Single-cell Transcriptome Sequencing Research Articles

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.

Single-cell transcriptomic sequencing analysis of mechanistic insights into the IFN-γ signaling pathway in different tumor cells.

This study aimed to investigate the relationship between the interferon-gamma (IFN-γ) pathway in different tumor microenvironments (TME) and patients' prognosis, as well as the regulatory mechanisms of this pathway in tumor cells. Using RNA-seq data from the TCGA database, we analyzed the predictive value of the IFN-γ pathway across various tumors. We employed a univariate Cox regression model to assess the prognostic significance of IFN-γ signaling in different tumor types. Additionally, we analyzed single-cell RNA sequencing (scRNA-seq) data from the Gene Expression Omnibus (GEO) database to examine the distribution characteristics of the IFN-γ pathway and explore its regulatory mechanisms, highlighting how IFN-γ influenced cellular interactions within the TME. Our analysis revealed a significant association between the IFN-γ pathway and adverse prognosis in pan-cancer tissues (P < 0.001). Interestingly, this correlation varied regarding positive and negative regulation across different tumor types. Through a detailed examination of scRNA-seq data, we found that the IFN-γ pathway exerted substantial regulatory effects on stromal and immune cells. In contrast, its expression and regulatory patterns in tumor cells exhibited diversity and heterogeneity. Further analysis indicated that the IFN-γ pathway not only enhanced the immunogenicity of tumor cells but also inhibited their proliferation. Cell-cell interaction analysis confirmed the pivotal role of the IFN-γ pathway within the overall regulatory network. Moreover, we identified HMGB2 (high mobility group box 2) in T cells as a potential key regulator of tumor cell proliferation. The IFN-γ pathway exhibited a dual function by both suppressing tumor cell proliferation and enhancing their immunogenicity, positioning it as a pivotal target for refined cancer diagnosis and cancer strategies.

Establishment and transcriptome analysis of single blastomere-derived cell lines from zebrafish

Maintaining chromosome euploidy in zebrafish embryonic cells is challenging because of the degradation of genomic integrity during cell passaging. In this study, we report the derivation of zebrafish cell lines from single blastomeres. These cell lines have a stable chromosome status attributed to BMP4 and exhibit continuous proliferation in vitro. Twenty zebrafish cell lines are successfully established from single blastomeres. Single-cell transcriptome sequencing analysis confirms the fidelity of gene expression profiles throughout long-term culturing of at least 45 passages. The long-term cultured cells are specialized into epithelial cells, exhibiting similar expression patterns validated by integrative transcriptomic analysis. Overall, this work provides a protocol for establishing zebrafish cell lines from single blastomeres, which can serve as valuable tools for in vitro investigations of epithelial cell dynamics in terms of life–death balance and cell fate determination during normal homeostasis.

Comparison among different preclinical models derived from the same patient with a non-functional pancreatic neuroendocrine tumor.

Pancreatic neuroendocrine tumors are the second most common tumors of the pancreas, and approximately half of patients are diagnosed with liver metastases. Currently, the improvement in the efficacy of relevant treatment methods is still limited. Therefore, there is an urgent need for in-depth research on the molecular biological mechanism of pancreatic neuroendocrine tumors. However, due to their relatively inert biology, preclinical models are extremely scarce. Here, the patient-derived organoid, and patient-derived xenograft were successfully constructed. These two models and the previously constructed cell line named SPNE1 all derived from the same patient with a grade 3 non-functional pancreatic neuroendocrine tumor, providing new tumor modeling platforms, and characterized using immunohistochemistry, whole-exome sequencing, and single-cell transcriptome sequencing. Combined with a tumor formation experiment in immunodeficient mice, we selected the model that most closely recapitulated the parental tumor. Overall, the patient-derived xenograft model most closely resembled human tumor tissue.

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).

