Multiple Immune-related Pathways Research Articles

Prognostic and immunological significance of tertiary lymphoid structures in nasopharyngeal carcinoma.

220 Background: Tertiary lymphoid structures (TLSs) are ectopic lymphoid tissues found in non-lymphoid organs, characterized by the presence of CD20+ B cell follicles adjacent to a T cell zone. Their presence has been associated with improved survival in various tumor types. However, the role of TLSs in nasopharyngeal carcinoma (NPC) remains unclear. In this study, we investigated the prognostic and immunological significance of TLSs in NPC patients. Methods: The investigation involved a discovery cohort (n = 145) comprising NPC patients diagnosed at Sun Yat-sen University Cancer Center between July 2010 and December 2016, with pretreatment primary tumor samples collected. An independent validation cohort included 150 NPC patients with gene expression profiles, among whom Hematoxylin & Eosin (H&E)-stained slides were available for 95 patients. The primary endpoint of this study is overall survival, defined as the duration of time from diagnosis until death from any cause. The presence of TLSs was assessed on H&E and CD20-stained slides. Analyses of tumor-infiltrating immune cell subsets, differentially expressed genes, and BCR/TCR clonotypes were performed to explore the association between TLSs and the anti-tumor immune response. A molecular definition of TLSs was proposed and assessed for its prognostic value across multiple cohorts. Results: The presence of TLSs was independently associated with a favorable overall survival (hazard ratio [HR], 0.26; 95% confidence interval [CI], 0.07-0.94; log-rank p = 0.04), surpassing the significance of B cell density in multivariate analysis (HR, 0.83; 95% CI, 0.38-1.79, log-rank p = 0.63). These findings were corroborated in the independent validation cohort. TLSs positively correlated with multiple immune-related pathways and a higher proportion of functionally mature immune cells. Notably, the counts of BCR and TCR clonotypes showed a significant positive correlation with the TLSs score. We proposed a TLSs gene signature and demonstrated its prognostic relevance across multiple cohorts, including patients treated with immune checkpoint blockade. Conclusions: TLSs exhibit heightened immune activity and indicate improved survival outcomes in NPC patients. A TLSs gene signature was proposed and demonstrated its prognostic relevance across multiple cohorts, indicating potential guidance for treatment decisions in cancer.

Integrating Multi-omics to Identify Age-Related Macular Degeneration Subtypes and Biomarkers

Age-related macular degeneration (AMD) is one of the most common causes of irreversible vision loss in the elderly. Its pathogenesis is likely multifactorial, involving a complex interaction of metabolic and environmental factors, and remains poorly understood. Previous studies have shown that mitochondrial dysfunction and oxidative stress play a crucial role in the development of AMD. Oxidative damage to the retinal pigment epithelium (RPE) has been identified as one of the major mediators in the pathogenesis of age-related macular degeneration (AMD). Therefore, this article combines transcriptome sequencing (RNA-seq) and single-cell sequencing (scRNA-seq) data to explore the role of mitochondria-related genes (MRGs) in AMD. Firstly, differential expression analysis was performed on the raw RNA-seq data. The intersection of differentially expressed genes (DEGs) and MRGs was performed. This paper proposes a deep subspace nonnegative matrix factorization (DS-NMF) algorithm to perform a multi-layer nonlinear transformation on the intersection of gene expression profiles corresponding to AMD samples. The age of AMD patients is used as prior information at the network’s top level to change the data distribution. The classification is based on reconstructed data with altered distribution. The types obtained significantly differ in scores of multiple immune-related pathways and immune cell infiltration abundance. Secondly, an optimal AMD diagnosis model was constructed using multiple machine learning algorithms for external and qRT-PCR verification. Finally, ten potential therapeutic drugs for AMD were identified based on cMAP analysis. The AMD subtypes identified in this article and the diagnostic model constructed can provide a reference for treating AMD and discovering new drug targets.

CPT1C-positive cancer-associated fibroblast facilitates immunosuppression through promoting IL-6-induced M2-like phenotype of macrophage

