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The effective treatment of advanced cervical cancer remains challenging. Herein, single‐nucleus RNA sequencing (snRNA‐seq) and SpaTial enhanced resolution omics‐sequencing (Stereo‐seq) are used to investigate the immunological microenvironment of cervical squamous cell carcinoma (CSCC). The expression levels of most immune suppressive genes in the tumor and inflammation areas of CSCC are not significantly higher than those in the non‐cancer samples, except for LGALS9 and IDO1. Stronger signals of CD56+ NK cells and immature dendritic cells are found in the hypermetabolic tumor areas, whereas more eosinophils, immature B cells, and Treg cells are found in the hypometabolic tumor areas. Moreover, a cluster of pro‐tumorigenic cancer‐associated myofibroblasts (myCAFs) are identified. The myCAFs may support the growth and metastasis of tumors by inhibiting lymphocyte infiltration and remodeling of the tumor extracellular matrix. Furthermore, these myCAFs are associated with poorer survival probability in patients with CSCC, predict resistance to immunotherapy, and might be present in a small fraction (< 30%) of patients with advanced cancer. Immunohistochemistry and multiplex immunofluorescence staining are conducted to validate the spatial distribution and potential function of myCAFs. Collectively, these findings enhance the understanding of the immunological microenvironment of CSCC and shed light on the treatment of advanced CSCC. The high heterogeneity in viral gene expression, immune response, and metabolism in cervical squamous cell carcinoma indicates that therapies targeting multiple biological processes may yield better curative effects. The discovery of pro‐tumorigenic cancer‐associated myofibroblasts (myCAFs) in clinical samples suggests that interventions on myCAFs might complement the current treatments for invasive cervical cancer.

INTRODUCTION Alzheimer's disease (AD), dementia with Lewy bodies (DLB), and Parkinson's disease (PD) represent a spectrum of neurodegenerative diseases (NDDs). Here, we performed the first direct comparison of their transcriptomic landscapes. METHODS We profiled whole transcriptomes of NDD cortical tissue by single‐nucleus RNA sequencing, using computational analyses to identify common and distinct differentially expressed genes (DEGs), pathways, vulnerable and disease‐driver cell subtypes, and altered cell‐to‐cell interactions. RESULTS The same inhibitory neuron subtype was depleted in both AD and DLB. Potentially disease‐driving neuronal cell subtypes were identified in both PD and DLB. Cell–cell communication was predicted to be increased in AD but decreased in DLB and PD. DEGs were most commonly shared across NDDs within inhibitory neuron subtypes. Overall, AD and PD showed greatest transcriptomic divergence, while DLB exhibited an intermediate signature. DISCUSSION These results may help explain the clinicopathological spectrum of these NDDs and provide unique insights into shared and distinct molecular mechanisms underlying pathogenesis. Highlights The same vulnerable inhibitory neuron subtype population was depleted in both Alzheimer's disease (AD) and dementia with Lewy bodies (DLB). Potentially disease‐driving neuronal cell subtypes were discovered in both Parkinson's disease (PD) and DLB. Cell–cell communication was predicted to be increased in AD but decreased in DLB and PD. Differentially expressed genes were most commonly shared across neurodegenerative diseases in inhibitory neuron types. AD and PD had the greatest transcriptomic divergence, with DLB showing an intermediate signature.

INTRODUCTION Using a single‐nucleus transcriptome derived from the dorsolateral prefrontal cortex of 424 Religious Orders Study and the Rush Memory and Aging Project (ROS/MAP) participants, we investigated the cell type–specific effect of ten vascular endothelial growth factor (VEGF) genes on Alzheimer's disease (AD) endophenotypes. METHODS Negative binomial mixed models were used for differential gene expression and association analysis with AD endophenotypes. VEGF‐associated intercellular communication was also profiled. RESULTS Higher microglia FLT1, endothelial FLT4, and oligodendrocyte VEGFB are associated with greater amyloid beta (Aβ) load, whereas higher VEGFB expression in inhibitory neurons is associated with lower Aβ load. Higher astrocyte NRP1 is associated with lower tau density. Higher microglia and endothelial FLT1 are associated with worse cognition performance. Endothelial and microglial FLT1 expression was upregulated in clinical AD patients compared to cognitively normal controls. Finally, AD cells showed a significant reduction in VEGF signaling compared to controls. DISCUSSION Our results highlight key changes in VEGF receptor expression in endothelial and microglial cells during AD, and the potential protective role of VEGFB in neurons. Highlights The prefrontal cortical expression of FLT1 and FLT4 was associated with worse cross‐sectional global cognitive function, longitudinal cognitive trajectories, and more Alzheimer's disease (AD) neuropathology. The associations between FLT1 or FLT4 and AD endophenotypes appear to be driven by endothelial and microglial cells. VEGFB expression seems to have opposing effects on the Aβ burden in AD depending on cell types, highlighting its potential protective role in neurons.

