CCR5 Expression Research Articles

Determination and analysis of gene expression involved in glucose metabolism under the conditions of the development of experimental diabetes of dexamethasone type (type 2 diabetes)

Type 2 diabetes is one of the most common and serious chronic diseases today, which has become a global health care problem. Type 2 diabetes accounts for about 90–95 % of all cases of diabetes and significantly affects patients’ quality life due to the development of numerous complications, such as cardiovascular diseases, chronic renal failure, retinopathy, neuropathy. Modern research in the field of diabetology pays considerable attention to the understanding of the genetic mechanisms of the pathogenesis of type 2 diabetes, which makes it possible to develop new approaches to its diagnosis and treatment. Individualization of laboratory diagnostics and treatment, which considers the genetic, metabolic and clinical characteristics of each patient, is a key direction in improving the effectiveness of the treatment of type 2 diabetes. Type 2 diabetes is a complex polygenic disease, that develops under the influence of both genetic and external factors, which requires an integrated approach to its diagnosis, treatment and prevention. Taking into account the constant increase in the prevalence of this disease, the relevance of scientific research in this area is beyond doubt. The development of new pharmacological agents, improvement of laboratory diagnostic strategies and individualization of treatment are key directions for overcoming the problem of type 2 diabetes and improving the quality of life of patients. The aim of the work: identification and analysis of genes panel, involved in glucose metabolism under the conditions of the development of experimental type 2 diabetes. Materials and methods. Analysis of the gene expression, involved in glucose metabolism was performed using the real-time reverse transcription polymerase chain reaction method CFX-96 Touch™ (Bio-Rad, USA) using the RT2Profiler™ PCR Array Rat Diabetes kit (QIAGEN, Germany). Results. Based on the results of the PCR study, the activity of the studied genes involved in glucose metabolism can be divided as follows: G6pc, Gpd1 – genes with high expression compared to the control group of animals; Ace, Acly, Foxg1, Foxp3, Gcgr, Gck, Gsk3b, Hmox1, Pygl, Snap23, Snap25 – genes with low expression compared to the control group of animals; Cebpa, Dpp4, Sell – genes in which no changes were detected in the samples in relation to the control group of animals; Ccr2, Fbp1, Gcg – genes, whose expression was not detected. Conclusions. The development of dexamethasone type 2 diabetes significantly (where ∆∆Ct <30) increases the expression of Gpd1 genes by 8 times and G6pc by 2 times compared to the control group of animals. During the development of type 2 dexamethasone diabetes, significantly (where ∆∆Ct <30) the Gsk3b and Hmox1 genes had a 17-fold low expression; Pygl at 11; Foxg1 in 7; Gck in 6; Ace and Foxp3 in 4; Acly in 3; Gcgr, Snap23, Snap25 in 2 times compared to the control group of animals. The expression of Ccr2, Fbp1, Gcg genes during the development of type 2 dexamethasone diabetes was not detected.

New insights into the immunomodulatory potential of sialic acid on monocyte-derived dendritic cells

Sialic acids at the cell surface of dendritic cells (DCs) play an important immunomodulatory role, and their manipulation enhances DC maturation, leading to heightened T cell activation. Particularly, at the molecular level, the increased stability of surface MHC-I molecules in monocyte-derived DCs (MoDCs) underpins an improved DC: T cell interaction. In this study, we focused on the impact of sialic acid remodelling by treatment with Clostridium perfringens sialidase on MoDCs' phenotypic and functional characteristics. Our investigation juxtaposes this novel approach with the conventional cytokine-based maturation regimen commonly employed in clinical settings.Notably, C. perfringens sialidase remarkably increased MHC-I levels compared to other sialidases having different specificities, supporting the idea that higher MHC-I is due to the cleavage of specific sialoglycans on cell surface proteins. Sialidase treatment induced rapid elevated surface expression of MHC-I, MHC-II and CD40 within an hour, a response not fully replicated by 48 h cytokine cocktail treatment. These increases were also observable 48 h post sialidase treatment. While CD86 and PD-L1 showed significant increases after 48 h of cytokine maturation, 48 h post sialidase treatment showed a higher increase in CD86 and shorter increase in PD-L1. CCR-7 expression was significantly increased 48 h after sialidase treatment but not significantly affected by cytokine maturation. Both treatments promoted higher secretion of the IL-12 cytokine. However, the cytokine cocktail induced a more pronounced IL-12 production. SNA lectin staining analysis demonstrated that the sialic acid profile is significantly altered by sialidase treatment, but not by the cytokine cocktail, which causes only slight sialic acid upregulation. Notably, the lipid-presenting molecules CD1a, CD1b and CD1c remained unaffected by sialidase treatment in MoDCs, a finding also further supported by experiments performed on C1R cells. Inhibition of endogenous sialidases Neu1 and Neu3 during MoDC differentiation did not affect surface MHC-I expression and cytokine secretion. Yet, sialidase activity in MoDCs was minimal, suggesting that sialidase inhibition does not significantly alter MHC-I-related functions. Our study highlights the unique maturation profile induced by sialic acid manipulation in MoDCs. These findings provide insights into the potential of sialic acid manipulation as a rapid immunomodulatory strategy, offering promising avenues for targeted interventions in inflammatory contexts.

