CXCL10 Gene Research Articles

Exploration of the Potential Mechanism of the Common Differentially Expressed Genes in Psoriasis and Atopic Dermatitis.

Backgrounds Psoriasis and atopic dermatitis are two common chronic inflammatory skin diseases that enormously deteriorate the psycho-physical and socio-economic condition of the patients. Although differential immune responses have been found to operate in the pathomechanisms of atopic dermatitis and psoriasis, the epidermal keratinocytes are the major targets in both diseases, and sometimes, they show similar clinical presentations. The skin barrier, itching, and inflammation are current and future treatment targets for both of them, but the relevant shared mechanisms of the two diseases are far from understood. Methods The differential analyses of GSE14905 (psoriasis) and GSE32924 (atopic dermatitis) deposited in GEO database were conducted and obtained their differential expressed genes. Moreover, PPI, functional modules, GO, and KEGG enrichment analyses were used for the further analysis. The mouse models of psoriasis and atopic dermatitis were established, and then, RT-qPCR and Western blotting assay were performed to check the abundant changes of hub genes. Results There are 732 differentially expressed genes in psoriasis versus nonlesional skin samples. Besides, 611 differentially expressed genes were identified in atopic dermatitis versus nonlesional skin data sets. Based on these differentially expressed genes, we predicted their joint and individual protein-protein interaction networks and functional modules in both psoriasis and atopic dermatitis. Through the PPI network of genes, we calculated the hub nodes and do the GO and KEGG enrichment analysis of overlapped genes of psoriasis and atopic dermatitis, which suggested there were some terms like “positive regulation of interleukin-12 production,” “centromeric region,” and “TNF signaling pathway.” Conclusion We constructed the predicted PPI networks and functional modules related to psoriasis and atopic dermatitis and distinguished the key candidate target genes CXCL8, STAT1, and MMP9 in the diagnosis and therapy of similar pathogenesis.

Suppression of CX3CL1 by miR-497-5p inhibits cell growth and invasion through inactivating the ERK/AKT pathway in NSCLC cells

ABSTRACT Non-small cell lung cancer (NSCLC) is the most common lung cancer with a highest mortality rate. MiR-497-5p has been reported as tumor suppressor in many cancers, but the role and mechanism of miR-497-5p in regulating NSCLC progression are still largely unknown in vitro and in vivo. Here, miR-497-5p was significantly downregulated in human NSCLC tissues and cell lines, compared with matched adjacent tissues and normal lung epithelial cell line. Then, miR-497-5p mimic and inhibitor were, respectively, transfected into human NSCLC cells A549 and H460, CCK-8 assay, transwell assay, and flow cytometry were used to detect the capacities of cell proliferation, invasion and apoptosis. MiR-497-5p negatively regulated proliferation and invasion of NSCLC cancer cells. MiR-497-5p was demonstrated to directly bound to 3’-UTR of CX3CL1 mRNA and post-transcriptionally suppressed its expression thus inactivating its downstream oncogenic pathway ERK/AKT. Moreover, transfection with short hairpin RNA (shRNA) against CX3CL1 decreased capacity of cell proliferation and invasion and promoted cell apoptosis in NSCLC cells. In addition, ERK inhibitor U0126 attenuated the promotion effect of miR-497-5p inhibitor on activation of ERK/AKT and cell proliferation and migration. Finally, overexpression of miR-497-5p substantially suppressed activation of the ERK/AKT pathway and tumor growth in tumor-bearing mice in vivo. Taken together, our findings showed that miR-497-5p is downregulated in human NSCLC tissues and cell lines, and it inhibited tumor growth and cell invasion by targeting CX3CL1 gene to inactivate the ERK/AKT pathway in NSCLC cells.

Airway epithelial interferon response to SARS-CoV-2 is inferior to rhinovirus and heterologous rhinovirus infection suppresses SARS-CoV-2 replication

