Objective To investigate the effects of extracellular vesicles(EVs)derived from synovial fluid of rheumatoid arthritis(RA)patients on angiogenesis of human umbilical vein endothelial cells,and to preliminarily explore the underlying mechanisms.Methods Synovial fluid samples of knee joint were collected from 20 patients with RA and 20 patients with osteoarthritis(OA)in this study.EVs were purified using ultracentrifugation.The morphology and size of EVs were observed by transmission electron microscopy and nanoparticle tracking analysis.CD9,CD63,cytochrome c(Cyt-c),vascular endothelial growth factor(VEGF),lysyl oxidase(LOX),matrix metalloproteinase 2(MMP2),tumour necrosis factor alpha(TNF-α),transforming growth factor beta1(TGF-β1)in EVs were detected using Western blot.Human umbilical vein endothelial cells(HUVEC)were treated with the EVs.The growth,migration and angiogenesis of HUVEC were observed by CCK8 assay,TranswellTM chamber assay,scratch test and matrigel angiogenesis assay,respectively.The effect of EVs on the PI3K/AKT pathway in HUVEC was assessed using Western blot.Results Both EVs from RA synovial fluid(RA-EVs)and OA synovial fluid(OA-EVs)were cup-shaped,mainly between 30-400 nm in diameter,expressing CD63 and CD9,but not Cyt-c.RA-EVs carried more VEGF,LOX,MMP2,TNF-α and TGF-β1 than OA-EVs.Compared with the OA-EVs intervention,RA-EVs significantly promoted the proliferation,migration,and angiogenesis of HUVECs,as well as upregulated PI3K/AKT phosphorylation.The inhibitor of PI3K suppressed angiogenesis induced by EVs.Conclusion EVs in synovial fluid of RA carried more cytokines and enzymes that related angiogenesis and inflammation.These EVs exert their pro-angiogenic effects by activating the PI3K/AKT pathway,then contributing to the pathological progression of RA.

Objective To explore the effects of oleanolic acid on NF-κB/caspase-9 signaling pathway in kidneys of rat with diabetic nephropathy.Methods The diabetic nephropathy model rats were established,and the model rats were randomly divided intooleanolic acid low,medium and high dose groups,metformin group,and model group,with another 12 healthy SD rats as control group.The levels of blood glucose were measured at weeks 1,2 and 3 post-drug administration,and blood lipid and 24h urine urinary microalbumin(UMA)were measured after entire drug administration.Furthermore,we also detected the renal histopathology of rats,apoptosis of renal tubular,glomerular cells,tumor necrosis factor-α(TNF-α),interleukin-6(IL-6)and protein expression of NF-κB/caspase-9 signaling pathway.Results Compared with the control group,the model group demonstrated higher levels of blood glucose at weeks 1,2 and 3,blood lipid,UMA,TNF-α and IL-6,higher proportion of apoptotic renal tubular and glomerular,and higher expressions of caspase-9 protein and p-NF-κB p65/NF-κB p65(P＜0.05).Compared with the model group,the pathological damage of renal tissue in the metformin group and oleanolic acid low,middle and high dose groups were alleviated,the levels of blood glucose at weeks 1,2 and 3,blood lipid,UMA,TNF-α and IL-6,the proportion of apoptotic renal tubular and glomerular,and the expressions of caspase-9 proteins and p-NF-κB p65/NF-κB p65 were decreased,and these indexes in oleanolic acid groups showed a dose-dependent manner(P＜0.05).Conclusion Oleanolic acid can improve the metabolism of glucose and lipid,reduce the pathological damage of renal tissue,inhibit the apoptosis of renal tubular and glomerular,and inhibit the NF-κB/caspase-9 pathway in diabetic nephropathy rats.

Objective To investigate the expression of dihydrolipoamide S-acetyltransferase(DLAT)in hepatocellular carcinoma(HCC)tissues and its impact on immunotherapy efficacy,aiming to identify a new biomarker for prognosis assessment and immunotherapy response prediction.Methods Bioinformatics methods were used to analyze the transcriptomic data,clinical pathological characteristics and survival information of HCC patients from TCGA database.The expression of DLAT in HCC and its correlation with clinical features were evaluated,along with its relationship with immune infiltration,immune checkpoint-related genes,and immunotherapy response.Results DLAT was highly expressed in HCC tissues and associated with poor prognosis(HR=1.63,P=0.006).The proportion of R1&R2 residual tumors were significantly higher in the DLAT high-expression group(4.1％vs 1.2％,P=0.011).Immune infiltration analysis revealed that high DLAT expression was negatively correlated with Th 17,DC cells,B cells and T cells,while positively correlated with T helper cells,Tcm and Tem cells.Furthermore,DLAT expression showed significant positive correlations with key immune checkpoint genes(PDCD1LG2,HAVCR2,TIGIT,and PVR),and TIDE algorithm predicted poor response to immune checkpoint inhibitor therapy in the DLAT high-expression group.Conclusion High expression of DLAT in HCC tissues is a risk factor for poor prognosis and may serve as a potential biomarker for prognostic assessment and immunotherapy response prediction in HCC patients.

