Recent advancements in cancer immunotherapy have improved patient outcomes, yet responses to immunotherapy remain moderate. Immunosenescence has been shown to contribute to the development and progression of various diseases; however, its specific role in solid tumors has not been fully delineated. Here we conducted a phase 2 clinical trial involving 51 patients with cancer undergoing neoadjuvant chemoimmunotherapy and applied single-cell RNA as well as TCR and BCR sequencing on tumor and blood samples to elucidate the immune cell perturbations. Our findings associate poor response with reduced levels of CCR7+ CD4+ naive T cells and CD27+ memory B cells, as well as higher expression of immunosenescence-related genes in T and B cell subsets. Using naturally aged mice and Ercc1-deficient mice (premature aging), we found that senolytics enhance the therapeutic efficacy of immunotherapy in multiple solid tumors by mitigating immunosenescence. Notably, we launched a phase 2 clinical trial (COIS-01) investigating the combination of senolytics with anti-PD-1 therapy. The results showed that the combination therapy achieved a 33.3% (95% confidence interval 16.6-54.7%) major pathological response rate with a low incidence of grade 3-4 adverse events (4.2%). These findings underscore the pivotal role of immunosenescence characteristics in influencing the effectiveness of immunotherapy and suggest a promising therapeutic efficacy along with a favorable safety for the combination of senolytics with anti-PD-1 therapy. ClinicalTrials.gov Identifier: OOC-001( NCT04718415 ) and COIS-01( NCT05724329 ).

Trichosanthin (TCS), a type I ribosome-inactivating protein, exerts its cytotoxic effects by inhibiting protein synthesis through depurination of 28S rRNA, resulting in apoptosis and cancer cell death. However, insufficient tumor specificity and limited cell-penetrating capabilities have restricted its applications. Herein, we engineered a recombinant TCS by inserting low-molecular-weight protamine (LMWP) and matrix metalloproteinase-selective peptide (MSP), thereby constructing a recombinant fusion protein (rTCS-LMWP-MSP, namely, rTLM) with enhanced tumor-targeting and cell-penetrating capabilities. Subsequently, manganese-doped calcium phosphate (MnCaP) nanoparticles were fabricated by bovine serum albumin (BSA)-templated mineralization to serve as a pH-responsive delivery system, which not only improves the biocompatibility of rTLM but also enables payload release activated by the acidic tumor microenvironment. Upon accumulation of BSA-MnCaP-rTLM in tumor tissues, the extracellular matrix metalloproteinase 2 (MMP2) could recognize and cleave MSP. This process not only enables tumor-targeting capability but also exposes the inserted LMWP to enhance cell-penetrating capability. When internalized by tumor cells, BSA-MnCaP could be degraded, leading to the release of rTCS-LMWP, which induces cell apoptosis. Simultaneously, the released Mn2+ ions catalyze the conversion of endogenous H2O2 into harmful hydroxyl radicals via a Fenton-like reaction, thus promoting the oxidative stress in tumor cells. Both in vitro and in vivo experiments confirmed the synergistic antitumor effects of BSA-MnCaP-rTLM. Our findings indicate that BSA-MnCaP-rTLM holds significant potential for effective cancer treatment.

Hexokinase 2 (HK2) is a critical rate-limiting enzyme in glycolysis, significantly linked to cancer metabolism and drug resistance. We hypothesized that targeting HK2 with Benserazide (BenZ), a known HK2 inhibitor, could potentiate the cytotoxicity of cisplatin (DDP) in non-small-cell lung cancer (NSCLC) cells. Our findings demonstrated that combination of DDP and BenZ significantly enhanced anticancer effects in NSCLC cells, either with or without acquired DDP resistance. In particular, BenZ effectively inhibited ATP and lactate productions. ATP supplementation mitigated the cytotoxicity of the combination treatment, which was restored when both the cisplatin efflux transporters, ATP7A and ATP7B, were simultaneously knocked down. Moreover, BenZ suppressed NF-κB activation and downregulated inflammatory gene expressions, including IκB and RELA phosphorylation, TNF-α, BCL-2, and IL-1β. These effects were attributed to inhibition of HK2 by BenZ leading to the suppression of IκBα phosphorylation. It has been shown that combined use of DDP and BenZ significantly inhibited tumor growth in NSCLC cell xenograft mouse models, while BenZ alleviated the DDP-induced nephrotoxicity. Our findings demonstrate that BenZ inhibits the kinase activity of HK2, reduces ATP level and cisplatin efflux, as well as alleviates NF-κB activation. These multifaceted actions enhance the anticancer efficacy of cisplatin in NSCLC cell models, indicating its potential as an effective adjunct therapy.