Expression of CCL2 signaling pathway genes in patients with periodontitis and atherosclerosis.

Periodontitis and atherosclerosis are chronic inflammatory diseases characterized by leukocyte infiltration. We investigated the expression of CCL4, CCR5, c-Jun, c-Fos, NF-κB, and CCL2 as well as the possible mechanism involved in the regulation of CCL2 in human periodontitis tissues and atherosclerotic aorta based on previous research on the CCL4/CCR5/c-Jun and c-Fos/CCL2 pathway leading to CCL2 expression in collagen-induced arthritis (CIA) rat. Sixty-five volunteers were recruited and the condition of their gingiva and coronary arteries were assessed. The subjects were divided into four groups: healthy control, chronic periodontitis (CP), coronary artery diseases (CAD), and noncoronary artery diseases (non-CAD). Total RNA was isolated from gingiva in periodontitis patients and control populations and from the aorta in patients with and without CAD. PCR was used to examine CCL4, CCR5, c-Jun, c-Fos, NF-κB, and CCL2 levels. The production of CCL2 in the gingiva and aorta was analyzed by immunostaining. PCR revealed that CCL4, CCR5, and CCL2 mRNA levels were increased in CP patients' gingivae and aortas from coronary artery bypass grafting (CABG) patients. Marked c-Jun, c-Fos, and NF-κB gene productions were detected in CP patients' gingivae but did not show statistical differences between the CAD and non-CAD groups. Stronger immunoreactivity against CCL2 was observed in periodontitis gingiva and aorta from CABG patients. Our findings suggest that the CCL4/CCR5/c-Jun and c-Fos/CCL2 pathways may be involved in CCL2 expression in periodontitis. CCL4, CCR5, and CCL2 might act as possible nodes to link the presence of periodontitis and atherosclerosis.

Long-term monocyte activation after coronary artery bypass grafting: an exploratory prospective observational study

Major surgery such as coronary artery bypass grafting (CABG) is associated with an increased post-operative risk of atherosclerotic cardiovascular events. Cells of the innate immune system can adopt a long-lasting pro-inflammatory and atherogenic phenotype after brief exposure to exogenous or endogenous inflammatory stimuli, a process called “trained immunity”. We hypothesized that the surgery-induced inflammation leads to sustained alterations in monocyte function, which promote the subsequent occurrence of cardiovascular events. Blood from 13 patients undergoing elective CABG was obtained before, 3-7 days (median 4) after, and 6-8 weeks (median 6) weeks after surgery. At 3-7 days postoperatively, circulating C-reactive protein (CRP) concentration, leukocyte counts and ex vivo Peripheral Blood Mononuclear Cell (PBMC) IL-6, TNFα and IL-1Ra production after stimulation (with various inflammatory stimuli) were significantly increased. Simultaneously, there was a reduction in monocyte HLA-DR expression. 6-8 weeks after CABG there was an ongoing systemic pro-inflammatory state with higher CRP concentrations, increased stimulated ex vivo PBMC IL-6 production, changes in monocytes subsets, and a higher expression of CCR2 on monocytes compared to baseline. In conclusion, CABG induces a persistent systemic inflammatory reaction with a sustained activated monocyte phenotype. This might contribute to the increased atherosclerotic cardiovascular event risk observed in cardiac surgery patients.

