Genes In CD4 Research Articles

Single-cell landscape of dynamic changes in CD8+ T cells, CD4+ T cells and exhausted T cells in hepatocellular carcinoma

Hepatocellular carcinoma has a high incidence and poor prognosis. In this study, we investigated the value of T-cell-related genes in prognosis by single-cell sequencing data in hepatocellular carcinoma. Twelve cases of hepatocellular carcinoma single-cell sequencing were included in the study. The high dimensional weighted gene co-expression network analysis (hdWGCNA) was used to identify gene modules associated with CD4+ T cells, CD8+ T cells and exhausted T cells. Altered signaling pathway activity in exhausted T cells was uncovered by the AUCell algorithm. xCELL, TIMER, QUANTISEQ, CIBERSORT and CIBERSORT-abs were performed to explore immune cell infiltration. Immune checkpoint inhibitor genes and TIDE methods were used to predict immunotherapy response. Finally, immunohistochemistry and real-time PCR were used to verify gene expression. The hdWGCNA algorithm identified 40 genes strongly associated with CD4+ T cells, CD8+ T cells and exhausted T cells. Seven genes were finally selected for transcriptome data modeling. The results of the three independent datasets suggested that the model had strong prognostic value. Model genes were critical factors influencing CD4+ T cell and CD8+ T cell infiltration in patients. The efficacy of PD-1 immunotherapy was higher in patients belonging to the low-risk group. Alterations in signaling pathways’ activity within exhausted T cells were crucial factors contributing to the decline in immune function. Differential expression of seven genes in CD8+ T cells, CD4+ T cells and exhausted T cells were key targets for improving immunotherapy response in HCC.

Combining Single-Cell RNA Sequencing and Mendelian Randomization to Explore Novel Drug Targets for Parkinson's Disease.

Neuroinflammation is a key pathological factor of PD, and T cells play a central role in neuroinflammatory progression. However, the causal effect of T cell-related genes on the risk of PD is still unclear. We explored single-cell RNA sequencing (scRNA-Seq) datasets of the peripheral blood T cells of PD patients and healthy controls, and screened the differentially expressed genes (DEGs) in the cytotoxic CD4 + T cells relative to the other T cell subsets. Pseudo-time series analysis, cell-cell communication analysis, and metabolic pathway analysis was performed for the cytotoxic CD4 + T cells. The DEGs were also functionally annotated through GO and KEGG pathway enrichment analyses. The MR approach was used to establish causal effects of the DEGs (exposure) on PD risk (outcome), and explore new drug targets for PD. The findings of MR analysis were further validated by Steiger filtering, bidirectional MR, Bayesian colocalization analysis, and phenotype scanning, and the GWAS data from an independent PD case-control cohort was used for external validation of the results. Finally, differences in gene expression between PD patients and healthy controls were further validated in scRNA-Seq and bulk transcriptome sequencing data. We found that increased expression of IL-32, GNLY, MT2A, and ARPC2 was significantly associated with a higher risk of PD. In contrast, the increase in ARRB2 was closely related to a lower risk of PD. IL32, GNLY, MT2A, ARRB2, and ARPC2 are the causal genes and potential drug targets of PD. Cytotoxic CD4 + T cells are likely the key effectors of PD-related neuroinflammation. These findings provide new insights into the pathogenesis and treatment options for PD, and further research and clinical trials based on the five potential drug targets and neuroinflammation are necessary.

Characterizing immune biomarkers and effector CD8+ T-cell exhaustion in pancreatic adenocarcinoma via single-cell RNA sequencing profiling