Combining single-cell spatial transcriptomics and molecular simulation to develop in vivo probes targeting the perineural invasion region of adenoid cystic carcinoma

Background and objectivesPerineural invasion (PNI) refers to the invasion, encasement, or penetration of tumor cells around or through nerves. Various malignant tumors, including pancreatic cancer, head and neck tumors, and bile duct cancer, exhibit the characteristic of PNI. Particularly, in head and neck-skull base tumors such as adenoid cystic carcinoma (ACC), PNI is a significant factor leading to incomplete surgical resection and postoperative recurrence. MethodsSpatial transcriptomic and single-cell transcriptomic sequencing were conducted on a case of ACC tissue with PNI to identify potential probes targeting PNI. The efficacy of the probes was validated through in vivo and in vitro experiments. ResultsSpatial transcriptomic and single-cell RNA sequencing revealed phenotypic changes in Schwann cells within the PNI region of ACC. Peptide probes were designed based on the antigen-presenting characteristics of Schwann cells in the PNI region, which are dependent on Major Histocompatibility Complex II (MHC-II) molecules. Successful validation in vitro and in vivo experiments confirmed that these probes can label viable Schwann cells in the PNI region, serving as a tool for dynamic in vivo marking of tumor invasion into nerves. ConclusionsPeptide probes targeting Schwann cells' MHC-II molecules have the potential to demonstrate the occurrence of PNI in patients with ACC

Single-cell sequencing of facial adipose tissue unveils FKBP5 as a therapeutic target for facial infiltrating lipomatosis

BackgroundFacial infiltrating lipomatosis is characterized by excessive growth of adipose tissue. Its etiology is associated with somatic phosphatidylinositol 3-kinase catalytic subunit alpha (PIK3CA) variants, but the specific mechanisms are not yet fully understood.MethodsWe collected facial adipose tissue from both FIL patients and non-FIL individuals, isolated the stromal vascular fraction (SVF) and performed single-cell transcriptome sequencing on these samples.ResultsWe mapped out the cellular landscape within the SVF, with a specific focus on a deeper analysis of fibro-adipogenic precursor cells (FAPs). Our analysis revealed that FAPs from FIL patients (FIL-FAPs) significantly overexpressed FK506 binding protein 51 (FKBP5) compared to FAPs from individuals without FIL. Further experiments indicated that FKBP5 is regulated by the PI3K-AKT signaling pathway. The overactivation of this pathway led to an increase in FKBP5 expression. In vitro experiments demonstrated that FKBP5 promoted adipogenic differentiation of FAPs, a process that could be hindered by FKBP5 knockdown or inhibition. Additionally, in vivo assessments confirmed FKBP5’s role in adipogenesis.ConclusionsThese insights into the pathogenesis of FIL underscore FKBP5 as a promising target for developing non-surgical interventions to manage the excessive adipose tissue growth in FIL.

Integration of single-cell and bulk RNA sequencing revealed immune heterogeneity and its association with disease activity in rheumatoid arthritis patients.

Rheumatoid arthritis (RA) is a chronic, inflammatory, systemic autoimmune disease characterized by cartilage, bone damage, synovial inflammation, hyperplasia, autoantibody production, and systemic features. To obtain an overall profile of the immune environment in RA patients and its association with clinical features, we performed single-cell transcriptome and T-cell receptor sequencing of mononuclear cells from peripheral blood (PBMC) and synovial fluid (SF) from RA patients, integrated with two large cohorts with bulk RNA sequencing for further validation and investigation. Dendritic cells (DCs) exhibited relatively high functional heterogeneity and tissue specificity in relation to both antigen presentation and proinflammatory functions. Peripheral helper T cells (TPHs) are likely to originate from synovial tissue, undergo activation and exhaustion, and are subsequently released into the peripheral blood. Notably, among all immune cell types, TPHs were found to have the most intense associations with disease activity. In addition, CD8 effector T cells could be clustered into two groups with different cytokine expressions and play distinct roles in RA development. By integrating single-cell data with bulk sequencing from two large cohorts, we identified interactions among TPHs, CD8 cells, CD16 monocytes, and DCs that strongly contribute to the proinflammatory local environment in RA joints. Of note, the swollen 28-joint counts exhibited a more pronounced association with this immune environment compared to other disease activity indexes. The immune environment alternated significantly from PBMCs to SF, which indicated that a series of immune cells was involved in proinflammatory responses in the local joints of RA patients. By integrating single-cell data with two large cohorts, we have uncovered associations between specific immune cell populations and clinical features. This integration provides a rapid and precise methodology for assessing local immune activation, offering valuable insights into the pathophysiological mechanisms at play in RA.