Over the past four decades, the relationship between viruses and gastrointestinal (GI) mucosal immunity has gained increasing attention due to frequent viral epidemics. This study explores current research trends and future directions in this field through bibliometric analysis. Literature from January 1, 1985, to December 31, 2024, was retrieved from the Web of Science Core Collection (WoSCC) on January 6, 2025. Analyses were conducted using VOSviewer, Citespace, and the Bibliometrix R package. A total of 4,842 publications were identified, with an annual growth rate of 8.65

Oral squamous cell carcinoma (OSCC) is the most common type of oral and maxillofacial malignancy. To ensure its accurate diagnosis and treatment, biomarker research is essential. Here, we aimed to analyze trends and research hot spots in OSCC biomarker studies over the past 30 years. Relevant articles were collected from the Web of Science Core Collection (WoSCC) covering the period from January 1, 1995, to December 31, 2024. Collaborative analyses were then conducted using VOSviewer and Scimago Graphica. CiteSpace software was used to perform dual-map overlay analyses, detect citation bursts, and generate timeline keyword clustering maps. A total of 6,764 English-language articles were retrieved and analyzed using bibliometric methods. The research was divided into three stages based on the rate of literature growth. China ranked first in total publications, while the United States (USA) led the field in collaboration and total citations. Shanghai Jiao Tong University in China produced the highest number of research publications, and Oral Oncol was the journal with the most articles. Keyword co-occurrence analysis revealed a multifaceted research landscape comprising five primary thematic clusters. In the past 5 years, keywords such as “tumor microenvironment,” “recurrent,” “liquid biopsy,” and “migration” have dominated the field, with “neutrophil-to-lymphocyte ratio” and “saliva” emerging as prominent research hot spots. Among specific molecules, P53, P16, and Cyclin D1 were the most extensively studied biomarkers. Additionally, molecules such as miR-21, miR-31, and COX-2 have attracted increasing attention in recent years. These findings may provide valuable insights for future research in this field.

Pharyngeal cartilage, derived from neural crest cells (NCCs), undergoes complex morphogenesis driven by signals from the pharyngeal endoderm. However, the molecular mechanisms governing NCC proliferation and differentiation in response to endoderm-derived signals remain poorly understood. Here, we investigate the role of klf5a, a zinc-finger transcription factor expressed in pharyngeal endodermal pouches, in zebrafish pharyngeal cartilage development. Knockdown of klf5a using morpholinos minimally affected cranial NCC specification and migration but significantly impaired their proliferation and differentiation in the pharyngeal region. Notably, klf5a deficiency reduced expression of fgfbp2b, a modulator of FGF signaling, in the pharyngeal endoderm. Co-injection of klf5a mRNA rescued the cartilage defects, but injection of fgfbp2b mRNA alone failed to restore normal cartilage morphogenesis, suggesting that fgfbp2b is not the sole mediator of klf5a's effects. These findings indicate that klf5a regulates endodermal signaling to direct NCC-derived pharyngeal cartilage formation, likely through multiple downstream targets including fgfbp2b. This study provides insights into the complex molecular network underlying craniofacial development and highlights potential therapeutic targets for craniofacial disorders.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) ravaged the world, and Coronavirus Disease 2019 (COVID-19) exhibited highly prevalent oral symptoms that had significantly impacted the lives of affected patients. However, the involvement of four human coronavirus (HCoVs), namely SARS-CoV-2, SARS-CoV, MERS-CoV, and HCoV-229E, in oral cavity infections remained poorly understood. We integrated single-cell RNA sequencing (scRNA-seq) data of seven human oral tissues through consistent normalization procedure, including minor salivary gland (MSG), parotid gland (PG), tongue, gingiva, buccal, periodontium and pulp. The Seurat, scDblFinder, Harmony, SingleR, Ucell and scCancer packages were comprehensively used for analysis. We identified specific cell clusters and generated expression profiles of SARS-CoV-2 and coronavirus-associated receptors and factors (SCARFs) in seven oral regions, providing direction for predicting the tropism of four HCoVs for oral tissues, as well as for dental clinical treatment. Based on our analysis, it appears that various SCARFs, including ACE2, ASGR1, KREMEN1, DPP4, ANPEP, CD209, CLEC4G/M, TMPRSS family proteins (including TMPRSS2, TMPRSS4, and TMPRSS11A), and FURIN, are expressed at low levels in the oral cavity. Conversely, BSG, CTSB, and CTSL exhibit enrichment in oral tissues. Our study also demonstrates widespread expression of restriction factors, particularly IFITM1-3 and LY6E, in oral cells. Additionally, some replication, assembly, and trafficking factors appear to exhibit broad oral tissues expression patterns. Overall, the oral cavity could potentially serve as a high-risk site for SARS-CoV-2 infection, while displaying a comparatively lower degree of susceptibility towards other HCoVs (including SARS-CoV, MERS-CoV and HCoV-229E). Specifically, MSG, tongue, and gingiva represent potential sites of vulnerability for four HCoVs infection, with the MSG exhibiting a particularly high susceptibility. However, the expression patterns of SCARFs in other oral sites demonstrate relatively intricate and may only be specifically associated with SARS-CoV-2 infection. Our study sheds light on the mechanisms of HCoVs infection in the oral cavity as well as gains insight into the characteristics and distribution of possible HCoVs target cells in oral tissues, providing potential therapeutic targets for HCoVs infection in the oral cavity.

