Lymphocyte Chemokine Research Articles

B cell lymphoma 6 promotes hepatocellular carcinoma progression by inhibiting tumor infiltrating CD4+T cell cytotoxicity through ESM1

Immunotherapy exhibited potential effects for advanced hepatocellular carcinoma, unfortunately, the clinical benefits are often countered by cancer adaptive immune suppressive response. Uncovering the mechanism how cancer cells evade immune surveillance would help to develop new immunotherapy approaches and combination therapy. In this article, by analyzing the transcriptional factors which modulate the differentially expressed genes between T cell infiltration high group and low group, we identified oncoprotein B cell lymphoma 6 (BCL6) suppresses the infiltration and activation of tumor infiltrating T lymphocytes, thus correlated with poorer clinical outcome. By using antibody deletion experiment, we further demonstrated that CD4+T cells but not CD8+T cells are the main lymphocyte population suppressed by Bcl6 to promote HCC development. Mechanistically, BCL6 decreases cancer cell expression of pro-inflammatory cytokines and T lymphocyte chemokines such as IL6, IL1F6, and CCL5. Moreover, BCL6 upregulates Endothelial cell-specific molecule 1 (ESM1) to inhibit T lymphocyte recruitment and activation possibly through ICAM-1/LFA-1 signaling pathway. Our findings uncovered an unappreciated paracrine mechanism how cancer cell-derived BCL6 assists cancer cell immune evasion, and highlighted the role of CD4+T cells in HCC immune surveillance.

Psoralen attenuates cigarette smoke extract-induced inflammation by modulating CD8+ T lymphocyte recruitment and chemokines via the JAK2/STAT1 signaling pathway

Chronic obstructive pulmonary disease (COPD) is a respiratory inflammatory disease. Psoralen (PSO) is the main pharmacological component identified from Bu-Shen-Fang-Chuan formula which has been traditionally used in treatment of COPD, yet its efficacy in COPD inflammation were unreported. In this study, we aimed to elucidate the anti-inflammatory potential of PSO in COPD and unravel the underlying mechanisms, focusing on T lymphocyte recruitment and the modulation of chemokines, namely monokine induced by interferon-gamma (CXCL9), interferon inducible protein 10 (CXCL10), and interferon inducible T-Cell alpha chemoattractant (CXCL11). In vitro, RAW264.7 was stimulated by interferon (IFN)-γ + cigarette smoke extract (CSE) and were treated with PSO (2.5, 5, 10 μM), then the levels of chemokines and the activation of Janus kinase (JAK)/Signal transducer and activator of transcription 1 (STAT1) pathway were analyzed by real time PCR and western blot. In vivo, a murine model was established by intraperitoneal injection of CSE on day 1, 8, 15, and 22, then treated with PSO (10 mg/kg). Our experiments in vitro illustrated that PSO reduced the levels of CXCL9, CXCL10, and CXCL11, and decreased the protein phosphorylation levels of JAK2 and STAT1. Additionally, PSO effectively improved inflammatory infiltration and decreased the proportion of CD8+ T cells in CSE-exposed mice. Furthermore, PSO reduced the levels of CXCL9, CXCL10, and CXCL11 in bronchoalveolar lavage fluid (BALF) and lung tissue, and decreased the protein phosphorylation levels of JAK2 and STAT1. In conclusion, our results revealed the therapeutic potential of PSO for COPD inflammation, possibly mediated through the regulation of CD8+ T cell recruitment and chemokines via the JAK2/STAT1 signaling pathway.

Abstract 6676: Exploratory biomarker analyses in phase 1 study of regorafenib, ipilimumab, and nivolumab (RIN) in chemotherapy resistant microsatellite stable (MSS) metastatic colorectal cancer (mCRC)

