ABSTRACT:Background:After ischemic stroke,neutrophils hyperactivate,increasing in number and worsening inflammation,causing neural damage.Prior scRNA-seq showed Lrg1 modulates cells subsentence to cerebral ischemiarep...ABSTRACT:Background:After ischemic stroke,neutrophils hyperactivate,increasing in number and worsening inflammation,causing neural damage.Prior scRNA-seq showed Lrg1 modulates cells subsentence to cerebral ischemiareperfusion injury,but its mechanism in regulating neutrophil accumulation/differentiation post-injury is unclear.Methods:Lrg1 knockout impact on neutrophil accumulation was assessed via immunofluorescence and western blot.Three-dimensional reconstruction of immunofluorescent staining analyzed cell-cell interactions among neutrophils and microglia.scRNA-seq of WT and Lrg1^(-/-)mice from GSE245386 and GSE279462 was conducted.Each group conducted oxidative phosphorylation scoring via Gene Set Enrichment Analysis(GSEA),while Metascape was employed to perform GO and KEGG enrichment analyses for elucidating functional mechanisms.CellChat exhibited cell-cell communication.Furthermore,alterations in microglial phagocytic activity were evaluated by immunostaining for CD68,a well-established marker of phagolysosomal activity in phagocytic cells.Brain energy metabolism was evaluated via glutamate dehydrogenase activity and ATP levels with ELISA,and enzyme expression was analyzed by immunofluorescence and western blot.Results:Lrg1 knockout decreased neutrophil accumulation and NET formation in mice.3D immunofluorescence reconstruction confirmed neutrophil co-localization with endothelial cells/microglia.scRNA-seq revealed that the oxidative phosphorylation score was significantly higher in the MCAO/R+WT group compared to both the Sham-operated+WT and Lrg1^(-/-)groups.Notably,the oxidative phosphorylation score was further elevated in the MCAO/R+Lrg1^(-/-)group.Immunostaining showed that Lrg1 knockout elevated CD68+lysosome expression post-MCAO/R,with TMEM119 colocalizing with these lysosomes.MCAO/R raised CD68 expression in ischemic brains,an effect further intensified by Lrg1 knockout.KEGG analysis linked differential genes to oxidative phosphorylation pathways.Validation in MCAO/R vs.sham groups revealed increased ROS production and reduced expression of complex enzymes I-V(NDUFB8,SDHB,UQCRC1,MTCO2,ATP5A1).Lrg1 intervention increased enzyme expression.Immunofluorescence and western blot in brain tissue showed similar patterns in microglia and enzymes I-V.Conclusions:Lrg1 knockout significantly enhances microglial phagocytic activity towards neutrophils subsequent to cerebral ischemia-reperfusion injury,through its regulatory effect on the oxidative phosphorylation pathway.This finding accentuates Lrg1 as a highly potential therapeutic target for intervening in and modulating post-ischemic inflammatory responses.展开更多
Gut microbes exhibit complex interactions with their hosts and shape an organism's immune system throughout its lifespan.As the largest secondary lymphoid organ,the spleen has a wide range of immunological functio...Gut microbes exhibit complex interactions with their hosts and shape an organism's immune system throughout its lifespan.As the largest secondary lymphoid organ,the spleen has a wide range of immunological functions.To explore the role of microbiota in regulating and shaping the spleen,we employ scRNA-seq and Stereo-seq technologies based on germ-free(GF)mice to detect differences in tissue size,anatomical structure,cell types,functions,and spatial molecular characteristics.We identify 18 cell types,9 subtypes of T cells,and 7 subtypes of B cells.Gene differential expression analysis reveals that the absence of microorganisms results in alterations in erythropoiesis within the red pulp region and congenital immune deficiency in the white pulp region.Stereo-seq results demonstrate a clear hierarchy of immune cells in the spleen,including marginal zone(MZ)macrophages,MZ B cells,follicular B cells and T cells,distributed in a well-defined pattern from outside to inside.However,this hierarchical structure is disturbed in GF mice.Ccr7 and Cxcl13 chemokines are specifically expressed in the spatial locations of T cells and B cells,respectively.We speculate that the microbiota may mediate the structural composition or partitioning of spleen immune cells by modulating the expression levels of chemokines.展开更多
The process of lymphatic metastasis was proved to be associated with podoplanin-expressing macrophages in breast cancer(BC).This study aimed to investigate the role of the M2 phenotype of tumor-associated macrophages ...The process of lymphatic metastasis was proved to be associated with podoplanin-expressing macrophages in breast cancer(BC).This study aimed to investigate the role of the M2 phenotype of tumor-associated macrophages and mine the key M2 macrophages-related genes for lymph node metastasis in BC.We downloaded the GSE158399 dataset from the Gene Expression Omnibus(GEO)database,which includes transcriptomic profiles of individual cells from primary tumors,negative lymph nodes(NLNs),and positive lymph nodes(PLNs)of breast cancer patients.The cell subsets were identified by clustering analysis after quality control of the scRNA-seq using Seurat.The activation and migration capability of M2 macrophages were evaluated with R package“GSVA”.The key M2 macrophages-related genes were screened from the differential expressed genes(DEGs)and M2 macrophages activation and migration gene sets collected