Psoriasis (Ps), a chronic inflammatory disease affecting approximately 2% of the global population, has been associated with an increased risk of liver cancer in observational studies. However, their causal relationships as well as underlying shared molecular mechanisms between Ps and liver cancer remain unclear. Using bidirectional Mendelian randomization analysis, we revealed that a genetic predisposition to liver cancer increased the risk of Ps in European and East Asian populations but not the other way around. Moreover, we analyzed three transcriptomic datasets of patients with Ps and liver cancer from open-source databases. Differentially expressed genes (DEGs) and disease-specific gene co-expression module analyses revealed that cell-cycle dysregulation was the shared mechanism of Ps and liver cancer. Moreover, we identified a rank-conservative gene signature shared between these two diseases, which demonstrated significance in diagnostic and prognostic predictions. These findings provided valuable insights into the interconnections between Ps and liver cancer, which may be helpful to guide therapeutic management.

Recent advancements in cancer immunotherapy have improved patient outcomes, yet responses to immunotherapy remain moderate. We conducted a Phase II clinical trial ([NCT04718415][1]) involving 51 cancer patients undergoing neoadjuvant chemoimmunotherapy and applied single-cell RNA and T/BCR sequencing on tumor and blood samples to elucidate the immune cell perturbations. Our findings associate poor response with reduced levels of CCR7+CD4 Naive T cells and CD27+ Memory B cells, as well as higher expression of immunosenescence-related genes in T and B cell subsets. Using naturally aged and Ercc1+/- transgenic aging mouse models, we found that senolytics enhance the therapeutic efficacy of immunotherapy in multiple solid tumors by mitigating tumor immunosenescence. Notably, we launched a Phase II clinical trial, COIS-01 ([NCT05724329][2]), which pioneers the combination of senolytics with anti-PD-1 therapy. The clinical results demonstrate that this therapeutic strategy is associated with a favorable safety profile and therapeutic efficacy, significantly mitigating adverse effects and alleviating immunosenescence. These findings underscore the pivotal role of immunosenescence characteristics in influencing the effectiveness of immunotherapy and suggest a promising therapeutic efficacy along with a beneficial safety assessment for the combination of senolytics with anti-PD-1 therapy. ### Competing Interest Statement The authors have declared no competing interest. ### Clinical Trial NCT05724329 ### Funding Statement This study was funded by the Joint Funds of the National Natural Science Foundation of China (U21A20381), the General Funds of the National Natural Science Foundation of China (82373452), the Guangdong Natural Science Funds for Distinguished Young Scholar (2022B1515020061), the Guangdong Basic and Applied Basic Research Foundation (2021A1515220138), the Guangzhou Basic Research Program Jointly Funded by Municipal Schools (Institutes) (202201020367), the Fundamental Research Funds for the Central Universities, Sun Yat-sen University (16ykpy10), the Fundamental Research Funds for the Central Universities, Sun Yat-sen University (19ykzd20), the General Funds of the National Natural Science Foundation of China (32071451), the Guangdong Provincial Pearl River Talents Program (2021QN02Y747), the Shenzhen Science and Technology Program (RCYX20210706092100003), and by the Shenzhen Medical Research Funds grant (A2303005). ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: Ethics Committee of Sun Yat-sen Memorial Hospital, Sun Yat-sen University gave ethical approval for this work. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes All data produced in the present study are available upon reasonable request to the authors. [1]: /lookup/external-ref?link_type=CLINTRIALGOV&access_num=NCT04718415&atom=%2Fmedrxiv%2Fearly%2F2024%2F10%2F15%2F2024.10.14.24315428.atom [2]: /lookup/external-ref?link_type=CLINTRIALGOV&access_num=NCT05724329&atom=%2Fmedrxiv%2Fearly%2F2024%2F10%2F15%2F2024.10.14.24315428.atom