Anti‐N‐methyl‐D‐aspartate receptor (NMDAR) encephalitis is a neuropsychiatric disease with variable clinical manifestations caused by NMDAR autoantibody. The underlying molecular underpinnings of this disease are rarely characterized on a genomic scale. Anti‐NMDAR encephalitis mainly affects the hippocampus, however, its effect on gene expression in hippocampal neurons is unclear at present. Here, we construct the active and passive immunization mouse models of anti‐NMDAR encephalitis, and use single‐nucleus RNA sequencing to investigate the diverse expression profile of neuronal populations isolated from different hippocampal regions. Dramatic changes in cell proportions and differentially expressed genes were observed in excitatory neurons of the dentate gyrus (DG) subregion. In addition, we found that ATP metabolism and biosynthetic regulators related genes in excitatory neurons of DG subregion were significantly affected. Kcnq1ot1 in inhibitory neurons and Meg3 in interneurons also changed. Notably, the latter two molecules exhibited opposite changes in different models. Therefore, the above genes were used as potential targets for further research on the pathological process of anti‐NMDAR encephalitis. These data involve various hippocampal neurons, which delineate a framework for understanding the hippocampal neuronal circuit and the potential molecular mechanisms of anti‐NMDAR encephalitis. The authors investigate the diverse neuronal expression profile in different hippocampal regions of the anti‐NMDAR encephalitis mouse model, and dramatic changes in cell proportions and differentially expressed genes were observed, such as ATP metabolism and biosynthetic regulators related genes.

Background Tumour‐induced skeletal muscle wasting in the context of cancer cachexia is a condition with profound implications for patient survival. The loss of muscle mass is a significant clinical obstacle and is linked to reduced tolerance to chemotherapy and increased frailty. Understanding the molecular mechanisms driving muscle atrophy is crucial for the design of new therapeutics. Methods Lewis lung carcinoma tumours were utilized to induce cachexia and muscle atrophy in mice. Single‐nucleus libraries of the tibialis anterior (TA) muscle from tumour‐bearing mice and their non‐tumour‐bearing controls were constructed using 10X Genomics applications following the manufacturer's guidelines. RNA sequencing results were analysed with Cell Ranger software and the Seurat R package. Oxygen consumption of mitochondria isolated from TA muscle was measured using an Oroboros O2k‐FluoRespirometer. Mouse primary myotubes were treated with a recombinant ectodysplasin A2 (EDA‐A2) protein to activate EDA‐A2 receptor (EDA2R) signalling and study changes in gene expression and oxygen consumption. Results Tumour‐bearing mice were sacrificed while exhibiting moderate cachexia. Average TA muscle weight was reduced by 11% (P = 0.0207) in these mice. A total of 12 335 nuclei, comprising 6422 nuclei from the control group and 5892 nuclei from atrophying muscles, were studied. The analysis of single‐nucleus transcriptomes identified distinct myonuclear gene signatures and a shift towards type IIb myonuclei. Muscle atrophy‐related genes, including Atrogin1, MuRF1 and Eda2r, were upregulated in these myonuclei, emphasizing their crucial roles in muscle wasting. Gene set enrichment analysis demonstrated that EDA2R activation and tumour inoculation led to similar expression patterns in muscle cells, including the stimulation of nuclear factor‐kappa B, Janus kinase–signal transducer and activator of transcription and transforming growth factor‐beta pathways and the suppression of myogenesis and oxidative phosphorylation. Muscle oxidative metabolism was suppressed by both tumours and EDA2R activation. Conclusions This study identified tumour‐induced transcriptional changes in muscle tissue at single‐nucleus resolution and highlighted the negative impact of tumours on oxidative metabolism. These findings contribute to a deeper understanding of the molecular mechanisms underlying muscle wasting.