ABSTRACT Cancer-associated fibroblasts (CAFs) exhibit remarkable phenotypic heterogeneity, with specific subsets implicated in immunosuppression in various malignancies. However, whether and how they attenuate anti-tumor immunity in gastric cancer (GC) remains elusive. CPT1C, a unique isoform of carnitine palmitoyltransferase pivotal in regulating fatty acid oxidation, is briefly indicated as a protumoral metabolic mediator in the tumor microenvironment (TME) of GC. In the present study, we initially identified specific subsets of fibroblasts exclusively overexpressing CPT1C, hereby termed them as CPT1C+CAFs. Subsequent findings indicated that CPT1C+CAFs fostered a stroma-enriched and immunosuppressive TME as they correlated with extracellular matrix-related molecular features and enrichment of both immunosuppressive subsets, especially M2-like macrophages, and multiple immune-related pathways. Next, we identified that CPT1C+CAFs promoted the M2-like phenotype of macrophage in vitro. Bioinformatic analyses unveiled the robust IL-6 signaling between CPT1C+CAFs and M2-like phenotype of macrophage and identified CPT1C+CAFs as the primary source of IL-6. Meanwhile, suppressing CPT1C expression in CAFs significantly decreased IL-6 secretion in vitro. Lastly, we demonstrated the association of CPT1C+CAFs with therapeutic resistance. Notably, GC patients with high CPT1C+CAFs infiltration responded poorly to immunotherapy in clinical cohort. Collectively, our data not only present the novel identification of CPT1C+CAFs as immunosuppressive subsets in TME of GC, but also reveal the underlying mechanism that CPT1C+CAFs impair tumor immunity by secreting IL-6 to induce the immunosuppressive M2-like phenotype of macrophage in GC.

Discovery and validation of molecular patterns and immune characteristics in the peripheral blood of ischemic stroke patients.

Stroke is a disease with high morbidity, disability, and mortality. Immune factors play a crucial role in the occurrence of ischemic stroke (IS), but their exact mechanism is not clear. This study aims to identify possible immunological mechanisms by recognizing immune-related biomarkers and evaluating the infiltration pattern of immune cells. We downloaded datasets of IS patients from GEO, applied R language to discover differentially expressed genes, and elucidated their biological functions using GO, KEGG analysis, and GSEA analysis. The hub genes were then obtained using two machine learning algorithms (least absolute shrinkage and selection operator (LASSO) and support vector machine-recursive feature elimination (SVM-RFE)) and the immune cell infiltration pattern was revealed by CIBERSORT. Gene-drug target networks and mRNA-miRNA-lncRNA regulatory networks were constructed using Cytoscape. Finally, we used RT-qPCR to validate the hub genes and applied logistic regression methods to build diagnostic models validated with ROC curves. We screened 188 differentially expressed genes whose functional analysis was enriched to multiple immune-related pathways. Six hub genes (ANTXR2, BAZ2B, C5AR1, PDK4, PPIH, and STK3) were identified using LASSO and SVM-RFE. ANTXR2, BAZ2B, C5AR1, PDK4, and STK3 were positively correlated with neutrophils and gamma delta T cells, and negatively correlated with T follicular helper cells and CD8, while PPIH showed the exact opposite trend. Immune infiltration indicated increased activity of monocytes, macrophages M0, neutrophils, and mast cells, and decreased infiltration of T follicular helper cells and CD8 in the IS group. The ceRNA network consisted of 306 miRNA-mRNA interacting pairs and 285 miRNA-lncRNA interacting pairs. RT-qPCR results indicated that the expression levels of BAZ2B, C5AR1, PDK4, and STK3 were significantly increased in patients with IS. Finally, we developed a diagnostic model based on these four genes. The AUC value of the model was verified to be 0.999 in the training set and 0.940 in the validation set. Our research explored the immune-related gene expression modules and provided a specific basis for further study of immunomodulatory therapy of IS.

Identifying immune-related prognostic biomarkers in osteosarcoma: Development and validation of the tumor immune microenvironment risk model based on analysis of TCGA.

Osteosarcoma is a rare and aggressive malignancy with limited effective therapeutic options. This study aimed to identify immune-related prognostic biomarkers and develop a prognostic model for osteosarcoma. We performed integrated analysis of transcriptomic data and immune cell infiltration profiles of 84 osteosarcoma samples from the Cancer Genome Atlas (TCGA) database. Time-dependent receiver operating characteristic (ROC) curve analysis was used to assess the prognostic value of the TIMErisk model. We also performed functional annotation and pathway enrichment analyses to explore the potential mechanisms underlying the TIMErisk model. We identified a seven-gene TIMErisk model (C2, APBB1IP, BST2, TRPV2, CCL5, GBP1, and F13A1) that was independently associated with overall survival of osteosarcoma patients. The TIMErisk model showed significant associations with immune cell infiltration and immunosuppressive gene expression. In addition, the TIMErisk model was associated with drug sensitivity, and we found that several immune checkpoint genes were significantly differentially expressed between high- and low-TIMErisk groups. Functional annotation and pathway enrichment analyses revealed that the TIMErisk model was associated with multiple immune-related pathways, including antigen processing and presentation, cytokine-cytokine receptor interaction, and T cell receptor signaling pathway. Our study identified a novel TIME-based prognostic model for osteosarcoma that incorporates immune-related genes and can be used to predict patient prognosis and response to immunotherapy. Our findings highlight the importance of the TIME microenvironment in osteosarcoma progression and suggest that immune-related biomarkers may have clinical significance in the management of osteosarcoma.

