Application of a single-cell-RNA-based biological-inspired graph neural network in diagnosis of primary liver tumors.

Single-cell technology depicts integrated tumor profiles including both tumor cells and tumor microenvironments, which theoretically enables more robust diagnosis than traditional diagnostic standards based on only pathology. However, the inherent challenges of single-cell RNA sequencing (scRNA-seq) data, such as high dimensionality, low signal-to-noise ratio (SNR), sparse and non-Euclidean nature, pose significant obstacles for traditional diagnostic approaches. The diagnostic value of single-cell technology has been largely unexplored despite the potential advantages. Here, we present a graph neural network-based framework tailored for molecular diagnosis of primary liver tumors using scRNA-seq data. Our approach capitalizes on the biological plausibility inherent in the intercellular communication networks within tumor samples. By integrating pathway activation features within cell clusters and modeling unidirectional inter-cellular communication, we achieve robust discrimination between malignant tumors (including hepatocellular carcinoma, HCC, and intrahepatic cholangiocarcinoma, iCCA) and benign tumors (focal nodular hyperplasia, FNH) by scRNA data of all tissue cells and immunocytes only. The efficacy to distinguish iCCA from HCC was further validated on public datasets. Through extending the application of high-throughput scRNA-seq data into diagnosis approaches focusing on integrated tumor microenvironment profiles rather than a few tumor markers, this framework also sheds light on minimal-invasive diagnostic methods based on migrating/circulating immunocytes.

Phenotype remodelling of HNSCC cells in the muscle invasion environment.

BACKGROUND: Tumour invading muscle in head and neck squamous cell carcinoma (HNSCC) is often associated with destructive growth and poor prognosis. However, the phenotypic functions and pathological mechanisms of muscle-invasive cancer cells in tumour progress remains unknown. In this study, we aimed to investigate the phenotypic functions of muscle-invasive cancer cells of HNSCC and their potential crosstalk with tumour microenvironment. METHODS: We obtained scRNA-seq data (SC) from GSE103322 (N = 18) and GSE181919 (N = 37), spatial RNA-seq data (ST) from GSE208253 and GSE181300 (N = 4), transcriptomics of human HNSCC samples from GSE42743 (N = 12) and GSE41613 (N = 97). Utilizing the TCGA-HNSC dataset, we conducted univariate and multivariate Cox analyses to investigate the prognostic impact of muscle-invasion in HNSCC, with validation in an additional cohort. Through Stutility and AUCell approaches, we identified and characterized muscle-invasive cancer cell clusters, including their functional phenotypes and gene-specific profiles. Integration of SC and ST data was achieved using Seurat analysis, multimodal intersection analysis, and spatial deconvolution. The results were further validated via in vitro and in vivo experiments. RESULTS: Our analyses of the TCGA-HNSC cohort revealed the presence of muscle-invasion was associated with a poor prognosis. By combining ST and SC, we identified muscle-invasive cancer cells exhibiting epithelial-to-mesenchymal transition (EMT) and myoepithelial-like transcriptional programs, which were correlated with a poor prognosis. Furthermore, we identified G0S2 as a novel marker of muscle-invasive malignant cells that potentially promotes EMT and the acquisition of myoepithelium-like phenotypes. These findings were validated through in vitro assays and chorioallantoic membranes experiments. Additionally, we demonstrated that G0S2-overexpressing cancer cells might attract human ECs via VEGF signalling. Subsequent in vitro and in vivo experiments revealed G0S2 plays key roles in promoting the proliferation and invasion of cancer cells. CONCLUSIONS: In this study, we profiled the transcriptional programs of muscle-invasive HNSCC cell populations and characterized their EMT and myoepithelial-like phenotypes. Furthermore, our findings highlight the presence of muscle-invasion as a predictive marker for HNSCC patients. G0S2 as one of the markers of muscle-invasive cancer cells is involved in HNSCC intravasation, probably via VEGF signalling.