Abstract Background: We recently reported an objective response rate of 36.4% in a phase I trial of RIN in MSS mCRC patients without liver metastases, while no response was noted in patients with liver metastases. Here we characterized kinetics in local and systemic immune responses associated with clinical outcomes, liver metastasis, and immune related toxicity during RIN treatment. Methods: Patients with chemotherapy-resistant MSS mCRC received RIN at (R) at 80 mg QD x 21 days in 4-week(w) cycle, (I) at 1mg/kg Q6w and (N) at 240 mg Q 2w (n=29). Needle biopsies were collected at baseline (n=8) and 4 weeks (n=6) after treatment. Blood samples were collected at baseline (n=29), 4 weeks (n=29) and 8 weeks (n=25) after treatment. Alterations in the tumor microenvironment (TME) were characterized by RNA sequencing. We analyzed the immune cell composition and activities in the blood, including 90 subpopulations of leukocytes and 29 activation/suppression/proliferation markers, with high-dimensional spectral flow cytometry. Results: The clinical responses are positively correlated with the induction of local anti-tumor immunity. Patients with better clinical outcomes exhibited higher levels of lymphocyte infiltration, chemokine production, and antigen-presentation potential in the local TME following one cycle of RIN treatment. The duration of progression-free survival (PFS) was correlated with a higher baseline STING signal in the TME. In the peripheral blood, the responders have more circulating CD4 T cells, B cells, and cDC1, while expressing lower immune activation markers, especially in T cells with type 1 immunity at baseline. At 4 weeks, immune activation and proliferation markers were higher in responders than non-responders. Interestingly, immune effectors (non-Tregs, CD45RA− CD8 T cells, NKT cells, and M1 monocytes) in the blood were proportionally reduced in responders at 4 weeks, coincident with their enrichment in the local TME. We also compared peripheral immune signatures between patients with and without liver metastases, liver metastases (n=7) were associated with a significantly lower ratio of CD4/CD8 T cells in the blood, a phenomenon of immunosenescence, both at baseline and after RIN treatment. Lastly, the occurrence of immune-related toxicities was closely correlated with the induction of immune activities in both acellular and cellular components in the circulation. Conclusions: Clinical benefit of RIN was correlated to more robust immune activation of the TME and peripheral blood following one cycle of treatment. A higher baseline STING signal in the TME was associated with longer PFS. Liver metastasis was associated with a lower ratio of peripheral CD4/CD8 T cells which may contribute to the resistance to immunotherapy in those patients. Citation Format: Jian Ye, Chongkai Wang, Colt Egelston, Weihua Guo, Peter P. Lee, Marwan Fakih. Exploratory biomarker analyses in phase 1 study of regorafenib, ipilimumab, and nivolumab (RIN) in chemotherapy resistant microsatellite stable (MSS) metastatic colorectal cancer (mCRC). [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6676.

Diminished PD-L1 regulation along with dysregulated T lymphocyte subsets and chemokine in ANCA-associated vasculitis.

ANCA-associated vasculitis (AAV) is a life-threatening disease characterized by small vessel inflammation and pathogenic self-directed antibodies. Programmed death-ligand 1 receptor (PD-1) and programmed cell death ligand-1 (PD-L1) are immune checkpoint molecules crucial for maintaining tolerance and immune homeostasis. After checkpoint inhibition therapy, development of various autoimmune diseases and immune-related adverse events (irAEs) have been observed. Here, we investigated the immunomodulatory roles of neutrophils through the expression of immune checkpoint molecule (PD-L1), migratory molecules (CXCR2), chemotactic chemokines (CXCL5) and other important molecules (BAFF and HMGB1) in development of AAV. We also scrutinized the immune mechanism responsible for development of pauci-immune crescentic GN (PICGN). We demonstrate for the first time that the frequency of PD-L1 expressing neutrophils was significantly reduced in AAV patients compared to healthy controls and correlated negatively with disease severity (BVASv3). Further, in renal biopsy, reduced PD-L1 immune checkpoint expression provides a microenvironment that unleashes uncontrolled activated CD4 + T cells, B cells, neutrophils and macrophages and ultimately causes engulfment of immune complexes leading to PICGN. Furthermore, during remission, reduced neutrophils PD-L1 and CXCR2 expression, increased neutrophils CXCL5 expression and increased peripheral effector memory T cells and increased HMGB1 and BAFF levels in serum, demonstrate the propensity for the persistence of sub-clinical inflammation, which could explain relapse, in this group of diseases.

Ascorbic acid induced TET2 enzyme activation enhances cancer immunotherapy efficacy in renal cell carcinoma.

Exploring the regulatory mechanism of PD-L1 in renal cancer is one of the key strategies to improve the response of renal cancer patients to checkpoint blockade therapy. In this study, the synergistic effect of ascorbic acid (vitamin C) supplementation and the impact of TET2 depletion on anti-PD-L1 therapy were determined in xenograft model experiments. Lymphocyte infiltration and chemokine expression were determined using flow cytometry and qRT-PCR. To determine the downstream targets of TET2, we performed hMeDip-seq and RNA-seq analyses. The molecular mechanism was further confirmed by hMeDip-qPCR, MeDip-qPCR, bisulfite sequencing, Western blotting, qRT-PCR and xenograft model experiments in vitro and in vivo. The present study demonstrated that ascorbic acid enhanced the efficacy of immunotherapy and that the loss of TET2 function enabled renal cancer cells to evade antitumor immunity. Ascorbic acid treatment significantly increased the intratumoral infiltration of T cells and the expression of cytokines and chemokines, while the loss of TET2 impaired the infiltration of T cells and the expression of cytokines and chemokines. TET2 was recruited to IRF1 by IFN-γ-STAT1 signaling, thereby maintaining IRF1 demethylation and ultimately inducing PD-L1 expression. These results suggest a new strategy of stimulating TET activity to improve immunotherapy for renal cell carcinoma.