Abstract B009: Dual PARP and EHMT1/2 inhibition induces an interferon response and anti-tumor immunity in ovarian cancer

Abstract Ovarian cancer has a high propensity to develop therapy resistance. Epigenetic therapies can sensitize ovarian cancer cells to first-line treatments such as platinum-based chemotherapy and Poly (ADP-ribose) polymerase inhibitors (PARPi). Particularly, the reduction of histone 3 lysine 9 dimethylation (H3K9me2) via a euchromatin histone methyltransferase inhibitor (EHMTi) has been shown to sensitize PARPi-resistant cells to PARPi therapy. Additionally, cell growth defects were observed in PARPi-resistant cells treated with dual PARPi/EHMTi treatment compared to DMSO or single agent treatments. To determine the mechanism by which dual therapy sensitizes cells and reduces cell viability, bulk RNA-sequencing was performed on several PARPi-resistant cell lines treated with DMSO, PARPi, EHMTi, or dual PARPi/EHMTi. Transcriptomic analyses revealed that dual therapy increased interferon (IFN) signaling-related genes compared to DMSO or single agent treatments. Additionally, expression of transposable element (TE) transcripts, which can form double-stranded RNA (dsRNA) and induce an IFN response, was also increased in dual treated cells compared to controls. To determine if the reduction in cell viability is dependent on TEs and IFN signaling (i.e., viral mimicry), CRISPRi technology was used to silence intracellular sensors of dsRNA, RIGI and MDA5. Cell growth assays of these silenced cells showed that the reduction in cell growth observed with dual treatment was partially dependent on MDA5 expression. Based on the observed increase in IFN signaling and T cell recruiting cytokines, we performed an animal study using an immunogenic, PARPi-resistant model of ovarian cancer to determine if this dual therapy has any anti-tumor effects. Indeed, we observed that single and dual EHMTi therapy significantly reduced tumor burden and that this reduction was partially dependent on CD8 T cells. Lastly, to determine if dual therapy also induces a viral mimicry response in the human tumor microenvironment, we created ex vivo cultures of tumors resected from patients with ovarian cancer. We then treated the ex vivo tissues with DMSO, PARPi, EHMTi, and dual PARPi/EHMTi, and performed bulk RNA-sequencing and multispectral immunohistochemistry. From these analyses, we found that dual therapy upregulated TE transcripts and multiple immune-related pathways, and increased T cell activity in the tumor microenvironment. Altogether, our data suggest that dual therapy activates viral mimicry and promotes anti-tumor immunity in both PARPi-resistant and -sensitive ovarian cancer. Citation Format: Lily Nguyen, Zachary L. Watson, Elizabeth R. Woodruff, Kimberly R. Jordan, Edward Chuong, Benjamin G. Bitler. Dual PARP and EHMT1/2 inhibition induces an interferon response and anti-tumor immunity in ovarian cancer [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr B009.

Abstract B024: Immune exhaustion and reversal of progestin-related immune modulation in adaptive resistance to levonorgestrel intrauterine device for treatment of atypical hyperplasia and early endometrial cancer

Abstract Background: A growing proportion of women diagnosed with atypical hyperplasia (AH) and early-stage endometrioid endometrial cancer (EEC) require nonsurgical treatment options. Our previously published phase II clinical trial of the levonorgestrel intrauterine device (LIUD) for AH and grade 1 EEC (G1EEC) demonstrated 83% response rate at 1 year; however, prospective long-term follow-up data and determinants for duration of response to LIUD are limited. Methods: Follow-up data from patients that participated in our prospective phase II trial of LIUD as a nonsurgical approach for AH and G1EEC were collected from medical records. Duration of response and time to relapse were evaluated. Spatial transcriptomic profiling from longitudinal biopsy samples from pre-treatment (n=5), on-treatment (n=5), and disease relapse lesions (n=3) were evaluated using digital spatial profiling (DSP, Nanostring GeoMX Whole Transcriptome Atlas). In silico cell type deconvolution was performed and pathway analyses was conducted using gene set variation analysis enrichment. Results: Of 43 participants exhibiting initial response to LIUD, 41 had follow-up data. Sixteen experienced disease relapse (39%) with a median time to relapse of 17 months after initial response. Clinical factors associated with shorter duration of response included age (HR 0.95, p=0.021), initial diagnosis of G1EEC (HR 3.51, p=0.014), premenopausal status (HR 3.85, p=0.039), and Hispanic ethnicity (HR 3.45, p=0.017). Pre-treatment IGF1 and IGF2 expression levels were positive predictors of duration of response (HR 0.44 and 0.45, respectively, p<0.05). DSP results revealed that LIUD increased NK cells (log2FC=46.13, FDR=0.004), and lymphocyte cytotoxicity signaling markers GZMB (log2FC=1.06, padj=0.023) and PRF1 (log2FC=1.62, padj=0.004), but NK cells were reduced in relapse lesions (log2FC=-55.96, FDR=0.02) as was PRF1 expression (log2FC=-1.87, padj=0.038). Multiple immune-related pathways (IFNα and IFNγ response, TGFβ signaling) were significantly upregulated in relapse lesions compared to matched pre-treatment lesions (FDR<0.05). IDO1 expression, a marker for immune exhaustion, was upregulated in relapse lesions compared to matched pre-treatment and on-treatment lesions (log2FC=2.56, FDR=0.01). Conclusions: While LIUD for treatment of AH or G1EEC has excellent initial response rates, upfront resistance, and relapse after initial response to LIUD impacts nearly half of patients. Our study shows progestin-related immunomodulatory effects, including increased NK cell infiltration and lymphocyte cytotoxicity. Immune mechanisms for relapse are implicated here, including reversal of progestin-related immunomodulatory effects (decreased NK cell infiltration and lymphocyte cytotoxicity) and increased immune exhaustion (prolonged IFNα and IFNγ response and increased IDO1 expression in relapse lesions). Thus, relieving immune exhaustion may increase durability of response to LIUD for treatment of AH or G1EEC. Citation Format: Mikayla B. Bowen, Brenda Melendez, Qian Zhang, Richard Yang, Bryan Fellman, Barrett Lawson, Naomi Adjei, Joseph Celestino, Tri Nguyen, Khalida Wani, Bhavana Singh, Christof Straub, Liang Zhang, Cheryl Tan, Felicia New, Diana Urbauer, Alexander Lazar, Karen Lu, Jennifer Wargo, Shannon N. Westin, Melinda S. Yates. Immune exhaustion and reversal of progestin-related immune modulation in adaptive resistance to levonorgestrel intrauterine device for treatment of atypical hyperplasia and early endometrial cancer [abstract]. In: Proceedings of the AACR Special Conference on Endometrial Cancer: Transforming Care through Science; 2023 Nov 16-18; Boston, Massachusetts. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(5_Suppl):Abstract nr B024.

