, Th1, Th2, and Th17 cell ratio from spleen were measured by flow cytometry. Correlation between NETs formation rate and Th1, Th2, and Th17 cell ratio was analyzed. Results:Compared with Control group, mice in EAT group had significantly increased thyroid inflammation scores, thyroiditis morbidity, TPOAb, TGAb levels, NETs formation rate, Th2(CD4 + IL-4 + or CD4 + IL-13 + )and Th17 cell proportions( P were <0.001, 0.002, 0.007, <0.001, <0.001, 0.003, 0.001, and 0.002, respectively), and significant decreased 1, 25(OH) 2D 3, Th1 cell proportions, Th1/Th2(CD4 + IL-4 + ), Th1/Th2(CD4 + IL-13 + ), and Th1/Th17 ratios( P were 0.010, 0.018, 0.010, 0.005, and 0.007, respectively). Compared with the EAT group, the VitD group had lower thyroid inflammation scores, TPOAb, TGAb levels, NETs formation rate, Th2(CD4 + IL-4 + or CD4 + IL-13 + ) and Th17 cell proportions( P were 0.044, 0.007, <0.001, 0.001, 0.014, 0.008, and 0.001, respectively), and significant higher Th1 cell ratio, Th1/Th2(CD4 + IL-13 + ) and Th1/Th17 ratio( P were 0.011, 0.009, and 0.003, respectively). The Th1/Th2(CD4 + IL-4 + ) was not significantly increased in VitD group compared with EAT group( P=0.174). NETs formation rate was positively correlated with Th2(CD4 + IL-4 + or CD4 + IL-13 + ) and Th17 cell proportion( r were 0.65, 0.59, and 0.61; and P were 0.004, 0.010, and 0.007, respectively), but not with Th1 cell proportion( r=-0.47, P=0.051). Conclusion:EAT mice were more prone to NETs formation. Active vitamin D may relieve immune imbalance with increased Th2 and Th17 cell ratio and decreased Th1 cell ratio by reducing the formation of NETs in EAT mice. Vitamin D played the protective role in thyroid by reducing thyroid pathological damage and thyroid autoantibody levels, and relived overall lymphocyte imbalance.

Expression of interleukin-36 and its modulation on the balance between regulatory T cells and Th17 cells in patients with myasthenia gravis / 中华神经科杂志

