Serum autophagy-related gene 5 level in stroke patients: correlation with CD4+ T cells and cognition impairment during a 3-year follow-up

Abstract Autophagy-related gene (ATG) 5 regulates blood lipids, chronic inflammation, CD4+ T-cell differentiation, and neuronal death and is involved in post-stroke cognitive impairment. This study aimed to explore the correlation of serum ATG5 with CD4+ T cells and cognition impairment in stroke patients. Peripheral blood was collected from 180 stroke patients for serum ATG5 and T helper (Th) 1, Th2, Th17, and regulatory T (Treg) cell detection via enzyme-linked immunosorbent assays and flow cytometry. The Mini-Mental State Examination (MMSE) scale was completed at enrollment, year (Y)1, Y2, and Y3 in stroke patients. Serum ATG5 was also measured in 50 healthy controls (HCs). Serum ATG5 was elevated in stroke patients compared to HCs (P<0.001) and was positively correlated to Th2 cells (P=0.022), Th17 cells (P<0.001), and Th17/Treg ratio (P<0.001) in stroke patients but not correlated with Th1 cells, Th1/Th2 ratio, or Treg cells (all P>0.050). Serum ATG5 (P=0.037), Th1 cells (P=0.022), Th17 cells (P=0.002), and Th17/Treg ratio (P=0.018) were elevated in stroke patients with MMSE score-identified cognition impairment vs those without cognition impairment, whereas Th2 cells, Th1/Th2 ratio, and Treg cells were not different between them (all P>0.050). Importantly, serum ATG5 was negatively linked with MMSE score at enrollment (P=0.004), Y1 (P=0.002), Y2 (P=0.014), and Y3 (P=0.001); moreover, it was positively related to 2-year (P=0.024) and 3-year (P=0.012) MMSE score decline in stroke patients. Serum ATG5 was positively correlated with Th2 and Th17 cells and estimated cognitive function decline in stroke patients.

Regulation of Immune Balance by Traditional Chinese Medicine in Treatment of Cough Variant Asthma： A Review / 中国实验方剂学杂志

Cough variant asthma （CVA） is a chronic respiratory disease with cough as its main symptom. The occurrence of CVA is closely related to non-specific airway inflammation， and its pathogenesis involves environmental， genetic， immune， and other factors. In recent years， the advantages of traditional Chinese medicine （TCM） in the treatment of CVA have attracted the attention of experts and scholars in China and abroad， especially its prominent role in regulating immune balance， relieving cough symptoms in CVA patients， and reducing recurrence. T Helper cells 1 （Th1）， T helper cells 2 （Th2）， T helper cells 17 （Th17）， and regulatory T cells （Treg） are derived from CD4+ T cells. Immune imbalance of Th1/Th2 and Th17/Treg is a new hotspot in the pathogenesis of CVA and a potential key target in the treatment of CVA by TCM. Th cell subsets are in dynamic balance under physiological conditions， maintaining respiratory immune homeostasis in which pro-inflammatory cytokines and anti-inflammatory cytokines are balanced. Immature helper T cells （Th0） can be differentiated into Th1， Th2， Th17， Treg， and other cell subsets due to cytokine types in the microenvironment in the stage of CVA maturation. The proliferation of Th2 cells leads to eosinophilic airway inflammation. Excessive differentiation of Th17 cells induces neutrophil airway inflammation. Th1/Th2 and Th17/Treg cells are mutually restricted in number and function， and the immune imbalance of Th1/Th2 and Th17/Treg is easy to aggravate the generation of inflammatory response. Restoring immune balance is particularly important for the airway anti-inflammatory therapy of CVA. In this paper， the imbalance of Th1/Th2 and Th17/Treg and the pathogenesis of CVA were systematically expounded. Meanwhile， the latest research on the regulation of immune imbalance by TCM compound， single TCM， and its effective ingredients in the treatment of CVA was reviewed. It provides ideas and references for revealing the scientific connotation of TCM regulating immune balance therapy of CVA， as well as the development of clinical treatment and basic research of CVA.