Common alphacoronaviruses and human rhinoviruses (HRV) induce type I and III interferon (IFN) responses important to limiting viral replication in the airway epithelium. In contrast, highly pathogenic betacoronaviruses including SARS-CoV-2 may evade or antagonize RNA-induced IFN I/III responses. In airway epithelial cells (AECs) from children and older adults we compared IFN I/III responses to SARS-CoV-2 and HRV-16, and assessed whether pre-infection with HRV-16, or pretreatment with recombinant IFN-β or IFN-λ, modified SARS-CoV-2 replication. Bronchial AECs from children (ages 6–18 years) and older adults (ages 60–75 years) were differentiated ex vivo to generate organotypic cultures. In a biosafety level 3 (BSL-3) facility, cultures were infected with SARS-CoV-2 or HRV-16, and RNA and protein was harvested from cell lysates 96 h. following infection and supernatant was collected 48 and 96 h. following infection. In additional experiments cultures were pre-infected with HRV-16, or pre-treated with recombinant IFN-β1 or IFN-λ2 before SARS-CoV-2 infection. In a subset of experiments a range of infectious concentrations of HRV-16, SARS-CoV-2 WA-01, SARS-CoV-2 Delta variant, and SARS-CoV-2 Omicron variant were studied. Despite significant between-donor heterogeneity SARS-CoV-2 replicated 100 times more efficiently than HRV-16. IFNB1, INFL2, and CXCL10 gene expression and protein production following HRV-16 infection was significantly greater than following SARS-CoV-2. IFN gene expression and protein production were inversely correlated with SARS-CoV-2 replication. Treatment of cultures with recombinant IFNβ1 or IFNλ2, or pre-infection of cultures with HRV-16, markedly reduced SARS-CoV-2 replication. In addition to marked between-donor heterogeneity in IFN responses and viral replication, SARS-CoV-2 (WA-01, Delta, and Omicron variants) elicits a less robust IFN response in primary AEC cultures than does rhinovirus, and heterologous rhinovirus infection, or treatment with recombinant IFN-β1 or IFN-λ2, reduces SARS-CoV-2 replication, although to a lesser degree for the Delta and Omicron variants.

Characterizing the Inflammatory Microenvironment in K14-HPV16 Transgenic Mice: Mast Cell Infiltration and MicroRNA Expression.

Simple SummaryK14-HPV16 transgenic mice have proved to be a useful model to study the carcinogenic cascade induced by HPV16, the tumor microenvironment and also the epigenetic and genetic factors associated with this type of malignancy. The aim of our study was to evaluate the infiltration of mast cells in two cutaneous regions with different severity of the lesions and to identify potential microRNAs that may regulate mast cell infiltration in this model. We were able to confirm that increased mast cell infiltration is associated with progression of HPV-induced lesions, and that miR-223-3p and miR-125b-5p might be assisting this process via the regulation of mast cell chemotactic proteins.High-risk human papillomavirus (HPV) is the etiologic agent of several types of cancer. Mast cells’ role as either a driving or opposing force for cancer progression remains controversial. MicroRNAs are dysregulated in several HPV-induced cancers, and can influence mast cell biology. The aim of this study was to evaluate mast cell infiltration and to identify microRNAs potentially regulating this process. Transgenic male mice (K14-HPV16; HPV+) and matched wild-type mice (HPV−) received 7,12-Dimethylbenz[a]anthracene (DMBA) (or vehicle) over 17 weeks. Following euthanasia, chest skin and ear tissue samples were collected. Mast cell infiltration was evaluated by immunohistochemistry. MicroRNAs associated with mast cell infiltration were identified using bioinformatic tools. MicroRNA and mRNA relative expression was evaluated by RT-qPCR. Immunohistochemistry showed increased mast cell infiltration in HPV+ mice (p < 0.001). DMBA did not have any statistically significant influence on this distribution. Ear tissue of HPV+ mice showed increased mast cell infiltration (p < 0.01) when compared with chest skin samples. Additionally, reduced relative expression of miR-125b-5p (p = 0.008, 2−ΔΔCt = 2.09) and miR-223-3p (p = 0.013, 2−ΔΔCt = 4.42) seems to be associated with mast cell infiltration and increased expression of target gene Cxcl10. These results indicate that HPV16 may increase mast cell infiltration by down-regulating miR-223-3p and miR-125b-5p.

Evaluation of Neutrophilic Receptors; CXCL8 and CXCR2 in Patients with Chronic Periodontitis Compared to Healthy Subjects by Real Time PCR Method