Objective:To investigate interleukin (IL)-36 expression in patients with myasthenia gravis (MG), and to study the modulatory function of IL-36 on regulatory T cells (Tregs) and Th17 cells in MG patients.Methods:Fifty-one MG patients (MG group) and 25 healthy controls (control group) were enrolled in this study in Xinxiang Central Hospital between July 2016 and August 2021. Peripheral blood was collected. Plasma and peripheral blood mononuclear cells (PBMCs) were isolated. Plasma IL-36α, IL-36β, IL-36γ, IL-36RA, IL-35, and IL-17 levels were measured by enzyme-linked immunosorbent assay. The percentages of Tregs and Th17 cells were measured by flow cytometry. Forkhead box protein P3 (FoxP3) and retinoid-related orphan receptor gamma t (RORγt) mRNA expressions were measured by real-time polymerase chain reaction. PBMCs or purified Tregs from MG patients were stimulated with recombinant IL-36β (5 ng/ml). Changes of Tregs and Th17 cell percentages, IL-35 and IL-17 secretions, FoxP3 and RORγt mRNA expressions, as well as immunosuppressive activity of Tregs were analyzed.Results:There were no statistically significant differences of IL-36α, IL-36γ, or IL-36RA between the control group and the MG group (all P>0.05). IL-36β level was notably higher in the MG group compared with the control group [(73.43±13.91) pg/ml vs (60.91±12.65) pg/ml, t=3.79, P<0.001]. Treg percentage [(4.67±1.33)% vs (6.32±1.81)%, t=4.48, P<0.001], IL-35 [(50.06±7.93) pg/ml vs (65.37±8.90) pg/ml, t=7.59, P<0.001] and FoxP3 mRNA expression (1.03±0.14 vs 1.57±0.46, t=7.78, P<0.001) was lower, while Th17 cell percentage [(1.05±0.15)% vs (0.94±0.21)%, t=2.61, P=0.011], IL-17 [(40.61±13.13) pg/ml vs (33.09±11.48) pg/ml, t=2.44, P=0.017] and RORγt mRNA expression (1.26±0.16 vs 1.03±0.13, t=6.08, P<0.001) was higher in the MG group ( P<0.05). There were no statistically significant differences of above indices between different genders, onset ages, afflicting with thymoma, or different Osserman types (all P>0.05). There were no statistically significant correlations between above indices and quantitative myasthenia gravis (QMG) score (all P>0.05). Recombinant IL-36β stimulation did not affect PBMCs proliferation in MG patients ( P=0.248), and reduced Tregs percentage [(3.05±0.66)% vs (4.18±1.07)%, t=4.23, P<0.001], IL-35 secretion [(48.12±10.93) pg/ml vs (56.96±13.73) pg/ml, t=2.36, P=0.023] and FoxP3 mRNA expression (0.99±0.17 vs 1.18±0.13, t=4.01, P<0.001), but did not affect Th17 cell percentage, IL-17 secretion or RORγt mRNA expression (all P>0.05). Recombinant IL-36β stimulation inhibited immunosuppressive activity of Tregs, which presented as enhanced cellular proliferation [(0.83±0.12)×10 5vs (0.69±0.15)×10 5, t=3.02, P=0.005] and reduced IL-35 secretion [(28.71±10.08) pg/ml vs (37.12±10.47) pg/ml, t=2.39, P=0.023]. Conclusion:Increased IL-36β contributed to the regulation of Tregs/Th17 cell balance probably through inhibition of Tregs function in MG patients.

Interleukin-38 suppresses the trans-differentiation of regulatory T cells to Th17 cells in patients with primary biliary cholangitis / 中华风湿病学杂志

Objective:To investigate the IL-38 level in patients with primary biliary cholangitis (PBC), and to assess the modulatory function of IL-38 on the phenotype shift from regulatory T cells (Tregs) to Th17 cells in PBC.Methods:Forty-six PBC patients and 24 controls were included. Serum IL-38, IL-35 and IL-17 levels were measured by enzyme-linked immunosorbent assay. CD4 +CD25 +CD127 dim/-Tregs and CD4 +IL-17A +Th17 cell proportions in peripheral blood mononuclear cells (PBMC) were assessed by flow cytometry. Forkhead box P3 (FoxP3) and retinoid acid receptor related orphan receptor γt (RORγt) mRNA expressions were semi-quantified by real-time PCR. Purified Tregs were stimulated with recombinant IL-38, and were cocultured with autologous PBMC. Tregs function was assessed by measuring cellular proliferation and cytokines expression in the supernatants. Tregs were also polarized for Th17 culture, and recombinant IL-38 was added for stimulation. The influence of IL-38 on trans-differentiation of Tregs into Th17 phenotype was investigated by measuring CCR4/CCR6 expression, IL-17 secretion and RORγt mRNA expression. Student′s t test or Mann-Whitney U test were used for comparison between groups. Results:Serum IL-38 level was significantly lower in PBC group compared with control group [68.02(48.51, 96.74) pg/ml vs 89.6(58.0, 265.5)pg/ml, Z=3.25, P=0.037]. Proportion of Tregs [(6.9±1.3)% vs (11.3±3.5)%, t=7.64, P<0.001], FoxP3 mRNA expression [(1.0±0.3) vs (1.9± 0.7), t=7.74, P<0.001] and serum IL-35 level [25.87(16.08, 37.26)pg/ml vs 30.99(24.81, 52.29)pg/ml, Z=2.02, P=0.047] were notably lower in PBC group, while Th17 cell proportion [(5.5±1.4)% vs (3.8±1.0)%, t=5.43, P<0.001)], RORγt mRNA expression [(1.4±0.6) vs (1.0±0.4), t=2.72, P=0.008] and serum IL-17 level [202.0(121.6, 311.6)pg/ml vs 104.5(79.8, 155.5)pg/ml, Z=4.43, P<0.001] were remarkably higher in PBC group. Purified Tregs were co-cultured with autologous PBMC. Cellular proliferation was increased in PBC group compared with control group [(6.5±1.40)×10 5vs (5.3±1.1)×10 5, t=2.49, P=0.020]. IL-35 and IL-10 secretions in the supernatants were lower in PBC group ( P<0.05), but interferon- γ was higher in PBC group ( P=0.011). Tregs were also polarized for Th17 culture. CCR4 and CCR6 mean fluorescence intensity (MFI), RORγt mRNA and IL-17 secretion were higher in PBC group compared with control group ( P<0.05). IL-38 stimulation enhanced the inhibitory activity of Tregs from both PBC group and control group, which presented as reduced cellular proliferation as well as enhanced IL-35 and IL-10 secretion ( P<0.05). IL-38 stimulation only suppressed transdifferentiation of Tregs into Th17 phenotype in PBC group, which presented as down-regulation of CCR4 MFI, CCR6 MFI and IL-17 secretion ( P<0.05). However, IL-38 did not affect Tregs trans-differentiation into Th17 phenotype in control group. Conclusion:IL-38 played an immune-protective role and suppressed inflammatory response in the disease progression of PBC. IL-38 inhibited transdifferentiation of Tregs into Th17 phenotype in PBC patients.