DOCK8 deficiency due to a deep intronic variant in two kindreds with hyper-IgE syndrome

Dedicator of cytokinesis 8 (DOCK8) deficiency underlies the majority of cases of patients with autosomal recessive form of the hyper-immunoglobulin E syndrome (HIES). Most DOCK8 mutations involve deletions and splice junction mutations that abrogate protein expression. However, a few patients whose presentation is reminiscent of DOCK8 deficiency have no identifiable mutations. Using Whole Exome Sequencing (WES), we identified a deep intronic homozygous DOCK8 variant located in intron 36 (c.4626 + 76 A > G) in two unrelated patients with features of HIES that resulted in an in-frame 75 base pair intronic sequence insertion in DOCK8 cDNA, resulting in a premature stop codon (p.S1542ins6Ter). This variant resulted in variable decrease in DOCK8 expression that was associated with impaired T cell receptor-triggered actin polymerization, decreased IL-6-induced STAT3 phosphorylation, reduced expression of the Th17 cell markers CCR6 and IL-17, and higher frequencies of GATA3+ T cells indicative of Th2 skewing. Our approach extends the reach of WES in identifying disease-related intronic variants. It highlights the role of non-coding mutations in immunodeficiency disorders, including DOCK8 deficiency, and emphasizes the need to explore these mutations in unexplained inborn errors of immunity.

Glucose metabolism controls monocyte homeostasis and migration but has no impact on atherosclerosis development in mice

Monocytes directly contribute to atherosclerosis development by their recruitment to plaques in which they differentiate into macrophages. In the present study, we ask how modulating monocyte glucose metabolism could affect their homeostasis and their impact on atherosclerosis. Here we investigate how circulating metabolites control monocyte behavior in blood, bone marrow and peripheral tissues of mice. We find that serum glucose concentrations correlate with monocyte numbers. In diet-restricted mice, monocytes fail to metabolically reprogram from glycolysis to fatty acid oxidation, leading to reduced monocyte numbers in the blood. Mechanistically, Glut1-dependent glucose metabolism helps maintain CD115 membrane expression on monocytes and their progenitors, and regulates monocyte migratory capacity by modulating CCR2 expression. Results from genetic models and pharmacological inhibitors further depict the relative contribution of different metabolic pathways to the regulation of CD115 and CCR2 expression. Meanwhile, Glut1 inhibition does not impact atherosclerotic plaque development in mouse models despite dramatically reducing blood monocyte numbers, potentially due to the remaining monocytes having increased migratory capacity. Together, these data emphasize the role of glucose uptake and intracellular glucose metabolism in controlling monocyte homeostasis and functions.

MerTK Signaling in Human Primary T cells Modulates Memory Potential and Improves Recall response.

Immune therapy using checkpoint inhibitors or adoptive cell transfer has revolutionized the treatment of several types of cancers. However, response to treatment is currently limited to a fraction of patients. Elucidation of immune modulatory mechanisms might optimize patient selection and present ways to modify anti-cancer immune responses. We recently discovered the expression and an important costimulatory role of TAM receptor MerTK signaling on activated human primary CD8+ T cells. Here we extend our study of the costimulatory role of MerTK expression in human CD8+ T cells. We uncover a clear link between MerTK expression and less differentiated Central Memory T cells based on an increased expression of CCR7, CD45RO, CD28, CD62L, and an altered metabolic profile. In addition, we observe an improved proliferative capacity and elevated expression of effector molecule IFNγ upon recall responses of MerTK-expressing cells in vitro. Finally, using gp100TCR-transduced T cells, we demonstrate how PROS1 treatment results in improved cytotoxicity and killing of tumors. Our findings describe a role of MerTK expression in T cells, which could be exploited in the search for improving immunotherapeutic approaches.

Single-cell sequencing technology to characterize stem T-cell subpopulations in acute T-lymphoblastic leukemia and the role of stem T-cells in the disease process.

Precursor T-cell acute lymphoblastic leukemia (Pre-T ALL) is a malignant neoplastic disease in which T-cells proliferate in the bone marrow. Single-cell sequencing technology could identify characteristic cell types, facilitating the study of the therapeutic mechanisms in Pre-T ALL. The single-cell sequencing data (scRNA-seq) of Pre-T ALL were obtained from public databases. Key immune cell subpopulations involved in the progression of Pre-T ALL were identified by clustering and annotating the cellular data using AUCell. Next, pseudo-temporal analysis was performed to identify the differentiation trajectories of immune cell subpopulations using Monocle. Copy number mutation landscape of cell subpopulations was characterized by inferCNV. Finally, cellphoneDB was used to analyze intercellular communication relationships. A total of 10 cellular subpopulations were classified, with Pre-T ALL showing a higher proportion of NK/T cells. NK/T cells were further clustered into two subpopulations. Stem T cells showed a high expression of marker genes related to hematopoietic stem cells, Naive T cells had a high expression of CCR7, CCR7, RCAN3, and NK cells high-expressed KLRD1, TRDC. The cell proliferation was reduced and the activation of T cell was increased during the differentiation of stem T cells to Naive T cells. We observed interaction between stem T cells with dendritic cells such as CD74-COPA, CD74-MIF as well as co-inhibition-related interactions such as LGALS9-HAVCR2, TGFB1-TGFBR3. Stem T cells were involved in the development of Pre-T-ALL through the regulatory effects of transcription factors (TFs) KLF2 and FOS and multiple ligand-receptor pairs.