Study of LY9 as a potential biomarker for prognosis and prediction of immunotherapy efficacy in lung adenocarcinoma.

Lymphocyte antigen 9 (LY9) participates in the development of several tumors and diseases but has not been reported yet in lung adenocarcinoma (LUAD). First, we analyzed the expression and prognostic value of LY9 in pan-cancer, including LUAD. Additionally, we conducted a correlation analysis of LY9 expression in LUAD with immune cell infiltration using the TIMER database and the CIBERSORT algorithm, and with immune checkpoints using the GEPIA database. Also, we constructed a potential ceRNA network for LY9. Furthermore, we explored LY9-related pathways by Gene Set Enrichment Analysis (GSEA). Finally, validation of differential expression at the mRNA level was obtained from the GEO database. We collected LUAD tissues for Quantitative Real-time PCR (qRT-PCR) to verify the expression of LY9, CD8, and CD4 and calculated the correlation between them. We also conducted immunohistochemistry (IHC) to verify the protein expression of LY9. Results showed that LY9 was highly expressed in various tumors, including LUAD. Besides, patients with high LY9 expression presented longer overall survival (OS) and more multiple lymphocyte infiltrations. The expression of LY9 in LUAD strongly and positively correlates with multiple immune cell infiltration and immune checkpoints. The functional enrichment analysis indicated that LY9 was involved in multiple immune-related pathways and non-small cell lung cancer. Moreover, a ceRNA regulatory network of LINC00943-hsa-miR-141-3p-LY9 might be involved. Finally, GSE68465 dataset confirmed differential expression of LY9 mRNA levels in LUAD and the qRT-PCR results verified LY9 had a strong and positive correlation with CD4 and CD8 T cells. Unfortunately, IHC did not detect the expression of LY9 protein level in tumor tissues and WB experiments validated the protein expression of LY9 in the OCI-AML-2 cell line. Therefore, we hypothesized that LY9 could serve as a potential, novel prognostic biomarker for LUAD and could predict immunotherapy efficacy at the mRNA level.

EPIGENETIC DIFFERENCE BETWEEN INSOMNIA GROUP AND NONINSOMNIA GROUP IN PARTICIPANTS WITH KNEE PAIN

Abstract Chronic knee pain is a common condition that often co-occurs with insomnia. Epigenetic modifications, such as DNA methylation, may play a role in the sleep-pain relationship. In this study, we aimed to identify differentially methylated CpGs/regions and enriched genomic pathways associated with insomnia in participants with knee pain (KP). We recruited 140 cognitively healthy middle to older-aged adults with self-reported KP and extracted DNA from peripheral blood. MethylationEPIC arrays were used for methylation data quantification, R package minfi was used for data preprocessing and quality control, and Ingenuity Pathway Analysis (IPA) was used to explore functional implications of differential methylation in pathways related to insomnia. Annotated genes within ±5kb of the putative differentially methylated regions (DMRs, p< 0.05) were subjected to the IPA analysis. There were no significant differences in sex or race between the two groups (p>0.05). We observed an over-representation of younger participants in the insomnia group (p=0.005) and also from the University of Florida (p=0.008). At raw p < 0.05 cutoff, we identified a total of 20,006 CpG probes, including 12,674 hypermethylated, and 7,332 hypomethylated CpG probes. Pathway analysis using IPA identified multiple immune-related pathways enriched with insomnia-related differentially methylated regions (DMRs), including the antigen presentation pathway (p=3.39E-08). Verapamil was identified as the most significant upstream regulator (p=8.8E-07). Our findings suggest the importance of epigenetic regulation of the immune system in the sleep-pain relationship and highlight the need for further research to understand the epigenetic contributions to sleep disorders in individuals with chronic pain.