Effects of Th17 and Treg cells with their balance on thyroid-associated ophthalmopathy / 国际眼科杂志(Guoji Yanke Zazhi)

Thyroid-associated ophthalmopathy(TAO)is an organ-specific autoimmune disease, which will cause a series of symptoms to significantly reduce the health level and life quality of patients. The pathogenesis of TAO has not been fully clarified. At present, there is a lack of unified and mature treatment scheme of it. Indeed, T-helper 17 lymphocyte(Th17)cells, regulatory T(Treg)cells and their imbalance are closely related to the immunological pathogenesis of TAO. It is currently believed that the cytokines secreted by Th17 cells can not only promote the inflammatory response of TAO and the fibrosis of orbital connective tissue, but also inhibit the adipogenic differentiation of TAO orbital connective tissue. In addition, Treg cells mainly exert immunosuppressive effect on TAO and delay the disease progression. At the same time, there is a dynamic balance relationship between Th17 and Treg cells, the imbalance of Th17/Treg cells can trigger the occurrence and development of TAO. This paper mainly expounds the influence mechanism of Th17, Treg cells and their balance on TAO, and analyzes the reasons for the differences between different research results, so as to provide some reference for the study of the pathogenesis and clinical treatment of TAO.

Mechanism research of the modified Xiaoyao Powder on inflammatory response of rats with syndrome of stagnation of liver qi and spleen deficiency of experimental autoimmune thyroiditis based on mirRNA326 regulating Th17 cells / 国际中医中药杂志