Background and purpose: Trichosanthin (TCS) is a plant-based ribosome-inactivating protein exhibiting a range of pharmacological properties, including abortifacient and anticancer. However, the routine clinical use in cancer treatment was hampered by its antigenicity. Hexokinase 2 (HK2) is a pivotal regulator of glycolysis, where aberrant expression is observed in many cancers. This study investigates the anticancer effects and mechanisms of TCS in combination with benserazide (Benz), a HK2 inhibitor, in Hela and SCC25 cancer models. Experimental approach: MTT, colony-formation and cell cycle assays were performed to assess the cytotoxic effects of TCS and Benz in HeLa and SCC25 cells. Seahorse assay, western blotting, flow cytometry analysis and RNA sequencing were employed to investigate the pharmacological effects of the combo treatment. SCC25 cell xenograft mouse model was established for in vivo efficacy study. Key results: Combined use of TCS and Benz exhibited synergistic anticancer effects in both Hela and SCC25 cell models. The observed synergistic effects were attributed to the modulation of glycolysis by targeting HK2, leading to reduced lactate production and increased ROS accumulation which further inhibited colony formation and cell cycle progression, as well as triggered apoptosis. Moreover, this combination effectively inhibited NFκB/ERK signalling pathways, which were found to be significantly activated upon single use of TCS. It was found that the combination significantly suppressed the tumour growth in SCC25 cell xenograft mouse model. Conclusion: Our findings suggested that targeting HK2 and modulating glycolysis may offer a promising avenue for improving the therapeutic outcomes of TCS-based anticancer treatments.

Individuals diagnosed with head and neck squamous cell carcinoma (HNSCC) experience a significant occurrence rate and are susceptible to premature spreading, resulting in a bleak outlook. Therapeutic approaches, such as chemotherapy, targeted therapy, and immunotherapy, may exhibit primary and acquired resistance during the advanced phases of HNSCC. There is currently no viable solution to tackle this issue. PANoptosis—a type of non-apoptotic cell death—is a recently identified mechanism of cellular demise that entails communication and synchronization among thermal apoptosis, apoptosis, and necrosis mechanisms. However, the extent to which PANoptosis-associated genes (PRG) contribute to the forecast and immune reaction of HNSCC remains mostly undisclosed. The present study aimed to thoroughly analyze the potential importance of PRG in HNSCC and report our discoveries. We systematically analyzed 19 PRG from previous studies and clinical data from HNSCC patients to establish a PAN-related signature and assess its prognostic, predictive potential. Afterward, the patient information was separated into two gene patterns that corresponded to each other, and the analysis focused on the connection between patient prognosis, immune status, and cancer immunotherapy. The PAN score was found to correlate with survival rates, immune systems, and cancer-related pathways. We then validated the malignant role of CD27 among them in HNSCC. In summary, we demonstrated the effectiveness of PAN.Score-based molecular clustering and prognostic features in predicting the outcome of HNSCC. The discovery we made could enhance our comprehension of the significance of PAN.Score in HNSCC and facilitate the development of more effective treatment approaches.