Aim: Pancreatic ductal adenocarcinoma (PDAC) is a leading cause of cancer-related mortality and is characterized by T-cell exhaustion, particularly in effector CD8+ T-cells. This exhaustion, driven by persistent immunosuppressive signals in the tumor microenvironment, impairs immune function and hinders effective immunotherapy. This study aimed to identify key exhaustion-related marker genes in CD8+ T-cells linked to PDAC and assess the potential of repurposing anti-inflammatory drugs to counteract T-cell exhaustion and enhance immune responses against PDAC. Methods: We employed a multi-omics approach, integrating single-cell RNA sequencing data with whole genome sequencing to identify dysregulated exhaustion-related immune markers in CD8+ T-cells in PDAC. We examined gene expression profiles and conducted functional enrichment analysis to evaluate their roles in immune exhaustion. We analyzed mutations in the shortlisted biomarkers from The Cancer Genome Atlas (TCGA) and performed in silico mutational analysis using Maestro to evaluate the impact of an IL7R mutation (K110N) on protein function. Virtual screening using a deep learning framework, GNINA, explored the inhibitory features of the anti-inflammatory drugs oxaprozin and celecoxib on IL7R. Results: Key dysregulated exhaustion-related immune markers were identified including PRF1, GZMA, CD8A, CD3D, NKG7, IL7R, and IL2RG. Pathway enrichment analysis indicated significant involvement in T-cell receptor signaling, Th1 and Th2 differentiation, and Th17 differentiation pathways, correlating with reported poor survival outcomes in PDAC patients. Mutational analysis of IL7R revealed a likely pathogenic mutation (K110N) located in the IL-7Ralpha fibronectin type III domain. Drug repurposing of oxaprozin and celecoxib showed favorable binding interactions with both wild and mutant IL7R proteins. Conclusions: The K110N mutation, despite not causing significant structural changes, may impact T-cell and B-cell homeostasis and development. Our findings suggest that oxaprozin and celecoxib could effectively inhibit T-cell exhaustion through favorable interactions with IL7R. Further clinical studies are necessary to validate the therapeutic potential of these anti-inflammatory drugs in enhancing immune responses in pancreatic cancer.

Heightened expression of type I interferon signaling genes in CD4+ T cells from acutely HIV-1-infected women is associated with lower viral loads.

Sex differences play a role in the pathogenesis of a number of viral diseases. In HIV-1, several studies have reported that chronically infected women have significantly lower plasma viremia than men, although the exact mechanism by which this occurs has yet to be identified. We have performed bulk RNA-seq experiments comparing gene expression between CD4+ T cells from acutely HIV-1-infected men and women in Zambia, because we observe lower viral load (VL) despite higher CD4+ T-cell activation in these women during acute/early infection. In a univariate analysis, we have identified a number of differentially expressed genes in naïve, central memory, and effector memory CD4 T cells of women with consistent elevated expression of genes linked to type 1 interferon (IFN) signaling. Moreover, after controlling for differences in VL and CD4+ T-cell count, genes within the type I IFN signaling pathway were further shown to be more highly expressed in women, whereas those genes more highly expressed in men showed no such enrichment. A subset of the genes highly expressed in women was further identified, including several involved in type I IFN signaling in response to viral infections (IRF7, DDX58, SAMHD1, OAS2, and TRIM14), that both are more highly expressed in CD4+ T cells from women and negatively correlated with VL, suggesting that they play a role in the comparative control of VL observed in women.

Convergent and divergent immune aberrations in COVID-19, post-COVID-19-interstitial lung disease, and idiopathic pulmonary fibrosis.

We aimed to study transcriptional and phenotypic changes in circulating immune cells associated with increased risk of mortality in COVID-19, resolution of pulmonary fibrosis in post-COVID-19-interstitial lung disease (ILD), and persistence of idiopathic pulmonary fibrosis (IPF). Whole blood and peripheral blood mononuclear cells (PBMCs) were obtained from 227 subjects with COVID-19, post-COVID-19 interstitial lung disease (ILD), IPF, and controls. We measured a 50-gene signature (nCounter, Nanostring) previously found to be predictive of IPF and COVID-19 mortality along with plasma levels of several biomarkers by Luminex. In addition, we performed single-cell RNA sequencing (scRNA-seq) in PBMCs (10x Genomics) to determine the cellular source of the 50-gene signature. We identified the presence of three genomic risk profiles in COVID-19 based on the 50-gene signature associated with low-, intermediate-, or high-risk of mortality and with significant differences in proinflammatory and profibrotic cytokines. Patients with COVID-19 in the high-risk group had increased expression of seven genes in CD14+HLA-DRlowCD163+ monocytic-myeloid-derived suppressive cells (7Gene-M-MDSCs) and decreased expression of 43 genes in CD4 and CD8 T cell subsets. The loss of 7Gene-M-MDSCs and increased expression of these 43 genes in T cells was seen in survivors with post-COVID-19-ILD. On the contrary, patients with IPF had low expression of the 43 genes in CD4 and CD8 T cells. Collectively, we showed that a 50-gene, high-risk profile, predictive of IPF and COVID-19 mortality is characterized by a genomic imbalance in monocyte and T-cell subsets. This imbalance reverses in survivors with post-COVID-19-ILD highlighting genomic differences between post-COVID-19-ILD and IPF.NEW & NOTEWORTHY Changes in the 50-gene signature, reflective of increase in CD14+HLA-DRlowCD163+ monocytes and decrease in CD4 and CD8 T cells, are associated with increased mortality in COVID-19. A reversal of this pattern can be seen in post-COVID-19-ILD, whereas its persistence can be seen in IPF. Modulating the imbalance between HLA-DRlow monocytes and T cell subsets should be investigated as a potential strategy to treat pulmonary fibrosis associated with severe COVID-19 and progressive IPF.