Objective: This study aimed to evaluate the levels of CXCL8 and its receptor (CXCR2) in gingival tissue neutrophils from patients with chronic periodontitis compared to periodontal healthy subjects using Real Time PCR method. Methods: The test group consisted of 21 patients with chronic periodontitis and the control group consisted of 18 healthy individuals. In both groups, gingival tissue samples were obtained during periodontal surgery.CXCL8 and CXCR2 RNA in tissue samples were examined by PCR method, and then the levels of genes expression were measured. Mann-Whitney U nonparametric test was used for statistical analysis. Results: CXCL8 gene expression in the gingival tissue of the test group with chronic periodontitis was significantly higher than the control group (p=0.028). CXCR2 gene expression in the gingival tissue of the test group with chronic periodontitis was significantly lower than the control group (p=0.043). In both test and control groups, there was a negative correlation between CXCL8 and CXCR2 gene expression. This correlation was statistically significant in the test group (p=0.001), but there was no significant correlation in the control group (p=0.431). Conclusion: The results of this present study suggested that the level of gene expression for CXCL8 was greater in patients with chronic periodontitis and CXCR2 was greater in healthy individuals. Although in people with chronic periodontitis, CXCR2 decreases slightly as CXCL8 levels increase.

Role of chemokines in early pregnancy loss.

The present study aimed to compare decidual protein levels and gene expression levels of chemokines between patients with early pregnancy loss and those with voluntary abortion. A total of 15 patients between 6 and 10 gestational weeks, who presented with negative fetal heartbeat to the obstetrics and gynecology outpatient clinics of Gaziosmanpasa Hospital (Yeni Yuzyil University, Istanbul, Turkey) and who had no additional systemic disease and 13 patients between 6 and 10 gestational weeks, who presented with positive fetal heartbeat for voluntary abortion were included in the present study. CX3CL1, CCL17, CXCR4, chemokine ligand 12 (CXCL12) and intercellular adhesion molecule (ICAM)5 protein expression levels were determined by ELISA and gene expression levels by reverse transcription-quantitative PCR in fresh materials recovered after therapeutic curettage. CX3CL1, CCL17, CXCR4, CXCL12 protein levels were significantly higher and ICAM protein level was significantly lower in pregrant women with missed abortion compared with those with voluntary abortion. While the amount of increase in mean CX3CL1, CCL17, CXCR4 and CXCL12 gene expression levels in the tissues of pregnant women with missed abortion was statistically higher than the pregnant women who underwent voluntary abortion, the amount of increase in ICAM5 gene expression was found to be lower (P<0.001) in those with missed abortion. In conclusion, the findings of the present study suggested that CCL17, CX3CL1, CXCL12, CXCR4 and ICAM5 may be associated with missed abortion and may play an important role in placental invasion and the continuation of pregnancy.

Blockade of the CXCR3/CXCL10 axis ameliorates inflammation caused by immunoproteasome dysfunction.

Immunoproteasomes regulate the degradation of ubiquitin-coupled proteins and generate peptides that are preferentially presented by MHC class I. Mutations in immunoproteasome subunits lead to immunoproteasome dysfunction, which causes proteasome-associated autoinflammatory syndromes (PRAAS) characterized by nodular erythema and partial lipodystrophy. It remains unclear, however, how immunoproteasome dysfunction leads to inflammatory symptoms. Here, we established mice harboring a mutation in Psmb8 (Psmb8-KI mice) and addressed this question. Psmb8-KI mice showed higher susceptibility to imiquimod-induced skin inflammation (IMS). Blockade of IL-6 or TNF-α partially suppressed IMS in both control and Psmb8-KI mice, but there was still more residual inflammation in the Psmb8-KI mice than in the control mice. DNA microarray analysis showed that treatment of J774 cells with proteasome inhibitors increased the expression of the Cxcl9 and Cxcl10 genes. Deficiency in Cxcr3, the gene encoding the receptor of CXCL9 and CXCL10, in control mice did not change IMS susceptibility, while deficiency in Cxcr3 in Psmb8-KI mice ameliorated IMS. Taken together, these findings demonstrate that this mutation in Psmb8 leads to hyperactivation of the CXCR3 pathway, which is responsible for the increased susceptibility of Psmb8-KI mice to IMS. These data suggest the CXCR3/CXCL10 axis as a new molecular target for treating PRAAS.

Butyrate Prevents Induction of CXCL10 and Non-Canonical IRF9 Expression by Activated Human Intestinal Epithelial Cells via HDAC Inhibition.