Astrocyte-specific expression of interleukin 23 leads to an aggravated phenotype and enhanced inflammatory response with B cell accumulation in the EAE model

BackgroundInterleukin 23 is a critical cytokine in the pathogenesis of multiple sclerosis. But the local impact of interleukin 23 on the course of neuroinflammation is still not well defined. To further characterize the effect of interleukin 23 on CNS inflammation, we recently described a transgenic mouse model with astrocyte-specific expression of interleukin 23 (GF-IL23 mice). The GF-IL23 mice spontaneously develop a progressive ataxic phenotype with cerebellar tissue destruction and inflammatory infiltrates with high amounts of B cells most prominent in the subarachnoid and perivascular space.MethodsTo further elucidate the local impact of the CNS-specific interleukin 23 synthesis in autoimmune neuroinflammation, we induced a MOG35-55 experimental autoimmune encephalomyelitis (EAE) in GF-IL23 mice and WT mice and analyzed the mice by histology, flow cytometry, and transcriptome analysis.ResultsWe were able to demonstrate that local interleukin 23 production in the CNS leads to aggravation and chronification of the EAE course with a severe paraparesis and an ataxic phenotype. Moreover, enhanced multilocular neuroinflammation was present not only in the spinal cord, but also in the forebrain, brainstem, and predominantly in the cerebellum accompanied by persisting demyelination. Thereby, interleukin 23 creates a pronounced proinflammatory response with accumulation of leukocytes, in particular B cells, CD4+ cells, but also γδ T cells and activated microglia/macrophages. Furthermore, transcriptome analysis revealed an enhanced proinflammatory cytokine milieu with upregulation of lymphocyte activation markers, co-stimulatory markers, chemokines, and components of the complement system.ConclusionTaken together, the GF-IL23 model allowed a further breakdown of the different mechanisms how IL-23 drives neuroinflammation in the EAE model and proved to be a useful tool to further dissect the impact of interleukin 23 on neuroinflammatory models.

Microenvironment-related prognostic genes in esophageal cancer

BackgroundEsophageal cancer is one of the most common malignant tumors. The role of tumor microenvironment in esophageal cancer is unclear.MethodsThe gene expression profiles and clinical data of 158 patients with esophageal cancer were extracted from The Cancer Genome Atlas database. Immune scores and stromal scores were calculated based on ESTIMATE algorithm. According to different immune/stromal scores, differentially expressed genes (DEGs) were identified. The function enrichment, protein interactions of shared DEGs and their associations with overall survival were analyzed.ResultsIn regard to the association of the immune/stromal scores and disease stage, pathological type and overall survival, only the stromal scores among the different stages were significantly different (P=0.015). In the high immune and stromal score groups, 603 shared up-regulated genes were found. The related function and pathways included regulation of lymphocyte activation, cytokine binding and chemokine signaling pathway. Protein-protein interaction analysis showed that ITGAM had the most connections, followed by CXCL10 and CCR2. High expression of 11 genes, including MS4A7, TMIGD3, MS4A4A, EVI2A, MS4A6A, FCER1G, AIF1, GNGT2, LCP2, DNAJC5B and RNASE6, were found to be associated with shorter overall survival.ConclusionsMicroenvironment-associated functions and pathways were analyzed in esophageal cancer, and 11 microenvironment-associated genes were correlated to poor prognoses. Further studies on these genes may be helpful to understand the tumor microenvironment and provide new therapies for esophageal cancer.

CXCL13/CXCR5 signalling is pivotal to preserve motor neurons in amyotrophic lateral sclerosis.