from MSigDB database.Our analysis identified three main cell types in primary tumors,NLNs,and PLNs:basal cells,luminal cells,and immune cell subsets.The further cell type classification of immune cell subsets indicated M2 macrophages accumulation in NLs and PLs.The GSVA enrichment scores for activation and migration capability were increased significantly in M2 macrophages from primary tumors than NLNs and PLNs(pvalue<0.001).Seven M2 macrophages activation-related and 15 M2 macrophages migration-related genes were significantly up-regulated in primary tumors than NLNs and PLNs.The proportion and GSVA enrichment scores for activation and migration of M2 macrophages may be potential markers for lymph node metastasis in breast cancer.Our study demonstrated that twenty-two up-regulated mRNA may be possible therapeutic targets for lymph node metastasis in breast cancer.展开更多
Single-cell RNA-sequencing(scRNA-seq)is a rapidly increasing research area in biomed-ical signal processing.However,the high complexity of single-cell data makes efficient and accurate analysis difficult.To improve th...Single-cell RNA-sequencing(scRNA-seq)is a rapidly increasing research area in biomed-ical signal processing.However,the high complexity of single-cell data makes efficient and accurate analysis difficult.To improve the performance of single-cell RNA data processing,two single-cell features calculation method and corresponding dual-input neural network structures are proposed.In this feature extraction and fusion scheme,the features at the cluster level are extracted by hier-archical clustering and differential gene analysis,and the features at the cell level are extracted by the calculation of gene frequency and cross cell frequency.Our experiments on COVID-19 data demonstrate that the combined use of these two feature achieves great results and high robustness for classification tasks.展开更多
The regulation of spermatogonial proliferation and apoptosis is of great significance for maintaining spermatogenesis.The single-cell RNA sequencing(scRNA-seq)analysis of the testis was performed to identify genes upr...The regulation of spermatogonial proliferation and apoptosis is of great significance for maintaining spermatogenesis.The single-cell RNA sequencing(scRNA-seq)analysis of the testis was performed to identify genes upregulated in spermatogonia.Using scRNAseq analysis,we identified the spermatogonia upregulated gene origin recognition complex subunit 6(Orc6),which is involved in DNA replication and cell cycle regulation;its protein expression in the human and mouse testis was detected by western blot and immunofluorescence.To explore the potential function of Orc6 in spermatogonia,the C18-4 cell line was transfected with control or Orc6 siRNA.Subsequently,5-ethynyl-2-deoxyuridine(EdU)and terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL)assays,flow cytometry,and western blot were used to evaluate its effects on proliferation and apoptosis.It was revealed that ORC6 could promote proliferation and inhibit apoptosis of C18-4 cells.Bulk RNA sequencing and bioinformatics analysis indicated that Orc6 was involved in the activation of wingless/integrated(Wnt)/β-catenin signaling.Western blot revealed that the expression ofβ-catenin protein and its phosphorylation(Ser675)were significantly decreased when silencing the expression of ORC6.Our findings indicated that Orc6 was upregulated in spermatogonia,whereby it regulated proliferation and apoptosis by activating Wnt/β-catenin signaling.展开更多
Background:Colorectal cancer(CRC)is a highly heterogeneous malignant tumor that significantly impacts clinical diagnosis and treatment.Single-cell RNA sequencing is an innovative method for exploring tumor heterogenei...Background:Colorectal cancer(CRC)is a highly heterogeneous malignant tumor that significantly impacts clinical diagnosis and treatment.Single-cell RNA sequencing is an innovative method for exploring tumor heterogeneity and understanding its role at cellular and genetic levels.Method:The colorectal cancer Single-cell RNA sequencing data were analysed on the immune.RNA-seq data in bulk form was utilized to assess the major genes of the immune cell subsets linked to CRC.We conducted an analysis of the abundance of immune cells in the microenvironment of CRC,and also performed weighted gene co-expression network analysis.Gene set enrichment analysis helped perform two analytical procedures of subtype groups.Furthermore,Least absolute shrinkage and selection operator regression was employed to analyse and screen for a gene signature.Finally,quantitative PCR Was performed to detect the expression levels of signature genes in CRC.Results:The Single-cell RNA sequencing(GSE146771)dataset was integrated to obtain 9 cell clusters.The Single-sample gene set enrichment analysis showed that the related gene expression of T-cell subsets of different functional statuses could vary greatly between patients with GSE146771.Immune cell analysis of TCGA-CRC indicated an improved overall survival rate for patients with elevated Th2 cell abundance.Five-gene signature(Risk Score=-0.205×CDC25C-0.231×GSTCD-0.010×KPNA2-0.002×KIF15-0.171×ORC1)was developed by weighted correlation network analysis,and lasso Cox regression.Then,the risk prediction efficacy of the signature was validated in four GSE datasets.Furthermore,the expression of five genes was reduced in CRC tissue by quantitative PCR.Conclusion:Five-gene signature based on CRC heterogeneity was developed as a prognosis predictor,which can serve as a potential treatment target.展开更多