Non-communicable diseases are increasing and have an underlying low-grade inflammation in common, which may affect gut health. To maintain intestinal homeostasis, unwanted epithelial activation needs to be avoided. This study compared the efficacy of butyrate, propionate and acetate to suppress IFN-γ+/−TNF-α induced intestinal epithelial activation in association with their HDAC inhibitory capacity, while studying the canonical and non-canonical STAT1 pathway. HT-29 were activated with IFN-γ+/−TNF-α and treated with short chain fatty acids (SCFAs) or histone deacetylase (HDAC) inhibitors. CXCL10 release and protein and mRNA expression of proteins involved in the STAT1 pathway were determined. All SCFAs dose-dependently inhibited CXCL10 release of the cells after activation with IFN-γ or IFN-γ+TNF-α. Butyrate was the most effective, completely preventing CXCL10 induction. Butyrate did not affect phosphorylated STAT1, nor phosphorylated NFκB p65, but inhibited IRF9 and phosphorylated JAK2 protein expression in activated cells. Additionally, butyrate inhibited CXCL10, SOCS1, JAK2 and IRF9 mRNA in activated cells. The effect of butyrate was mimicked by class I HDAC inhibitors and a general HDAC inhibitor Trichostatin A. Butyrate is the most potent inhibitor of CXCL10 release compared to other SCFAs and acts via HDAC inhibition. This causes downregulation of CXCL10, JAK2 and IRF9 genes, resulting in a decreased IRF9 protein expression which inhibits the non-canonical pathway and CXCL10 transcription.

Lack of Association between IFN-γ, CXCL10 and TGF-β1 Gene Polymorphisms and Liver Complication in HIV-infected Thais.

Objective:Chronic liver disease has become a leading cause of illness and death in people living with HIV and the production of the cytokines IFN-γ and TGF-β1, and chemokine CXCL10 during chronic inflammation contributes to liver disease progression in HIV patients under long-term anti-retroviral therapy. This study aimed to examine association of IFN-γ +874T/A, CXCL10 G-201A and C-1596T, and TGF-β1 -509C/T single nucleotide polymorphisms (SNPs) with liver complications in the HIV-infected Thais. Methods:A cross-sectional study was conducted in 200 Thai HIV patients who were evaluated for transaminitis and significant liver fibrosis by fibrosis-4 score (FIB-4), and genotypes for IFN-γ +874T/A, CXCL10 G-201A and C-1596T, and TGF-β1 -509C/T SNPs using PCR-based methods. Result:There were high rates of transaminitis (30.1%) and significant liver fibrosis assessed by FIB-4 score > 1.45 (18.8%) in this group of patients, mostly under anti-retroviral therapy (73.0%). The genotypes and alleles of IFN-γ +874T/A, CXCL10 G-201A and C-1596T, and TGF-β1 -509C/T SNPs were not associated with either transaminitis or FIB-4 score > 1.45 (p > 005). Logistic regression analysis identified age and gender as risk factors, and CD4+ cell count higher than 350 cells/ul as a protective factor of liver fibrosis in this study group.Conclusion:The IFN-γ +874T/A, CXCL10 G-201A and C-1596T, and TGF-β11 -509C/T SNPs were not significantly associated with liver complication in HIV-infected Thais, mostly under ART.

IL1B gene variants, but not TNF, CXCL8, IL6 and IL10, modify the course of cystic fibrosis in Polish patients.

Background: The main aim of this study was to evaluate whether selected polymorphic variants in genes from the inflammatory pathway can be predictors of pulmonary or digestive manifestation of cystic fibrosis, as well as of severity of lung disease. Materials and methods: Using pyrosequencing and sequencing we have genotyped 12 variants in TNF (rs361525, rs1800629), CXCL8 (rs4073, rs2227306, rs2227307, rs188378669), IL1B (rs16944, rs1143634, rs1142639, rs1143627), IL6 (rs1800795) and IL10 (rs1800896) genes in a cohort of 55 Polish patients with diagnosed cystic fibrosis and controls. In our study group, a pulmonary manifestation of disease revealed 44 of subjects (80%), and digestive symptoms dominated in 11 (20%) of analyzed individuals. Severe lung dysfunction has occurred in 20 (36.4%) of patients. Results: We proved, that two promoter variants of IL1B, rs1143627 (c.-118G &gt; A) and rs16944 (c.-598T &gt; C) are presented significantly more often in patients with severe character of lung disease compared to mild (82.5% vs. 62.8%, p-value 0.030, and 87.5% vs. 64.3%, p-value 0.008, respectively) in cystic fibrosis course. Haplotype AC formed by both changes had also a higher frequency (80%) in patients with severe course compared to the mild character (61.4%) of disease. However, the frequency of promoter variant TNF c.-308C &gt; T (rs1800629) was presented at a significantly lower level in the patient’s group compared to healthy controls (2.7% vs. 15%, p-value 0.001). Furthermore, the presence of methicillin-resistant Staphylococcus aureus significantly correlated with the lower FEV1% in patients (p-value 0.01). Conclusions: Genetic variants, rs1143627 and rs16944, of IL1B are promising candidates as predictors of the severe character of lung disease in Polish patients with cystic fibrosis.