Objective To analyze the clinical and immunological characteristics of a rare case of CHARGE syndrome,we summarize the genotype and phenotype in the Chinese patient population,and explore the underlying immunopathogenic mechanisms.Methods Clinical data from a pediatric patient with CHARGE syndrome were collected and analyzed.A comprehensive analysis of the Chinese patient population was conducted.Gene analysis and immunological characterization were performed using flow cytometry,deep sequencing,and quantitative PCR.Results The proband was a premature female infant whose primary clinical manifestations included congenital heart disease,recurrent respiratory infections,respiratory failure,airway dysplasia,hearing impairment,and bilateral choroidal coloboma.Whole-exome sequencing revealed a de novo heterozygous nonsense mutation in the CHD7 gene,c.5122C＞T(p.Gln1708Ter),classified as pathogenic according to ACMG criteria.Immunological studies indicated impaired thymic output of T cells,significant alterations in the number and proportion of CD8+T cell subsets,increased apoptosis,and defective activation and production of key effector cytokines such as IFN-γ by CD8+T cells.However,no significant abnormalities were observed in peripheral lymphocyte proliferation.Conclusion CHARGE syndrome is a rare autosomal dominant genetic disorder primarily caused by mutations in the CHD7 gene.The main clinical features include ocular defects,cardiac disease,choanal atresia/cleft lip and palate,growth retardation,gonadal hypoplasia,and ear anomalies.This case study suggests that CHARGE syndrome is associated with abnormalities in the development,apoptosis,and effector functions of immune cells.

Objective To determine the pathogenicity of a novel mutation(c.109_111del)in DKC1 gene of an adult patient,and to analyze the clinical phenotype,immunophenotype and telomere length,so as to provide clues for early clinical identification and diagnosis.Methods The clinical data and peripheral blood samples of the patient were collected for genetic testing and family analysis.The lymphocyte subsets of the patient were detected by Flow cytometry,and the telomere length of the patient and healthy controls were detected by Flow-FISH.Results The main clinical manifestations of the patient were mucocutaneous triad,bone marrow failure and infection.The telomere length of lymphocytes in the patient was significantly shorter than that of healthy controls of the same age,and the absolute value and percentage of lymphocyte subsets were abnormal.Conclusion The clinical manifestations of DC patients are diverse.Flow-FISH detection of telomere length is helpful for early diagnosis of DC patients.

Calprotectin,called MRP8/14 or S100A8/A9,is a heterodimer formed by two proteins,S100A8 and S100A9.It is mainly produced by activated monocytes and neutrophils in circulation and in inflamed tissues,thereby promoting the inflammatory response in vivo and in vitro as an important innate immune proinflammatory factor.Calprotectin is elevated in many inflammatory conditions and can be detected in blood,urine,faeces,cerebrospinal fluid,salivary synovial fluid and colon biopsy tissue.Due to its immunological features,calprotectin plays an important role in tumor,autoimmune disease,infection and other diseases.In recent years,more and more studies have been conducted on the relationship between calprotectin and rheumatic diseases,and the mechanism has also been gradually deepened.Many clinical trials have shown that calprotectin is correlated with rheumatic diseases in terms of reflecting disease activity or predicting prognosis.In this review,we will systematically summarize the relationship between calprotectin and some rheumatic diseases and its clinical progresses.