近年来,单细胞RNA测序技术(scRNA-seq)已经被广泛使用。使用scRNA-seq技术获取的单细胞RNA数据集日益增多,数据集中的细胞数目和基因数目也不断提高。随着数据集维度的提升,支撑聚类分析所需的数据量和计算量呈指数形式剧增,传统的单细...近年来,单细胞RNA测序技术(scRNA-seq)已经被广泛使用。使用scRNA-seq技术获取的单细胞RNA数据集日益增多,数据集中的细胞数目和基因数目也不断提高。随着数据集维度的提升,支撑聚类分析所需的数据量和计算量呈指数形式剧增,传统的单细胞聚类分析方法难以应对高维度、高噪声的scRNA-seq数据。基于对抗约束自编码器的scRNA-seq数据聚类模型(Adversarially Constrained ScRNA-seq data Clustering,ACSC)通过引入自编码器的方法,对原始的高维度稀疏数据集进行降维,能够在公开单细胞数据集上具有更好的聚类准确率。展开更多
In vertebrates,bony fishes possess not only innate immune cells but also T and B cells that are equivalent to those in mammals.However,the precise sub-cluster of immune cells in teleost fish remains largely unknown.He...In vertebrates,bony fishes possess not only innate immune cells but also T and B cells that are equivalent to those in mammals.However,the precise sub-cluster of immune cells in teleost fish remains largely unknown.Herein,we developed a dynamic bacterial infection model in turbot(Scophthalmus maximus)and created a fish immune cell landscape(FICL)for a primary lymphoid organ(head kidney),a secondary lymphoid organ(spleen),and barrier tissues(gill and posterior intestine).Moreover,through comprehensive characterization of the expression profiles of 16 clusters,including dendritic cells-like(DCs-like),macrophages(Mos),neutrophils,NK cells,as well as 12 sub-clusters of T and B cells,we found that CD8+CTLs,CD4-CD8-T,Th17 and ILC3-2 like cells posssa bifunctional role associated with cytotoxicity and immunoregulation during bacterial infection.To our knowledge,these results could provide a useful resource for a better understanding of immune cells in teleost fish and could act as a comprehensive knowledge base for assessing the evolutionary mechanism of adaptive immunity in vertebrates.展开更多
Plant defense responses involve several biological processes that allow plants to fight against pathogenic attacks.How these different processes are orchestrated within organs and depend on specific cell types is poor...Plant defense responses involve several biological processes that allow plants to fight against pathogenic attacks.How these different processes are orchestrated within organs and depend on specific cell types is poorly known.Here,using single-cell RNA sequencing(scRNA-seq)technology on three independent biological replicates,we identified several cell populations representing the core transcriptional responses of wild-type Arabidopsis leaves inoculated with the bacterial pathogen Pseudomonas syringae DC3000.Among these populations,we retrieved major cell types of the leaves(mesophyll,guard,epidermal,companion,and vascular S cells)with which we could associate characteristic transcriptional reprogramming and regulators,thereby specifying different cell-type responses to the pathogen.Further analyses of transcriptional dynamics,on the basis of inference of cell trajectories,indicated that the different cell types,in addition to their characteristic defense responses,can also share similar modules of gene reprogramming,uncovering a ubiquitous antagonism between immune and susceptible processes.Moreover,it appears that the defense responses of vascular S cells,epidermal cells,and mesophyll cells can evolve along two separate paths,one converging toward an identical cell fate,characterized mostly by lignification and detoxification functions.As this divergence does not correspond to the differentiation between immune and susceptible cells,we speculate that this might reflect the discrimination between cellautonomous and non-cell-autonomous responses.Altogether our data provide an upgraded framework to describe,explore,and explain the specialization and the coordination of plant cell responses upon pathogenic challenge.展开更多
Recent advances of single-cell RNA sequencing(scRNA-seq)technologies have led to extensive study of cellular heterogeneity and cell-to-cell variation.However,the high frequency of dropout events and noise in scRNA-seq...Recent advances of single-cell RNA sequencing(scRNA-seq)technologies have led to extensive study of cellular heterogeneity and cell-to-cell variation.However,the high frequency of dropout events and noise in scRNA-seq data confounds the accuracy of the downstream analysis,i.e.clustering analysis,whose accuracy depends heavily on the selected feature genes.Here,by deriving an entropy decomposition formula,we propose a feature selection method,i.e.an intrinsic entropy(IE)model,to identify the informative genes for accurately clustering analysis.Specifically,by eliminating the‘noisy’fluctuation or extrinsic entropy(EE),we extract the IE of each gene from the total entropy(TE),i.e.TE=IE+EE.We show that the IE of each gene actually reflects the regulatory fluctuation of this gene in a cellular process,and thus high-IE genes provide rich information on celltype or state analysis.To validate the performance of the high-IE genes,we conduct computational analysis on both simulated datasets and real single-cell datasets by comparing with other representative methods.The results show that our IE model is not only broadly applicable and robust for different clustering and classification methods,but also sensitive for novel cell types.Our results also demonstrate that the intrinsic entropy/fluctuation of a gene serves as information rather than noise in contrast to its total entropy/fluctuation.展开更多