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.

Isolation and characterization of normal mandibular periosteal stem cells from human and macaca mulatta and cross-species single-cell analysis

Objective: To investigate the presence of a distinct stem cell populations different from mesenchymal stem cells in the mandibular periosteum of both human and non-human primates (macaca mulatta), to explore its properties during intramembranous osteogenesis and to establish standard protocols for the isolation, culturing and expanding of mandibular periosteal stem cells (PSC) distinguished from other PSCs in other anatomical regions. Methods: Periosteum was harvested from the bone surface during flap bone removal in patients aged 18-24 years undergoing third molar extraction and from the buccal side of the mandibular premolar region of 6-year-old macaca mulatta respectively, and then subjected to single-cell sequencing using the Illumina platform Novaseq 6000 sequencer. Cross-species single-cell transcriptome sequencing results were compared using homologous gene matching. PSC were isolated from primary tissues using two digestion methods with body temperature and low temperature, and their surface markers (CD200, CD31, CD45 and CD90) were identified by cell flow cytometry. The ability of cell proliferation and three-lineage differentiation of PSC expanded to the third generation in vitro in different species were evaluated. Finally, the similarities and differences in osteogenic properties of PSC and bone marrow mesenchymal stem cells (BMSC) were compared. Results: The single-cell sequencing results indicated that 18 clusters of cell populations were identified after homologous gene matching for dimensionality reduction, and manual cellular annotation was conducted for each cluster based on cell marker databases. The comparison of different digestion protocols proved that the low-temperature overnight digestion protocol can stably isolate PSC from the human and m. mulatta mandibular periosteum and the cells exhibited a fibroblast-like morphology. This research confirmed that PSC of human and m. mulatta had similar proliferation capabilities through the cell counting kit-8 assay. Flow cytometry analysis was then used to identify the cells isolated from the periosteum expressed CD200(+), CD31(-), CD45(-), CD90(-). Then, human and m. mulatta PSC were induced into osteogenesis, adipogenesis, and chondrogenesis to demonstrate their corresponding multi-lineage differentiation capabilities. Finally, comparison with BMSC further clarified the oesteogenesis characteristics of PSC. The above experiments proved that the cells isolated from the periosteum were peiosteal cells with characteristics of stem cells evidenced by their cell morphology, proliferation ability, surface markers, and differentiation ability, and that this group of PSC possessed characteristics different from traditional mesenchymal stem cells. Conclusions: In this study, normal mandibular PSC from humans and m. mulatta were stably isolated and identified for the first time, providing a cellular foundation for investigating the mechanism of mandibular intramembranous osteogenesis, exploring ideal non-human primate models and establishing innovative strategies for clinically mandibular injury repair.

P-682 Malic enzyme 1 in granulosa cells promotes ovarian aging through inhibiting the AMPK-FoXO1-IDH2 signaling pathway