Regulatory mechanism of downregulation of SOD1 expression on cardiomyocyte function.

Many diseases are clinically related to oxidative stress. Obstructive sleep apnea (OSA) is a common disease with oxidative stressin clinical practice, which is mostly associated with cardio-cerebrovascular diseases. It has been shown that the level of oxidative stress increases and the level of antioxidant copper zinc superoxide dismutase (SOD1) decreases in intermittent hypoxia (IH). SOD1 is one of the key antioxidant enzymes in organisms, and it can also be used as a signal transmission controller. Its abnormal expression further affects organ functions, but the specific mechanism is not yet fully clear. We downregulated the SOD1 gene in H9C2 cell line, using high-throughput RNA sequencing (RNA-seq) to find differentially expressed genes (DEGs) related to cardiomyocyte function by using GO and KEGG databases to annotate, enrich and analyze the metabolic pathways of DEGs. Through the analysis of these functional gene changes, we can understand the regulation of SOD1 downregulation on cardiomyocyte function. The results found 213 DEGs, of which 135 genes were upregulated and 78 genes were downregulated. The upregulated DEGs were mainly enriched in biological processes such as transcriptional regulation and metabolism. The expression levels of EGR1 and NR1D1 exceeded 1 in the samples. EGR1 was reported to be involved in oxidative stress and cardiac hypertrophy, and NR1D1played an important regulatory role in regulating inflammatory responses and reducing ROS production. The biological processes involved in downregulated DEGs mainly involve metabolism and redox processes. Among them, SCD1 and CCL2 genes were highly expressed among the genes involved in the redox process involved in SOD1. SCD1 is an important player in the regulation of cardiometabolic processes; downregulation of CCL2 reduces atherosclerosis. We found that the TNF signaling pathway, NOD-like receptor signaling pathway, and chemokine signaling pathway, which were enriched in KEGG analysis, were all associated with inflammation, and the CXCL1 and CCL7 genes are all related to inflammation. The gene and signaling pathways involved in oxidative stress and inflammatory response process regulated by SOD1 were demonstrated. SOD1 may affect the function of the heart by affecting myocardial contraction, inflammation, lipid metabolism, and other pathways. It is inferred that they may also play a role in the process of OSA-related myocardial injury, which is worthy of attention and further study.

Transcriptional activity analysis of the immune response genes in the peripheral blood of patients with comorbid acute urticaria and lyme borreliosis

Acute urticaria (AU) and Lyme borreliosis (LB) are known to alter the transcriptional profile of blood cells. The nature of changes in the transcriptional activity of genes of the innate and adaptive immune system in the peripheral blood in patients with comorbid AU and LB is unknown. In our study, we applied a pathway-specific PCR array (Human Innate &amp; Adaptive Immune Responses RT2 Profiler PCR Array, QIAGEN, Germany) to detect and verify the innate and adaptive immune responses of pathway-focused genes expression in the blood of patients with a comorbid course of these pathologies. It was found that in patients with comorbid AU and Lyme disease, transcriptional induction of a number of genes of the innate immune system in PBMC was observed, in particular: TLR2, NOD2, NLRP3, APCS, complement component 3 (C3), CD14, CD86 compared with patients suffering from acute urticaria. These changes were accompanied by an increased transcriptional activity of systemic proinflammatory cytokines IL1B, IL6, IFNG and its receptor IFNGR1, TNF, ligands, and chemokine receptors CXCL10, CXCR3, CCR5, tyrosine kinase JAK2 and transcription factors STAT1 and TBX21. At the same time, the comorbid course of acute urticaria and Lyme disease led to the repression of the transcriptional activity of the CD80, IL4, and CXCL8 genes. The comorbid course of acute urticaria and Lyme borreliosis is accompanied by activation of the transcriptional activity of genes of the innate immune system and proinflammatory cytokines.