The increasing emergence of the time-series single-cell RNA sequencing(scRNA-seq)data,inferring developmental trajectory by connecting transcriptome similar cell states(i.e.,cell types or clusters)has become a major c...The increasing emergence of the time-series single-cell RNA sequencing(scRNA-seq)data,inferring developmental trajectory by connecting transcriptome similar cell states(i.e.,cell types or clusters)has become a major challenge.Most existing computational methods are designed for individual cells and do not take into account the available time series information.We present IDTI based on the Increment of Diversity for Trajectory Inference,which combines time series information and the minimum increment of diversity method to infer cell state trajectory of time-series scRNA-seq data.We apply IDTI to simulated and three real diverse tissue development datasets,and compare it with six other commonly used trajectory inference methods in terms of topology similarity and branching accuracy.The results have shown that the IDTI method accurately constructs the cell state trajectory without the requirement of starting cells.In the performance test,we further demonstrate that IDTI has the advantages of high accuracy and strong robustness.展开更多
Background:Accumulating researchers have recognized mitophagy as a key player in tumors,but few studies have investigated its role in the tumor microenvironment(TME).Advances in the technology of single-cell RNA seque...Background:Accumulating researchers have recognized mitophagy as a key player in tumors,but few studies have investigated its role in the tumor microenvironment(TME).Advances in the technology of single-cell RNA sequencing(scRNA-seq)have allowed unveiling the concealed features of the TME at cellular resolution.This study aimed to elucidate the role of mitophagy within the TME of colorectal cancer(CRC)and to establish a mitophagy-mediated risk model.Methods:We assessed mitophagy-related pathway activities at both single-cell and tissue levels.Subsequently,an unsupervised clustering algorithm was employed to identify mitophagy-mediated subtypes.Furthermore,we developed a mitophagy-mediated risk signature(MMRS)using least absolute shrinkage and selection operator(LASSO)Cox analysis and constructed a MMRS model incorporating the risk score and clinical variables.Subsequently,we used quantitative reverse transcription polymerase chain reaction analysis to verify the expression of the screened genes.Results:We retrieved and annotated a total of 14,719 cells from eight samples in the scRNA-seq GSE132465 data set.The activities of mitophagy-related pathways were uniformly upregulated in cancer cells.Integrating with bulk RNA-seq data,we identified two mitophagy-mediated clusters(C1 and C2)with distinct characteristics and prognoses.C2 was identified as a mitophagy-high cluster.Then,we developed a five-gene MMRS via LASSO Cox analysis in The Cancer Genome Atlas(TCGA)cohort.We utilized the GSE39582 cohort to validate the efficacy of our model.The expression of CX3CL1 and INHBB was upregulated in CRC tissues.Conclusions:The present study identified two mitophagy-mediated CRC subtypes with distinct features.Our MMRS may provide potential therapeutic strategies for CRC.The findings of our work offer novel insights into the involvement of mitophagy in CRC.展开更多
Objective:To investigate the substantial changes in cell types,pathways,and cell-cell interactions occurring in the irradiation-induced alopecia and dermatitis(IRIAD)mouse model and to identify potential targets for p...Objective:To investigate the substantial changes in cell types,pathways,and cell-cell interactions occurring in the irradiation-induced alopecia and dermatitis(IRIAD)mouse model and to identify potential targets for patients experiencing skin adverse reactions to radiotherapy.Methods:Mice were irradiated at 15 Gy,targeting the head and neck region.After a 14-day interval,living cells were extracted from both wild-type(WT)mice and irradiated mice for single-cell RNA sequencing(scRNA-seq).The scRNA-seq data,retrieved from the GEO database(GSE201447),underwent stringent quality control using the Seurat(v4.3.0)R package.Cell type annotation relied on previously reported typical markers and CellMarker 2.0.Differentially expressed genes were calculated to perform gene ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analyses.Cell-cell interactions were evaluated using the Cellchat R package.Results:The application of single-cell RNA sequencing(scRNA-seq)enabled a comprehensive characterization of the intricate cellular composition of both wild-type(WT)and irradiated mice skin.Remarkably,cells within irradiated mice skin exhibited a significant alteration in the intensity of cell-cell interactions compared to their wild-type counterparts.This change in interaction intensity was observed across various cell types,including fibroblast cells,endothelial cells,and dendritic cells.Importantly,these"interacting cells"shared common signaling pathways,notably the upregulation of the IL-17 pathway following irradiation.Conclusions:The modification of intercellular communication induced by irradiation primarily involves fibroblast cells,endothelial cells,and various types of immune cells.This investigation provides a novel perspective on potential targets and holds promise for enhancing the clinical management of IRIAD.展开更多
基金supported by the Foundation Project:National Natural Science Foundation of China(Nos.:82460249,82100417,81760094)The Foundation of Jiangxi Provincial Department of Science and Technology Outstanding Youth Fund Project(20242BAB23080).