Effect of Buzhong Yiqitang on Th17/Treg Immune Imbalance and Notch1 Signaling Pathway in AIT Mice / 中国实验方剂学杂志

ObjectiveTo explore the effect of Buzhong Yiqitang on the immune imbalance of helper T cell 17 （Th17）/regulatory T cell （Treg） and Notch1 signaling pathway in mice with autoimmune thyroiditis （AIT）. MethodA total of 60 8-week-old NOD.H-2h4 mice were randomly divided into the normal group， model group， western medicine group （selenium yeast tablet， 32.5 mg·kg-1）， and low-dose （4.78 g·kg-1·d-1）， middle-dose （9.56 g·kg-1·d-1）， and high-dose （19 g·kg-1·d-1） Buzhong Yiqitang groups， with 10 mice in each group. The normal group was fed with distilled water， and the other groups were fed with water containing 0.05% sodium iodide for eight weeks. After the animal model of AIT was formed spontaneously， the mice were killed under anesthesia after intragastric administration for eight weeks. Serum anti-thyroglobulin antibodies （TGAb）， thyroid-stimulating hormone （TSH）， free triiodothyronine （FT3）， and free thyroid hormone （FT4） were detected by enzyme-linked immunosorbent assay （ELISA）， and thyroid tissue changes were observed by hematoxylin-eosin （HE） staining. The mRNA and protein expressions of retinoid-related orphan receptor-γt （RORγt）， interleukin （IL）-17， forkhead box P3 （FoxP3）， IL-10， Notch1， and hair division-related enhancer 1 （Hes1） in thyroid tissue were detected by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot， respectively. ResultCompared with the normal group， the thyroid structure of the model group was severely damaged， and lymphocytes were infiltrated obviously. The levels of serum TGAb， FT3， and FT4 contents were significantly increased， and TSH content was significantly decreased （P<0.01）. The mRNA and protein expression levels of RORγt， IL-17， Notch1， and Hes1 were significantly increased， while those of FoxP3 and IL10 were significantly decreased in the model group （P<0.01）. Compared with the model group， thyroid structural damage and lymphocyte infiltration were improved in the treatment groups， and serum TGAb， FT3， and FT4 contents were significantly decreased. TSH content was increased， and mRNA and protein expression levels of RORγt， IL-17， Notch1， and Hes1 were decreased. mRNA and protein expression levels of FoxP3 and IL-10 were increased to different degrees （P<0.05， P<0.01）， and the middle-dose Buzhong Yiqitang group had the most significant intervention effect. ConclusionBuzhong Yiqitang can alleviate the thyroid structural damage in AIT mice， and its mechanism may be related to improving the abnormal differentiation of Th17/Treg immune cells and inhibiting the activation of the Notch1 signaling pathway.

Shaoyaotang Alleviates Ulcerative Colitis by Regulating Th17/Treg Balance Through IL-6/STAT3 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo explore the role of interleukin-6 （IL-6）/signal transducer and activator of transcription 3 （STAT3） pathway in the balance of T helper 17 （Th17）/regulatory T （Treg） cells in ulcerative colitis （UC） with internal dampness-heat accumulation syndrome and the intervention mechanism of Shaoyaotang. MethodA total of 60 SD rats were randomized into blank group （equivalent volume of normal saline）， model group （equivalent volume of normal saline）， western medicine control group （0.42 g·kg-1 mesalazine）， and low-dose （11.1 g·kg-1）， medium-dose （22.2 g·kg-1）， and high-dose （44.4 g·kg-1） Shaoyaotang groups. UC with internal dampness-heat accumulation syndrome was induced in rats with the compound method except for the blank group. The administration lasted 14 days for each group. At 24 h after the last administration， rats were killed and the spleen and colon tissues were separated. The histopathological changes of colon were observed based on hematoxylin and eosin （HE） staining and the levels of interleukin-17 （IL-17） and transforming growth factor-β1 （TGF-β1） in colon tissue were detected by immunohistochemistry （IHC）. Flow cytometry was employed to determine the levels of Th17/Treg cells in the spleen， and Western blot to measure the levels of IL-6 and STAT3 proteins in colon tissue. ResultCompared with the blank group， the model group had lesions such as congestion and erosion， low percentage of spleen Treg cells （P<0.01）， high percentage of Th17 cells （P<0.01）， and high levels of IL-6 and STAT3 proteins in colon tissue （P<0.01）. Compared with the model group， the administration groups showed alleviation of colon injury， high percentage of spleen Treg cells （P<0.05， P<0.01）， low percentage of Th17 cells （P<0.01）， and low levels of IL-6 and STAT3 proteins in colon tissue （P<0.01）. ConclusionShaoyaotang regulates the balance of Th17/Treg by inhibiting the IL-6/STAT3 pathway， thereby relieving the pathological damage of UC rats with internal dampness-heat accumulation syndrome and affecting their immune function.