STYK1 mediates NK cell anti-tumor response through regulating CCR2 and trafficking

The serine/threonine/tyrosine kinase 1 (STYK1) is a receptor protein-tyrosine kinase (RPTK)-like molecule that is detected in several human organs. STYK1 plays an important role in promoting tumorigenesis and metastasis in various cancers. By analyzing the expression of RTKs in immune cells in the database of 2013 Immunological Genome Project, we found that STYK1 was principally expressed in NK cells. In order to investigate the function of STYK1, we used CRISPR/Cas9 technology to generate STYK1-deleted mice, we found STYK1 deletion mice have normal number, development, and function of NK cells in spleen and bone marrow in tumor-free resting state. To examine the tumor surveillance of STYK1 in vivo, we utilized a variety of tumor models, including NK cell-specific target cell (ß2M and RMA-S) clearance experiments in vivo, subcutaneous and intravenous injection of B16F10 melanoma model, and the spontaneous breast cancer model MMTV-PyMT. Surprisingly, we discovered that deletion of the oncogenic STYK1 promoted the four-model tumor progression, and we observed a reduction of NK cell accumulation in the tumor tissues of STYK1 deletion mice compared to WT mice. In order to study the mechanism of STYK1 in NK, RNA sequence of STYK1−/− and WT NK have unveiled a disparity in the signaling pathways linked to migration and adhesion in STYK1−/− NK cells. Further analysis of chemokine receptors associated with NK cell migration revealed that STYK1-deficient NK cells exhibited a significant reduction in CCR2 expression. The STYK1 expression was negatively associated with tumor progression in glioma patients. Overall, our study found the expression of STYK1 in NK cell mediates NK cell anti-tumor response through regulating CCR2 and infiltrating into tumor tissue.

Hemodiafiltration May Be Associated with Senescence-Related Phenotypic Alterations of Lymphocytes, Which May Predict Mortality in Patients Undergoing Dialysis.

Senescence-resembling alterations on the lymphocytes of patients undergoing dialysis have been widely described. However, the pathophysiology behind these phenomena has not been clarified. In this study, we examined the impact of dialysis prescription on T and B lymphocytes, in patients undergoing dialysis.: T and B cell subsets were determined with flow cytometry in 36 patients undergoing hemodialysis and 26 patients undergoing hemodiafiltration, according to the expression of CD45RA, CCR7, CD31, CD28, CD57, and PD1 for T cells, and IgD and CD27 for B cells. The immune phenotype was associated with dialysis modality, hemofiltration volume, and mortality. Compared with hemodialysis, patients undergoing hemodiafiltration had a significantly decreased percentage of CD4+CD28-CD57- T cells [3.8 (2.4-5.3) vs. 2.1 (1.3-3.3)%, respectively, p = 0.002] and exhausted CD4+ T cells [14.1 (8.9-19.4) vs. 8.5 (6.8-11.7)%, respectively, p = 0.005]. Additionally, the hemofiltration volume was negatively correlated with CD8+ EMRA T cells (r = -0.46, p = 0.03). Finally, the increased exhausted CD4+ T cell percentage was associated with increased all-cause mortality in patients undergoing dialysis, independent of age. Hemodiafiltration, especially with high hemofiltration volume, may have beneficial effects on senescence-related immune phenotypes. Immune phenotypes may also be a predicting factor for mortality in patients undergoing dialysis.