文摘ABSTRACT:Background:After ischemic stroke,neutrophils hyperactivate,increasing in number and worsening inflammation,causing neural damage.Prior scRNA-seq showed Lrg1 modulates cells subsentence to cerebral ischemiareperfusion injury,but its mechanism in regulating neutrophil accumulation/differentiation post-injury is unclear.Methods:Lrg1 knockout impact on neutrophil accumulation was assessed via immunofluorescence and western blot.Three-dimensional reconstruction of immunofluorescent staining analyzed cell-cell interactions among neutrophils and microglia.scRNA-seq of WT and Lrg1^(-/-)mice from GSE245386 and GSE279462 was conducted.Each group conducted oxidative phosphorylation scoring via Gene Set Enrichment Analysis(GSEA),while Metascape was employed to perform GO and KEGG enrichment analyses for elucidating functional mechanisms.CellChat exhibited cell-cell communication.Furthermore,alterations in microglial phagocytic activity were evaluated by immunostaining for CD68,a well-established marker of phagolysosomal activity in phagocytic cells.Brain energy metabolism was evaluated via glutamate dehydrogenase activity and ATP levels with ELISA,and enzyme expression was analyzed by immunofluorescence and western blot.Results:Lrg1 knockout decreased neutrophil accumulation and NET formation in mice.3D immunofluorescence reconstruction confirmed neutrophil co-localization with endothelial cells/microglia.scRNA-seq revealed that the oxidative phosphorylation score was significantly higher in the MCAO/R+WT group compared to both the Sham-operated+WT and Lrg1^(-/-)groups.Notably,the oxidative phosphorylation score was further elevated in the MCAO/R+Lrg1^(-/-)group.Immunostaining showed that Lrg1 knockout elevated CD68+lysosome expression post-MCAO/R,with TMEM119 colocalizing with these lysosomes.MCAO/R raised CD68 expression in ischemic brains,an effect further intensified by Lrg1 knockout.KEGG analysis linked differential genes to oxidative phosphorylation pathways.Validation in MCAO/R vs.sham groups revealed increased ROS production and reduced expression of complex enzymes I-V(NDUFB8,SDHB,UQCRC1,MTCO2,ATP5A1).Lrg1 intervention increased enzyme expression.Immunofluorescence and western blot in brain tissue showed similar patterns in microglia and enzymes I-V.Conclusions:Lrg1 knockout significantly enhances microglial phagocytic activity towards neutrophils subsequent to cerebral ischemia-reperfusion injury,through its regulatory effect on the oxidative phosphorylation pathway.This finding accentuates Lrg1 as a highly potential therapeutic target for intervening in and modulating post-ischemic inflammatory responses.
基金funded by the National Natural Science Foundation of China(81700436)the Science Technology and Innovation Committee of Shenzhen Municipality,China(SGDX20190919142801722)。
文摘Gut microbes exhibit complex interactions with their hosts and shape an organism's immune system throughout its lifespan.As the largest secondary lymphoid organ,the spleen has a wide range of immunological functions.To explore the role of microbiota in regulating and shaping the spleen,we employ scRNA-seq and Stereo-seq technologies based on germ-free(GF)mice to detect differences in tissue size,anatomical structure,cell types,functions,and spatial molecular characteristics.We identify 18 cell types,9 subtypes of T cells,and 7 subtypes of B cells.Gene differential expression analysis reveals that the absence of microorganisms results in alterations in erythropoiesis within the red pulp region and congenital immune deficiency in the white pulp region.Stereo-seq results demonstrate a clear hierarchy of immune cells in the spleen,including marginal zone(MZ)macrophages,MZ B cells,follicular B cells and T cells,distributed in a well-defined pattern from outside to inside.However,this hierarchical structure is disturbed in GF mice.Ccr7 and Cxcl13 chemokines are specifically expressed in the spatial locations of T cells and B cells,respectively.We speculate that the microbiota may mediate the structural composition or partitioning of spleen immune cells by modulating the expression levels of chemokines.
文摘The process of lymphatic metastasis was proved to be associated with podoplanin-expressing macrophages in breast cancer(BC).This study aimed to investigate the role of the M2 phenotype of tumor-associated macrophages and mine the key M2 macrophages-related genes for lymph node metastasis in BC.We downloaded the GSE158399 dataset from the Gene Expression Omnibus(GEO)database,which includes transcriptomic profiles of individual cells from primary tumors,negative lymph nodes(NLNs),and positive lymph nodes(PLNs)of breast cancer patients.The cell subsets were identified by clustering analysis after quality control of the scRNA-seq using Seurat.The activation and migration capability of M2 macrophages were evaluated with R package“GSVA”.The key M2 macrophages-related genes were screened from the differential expressed genes(DEGs)and M2 macrophages activation and migration gene sets collected from MSigDB database.Our analysis identified three main cell types in primary tumors,NLNs,and PLNs:basal cells,luminal cells,and immune cell subsets.The further cell type classification of immune cell subsets indicated M2 macrophages accumulation in NLs and PLs.The GSVA enrichment scores for activation and migration capability were increased significantly in M2 macrophages from primary tumors than NLNs and PLNs(pvalue<0.001).Seven M2 macrophages activation-related and 15 M2 macrophages migration-related genes were significantly up-regulated in primary tumors than NLNs and PLNs.The proportion and GSVA enrichment scores for activation and migration of M2 macrophages may be potential markers for lymph node metastasis in breast cancer.Our study demonstrated that twenty-two up-regulated mRNA may be possible therapeutic targets for lymph node metastasis in breast cancer.
