Introduction:Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne pathogen causing severe hemorrhagic disease with high mortality. The viral glycoprotein Gc mediates membrane fusion and represents a key target of CD8⁺ T-cell responses. However, systematic identification and comprehensive evaluation of pan-MHC-I-restricted Gc epitopes remain limited. Methods:An integrated immunoinformatics workflow incorporating five prediction algorithms (IEDB, NetMHCpan4.1, SMMPMBEC, SYFPEITHI, and Rankpep) was applied to identify high-affinity 9-mer epitopes from the CCHFV Gc protein restricted by human HLA-I and murine H2 alleles. Immunogenicity, conservation, toxicity, and allergenicity were assessed using established computational tools. Peptide-MHC interactions were further examined by molecular docking and molecular dynamics simulations. Selected epitopes were experimentally validated by ELISpot assays in BALB/c and SJL mice immunized with a Gc-based DNA vaccine. Results:Ninety-four human HLA-I and thirty-seven murine H2-restricted dominant epitopes were predicted. Among these, 21 epitopes exhibited high binding affinity, favorable immunogenicity, and broad conservation across viral strains. Most candidates showed low predicted toxicity and allergenicity. Structural analyses supported stable peptide-MHC interactions. ELISpot assays confirmed that several epitopes, including APFILLILF and FVKWKVEYI, elicited significant IL-2 responses, indicating functional T-cell activation. Conclusion:This study provides a systematic framework for identifying conserved and immunogenic pan-MHC-I-restricted epitopes within the CCHFV Gc protein. The validated candidates may support the rational design of multi-epitope vaccines and contribute to understanding cellular immune responses against CCHFV.

Cervical cancer is one of the major gynecological malignancies worldwide, and tumor immune escape is essential for its progression. Accumulating evidence indicates that BAP31 participates in tumor development and immune regulation, but its specific role in the immune escape of cervical cancer has not been clarified. We found that BAP31 expression was obviously increased in cervical cancer tissues, and high BAP31 expression was tightly associated with poor prognosis in patients. Cell function experiments showed that silencing BAP31 could effectively recover the immune activity of dendritic cells. Mechanistic studies revealed that TGF-β acts as a downstream molecule of BAP31. BAP31 binds to TGF-β at the asparagine 381 (Asn-381) residue, accelerates the transport of TGF-β from the endoplasmic reticulum to the Golgi apparatus, and promotes its intracellular maturation and extracellular secretion. Rescue experiments confirmed that TGF-β could block the maturation of dendritic cells, reduce their migration capability and strengthen their endocytic activity. In summary, the BAP31-TGF-β axis induces dendritic cell dysfunction and further mediates immune escape in cervical cancer. This study uncovers a new mechanism of cervical cancer immune escape, and suggests that targeting the BAP31-TGF-β axis may be an effective strategy for immunotherapy against cervical cancer.

Objective To investigate the expression characteristics of the interleukin 2-inducible T-cell kinase (ITK) gene in lung adenocarcinoma (LUAD), its clinical prognostic value, and its potential role in the tumor microenvironment. Methods ITK expression data were obtained from the TCGA and GEO datasets (GSE75037, GSE32863) to analyze its differential expressions between LUAD and adjacent normal tissues. The TCGA-LUAD cohort was used for survival analysis, clinicopathological correlation analysis, and to construct an integrated nomogram prognostic model combining ITK expression with clinical features via Cox regression. The TIMER and CIBERSORT algorithms were employed to assess the correlation between ITK expression and immune cell infiltration. GSEA enrichment analysis was used to compare pathway differences between the high and low ITK expression groups. Results ITK expression was significantly lowered in multiple cancers including LUAD, and its low expression was associated with poor prognosis of patients. Clinical analysis revealed that ITK expression was significantly correlated with tumor Stage grade and T stage. The constructed nomogram model demonstrated good predictive performance (C-index=0.67). High ITK expression showed a significant positive correlation with increased infiltration of anti-tumor immune cells such as CD8+ T cells and M1 macrophages. Pathway enrichment analysis indicated that the high ITK expression group was primarily enriched in immune-related pathways like inflammatory response and interferon-gamma signaling, whereas the low expression group was enriched in metabolic reprogramming processes such as fatty acid metabolism and glycolysis. Conclusion ITK is a potential independent prognostic biomarker in LUAD. Its loss of expression may be associated with suppressed tumor immune microenvironment and activated metabolic reprogramming, thereby promoting tumor progression. ITK may serve as a novel target for prognostic assessment and immunotherapy research in LUAD.