Breaking New Ground: Unraveling the USP1/ID3/E12/P21 Axis in Vascular Calcification.

Vascular calcification (VC) poses significant challenges in cardiovascular health. This study employs single-cell transcriptome sequencing to dissect cellular dynamics in this process. We identify distinct cell subgroups, notably in vascular smooth muscle cells (VSMCs), and observe differences between calcified atherosclerotic cores and adjacent regions. Further exploration reveals ID3 as a key gene regulating VSMC function. In vitro experiments demonstrate ID3's interaction with USP1 and E12, modulating cell proliferation and osteogenic differentiation. Animal models confirm the critical role of the USP1/ID3/E12/P21 axis in VC. This study sheds light on a novel regulatory mechanism, offering potential therapeutic targets.

UCN-Centric Prognostic Model for Predicting Overall Survival and Immune Response in Colorectal Cancer.

Colorectal cancer (CRC), a prevalent malignancy, ranks third in global incidence and second in mortality rates. Despite advances in screening methods such as colonoscopy, the accurate diagnosis of CRC remains challenging due to the absence of reliable biomarkers. This study aimed to develop a robust prognostic model for precise CRC outcome prediction. Employing weighted co-expression network analysis (WGCNA) and Cox regression analysis on data from The Cancer Genome Atlas (TCGA), we identified a panel of 12 genes strongly associated with patient survival. This gene panel facilitated accurate CRC outcome predictions, which is also validated via the external validation cohort GSE17536. We conducted further investigations into the key gene, urocortin (UCN), using single-cell transcriptomic data and immune infiltration analysis in CRC patients. Our results revealed a significant correlation between high UCN expression and the reduced prevalence of key immune cells, including B cells, CD4+ cytotoxic T cells, CD8+ T cells, and NKT cells. Functional experiments showed that UCN gene interference in the CRC cell lines significantly decreased cancer cell proliferation, underscoring UCN's role in intestinal immunity modulation. The UCN-centric prognostic model developed enhances prognosis prediction accuracy and offers critical insights for CRC diagnosis and therapeutic interventions.

A Strategy Involving Microporous Microneedles Integrated with CAR-TREM2-Macrophages for Scar Management by Regulating Fibrotic Microenvironment.

Dipeptidyl peptidase 4 (DPP4) positive fibroblasts play a pivotal role in scar development following skin injury. Heterogeneous vascular endothelial cells (ECs) within scarred areas retain the capacity to drive tissue regeneration and repair. Simultaneously, TREM2 macrophages play a crucial role in the progression and resolution of fibrosis by engaging in mutual regulation with ECs. However, effective strategies to inhibit scar formation through multi-factor regulation of the scar microenvironment remain a challenge. Here, CAR-TREM2-macrophages (CAR-TREM2-Ms) capable of targeting DPP4+ fibroblasts and modulating ECs subtype within the scar microenvironment are engineered to effectively prevent scarring. Hydrogel microporous microneedles (mMNs) are employed to deliver CAR-TREM2-Ms, which can effectively alleviate scar. Single-cell transcriptome sequencing (scRNA-seq) analysis reveals that CAR-TREM2-Ms can modify ECs fibrotic phenotype and regulate fibrosis by suppressing the profibrotic gene leucine-rich-alpha-2-glycoprotein 1 (Lrg1). In vitro experiments further demonstrate that CAR-TREM2-Ms improve the scar microenvironment by phagocytosing DPP4+ fibroblasts and suppressing TGFß secretion. This, in turn, inhibits the phenotypic conversion of LRG1 ECs and provides multifactorial way of alleviating scars. This study uncovers the evidence that mMNs attached to CAR-TREM2-Ms may exert vital influences on skin scarring through the regulation of the skin scar microenvironment, providing a promising approach for treating posttraumatic scarring.

Single-Cell WGCNA Combined with Transcriptome Sequencing to Study the Molecular Mechanisms of Inflammation-Related Ferroptosis in Myocardial Ischemia-Reperfusion Injury.