文摘Single-cell RNA-sequencing(scRNA-seq)is a rapidly increasing research area in biomed-ical signal processing.However,the high complexity of single-cell data makes efficient and accurate analysis difficult.To improve the performance of single-cell RNA data processing,two single-cell features calculation method and corresponding dual-input neural network structures are proposed.In this feature extraction and fusion scheme,the features at the cluster level are extracted by hier-archical clustering and differential gene analysis,and the features at the cell level are extracted by the calculation of gene frequency and cross cell frequency.Our experiments on COVID-19 data demonstrate that the combined use of these two feature achieves great results and high robustness for classification tasks.
基金This work was supported by the National Key Research and Development Program of China(No.2022YFC2702700)the National Natural Science Foundation of China(No.82171597)Clinical Research Plan of Shanghai Hospital Development Center(No.SHDC2020CR3077B).
文摘The regulation of spermatogonial proliferation and apoptosis is of great significance for maintaining spermatogenesis.The single-cell RNA sequencing(scRNA-seq)analysis of the testis was performed to identify genes upregulated in spermatogonia.Using scRNAseq analysis,we identified the spermatogonia upregulated gene origin recognition complex subunit 6(Orc6),which is involved in DNA replication and cell cycle regulation;its protein expression in the human and mouse testis was detected by western blot and immunofluorescence.To explore the potential function of Orc6 in spermatogonia,the C18-4 cell line was transfected with control or Orc6 siRNA.Subsequently,5-ethynyl-2-deoxyuridine(EdU)and terminal deoxynucleotidyl transferase dUTP nick end labeling(TUNEL)assays,flow cytometry,and western blot were used to evaluate its effects on proliferation and apoptosis.It was revealed that ORC6 could promote proliferation and inhibit apoptosis of C18-4 cells.Bulk RNA sequencing and bioinformatics analysis indicated that Orc6 was involved in the activation of wingless/integrated(Wnt)/β-catenin signaling.Western blot revealed that the expression ofβ-catenin protein and its phosphorylation(Ser675)were significantly decreased when silencing the expression of ORC6.Our findings indicated that Orc6 was upregulated in spermatogonia,whereby it regulated proliferation and apoptosis by activating Wnt/β-catenin signaling.
基金supported by the Guangzhou Science and Technology Plan Project(No.202201010786&2023A04J1129)the Basic Research Project of Guangzhou Municipal School(Hospital),(No.202201020483)+4 种基金the Guangdong Second Provincial General Hospital(No.3DA2021015)Doctoral workstation foundation of Guangdong Second Provincial General Hospital(2021BSGZ018)the science foundation of Guangdong Second Provincial General Hospital(TJGC-2021007)Guangdong Medical Scientific Research(grant No.B2023038)National Natural Science Foundation of China(No.82302640).
文摘Background:Colorectal cancer(CRC)is a highly heterogeneous malignant tumor that significantly impacts clinical diagnosis and treatment.Single-cell RNA sequencing is an innovative method for exploring tumor heterogeneity and understanding its role at cellular and genetic levels.Method:The colorectal cancer Single-cell RNA sequencing data were analysed on the immune.RNA-seq data in bulk form was utilized to assess the major genes of the immune cell subsets linked to CRC.We conducted an analysis of the abundance of immune cells in the microenvironment of CRC,and also performed weighted gene co-expression network analysis.Gene set enrichment analysis helped perform two analytical procedures of subtype groups.Furthermore,Least absolute shrinkage and selection operator regression was employed to analyse and screen for a gene signature.Finally,quantitative PCR Was performed to detect the expression levels of signature genes in CRC.Results:The Single-cell RNA sequencing(GSE146771)dataset was integrated to obtain 9 cell clusters.The Single-sample gene set enrichment analysis showed that the related gene expression of T-cell subsets of different functional statuses could vary greatly between patients with GSE146771.Immune cell analysis of TCGA-CRC indicated an improved overall survival rate for patients with elevated Th2 cell abundance.Five-gene signature(Risk Score=-0.205×CDC25C-0.231×GSTCD-0.010×KPNA2-0.002×KIF15-0.171×ORC1)was developed by weighted correlation network analysis,and lasso Cox regression.Then,the risk prediction efficacy of the signature was validated in four GSE datasets.Furthermore,the expression of five genes was reduced in CRC tissue by quantitative PCR.Conclusion:Five-gene signature based on CRC heterogeneity was developed as a prognosis predictor,which can serve as a potential treatment target.