Transcriptomic HIV-1 reservoir profiling reveals a role for mitochondrial functionality in HIV-1 latency.

Identifying cellular and molecular mechanisms maintaining HIV-1 latency in the viral reservoir is crucial for devising effective cure strategies. Here we developed an innovative flow cytometry-fluorescent in situ hybridization (flow-FISH) approach for direct ex vivo reservoir detection without the need for reactivation using a combination of probes detecting abortive and elongated HIV-1 transcripts. Our flow-FISH assay distinguished between HIV-1-infected CD4+ T cells expressing abortive or elongated HIV-1 transcripts in PBMC from untreated and ART-treated PWH from the Amsterdam Cohort Studies. This flow-FISH method was employed to isolate CD4+ T cells expressing abortive or elongated HIV-1 transcripts from five ART-naïve PWH for transcriptomic analysis by 3' RNA sequencing. Supervised cluster analysis identified several differentially expressed mitochondrial genes in infected CD4+ T cells with abortive HIV-1 transcripts compared to cells containing elongated HIV-1 transcripts. Notably, enhancing mitochondrial function induced HIV-1 transcription in PBMC from PWH. Our data strongly suggests that cellular metabolism is involved in maintaining HIV-1 latency and show that improving mitochondrial functions induces HIV-1 transcriptional activity in PWH. These findings underline the relevance of metabolic regulation in HIV-1 infection, and support the development of strategies modulating immunometabolism to target viral latency.

Association Between Common Variants in the LAG3/CD4 Genes and Risk for Essential Tremor.

Many clinical, neuroimaging, neuropathological, epidemiological, and genetic data suggest a relationship between essential tremor (ET) and Parkinson's disease (PD). Several hypothesis-based gene association studies attempted to find a genetic association between these diseases. Recent case-control association studies in Chinese and Spanish populations showed a marginal association between the CD4 rs1922452 and CD4 rs951818 single nucleotide variants (SNVs) and the risk of PD. The proteins encoded by the CD4 and LAG3 genes have an important role in modulating inflammatory responses, and some recent data associated inflammatory markers to ET. This study investigates a possible association between the most common SNVs in the LAG3/CD4 genes and the risk of ET in the Spanish Caucasian population. We genotyped 267 patients diagnosed with familial ET and 270 age- and sex-matched controls using specific TaqMan assays for CD4 rs1922452, CD4 rs951818, and LAG3 rs870849 variants. We found a decreased risk for ET in carriers of the LAG3 rs870849 C/C genotype and the LAG3 rs870849C allelic variant exclusively in men. The mean age of onset of ET was not related to any of the variants studied. These data suggest no association of the gene variants studied with the overall risk for ET, except for a slight decrease in risk in male ET patients carrying the variant LAG3 rs870849C. However, such an association lost significance after correcting for multiple comparisons.

Comprehensive analysis of differentially expressed genes in toll-like receptor signalling pathway: Insights into new-onset microscopic polyangiitis.