Purpose: Myocardial ischemia-reperfusion injury (MIRI) is characterized by inflammation and ferroptosis, but the precise mechanisms remain unknown. This study used single-cell transcriptomics technology to investigate the changes in various cell subtypes during MIRI and the regulatory network of ferroptosis-related genes and immune infiltration. Methods: Datasets GSE146285, GSE83472, GSE61592, and GSE160516 were obtained from Gene Expression Omnibus. Each cell subtype in the tissue samples was documented. The Seurat package was used for data preprocessing, standardization, and clustering. Cellphonedb was used to investigate the ligand-receptor interactions between cells. The hdWGCNA analysis was used to create a gene co-expression network. GSVA and GSEA were combined to perform functional enrichment and pathway analysis on the gene set. Furthermore, characteristic genes of the disease were identified using Lasso regression and SVM algorithms. Immune cell infiltration analysis was also performed. MIRI rat models were created, and samples were taken for RT-qPCR and Western blot validation. Results: The proportion of MIRI samples in the C2, C6, and C11 subtypes was significantly higher than that of control samples. Three genes associated with ferroptosis (CD44, Cfl1, and Zfp36) were identified as MIRI core genes. The expression of these core genes was significantly correlated with mast cells and monocyte immune infiltrating cells. The experimental validation confirmed the upregulation of Cd44 and Zfp36 expression levels in MIRI, consistent with current study trends. Conclusion: This study used single-cell transcriptomics technology to investigate the molecular mechanisms underpinning MIRI. Numerous important cell subtypes, gene regulatory networks, and disease-associated immune infiltration were also discovered. These findings provide new information and potential therapeutic targets for MIRI diagnosis and treatment.

Human umbilical cord mesenchymal stem cell-derived exosomes mitigate diabetic nephropathy via enhancing M2 macrophages polarization.

Background and objectives: Exosomes, which are small nanoscale vesicles capable of secretion, have garnered significant attention in recent years because of their therapeutic potential, particularly in the context of kidney diseases. Notably, human umbilical cord mesenchymal stem cell-derived exosomes (hucMSC-Exos) are emerging as promising targeted therapies for renal conditions. The aim of this study was to investigate the therapeutic effects of hucMSC-Exos on diabetic kidney disease (DKD) both in vivo and in vitro. Additionally, this study seeks to elucidate cellular and molecular differentials, as well as the expression of relevant signaling pathways, through single-cell RNA sequencing. This endeavor was designed to enhance our understanding of the connection between hucMSC-Exos and the pathogenesis of DKD. Methods and results: The study commenced with the extraction and characterization of hucMSC-Exos, including the determination of their concentrations. Animal experiments were conducted to evaluate the therapeutic potential of hucMSC-Exos in a DKD mouse model. Subsequently, single-cell sequencing was employed to investigate the molecular mechanisms underlying the efficacy of extracellular vesicles in ameliorating DKD. These findings were further substantiated by cell-based experiments. Importantly, the results indicate that hucMSC-Exos can impede the progression of DKD in mice, with macrophage activation playing a pivotal role in this process. Conclusions: The in vivo experiments conclusively established hucMSC-Exos as a pivotal component in preserving renal function and retarding the progression of DKD. Our utilization of single-cell sequencing technology, in conjunction with in vivo and in vitro experiments, provides compelling evidence that M2 macrophages are instrumental in enhancing the amelioration of diabetic nephropathy.

Single-cell spatial transcriptomics in cardiovascular development, disease, and medicine.