Background: Lung cancer remains a major global health burden. RNA-binding proteins (RBPs) play crucial roles in post-transcriptional gene regulation, and their dysregulation is frequently implicated in tumorigenesis. The present study aimed to elucidate the molecular network governed by the highly expressed RBP TIMELESS in lung adenocarcinoma (LUAD) and determine its mechanistic role in LUAD progression. Methods: The Cancer Genome Atlas-LUAD, Gene Expression Omnibus, and single-cell RNA sequencing datasets were analyzed to identify aberrantly expressed RBP genes. The RBP gene TIMELESS exhibited the most significant effect on LUAD cell death and was selected for further study. Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation sequencing and RNA sequencing were employed to identify ferroptosis-related targets directly bound by TIMELESS. Molecular mechanisms underlying the TIMELESS-mediated regulation of ferroptosis in LUAD were investigated via immunoprecipitation–mass spectrometry, glutathione S-transferase pull-down, immunofluorescence–fluorescence in situ hybridization, RNA immunoprecipitation, poly(A)-tail, and RNA stability assays. In an orthotopic lung cancer mouse model treated with erastin (a ferroptosis inducer) and programmed cell death protein 1 (PD-1) blockade, the role of TIMELESS in therapeutic response was assessed via flow cytometry and multiplex immunofluorescence (mIF). Infiltrating immune cells in LUAD were analyzed by tissue microarrays (TMAs) via mIF. Results: TIMELESS significantly affected LUAD cell proliferation and death, and TIMELESS knockdown significantly enriched RNA-binding and ferroptosis pathways. Transferrin (TF) was identified as a direct TIMELESS target governing ferroptosis. TIMELESS was revealed to bind Ccr4-Not transcription complex subunit 3 (CNOT3) to promote TF mRNA degradation. TIMELESS depletion combined with erastin and PD-1 blockade enhances efficacy, prolongs survival, increases T cell and M1 macrophage infiltration, and reduces M2 macrophage infiltration. Further, high TIMELESS expression was inversely correlated with ferroptosis marker 4-hydroxynonenal but positively correlated with programmed cell death ligand 1 (PD-L1), reduced T cell and M1 macrophage infiltration, and increased M2 macrophage infiltration. Conclusions: TIMELESS recruits CNOT3 to accelerate TF mRNA degradation, thereby suppressing ferroptosis and promoting LUAD growth. These findings suggest that the TIMELESS/TF regulatory axis may be a promising therapeutic target for LUAD.

CAR-T cell therapy represents a major breakthrough in hematological malignancies. However, the trade-off between insufficient treatment persistence and treatment-related toxicities (especially cytokine release syndrome, CRS) still limits its wider clinical application. Our review aims to reframe the understanding of this central dilemma. It posits that the dynamic imbalance of intracellular signaling networks and external inflammation in the tumor microenvironment (TME) are the causes of insufficient persistence of CAR-T cells and the severe CRS. The intracellular signaling network and the influence of the external TME jointly regulate CAR-T cell persistence and inflammatory response. We explore strategies for designing CAR-T cells that simultaneously enhance persistence and mitigate severe CRS risk. The next-generation goal is to achieve an optimal therapeutic outcome in which CAR-T cells exhibit sustained antitumor efficacy alongside a favorable safety profile. The central thesis posits that persistence and safety do not have to be chosen one at the expense of the other. Rather, the relationship between persistence and CRS exists in harmony. Through the precise modulation, it is feasible to simultaneously enhance CAR-T cell persistence while effectively mitigating CRS. This review paints a blueprint for the next-generation CAR-T cell therapies that are both more persistent and inherently safer.

