Objective:To analyze the characteristics of CD4 + T cell subsets and cytokines in patients with mixed connective tissue disease (MCTD) and the correlation of MCTD disease activity, laboratory data, and clinical symptoms with cytokines. Methods:A total of 48 MCTD patients (including 24 newly diagnosed patients and 24 treated patients) were enrolled from the Department of Rheumatology and Immunology, the Second Hospital of Shanxi Medical University from 2018 to 2021. Meanwhile, 49 healthy subjects who underwent physical examination were recruited (healthy control group). The absolute counts of CD4 + T cell subsets in peripheral blood samples were analyzed by flow cytometry. The levels of serum cytokines were detected by flow bead array. Analysis of variance and Mann-Whitney U test were used to compare the differences between groups. Pearson or Spearman correlation analysis was used for correlation analysis. Logistic regression analysis was used to analyze related factors. The receiver operating characteristic curve was used to detect the best cut-off value and effectiveness. Results:The absolute counts of Th1 ( P<0.01), Th2 ( P<0.01) and Treg cells ( P<0.01) in the newly diagnosed MCTD patients and the treated MCTD patients were lower than those in the healthy subjects. The levels of cytokines (IL-2, IL-4, IL-6, IL-10, IL-17, IFN-γ, and TNF-α) in the two MCTD groups were higher than those in the healthy control group ( P<0.01). Further analysis revealed that the cardiac enzymes in MCTD patients included creatine kinase, creatine kinase-MB, aspartate aminotransferase, α-hydroxybutyrate dehydrogenase, and lactate dehydrogenase were positively correlated with cytokines ( P<0.05). In addition, it was found that IL-2 was positively correlated with erythrocyte sedimentation rate ( r=0.477, P<0.001), but it was negatively correlated with complement C3 ( r=-0.546, P=0.002) and complement C4 ( r=-0.422, P=0.02). IL-10 was correlated with the myositis symptoms in MCTD patients and the area under the receiver operator characteristic curve was 0.745 (95% CI: 0.576-0.915, P<0.05). Conclusions:This study provides insights into the unique immunological characteristics of CD4 + T lymphocyte subsets and cytokines in patients with MCTD, and also reveals a close correlation between cytokines and cardiac enzymes in MCTD patients. IL-2 has been shown to be associated with disease activity in MCTD patients. The level of IL-10 may be related to the occurrence of myositis symptoms in MCTD.

Objective To observe the effects of Dihong Prescription on glomerular mesangial cells of mice cultured with high glucose;To explore its mechanism in the treatment of diabetic nephropathy.Methods The TargetScan database was used to predict the miRNA regulating bispecific phosphatase 1(DUSP1),and the double luciferase was used to verify the targeting relationship between miR-133b and DUSP1.The glomerular mesangial cells of mice were cultured in vitro and divided into control group,high glucose group and Dihong Prescription low-,medium-and high-dosage groups,and was given corresponding intervention.CCK-8 was used to detect cell viability,TUNEL staining was used to observe the apoptosis of cells,flow cytometry was used to determine the expression of reactive oxygen species(ROS),Western blot was used to detect the protein expressions of phosphorylated extracellular regulatory protein kinase 1/2(p-ERK1/2),ERK1/2,phosphorylated c-Jun amino-terminal kinase(p-JNK),JNK,p-p38,p38,mitochondrial dynamoculture-associated protein 1(Drp1)and mitochondrial fission protein 1(Fis1),transmission electron microscopy was used to observe the ultrastructure of mitochondria.Results The double luciferase demonstrated that miR-133b could target the regulation of DUSP1.Compared with the control group,the cell viability significantly decreased in the high glucose group(P<0.01),the apoptosis rate significantly increased(P<0.01),the content of ROS significantly increased(P<0.01),and the protein expressions of p-ERK1/2/ERK1/2,p-p38/p38,p-JNK/JNK,Drp1 and Fis1 significantly increased(P<0.01).Compared with the high glucose group,the cell viability significantly increased in Dihong Prescription medium-and high-dosage groups(P<0.01),the apoptosis rate significantly decreased(P<0.01),the content of ROS significantly decreased(P<0.01),and the protein expressions of p-ERK1/2/ERK1/2,p-p38/p38,p-JNK/JNK,Drp1 and Fis1 significantly decreased(P<0.01).Transmission electron microscopy showed that large number of autophagosomes,autophagosomes,mitochondria vacuolation and cristae fusion were observed in the high glucose group;the mitochondrial damage caused by high glucose was alleviated in Dihong Prescription low-,medium-and high-dosage groups.Conclusion Dihong Prescription can improve oxidative stress injury in glomerular mesangial cells of mice cultured with high glucose,and its mechanism may be related to the regulation of mitochondrial function and miR-133b/DUSP1/MAPK pathway.