Objective:To explore the mechanism of modified Xiaoyao Powder on inflammatory response of rats with syndrome of stagnation of liver qi and spleen deficiency of experimental autoimmune thyroiditis (EAT) from the perspective of differentiation of microrna 326 (miR326) regulating Th17 cell.Methods:48 rats were randomly divided into normal group (12 rats) and model group (36 rats) respectively and they were immunized twice a week with high iodine water combined with subcutaneous injection of thyroglobulin. From the fifth to eighth weeks, 36 rats were immunized once a week. From the fifth week, the model group with liver depression and spleen deficiency syndrome of Traditional Chinese Medicine was reproduced with chronic restraint stress, excessive fatigue and eating incoherence methods. The modelrats were randomly divided into model group, Xiaoyao Powder group and Jinshuibao group. Rats in Xiaoyao Powder group were gavaged with 13.63 g/(kg·d) Xiaoyao Powder modified granule suspension, and rats in Jinshuibao group were gavaged with 477 mg/(kg·d) Jinshuibao suspension, twice a day, for 8 weeks.The levels of serum FT3, FT4, TSH, TGAb and TPOAb were detected by ELISA; the expression of miR326, IL-17 mRNA, IL-4 mRNA and IFN-γ mRNA were detected by PCR. The expression of Ets-1 protein in thyroid tissue was detected by Wes method, and the proportion of CD4 + IFNγ + T cells, CD4 + IL-4 + T cells and CD4 + IL-17 + T cells were detected by flow cytometry, HE staining was used to detect the pathological manifestations of thyroid tissue in each group. Results:Compared with the model group, the serum TSH [(3 328.88±724.45) pg/ml vs. (1 900.25±203.91) pg/ml] in Xiaoyao Powder group increased ( P<0.01), TGAb [(63.60±9.01) IU/ml vs. (96.19±10.74) IU/ml] and TPOAb [(6.84±1.45) IU/ml vs. (11.62±2.06) IU/ml] decreased ( P<0.01), and the expression of miR326 (3.57±0.57 vs. 7.63±0.90),IL-17 mRNA (6.71±0.97 vs. 13.02±1.18) significantly decreased ( P<0.01), the expression of Ets-1 (0.71±0.40 vs. 0.39±0.02) significantly increased ( P<0.01), the ratio of CD4 +IFN-γ + T cell [(13.10±2.23)% vs. (20.7±2.07)%], CD4 +IL-17 + T cell ratio [(18.90±1.31)% vs. (25.1±1.03)%] significantly decreased ( P<0.01), and thyroid histopathology changed significantly. Conclusion:Modified Xiaoyao Powder could regulate the expression of target protein Ets-1 upward, inhibit the differentiation of Th17 cells and further reduce the expression of IL-17 mRNA by regulating the expression of mir-326 downward in the thyroid tissue of EAT rats, so as to improve the inflammatory response of rats with liver depression and spleen deficiency.

The role of autophagy in the Treg/Th17 cell imbalance in mice with acute lung injury / 中华急诊医学杂志

Objective:To investigate the effect of autophagy on the Treg/Th17 cell imbalance in mice with acute lung injury (ALI).Methods:Twenty-four male SD mice were randomly divided into the sham operation group (S group), sepsis group (Sep group) and autophagy inhibitor 3-methyladenine group (Sep +3-MA group). ALI model was prepared by LPS tracheal dripping method. The mouse pathological injury score mice were evaluated under light microscopy and the W/D ratio was calculated. The counts of Th17 cells and Treg cells in tracheoalveolar lavage fluid (BALF) of mice and the levels of related cytokines were detected by flow cytometry. The expressions of LC3-Ⅱ, Beclin-1 and p62 in Th17 cells and Treg cells in BALF were determined by Western blot.Results:CCompared with the S group, the lung histopathological score and W/D ratio of the Sep group and Sep+3-MA group increased ( P<0.05). Compared with the Sep group, the count of Th17 cells in BALF of the Sep +3-MA group decreased, while the count of Treg cells increased significantly with the progression of sepsis( P<0.05), and the levels of IL-17, IL-10 and TNF-α were significantly decreased ( P<0.05). TGF-β1 levels increased in the early stages of sepsis, but decreased significantly with the progression of sepsis( P<0.05). Compared with the Sep group, LC3-Ⅱ expression in BALF Th17 cells and Treg cells of the Sep+3-MA group showed a downward trend, but there was no statistical difference, while Beclin-1 expression significantly decreased ( P<0.05), and the expression of p62 significantly increased ( P<0.05). Conclusions:Abnormal activation of autophagy in Th17 cells and Treg cells is involved in the immune imbalance of Th17/Treg cells in ALI with sepsis. Inhibition of autophagy can restore the functions of Th17 cells and Treg cells, and improve the imbalance of Th17/Treg by inhibiting autophagy may become a new idea to control the pathogenesis and progression of immune disorders with sepsis.