Transcriptome regulation of extracellular vesicles derived from porcine uterine flushing fluids during peri-implantation on endometrial epithelial cells and embryonic trophoblast cells

The extracellular vesicles (EVs) in uterine fluids play a vital role in embryo implantation by mediating intrauterine communication between conceptus and maternal endometrium in pigs. However, the regulatory mechanism of EVs in uterine fluids is largely unclear. In order to understand the effect of EVs in uterine flushing fluids (UFs) during embryo implantation on endometrial epithelial cells (EECs) and embryonic trophoblast cells (PTr2 cells). The UFs-EVs on day 13 of pregnancy (D13) were added to the culture medium of EECs and PTr2 cells. It was found that PKH-67 labeled UFs-EVs could be taken up in EECs and PTr2 cells. Transcriptome sequencing analysis showed that a total of 1793 and 6279 genes were differentially expressed in the EECs and PTr2 cells after the treatment of UFs-EVs on D13, respectively. Among these genes, real-time quantitative PCR (RT-qPCR) results indicated that ID2, ITGA5, CXCL10 and CXCL11 genes were differentially expressed in both EECs and PTr2 cells after treatment. Bioinformatics analysis showed that the differentially expressed (DE) genes in EECs and PTr2 cells after treatment are involved in immune regulation, cell migration, cell adhesion and the secretion and uptake of EVs. Our research offers novel insight into the regulation mechanism of UFs-EVs on D13 in EECs and PTr2 cells.

A Rationale for the Activity of Bone Target Therapy and Tyrosine Kinase Inhibitor Combination in Giant Cell Tumor of Bone and Desmoplastic Fibroma: Translational Evidences

Giant cell tumor of bone (GCTB) and desmoplastic fibroma (DF) are bone sarcomas with intermediate malignant behavior and unpredictable prognosis. These locally aggressive neoplasms exhibit a predilection for the long bone or mandible of young adults, causing a severe bone resorption. In particular, the tumor stromal cells of these lesions are responsible for the recruiting of multinucleated giant cells which ultimately lead to bone disruption. In this regard, the underlying pathological mechanism of osteoclastogenesis processes in GCTB and DF is still poorly understood. Although current therapeutic strategy involves surgery, radiotherapy and chemotherapy, the benefit of the latter is still debated. Thus, in order to shed light on these poorly investigated diseases, we focused on the molecular biology of GCTB and DF. The expression of bone-vicious-cycle- and neoangiogenesis-related genes was investigated. Moreover, combining patient-derived primary cultures with 2D and 3D culture platforms, we investigated the role of denosumab and levantinib in these diseases. The results showed the upregulation of RANK-L, RANK, OPN, CXCR4, RUNX2 and FLT1 and the downregulation of OPG and CXCL12 genes, underlining their involvement and promising role in these neoplasms. Furthermore, in vitro analyses provided evidence for suggesting the combination of denosumab and lenvatinib as a promising therapeutic strategy in GCTB and DF compared to monoregimen chemotherapy. Furthermore, in vivo zebrafish analyses corroborated the obtained data. Finally, the clinical observation of retrospectively enrolled patients confirmed the usefulness of the reported results. In conclusion, here we report for the first time a molecular and pharmacological investigation of GCTB and DF combining the use of translational and clinical data. Taken together, these results represent a starting point for further analyses aimed at improving GCTB and DF management.

Aggregatibacter actinomycetemcomitans Outer Membrane Proteins 29 and 29 Paralogue Induce Evasion of Immune Response.

Aggregatibacter actinomycetemcomitans (Aa) is abundant within the microbial dysbiotic community of some patients with periodontitis. Aa outer membrane protein 29 (OMP29), a member of the OMPA family, mediates the invasion of Aa to gingival epithelial cells (GECs). This study evaluated the effect of OMP29 and its paralogue OMP29par on the response of GECs to Aa. The omp29 or/and omp29par deletion mutants AaΔ29, AaΔ29P, and AaΔ29Δ29P were constructed, and recombinant Aa OMP29His was obtained. Microarray analysis and the evaluation of cxcl-8 gene expression were performed to examine the response of GECs line OBA-09 to Aa and its mutants. The expression of cxcl-8 and its product CXCL-8 was examined in LPS-stimulated OBA-09 cells with Aa OMP29His. Proteomics analysis showed that the deletion of omp29 led to overexpression of both OMP29par and another membrane protein OMP39, the expression of which was further increased in AaΔ29Δ29P. OBA-09 cells challenged with AaΔ29Δ29P exhibited a higher expression of cxcl-8 in comparison to wildtype Aa strain AaD7S or single-deletion mutants AaΔ29 or AaΔ29P. LPS-stimulated OBA-09 cells challenged with Aa OMP29His showed reduced expressions of cxcl-8 and its product CXCL-8. OBA-09 cells challenged with AaΔ29Δ29P in comparison to Aa strain AaD7S resulted in higher expressions of genes involved in apoptosis and inflammatory response such as bcl2, birc3, casp3, c3, ep300, fas, fosb, grb2, il-1α, il-1β, il-6, cxcl-8, nr3c1, prkcq, socs3, and tnfrsf1β and reduced expressions of cd74, crp, faslg, tlr1, and vcam1. The results suggested a novel strategy of Aa, mediated by OMP29 and OMP29par, to evade host immune response by inhibiting CXCL-8 expression and modulating the genes involved in apoptosis and inflammatory response in GECs. Pending further confirmation, the strategy might interfere with the recruitment of neutrophils and dampen the host inflammatory response, leading to a more permissive subgingival niche for bacterial growth.