Objective To observe the effects of Dihong Prescription on glomerular mesangial cells of mice cultured with high glucose;To explore its mechanism in the treatment of diabetic nephropathy.Methods The TargetScan database was used to predict the miRNA regulating bispecific phosphatase 1(DUSP1),and the double luciferase was used to verify the targeting relationship between miR-133b and DUSP1.The glomerular mesangial cells of mice were cultured in vitro and divided into control group,high glucose group and Dihong Prescription low-,medium-and high-dosage groups,and was given corresponding intervention.CCK-8 was used to detect cell viability,TUNEL staining was used to observe the apoptosis of cells,flow cytometry was used to determine the expression of reactive oxygen species(ROS),Western blot was used to detect the protein expressions of phosphorylated extracellular regulatory protein kinase 1/2(p-ERK1/2),ERK1/2,phosphorylated c-Jun amino-terminal kinase(p-JNK),JNK,p-p38,p38,mitochondrial dynamoculture-associated protein 1(Drp1)and mitochondrial fission protein 1(Fis1),transmission electron microscopy was used to observe the ultrastructure of mitochondria.Results The double luciferase demonstrated that miR-133b could target the regulation of DUSP1.Compared with the control group,the cell viability significantly decreased in the high glucose group(P<0.01),the apoptosis rate significantly increased(P<0.01),the content of ROS significantly increased(P<0.01),and the protein expressions of p-ERK1/2/ERK1/2,p-p38/p38,p-JNK/JNK,Drp1 and Fis1 significantly increased(P<0.01).Compared with the high glucose group,the cell viability significantly increased in Dihong Prescription medium-and high-dosage groups(P<0.01),the apoptosis rate significantly decreased(P<0.01),the content of ROS significantly decreased(P<0.01),and the protein expressions of p-ERK1/2/ERK1/2,p-p38/p38,p-JNK/JNK,Drp1 and Fis1 significantly decreased(P<0.01).Transmission electron microscopy showed that large number of autophagosomes,autophagosomes,mitochondria vacuolation and cristae fusion were observed in the high glucose group;the mitochondrial damage caused by high glucose was alleviated in Dihong Prescription low-,medium-and high-dosage groups.Conclusion Dihong Prescription can improve oxidative stress injury in glomerular mesangial cells of mice cultured with high glucose,and its mechanism may be related to the regulation of mitochondrial function and miR-133b/DUSP1/MAPK pathway.

Objective:To analyze the characteristics of CD4 + T cell subsets and cytokines in patients with mixed connective tissue disease (MCTD) and the correlation of MCTD disease activity, laboratory data, and clinical symptoms with cytokines. Methods:A total of 48 MCTD patients (including 24 newly diagnosed patients and 24 treated patients) were enrolled from the Department of Rheumatology and Immunology, the Second Hospital of Shanxi Medical University from 2018 to 2021. Meanwhile, 49 healthy subjects who underwent physical examination were recruited (healthy control group). The absolute counts of CD4 + T cell subsets in peripheral blood samples were analyzed by flow cytometry. The levels of serum cytokines were detected by flow bead array. Analysis of variance and Mann-Whitney U test were used to compare the differences between groups. Pearson or Spearman correlation analysis was used for correlation analysis. Logistic regression analysis was used to analyze related factors. The receiver operating characteristic curve was used to detect the best cut-off value and effectiveness. Results:The absolute counts of Th1 ( P<0.01), Th2 ( P<0.01) and Treg cells ( P<0.01) in the newly diagnosed MCTD patients and the treated MCTD patients were lower than those in the healthy subjects. The levels of cytokines (IL-2, IL-4, IL-6, IL-10, IL-17, IFN-γ, and TNF-α) in the two MCTD groups were higher than those in the healthy control group ( P<0.01). Further analysis revealed that the cardiac enzymes in MCTD patients included creatine kinase, creatine kinase-MB, aspartate aminotransferase, α-hydroxybutyrate dehydrogenase, and lactate dehydrogenase were positively correlated with cytokines ( P<0.05). In addition, it was found that IL-2 was positively correlated with erythrocyte sedimentation rate ( r=0.477, P<0.001), but it was negatively correlated with complement C3 ( r=-0.546, P=0.002) and complement C4 ( r=-0.422, P=0.02). IL-10 was correlated with the myositis symptoms in MCTD patients and the area under the receiver operator characteristic curve was 0.745 (95% CI: 0.576-0.915, P<0.05). Conclusions:This study provides insights into the unique immunological characteristics of CD4 + T lymphocyte subsets and cytokines in patients with MCTD, and also reveals a close correlation between cytokines and cardiac enzymes in MCTD patients. IL-2 has been shown to be associated with disease activity in MCTD patients. The level of IL-10 may be related to the occurrence of myositis symptoms in MCTD.

Systemic lupus erythematosus (SLE) shows a significant gender difference. In addition to X chromosome inactivation (XCI) abnormalities, which may lead to the gender difference in SLE, studies have found that estrogen plays a key role in regulating Th17/Treg immune balance in SLE. Estrogen indirectly affects the quantity and function of Th17 and Treg cells by acting on B cells. In this process, the mutual influence and the interaction between B and T cells promote the development of SLE. Recent studies have reported gender differences in intestinal microbiota, which may lead to sex-dependent genetic susceptibility and epigenetic changes in autoimmune diseases represented by SLE. The interaction between estrogen and intestinal microbiota in SLE affects the immune balance of Th17/Treg cells. This paper mainly reviews the way estrogen works, and the mechanisms by which estrogen regulates Th17/Treg immune balance and the interaction between B and T cells in SLE, hoping to provide new targets for new therapeutic strategies such as selective estrogen receptor modulators.

Cyclic GMP-AMP synthase (cGAS)-stimulator of interferon gene (STING) pathway, as an important part of the innate immune system, is the main pathway for cytoplasmic DNA recognition and cGAS can be triggered by a variety of cytoplasmic dsDNA. This pathway has become an important bridge connecting autoimmunity, aseptic inflammatory response and cell aging. In recent years, cGAS-STING pathway has attracted increasing attention in autoimmune diseases. In rheumatoid arthritis (RA), neutrophil extracellular traps (NETs) induce typeⅠ interferon response and accelerate the production of anti-citrullinated peptide antibody (ACPA) through the cGAS-STING pathway. In addition, the cGAS-STING pathway also participates in synovitis, bone destruction and RA progression by promoting the proliferation and activation of fibroblast-like synovitis cells and the polarization of M1 macrophages. Inhibition of the cGAS-STING pathway or its downstream signaling pathway can reduce synovial inflammation in RA, suggesting that cGAS-STING pathway may be a potential therapeutic target for RA.