Partial reprogramming (pulsed expression of reprogramming transcription factors) ameliorates multiple tissue functions in aged mice; however, its impact on peripheral nerve regeneration remains largely unexplored. In this study, the temporal dynamics of Schwann cells following sciatic nerve injury in young and aged rats are systematically examined using single‐cell transcriptomics to identify a Runx2+ cell population highly enriched with stress granules as transitional homeostatic cells during Schwann cell differentiation. It is found that pathological accumulation of this cluster during axonal regeneration constitutes a critical contributing factor to impaired neural repair in aging. Intriguingly, partial reprogramming enhances axonal regeneration and attenuates senescence‐associated phenotypes and functional deficits in aged Schwann cells, demonstrating that partial reprogramming promotes peripheral nerve regeneration through Schwann cell rejuvenation. Mechanistically, aged Schwann cells exhibit a stress granule homeostatic imbalance, characterized by compromised formation and impaired degradation, which is effectively reset by partial reprogramming. Importantly, this homeostatic resetting ameliorated the pathological aggregation of Runx2+ Schwann cells during nerve repair in aged rats. The findings reveal that dysregulated stress granule homeostasis drives the pathological accumulation of Runx2+ Schwann cells, representing a key mechanism underlying age‐related axonal regeneration deficits in peripheral nerve repair. This study establishes that partial reprogramming can restore this critical cellular homeostasis and enhance peripheral nerve regeneration during aging. In this study, it is discovered that pathological accumulation of Runx2+ Schwann cell clusters​ during axonal regeneration is a key factor impairing nerve repair in aging. This pathology is associated with ​dysregulation of stress granule homeostasis. Partial reprogramming enhances axonal regeneration and restores stress homeostasis in aged Schwann cells, thus demonstrating that partial reprogramming promotes peripheral nerve regeneration through ​Schwann cell rejuvenation.

Tumor heterogeneity and its drivers impair tumor progression and cancer therapy. Single‐cell RNA sequencing is used to investigate the heterogeneity of tumor ecosystems. However, most methods of scRNA‐seq amplify the termini of polyadenylated transcripts, making it challenging to perform total RNA analysis and somatic mutation analysis.Therefore, a high‐throughput and high‐sensitivity method called snHH‐seq is developed, which combines random primers and a preindex strategy in the droplet microfluidic platform. This innovative method allows for the detection of total RNA in single nuclei from clinically frozen samples. A robust pipeline to facilitate the analysis of full‐length RNA‐seq data is also established. snHH‐seq is applied to more than 730 000 single nuclei from 32 patients with various tumor types. The pan‐cancer study enables it to comprehensively profile data on the tumor transcriptome, including expression levels, mutations, splicing patterns, clone dynamics, etc. New malignant cell subclusters and exploring their specific function across cancers are identified. Furthermore, the malignant status of epithelial cells is investigated among different cancer types with respect to mutation and splicing patterns. The ability to detect full‐length RNA at the single‐nucleus level provides a powerful tool for studying complex biological systems and has broad implications for understanding tumor pathology. In this study, snHH‐seq, a high‐throughput, and high‐sensitivity snRNA‐seq method, is applied to pan‐cancer samples from 32 patients, comprising > 700 000 nuclei from various cancer types. The comprehensive analysis of gene expression, somatic mutations, splicing patterns, and clonal behavior shows broad implications for understanding tumor pathology.