Pharmacological inhibition of Kv1.3 channel impairs TLR3/4 activation and type I IFN response and confers protection against Listeria monocytogenes infection

Emerging data have demonstrated the critical roles of potassium efflux in the innate immune system. However, the role of potassium efflux in TLR3/4 activation and type I interferon (IFN) responses are not well elucidated. In the present study, we found potassium efflux is essential for TLR3/4 signaling, which mediates the expression of IFN and its inducible gene Cxcl10 and proinflammatory cytokine gene TNF-α. Furthermore, pharmacological inhibition of Kv1.3 channel (PAP-1), but not Kir2.1, KCa3.1 or TWIK2, attenuated TLR3/4 receptor activation in macrophages. Mechanistically, PAP-1 suppressed LPS-induced inflammatory function through marked suppressing the activation of JNK mitogen-activated protein kinase (MAPK) and p65 subunit of nuclear factor-kB (NF-kB). Notably, PAP-1 effectively protected mice against Listeria monocytogenes induced infection. Our findings reveal that potassium efflux mediated by the Kv1.3 channel is essential for TLR3/4 activation and suggest that pharmacological inhibition of Kv1.3 may help to treat type I IFN related autoimmune diseases and bacterial infections.

Single-Cell Transcriptome Analysis of Chronic Antibody-Mediated Rejection After Renal Transplantation.

Renal transplantation is currently the most effective treatment for end-stage renal disease. However, chronic antibody-mediated rejection (cABMR) remains a serious obstacle for the long-term survival of patients with renal transplantation and a problem to be solved. At present, the role and mechanism underlying immune factors such as T- and B- cell subsets in cABMR after renal transplantation remain unclear. In this study, single-cell RNA sequencing (scRNA-seq) of peripheral blood monocytes (PBMCs) from cABMR and control subjects was performed to define the transcriptomic landscape at single-cell resolution. A comprehensive scRNA-seq analysis was performed. The results indicated that most cell types in the cABMR patients exhibited an intense interferon response and release of proinflammatory cytokines. In addition, we found that the expression of MT-ND6, CXCL8, NFKBIA, NFKBIZ, and other genes were up-regulated in T- and B-cells and these genes were associated with pro-inflammatory response and immune regulation. Western blot and qRT-PCR experiments also confirmed the up-regulated expression of these genes in cABMR. GO and KEGG enrichment analyses indicated that the overexpressed genes in T- and B-cells were mainly enriched in inflammatory pathways, including the TNF, IL-17, and Toll-like receptor signaling pathways. Additionally, MAPK and NF-κB signaling pathways were also involved in the occurrence and development of cABMR. This is consistent with the experimental results of Western blot. Trajectory analysis assembled the T-cell subsets into three differentiation paths with distinctive phenotypic and functional prog rams. CD8 effector T cells and γδ T cells showed three different differentiation trajectories, while CD8_MAI T cells and naive T cells primarily had two differentiation trajectories. Cell-cell interaction analysis revealed strong T/B cells and neutrophils activation in cABMR. Thus, the study offers new insight into pathogenesis and may have implications for the identification of novel therapeutic targets for cABMR.

Identification of Liver Immune Microenvironment-Related Hub Genes in Liver of Biliary Atresia.