Cardiovascular diseases (CVDs) impose a significant burden worldwide. Despite the elucidation of the etiology and underlying molecular mechanisms of CVDs by numerous studies and recent discovery of effective drugs, their morbidity, disability, and mortality are still high. Therefore, precise risk stratification and effective targeted therapies for CVDs are warranted. Recent improvements in single-cell RNA sequencing and spatial transcriptomics have improved our understanding of the mechanisms and cells involved in cardiovascular phylogeny and CVDs. Single-cell RNA sequencing can facilitate the study of the human heart at remarkably high resolution and cellular and molecular heterogeneity. However, this technique does not provide spatial information, which is essential for understanding homeostasis and disease. Spatial transcriptomics can elucidate intracellular interactions, transcription factor distribution, cell spatial localization, and molecular profiles of mRNA and identify cell populations causing the disease and their underlying mechanisms, including cell crosstalk. Herein, we introduce the main methods of RNA-seq and spatial transcriptomics analysis and highlight the latest advances in cardiovascular research. We conclude that single-cell RNA sequencing interprets disease progression in multiple dimensions, levels, perspectives, and dynamics by combining spatial and temporal characterization of the clinical phenome with multidisciplinary techniques such as spatial transcriptomics. This aligns with the dynamic evolution of CVDs (e.g., "angina-myocardial infarction-heart failure" in coronary artery disease). The study of pathways for disease onset and mechanisms (e.g., age, sex, comorbidities) in different patient subgroups should improve disease diagnosis and risk stratification. This can facilitate precise individualized treatment of CVDs.

The single-cell transcriptome of mTECs and CD4+ thymocytes under adhesion revealed heterogeneity of mTECs and a network controlled by Aire and lncRNAs.

To further understand the impact of deficiency of the autoimmune regulator (Aire) gene during the adhesion of medullary thymic epithelial cells (mTECs) to thymocytes, we sequenced single-cell libraries (scRNA-seq) obtained from Aire wild-type (WT) (Airewt/wt ) or Aire-deficient (Airewt/mut ) mTECs cocultured with WT single-positive (SP) CD4+ thymocytes. Although the libraries differed in their mRNA and long noncoding RNA (lncRNA) profiles, indicating that mTECs were heterogeneous in terms of their transcriptome, UMAP clustering revealed that both mTEC lines expressed their specific markers, i.e., Epcam, Itgb4, Itga6, and Casp3 in resting mTECs and Ccna2, Pbk, and Birc5 in proliferative mTECs. Both cocultured SP CD4+ thymocytes remained in a homogeneous cluster expressing the Il7r and Ccr7 markers. Comparisons of the two types of cocultures revealed the differential expression of mRNAs that encode transcription factors (Zfpm2, Satb1, and Lef1), cell adhesion genes (Itgb1) in mTECs, and Themis in thymocytes, which is associated with the regulation of positive and negative selection. At the single-cell sequencing resolution, we observed that Aire acts on both Aire WT and Aire-deficient mTECs as an upstream controller of mRNAs, which encode transcription factors or adhesion proteins that, in turn, are posttranscriptionally controlled by lncRNAs, for example, Neat1, Malat1, Pvt1, and Dancr among others. Under Aire deficiency, mTECs dysregulate the expression of MHC-II, CD80, and CD326 (EPCAM) protein markers as well as metabolism and cell cycle-related mRNAs, which delay the cell cycle progression. Moreover, when adhered to mTECs, WT SP CD4+ or CD8+ thymocytes modulate the expression of cell activation proteins, including CD28 and CD152/CTLA4, and the expression of cellular metabolism mRNAs. These findings indicate a complex mechanism through which an imbalance in Aire expression can affect mTECs and thymocytes during adhesion.

Single-cell transcriptional atlas of tumor-associated macrophages in breast cancer.