DNALI1 is a prognosis-related biomarker and correlates with immune infiltrates in low grade glioma.

Dynein axonemal light intermediate chain 1 (DNALI1) is a component of axonemal dyneins and its role in cancer progression is not known. The influence of DNALI1 expression on the prognosis of low-grade gliomas (LGG) and the possible mechanisms of DNALI1 in promoting the progression of LGG was investigated by applying multiple bioinformatics analyses using datasets from TCGA, GTEx, CPTAC, and CGGA. The expression of DNALI1 in different tumor tissues including LGG was investigated. GO functional annotation, KEGG pathway analysis, and GSEA enrichment analysis were performed. The correlation between DNALI1 and prognosis, tumor microenvironment (TME) and immune checkpoints in LGG were assessed. DNALI1 is mainly expressed in malignant cells in the TME of LGG and positively correlated with the development of LGG. DNALI1 expression is negatively correlated with isocitrate dehydrogenase (IDH) mutations and 1p/19q co-deletion. High DNALI1 expression is associated with poor prognosis in LGG. DNALI1 may promote LGG progression through multiple immune-related pathways. The expression of DNALI1 is positively correlated with the infiltration of certain types of immune cells and the expression of some immune checkpoints. DNALI1 is a potential prognostic marker for LGG, and high expression of DNALI1 may play an important role in maintaining the immunosuppressive microenvironment of LGG.

Chromosome-level genome assembly and population genomic analysis provide novel insights into the immunity and evolution of Sogatella furcifera

Sogatella furcifera is a destructive agricultural pest causing large threats to rice production in China and Southeast Asian countries. Despite recent breakthroughs in long-read sequencing, high quality genomic data are very limited in S. furcifera. In present study, a chromosome-level assembly of the S. furcifera genome was completed (0.64 GB), comprising 15 chromosomes covered 95.04% of the estimated genome size, along with other 624 small scaffolds making up the remaining 4.96% of the genome of S. furcifera. A total of 24,669 protein-coding genes, 1211 long noncoding RNA and 7595 circular RNA transcripts were predicted in this study. Comparative genomic analysis revealed rapidly evolved genes were associated with multiple immune-related pathways in S. furcifera. Genome resequencing of 44 individuals from 12 geographic populations revealed frequent gene flow among populations. The systemic genomic analysis will provide more insights into the understanding of the immunity and evolutionary adaptation of S. furcifera.

Independent prognostic biomarker FERMT3 associated with immune infiltration and immunotherapy response in glioma

Background Adult glioma progresses rapidly and has a poor clinical outcome. The focal adhesion protein Kindlin-3 (encoded by the FERMT3 gene) participates in tumor development, drug resistance, and progression. However, the relationship between Kindlin-3 and glioma prognosis or immune microenvironment is poorly understood. Methods We comprehensively analyzed the expression, prognostic value, mutation landscape, functional enrichment, immune infiltration, and therapeutic role of FERMT3 in glioma using multiple datasets and validated Kindlin-3 expression in clinical tissue specimens by immunohistochemistry and multiple immunofluorescence staining. Results FERMT3 is an independent predictor of glioma prognosis and is highly expressed in glioblastoma tissues. Functional enrichment analyses indicated that FERMT3 participates in multiple immune-related pathways such as immune response and cytokine production. Furthermore, FERMT3 expression was positively correlated with the infiltration of several immune cells, immune scores, and the expression of genes related to immune checkpoints. Further analyses revealed that overexpression of FERMT3 was linked to a better response to anti-PD1 therapy. Data from single-cell RNA-seq reveal that FERMT3 was largely expressed in microglial cells and tissue-resident macrophages. Multiple immunofluorescence staining confirmed the overexpression of Kindlin-3 in the glioma-associated microglia/macrophages (GAMs). Conclusion The findings of this study provide a new perspective on the role of Kindlin-3 in glioma and may have a significant impact on the discovery of novel biomarkers and targeting of GAMs in the future.

Featured immune characteristics of COVID-19 and systemic lupus erythematosus revealed by multidimensional integrated analyses.