We aimed to elucidate the potential contributions of the toll-like receptor (TLR) signalling pathway and identify promising candidates for new-onset microscopic polyangiitis (MPA) using integrated bioinformatics analysis. A PCR array was used to determine the expression profiles of TLR signalling-related genes in CD4+T lymphocytes of individuals with new-onset MPA and healthy controls. Four genes were selected for validation through real-time quantitative polymerase chain reaction (RT-qPCR). Followed by functional enrichment and pathway analysis, we identified the hub genes with cytoHubba. The differentially expressed miRNAs of the target genes were subsequently predicted and visualized via Cytoscape. Finally, these candidates were validated and evaluated at the expression level and for diagnostic value in public databases. Nineteen differentially expressed genes were screened, and the levels of the validated genes detected using RT‒qPCR were consistent with the findings obtained through the PCR array. The significantly enriched signalling pathways involved were TLR signalling pathway, IL-17 signalling pathway, and NF-κB signalling pathway. Nine hub genes and nine key miRNAs were identified. Furthermore, analysis of three distinct gene expression datasets validated several key genes (TLR4, MYD88, IRF1, CXCL10, CXCL8, and CSF2), showing significant differences between groups and strong diagnostic value, especially TLR4, MYD88, and IRF1. Interestingly, in contrast to the validation results, our results showed that CXCL10 and CXCL8 expression levels were markedly lower, but CSF2 was highly expressed in patients with MPA compared to controls. Aberrant expression of TLRs may occur in CD4+ T lymphocytes of patients with new-onset MPA, offering insights into the pathogenesis as well as potential biomarkers and novel therapeutic targets.

Kinetics of viral hemorrhagic septicemia virus (VHSV) and immune response in olive flounder (Paralichthys olivaceus) at altered temperature

Viral hemorrhagic septicemia virus (VHSV) is a pathogen that causes hemorrhagic septicemia in olive flounder at water temperatures below 15 °C, leading to symptoms such as abdominal swelling due to ascites and muscle hemorrhaging, and in severe cases, mortality. In this study, we investigated the proliferation of NCCs, the transcriptional analysis of CD4 and CD8, and the expression of pro-inflammatory cytokines (IL-1β, IL6, TNFα) and the cytokines (IL12, IL15, IFN-1β, IFNγ) involved in cytotoxic cell activation in the kidney of olive flounder during VHSV infection at suboptimal temperature (17 °C) and following a shift to optimal temperature (10 °C). Following viral infection, the population of NCCs and CD8 gene expression steadily increased. Meanwhile, IL12, IL15, and IFN-1β levels exhibited an early surge, reaching their peak within 1–2 days post-challenge at 17 °C. Notably, in olive flounder that were re-challenged with VHSV at 10 °C after an initial challenge at 17 °C, there was a rapid and robust expansion of NCCs and increased CD4 and CD8 gene expression, demonstrating the mounting of immune memory by NCCs and an efficient adaptive immune response from CD4+ and CD8+ T cells against VHSV infection.

Single-cell RNA sequencing uncovers molecular mechanisms of intravenous immunoglobulin plus methylprednisolone in Kawasaki disease: attenuated monocyte-driven inflammation and improved NK cell cytotoxicity.

Intravenous immunoglobulin (IVIG) plus methylprednisolone as initial intensive therapy or additional therapy in Kawasaki disease (KD) has been used in clinical practice. However, its molecular and cellular mechanism is unclear. We performed single-cell analysis on 14 peripheral blood mononuclear cell (PBMC) samples obtained from 7 KD patients who received either IVIG monotherapy or IVIG plus methylprednisolone therapy. This encompassed 4 samples from KD patients collected before and after IVIG treatment, as well as 3 samples from KD patients before and after IVIG plus methylprednisolone therapy. Both IVIG monotherapy and IVIG plus methylprednisolone therapy can increase lymphocyte counts (e.g. CD4+T, CD8+T, and gdT cells) to address lymphopenia. They can also decrease monocyte counts and repress the expression of S100A12, NLRP3, and genes associated with immune-cell migration in monocytes. IVIG combined with methylprednisolone downregulates more monocyte-driven inflammatory pathways than IVIG alone. Additionally, this combination uniquely enhances NK cell cytotoxicity by modulating receptor homeostasis, while significantly upregulating interferon-related genes in CD4+ T cells, CD8+ T cells, and B cells, particularly type I interferons. The combination of IVIG with methylprednisolone attenuated monocyte-driven inflammation and improved NK cell cytotoxicity which might provide clues for pediatricians to consider treatment options for children with KD. Whether the monocyte-driven hyperinflammatory state and NK cell function can be indicators for the clinical choice of IVIG with methylprednisolone therapy in KD needs further investigation.