CRISPR screens unveil nutrient-dependent lysosomal and mitochondrial nodes impacting intestinal tissue-resident memory CD8+ T cell formation

Nutrient availability and organelle biology direct tissue homeostasis and cell fate, but how these processes orchestrate tissue immunity remains poorly defined. Here, using invivo CRISPR-Cas9 screens, we uncovered organelle signaling and metabolic processes shaping CD8+ tissue-resident memory T (TRM) cell development. TRM cells depended on mitochondrial translation and respiration. Conversely, three nutrient-dependent lysosomal signaling nodes-Flcn, Ragulator, and Rag GTPases-inhibited intestinal TRM cell formation. Depleting these molecules or amino acids activated the transcription factor Tfeb, thereby linking nutrient stress to TRM programming. Further, Flcn deficiency promoted protective TRM cell responses in the small intestine. Mechanistically, the Flcn-Tfeb axis restrained retinoic acid-induced CCR9 expression for migration and transforming growth factor β (TGF-β)-mediated programming for lineage differentiation. Genetic interaction screening revealed that the mitochondrial protein Mrpl52 enabled early TRM cell formation, while Acss1 controlled TRM cell development under Flcn deficiency-associated lysosomal dysregulation. Thus, the interplay between nutrients, organelle signaling, and metabolic adaptation dictates tissue immunity.

Host genetic and immune factors drive evasion of HIV-1 pathogenesis in viremic non-progressors.

Viremic non-progressors (VNPs) represent an exceptional and uncommon subset of people with HIV-1, characterized by the remarkable preservation of normal CD4+ Tcell counts despite uncontrolled viral replication-a trait reminiscent of natural hosts of simian immunodeficiency virus. The mechanisms orchestrating evasion from HIV-1 pathogenesis in human VNPs remain elusive, primarily due to the absence of integrative studies. We implemented a novel single-cell and multiomics approach to comprehensively characterize viral, genomic, transcriptomic, and metabolomic factors driving this exceedingly rare disease phenotype in 16 VNPs and 29 HIV+ progressors. Genetic predisposition to the VNP phenotype was evidenced by a higher prevalence of CCR5Δ32 heterozygosity, which was associated with lower levels of CCR5 expression and a lower frequency of infected cells in peripheral circulation. We also observed reduced levels of plasma markers of intestinal disruption and attenuated interferon responses in VNPs. These factors potentially drive the other phenotypic traits of immune preservation in this population, including the unaltered tryptophan metabolic profile, reduced activation of cytotoxic lymphocytes, and reduced bystander CD4+ Tcell apoptosis. In summary, our comprehensive analysis identified intricate factors collectively associated with the unique immunovirological equilibrium in VNPs, shedding light on potential avenues for therapeutic exploration in managing HIV pathogenesis. The work was supported by funding from the Spanish Ministry of Science and Innovation and the National Institutes of Health (NIH).

The Long COVID: Further Advances in Our Understanding about the Role of Specific Chemokines (CCR5, CCR6, CCR9 and CCL3) in Pediatrics

Introduction: Long COVID, also known as post-COVID-19 syndrome (PCS), is a condition where individuals continue to experience symptoms for weeks or months after recovering from COVID-19. The aim of this study was to evaluate the long-term effects of severe COVID-19 on promotor methylation and expression genes including CCR5, CCR6, CCR9 and CCL3 in children. Material and Methods: Clinical data and blood samples from 94 long COVID patients and 25 healthy subjects were collected. The control group was age-matched. Promotor methylation and mRNA expression of CCR5, CCR6, CCR9 and CCL3 genes in these patients and control group were assessed through methylation-specific PCR and Real-time PCR assay. Results: Our result indicated that promotor of CCR5 (p = 0.01) and CCL3 (p = 0.006) in long COVID children were hyper-methylated compared to healthy control group. Subsequently CCR5 and CCL3 transcript was decreased compared to control group (p = 0.01). In addition, CCL3 transcript in children with long COVID was decreased compared to control group (p = 0.008). On the other hand, we did not observe any significant modification in the transcript levels of CCR6 (p = 0.7) and CCR9 (p = 0.46) in children with long COVID in comparison to all control groups. Conclusion: The CCR5 and CCL3 promoter region DNA methylation and the subsequent decrease in the expression of these genes were possibly correlated with long COVID occurrence in children. Our study revealed additional data on the SARS-CoV-2 mediated inflammatory response. conclusion: The CCR5 and CCL3 promoter region DNA methylation and the subsequent decrease in the expression of these genes were possibly correlated with long COVID occurrence in children. Our study revealed additional data on the SARS-CoV-2 mediated inflammatory response.

Loss of CCL28 and CXCL17 Expression and Increase in CCR1 Expression May Be Related to Malignant Transformation of LGBLEL into Lymphoma.