Immunoregulatory effects of miR-31-5p overexpression on peripheral blood Th17 cell in a rabbit model of autoimmune dry eye / 中华实验眼科杂志

Objective:To investigate the immunoregulatory effects of lentivirus-mediated microRNA (miR)-31-5p overexpression on peripheral blood T helper cell 17 (Th17) in a rabbit model of autoimmune dry eye.Methods:The miR-31-5p recombinant lentiviral vector was constructed.Lentivirus overexpressing miR-31-5p and its control virus were packaged.The concentration measurement and lentiviral titer determination were carried out.A rabbit model of autoimmune dry eye was established and the peripheral blood mononuclear cells (PBMC) of the rabbits were isolated.PBMC infected with miR-31-5p and negative control lentivirus particles were assigned as the miR-31-5p overexpression group and control group, respectively.The miR-31-5p expression level was detected using quantitative real-time PCR (qRT-PCR). Then PBMC in the two groups were co-cultured with γ-ray irradiated lacrimal gland epithelial cells.The expressions of Th17 cell related transcription factor retinoic acid-receptor-related orphan receptor C (RORC) and interleukin-17 (IL-17) mRNA, IL-1β, IL-6 and IL-23 were determined by qRT-PCR.The IL-17 protein expression level was detected by Western blot.The use and care of animals complied with Regulation for the Administration of Affair Concerning Experiment Animals by State Science and Technology Commission.The study protocol was approved by the Ethics Committee of Tianjin Medical University Eye Hospital (No.TJYY20201221036).Results:The construction of the miR-31-5p recombinant lentiviral vector was verified by DNA sequencing.The lentiviral titer of lentivirus overexpressing miR-31-5p and control lentivirus particles was 3.82×10 7 TU/ml and 3.50×10 7 TU/ml, respectively.The miR-31-5p relative expression level of PBMC was significantly increased in miR-31-5p overexpression group in comparison with control group, showing a statistically significant difference ( t=-9.696, P<0.001). When PBMC were co-cultured with lacrimal gland epithelial cells in vitro, the relative expression levels of RORC and IL-17 mRNA in miR-31-5p overexpression group were 0.33±0.03 and 0.28±0.09, which were significantly decreased in comparison with 1.00±0.00 and 1.00±0.00 in control group, with statistically significant differences between them ( t=46.256, 13.810; both at P<0.05). The relative expression level of IL-17 protein in miR-31-5p overexpression group was significantly reduced than control group ( t=4.977, P=0.008). The relative expression levels of IL-1β, IL-6 and IL-23 mRNA were significantly lower in miR-31-5p overexpression group than control group ( t=220.076, 6.641, 13.271; all at P<0.05). Conclusions:The overexpression of miR-31-5p can inhibit the Th17-immune response via down-regulating the expression of IL-6, IL-1β and IL-23.

The immune modulation of Th17 and its related cytokines in inflammatory bowel diseases / 中国医师杂志

Inflammatory bowel diseases (IBD) are a kind of non-specific inflammatory disease that occurs in gastrointestinal tract. Abnormal immune regulation is a key factor in its pathogenesis. The acquired immune regulation is mediated by helper T cells (Th), which is reported to play an important role in the pathogenesis of IBD. Th17 is a subtype of CD4 + T cells that could specifically produce interleukin-17 (IL-17) and other related cytokines. In this paper, we review the immune modulation of Th17 and its related cytokines in IBD.

Effects of hepatitis B virus on Th17, Treg and Th17/Treg ratio in different alanine aminetransferase stages / 北京大学学报(医学版)

