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In recent years, data-driven inference of cell-cell communication has helped reveal coordinated biological processes across cell types. Here, we integrate two tools, LIANA and Tensor-cell2cell, which, when combined, can deploy multiple existing methods and resources to enable the robust and flexible identification of cell-cell communication programs across multiple samples. In this work, we show how the integration of our tools facilitates the choice of method to infer cell-cell communication and subsequently perform an unsupervised deconvolution to obtain and summarize biological insights. We explain how to perform the analysis step by step in both Python and R and provide online tutorials with detailed instructions available at https://ccc-protocols.readthedocs.io/. This workflow typically takes ∼1.5 h to complete from installation to downstream visualizations on a graphics processing unit-enabled computer for a dataset of ∼63,000 cells, 10 cell types, and 12 samples.

BACKGROUND: A traditional view is that stem cells (SCs) divide slowly. Meanwhile, both embryonic and pluripotent SCs display a shorter cell cycle duration (CCD) in comparison to more committed progenitors (CPs). METHODS: We examined the in vitro proliferation and cycling behavior of somatic adult human cells using live cell imaging of passage zero keratinocytes and single-cell RNA sequencing. RESULTS: We found two populations of keratinocytes: those with short CCD and protracted near exponential growth, and those with long CCD and terminal differentiation. Applying the ergodic principle, the comparative numbers of cycling cells in S phase in an enriched population of SCs confirmed a shorter CCD than CPs. Further, analysis of single-cell RNA sequencing of cycling adult human keratinocyte SCs and CPs indicated a shortening of both G1 and G2M phases in the SC. CONCLUSIONS: Contrary to the pervasive paradigm, SCs progress through cell cycle more quickly than more differentiated dividing CPs. Thus, somatic human adult keratinocyte SCs may divide infrequently, but divide rapidly when they divide. Additionally, it was found that SC-like proliferation persisted in vitro.

Background: Recent advancements in contemporary therapeutic approaches have increased the survival rates of lung cancer patients; however, the long-term benefits remain constrained, underscoring the pressing need for novel biomarkers. Surfactant-associated 3 (SFTA3), a long non-coding RNA predominantly expressed in normal lung epithelial cells, plays a crucial role in lung development. Nevertheless, its function in lung adenocarcinoma (LUAD) remains inadequately understood. Methods: Single-cell RNA sequencing data were utilized to identify novel cancer cell-intrinsic gene signatures associated with the progression of LUAD, and their roles in LUAD were comprehensively analyzed. Serum samples were collected to quantify the expression levels of SFTA3 in LUAD patients. Furthermore, a series of biological experiments, including cell viability assays, scratch wound healing assays, and colony formation assays, were conducted to demonstrate the tumor-suppressive effects of SFTA3. RNA sequencing was performed to elucidate the molecular mechanisms underlying the role of SFTA3 in lung cancer cells. Results: We constructed a prognostic model comprising eight genes: ALDOA, ATP5MD, SERPINH1, SFTA3, SLK, U2SURP, SCGB1A1, and SCGB1A3. The model effectively stratified patients into high- and low-risk categories, revealing that low-risk patients experienced superior clinical outcomes, exhibited an immunologically hot tumor microenvironment (TME), and had a greater probability of responding to immunotherapy. In contrast, the high-risk group exhibited a cold TME and may benefit more from chemotherapy. Furthermore, our study revealed that a progressive decrease in SFTA3 expression in cancer cells was correlated with tumor advancement. Notably, the serum levels of SFTA3 significantly decreased in patients with LUAD, suggesting its potential utility in liquid biopsy for LUAD diagnosis. Additionally, the knockdown of SFTA3 enhances the proliferation and migration of lung cancer cells, whereas its overexpression inhibits these phenotypes. The epithelial-mesenchymal transition pathway was significantly enriched following SFTA3 silencing, suggesting that SFTA3 may impact tumor progression by modulating this process. We also identified key transcription factors and epigenetic mechanisms implicated in the downregulation of SFTA3 in LUAD. Conclusion: We developed a robust prognostic model and identified SFTA3 as a novel biomarker with potential applications in the diagnosis, prognosis, and personalized treatment of LUAD. Additionally, our findings offer new insights into the mechanisms underlying LUAD tumorigenesis and immune evasion. [ABSTRACT FROM AUTHOR]