Immunoprotective effect of recombinant Lactobacillus plantarum expressing largemouth bass virus MCP on largemouth bass

Largemouth bass virus (LMBV) is an infectious pathogen that causes high mortality rates in largemouth bass, and outbreaks of this virus can significantly harm the aquaculture industry. Currently, no vaccine has been developed that can effectively prevent the transmission of LMBV. In this study, we constructed a recombinant Lactobacillus plantarum (L. plantarum) strain capable of expressing the MCP gene of LMBV and displaying this protein on its surface; then, we evaluated the immunoprotective effect of this recombinant bacterium on largemouth bass. Western blotting, immunofluorescence, and flow cytometry confirmed that MCP was successfully expressed and anchored on the surfaces of NC8 cells. Immunization of largemouth bass with NC8-pSIP409-pgsA′-MCP via the oral feeding route induced CD4, CD8, IL-1β, and IL-6 gene expression. In addition, NC8-pSIP409-pgsA′-MCP at different CFUs increased the survival of largemouth bass after LMBV infection; in particular, NC8-pSIP409-pgsA′-MCP (109 CFU) resulted in approximately 30 % survival. NC8-pSIP409-pgsA′-MCP immunization alleviated the pathological changes in the liver and spleen, exerting a more advantageous protective effect. These data suggest that the recombinant L. plantarum strain NC8-pSIP409-pgsA′-MCP can increase the resistance of largemouth bass to LMBV infection and that this strain is a promising candidate oral vaccine for the prevention of LMBV infection.

Similar humoral responses but distinct CD4+ T cell transcriptomic profiles in older adults elicited by MF59 adjuvanted and high dose influenza vaccines.

Older age (≥ 65 years) is associated with impaired responses to influenza vaccination, leading to the preferential recommendation of MF59-adjuvanted (MF59Flu) or high-dose (HDFlu) influenza vaccines for this age group in the United States. Herein, we characterized transcriptomic profiles of CD4+ T cells isolated from 234 recipients (≥ 65 years) of either MF59Flu or HDFlu vaccine, prior to vaccination and 28 days thereafter. We identified 412 and 645 differentially expressed genes (DEGs) in CD4+ T cells of older adults after receiving MF59Flu and HDFlu, respectively. DEGs in CD4+ T cells of MF59Flu recipients were enriched in 14 KEGG pathways, all of which were downregulated. DEGs in CD4+ T cells of HDFlu recipients were enriched in 11 upregulated pathways and 20 downregulated pathways. CD4+ T cells in both vaccine groups shared 50 upregulated genes and 75 downregulated genes, all of which were enriched in 7 KEGG pathways. The remaining 287 and 520 DEGs were specifically associated with MF59Flu and HDFlu, respectively. Unexpectedly, none of these DEGs was significantly correlated with influenza A/H3N2-specific HAI titers, suggesting these DEGs at the individual level may have a limited role in protection against influenza. Our findings emphasize the need for further investigation into other factors influencing immunity against influenza in older adults.

Gene regulatory network inference from CRISPR perturbations in primary CD4+ T cells elucidates the genomic basis of immune disease

The effects of genetic variation on complex traits act mainly through changes in gene regulation. Although many genetic variants have been linked to target genes in cis, the trans-regulatory cascade mediating their effects remains largely uncharacterized. Mapping trans-regulators based on natural genetic variation has been challenging due to small effects, but experimental perturbations offer a complementary approach. Using CRISPR, we knocked out 84 genes in primary CD4+ Tcells, targeting inborn error of immunity (IEI) disease transcription factors (TFs) and TFs without immune disease association. We developed a novel gene network inference method called linear latent causal Bayes (LLCB) to estimate the network from perturbation data and observed 211 regulatory connections between genes. We characterized programs affected by the TFs, which we associated with immune genome-wide association study (GWAS) genes, finding that JAK-STAT family members are regulated by KMT2A, an epigenetic regulator. These analyses reveal the trans-regulatory cascades linking GWAS genes to signaling pathways.