BACKGROUND: The internal heterogeneity of breast cancer, notably the tumor microenvironment (TME) consisting of malignant and non-malignant cells, has been extensively explored in recent years. The cells in this complex cellular ecosystem activate or suppress tumor immunity through phenotypic changes, secretion of metabolites and cell-cell communication networks. Macrophages, as the most abundant immune cells within the TME, are recruited by malignant cells and undergo phenotypic remodeling. Tumor-associated macrophages (TAMs) exhibit a variety of subtypes and functions, playing significant roles in impacting tumor immunity. However, their precise subtype delineation and specific function remain inadequately defined. METHODS: The publicly available single-cell transcriptomes of 49,141 cells from eight breast cancer patients with different molecular subtypes and stages were incorporated into our study. Unsupervised clustering and manual cell annotation were employed to accurately classify TAM subtypes. We then conducted functional analysis and constructed a developmental trajectory for TAM subtypes. Subsequently, the roles of TAM subtypes in cell-cell communication networks within the TME were explored using endothelial cells (ECs) and T cells as key nodes. Finally, analyses were repeated in another independent publish scRNA datasets to validate our findings for TAM characterization. RESULTS: TAMs are accurately classified into 7 subtypes, displaying anti-tumor or pro-tumor roles. For the first time, we identified a new TAM subtype capable of proliferation and expansion in breast cancer-TUBA1B+ TAMs playing a crucial role in TAMs diversity and tumor progression. The developmental trajectory illustrates how TAMs are remodeled within the TME and undergo phenotypic and functional changes, with TUBA1B+ TAMs at the initial point. Notably, the predominant TAM subtypes varied across different molecular subtypes and stages of breast cancer. Additionally, our research on cell-cell communication networks shows that TAMs exert effects by directly modulating intrinsic immunity, indirectly regulating adaptive immunity through T cells, as well as influencing tumor angiogenesis and lymphangiogenesis through ECs. CONCLUSIONS: Our study establishes a precise single-cell atlas of breast cancer TAMs, shedding light on their multifaceted roles in tumor biology and providing resources for targeting TAMs in breast cancer immunotherapy.

Single-Cell Transcriptomics Reveals Early Effects of Ionizing Radiation on Bone Marrow Mononuclear Cells in Mice.

Ionizing radiation exposure can cause damage to diverse tissues and organs, with the hematopoietic system being the most sensitive. However, limited information is available regarding the radiosensitivity of various hematopoietic cell populations in the bone marrow due to the high heterogeneity of the hematopoietic system. In this study, we observed that granulocyte-macrophage progenitors, hematopoietic stem/progenitor cells, and B cells within the bone marrow showed the highest sensitivity, exhibiting a rapid decrease in cell numbers following irradiation. Nonetheless, neutrophils, natural killer (NK) cells, T cells, and dendritic cells demonstrated a certain degree of radioresistance, with neutrophils exhibiting the most pronounced resistance. By employing single-cell transcriptome sequencing, we investigated the early responsive genes in various cell types following irradiation, revealing that distinct gene expression profiles emerged between radiosensitive and radioresistant cells. In B cells, radiation exposure led to a specific upregulation of genes associated with mitochondrial respiratory chain complexes, suggesting a connection between these complexes and cell radiosensitivity. In neutrophils, radiation exposure resulted in fewer gene alterations, indicating their potential for distinct mechanisms in radiation resistance. Collectively, this study provides insights into the molecular mechanism for the heterogeneity of radiosensitivity among the various bone marrow hematopoietic cell populations.

Insights into left-right asymmetric development of chicken ovary at the single-cell level.

Avian ovaries develop asymmetrically apart from prey birds, with only the left ovary growing more towards functional organ. Here, we analyze over 135,000 cells from chick's left and right ovaries at six distinct embryonic developmental stages utilizing single-cell transcriptome sequencing. We delineate gene expression patterns across 15 cell types within these embryo ovaries, revealing side-specific development. The left ovaries exhibit cortex cells, zygotene germ cells, and transcriptional changes unique to the left side. Differential gene expression analysis further identifies specific markers and pathways active in these cell types, highlighting the asymmetry in ovarian development. A fine-scale analysis of the germ cell meiotic transcriptome reveals seven distinct clusters with gene expression patterns specific to various meiotic stages. The study also identifies signaling pathways and intercellular communications, particularly between pre-granulosa and germ cells. Spatial transcriptome analysis shows the asymmetry, demonstrating cortex cells exclusively in the left ovary, modulating neighboring cell types through putative secreted signaling molecules. Overall, this single-cell analysis provides insights into the molecular mechanisms of the asymmetric development of avian ovaries, particularly the significant role of cortex cells in the left ovary.