BACKGROUND: Angiogenesis is the main treatment target of cardiovascular diseases. Bone morphogenetic protein 2 can modulate angiogenesis, but the regulatory effect of endothelial cell-specific bone morphogenetic protein 2 on angiogenesis is unclear OBJECTIVE: To investigate the effect of endothelial-specific bone morphogenetic protein 2 on angiogenesis. METHODS: (1) Bioinformatics analysis: Cellular expression specificity and abundance of bone morphogenetic protein 2 were meta-analyzed by the PanglaoDB single-cell transcriptome database. The endothelial cell transcriptome sequencing dataset of the mouse hindlimb model and endocardial transcriptome dataset of mice overexpressing bone morphogenetic protein 2 were reanalyzed to evaluate the effect of endothelial cell bone morphogenetic protein 2 on the angiogenesis pathway. (2) Validation in vivo: After establishing the mouse hindlimb model, we compared the blood perfusion between the affected and sham limb at 7, 14, and 21 days. The expression of the colocation of bone morphogenetic protein 2 and CD31 was explored by immunofluorescence and immunohistochemical staining. (3) Validation in vitro: The cultured human umbilical vein endothelial cells in vitro were divided into a control group, a hypoxia group, and a bone morphogenetic protein 2 inhibitor Noggin intervention group. After being cultured for 24 hours, the angiogenesis of endothelial cells in each group was observed. RESULTS AND CONCLUSION: (1) Endothelial cells are an important cell subgroup expressing bone morphogenetic protein 2. Both in the mouse hindlimb ischemia model and endocardial cells overexpressing bone morphogenetic protein 2, bone morphogenetic protein 2 was significantly up-regulated, and the angiogenesis pathway was significantly activated. (2) In the mouse hindlimb model, bone morphogenetic protein 2-positive blood vessels around neoangiogenesis increased significantly at 7 days of ischemia (P < 0.05), and decreased significantly after 2 weeks of ischemia (P < 0.001). (3) In umbilical vein endothelial cells cultured in vitro, after hypoxic intervention, the migration and sprouting of endothelial cells increased significantly, and the expression of angiogenesis factors vascular endothelial growth factor and platelet-derived growth factor was significantly increased. Noggin significantly reduced hypoxiainduced endothelial cell angiogenesis (P < 0.001) and down-regulated the expression of vascular endothelial growth factor and platelet-derived growth factor (P < 0.01). (4) These findings verify that endothelial cell-specific bone morphogenetic protein 2 can regulate angiogenesis, and targeting endothelial cell bone morphogenetic protein 2 is a promising way to improve angiogenesis. [ABSTRACT FROM AUTHOR]

Reliable methods for detecting and analyzing gene expression are necessary tools for understanding development and investigating biological responses to genetic and environmental perturbation. With its fully sequenced genome, invariant cell lineage, transparent body, wiring diagram, detailed anatomy, and wide array of genetic tools, Caenorhabditis elegans is an exceptionally useful model organism for linking gene expression to cellular phenotypes. The development of new techniques in recent years has greatly expanded our ability to detect gene expression at high resolution. Here, we provide an overview of gene expression methods for C. elegans , including techniques for detecting transcripts and proteins in situ, bulk RNA sequencing of whole worms and specific tissues and cells, single-cell RNA sequencing, and high-throughput proteomics. We discuss important considerations for choosing among these techniques and provide an overview of publicly available online resources for gene expression data. [ABSTRACT FROM AUTHOR]