Background: Biliary atresia (BA) is one of the most common and fatal abnormalities of newborns. Increasing evidences indicated that immunology was the critical part of the etiology. This research used a public gene expression database to explore the immune microenvironment of BA liver.Methods: The gene expression profiles GSE46960, GSE159720, and GSE15235, containing BA and normal liver gene expression data, were obtained from the Expression Omnibus Gene. We applied CIBERSORTx to quantify 22 subsets of immune cells in BA liver. The differentially expressed genes (DEGs) and immune cells were used to further explore their relationship with liver fibrosis and the inflammation status of BA.Results: The expression of immune-related genes CXCL6, CXCL8, CXCL10, CCL20, IL32, TGFB2, SPP1, and SLIT2 was significantly different between BA and normal liver, among which CXCL8 was the hub gene. Six of 22 immune cell proportions were significantly different between BA and normal liver. Specifically, M0 macrophages and resting memory CD4+ T cells were upregulated in BA liver compared with normal liver. Meanwhile, monocytes, resting natural killer (NK) cells, plasma cells, and regulatory T (Treg) cells were downregulated. A further correlation analysis revealed that SLIT2 and CXCL6 owned high positive correlation coefficients with fibrosis grade, while the proportion of resting NK cells was negatively correlated. Proportions of resting CD4+ memory T cells were strongly related to the inflammation grade of BA liver.Conclusion: Biliary atresia is a disease strongly correlated with immune response. Our results might provide a clue for further exploration of BA etiology, which may promote a potential prediction model based on immune infiltration features.

STING Signaling Drives Production of Innate Cytokines, Generation of CD8+ T Cells and Enhanced Protection Against Trypanosoma cruzi Infection.

A variety of signaling pathways are involved in the induction of innate cytokines and CD8+ T cells, which are major players in protection against acute Trypanosoma cruzi infection. Previous data have demonstrated that a TBK-1/IRF3-dependent signaling pathway promotes IFN-β production in response to Trypanosoma cruzi, but the role for STING, a main interactor of these proteins, remained to be addressed. Here, we demonstrated that STING signaling is required for production of IFN-β, IL-6, and IL-12 in response to Trypanosoma cruzi infection and that STING absence negatively impacts activation of IRF-dependent pathways in response to the parasite. We reported no significant activation of IRF-dependent pathways and cytokine expression in RAW264.7 macrophages in response to heat-killed trypomastigotes. In addition, we showed that STING is essential for T. cruzi DNA-mediated induction of IFN-β, IL-6, and IL-12 gene expression in RAW264.7 macrophages. We demonstrated that STING-knockout mice have significantly higher parasitemia from days 5 to 8 of infection and higher heart parasitism at day 13 after infection. Although we observed similar heart inflammatory infiltrates at day 13 after infection, IFN-β, IL-12, CXCL9, IFN-γ, and perforin gene expression were lower in the absence of STING. We also showed an inverse correlation between parasite DNA and the expression of CXCL9, IFN-γ, and perforin genes in the hearts of infected animals at day 13 after infection. Finally, we reported that STING signaling is required for splenic IFN-β and IL-6 expression early after infection and that STING deficiency results in lower numbers of splenic parasite-specific IFN-γ and IFN-γ/perforin-producing CD8+ T cells, indicating a pivotal role for STING signaling in immunity to Trypanosoma cruzi.

Local Injection of Hydroxyapatite Electret Ameliorated Infarct Size After Myocardial Infarction.

Background: Previous studies showed that hydroxyapatite electret (HAE) accelerates the regeneration of vascular endothelial cells and angiogenesis. This study investigated the effects of HAE in myocardial infarction (MI) model mice. Methods and Results: MI was induced in mice by ligating the left anterior descending artery. Immediately after ligation, HAE, non-polarized hydroxyapatite (HAN), or water (control) was injected into the infarct border myocardium. Functional and histological analyses were performed 2 weeks later. Echocardiography revealed that HAE injection preserved left ventricular systolic function and the wall thickness of the scar, whereas HAN-injected mice had impaired cardiac function and thinning of the wall, similar to control mice. Histological assessment showed that HAE injection significantly attenuated the length of the scar lesion. There was significant accumulation of CD31-positive cells and increased expression of vascular endothelial growth factor (Vegf), intercellular adhesion molecule-1 (Icam1), vascular cell adhesion molecule-1 (Vcam1), hypoxia-inducible factor-1α (Hif1a), and C-X-C motif chemokine ligand 12 (Cxcl12) genes in the infarct border zone of HAE-injected mice. These effects were not induced by HAN injection. Anti-VEGFR2 antibody canceled the beneficial effect of HAE. In vitro experiments in a human cardiovascular endothelial cell line showed that HAE dose-dependently increased VEGFA expression. Conclusions: Local injection of HAE attenuated infarct size and improved cardiac function after MI, probably due to angiogenesis. The electric charge of HAE may stimulate angiogenesis via HIF1α-CXCL12/VEGF signaling.