BackgroundCXCL13 is a B and T lymphocyte chemokine that mediates neuroinflammation through its receptor CXCR5. This chemokine is highly expressed by motoneurons (MNs) in Amyotrophic Lateral Sclerosis (ALS) SOD1G93A (mSOD1) mice during the disease, particularly in fast-progressing mice. Accordingly, in this study, we investigated the role of this chemokine in ALS.MethodsWe used in vitro and in vivo experimental paradigms derived from ALS mice and patients to investigate the expression level and distribution of CXCL13/CXCR5 axis and its role in MN death and disease progression. Moreover, we compared the levels of CXCL13 in the CSF and serum of ALS patients and controls.FindingsCXCL13 and CXCR5 are overexpressed in the spinal MNs and peripheral axons in mSOD1 mice. CXCL13 inhibition in the CNS of ALS mice resulted in the exacerbation of motor impairment (n = 4/group;Mean_Diff.=27.81) and decrease survival (n = 14_Treated:19.2 ± 1.05wks, n = 17_Controls:20.2 ± 0.6wks; 95% CI: 0.4687–1.929). This was corroborated by evidence from primary spinal cultures where the inhibition or activation of CXCL13 exacerbated or prevented the MN loss. Besides, we found that CXCL13/CXCR5 axis is overexpressed in the spinal cord MNs of ALS patients, and CXCL13 levels in the CSF discriminate ALS (n = 30) from Multiple Sclerosis (n = 16) patients with a sensitivity of 97.56%.InterpretationWe hypothesise that MNs activate CXCL13 signalling to attenuate CNS inflammation and prevent the neuromuscular denervation. The low levels of CXCL13 in the CSF of ALS patients might reflect the MN dysfunction, suggesting this chemokine as a potential clinical adjunct to discriminate ALS from other neurological diseases.FundingVaccinex, Inc.; Regione Lombardia (TRANS-ALS)

Intake of Fermented Dairy Products Induces a Less Pro-Inflammatory Postprandial Peripheral Blood Mononuclear Cell Gene Expression Response than Non-Fermented Dairy Products: A Randomized Controlled Cross-Over Trial.

It is aimed to investigate how intake of high-fat meals composed of different dairy products with a similar fat content affects postprandial peripheral blood mononuclear cell (PBMC) expression of inflammation-related genes, as well as circulating inflammatory markers and metabolites. Healthy subjects (n = 47) consume four different high-fat meals composed of either butter, cheese, whipped cream, or sour cream in a randomized controlled cross-over study. Fasting and postprandial PBMC gene expression, plasma metabolites, and circulating inflammatory markers are measured. Using a linear mixed model, it is found that expression of genes related to lymphocyte activation, cytokine signaling, chemokine signaling, and cell adhesion is differentially altered between the four meals. In general, intake of the fermented products cheese and sour cream reduces, while intake of the non-fermented products butter and whipped cream increases, expression of these genes. Plasma amino acid concentrations increase after intake of cheese compared to the other meals, and the amino acid changes correlate with several of the differentially altered genes. Intake of fermented dairy products, especially cheese, induces a less inflammatory postprandial PBMC gene expression response than non-fermented dairy products. These findings may partly explain inconsistent findings in studies on health effects of dairy products.

The Stat3 inhibitor, S3I-201, downregulates lymphocyte activation markers, chemokine receptors, and inflammatory cytokines in the BTBR T+ Itpr3tf/J mouse model of autism

Autism is a complex neurodevelopmental disorder with a high incidence rate. It is characterized by deficits in communication, a lack of social skills, cognitive inflexibility, and stereotypical behaviors. Autism has been gradually increasing in children over the past several years, without the existence of an effective treatment. BTBR T+ Itpr3tf/J (BTBR) mice serve as an accepted model to evaluate autistic-like behaviors as they display core behavioral symptoms displayed in autism. Previous findings showed that S3I-201, a selective Stat3 inhibitor, can be used to treat neuroinflammation disorders. Previously, we showed that S3I-201 treatment has therapeutic effects on autism-like behaviors, and Th1/Th17 and regulatory T cells in BTBR mice. The objective of the present study was to further explore the role of S3I-201 in BTBR mice, and this was performed by investigating the effects of S3I-201 treatment on lymphocyte activation markers (CD4+CD25+ and CD4+CD69+), chemokine receptors (CD4+CCR6+, CD4+CCR7+, CD4+CXCR4+, and CD4+CXCR5+), and proinflammatory cytokines (CD4+IL-6+ and CD4+TNF-α+) in the spleen cells of BTBR and C57BL/6 (C57) mice. The mRNA and protein expression levels of CD69, CCR6, CCR7, CXCR4, CXCR5, IL-1β, IL-6, and TNF-α were examined in the brain tissues, and in BTBR mice, a significant decrease in CD25, CD69, CCR6, CCR7, CXCR4, CXCR5, IL-6, and TNF-α producing CD4+ T cells was observed. The present findings suggest that treatment with S3I-201 may be a therapeutic approach to improve immune abnormalities in a subgroup of autistic subjects.