OBJECTIVE@#To evaluate the effects of hepatitis B virus (HBV) on helper T lymphocytes 17 (Th17), regulatory T lymphocyte (Treg) and Th17/Treg ratio in chronic hepatitis B patients in different alanine aminetransferase (ALT) stages.@*METHODS@#In the study, 336 chronic hepatitis B patients in the first hospital of Lanzhou University were analyzed. The hepatitis B antigen antibody parameters were measured by chemiluminescence immunoassay analyzer, the liver function parameters were measured by automatic biochemical analyzer, the HBV loads were measured by quantitative PCR, Th17, Treg and Th17/Treg ratios were detected by flow cytometry. Among them, 111 cases (ALT < 40 U/L) of ALT were normal hepatitis B, 108 cases of chronic hepatitis B with ALT above normal upper limit and < 2 times higher (40 U/L≤ALT < 80 U/L), and 117 cases of chronic hepatitis B with ALT above 2 times normal upper limit (80 U/L≤ALT). According to the viral load, they were divided into low replication group with HBV DNA < 4.0 lg copies/mL, medium replication group with 4.0 lg copies/mL≤HBV DNA < 6.0 lg copies/mL and high replication group with HBV DNA ≥ 6.0 lg copies / mL. Dunnett T3 variance analysis were used to analyze the effects of HBV on Th17, Treg and Th17/Treg ratio in the chronic hepatitis B patients in different ALT stages. The changes of virological and immunological indexes before and after treatment were observed for 24 weeks of antiviral therapy in the hepatitis B patients with ALT≥double upper limit of normal group.@*RESULTS@#In the ALT normal group, different virus load HBV had minor effects on Th17, Treg and Th17/Treg ratio. In the ALT≥2 times upper limit of normal group, with the virus load increased, Th17 (3.18%±0.79% in low replication group, 3.78%±0.92% in medium replication group and 4.57%±1.15% in high replication group), Treg cells (5.52%±1.58% in low replication group, 5.89%±1.84% in medium replication group and 6.37%±2.35% in high replication group) and their ratio Th17/Treg (0.57±0.25 in low replication group, 0.65±0.29 in medium replication group and 0.73±0.36 in high replication group) were significantly increased (P < 0.05). After entecavir treatment 24 weeks, the patient' s HBV-DNA decreased significantly, Th17 (3.89%±1.02% vs. 2.06%±0.46%), Treg (6.02%±2.03% vs. 5.06%±1.25%), Th17/Treg ratio (0.65±0.28 vs. 0.41±0.14) decreased significantly (P < 0.05).@*CONCLUSION@#Investigation on the effects of HBV on Th17 and Treg cells and their ratios in different ALT states can clarify the effects of HBV on the body from the immunological perspective and can further understand the ALT grouping for antiviral treatment theoretical significance, which is helpful for clinical treatment.

Association of signal transducer and activator of transcription 3 and signal transducer and activator of transcription 5 with regulatory T cell/T helper 17 cell balance in chronic hepatitis B / 临床肝胆病杂志

The immune mechanism of chronic hepatitis B (CHB) persistent infection is closely associated with T cells, and the development of T cells requires the coordination of a variety of cytokines. The proteins of the signal transducer and activator of transcription (STAT) family are mainly involved in the signal transduction of cytokines, and STAT5a/b and STAT3 play an important role in the differentiation and development of regulatory T cells (Treg) and T helper 17 cells (Th17). This article analyzes the association of STAT3 and STAT5 with Treg/Th17 balance in CHB and investigates the chronicity of hepatitis B virus infection and the regulatory mechanism of liver inflammation.

Effects of astragaloside Ⅳ on inflammatory response and percentage of peripheral blood Th17 cells in mice with ulcerative colitis / 中国中药杂志