Coronavirus disease 2019 (COVID-19) shares similar immune characteristics with autoimmune diseases like systemic lupus erythematosus (SLE). However, such associations have not yet been investigated at the single-cell level. We integrated and analyzed RNA sequencing results from different patients and normal controls from the GEO database and identified subsets of immune cells that might involve in the pathogenesis of SLE and COVID- 19. We also disentangled the characteristic alterations in cell and molecular subset proportions as well as gene expression patterns in SLE patients compared with COVID-19 patients. Key immune characteristic genes (such as CXCL10 and RACK1) and multiple immune-related pathways (such as the coronavirus disease-COVID-19, T-cell receptor signaling, and MIF-related signaling pathways) were identified. We also highlighted the differences in peripheral blood mononuclear cells (PBMCs) between SLE and COVID-19 patients. Moreover, we provided an opportunity to comprehensively probe underlying B-cell‒cell communication with multiple ligand-receptor pairs (MIF-CD74+CXCR4, MIF-CD74+CD44) and the differentiation trajectory of B-cell clusters that is deemed to promote cell state transitions in COVID-19 and SLE. Our results demonstrate the immune response differences and immune characteristic similarities, such as the cytokine storm, between COVID-19 and SLE, which might pivotally function in the pathogenesis of the two diseases and provide potential intervention targets for both diseases.

Single‐Cell RNA Sequencing of Peripheral Blood Reveals Immune Cell Signatures in Alzheimer’s Disease

AbstractBackgroundAlzheimer’s disease (AD) pathogenesis involves various immune‐related phenomena both in central nervous system and peripheral blood. However, the characteristics of peripheral immune cells is poorly defined in AD at a single cell level.MethodWe profile 36,849 peripheral blood mononuclear cells (PBMC) from AD patients with amyloid positive (Aβ+) and normal controls with amyloid negative (Aβ‐) by 5' single‐cell transcriptome and immune repertoire sequencing with cell ranger standard analysis procedure. Based on the gene expression profile, immune cells were clustered and visualized by t‐distributed stochastic neighbour embedding (tSNE). In addition, we described the features of T cell receptor (TCR) and B cell receptor (BCR) repertoire from clonotypes, V gene and J gene skewed usage, amino acid length, and diversity of complementarity determining region 3 (CDR3).ResultWe revealed five immune cell subsets, CD4+ T cells, CD8+ T cells, B cells, natural killer cells, and monocytes‐macrophages cells, disentangled the characteristic alterations of cell subset proportion and gene expression patterns in AD. A total of 31 cell‐type‐specific key genes, comprising abundant human leucocyte antigen genes, and multiple immune related pathways were identified by protein‐protein interaction network and pathway enrichment analysis. We also found High‐frequency amplification clonotypes in T cells and B cells and the decreased diversity in T cells were detected in AD.ConclusionWe found abnormal immune infiltration and complex TCR and BCR alterations were presented in AD peripheral blood. As clone amplification suggested the activation of adaptive immune response against specific antigens, we speculated peripheral adaptive immune response, especially mediated by T cell, may have a role in the pathogenesis of AD. This finding may also contribute to further research regarding disease mechanism and development of immune related biomarkers or therapy.

Transcriptome profiling and differential expression analysis of altered immune-related genes in goldfish (Carassius auratus) infected with Aeromonas hydrophila

Goldfish (Carassius auratus) have been employed as a model organism to investigate the innate immune system and host-pathogen interactions. A Gram-negative bacterium called Aeromonas hydrophila has been found to cause mass mortality due to infection in a wide variety of fish species in the aquatic system. In this study, damages in Bowman's capsule, inflammatory tubular (proximal and distilled convoluted) structure, and glomerular necrosis were observed in A. hydrophila-infected head kidney of goldfish. To increase the better understanding of immune mechanisms of host defense against A. hydrophila, we performed a transcriptome analysis in head kidney of goldfish at 3 and 7 days of post-infection (dpi). Comparing to the control group, 4638 and 2580 differentially expressed genes (DEGs) were observed at 3 and 7 dpi, respectively. The DEGs were subsequently enriched in multiple immune-related pathways including Protein processing in endoplasmic reticulum, Insulin signaling pathway, and NOD-like receptor signaling pathway. The expression profile of immune-related genes such as TRAIL, CCL19, VDJ recombination-activating protein 1-like, Rag-1, and STING was validated by qRT-PCR. Furthermore, the levels of immune-related enzyme (LZM, AKP, SOD, and CAT) activities were examined at 3 and 7 dpi. The knowledge gained from the current study will be helpful for better understanding of early immune response in goldfish after A. hydrophila challenge, which will aid in future research on prevention strategies in teleost.