Cigarette smoke exposure worsens acute lung injury in antibiotic-treated bacterial pneumonia in mice.

Evidence is accumulating that exposure to cigarette smoke (CS) increases the risk of developing acute respiratory distress syndrome (ARDS). Streptococcus pneumoniae is the most common cause of bacterial pneumonia, which in turn is the leading cause of ARDS. Chronic smokers have increased rates of pneumococcal colonization and develop more severe pneumococcal pneumonia than nonsmokers; yet mechanistic connections between CS exposure, bacterial pneumonia, and ARDS pathogenesis remain relatively unexplored. We exposed mice to 3 wk of moderate whole body CS or air, followed by intranasal inoculation with an invasive serotype of S. pneumoniae. CS exposure alone caused no detectable lung injury or bronchoalveolar lavage (BAL) inflammation. During pneumococcal infection, CS-exposed mice had greater survival than air-exposed mice, in association with reduced systemic spread of bacteria from the lungs. However, when mice were treated with antibiotics after infection to improve clinical relevance, the survival benefit was lost, and CS-exposed mice had more pulmonary edema, increased numbers of BAL monocytes, and elevated monocyte and lymphocyte chemokines. CS-exposed antibiotic-treated mice also had higher serum surfactant protein D and angiopoietin-2, consistent with more severe lung epithelial and endothelial injury. The results indicate that acute CS exposure enhances the recruitment of immune cells to the lung during bacterial pneumonia, an effect that may provide microbiological benefit but simultaneously exposes the mice to more severe inflammatory lung injury. The inclusion of antibiotic treatment in preclinical studies of acute lung injury in bacterial pneumonia may enhance clinical relevance, particularly for future studies of current or emerging tobacco products.

Correlations between CXCL13, IL-24 genes and wrist arthritis.

To investigate the relationship between B lymphocyte chemokine 1 (CXCL13) and interleukin-24 (IL-24) gene and wrist arthritis. A total of 122 cases of patients with wrist arthritis treated in our hospital from May 2013 to April 2016 were randomly selected as wrist arthritis group, while 120 normal subjects were selected as normal control group. Venous blood was collected from all patients in normal control group and wrist arthritis group, respectively. Rheumatoid factor (RF), human C-reactive protein (CRP), and erythrocyte sedimentation rate (ESR) in venous blood were measured. The visual analogue scale (VAS) score was used to statistically analyze the pain of subjects in normal control and wrist arthritis groups; the wrist flexion and extension activities of subjects in normal control group and wrist arthritis group were measured. The expressions of CXCL13 and IL-24 mRNA in synovial tissue of normal control group and wrist arthritis group were detected by reverse transcription-polymerase chain reaction (RT-PCR). Western blotting was used to detect the expressions of CXCL13 and IL-24 in normal control group and wrist arthritis group. The levels of CRP, RF, and ESR in the normal control group were within the normal range, but the levels of CRP, RF, and ESR in the wrist arthritis group were significantly higher than those in the normal control group. VAS scores and joint flexion extension activities in the normal control group were at normal levels. The VAS score of wrist arthritis group was significantly higher than that of the normal control group, and the joint flexion extension activities were significantly lower than that in the normal control group. The results of RT-PCR showed that the expression of CXCL13 mRNA in synovial tissue of wrist arthritis was significantly higher than that in the normal control group, while the expression of IL-24 mRNA in synovial tissue of wrist arthritis was significantly lower than that in normal control tissues. Western blotting showed that the expression of CXCL13 in synovial tissue of wrist arthritis was significantly higher than that in the normal control group, while the expression of IL-24 in synovial tissue of wrist arthritis was significantly lower than that in normal control groups. Analysis of variance showed that the expressions of CXCL13 and IL-24 in the normal control group and wrist arthritis group had statistically significant differences (p<0.01). The abnormal expressions of CXCL13 and IL-24 are closely related to the occurrence and development of wrist arthritis. This study shows that CXCL13 and IL-24 have important research values in wrist arthritis. CXCL13 and IL-24 expressions can be used as new indicators of the diagnosis and treatment of wrist arthritis.