To investigate the differential expression of the chemokine signaling pathway in lacrimal gland benign lymphoepithelial lesion (LGBLEL) and lacrimal lymphoma, providing insights into the mechanisms underlying malignant transformation and aiding clinical differentiation. Transcriptome analysis was conducted on patients with LGBLEL, lymphoma, and orbital cavernous hemangioma (CH). Three cases of LGBLEL and three cases of lymphoma were randomly selected as control and experimental groups, respectively. A real-time quantitative polymerase chain reaction (RT-qPCR) was used to validate genes associated with the chemokine signaling pathway. Immunohistochemical (IHC) staining and quantitative Western blotting (WB) were performed for precise protein quantification. Transcriptome analysis revealed differential expression of the chemokine signaling pathway between the LGBLEL and lymphoma groups, identifying ten differentially expressed genes: CCL17, VAV2, CXCR5, NRAS, HCK, RASGRP2, PREX1, GNB5, ADRBK2, and CCL22. RT-qPCR showed that, compared to the lymphoma group, the LGBLEL group had significantly higher expression of CCL28, CXCL17, HCK, GNB5, NRAS, and VAV2 (p = 0.001, <0.001, <0.001, <0.001, =0.020, <0.001, respectively) and lower expression of CCR1 (p = 0.002). IHC staining and quantitative analysis confirmed significant differences in protein expression between the groups for CCL28, CCR1, CXCL17, HCK, GNB5, NRAS, and VAV2 (p = 0.003, 0.011, 0.001, 0.024, 0.005, 0.019, and 0.031, respectively). While IHC provided localization, WB offered greater precision. WB revealed that, compared to the lymphoma group, the LGBLEL group exhibited significantly higher expression of CCL28, CXCL17, HCK, GNB5, NRAS, and VAV2 (p = 0.012, 0.005, 0.009, 0.011, 0.008, and 0.003, respectively) and lower expression of CCR1 (p = 0.014). The chemokine signaling pathway plays a role in the malignant transformation of LGBLEL. The decreased expression of CCL28 and CXCL17, coupled with the increased expression of CCR1, may be linked to the progression of LGBLEL into lymphoma.

NEIL3 Upregulated by TFAP2A Promotes M2 Polarization of Macrophages in Liver Cancer via the Mediation of Glutamine Metabolism

Introduction: Tumor-associated macrophages, which are part of the tumor microenvironment, are a major factor in cancer progression. However, a complete understanding of the regulatory mechanism of M2 polarization of macrophages (Mø) in liver cancer is yet to be established. This study aimed to investigate the potential mechanism by which NEIL3 influenced M2 Mø polarization in liver cancer. Methods: Bioinformatics analysis analyzed NEIL3 expression and its enriched pathways in liver cancer tissue, as well as its correlation with pathway genes. The upstream transcription factor of NEIL3, TFAP2A, was predicted and its expression in liver cancer tissue was analyzed. The binding relationship between the two was analyzed by dual-luciferase reporter and chromatin immunoprecipitation experiments. qRT-PCR assessed NEIL3 and TFAP2A levels in liver cancer cells. Cell viability was detected by CCK-8, while CD206 and CD86 expression was detected by immunofluorescence. IL-10 and CCR2 expressions were assessed using qRT-PCR, and M2 Mø quantity was detected using flow cytometry. Reagent kits tested glutamine (Gln) consumption, α-ketoglutarate, and glutamate content, as well as NADPH/NADP+ and GSH/GSSG ratios. Expression of Gln transport proteins was detected using Western blot. An animal model was established to investigate the influence of NEIL3 expression on liver cancer growth. Results: NEIL3 was highly expressed in liver cancer and promoted Mø M2 polarization through Gln metabolism. TFAP2A was identified as the upstream transcription factor of NEIL3 and was highly expressed in liver cancer. Rescue experiments presented that overexpression of NEIL3 reversed the suppressive effect of TFAP2A knockdown on Mø M2 polarization in liver cancer. In vivo experiments demonstrated that the knockdown of NEIL3 could significantly repress the growth of xenograft tumors. Conclusion: This study suggested that the TFAP2A/NEIL3 axis promoted Mø M2 polarization through Gln metabolism, providing a theoretical basis for immune therapy targeting the liver cancer TME.

Differential expression of HIV target cells CCR5 and α4β7 in tissue resident memory CD4 T cells in endocervix during the menstrual cycle of HIV seronegative women.