As a bridge between humoral and cellular immunity in the body, CD4+ T cells play an important role in the clearance of Hantaan virus (HTNV) infection. The envelope-anchored glycoprotein (GP) is responsible for orchestrating host cell entry and fusion. In the bioinformatics stage of this study, we assessed 22 dominant B-cell epitopes and 191 T-cell epitopes (including MHC-I and MHC-II restricted) covering MHC genotype frequencies ≥30% in various regions worldwide, using multiple prediction algorithms and HLA population-frequency criteria. Through affinity analysis and molecular docking, the relationship between nonapeptide determinants and 15-mer epitopes suggested that consecutive CD4+ T-cell epitopes may share a nonapeptide that plays a major binding role in multiple HLA genotypes. Sequence alignment of the reported viral variants showed that affinity changes of non-conserved epitopes were tiny in either humoral or cellular immunity. Toxicity and allergenicity screening identified 23 peptides as safe and available to the body. Based on these bioinformatics predictions, we selected candidate peptides for experimental validation. In the wet-lab stage, specialized immunological assays (ELISA and ELISpot) were performed using human peripheral blood samples from vaccinated volunteers, and anti-HTNV humoral and cellular immune epitopes were validated. The results are strongly representative in a certain population and HLA genotypes. Our research is grounded in HTNV GP and provides a theoretical basis for the use of epitopes in the prevention and control of HTNV. More importantly, studies of T-cell and B-cell epitopes from multiple dimensions and perspectives would further deepen the understanding of peptide-based immunotherapy against emerging viral infections and accelerate the development of global public health services.

As essential amino acids, branched-chain amino acids (BCAAs) have attracted much attention in the field of tumor metabolism in recent years. This article reviews the metabolic characteristics of BCAAs in tumors and discusses their involvement in tumorigenesis and development through regulating signaling pathways, affecting the immune microenvironment, and promoting tumor metastasis. Research has shown that abnormal BCAA metabolism is closely related to the occurrence, progression, and prognosis of various tumors. By regulating BCAA metabolism, it is expected to provide new targets and strategies for tumor treatment. Indepth research on the specific mechanisms of BCAAs in tumors will provide more evidence for precision cancer treatment.

During viral infection, CD4+ helper T-cell is indispensable for the establishment of the humoral immune protection, CTL activation, and even long-term memory response. It requires MHC-II molecules in viral structural antigens process and presentation. HTNV NP epitopes exhibited high affinity to both of HLA-II superfamilies and H-2-I genes in the present study. Immunogenicity and conservation analyses identified 34 selective epitopes, later validated by molecular docking (MD) with MHC-II structures. NP 15-mer peptides and MHC-II haplotypes were found to interact bidirectionally through hierarchical clustering. In brief, epitopes that exhibit immunoreactivities for a wide range of MHC-II molecules reflect the biomedical practice of vaccination, while haplotype clusters reflect individual differences in T-cell antigen presentation. Then, 11 HTNV variants showed three amino acid substitutions in three epitopes, with little impact on their pan-HLA-II immunoreactivity. Safety analyses indicated that the 34 selective epitopes exhibit favorable safety profiles for potential applications. Finally, we validated the immunogenicity of the selective epitopes using ELISA, ELISpot, and flow cytometry. In conclusion, our work provides a comprehensive assessment of the pan-MHC-II immunoreactivity of HTNV NP and lays the theoretical and technical foundations for the development of protective epitope vaccines in the context of population immunity.

Background: Breast cancer remains a major global health threat to women. While current therapies exist, their limitations necessitate novel strategies. Melatonin, an endogenous circadian regulator, has shown anti-tumor potential, but its mechanisms from a circadian perspective require further exploration. Methods: The anti-tumor effects of melatonin were evaluated through cell proliferation, colony formation, and apoptosis assays. Through data analysis and experimental verification at the RNA and protein levels, the regulatory effect of it on the core clock gene BMAL1 was studied. The role of BMAL1 in mediating melatonin’s suppression of glucose metabolism was assessed by measuring glucose uptake and lactate production. Downstream effector molecules of BMAL1 were identified through molecular interaction and transcriptional regulation analyses. Results: Melatonin significantly inhibited breast cancer cell proliferation and colony formation and induced apoptosis. Mechanistically, it upregulates the core clock gene BMAL1, which suppresses glucose metabolism. ALDH3A1 was identified as a key downstream target of BMAL1, defining a novel “melatonin-BMAL1-ALDH3A1” axis. In vivo studies confirmed that this axis effectively inhibits tumor growth without apparent toxicity, and SR8278 also shows a synergistic effect when used in combination with melatonin. Conclusions: Our findings elucidate the role of the “melatonin-BMAL1-ALDH3A1” axis in combating breast cancer, offering a new direction for treatment and laying the groundwork for developing precision chronotherapy-based combination regimens.