Renal allograft rejection, lymphocyte infiltration, and de novo donor-specific antibodies in a novel model of non-adherence to immunosuppressive therapy

BackgroundNon-adherence has been associated with reduced graft survival. The aim of this study was to investigate the immunological mechanisms underlying chronic renal allograft rejection using a model of non-adherence to immunosuppressive therapy. We used a MHC (major histocompatibility complex) -mismatched rat model of renal transplantation (Brown Norway to Lewis), in which rats received daily oral cyclosporine A. In analogy to non-adherence to therapy, one group received cyclosporine A on alternating days only. Rejection was histologically graded according to the Banff classification. We quantified fibrosis by trichrome staining and intra-graft infiltration of T cells, B cells, and monocytes/macrophages by immunohistochemistry. The distribution of B lymphocytes was assessed using immunofluorescence microscopy. Intra-graft chemokine, chemokine receptor, BAFF (B cell activating factor belonging to the TNF family), and immunoglobulin G transcription levels were analysed by RT-PCR. Finally, we evaluated donor-specific antibodies (DSA) and complement-dependent cytotoxicity using flow cytometry.ResultsAfter 28 days, cellular rejection occurred during non-adherence in 5/6 animals, mixed with humoral rejection in 3/6 animals. After non-adherence, the number of T lymphocytes were elevated compared to daily immunosuppression. Monocyte numbers declined over time. Accordingly, lymphocyte chemokine transcription was significantly increased in the graft, as was the transcription of BAFF, BAFF receptor, and Immunoglobulin G. Donor specific antibodies were elevated in non-adherence, but did not induce complement-dependent cytotoxicity.ConclusionCellular and humoral rejection, lymphocyte infiltration, and de novo DSA are induced in this model of non-adherence.

Selective Expression of CCR10 and CXCR3 by Circulating Human Herpes Simplex Virus-Specific CD8 T Cells

Herpes simplex virus (HSV) infection is restricted to epithelial cells and neurons and is controlled by CD8 T cells. These cells both traffic to epithelial sites of recurrent lytic infection and to ganglia and persist at the dermal-epidermal junction for up to 12 weeks after lesion resolution. We previously showed that cutaneous lymphocyte-associated antigen (CLA), a functional E-selectin ligand (ESL), is selectively expressed on circulating HSV-2-specific CD8 T cells. CLA/ESL mediates adhesion of T cells to inflamed vascular endothelium. Later stages in T-cell homing involve chemokines (Ch) and lymphocyte chemokine receptors (ChR) for vascular wall arrest and diapedesis. Several candidate ChR have been implicated in skin homing. We measured cell surface ChR on HSV-specific human peripheral blood CD8 T cells and extended our studies to HSV-1. We observed preferential cell surface expression of CCR10 and CXCR3 by HSV-specific CD8 T cells compared to CD8 T cells specific for control viruses, Epstein-Barr virus (EBV) and cytomegalovirus (CMV), and compared to bulk memory CD8 T cells. CXCR3 ligand mRNA levels were selectively increased in skin biopsy specimens from persons with recurrent HSV-2, while the mRNA levels of the CCR10 ligand CCL27 were equivalent in lesion and control skin. Our data are consistent with a model in which CCL27 drives baseline recruitment of HSV-specific CD8 T cells expressing CCR10, while interferon-responsive CXCR3 ligands recruit additional cells in response to virus-driven inflammation.IMPORTANCE HSV-2 causes very localized recurrent infections in the skin and genital mucosa. Virus-specific CD8 T cells home to the site of recurrent infection and participate in viral clearance. The exit of T cells from the blood involves the use of chemokine receptors on the T-cell surface and chemokines that are present in infected tissue. In this study, circulating HSV-2-specific CD8 T cells were identified using specific fluorescent tetramer reagents, and their expression of several candidate skin-homing-associated chemokine receptors was measured using flow cytometry. We found that two chemokine receptors, CXCR3 and CCR10, are upregulated on HSV-specific CD8 T cells in blood. The chemokines corresponding to these receptors are also expressed in infected tissues. Vaccine strategies to prime CD8 T cells to home to HSV lesions should elicit these chemokine receptors if possible to increase the homing of vaccine-primed cells to sites of infection.