文摘近年来,单细胞RNA测序技术(scRNA-seq)已经被广泛使用。使用scRNA-seq技术获取的单细胞RNA数据集日益增多,数据集中的细胞数目和基因数目也不断提高。随着数据集维度的提升,支撑聚类分析所需的数据量和计算量呈指数形式剧增,传统的单细胞聚类分析方法难以应对高维度、高噪声的scRNA-seq数据。基于对抗约束自编码器的scRNA-seq数据聚类模型(Adversarially Constrained ScRNA-seq data Clustering,ACSC)通过引入自编码器的方法,对原始的高维度稀疏数据集进行降维,能够在公开单细胞数据集上具有更好的聚类准确率。
基金supported by the National Natural Science Foundation of China[Grant Nos.32025038(Q.L.),32122090(D.Y.)]and the Natural Science Foundation of Shanghai[Grant No.20ZR1415500(D.Y.)].
文摘In vertebrates,bony fishes possess not only innate immune cells but also T and B cells that are equivalent to those in mammals.However,the precise sub-cluster of immune cells in teleost fish remains largely unknown.Herein,we developed a dynamic bacterial infection model in turbot(Scophthalmus maximus)and created a fish immune cell landscape(FICL)for a primary lymphoid organ(head kidney),a secondary lymphoid organ(spleen),and barrier tissues(gill and posterior intestine).Moreover,through comprehensive characterization of the expression profiles of 16 clusters,including dendritic cells-like(DCs-like),macrophages(Mos),neutrophils,NK cells,as well as 12 sub-clusters of T and B cells,we found that CD8+CTLs,CD4-CD8-T,Th17 and ILC3-2 like cells posssa bifunctional role associated with cytotoxicity and immunoregulation during bacterial infection.To our knowledge,these results could provide a useful resource for a better understanding of immune cells in teleost fish and could act as a comprehensive knowledge base for assessing the evolutionary mechanism of adaptive immunity in vertebrates.
基金supported by INRAE funding(SCANNER project,BAP Department)the support of Saclay Plant Sciences-SPS(ANR-17-EUR-0007).
文摘Plant defense responses involve several biological processes that allow plants to fight against pathogenic attacks.How these different processes are orchestrated within organs and depend on specific cell types is poorly known.Here,using single-cell RNA sequencing(scRNA-seq)technology on three independent biological replicates,we identified several cell populations representing the core transcriptional responses of wild-type Arabidopsis leaves inoculated with the bacterial pathogen Pseudomonas syringae DC3000.Among these populations,we retrieved major cell types of the leaves(mesophyll,guard,epidermal,companion,and vascular S cells)with which we could associate characteristic transcriptional reprogramming and regulators,thereby specifying different cell-type responses to the pathogen.Further analyses of transcriptional dynamics,on the basis of inference of cell trajectories,indicated that the different cell types,in addition to their characteristic defense responses,can also share similar modules of gene reprogramming,uncovering a ubiquitous antagonism between immune and susceptible processes.Moreover,it appears that the defense responses of vascular S cells,epidermal cells,and mesophyll cells can evolve along two separate paths,one converging toward an identical cell fate,characterized mostly by lignification and detoxification functions.As this divergence does not correspond to the differentiation between immune and susceptible cells,we speculate that this might reflect the discrimination between cellautonomous and non-cell-autonomous responses.Altogether our data provide an upgraded framework to describe,explore,and explain the specialization and the coordination of plant cell responses upon pathogenic challenge.
基金supported by grants from the National Key R&D Program of China(2017YFA0505500)the National Natural Science Foundation of China(31930022,12131020,12026608,and 31771476)+1 种基金the Strategic Priority Research Program of the Chinese Academy of Sciences(XDB38040400)JST Moonshot R&D(JPMJMS2021).
文摘Recent advances of single-cell RNA sequencing(scRNA-seq)technologies have led to extensive study of cellular heterogeneity and cell-to-cell variation.However,the high frequency of dropout events and noise in scRNA-seq data confounds the accuracy of the downstream analysis,i.e.clustering analysis,whose accuracy depends heavily on the selected feature genes.Here,by deriving an entropy decomposition formula,we propose a feature selection method,i.e.an intrinsic entropy(IE)model,to identify the informative genes for accurately clustering analysis.Specifically,by eliminating the‘noisy’fluctuation or extrinsic entropy(EE),we extract the IE of each gene from the total entropy(TE),i.e.TE=IE+EE.We show that the IE of each gene actually reflects the regulatory fluctuation of this gene in a cellular process,and thus high-IE genes provide rich information on celltype or state analysis.To validate the performance of the high-IE genes,we conduct computational analysis on both simulated datasets and real single-cell datasets by comparing with other representative methods.The results show that our IE model is not only broadly applicable and robust for different clustering and classification methods,but also sensitive for novel cell types.Our results also demonstrate that the intrinsic entropy/fluctuation of a gene serves as information rather than noise in contrast to its total entropy/fluctuation.