Clinical Efficacy and Mechanism of Erzhi Tiangui Prescription in Treatment of Patients with Repeated Implantation Failure of Kidney Deficiency Syndrome Based on sPD-1 and Th17/Treg Cytokines / 中国实验方剂学杂志

ObjectiveTo investigate the clinical efficacy and possible mechanism of Erzhi Tiangui prescription on repeated implantation failure （RIF） of kidney deficiency syndrome. MethodSeventy patients with RIF of kidney deficiency syndrome who underwent natural cycle frozen-thawed embryo transfer （FET） in the Reproductive and Genetic Center of the Affiliated Hospital of Shandong University of Traditional Chinese Medicine were enrolled and randomly divided into a treatment group （35 cases） and a control group （35 cases）. Patients in the treatment group took oral Erzhi Tiangui prescription from the third day of each menstrual cycle two months before the FET cycle and continued to take it until the day of transplantation from the third day of the menstrual cycle in the month of transplantation. Those in the control group did not accept traditional Chinese medicine （TCM）. In addition，10 patients who successfully achieved clinical pregnancy after the first natural cycle FET were screened from the reproductive medical record bank of this hospital and assigned to the normal group. Peripheral blood samples of patients in the three groups on the day of embryo transfer were collected from the specimen bank of the Reproductive and Genetic Center. Serum soluble programmed death molecule-1 （sPD-1），soluble programmed death molecule-ligand 1 （sPD-L1），transforming growth factor-β （TGF-β），interleukin-17 （IL-17）， and interleukin-10 （IL-10） levels were measured by enzyme-linked immunosorbent assay （ELISA）. The changes in kidney deficiency syndrome scores， the final biochemical pregnancy rates， clinical pregnancy rates， and embryo implantation rates of the treatment group and the control group before and after treatment were observed. ResultCompared with the normal group，the model group showed increased serum levels of sPD-1 and IL-17（r=0.347，P<0.05），decreased levels of IL-10 and TGF-β （P<0.01），and non-significant change in sPD-L1 level. Serum sPD-1 was positively correlated with IL-17 （P<0.05） and negatively correlated with IL-10（r=-0.521，P<0.01） and TGF-β（r=-0.457，P<0.01） in RIF patients with kidney deficiency syndrome. After TCM treatment，compared with the control group， the treatment group showed improved TCM syndrome score （P<0.05） and increased clinical pregnancy rate and embryo transfer rate（P<0.05），but there was no statistically significant difference in the biochemical pregnancy rate between the two groups. ConclusionAbnormal expression of sPD-1 in patients with RIF of kidney deficiency syndrome breaks the balance of T helper 17 （Th17）/regulatory T cell （Treg），which is not conducive to embryo implantation and pregnancy maintenance. Erzhi Tiangui prescription，a TCM for tonifying the kidney，can significantly improve the symptoms of kidney deficiency in patients with RIF of kidney deficiency syndrome，reduce the concentrations of sPD-1 and IL-17 in the peripheral serum，increase the levels of TGF-β and IL-10，regulate the peripheral Th17/Treg immune balance，and increase the implantation rate and clinical pregnancy rate，which has a high clinical value.