Ovarian hormones are known to modulate the immune system in the female genital tract (FGT). We sought to define the impact of the menstrual cycle on the mucosal HIV target cell levels, and tissue-resident CD4 T cells. Here, we characterized the distribution, phenotype, and function of CD4 T cells with special emphasis on HIV target cells (CCR5+ and α4β7+) as well as tissue-resident memory (TRM; CD69+ and CD103+) CD4 T cells in FGT of cycling women. Peripheral blood and Endocervical cells (EC-collected from cytobrush) were collected from 105 healthy women and performed multicolor flow cytometry to characterize the various subsets of CD4 T cells. Cervicovaginal lavage (CVL) were collected for cytokine analysis and plasma were collected for hormonal analysis. All parameters were compared between follicular and luteal phase of menstrual cycle. Our findings revealed no significant difference in the blood CD4 T cell subsets between the follicular and luteal phase. However, in EC, the proportion of several cell types was higher in the follicular phase compared to the luteal phase of menstrual cycle, including CCR5+α4β7-cells (p=0.01), CD69+CD103+ TRM (p=0.02), CCR5+CD69+CD103+ TRM (p=0.001) and FoxP3+ CD4 T cells (p=0.0005). In contrast, α4β7+ CCR5- cells were higher in the luteal phase (p=0.0004) compared to the follicular phase. In addition, we also found that hormonal levels (P4/E2 ratio) and cytokines (IL-5 and IL-6) were correlated with CCR5+ CD4 T cells subsets during the follicular phase of the menstrual cycle. Overall, these findings suggest the difference in the expression of CCR5 and α4β7 in TRM CD4 T cell subsets in endocervix of HIV seronegative women between the follicular and luteal phase. Increase in the CCR5+ expression on TRM subsets could increase susceptibility to HIV infection during follicular phase of the menstrual cycle.

Changes in conjunctival mononuclear phagocytes and suppressive activity of regulatory macrophages in desiccation induced dry eye

PurposeTo evaluate the effects of dry eye on conjunctival immune cell number and transcriptional profiles with attention to mononuclear phagocytes. MethodsExpression profiling was performed by single-cell RNA sequencing on sorted conjunctival immune cells from non-stressed and C57BL/6 mice subjected to desiccating stress (DS). Monocle 3 modeled cell trajectory, scATAC-seq assessed chromatin accessibility and IPA identified canonical pathways. Inflammation and goblet cells were measured after depletion of MRC1+ MΦs with mannosylated clodronate liposomes. ResultsMononuclear phagocytes (monocytes, MΦs, DCs) comprised 72 % of immune cells and showed the greatest changes with DS. Distinct DS induced gene expression patterns were seen in phagocytes classified by expression of Ccr2 and [Timd4, Lyve1, Folr2 (TLR)]. Expression of phagocytosis/efferocytosis genes increased in TLF+CCR2- MΦs. Monocytes showed the highest expression of Ace, Cx3cr1, Vegfa, Ifngr1,2, and Stat1 and TLF−CCR2+ cells expressed higher levels of inflammatory mediators (Il1a, Il1b, Il1rn, Nfkb1, Ccl5, MHCII, Cd80, Cxcl10, Icam1). A trajectory from monocyte precursors branched to terminate in regulatory MΦs or in mDCs via transitional MΦ and cDC clusters. Activated pathways in TLF+ cells include phagocytosis, PPAR/RXRα activation, IL-10 signaling, alternate MΦ activation, while inflammatory pathways were suppressed. Depletion of MRC1+ MΦs increased IL-17 and IFN-γ expression and cytokine-expressing T cells, reduced IL-10 and worsened goblet loss. ConclusionsDryness stimulates distinct gene expression patterns in conjunctival phagocytes, increasing expression of regulatory genes in TLF+ cells regulated in part by RXRα, and inflammatory genes in CCR2+ cells. Regulatory MΦs depletion worsens DS induced inflammation and goblet cell loss.

Blockade of the CCR3 receptor reduces neutrophil recruitment to the lung during acute inflammation.