Objective The aim of this study was to clarify the role and mechanism of Pseudomonas aeruginosa vaccine subunit OprI in the fusion protein vaccine rePO(PcrV-OprI).Methods The in vitro stability of rePO,PcrV and OprI at 4 ℃,25 ℃,and 37 ℃ was examined.After immunizing mice with rePO,OprI and PcrV,respectively,the specific antibody potency in serum and the proportion of cells secreting IFN-γ and IL-4 in the spleen were examined;Additionally,detection of the levels of protein uptake by DC2.4 cells in vitro using laser confocal microscopy and flow cytometry,and their ability to promote the maturation of mouse bone marrow-derived dendritic cells(BMDC).Results The heat stability of fusion protein rePO was significantly better than that of PcrV.The induced anti-PcrV IgG and anti-OprI IgG potency of rePO was significantly higher than that of monomeric PcrV and OprI.Additionally,the number of cells secreting IFN-γ and IL-4 induced by immunization with rePO was significantly higher than that of PcrV and OprI.The uptake rate of fusion protein rePO by DC2.4 cells was significantly higher than that of PcrV and OprI.Furthermore,rePO promoted the maturation of mouse BMDC more effectively than PcrV and OprI.Conclusion OprI in the fusion protein rePO can significantly improve its thermal stability and immunogenicity,which lays the foundation for the successful development of Pseudomonas aeruginosa vaccine.

Objective To explore the effects of hydroxytyrosol-mediated macrophage polarization on burn wound healing in rats.Methods SD rats were randomly divided into burn group,hydroxytyrosol group,hydroxytyrosol+ov-NC group and hydroxytyrosol+ov-S100A9 group,with 10 rats in each group.Burn model was established by metal weight burn method.Wound healing rate was calculated;HE staining was used to observe the pathological changes of wound skin tissue in rats;immunofluorescence was used to detect the ratios of M1 and M2 macrophages in the wound skin of rats;ELISA was used to detect the levels of macrophage inflammatory factors in the wound skin of rats;Western blot was used to detect the expression of S100A9/TLR4/NF-κB signal pathway in wound skin of rats.Results Compared with the burn group,hydroxytyrosol group and hydroxytyrosol+ov-NC group demonstrated higher wound healing ability and alleviated pathological damage,lower levels of M1 type macrophages and related factors,higher levels of M2 type macrophages and related factors,and lower expression of S100A9,TLR4 and p-NF-κB p65 proteins.Compared with hydroxytyrosol group and hydroxytyrosol+ov-NC group,S100A9 overexpression reduced the wound healing ability and aggravated the pathological damage,increased M1 type macrophages and related factors,decreased M2 type macrophages and related factors,and increased the expression of S100A9,TLR4 and p-NF-κB p65 proteins in hydroxytyrosol+ov-S100A9 group.Conclusion Hydroxytyrosol promotes wound healing in burn rats by down-regulating S100A9,and its mechanism may be related to inhibiting the activation of TLR4/NF-κB signaling pathway and improving the imbalance of M1/M2 macrophages.

Cancer is one of the major diseases of high morbidity and mortality worldwide,and its therapeutic approaches are facing great challenges.Immunotherapy,especially the activation of innate immunity represented by the cGAS-STING signaling pathway,is the current research hotspot in tumor immunotherapy.Activation of innate immune response by gas therapy is the latest development in tumor therapeutic approaches,especially the use of gas signaling molecules(NOx CO,H2S and SO2)to activate the cGAS-STING signaling pathway to induce intrinsic immunity of the organism,which leads to anti-tumor immunotherapy.Although intrinsic immunity activated by gas signaling molecules plays an important role in tumor immunotherapy,few reviews have been reported on its association with the cGAS-STING signaling mechanism.In this paper,we will comprehensively describe how gas signaling molecules damage the mitochondrial matrix and DNA damage through oxidative/nitrosative stress,thereby activating the cGAS-STING signaling pathway and triggering the innate immune cascade,aiming to summarize the process of activation of anti-tumor immune effects by gas signaling molecules,and to provide more references for the gas therapies in the future anti-tumor immunity research.

The occurrence and development of a variety of retinal diseases are related to inflammatory responses,and various inflammatory cells play an important role in retinal damage,which can lead to vision impairment,vision loss,and blindness.Microglia are resident immune cells in the retina,distributed in the inner layer of the retina.They mainly maintain the normal homeostasis of the retina,regulate the apoptosis of neurons,and play an immune surveillance role in the retina.Under inflammatory stimulation,microglia in the retina are activated,secrete a variety of inflammatory factors,engulf neurons and photoreceptors,and destroy the blood-retinal barrier,aggravating retinal damage.This article reviews the physiological function of microglia and the changes in microglia under the inflammatory effects of various retinal diseases.It also discusses how to inhibit microglia from damaging the retina and promote microglia to control retinal inflammation,thereby providing a basis for the clinical treatment of various retinal diseases.