Novel m7G-related lncRNA signature for predicting overall survival in patients with gastric cancer

Presenting with a poor prognosis, gastric cancer (GC) remains one of the leading causes of disease and death worldwide. Long non-coding RNAs (lncRNAs) regulate tumor formation and have been long used to predict tumor prognosis. N7-methylguanosine (m7G) is the most prevalent RNA modification. m7G-lncRNAs regulate GC onset and progression, but their precise mechanism in GC is unclear. The objective of this research was the development of a new m7G-related lncRNA signature as a biomarker for predicting GC survival rate and guiding treatment. The Cancer Genome Atlas database helped extract gene expression data and clinical information for GC. Pearson correlation analysis helped point out m7G-related lncRNAs. Univariate Cox analysis helped in identifying m7G-related lncRNA with predictive capability. The Lasso-Cox method helped point out seven lncRNAs for the purpose of establishing an m7G-related lncRNA prognostic signature (m7G-LPS), followed by the construction of a nomogram. Kaplan–Meier analysis, univariate and multivariate Cox regression analysis, calibration plot of the nomogram model, receiver operating characteristic curve and principal component analysis were utilized for the verification of the risk model’s reliability. Furthermore, q-PCR helped verify the lncRNAs expression of m7G-LPS in-vitro. The study subjects were classified into high and low-risk groups based on the median value of the risk score. Gene enrichment analysis confirmed the constructed m7G-LPS’ correlation with RNA transcription and translation and multiple immune-related pathways. Analysis of the clinicopathological features revealed more progressive features in the high-risk group. CIBERSORT analysis showed the involvement of m7G-LPS in immune cell infiltration. The risk score was correlated with immune checkpoint gene expression, immune cell and immune function score, immune cell infiltration, and chemotherapy drug sensitivity. Therefore, our study shows that m7G-LPS constructed using seven m7G-related lncRNAs can predict the survival time of GC patients and guide chemotherapy and immunotherapy regimens as biomarker.

LRP1B mutation is associated with tumor immune microenvironment and progression-free survival in lung adenocarcinoma treated with immune checkpoint inhibitors.

Only a fraction of lung adenocarcinoma (LUAD) patients are eligible for immunotherapy. The identification of biomarkers for immunotherapy is crucial to improve patient outcomes. This study aims to systemically analyze LRP1B mutation and its association with the tumor immune microenvironment (TIME) and immunotherapy. A cohort of immune checkpoint inhibitors (ICIs)-treated LUAD patients was analyzed to assess the association of LRP1B mutation with immunotherapy prognosis. Another cohort of LUAD patients with genetic and transcriptomic data was also obtained from The Cancer Genome Atlas (TCGA). By investigating the ICIs and the TCGA-LUAD cohorts, we compared the differences in mutation profiles, immunogenicity, TIME, and DNA damage repair (DDR) mutations between the LRP1B-mutated and LRP1B wild-type groups. Additionally, we performed multiplex immunohistochemistry (mIHC) to validate the differences in the tumor microenvironment. Our results revealed that LRP1B mutation is associated with multiple immune-related pathways. Analysis of TIME indicated that LUAD patients with LRP1B mutation expressed significant levels of genes involved in antigen presentation, cytotoxicity, chemokines, and pro-inflammatory mediators, whereas a few immune checkpoint genes were highly expressed in the LRP1B-mutated group as well. Cell-type Identification by Estimating Relative Subsets of RNA Transcripts (CIBERSORT) analysis indicated that LRP1B-mutated LUAD patients had higher infiltration of active immune cells. Multiplex IHC analysis showed that LRP1B-mutated LUAD patients had elevated programmed death ligand-1 (PD-L1) expression and immune cell infiltration. Patients with LRP1B mutation had higher tumor mutation burden, neoantigens, as well as more mutated genes in the DDR-related pathways. Finally, LRP1B-mutated LUAD patients showed a significant prolongation of progression-free survival (PFS) in the ICIs cohort and could be effectively predicted by our constructed nomogram. Our study suggests that LRP1B mutation is associated with higher immune cell infiltration and elevated immune gene expression in TIME and potentially serves as a prognostic biomarker for LUAD patients treated with ICIs.