Neutrophils represent one of the host's first lines of defense against invading pathogens. However, an aberrant activation can cause damage to the host. In the case of respiratory infections with viral or bacterial pathogens, one of the most common complications is the development of acute respiratory distress syndrome, in which neutrophil infiltration into the lung is a hallmark. Neutrophils gain expression of chemokine receptors under inflammatory conditions, and their activation can amplify the neutrophil responses. Earlier studies showed that neutrophils recruited to the lung mucosa during bacterial infection upregulate expression of CCR3 and ex vivo stimulation of CCR3 results in an increased neutrophil activation. Therefore, the modulation of effector functions or migration of neutrophils to target sites through chemokine receptors constitutes an opportunity for pharmacological intervention. We aimed to determine whether the blockade of the CCR3 using the specific antagonist SB-328437 reduces neutrophil recruitment and inflammation in the lung in the lipopolysaccharide (LPS)-induced lung injury model and influenza infection in mice. We found that neutrophils acquire CCR3 expression in the lung alveolar space. The intraperitoneal administration of SB-328437 reduced neutrophil recruitment to the lung alveolar space and reduced tissue damage in both the LPS-induced lung injury model and influenza infection. Moreover, treatment with SB-328437 reduced the percentage of neutrophils producing TNFα and neutrophil activation in the alveolar space. Together, these data suggest that CCR3 blockade might be a pharmacological strategy to prevent the aberrant neutrophil activation that results detrimental for the host but preserves sufficient effector response to control the pathogen.

Changes in the Release of Endothelial Extracellular Vesicles CD144+, CCR6+, and CXCR3+ in Individuals with Acute Myocardial Infarction.

Acute myocardial infarction (AMI) results from vulnerable plaque rupture, causing ischemic cardiomyocyte necrosis and intense inflammation. Paradoxically, this inflammation releases factors that aid heart repair. Recent findings suggest a role for extracellular vesicles (EVs) in intercellular communication during post-AMI cardiac repair. However, EVs' tissue origin and chemokine profile in the blood of patients with AMI remains unclear. This study characterized the tissue origin and chemokine receptor profile of EVs in the coronary and peripheral blood of patients with AMI. The results reveal that vesicles isolated from coronary and peripheral blood plasma are enriched in tetraspanin (CD9) and express CD81+, CD90+, and CD144+. The vesicle size ranged between 145 and 162 nm, with the control group exhibiting smaller vesicles (D10) than the AMI group. Furthermore, all vesicles expressed CCR6 and CXCR3, whereas a small percentage expressed CCR4. In addition, a decrease in CXCR3 and CCR6 expression was observed in coronary and peripheral AMI blood vesicles compared with the control; however, no difference was found between AMI coronary and AMI peripheral blood vesicles. In conclusion, our study demonstrates, for the first time, changes in the number of extracellular vesicles expressing CD144+, CXCR3, and CCR6 in the peripheral circulation of patients with AMI. Extracellular vesicles present in the circulation of patients with AMI hold excellent promise as a potential diagnostic tool.

Identification of common signature genes and pathways underlying the pathogenesis association between nonalcoholic fatty liver disease and heart failure.

Non-alcoholic fatty liver disease (NAFLD) and heart failure (HF) are related conditions with an increasing incidence. However, the mechanism underlying their association remains unclear. This study aimed to explore the shared pathogenic mechanisms and common biomarkers of NAFLD and HF through bioinformatics analyses and experimental validation. NAFLD and HF-related transcriptome data were extracted from the Gene Expression Omnibus (GEO) database (GSE126848 and GSE26887). Differential analysis was performed to identify common differentially expressed genes (co-DEGs) between NAFLD and HF. Gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and gene set enrichment analysis (GSEA) were conducted to explore the functions and regulatory pathways of co-DEGs. Protein-protein interaction (PPI) network and support vector machine-recursive feature elimination (SVM-RFE) methods were used to screen common key DEGs. The diagnostic value of common key DEGs was assessed by receiver operating characteristic (ROC) curve and validated with external datasets (GSE89632 and GSE57345). Finally, the expression of biomarkers was validated in mouse models. A total of 161 co-DEGs were screened out in NAFLD and HF patients. GO, KEGG, and GSEA analyses indicated that these co-DEGs were mainly enriched in immune-related pathways. PPI network revealed 14 key DEGs, and SVM-RFE model eventually identified two genes (CD163 and CCR1) as common key DEGs for NAFLD and HF. Expression analysis revealed that the expression levels of CD163 and CCR1 were significantly down-regulated in HF and NAFLD patients. ROC curve analysis showed that CD163 and CCR1 had good diagnostic values for HF and NAFLD. Single-gene GSEA suggested that CD163 and CCR1 were mainly engaged in immune responses and inflammation. Experimental validation indicated unbalanced macrophage polarization in HF and NAFLD mouse models, and the expression of CD163 and CCR1 were significantly down-regulated. This study identified M2 polarization impairment characterized by decreased expression of CD163 and CCR1 as a common pathogenic pathway in NAFLD and HF. The downregulation of CD163 and CCR1 may reflect key pathological changes in the development and progression of NAFLD and HF, suggesting their potential as diagnostic and therapeutic targets.