An exploratory genome-wide analysis of genetic risk for alcoholic hepatitis

Objectives: To elucidate the genetic variability between heavy drinkers with and without alcoholic hepatitis (AH).Materials and methods: An exploratory genome-wide association study (GWAS; NCT02172898) was conducted comparing 90 AH cases with 93 heavy drinking matched controls without liver disease in order to identify variants or genes associated with risk for AH. Individuals were genotyped using the multi-ethnic genotyping array, after which the data underwent conventional quality control. Using bioinformatics tools, pathways associated with AH were explored on the basis of individual variants, and based on genes with a higher ’burden’ of functional variation.Results: Although no single variant reached genome-wide significance, an association signal was observed for PNPLA3 rs738409 (p = .01, OR 1.9, 95% CI 1.1–3.1), a common single nucleotide polymorphism that has been associated with a variety of liver-related pathologies including alcoholic cirrhosis. Using the improved gene set enrichment analysis for GWAS tool, it was shown that, based on the single variants’ trait-association p-values, multiple pathways were associated with risk for AH with high confidence (false discovery rate [FDR] < 0.05), including several pathways involved in lymphocyte activation and chemokine signaling, which coincides with findings from other research groups. Several Tox Functions and Canonical Pathways were highlighted using Ingenuity Pathway Analysis, with an especially conspicuous role for pathways related to ethanol degradation, which is not surprising considering the phenotype of the genotyped individuals.Conclusion: This preliminary analysis suggests a role for PNPLA3 variation and several gene sets/pathways that may influence risk for AH among heavy drinkers.

Super Cross-Presentation of Tumor Antigens to Elicit Anti-Lymphoma Immunity By Synthetic Design of an Anti-Phosphatidylserine Bridge Protein

Cell loss by apoptosis is a common feature in tumors. Dying tumor cells induce immune tolerance within the tumor microenvironment largely through highly conserved homeostatic clearance programs that restore tissue immune homeostasis and contribute to the formation of an immunosuppressive niche. The translocation of phosphatidylserine (PS) on cellular membranes, during the initial phases of apoptosis, functions as a recognition and removal signal that limits the immunogenicity of cell death. We examined whether altering clearance of dying cancer cells to elicit inflammatory turnover potentiates immune responses against lymphoma cells. To remove inhibitory signals in the homeostatic clearance pathway we utilized a molecular bridge scaffold to engineer a modified phosphatidylserine bridge protein (FA58C2-hIgG1 or C2-hIgG1) that works as a bridge between apoptotic cells expressing aminophospholipids and phagocytes bearing Fc receptors. In vitro administration of C2-hIgG1 to murine bone marrow derived macrophages promotes engulfment of apoptotic murine lymphoma cells (38C13 cell line) and ablates the secretion of the anti-inflammatory cytokine interleukin-10 (IL-10) and suppression of pro-inflammatory cytokines tumor necrosis factor (TNF-α) and IL-12p40 to the presence of apoptotic cells. Similarly, uptake of C2-hIgG1 treated lymphoma cells triggers upregulation of the costimulatory markers CD80, CD86, and MHC class II on macrophages and promotes secretion of Th1-recruiting lymphocyte chemokines CXCL9, CXCL10, and CCL5. Accordingly, in vivo administration of C2-hIgG1 partially restores immune responses to dead lymphoma cells in antigen cross presentation assays and promotes recruitment and retention of tumor antigen specific CD8+ T cells, dendritic cells, and natural killer cells into tumors. These effects combine to elicit anti-lymphoma immunity, improve responses to immune checkpoint inhibitors, and enhance the effectiveness of adoptive T cell transfers using engineered T Cell Receptors (TCRs) but not CD19-directed chimeric antigen receptor engineered (CAR-T) T cells. DisclosuresNo relevant conflicts of interest to declare.

The effect of sex on immune cells in healthy aging: elderly women have more robust natural killer (NK) lymphocytes than do elderly men

Abstract Gender differences in blood leukocytes and immune responses have been reported, but there is limited knowledge of gender differences in elderly humans. We compared monocyte and lymphocyte subsets, along with soluble immune mediators in healthy men and women over the age of 70. We also measured natural killer (NK) lymphocyte cytotoxic granule exocytosis, chemokine synthesis, and cytokine synthesis in response to a variety of stimuli. We found that elderly women had significantly more circulating B cells than men, whereas men had more CD4 central memory T cells. Men also had higher monocyte levels. Women had a higher ratio of immature CD56bright NK cells to mature CD56dim NK cells, indicating a gender difference in NK cell maturation in the elderly. Moreover, compared with NK cells from men, mature NK cells from women had more vigorous cytotoxic granule responses to K562 leukemia cells and IFN-g responses to NKp46 crosslinking. Female mature NK cells were also more likely to produce MIP-1b than male mature NK cells in response to a variety of stimuli. We also measured soluble factors that affect immune cells or are produced by immune cells. Plasma adiponectin levels were higher in women, plasma levels of retinol-binding protein 4 were higher in men, but there was no significant gender difference in C-reactive protein, IL-15, or sphingosine-1-phosphate. These data support the hypothesis that gender influences immunity in general, and NK cell activity in particular, in elderly humans.