This study aimed to investigate the anti-inflammatory effect of astragaloside Ⅳ in mice with ulcerative colitis(UC) and its effect on the percentage of peripheral blood T helper(Th17) cells. Following the establishment of UC mouse model with 2% sodium dextran sulfate(DSS), mice in the positive control group and low-and high-dose astragaloside Ⅳ groups were treated with corresponding drugs by gavage. Disease activity index(DAI) was calculated, and serum interleukin-17(IL-17), tumor necrosis factor-α(TNF-α), and transforming growth factor-β(TGF-β) levels were assayed by ELISA. The pathological changes in colon tissue were observed by HE staining, and Th17/regulatory T cells(Treg) ratio in the peripheral blood was determined by flow cytometry. Western blot was conducted for detecting the relative protein expression levels of forkhead box protein P3(Foxp3) and retinoic acid-related orphan nuclear receptor γT(ROR-γt). The findings demonstrated that in normal mice, the colonic structure was intact. The goblet cells were not reduced and the glands were neatly arranged, with no mucosal erosion, bleeding, or positive cell infiltration. In the model group, the colonic mucosal structure was seriously damaged, manifested as disordered arrangement or missing of glands, vascular dilatation, congestion, and massive inflammatory cell infiltration. The pathological injury of colon tissue was alleviated to varying degrees in drug treatment groups. Compared with the normal group, the model group exhibited elevated percentage of Th17 cells, increased IL-17 and TNF-α content, up-regulated relative ROR-γt protein expression, lowered TGF-β, reduced percentage of Treg cells, and down-regulated relative Foxp3 protein expression. The comparison with the model group showed that DAI score, pathological score, percentage of Th17 cells, IL-17 and TNF-α content, and relative ROR-γt protein expression in the positive control group, low-dose astragaloside Ⅳ group, and high-dose astragaloside Ⅳ group were decreased, while TGF-β content, percentage of Treg cells, and relative Foxp3 protein expression were increased. The DAI score, pathological score, percentage of Th17 cells, IL-17 and TNF-α content, and relative ROR-γt protein expression in the low-dose astragaloside Ⅳ group were higher than those in the positive control group, whereas the content of TGF-β, percentage of Treg cells, and relative Foxp3 protein expression were lower. DAI score, pathological score, percentage of Th17 cells, IL-17 and TNF-α content, relative ROR-γt protein expression in the high-dose astragaloside Ⅳ group declined in contrast to those in the low-dose astragaloside Ⅳ group, while the TGF-β content, percentage of Treg cells, and relative Foxp3 protein expression rose. There was no significant difference between the positive control group and the high-dose astragaloside Ⅳ group. Astragaloside Ⅳ is able to inhibit inflammatory response and diminish the percentage of Th17 cells in mice with UC.

Research progress in Th17 cells and the relevant cytokines in Graves ' ophthalmopathy / 中南大学学报(医学版)

Graves' ophthalmopathy is the most common clinical orbital disease, and T helper (Th) cells play an important role in the development of Graves' ophthalmopathy. Th17 cells are a major subpopulation of Th cells and abnormally highly expressed in patients with Graves' ophthalmopathy. Th17 cells and the related cytokines interleukin (IL)-17A, IL-21 and IL-23 are involved in regulating the inflammatory response, fibrosis and adipogenesis. Th17 cells are unstable and exhibit a degree of plasticity, and they can differentiate into IL-17A and interferon (IFN)-γ dual-producing Th17.1 cells, which exacerbate the pathogenicity of Th17 cells. In addition, Th17 cells and the relevant factors are strongly associated with disease activity and severity in Graves' ophthalmopathy.

Effect of interleukin-23R overexpression on Th17/Treg balance in experimental autoimmune uveitis mice / 中华眼底病杂志