• NT5E and TMEM158 as biomarkers of IDD associated with necroptosis. • The key cells in NT5E and TMEM158 were fibrochondrocytes and inflammatory chondrocytes, respectively, suggesting a potential role for chondrocytes in IDD. • TMEM158 and NT5E are co-regulated by GATA2 and FOXL1 to influence IDD development and progression. Intervertebral disc degeneration (IDD) is associated with back pain; back pain is a world-wide contributor to poor quality of life, while necroptosis has the characteristics of necroptosis and apoptosis, however, its role in IDD is still unclear. Therefore, the aim of this study was to identify biomarkers associated with necroptosis in IDD. To explore biomarkers associated with necroptosis in IDD, reveal the pathogenesis of IDD, as well as provide new directions for the diagnosis and treatment of this disease. Retrospective cohort study. Our study employs scRNA-seq coupled with MR analysis to investigate the causal relationship between necroptosis and IDD, laying a foundational groundwork for unveiling the intricate pathogenic mechanisms of this condition. Data quality control and normalisation was executed in single-cell dataset, GSE205535. Then, different cell types were obtained by cell annotation through marker genes. Subsequently, chi-square test was employed to assess the distribution difference of different cell types between IDD and control to screen key cells. AUCell was applied to calculate necroptosis-related genes (NRGs) scores of all cell types, further key cells were divided into high and low NRGs groups according to the median AUC scores of different cell types. Afterwards, the differentially expressed genes (DEGs) within the 2 score groups were screened. Then, the genes that had causal relationship with IDD were selected as biomarkers by univariate and multivariate Mendelian randomization (MR) analysis. Finally, the expression of biomarkers in different cell types and pseudo-time analysis was analyzed separately. In GSE205535, 16 different cell populations identified by UMAP cluster analysis were further annotated to 8 cell types using maker genes. Afterwards, 53 DEGs were screened between the high and low NRGs groups. In addition, 9 genes with causal relationship with IDD were obtained by univariate MR analysis, further multivariate MR analysis proved that NT5E and TMEM158 had a direct causal relationship with IDD, which were used as biomarkers in this study. This study not only found that the expression levels of NT5E and TMEM158 were higher in IDD group, but also found that fibrochondrocytes and inflammatory chondrocytes were the key cells of NT5E and TMEM158 , respectively. In the end, the biomarkers had the same expression trend in the quasi-time series, and both of them from high to low and then increased. NT5E and TMEM158 , as biomarkers of necroptotic apoptotic IDD, were causally associated with IDD. The understanding of chondrocytes as key cells provides new perspectives for deeper elucidation of the pathogenesis of IDD, improved diagnostic methods, and the development of more effective treatments. These findings are expected to provide a more accurate and personalised approach to clinical diagnosis and treatment, thereby improving the prognosis and quality of life of patients with IDD. [ABSTRACT FROM AUTHOR]

Dilated cardiomyopathy (DCM) is a significant cause of heart failure that requires heart transplantation. Fibroblasts play a central role in the fibro-inflammatory microenvironment of DCM. However, their cellular heterogeneity and interaction with immune cells have not been well identified. An integrative analysis was conducted on single-cell RNA sequencing (ScRNA-Seq) data from human left ventricle tissues, which comprised 4 hearts from healthy donors and 6 hearts with DCM. The specific antigen-presenting fibroblast (apFB) was explored as a subtype of fibroblasts characterized by expressing MHCII genes, the existence of which was confirmed by immunofluorescence staining of 3 cardiac tissues from DCM patients with severe heart failure. apFB highly expressed the genes that response to IFN-γ, and it also have a high activity of the JAK-STAT pathway and the transcription factor RFX5. In addition, the analysis of intercellular communication between apFBs and CD4+T cells revealed that the anti-inflammatory ligand-receptor pairs TGFB-TGFR, CLEC2B-KLRB1, and CD46-JAG1 were upregulated in DCM. The apFB signature exhibited a positive correlation with immunosuppression and demonstrated diagnostic and prognostic value when evaluated using a bulk RNA dataset comprising 166 donors and 166 DCM samples. In conclusion, the present study identified a novel subpopulation of fibroblasts that specifically expresses MHCII-encoding genes. This specific apFBs can suppress the inflammation occurring in DCM. Our findings further elucidate the composition of the fibro-inflammatory microenvironment in DCM, and provide a novel therapeutic target. [ABSTRACT FROM AUTHOR]