Serum amyloid A induces T lymphocyte migration through chemokine receptor CXCR3

Abstract Serum amyloid A (SAA) is a major acute-phase protein with cytokine/chemokine-like properties. It can be expressed at sites of inflammation and induce the migration of immune cells including T lymphocytes to the inflammatory sites. SAA plasma concentration can be 1000 times higher in patients with autoimmune diseases such rheumatoid arthritis, than in healthy people. However, its precise role in inflammation and infection remains unclear. SAA is known to bind to several receptors such as FPRL1, TLR2, and TLR4, etc. It is possible that SAA also use other chemokine receptors to recruit immune cells during the inflammation. In our previous study, we used a mathematical approach called Natural Vector Method (NVM) to analyze the genetic information of all major human chemokines and SAA and found that chemokine receptor CXCR3 may be a new receptor for SAA. We reported here that SAA indeed can induce MAP kinase ERK activation in the CXCR3 over-expressed cell line, and trigger the receptor-mediated intracellular calcium mobilization. Our chemotaxis experiments also showed that SAA promotes CXCR3-overexpressed cell migration. These findings indicate that SAA may use lymphocyte chemokine receptor CXCR3 to recruit activated lymphocytes to the inflammation sites. We will further investigate the signaling pathways involved in the CXCR3 mediated SAA-induced cell migration. We hope that our research work not only helps to understand the function of SAA, but also introduces a new way of finding receptors for the endogenous molecules. [This study is supported by NSF grant DMS-1119612 and NIH grant 1SC3GM098180 awarded to Rong L He]

The effect of sex on immune cells in healthy aging: Elderly women have more robust natural killer lymphocytes than do elderly men

Immune gender differences have been reported, but are little studied in elderly humans. We compared monocyte and lymphocyte subsets, along with soluble immune mediators in healthy men and women over the age of 70. We also measured natural killer (NK) lymphocyte cytotoxic granule exocytosis, chemokine synthesis, and cytokine synthesis in response to a variety of stimuli. Elderly women had significantly more circulating B cells than men, whereas men had more CD4 central memory T cells and higher monocyte levels. Plasma adiponectin levels were higher in women, plasma retinol-binding protein 4 levels were higher in men, but there were no significant gender differences in C-reactive protein, IL-15, or sphingosine-1-phosphate. Women had a higher ratio of immature CD56bright NK cells to mature CD56dim NK cells, indicating a gender difference in NK cell maturation in the elderly. Comparing sexes, female mature NK cells had more vigorous cytotoxic granule responses to K562 leukemia cells and IFN-γ responses to NKp46 crosslinking. Moreover, female NK cells were more likely to produce MIP-1β in response to a variety of stimuli. These data show that gender influences NK cell activity in elderly humans.

Healthy human second‐trimester fetal skin is deficient in leukocytes and associated homing chemokines

The lack of immune cells in mid-gestational fetal skin is often mentioned as a key factor underlying scarless healing. However, the scarless healing ability is conserved until long after the immune system in the fetus is fully developed. Therefore, we studied human second-trimester fetal skin and compared the numbers of immune cells and chemokine levels from fetal skin with adult skin. By using immunohistochemistry, we show that healthy fetal skin contains significant lower numbers of CD68(+) -macrophages, Tryptase(+) -mast cells, Langerin(+) -Langerhans cells, CD1a(+) -dendritic cells, and CD3(+) -T cells compared to adult skin. Staining with an early lineage leukocyte marker, i.e., CD45, verified that the number of CD45(+) -immune cells was indeed significantly lower in fetal skin but that sufficient numbers of immune cells were present in the fetal lymph node. No differences in the vascular network were observed between fetal and adult skin. Moreover, significant lower levels of lymphocyte chemokines CCL17, CCL21, and CCL27 were observed in fetal skin. However, levels of inflammatory interleukins such as IL-6, IL-8, and IL-10 were undetectable and levels of CCL2 were similar in healthy fetal and adult skin. In conclusion, this study shows that second-trimester fetal skin contains low levels of immune cells and leukocyte chemokines compared to adult skin. This immune cell deficiency includes CD45(+) leukocytes, despite the abundant presence of these cells in the lymph node. The immune deficiency in healthy second-trimester fetal skin may result in reduced inflammation during wound healing, and could underlie the scarless healing capacities of the fetal skin.