Identification of senescent cell subpopulations by CITE-seq analysis.

Cellular senescence, a state of persistent growth arrest, is closely associated with aging and age-related diseases. Deciphering the heterogeneity within senescent cell populations and identifying therapeutic targets are paramount for mitigating senescence-associated pathologies. In this study, proteins on the surface of cells rendered senescent by replicative exhaustion and by exposure to ionizing radiation (IR) were identified using mass spectrometry analysis, and a subset of them was further studied using single-cell CITE-seq (Cellular Indexing of Transcriptomes and Epitopes by Sequencing) analysis. Based on the presence of proteins on the cell surface, we identified two distinct IR-induced senescent cell populations: one characterized by high levels of CD109 and CD112 (cluster 3), the other characterized by high levels of CD112, CD26, CD73, HLA-ABC, CD54, CD49A, and CD44 (cluster 0). We further found that cluster 0 represented proliferating and senescent cells in the G1 phase of the division cycle, and CITE-seq detection of cell surface proteins selectively discerned those in the senescence group. Our study highlights the heterogeneity of senescent cells and underscores the value of cell surface proteins as tools for distinguishing senescent cell programs and subclasses, paving the way for targeted therapeutic strategies in disorders exacerbated by senescence.

Integrative transcriptome analysis reveals the molecular events underlying impaired T-cell responses in EGFR-mutant lung cancer.

EGFR mutations are critical oncogenic drivers in lung adenocarcinoma (LUAD). However, the mechanisms by which they impact the tumor microenvironment (TME) and tumor immunity are unclear. Furthermore, the reasons underlying the poor response of EGFR-mutant (EGFR-MU) LUADs to immunotherapy with PD-1/PD-L1 inhibitors are unknown. Utilizing single-cell RNA (sc-RNA) and bulk RNA sequencing datasets, we conducted high-dimensional weighted gene coexpression network analysis to identify key genes and immune-related pathways contributing to the immunosuppressive TME. EGFR-MU cancer cells downregulated MHC class I genes to evade CD8+ cytotoxic T cells, expressed substantial levels of MHC class II molecules, and engaged with CD4+ regulatory T cells (Tregs). EGFR-MU tumors may recruit Tregs primarily through the CCL17/CCL22/CCR4 axis, leading to a Treg-enriched TME. High levels of MHC class II-positive cancer-associated fibroblasts and tumor endothelial cells were found within EGFR-MU tumors. Owing to the absence of costimulatory factors, they may inhibit rather than activate the tumor antigen-specific CD4+ T-cell response, contributing further to immune suppression. Multiplex immunohistochemistry analyses in a LUAD cohort confirmed increased expression of MHC class II molecules in cancer cells and fibroblasts in EGFR-MU tumors. Our research elucidates the highly immunosuppressive TME in EGFR-MU LUAD and suggests potential targets for effective immunotherapy.

Function of unconventional T cells in oral lichen planus revealed by single-cell RNA sequencing.

OBJECTIVE: We intended to map the single-cell profile of OLP, explore the molecular characteristics of unconventional T cells in OLP tissues. METHODS: Buccal mucosa samples from OLP patients and healthy individuals were used to prepare single-cell suspension. Single-cell RNA sequencing was used to analyze the proportion of all the cells, and the molecular characteristics of unconventional T cells. Immunohistochemical staining was used to detect the expression of unconventional T cells marker genes. RESULTS: The cell clusters from buccal mucosa were categorized into immune cells, fibroblasts, endothelial cells, and epithelial cells. Unconventional T cells with phenotype of CD247+TRDC+NCAM1+ were identified. Immunohistochemical staining revealed higher expression of unconventional T cell marker genes in OLP tissue, predominantly in the lamina propria. In OLP, unconventional T cells are in a unique stress response state, exhibited enhanced NF-κB signaling and apoptosis inhibition, enhanced heat shock protein genes expression, weakened cytotoxic function. A large number of ligand-receptor pairs were found between unconventional T cells and other cells, particularly with fibroblasts and endothelial cells. CONCLUSIONS: This study mapped the single-cell profile of OLP, delineated the molecular characteristics of unconventional T cells in OLP, and uncovered that these unconventional T cells are in a stress response state.