Cellular and molecular mechanisms of Notch signal in pulmonary microvascular endothelial cells after acute lung injury

Abstract This study focused on the effect and mechanism of Notch signal on pulmonary microvascular endothelial cells (PMVECs) following acute lung injury. PMVECs were cultured in vitro and randomly divided into eight groups. Grouping was based on whether cells were co-cultured with T cells (splenic CD4+T cells were isolated using MACS microbeads) and the level of Notch expression: Normal group and Normal+T cells group, Model group and Model+T cells group, Notch low-expression group and Notch low-expression+T cells group, and Notch overexpression group and Notch overexpression+T cells group. Except for the Normal group and Normal+T cells group, all other groups were treated with 500 μL lipopolysaccharide (1 μg/mL). The expression of VE-cadherin and Zo-1 protein in the Model group (with or without T cells) was lower than that in the normal group (with or without T cells), their expression in the Notch low-expression group (with or without T cells) was significantly increased, and their expression in the Notch overexpression group (with or without T cells) was significantly decreased. Compared with the normal+T cells group, the number of Treg cells in the Notch low-expression+T cells group decreased significantly (P<0.01). The number of Th17 cells in the Notch overexpression+T cells group was higher than that in the Model+T cells group (P<0.01), while the number of Treg cells decreased (P<0.01). Our results demonstrated that activated Notch signal can down-regulate the expression of the tight junction proteins VE-Cadherin and Zo-1 in PMVECs and affect Th17/Treg immune imbalance. Autophagy was discovered to be involved in this process.

Influence of matrine on Th17/Treg balance in rats with inflammatory bowel disease by regulating IL-6/STAT3/NF-κB signaling pathway / 西安交通大学学报(医学版)

【Objective】 To investigate the influence of matrine (MT) on the balance of T helper cell 17 (Th17)/regulatory T cells (Treg) in rats with inflammatory bowel disease by regulating interleukin-6 (IL-6)/signal transducer and activator of transcription 3 (STAT3)/nuclear transcription factor-κB (NF-κB) pathway. 【Methods】 SD rats were grouped into control check group (CK group), model group, low-dose MT group (MT-L group, 50 mg/kg), medium-dose MT group (MT-M group, 100 mg/kg), high-dose MT group (MT-H group, 200 mg/kg), mesalazine group (MSLM group, 0.42 g/kg), and MT-H+rIL-6 (IL-6 activator) group (200 mg/kg+0.05 mg/kg) according to the random number table method, with 18 in each group. Except for the CK group, the rats in other groups all received with 5% trinitrobenzenesulfonic acid (20 mg/kg) buffer solution mixed with 50% ethanol at a ratio of 1∶1 and then enema to construct a rat model of inflammatory bowel disease. After the successful modeling, they were treated with drug administration once a day for 7 weeks. The body weight of rats was measured at 1, 3, 5, and 7 weeks of administration; the changes of colon length of rats in each group were compared; HE staining was used to detect the pathological damage of rat colon tissue; the levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-17 and IL-10 in serum of rats were detected by ELISA; the proportions of Th17 and Treg cells in peripheral blood of rats were detected by flow cytometry; Western blottingt was performed to detect the protein expression of retinoic acid-related orphan receptor γt (RORγt), forkhead box protein P3 (Foxp3), IL-6, p-STAT3, and p-NF-κB p65 in rat colon tissue. 【Results】 Compared with the CK group, the colon tissue of the model group was severely damaged by pathology, and the body weight (at 3, 5, and 7 weeks), the level of IL-10, the proportion of Treg cell, and the expression of Foxp3 protein were decreased, the colon length shortened, the levels of TNF-α, IL-17, the proportions of Th17 cell, Th17/Treg ratio, and the protein expression of RORγt, IL-6, p-STAT3, and p-NF-κB p65 increased (P<0.05). Compared with the model group, the corresponding indicators of the MT-L group, MT-M group, MT-H group, and MSLM group had the opposite trends (P<0.05); rIL-6 attenuated the promoting effect of high-dose MT on Th17/Treg balance in inflammatory bowel disease rats. 【Conclusion】 MT may promote Th17/Treg balance in inflammatory bowel disease rats by inhibiting IL-6/STAT3/NF-κB signaling pathway.