A notable number of locally advanced cervical carcinoma (LACC) patients experience local or distant disease relapse following radiotherapy. The contribution of tumor microenvironment (TME) to tumor recurrence at different sites remains unclear. Here, single‐nucleus RNA sequencing data from 28 pre‐ and on‐treatment LACC samples from patients with different disease relapse patterns is analyzed. The findings revealed opposing alterations in the expression levels of the cellular senescence pathway after radiotherapy in patients with local and distant relapses. In contrast, an increase in the expression of the epithelial‐mesenchymal transition module after radiotherapy in both relapse groups is observed. Cell–cell interactions, drug‐target expression analyses in malignant cells after radiation, and multiplex immunofluorescence of tumor tissue identified interleukin‐1 receptor type I (IL1R1) as a potential therapeutic target. It is demonstrated that combining the IL1R1 inhibitor anakinra with radiation can mitigate the effects of radiation on tumor cells. This study highlights the distinct roles of cellular senescence and EMT in tumor recurrence. The relapse rate for locally advanced cervical cancer (LACC) patients after radiotherapy is 20%–40%. This study shows that the cellular senescence pathway in malignant cells varies among relapse patterns, while EMT module expression is consistently elevated in all relapse patterns. Additionally, the cellular senescence status of fibroblasts ubiquitously increased following radiotherapy.

Penile squamous cell carcinoma (PSCC) is a highly aggressive malignancy without effective treatment due to limited knowledge of its development and tumor microenvironment (TME). In this study, single‐nucleus RNA sequencing (snRNA‐seq) and high‐resolution spatial transcriptomics are employed to comprehensively investigate the development trajectories and the TME. The results revealed that PSCC cells mimicked the differentiation and tissue organization of normal penile epithelium, independent of the human papillomavirus (HPV) infection status. Notably, a spatial subtype, Tum_1, appeared at early stage of tumor differentiation and in tumor–normal boundary regions. This subtype exhibited enhanced basal‐like and stemness features and showed high LAMC2 expression, which activated laminin‐integrin signaling via ITGA6/ITGB4, promoting tumor invasiveness. Furthermore, the results indicated that HPV‐positive basal stem‐like neoplasms dampened the immune function of T cells and macrophages, promoting an immunosuppressive environment that facilitates tumor progression. Supporting this, the patients with head and neck squamous cell carcinoma and lung squamous cell carcinoma who have high expression of HPV‐positive Tum_1 signatures derived greater benefit from PD‐1 blockade therapy. In summary, the findings provide a comprehensive spatial landscape of the PSCC TME and suggest potential treatment approaches targeting laminin‐integrin interaction and immunotherapy, especially in HPV‐positive patients. This study uses single‐nucleus RNA sequencing and spatial transcriptomics to investigate penile squamous cell carcinoma (PSCC). It reveals that PSCC tumor cells mimic normal penile epithelium differentiation, independent of HPV status. The Tum_1 subtype shows basal stem‐like characteristics and promotes invasiveness. HPV‐positive basal stem‐like tumors exhibit an immunosuppressive microenvironment, suggesting potential for personalized treatments based on HPV and tumor immune microenvironment.

Systemic metabolic dysregulation in sepsis critically impacts patient survival. To better understand its onset, untargeted serum metabolomics and lipidomics are analyzed from 152 presymptomatic patients undergoing major elective surgery, and identified key metabolites, including serine and aminoadipic acid, that differentiate postoperative uncomplicated infection from sepsis. Using single‐nucleus RNA sequencing data from an in vivo mouse model of sepsis, tissue‐independent down‐regulation and tissue‐specific differences of serine and energy‐related genes including key module roles for the mitochondria‐linked genes, Cox4i1, Cox8a, and Ndufa4 are identified. Finally, serine‐dependent metabolic shifts, especially in the liver, are revealed by using 12C/13C murine data with labeled serine, and link altered activity of the serine hydroxymethyltransferase (SHMT) cycle with perturbed purine metabolism during sepsis. This study demonstrates the close interrelationship between early metabolite changes and mitochondrial dysfunction in sepsis, improves the understanding of the underlying pathophysiology, and highlights metabolic targets to prospectively treat presymptomatic, but at‐risk, patients. This study shows that patients at risk of sepsis have a distinct metabolite and lipid signature, including serine and aminoadipic acid, in their serum before clinical diagnosis. A mouse model of sepsis with a compatible serum signature reveals underlying metabolic changes, including mitochondrial adaptation, altered serine‐dependent purine metabolism, and key gene factors as potential drug targets.