HAO2 protects from proximal tubular cells injured in rats with chronic kidney disease by promoting fatty acid metabolic processes.

The complex pathogenesis of kidney disease is closely related to the diversity of kidney intrinsic cells. In this study, single-cell transcriptome sequencing technology was used to sequence and analyze blood and kidney tissue cells in normal control rats and rats with chronic kidney disease (CKD), focusing on key cell populations and functional enrichment to explore the pathogenesis of CKD. Oil red O staining and enzyme-linked immunosorbent assay (ELISA) were used to detect lipid droplets and free fatty acid (FFA). Quantitative real-time polymerase chain reaction (RT-PCR), western blot (WB) were used to verify the differential gene hydroxyacid oxidase 2 (HAO2) and fatty acid metabolic process in tissue to ensure the reliability of single-cell sequencing results. We successfully established a single-cell transcriptome atlas of blood and kidney tissue in rats with CKD, which were annotated into 14 cell subsets (MPCs, PT, Tc, DCT, B-IC, A-IC, CNT, ALOH, BC, Neu, Endo, Pla, NKT, Baso) according to marker gene, and the integrated single-cell atlas of rats showed a significant increase and decrease of MPCs and PTs in the CKD group, respectively. Functional analysis found extensive enrichment of metabolic-related pathways in PT cells, includes fatty acid metabolic process, cellular amino acid metabolic process and generation of precursor metabolites and energy. Immunohistochemical experiments determined that the differential gene HAO2 was localized in the renal tubules, and its expression was significantly reduced in CKD group compared with control, and oil red O staining showed that lipid droplets increased in the CKD group, after overexpression of HAO2 the lipid droplets was inhibited. ELISA assay showed that ATP content decreased in the CKD group and FFA increased in the CKD group. Moreover, the mitochondrial membrane potential of the cells in the OE-HAO2 group was significantly increased compared with OE-NC. The acyl-CoA oxidase 1(ACOX1), peroxisome proliferator-activated receptor alpha (PPARα), peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) were decreased in the CKD group, while genes and proteins were increased after overexpression of HAO2, and the AMP-activated protein kinase (AMPK) phosphorylated proteins were increased, the acetyl-CoA carboxylase (ACC) phosphorylated proteins were decreased, reversely. Therefore, HAO2 may be an important regulator of fatty acid metabolic processes in CKD, and overexpression of HAO2 can enhance fatty acid metabolism by promoting fatty acid oxidation (FAO) pathway.

The heterogeneity of erythroid cells: insight at the single-cell transcriptome level.

Erythroid cells, the most prevalent cell type in blood, are one of the earliest products and permeate through the entire process of hematopoietic development in the human body, the oxygen-transporting function of which is crucial for maintaining overall health and life support. Previous investigations into erythrocyte differentiation and development have primarily focused on population-level analyses, lacking the single-cell perspective essential for comprehending the intricate pathways of erythroid maturation, differentiation, and the encompassing cellular heterogeneity. The continuous optimization of single-cell transcriptome sequencing technology, or single-cell RNA sequencing (scRNA-seq), provides a powerful tool for life sciences research, which has a particular superiority in the identification of unprecedented cell subgroups, the analyzing of cellular heterogeneity, and the transcriptomic characteristics of individual cells. Over the past decade, remarkable strides have been taken in the realm of single-cell RNA sequencing technology, profoundly enhancing our understanding of erythroid cells. In this review, we systematically summarize the recent developments in single-cell transcriptome sequencing technology and emphasize their substantial impact on the study of erythroid cells, highlighting their contributions, including the exploration of functional heterogeneity within erythroid populations, the identification of novel erythrocyte subgroups, the tracking of different erythroid lineages, and the unveiling of mechanisms governing erythroid fate decisions. These findings not only invigorate erythroid cell research but also offer new perspectives on the management of diseases related to erythroid cells.