基金the National Natural Science Foundation of China(62061034,62171241)the key technology research program of Inner Mongolia Autonomous Region(2021GG0398)the Science and Technology Leading Talent Team in Inner Mongolia Autonomous Region(2022LJRC0009).
文摘The increasing emergence of the time-series single-cell RNA sequencing(scRNA-seq)data,inferring developmental trajectory by connecting transcriptome similar cell states(i.e.,cell types or clusters)has become a major challenge.Most existing computational methods are designed for individual cells and do not take into account the available time series information.We present IDTI based on the Increment of Diversity for Trajectory Inference,which combines time series information and the minimum increment of diversity method to infer cell state trajectory of time-series scRNA-seq data.We apply IDTI to simulated and three real diverse tissue development datasets,and compare it with six other commonly used trajectory inference methods in terms of topology similarity and branching accuracy.The results have shown that the IDTI method accurately constructs the cell state trajectory without the requirement of starting cells.In the performance test,we further demonstrate that IDTI has the advantages of high accuracy and strong robustness.
基金supported by Traditional Chinese Medicine Bureau of Guangdong Province[grant no.20221092]the Sixth Affiliated Hospital of Sun Yat-sen University,1010 Clinical Research Project[grant no.1010PY(2020)-28].
文摘Background:Accumulating researchers have recognized mitophagy as a key player in tumors,but few studies have investigated its role in the tumor microenvironment(TME).Advances in the technology of single-cell RNA sequencing(scRNA-seq)have allowed unveiling the concealed features of the TME at cellular resolution.This study aimed to elucidate the role of mitophagy within the TME of colorectal cancer(CRC)and to establish a mitophagy-mediated risk model.Methods:We assessed mitophagy-related pathway activities at both single-cell and tissue levels.Subsequently,an unsupervised clustering algorithm was employed to identify mitophagy-mediated subtypes.Furthermore,we developed a mitophagy-mediated risk signature(MMRS)using least absolute shrinkage and selection operator(LASSO)Cox analysis and constructed a MMRS model incorporating the risk score and clinical variables.Subsequently,we used quantitative reverse transcription polymerase chain reaction analysis to verify the expression of the screened genes.Results:We retrieved and annotated a total of 14,719 cells from eight samples in the scRNA-seq GSE132465 data set.The activities of mitophagy-related pathways were uniformly upregulated in cancer cells.Integrating with bulk RNA-seq data,we identified two mitophagy-mediated clusters(C1 and C2)with distinct characteristics and prognoses.C2 was identified as a mitophagy-high cluster.Then,we developed a five-gene MMRS via LASSO Cox analysis in The Cancer Genome Atlas(TCGA)cohort.We utilized the GSE39582 cohort to validate the efficacy of our model.The expression of CX3CL1 and INHBB was upregulated in CRC tissues.Conclusions:The present study identified two mitophagy-mediated CRC subtypes with distinct features.Our MMRS may provide potential therapeutic strategies for CRC.The findings of our work offer novel insights into the involvement of mitophagy in CRC.
基金National Natural Science Foundation of China(82373525,31971165,82173465)Leading Talents Program of Gusu District,China(ZXL2022454,LC)+1 种基金Jiangsu Provincial Outstanding Postdoctoral Program,China(2023ZB254,MT-J)Youth Fund of Jiangsu Provincial Natural Science Foundation,China(BK20230490,MT-J).
文摘Objective:To investigate the substantial changes in cell types,pathways,and cell-cell interactions occurring in the irradiation-induced alopecia and dermatitis(IRIAD)mouse model and to identify potential targets for patients experiencing skin adverse reactions to radiotherapy.Methods:Mice were irradiated at 15 Gy,targeting the head and neck region.After a 14-day interval,living cells were extracted from both wild-type(WT)mice and irradiated mice for single-cell RNA sequencing(scRNA-seq).The scRNA-seq data,retrieved from the GEO database(GSE201447),underwent stringent quality control using the Seurat(v4.3.0)R package.Cell type annotation relied on previously reported typical markers and CellMarker 2.0.Differentially expressed genes were calculated to perform gene ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analyses.Cell-cell interactions were evaluated using the Cellchat R package.Results:The application of single-cell RNA sequencing(scRNA-seq)enabled a comprehensive characterization of the intricate cellular composition of both wild-type(WT)and irradiated mice skin.Remarkably,cells within irradiated mice skin exhibited a significant alteration in the intensity of cell-cell interactions compared to their wild-type counterparts.This change in interaction intensity was observed across various cell types,including fibroblast cells,endothelial cells,and dendritic cells.Importantly,these"interacting cells"shared common signaling pathways,notably the upregulation of the IL-17 pathway following irradiation.Conclusions:The modification of intercellular communication induced by irradiation primarily involves fibroblast cells,endothelial cells,and various types of immune cells.This investigation provides a novel perspective on potential targets and holds promise for enhancing the clinical management of IRIAD.