IL-7Rα on CD4+ T cells is required for their survival and the pathogenesis of experimental autoimmune encephalomyelitis

BackgroundThe IL-7 receptor alpha (IL-7Rα) binds both IL-7 and thymic stromal lymphopoietin (TSLP). IL-7Rα is essential for the development and survival of naive CD4+ T cells and their differentiation to effector/memory CD4+ T cells. Mice lacking IL-7Rα have severe lymphopenia and are resistant to experimental autoimmune encephalomyelitis (EAE), a model for multiple sclerosis. However, it has been reported that IL-7Rα on peripheral CD4+ T cells is disposable for their maintenance and EAE pathogenesis, which does not align with the body of knowledge on the role of IL-7Rα in the biology of CD4+ T cells. Given that a definitive study on this important topic is lacking, we revisited it using a novel approach, an inducible knockout of the IL-7Rα gene in CD4+ T cells.MethodsWe generated Il7rafl/fl/CD4CreERT2 double transgenic mouse line (henceforth CD4ΔIl7ra), susceptible to tamoxifen-induced knockout of the IL-7Rα gene in CD4+ T cells. CD4ΔIl7ra mice were immunized with MOG35 − 55 for EAE induction and monitored for disease development. The expression of IL-7Rα, CD4+ T cell numbers, and MOG35 − 55-specific CD4+ T cell response was evaluated in the central nervous system (CNS) and lymphoid tissues by flow cytometry. Additionally, splenocytes of CD4ΔIl7ra mice were stimulated with MOG35 − 55 to assess their proliferative response and cytokine production by T helper cells.ResultsLoss of IL-7Rα from the surface of CD4+ T cells in CD4ΔIl7ra mice was virtually complete several days after tamoxifen treatment. The loss of IL-7Rα in CD4+ T cells led to a gradual and substantial decrease in their numbers in both non-immunized and immunized CD4ΔIl7ra mice, followed by slow repopulation up to the initial numbers. CD4ΔIl7ra mice did not develop EAE. We found a decrease in the total numbers of TNF-, IFN-γ-, IL-17 A-, and GM-CSF-producing CD4+ T cells and regulatory T cells in the spleens and CNS of immunized CD4ΔIl7ra mice. Tracking MOG35 − 55-specific CD4+ T cells revealed a significant reduction in their numbers in CD4ΔIl7ra mice and decreased proliferation and cytokine production in response to MOG35 − 55.ConclusionOur study demonstrates that IL-7Rα on peripheral CD4+ T cells is essential for their maintenance, immune response, and EAE pathogenesis.

Effects of Zinc Oxide Nanoparticles (ZnO NPs) on Growth, Immune Responses and Histopathological Alterations in Asian Seabass (Lates calcarifer, Bloch 1790) under Low-Salinity Conditions.

In the present study, Asian seabass (Lates calcarifer, Bloch) fingerings were used as an animal model to investigate the toxicological effects of zinc oxide nanoparticles (ZnO NPs) under 5 ppt estuarine conditions. The fish were exposed to 0, 1, 5 or 50 ppm ZnO NPs for 8 weeks. It was found that ZnO NP concentrations of 5-50 ppm negatively affected several growth rate parameters, such as the weight and total length of the fish. Additionally, 5 and 50 ppm ZnO NPs led to 32.55% and 100% mortality, respectively, after 8 weeks after exposure (WAE). Furthermore, compared with the control, exposure to 1-50 ppm ZnO NPs strongly affected hematological indices, such as total blood cells, red blood cells, leukocytes and hematocrit, and suppressed lysozyme activity, superoxide anion production and bactericidal activity. High Zn concentrations accumulated in the head kidney, gills and liver, whereas low levels were detected in the gut, skin and muscle. Expression analysis of immune-related genes via quantitative real-time RT-PCR revealed that 5 and 50 ppm ZnO NPs significantly upregulated the cc and cd4 genes at 1 WAE. In contrast, 50 ppm ZnNPs downregulated the expression levels of the cd8, cc, hsp70, hsp90, tcrα, lyz and igmh genes at 1 WAE (p < 0.05). Finally, at 8 WAE, histopathological analysis revealed that 5 and 50 ppm ZnO NPs severely induced alterations in the head kidney, gills and liver.