Leucine-rich repeat containing protein 59 (LRRC59), a ribosome binding protein located on the endoplasmic reticulum and the nuclear envelope. Although it has been found in blood plasma and linked to the development of a few cancers, the function of LRRC59 is still largely unknown and there is no systematic investigation on its role in various human cancers. We performed a multi-omics data analysis to investigate the expression of LRRC59 in human tumors and its correlation with clinical prognosis, gene set enrichment, mutation status, and immune infiltration in cancers using the TCGA, GTEx, GEPIA2, HPA, UALCAN, Timer2, GTBAdb, cBioPortal database and R packages. In the majority of TCGA tumors, LRRC59 was expressed significantly differently (up-regulated in 25 and down-regulated in 4 cancer types). High LRRC59 expression has been linked to worse prognosis in several malignancies. In numerous carcinomas, the expression of LRRC59 was associated clinicopathological stages. The LRRC59 regulation network was mainly involved in the pathways related to endoplasmic reticulum homeostatic and cell proliferation. In addition, the expression of LRRC59 is also strongly associated with the immune cell infiltration. LRRC59 could also predicts the response to immunotherapy. LRRC59 is a potential valuable biomarker not only for diagnostic and prognostic, but also for immunotherapy in most cancers.

The vulnerability of the oral cavity to SARS-CoV-2 infection is well-known, and cancer patients are at a higher risk of COVID-19, emphasizing the need to prioritize this patient population. Head and neck squamous cell carcinoma (HNSCC) is one of the most common malignant cancers associated with early metastasis and poor prognosis. It has been established that cancerous tissues express Cathepsin L (CTSL), a proteinase that regulates cancer progression and SARS-CoV-2 entry. Therefore, it is essential to evaluate the correlation between disease outcomes and CTSL expression in cancer tissues and predict the susceptibility of cancer patients to SARS-CoV-2. In this study, we used transcriptomic and genomic data to profile CTSL expression in HNSCC and developed a CTSL signature that could reflect the response of HNSCC patients to chemotherapy and immunotherapy. Additionally, we investigated the relationship between CTSL expression and immune cell infiltration and established CTSL as a potential carcinogenic factor for HNSCC patients. These findings could aid in understanding the mechanisms underlying the increased susceptibility of HNSCC patients to SARS-CoV-2 and contribute to the development of therapy for both HNSCC and COVID-19.

Pyruvate dehydrogenase kinase 1 (PDK1) is an important metabolic enzyme which is often overexpressed in many types of cancers, including non-small-cell lung cancers (NSCLC). Targeting PDK1 appears to be an attractive anticancer strategy. Based on a previously reported moderate potent anticancer PDK1 inhibitor, 64, we developed three dichloroacetophenone biphenylsulfone ethers, 30, 31 and 32, which showed strong PDK1 in-hibitions of 74%, 83% and 72% at 10 mu M, respectively. Then we investigated the anticancer effects of 31 in two NSCLC cell lines, namely, NCI-H1299 and NCI-H1975. It was found that 31 exhibited sub-micromolar cancer cell IC50s, suppressed colony formation, induced mitochondrial membrane potential depolarization, triggered apoptosis, altered cellular glucose metabolism, with concomitant reductions in extracellular lactate levels and enhanced the generation of reactive oxygen species in NSCLC cells. Moreover, 31 significantly suppressed the tumor growth in an NCI-H1975 mouse xenograft model, outperforming the anticancer effects of 64. Taken together our results suggested that inhibition of PDK1 via dichloroacetophenone biphenylsulfone ethers may provide a novel direction leading to an alternative treatment option in NSCLC therapy.

Trichosanthin (TCS) is a type I ribosome-inactivating protein extracted from the tuberous root of the plant Trichosanthes. TCS shows promising potential in clinical drug abortion, anti-tumor and immunological regulation. However, the molecular mechanisms of its anti-tumor and immune regulation properties are still not well discovered. In the present study, we investigated the anti-tumor activity of TCS in hepatocellular carcinoma (HCC), both in vitro and in vivo. Both HCC cell lines and xenograft tumor tissues showed considerable growth inhibition after they were treated with TCS. TCS provoked caspase-mediated apoptosis in HCC cells and xenograft tumor tissues. The recruitment of CD8+ T cells to HCC tissues and the expression of chemokines, CCL2 and CCL22, were promoted upon TCS treatment. In addition, TCS induced an upregulation of Granzyme B (GrzB), TNF-α and IFN-γ in HCC tissues, which are the major cytotoxic mediators produced by T cells. Furthermore, TCS also resulted in an increase of mannose-6-phosphate receptor (M6PR), the major receptor of GrzB, in HCC tissues. In summary, these results suggest that TCS perhaps increases T-cell immunity via promoting the secretion of chemokines and accelerating the entry of GrzB to HCC cells, which highlights the potential role of TCS in anti-tumor immunotherapy.