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.

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.

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.

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 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.

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 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.

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.

Effect of mechanical stimulation by rubbing abdomen by interfering with Th17 cell function on intestinal homeostasis in IBS-C model mice / 国际生物医学工程杂志

Objective:To investigate the possible mechanism of rubbing abdomen to regulate intestinal homeostasis in irritable bowel syndrome with constipation (IBS-C) mouse models.Methods:IBS-C mouse models of intestinal immune dysfunction were established using trinitrobenzene sulfonic acid (TNBS)-induced C57BL/J6 male mice. Thirty C57BL/J6 mice were randomly divided into three groups: the control group, the model group, and the mogul group. After 7 days of modeling, mice in the mogul group were given a mogul mechanical stimulation intervention once per day for 2 weeks, while mice in the control and model groups were not given any intervention. Changes in serum levels of interleukin-6 (IL-6) and IL-17A were detected by enzyme-linked immune sorbent assay (ELISA) and immunohistochemistry. The gene expression and protein levels of IL-17A and IL-23 were detected by quantitative real-time fluorescence PCR (qRT-PCR) and Western Blot, respectively. The morphological changes were observed by HE staining. The CD44 and CD62L expression changes were observed by immunofluorescence staining.Results:Compared with the model group, the levels of IL-6 and IL-17A in the serum of mice in the mogul group were decreased, and the expression of IL-6 and IL-7A in the tissues was down-regulated (all P<0.001). In addition, the gene expression and protein expression levels of IL-17A and IL-13 in the tissues of mice in the mogul group were decreased (all P<0.001). HE staining results showed that the mogul mechanical stimulation intervention could repair colonic tissues and reduce the inflammatory response. Immunofluorescence staining results showed that mogul mechanical stimulation intervention could downregulate the expression of CD44 but had no modulating effect on the expression of CD62L. Conclusions:Rubbing abdomen can improve intestinal homeostasis in IBS-C model mice by regulating changes in Th17 cell function.

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.

Effect of Asiaticoside on Expression of Th17/Treg Cells in DBA/1 Mice with Collagen-Induced Arthritis / 中国实验方剂学杂志

ObjectiveTo observe the effect of asiaticoside （AC） on the expression of T helper 17 （Th17） cells and regulatory T （Treg） cells in DBA/1 mice with collagen-induced arthritis （CIA）. MethodMale SPF DBA/1 mice were randomized into six groups according to body weight： control group， CIA group， methotrexate group （MTX group， ip， 0.5 mg·kg-1）， and AC low-， medium-， and high-dose groups （ig， 5， 15， 45 mg·kg-1， respectively）. Modeling was performed in rats other than the control group. To be specific， they were immunized with bovine type Ⅱ collagen and complete Freund's adjuvant on the first day and with bovine type Ⅱ collagen and incomplete Freund's adjuvant on the 21st day. Administration began on the day of the second immunization， once a day for 28 days. On the 49th day， related tissues were collected. Then， hematoxylin-eosin （HE） staining was performed to observe the pathological changes of the joints. Immunohistochemical method was used to detect the expression of interleukin-17 （IL-17） and forkhead box protein-3 （FoxP3）， the markers of Th17 and Treg cells， respectively， immunofluorescence double staining the expression of IL-17 and FoxP3 in CD4+T cells of mouse joint tissue， and flow cytometry the proportions of Th17 and Treg cells in mouse lymph nodes. ResultCompared with the control group， CIA group demonstrated joint disorder， damage of articular cartilage and bone， severe bone erosion （P<0.01）， increase in stained CD4 and IL-17 and the integral absorbance （IA） （P<0.01）， decrease in stained FoxP3 and the IA （P<0.01）， rise of Th17/Treg ratio （P<0.01）， elevation of Th17 expression in mouse lymph nodes （P<0.01）， and reduction in Treg expression （P<0.01）. Compared with CIA group， MTX group and three AC groups showed normal joints， alleviated bone erosion and damage， intact and smooth joint surface， and decrease in stained IL-17 and IA （P<0.05， P<0.01）， and MTX group and AC medium-dose and high-dose groups registered decrease in stained CD4 and IA （P<0.01） and reduction in Th17/Treg ratio （P<0.05， P<0.01）. Moreover， AC medium-dose and high-dose groups showed rise in stained FoxP3 and IA （P<0.05， P<0.01）. In the lymph nodes of mice， decrease in expression of Th17 cells （P<0.05， P<0.01） and the increase in expression of Treg cells （P<0.05， P<0.01） were observed in all the three AC group. ConclusionAC can regulate Th17/Treg balance by inhibiting the expression of Th17 cells and promoting the expression of Treg cells in CIA mice.