Abstract Study question Reduced levels of oxidative phosphorylation in granulosa cells are a key factor in regulating ovarian aging, but the upstream regulatory mechanisms are unknown. Summary answer Our study revealed that elevated malic enzyme 1 in granulosa cells promotes ovarian aging by inhibiting oxidative phosphorylation levels through the AMPK-FoXO1-IDH2 signaling pathway. What is known already With ovarian aging, the ovarian granulosa cells appear hallmarks of high apoptosis activity and low-proliferative activity, which lead to a reduction in ovarian reserve and a decrease in hormone synthesis, resulting in female infertility and reduced quality of life. And the decline of isocitrate dehydrogenase, IDH, in non-human primate ovarian granulosa cells is a key causative factor for the decline in the level of oxidative phosphorylation in granulosa cells. Recently, researchers reported that malic enzyme 1, which encodes a cytosolic, NADP-dependent Enzyme that generates NADPH for fatty acid biosynthesis inhibits IDH2 expression in tumor cells. Study design, size, duration Utilizing clinical samples, cells and animal models, with the help of research tools such as biochemical experiments, gene editing and single-cell transcriptome sequencing, at different levels, including molecular, cellular, animal and human samples, we systematically analyzed the characteristic changes and key molecules in granulosa cells along with ovarian aging and reveal the intracellular molecular pathways through which ME1 and IDH2 regulate the process of ovarian aging for 1 year. Participants/materials, setting, methods We first apply single-cell transcriptome sequencing analysis of young and old mouse ovaries to reveal cellular subpopulation changes and transcriptomic hallmarks of ovarian granulosa cell with aging. And then we validate the biological function and revealing the molecular mechanism to regulate ovarian aging by using physiological ovarian aging mouse model, clinical ovarian granulosa cells from young and old female and human ovarian granulosa cell line KGN utilizing technologies of protein immunology, qPCR and CRISPR/Cas9. Main results and the role of chance Ovarian single-cell transcriptome sequencing data from young and old mice showed that ovarian aging was accompanied by a dramatic decrease in the number of granulosa cells and drastic changes in the proportions of different subpopulations. The proportion of mitotically active granulosa cells was significantly lower in aging ovaries than in young mouse ovaries, and GO enrichment analysis showed that granulosa cell cycle, antioxidant and hormone synthesis-related signaling pathways were significantly down-regulated in aging ovaries. Using immunoblotting, immunofluorescence and qPCR, we demonstrated that the expression of ME1 in granulosa cells of aging mice and human ovaries was significantly elevated, while the expression of IDH2 was significantly decreased. Performing the overexpression and knockdown of ME1 in human ovarian granulosa cell line KGN, which further demonstrated that ME1 could inhibit the transcriptionally regulated IDH2 expression of downstream genes, such as FoXO1, by suppressing the phosphorylation of AMPK. The decrease of IDH2 in mitochondria reduces the level of oxidative phosphorylation in granulosa cells, resulting in the increase of intracellular reactive oxygen species, activation of the intracellular p53-p21 signaling pathway, inhibition of the cell cycle, and promotion of apoptosis, which in turn impairs the function of granulosa cells to synthesize hormones and promotes ovarian aging. Limitations, reasons for caution Malic enzyme 1, which links the glycolytic cycle to the citric acid cycle, expressing in other ovarian cells including oocytes, and the regulation of its expression is complex. Future studies will focus on its biological functions in other ovarian cells and its molecular mechanism in regulating ovarian aging. Wider implications of the findings Our study suggests that granulosa cell ME1 and IDH2 co-regulate oxidative phosphorylation levels to regulate ovarian aging, and that ME1 and IDH2 may serve as potential targets for the finding and treatment of premature ovarian aging and delaying ovarian aging in women. Trial registration number not applicable

The correlation between TRIM28 expression and immune checkpoints in CRPC.

This study delves into the unexplored realm of castration-resistant prostate cancer (CRPC) by investigating the role of TRIM28 and its intricate molecular mechanisms using high-throughput single-cell transcriptome sequencing and advanced bioinformatics analysis. Our comprehensive examination unveiled dynamic TRIM28 expression changes, particularly in immune cells such as macrophages and CD8+ T cells within CRPC. Correlation analyses with TCGA data highlighted the connection between TRIM28 and immune checkpoint expression and emphasized its pivotal influence on the quantity and functionality of immune cells. Using TRIM28 knockout mouse models, we identified differentially expressed genes and enriched pathways, unraveling the potential regulatory involvement of TRIM28 in the cGAS-STING pathway. Invitro, experiments further illuminated that TRIM28 knockout in prostate cancer cells induced a notable anti-tumor immune effect by inhibiting M2 macrophage polarization and enhancing CD8+ T cell activity. This impactful discovery was validated in an insitu transplant tumor model, where TRIM28 knockout exhibited a deceleration in tumor growth, reduced proportions of M2 macrophages, and enhanced infiltration of CD8+ T cells. In summary, this study elucidates the hitherto unknown anti-tumor immune role of TRIM28 in CRPC and unravels its potential regulatory mechanism via the cGAS-STING signaling pathway. These findings provide novel insights into the immune landscape of CRPC, offering promising directions for developing innovative therapeutic strategies.