A global view of altered ligand-receptor interactions in bone marrow aging based on single-cell sequencing.

Altered cell-cell communication is a hallmark of aging, but its impact on bone marrow aging remains poorly understood. Based on a common and effective pipeline and single-cell transcriptome sequencing, we detected 384,124 interactions including 2575 ligand-receptor pairs and 16 non-adherent bone marrow cell types in old and young mouse and identified a total of 5560 significantly different interactions, which were then verified by flow cytometry and quantitative real-time PCR. These differential ligand-receptor interactions exhibited enrichment for the senescence-associated secretory phenotypes. Further validation demonstrated supplementing specific extracellular ligands could modify the senescent signs of hematopoietic stem cells derived from old mouse. Our work provides an effective procedure to detect the ligand-receptor interactions based on single-cell sequencing, which contributes to understand mechanisms and provides a potential strategy for intervention of bone marrow aging.

Single-cell transcriptome sequencing partially revealed the changes of T cells in the early stage of aging kidney.

Aging is a gradual, inevitable physiologic process. The organ aging is related to the persistence of chronic inflammation, but the understanding of inflammatory state during renal aging is lacking currently. Single-cell transcriptome sequencing was performed on aging mouse kidney to reveal the molecular phenotype and composition changes of different cell types. In the early stage of aging, immune cells such as T, B cells and mononuclear macrophages increased in kidney. The molecular state of T cells in aging kidney changed and polarized. Among them, we identified a group of GZMK+ CD8 + T cells with high expression of Eomes, Pdcd1 and Ifng and a group of Il17a+ T cells with high expression of Il17a and Il23r. Moreover, the cytokines and inflammations can aggravate tissue damage eventually. Furthermore, we found the interaction between different types of epithelial cells and T cells increased during the renal aging. These results identify the changes of T cells in the early stage of aging kidney and suggest that GZMK+CD8+ T cells might be a potential target to ameliorate age-associated dysfunctions of kidney(Graphical Abstract).

Integrated analyses of single-cell transcriptome and Mendelian randomization reveal the protective role of FCRL3 in multiple sclerosis.

Background: Multiple sclerosis (MS) represents a multifaceted autoimmune ailment, prompting the development and widespread utilization of numerous therapeutic interventions. However, extant medications for MS have proven inadequate in mitigating relapses and halting disease progression. Innovative drug targets for preventing multiple sclerosis are still required. The objective of this study is to discover novel therapeutic targets for MS by integrating single-cell transcriptomics and Mendelian randomization analysis. Methods: The study integrated MS genome-wide association study (GWAS) data, single-cell transcriptomics (scRNA-seq), expression quantitative trait loci (eQTL), and protein quantitative trait loci (pQTL) data for analysis and utilized two-sample Mendelian randomization study to comprehend the causal relationship between proteins and MS. Sequential analyses involving colocalization and Phenome-wide association studies (PheWAS) were conducted to validate the causal role of candidate genes. Results: Following stringent quality control preprocessing of scRNA-seq data, 1,123 expression changes across seven peripheral cell types were identified. Among the seven most prevalent cell types, 97 genes exhibiting at least one eQTL were discerned. Examination of MR associations between 28 proteins with available index pQTL signals and the risk of MS outcomes was conducted. Co-localization analyses and PheWAS indicated that FCRL3 may exert influence on MS. Conclusion: The integration of scRNA-seq and MR analysis facilitated the identification of potential therapeutic targets for MS. Notably, FCRL3, implicated in immune function, emerged as a significant drug target in the deCODE databases. This research underscores the importance of FCRL3 in MS therapy and advocates for further investigation and clinical trials targeting FCRL3.