Immunomodulatory Mechanism of Kangxian Yixin Prescriptions on Autoimmune Injury Mice Based on Th17/Treg Cell Balance / 中国实验方剂学杂志

ObjectiveTo investigate the immunomodulatory mechanism of Kangxian Yixin prescription （KYP） on autoimmune injury mice and its relationship with the T helper 17 （Th17）/regulatory T cell （Treg） balance. MethodSixty healthy 8-week-old male BALBc mice were randomly divided into a normal group and an experimental group at a ratio of 1∶5. On the 0th， 7th， and 28th days， 0.2 mL of porcine cardiac myosin emulsion （containing 0.1 mg of porcine cardiac myosin） was subcutaneously injected into the groin， armpit， and back of the mice in the experimental group to induce an animal model of myocardial immune injury. Mice with myocardial immune injury were randomly divided into a model group （Model）， a KYP group （20.4 g·kg-1·d-1， ig）， and a valsartan group （12 mg·kg-1·d-1， ig）. Mice in the control group and the model group received the same amount of normal saline by gavage. After four weeks of intervention， the heart tissues were collected. Hematoxylin-eosin （HE） staining and Masson staining were used to detect pathological damage in heart tissues. Enzyme-linked immunosorbent assay （ELISA） was used to detect the expression of type B-type natriuretic peptide （BNP）， anti-cardiac antibody， interleukin-17 （IL-17）， and interleukin-10 （IL-10） in the serum of mice， and the expression levels of Th17 cells and Tregs in the spleen were detected by flow cytometry. The protein expression of B-cell lymphoma-2 （Bcl-2） and Bcl-2-associated X （Bax） in heart tissues was detected by Western blot， and the mRNA expression of retinoid-related orphan receptor gamma t （RORγt） and forkhead box P3 （FoxP3） in the spleen was detected by quantitative real-time polymerase chain reaction （Real-time PCR）. ResultCompared with the control group， the model group showed worsened pathological damage in heart tissues， elevated serum levels of BNP， anti-myocardial antibody， and IL-17， decreased serum expression of IL-10 （P<0.05）， increased expression of Th17 cells and reduced expression of Tregs in spleen tissues （P<0.05）， increased protein expression of Bax， diminished Bcl-2 protein expression， elevated Bax/Bcl-2 ratio， up-regulated mRNA expression of RORγt， dwindled mRNA expression of FoxP3， and elevated ratio of RORγt/FoxP3 （P<0.05）. Compared with the model group， the KYP group and the valsartan group displayed relieved pathological damage in heart tissues， decreased serum expression of BNP， anti-myocardial antibody， and IL-17， increased serum expression of IL-10 （P<0.05）， reduced expression of Th17 cells and increased Tregs in spleen tissues （P<0.05）， dwindled protein expression of Bax and elevated protein expression of Bcl-2 in heart tissues （P<0.05）， diminished Bax/Bcl-2 ratio， reduced mRNA expression of RORγt， up-regulated FoxP3， and down-regulated ratio of RORγt/FoxP3 （P<0.05）. ConclusionKYP may improve myocardial immune damage by regulating the Th17/Treg cell balance.