Objective:To investigate the effect of interleukin (IL)-23 receptor (IL-23R) overexpression on the balance of T helper 17 (Th17 cells)/regulatory T cells (Treg cells) in experimental autoimmune uveitis (EAU) mice.Methods:Twelve 8-week-old female C57BL/6J mice were randomly divided into LV-Ctrl group and LV-IL-23R group, with 6 mice in each group. Two groups of mice were injected with LV-Ctrl and LV-IL-23R lentiviruses through the tail vein, respectively; 7 days after injection, the EAU mouse model was established by active immunization with vitamin A-binding protein 1-20 between photoreceptors. Starting from 13 days after immunization, the fundus of the mice was observed by indirect ophthalmoscopy every 2 days and clinical scores were performed; 30 days after immunization, hematoxylin-eosin staining was used to observe the histopathological changes of mouse retina. The levels of IL-17 in serum of the two groups of mice were detected by enzyme-linked immunosorbent assay; the proportion of Th17 cells and Treg cells was detected by flow cytometry. The relative mRNA expression of IL-23R, IL-17, retinoic acid-related orphan receptor γt (RORγt), IL-10 and forkhead transcripyion factor p3 (Foxp3) were detected by real-time quantitative polymerase chain reaction. Comparisons between groups were performed using repeated measures analysis of variance, independent samples Mann-Whitney U test, and independent samples t test. Results:Compared with the LV-Ctrl group, the retinal inflammatory reaction of the LV-IL-23R group was more severe. At 13 days after immunization, there was no significant difference in fundus inflammation scores between LV-IL-23R group and LV-Ctrl group ( t=-2.001, P=0.058); 15-29 days after immunization. The fundus inflammation scores of LV-IL-23R group were higher than those of LV-Ctrl group, and the difference was statistically significant ( t=-4.429,-6.578, -7.768, -10.183, -6.325, -7.304, -4.841, -6.872; P<0.001). Histopathological examination showed that the infiltration of inflammatory cells in the fundus increased, the retinal structure was damaged more seriously, and the histopathological score was significantly increased, and the difference was statistically significant ( t=-4.339, P=0.001). Compared with the LV-Ctrl group, the relative expression of IL-23R mRNA in the spleen of the LV-IL-23R group was significantly increased, and the difference was statistically significant ( Z=2.087, P=0.037). The relative expression of IL-17 and RORγt mRNA increased, while the relative expression of IL-10 and Foxp3 mRNA decreased, and the differences were statistically significant ( t=-6.313,-5.922, 4.844, 7.572; P=0.003, 0.004, 0.008, 0.002). Compared with the LV-Ctrl group, the level of IL-17 in the serum of the mice in the LV-IL-23R group was significantly increased, and the difference was statistically significant ( t=-5.423, P=0.002); the proportion of Th17 cells in the spleen and lymph nodes was significantly increased, whereas, the proportion of Treg cells was significantly reduced, and the difference was statistically significant ( t=-4.290, 3.700; P=0.002, 0.006). Conclusion:IL-23R overexpression can promote Th17/Treg imbalance in EAU mice, and aggravate the clinical and pathological manifestations of EAU.

Role of glycolysis in Th17 cell differentiation in autoimmune diseases / 中华微生物学和免疫学杂志

T helper 17 (Th17) cells are closely associated with the pathogenesis of several autoimmune and inflammatory diseases, and selective suppression of Th17 cell production and pathogenicity is an effective strategy for the treatment of these diseases. There is growing evidence that cellular metabolism is associated with the development of autoimmune diseases and determines the differentiation and effector functions of Th17 cells, which undergoes a metabolic reorganization during differentiation from an oxidative phosphorylation-based catabolism to a glucose-based anabolic metabolism in the initial T cells. This paper focused on reviewing recent findings regarding the importance of metabolism in T cell differentiation and autoimmune diseases, especially in Th17 cells, and discussing the regulatory mechanisms of glycolysis in Th17 cell differentiation. This review summarized the regulation of metabolism on T-cell activation and differentiation, revealed metabolic targets with specific regulation on Th17 cells, and provided reference for finding potential therapeutic targets for Th17 cell-mediated autoimmune diseases.

Mechanism of bigelovin in treatment of imiquimod-induced psoriasiform mouse model / 中国药理学通报

Aim To investigate the effects of bigelovin on mouse model of imiquimod-induced psoriatic itch and its mechanism. Methods Psoriasis-like mouse model was established by applying imiquimod cream on the back skin of mouse. Psoriasis area and severity index, pathological changes, the expression levels of inflammatory factors and related molecular biological data were used as effect indicators. The changes of the above parameters were observed after administration of different concentrations of bigelovin. Then the possible mechanism of the effects was further analysed.Results Compared with the model group, bigelovin significantly decreased the symptoms of skin lesions and reduced the PASI score. Bigelovin alleviated epidermal thickening and reduced the expression of Ki67 in a dose-dependent manner. The expression levels of inflammatory factors were reduced in both skin and serum.The percentage of Th17 cells was reduced and the percentage of Treg cells was increased in the lymph node.In addition, bigelovin also inhibited the phosphorylation of P65 protein and significantly reduced the nuclear localization of P65, suggesting that bigelovin might inhibit the activation of P65 protein. Conclusions The effect of bigelovin on improving the signs and symptoms of imiquimod-induced psoriasis mice may be related to the inhibition of P65 protein phosphorylation in keratinocytes.