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.

Cancer-Associated Endocrine Cells Participate in Pancreatic Carcinogenesis

Background & AimsThe pancreas is composed of endocrine and exocrine parts, and its interlacing structure indicates potential interaction between endocrine and exocrine cells. Although the tumor microenvironment of pancreatic ductal adenocarcinoma (PDAC) has been well characterized, the role of pancreatic endocrine cells during carcinogenesis is relatively understudied. MethodsWe depicted the changes of endocrine cells in PDAC by single-cell transcriptome sequencing, spatial transcriptome sequencing and multiplex immunohistochemistry. After that, the interaction between pancreatic carcinogenesis and endocrine changes was explored in orthotopic transplantation mice, KC mice and KPC mice. Finally, we proved the mechanism of the interaction between endocrine and exocrine parts of the pancreas through islet isolation, co-culture in vitro and co-injection in vivo. ResultsWe found that pancreatic endocrine cells displayed significantly different transcriptomic characteristics and increased interaction with exocrine part in PDAC. Specifically, among all the changes, pancreatic polypeptide positive (PPY+) cells showed a sharp increment accompanied with the progression of the cancer lesion, which might be derived from the transdifferentiation of α and β cells. Interestingly, it was proved that PDAC cells were able to induce the transdifferentiation of pancreatic α cells and β cells into GCG+PPY+ and INS+PPY+ double-positive cells, which further promoted carcinogenesis and development of PDAC in a paracrine-dependent manner and formed a reciprocal interaction. ConclusionsOur study systematically maps the alteration of pancreatic endocrine cells in PDAC and elucidates the potential endocrine-exocrine interaction mechanisms during PDAC carcinogenesis. Meanwhile, we first time define and characterize cancer-associated endocrine cells (CAEs), thereby further broadening the composition of PDAC microenvironment.

The relationship between psoriasis and vitiligo: From a comprehensive study.

Both psoriasis and vitiligo are autoimmune skin diseases. Previous observational studies have indicated a relationship between the two conditions, and simultaneous onset of both diseases poses increased health risks to patients. However, limited research has explored the causal relationship between psoriasis and vitiligo. To investigate whether a causal association exists between psoriasis and vitiligo. A case of Chinese patients diagnosed with psoriasis and vitiligo has been reported. Transcriptome sequencing was performed on normal, psoriasis, vitiligo, and co-morbid skin tissues of the patients, and single-cell transcriptome sequencing was conducted on the co-morbid skin tissues. A comprehensive Mendelian randomization analysis of Genome-wide association studies (GWAS) was performed on a cohort of 261 018 European individuals with psoriasis from the IEU Open GWAS Project and vitiligo from the National Institutes of Health (NIH) Database of Genotypes and Phenotypes. Case report and transcriptome results showed that skin tissue with vitiligo combined with psoriasis exhibited both vitiligo and psoriasis. Single-cell transcriptome sequencing results showed that in comparison to normal skin and psoriatic skin, the proportions of CD8+ T cells, natural killer cells, naive T cells, T helper cells 17, regulatory T cells, conventional type 1 dendritic cells, Conventional type 2 dendritic cells, and plasmacytoid dendritic cells were all increased in skin tissue with vitiligo combined with psoriasis. Mendelian randomization analysis included 4510 patients with psoriasis and 4680 patients with vitiligo. The results showed no causal relationship between vitiligo and psoriasis in the forward direction (p=0.192; odds ratio [OR], 1.059; 95% confidence interval [CI], 0.971-1.155) or in the reverse direction (p=0.459; OR, 0.927; 95% CI, 0.757-1.134). This study suggests that the association between psoriasis and vitiligo may be closely related to immunity, however, Mendelian randomization studies do not support a causal relationship. These findings hold significant implications for clinicians aiming to enhance their understanding and treatment approaches for psoriasis and vitiligo.