Single-cell RNA sequencing analysis of peripheral blood mononuclear cells in PD-1-induced renal toxicity in patients with lung cancer

BackgroundAlthough the patient survival rate for many malignancies has been improved with immune checkpoint inhibitors (ICIs), some patients experience various immune-related adverse events (irAEs). IrAEs impact several organ systems, including the kidney. With anti-programmed cell death protein 1 (PD-1) therapy (pembrolizumab), kidney-related adverse events occur relatively rarely compared with other irAEs. However, the occurrence of AKI usually leads to anti-PD-1 therapy interruption or discontinuation. Therefore, there is an urgent need to clarify the mechanisms of renal irAEs (R-irAEs) to facilitate early management. This study aimed to analyse the characteristics of peripheral blood mononuclear cells (PBMCs) in R-irAEs.MethodsPBMCs were collected from three patients who developed R-irAEs after anti-PD-1 therapy and three patients who did not. The PBMCs were subjected to scRNA-seq to identify cell clusters and differentially expressed genes (DEGs). Kyoto Encyclopedia of Genes and Genomes (KEGG) and gene ontology (GO) enrichment analyses were performed to investigate the most active biological processes in immune cells.ResultsFifteen cell clusters were identified across the two groups. FOS, RPS26, and JUN were the top three upregulated genes in CD4+ T cells. The DEGs in CD4+ T cells were enriched in Th17 differentiation, Th1 and Th2 cell differentiation, NF-kappa B, Nod-like receptor, TNF, IL-17, apoptosis, and NK cell-mediated cytotoxicity signaling pathways. RPS26, TRBV25-1, and JUN were the top three upregulated genes in CD8+ T cells. The DEGs in CD8+ T cells were enriched in Th17 cell differentiation, antigen processing and presentation, natural killer cell-mediated cytotoxicity, the intestinal immune network for IgA production, the T-cell receptor signalling pathway, Th1 and Th2 cell differentiation, the phagosome, and cell adhesion molecules.ConclusionsIn conclusion, R-irAEs are associated with immune cell dysfunction. DEGs and their enriched pathways identified in CD4+ T cells and CD8+ T cells play important roles in the development of renal irAEs related to anti-PD-1 therapy. These findings offer fresh perspectives on the pathogenesis of renal damage caused by anti-PD-1 therapy.

The Oncoprotein Fra-2 Drives the Activation of Human Endogenous Retrovirus Env Expression in Adult T-Cell Leukemia/Lymphoma (ATLL) Patients.

Human endogenous retroviruses (HERVs) are retroviral sequences integrated into 8% of the human genome resulting from ancient exogenous retroviral infections. Unlike endogenous retroviruses of other mammalian species, HERVs are mostly replication and retro-transposition defective, and their transcription is strictly regulated by epigenetic mechanisms in normal cells. A significant addition to the growing body of research reveals that HERVs' aberrant activation is often associated with offsetting diseases like autoimmunity, neurodegenerative diseases, cancers, and chemoresistance. Adult T-cell leukemia/lymphoma (ATLL) is a very aggressive and chemoresistant leukemia caused by the human T-cell leukemia virus type 1 (HTLV-1). The prognosis of ATLL remains poor despite several new agents being approved in the last few years. In the present study, we compare the expression of HERV genes in CD8+-depleted PBMCs from HTLV-1 asymptomatic carriers and patients with acute ATLL. Herein, we show that HERVs are highly upregulated in acute ATLL. Our results further demonstrate that the oncoprotein Fra-2 binds the LTR region and activates the transcription of several HERV families, including HERV-H and HERV-K families. This raises the exciting possibility that upregulated HERV expression could be a key factor in ATLL development and the observed chemoresistance, potentially leading to new therapeutic strategies and significantly impacting the field of oncology and virology.

Tyrosine Protein Kinase SYK-Related Gene Signature in Baseline Immune Cells Associated with Adjuvant Immunotherapy-Induced Immune-Related Adverse Events in Melanoma.

Immune checkpoint inhibition (ICI) shows benefits in adjuvant (AT) and neoadjuvant melanoma treatments. However, ICI frequently induces severe immune-related adverse events (irAE). Unlike metastatic disease, in which irAEs are a clinical trade-off for treatment that improves survival, the toxicity burden from ICI in the AT setting is a substantial clinical problem urging for irAE-predictive biomarkers. We assessed postsurgical, pre-ICI treatment peripheral CD4+ and CD8+ T cells from clinical trial patients (CheckMate 915) treated with AT nivolumab (n = 130) or ipilimumab/nivolumab (COMBO, n = 82). Performing RNA sequencing differential gene expression analysis, we tested baseline differences associated with severe (grades 3-5) irAEs and constructed an irAE-predictive model using least absolute shrinkage and selection operator-regularized logistic regression. The analysis of predicted protein-protein interactions among differentially expressed genes in peripheral CD4+ cells revealed significant enrichment of the spleen tyrosine kinase (SYK) pathway, associated with severe irAEs in COMBO-treated patients. This gene expression signature predicted severe-irAE COMBO patients (χ2P value = 0.001) with 73% accuracy and was independent of disease recurrence (P = 0.79). The irAE-predictive model incorporating this gene expression signature demonstrated 82% accuracy (χ2P value = 8.91E-06). We identified baseline gene expression differences in key immune pathways of peripheral blood T cells from COMBO-treated patients with grades 3 to 5 irAEs and defined a SYK-related gene signature correctly identifying ∼60% of COMBO-treated patients with grades 3 to 5 irAEs. This finding aligns with our previous work linking anti-CTLA4 irAEs with a germline variant associated with high SYK expression. This gene signature may serve as a baseline biomarker of severe grade 3 to 5 irAE risk, which is especially important in AT treatment.