Identification of an unfolded protein response-related signature for predicting the prognosis of pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive lethal malignancy. An effective prognosis prediction model is urgently needed for treatment optimization. The differentially expressed unfolded protein response (UPR)‒related genes between pancreatic tumor and normal tissue were analyzed using the TCGA-PDAC dataset, and these genes that overlapped with UPR‒related prognostic genes from the E-MTAB-6134 dataset were further analyzed. Univariate, LASSO and multivariate Cox regression analyses were applied to establish a prognostic gene signature, which was evaluated by Kaplan‒Meier curve and receiver operating characteristic (ROC) analyses. E‒MTAB‒6134 was set as the training dataset, while TCGA-PDAC, GSE21501 and ICGC-PACA-AU were used for external validation. Subsequently, a nomogram integrating risk scores and clinical parameters was established, and gene set enrichment analysis (GSEA), tumor immunity analysis and drug sensitivity analysis were conducted. A UPR-related signature comprising twelve genes was constructed and divided PDAC patients into high- and low-risk groups based on the median risk score. The UPR-related signature accurately predicted the prognosis and acted as an independent prognostic factor of PDAC patients, and the AUCs of the UPR-related signature in predicting PDAC prognosis at 1, 2 and 3 years were all more than 0.7 in the training and validation datasets. The UPR-related signature showed excellent performance in outcome prediction even in different clinicopathological subgroups, including the female (p<0.0001), male (p<0.0001), grade 1/2 (p<0.0001), grade 3 (p=0.028), N0 (p=0.043), N1 (p<0.001), and R0 (p<0.0001) groups. Furthermore, multiple immune-related pathways were enriched in the low-risk group, and risk scores in the low-risk group were also associated with significantly higher levels of tumor-infiltrating lymphocytes (TILs). In addition, DepMap drug sensitivity analysis and our validation experiment showed that PDAC cell lines with high UPR-related risk scores or UPR activation are more sensitive to floxuridine, which is used as an antineoplastic agent. Herein, we identified a novel UPR-related prognostic signature that showed high value in predicting survival in patients with PDAC. Targeting these UPR-related genes might be an alternative for PDAC therapy. Further experimental studies are required to reveal how these genes mediate ER stress and PDAC progression.

Bioinformatic analysis of potential biomarkers and mechanisms of immune infiltration in mitral regurgitation complicated by atrial fibrillation.

Mitral regurgitation (MR) is one of the most prevalent valvular diseases. Degenerated MR-induced volume overload leads to left atrial enlargement and eventually, atrial fibrillation (AF). AF has a negative effect on patient prognosis despite recent advances in minimal invasive transcatheter devices for valve surgery. However, more effective strategies aimed at precisely treating from pathophysiology and genetic perspective are scarce. The gene expression datasets, GSE109744 and GSE79768, were obtained from the Gene Expression Omnibus database and analyzed to identify the differentially expressed genes (DEGs) in patients with mitral value prolapse (MVP) and AF. Subsequently, we predicted the extensive miRNA targets, and the protein-protein interaction (PPI) and miRNA-target gene regulatory networks were established. Functional enrichment analyses were performed for the DEGs. In addition, the co-expressed DEGs coupled with their predicted miRNAs and disease phenotypes involved in MVP and AF were assessed. Finally, the immune infiltration in both datasets was examined. A total of 491 and 180 DEGs were identified in the mitral valve and left atrial specimens, respectively. From these, 11 integrated co-expressed DEGs were identified, namely, PRG4, GPR34, RELN, CA3, IL1B, EPHA3, CHGB, TCEAL2, B3GALT2, ASB11, and CRISPLD1. The enriched Gene Ontology terms and KEGG pathways associated with the DEGs were determined, and the top 10 hub genes and top 3 gene clusters were selected from the PPI network. A prediction of target miRNAs was performed based on the co-expressed DEGs. The enrichment of the co-expressed DEGs suggested that immune and inflammatory responses might be involved in the disease development through multiple immune related pathways, including the interaction of cytokines and chemokines. Notably, this result was consistent with the immune infiltration analysis since the proportions of naïve B cells and memory B cells were significantly different in MVP and AF tissues compared to normal tissues. MR and AF are related, and 11 co-expressed DEGs were found to be significantly associated with MVP with AF, and indeed, these may represent novel biomarkers. Several immune cells were found to contribute to the process of MVP and AF via diverse mechanisms, in particular, antigen-presenting cells.

Transcriptome profiling and differential expression analysis of the immune-related genes during the acute phase of infection with Photobacterium damselae subsp. damselae in silver pomfret (Pampus argenteus)

Silver pomfret has been widely cultured in China due to its high economic value. Photobacterium damselae subsp. damselae (PDD) is a Gram-negative bacterium that has been shown to infect many fish species. To increase knowledge of the molecular mechanisms of the host defense against PDD, we conducted transcriptome analysis of head kidney in silver pomfret at 24 h and 72 h post-infection (hpi) via Illumina sequencing. The de novo assembly resulted in the identification of 79,063 unigenes, with 59,386 (75.11%) successfully annotated in public databases (NR, NT, KO, Swiss-Prot, Pfam, GO, and KOG databases). Comparison of gene expression profiles between PBS-injected fish (sham control) and PDD-challenged fish revealed 329 and 570 differentially expressed genes (DEGs) were screened at 24 hpi and 72 hpi, respectively. The DEGs were enriched in multiple immune-related pathways such as Hepatitis C, Gastric acid secretion, CAMs and Leukocyte transendothelial migration pathways, Primary immunodeficieny, ECM-receptor interaction, PI3K-Akt signaling pathway. The data obtained in the present study offers valuable information for acute immune response of silver pomfret challenged with PDD, which will facilitate further investigations on strategies against Photobacterium spp. infection in teleosts.