SFINN: inferring gene regulatory network from single-cell and spatial transcriptomic data with shared factor neighborhood and integrated neural network.

The rise of single-cell RNA sequencing (scRNA-seq) technology presents new opportunities for constructing detailed cell type-specific gene regulatory networks (GRNs) to study cell heterogeneity. However, challenges caused by noises, technical errors, and dropout phenomena in scRNA-seq data pose significant obstacles to GRN inference, making the design of accurate GRN inference algorithms still essential. The recent growth of both single-cell and spatial transcriptomic sequencing data enables the development of supervised deep learning methods to infer GRNs on these diverse single-cell datasets. In this study, we introduce a novel deep learning framework based on shared factor neighborhood and integrated neural network (SFINN) for inferring potential interactions and causalities between transcription factors and target genes from single-cell and spatial transcriptomic data. SFINN utilizes shared factor neighborhood to construct cellular neighborhood network based on gene expression data and additionally integrates cellular network generated from spatial location information. Subsequently, the cell adjacency matrix and gene pair expression are fed into an integrated neural network framework consisting of a graph convolutional neural network and a fully-connected neural network to determine whether the genes interact. Performance evaluation in the tasks of gene interaction and causality prediction against the existing GRN reconstruction algorithms demonstrates the usability and competitiveness of SFINN across different kinds of data. SFINN can be applied to infer GRNs from conventional single-cell sequencing data and spatial transcriptomic data. SFINN can be accessed at GitHub: https://github.com/JGuan-lab/SFINN.

Single-cell transcriptome dissecting the microenvironment remodeled by PD1 blockade combined with photodynamic therapy in a mouse model of oral carcinogenesis.

Oral squamous cell carcinoma (OSCC) stands as a predominant and perilous malignant neoplasm globally, with the majority of cases originating from oral potential malignant disorders (OPMDs). Despite this, effective strategies to impede the progression of OPMDs to OSCC remain elusive. In this study, we established mouse models of oral carcinogenesis via 4-nitroquinoline 1-oxide induction, mirroring the sequential transformation from normal oral mucosa to OPMDs, culminating in OSCC development. By intervening during the OPMDs stage, we observed that combining PD1 blockade with photodynamic therapy (PDT) significantly mitigated oral carcinogenesis progression. Single-cell transcriptomic sequencing unveiled microenvironmental dysregulation occurring predominantly from OPMDs to OSCC stages, fostering a tumor-promoting milieu characterized by increased Treg proportion, heightened S100A8 expression, and decreased Fib_Igfbp5 (a specific fibroblast subtype) proportion, among others. Notably, intervening with PD1 blockade and PDT during the OPMDs stage hindered the formation of the tumor-promoting microenvironment, resulting in decreased Treg proportion, reduced S100A8 expression, and increased Fib_Igfbp5 proportion. Moreover, combination therapy elicited a more robust treatment-associated immune response compared with monotherapy. In essence, our findings present a novel strategy for curtailing the progression of oral carcinogenesis.

Single-cell transcriptome sequencing reveals SPP1-CD44-mediated macrophage-tumor cell interactions drive chemoresistance in TNBC.

Triple-negative breast cancer (TNBC) is often considered one of the most aggressive subtypes of breast cancer, characterized by a high recurrence rate and low overall survival (OS). It is notorious for posing challenges related to drug resistance. While there has been progress in TNBC research, the mechanisms underlying chemotherapy resistance in TNBC remain largely elusive. We collect single-cell RNA sequencing (scRNA-seq) data from five TNBC patients susceptible to chemotherapy and five resistant cases. Comprehensive analyses involving copy number variation (CNV), pseudotime trajectory, cell-cell interactions, pseudospace analysis, as well as transcription factor and functional enrichment are conducted specifically on macrophages and malignant cells. Furthermore, we performed validation experiments on clinical samples using multiplex immunofluorescence. We identified a subset of SPP1+ macrophages that secrete SPP1 signals interacting with CD44 on malignant cell surfaces, potentially activating the PDE3B pathway within malignant cells via the integrin pathway, leading to chemotherapy resistance. The abnormally enhanced SPP1 signal between macrophages and malignant cells may serve as a factor promoting chemotherapy resistance in TNBC patients. Therefore, SPP1+ macrophages could potentially serve as a therapeutic target to reduce chemotherapy resistance.