Immunogenic cell death-related genes predict prognosis and response to immunotherapy in lung squamous cell carcinoma.

Lung squamous cell carcinoma (LUSC) is a malignancy with limited therapeutic options. Immunogenic cell death (ICD) has the potential to enhance the efficacy of cancer therapy by triggering immune responses. We aimed to explore the potential of ICD-based classification in predicting prognosis and response to immunotherapy for LUSC. RNA-seq information and clinical data of LUSC patients were obtained from The Cancer Genome Atlas (TCGA) dataset. ICD-related gene expressions in LUSC samples were analyzed by consensus clustering. Subsequently, differentially expressed genes (DEGs) between different ICD-related subsets were analyzed. Tumor mutation burden, immune cell infiltration, and survival analyses were conducted between different ICD subsets. Finally, an ICD-related risk signature was constructed and evaluated in LUSC patients, and the immunotherapy responses based on the gene expressions were also forecasted. ICD-high and ICD-low groups were defined, and 1466 DEGs were identified between the two subtypes. These DEGs were mainly enriched in collagen-containing extracellular matrix, cytokine-cytokine receptor interaction, the PI3K-Akt signaling pathway, and neuroactive ligand-receptor interaction. Furthermore, the ICD-low group exhibited a favorable prognosis, enhanced TTN and MUC16 mutation frequencies, increased infiltrating immune cells, and downregulated immune checkpoint expressions. Furthermore, we demonstrated that an ICD-related model (based on CD4, NLRP3, NT5E, and TLR4 genes) could forecast the prognosis of LUSC, and ICD risk scores were lower in the responder group. In summary, the predicted values of ICD-related genes (CD4, NLRP3, NT5E, and TLR4) for the prognosis and response to immunotherapy in LUSC were verified in the study, which benefits immunotherapy-based interventions for LUSC patients.

Validation of reliable reference genes for qPCR of CD4+ Tcells exposed to compressive strain.

For accurate interpretation of quantitative real-time PCR (qPCR) data, stable reference genes are essential for normalization of target genes. To date, there is no information on reliable housekeeping genes in CD4+ Tcells in athree-dimensional (3D) matrix under pressure stimulation. This in vitro study describes for the first time amethod for pressure stimulation of CD4+ Tcells in a3D matrix in the context of orthodontic tooth movement (OTM) and identifies aset of reliable reference genes. CD4+ Tcells were isolated from murine spleen and activated with anti-CD3/-CD28 Dynabeads (Thermo Fisher, Langenselbold, Germany) on standard cell culture plates or in 3D scaffolds with or without compressive strain. Expression stability of nine potential reference genes was examined using four mathematical algorithms. Gene expression of Il2 was normalized to all potential reference genes to highlight the importance of correct normalization. Cell proliferation and the expression of the surface markers CD25 and CD69 were also determined. The 3D matrix did not inhibit proliferation after immunological activation of Tcells and embedded the cells sufficiently to expose them to pressure load. Expression of ubiquitin C (Ubc) and hypoxanthine phosphoribosyltransferase (Hprt) was the most stable under all conditions tested. Acombination of these two genes was suitable for normalization of qPCR data. Normalization of Il2 gene expression showed highly variable results depending on the reference gene used. Pressure reduced cell proliferation and the number of CD69-positive Tcells. This study provides abasis for performing valid and reliable qPCR experiments with CD4+ Tcells cultured in 3D scaffolds and exposed to compressive forces simulating OTM.