This study aimed to investigate the effects of four alcoholic beverages on enamel erosion. Fifty enamel specimens were randomly allocated into the following five groups (n=10): group 1, water as negative control; group 2, red wine; group 3, white wine; group 4, distilled spirit; and group 5, beer. The specimens were immersed in the respective solution for a 16 h demineralization, followed by an 8 h remineralization in artificial saliva. Cyclic de- and re-mineralization were performed for 8 days. Surface roughness, microhardness and morphology of the enamel specimens were studied after the cycling. The results were analyzed by One-way ANOVA and Dunnett's post-hoc test (p<0.05). All investigated beverages showed an erosive effect on enamel. White wine had the highest erosive potential whereas distilled spirit had the least.

Coronavirus disease 2019 (COVID-19) was reported to be associated with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection, and patients present mostly with respiratory symptoms. There have been an increasing number of reports on oral manifestations, and some of these signs are informative in terms of identifying SARS-CoV-2 infection. The goal of present study was to review and synthesize the clinical characteristics and underlying mechanisms of COVID-19 oral manifestations, as well as to evaluate the factors influencing SARS-CoV-2 infectivity, in order to conduct further in-depth investigations and help clinicians diagnose COVID-19 patients exhibiting oral symptoms.

Background The centrosome is one of the most important non-membranous organelles regulating microtubule organization and progression of cell mitosis. The coiled-coil alpha-helical rod protein 1 (CCHCR1, also known as HCR) gene is considered to be a psoriasis susceptibility gene, and the protein is suggested to be localized to the P-bodies and centrosomes in mammalian cells. However, the exact cellular function of HCR and its potential regulatory role in the centrosomes remain unexplored. Results We found that HCR interacts directly with astrin, a key factor in centrosome maturation and mitosis. Immunoprecipitation assays showed that the coiled-coil region present in the C-terminus of HCR and astrin respectively mediated the interaction between them. Astrin not only recruits HCR to the centrosome, but also protects HCR from ubiquitin-proteasome-mediated degradation. In addition, depletion of either HCR or astrin significantly reduced centrosome localization of CEP72 and subsequent MCPH proteins, including CEP152, CDK5RAP2, and CEP63. The absence of HCR also caused centriole duplication defects and mitotic errors, resulting in multipolar spindle formation, genomic instability, and DNA damage. Conclusion We conclude that HCR is localized and stabilized at the centrosome by directly binding to astrin. HCR are required for the centrosomal recruitment of MCPH proteins and centriolar duplication. Both HCR and astrin play key roles in keeping normal microtubule assembly and maintaining genomic stability.

Stably Expressed Genes (SEGs) are a set of genes with invariant expression. Identification of SEGs, especially among both healthy and diseased tissues, is of clinical relevance to enable more accurate data integration, gene expression comparison and biomarker detection. However, it remains unclear how many global SEGs there are, whether there are development-, tissue- or cell-specific SEGs, and whether diseases can influence their expression. In this research, we systematically investigate human SEGs at single-cell level and observe their development-, tissue- and cell-specificity, and expression stability under various diseased states. A hierarchical strategy is proposed to identify a list of 408 spatial-temporal SEGs. Development-specific SEGs are also identified, with adult tissue-specific SEGs enriched with the function of immune processes and fetal tissue-specific SEGs enriched in RNA splicing activities. Cells of the same type within different tissues tend to show similar SEG composition profiles. Diseases or stresses do not show influence on the expression stableness of SEGs in various tissues. In addition to serving as markers and internal references for data normalization and integration, we examine another possible application of SEGs, i.e., being applied for cell decomposition. The deconvolution model could accurately predict the fractions of major immune cells in multiple independent testing datasets of peripheral blood samples. The study provides a reliable list of human SEGs at the single-cell level, facilitates the understanding on the property of SEGs, and extends their possible applications.