Integrative Pan-Cancer Analysis Reveals the Oncogenic Role of MND1 and Validation of MND1's Role in Breast Cancer.

Meiotic nuclear division 1 (MND1) is a meiosis-specific protein that promotes lung adenocarcinoma progression. However, its expression and biological function across cancers remain largely unexplored. The expression, prognostic significance, mutation status, and methylation profile of MND1 in various cancers were comprehensively analyzed using the TIMER, GTEX, Kaplan-Meier plotter, cBioPortal, and GSCA databases. Additionally, we constructed a PPI network, enrichment analysis and single-cell transcriptomic sequencing to elucidate the underlying mechanism of MND1. Furthermore, we investigated the association between MND1 expression and drug sensitivity using CellMiner. Moreover, we also explored the correlation between MND1 expression and immune infiltration. Finally, we validated the functional role of MND1 in breast cancer through IHC staining, CCK8, EdU, colony formation, and flow cytometry assays. MND1 has been reported to be highly expressed in Pan-cancer, High MND1 expression was significantly associated with poor prognosis in cancers. Additionally, MND1 mutation frequency is high in most cancers, and its expression correlates with methylation. Furthermore, MND1 expression significantly correlates with immune checkpoint blockade (ICB) markers, including PD-L1, PD-1, and CTLA-4. The PPI network reveals interactions between MND1 and PSMC3IP, BRCA1, and BRCA2. Enrichment analysis and single-cell sequencing indicate that MND1 positively correlates with cell cycle. ROC curve reveals favorable diagnostic efficacy of MND1 in breast cancer. In vitro, MND1 overexpression promotes breast cancer cell proliferation and increases the expression of key cell cycle regulators (CDK4, CDK6, and cyclin D3), accelerating the G1/S phase transition and leading to abnormal breast cancer cell proliferation. The immunohistochemical analysis revealed a robust expression of MND1 in breast cancer tissues, exhibiting a significant positive correlation with PD-L1 and FOXP3. MND1 is an oncogene and may serve as a biomarker for cancer prognosis and immunotherapy. Targeting MND1 may be a potential tumor treatment strategy.

RNA-binding protein AZGP1 inhibits epithelial cell proliferation by regulating the genes of alternative splicing in COPD

BackgroundChronic Obstructive Pulmonary Disease (COPD) is characterized by high morbidity, disability, and mortality rates worldwide. RNA-binding proteins (RBPs) might regulate genes involved in oxidative stress and inflammation in COPD patients. Single-cell transcriptome sequencing (scRNA-seq) offers an accurate tool for identifying intercellular heterogeneity and the diversity of immune cells. However, the role of RBPs in the regulation of various cells, especially AT2 cells, remains elusive. Materials and methodsA scRNA-seq dataset (GSE173896) and a bulk RNA-seq dataset acquired from airway tissues (GSE124180) were employed for data mining. Next, RNA-seq analysis was performed in both COPD and control patients. Differentially expressed genes (DEGs) were identified using criteria of fold change (FC ≥ 1.5 or ≤ 1.5) and P value ≤ 0.05. Lastly, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and alternative splicing identification analyses were carried out. ResultsRBP genes exhibited specific expression patterns across different cell groups and participated in cell proliferation and mitochondrial dysfunction in AT2 cells. As an RBP, AZGP1 expression was upregulated in both the scRNA-seq and RNA-seq datasets. It might potentially be a candidate immune biomarker that regulates COPD progression by modulating AT2 cell proliferation and adhesion by regulating the expression of SAMD5, DNER, DPYSL3, GBP5, GBP3, and KCNJ2. Moreover, AZGP1 regulated alternative splicing events in COPD, particularly DDAH1 and SFRP1, holding significant implications in COPD. ConclusionRBP gene AZGP1 inhibits epithelial cell proliferation by regulating genes participating in alternative splicing in COPD.