RNA-binding proteins (RBPs), particularly IGF2BP3, play critical but underexplored roles in lung adenocarcinoma (LUAD). This study investigated IGF2BP3's clinical and functional significance using single-cell/RNA sequencing, validated by qPCR, Western blot, and immunohistochemistry. The results show IGF2BP3 was significantly upregulated in LUAD tissues and associated with advanced-stage, larger tumors, lymph node metastasis, and poor prognosis. A prognostic nomogram confirmed its independent predictive value. Functionally, IGF2BP3 knockdown suppressed proliferation, and induced G2/M arrest and apoptosis. GSEA linked high IGF2BP3 to cell cycle activation and low expression to metabolic pathways. Notably, high IGF2BP3 correlated with immune evasion markers (downregulated CD4+ effector T cells, upregulated Th2 cells), while TIDE analysis suggested a better immunotherapy response in low-expressing patients. Drug screening identified BI-2536 as a potential therapy for low-IGF2BP3 cases, supported by strong molecular docking affinity (-7.55 kcal/mol). These findings establish IGF2BP3 as a key driver of LUAD progression and a promising target for immunotherapy and precision medicine.

Primary liver cancer, with 906,000 new cases, ranks sixth in growth among malignant tumors. Additionally, with 830,000 deaths, it ranks third in terms of mortality (1). Hepatocellular carcinoma (HCC), which is the most important form of primary liver cancer, accounts for approximately 90% of liver cancer cases (2). A variety of risk factors can contribute to the development of primary liver cancer, including hepatitis B virus (HBV) infection, hepatitis C virus (HCV) infection, fibrotic chronic liver damage, aflatoxin B1, and excessive alcohol consumption (3,4). The progression of HCC evolves continuously from dysplastic lesions with minor genetic mutations to the late stages of HCC, displaying significant molecular heterogeneity involving numerous molecules (5). The extensive tumor heterogeneity across multiple stages of HCC development hinders patient stratification for effective treatment (6). Therefore, exploring the tumor heterogeneity of HCC would help stratify patients for effective treatment.The tumor transformation of HCC usually originates from hepatocytes and progenitor cells, both of which are epithelial cell types. The plasticity changes in these epithelial cells commonly known as epithelial-to-mesenchymal transition (EMT) increase the complexity of cellular heterogeneity (6). The EMT program in cancer cells can be transiently or stably activated to varying extents during invasion and metastasis. High expression of adhesion molecules can increase cell migration ability and invasiveness. A significant body of evidence illustrates that the EMT plays an important role in cancer invasion and metastasis (7)(8)(9)(10). By analyzing various EMT phenotypes in malignant epithelial hepatocytes, researchers could estimate the complexity and cellular heterogeneity of HCC. Few studies have investigated several EMT markers in a large number of biopsies, making it difficult to identify the occurrence of EMT based on a single marker alone (11). E-cadherin together with occludin or cytokeratins represents the most commonly used markers for epithelial features, while Ncadherins and vimentin are markers for mesenchymal features (8). Hence, in this study, E-cadherin, N-cadherins, and vimentin (EMT triplet) were selected to characterize the occurrence of EMT. B-cell receptor-associated protein 31 (BAP31) is named for its association with the B-cell receptor component immunoglobulin D. BAP31 exhibits an apparent molecular weight of 31 kDa, a characteristic that is clearly observed on a denaturing electrophoresis gel (12). BAP31 has been identified as a cancer antigen (13). It is overexpressed in cancer tissues compared to healthy adjacent tissues, and it holds promise as a prognostic biomarker for several different types of cancer (14)(15)(16)(17). In our previous study, using mIHC and multispectral imaging techniques, we demonstrated that BAP31 promotes cell proliferation by interacting with Serpin Family E Member 2 (SERPINE2) in hepatocellular carcinoma (HCC) (18). Only a few studies have reported on the role of BAP31 in activating invasion and metastasis. Recent studies have demonstrated that BAP31 can induce epithelial-mesenchymal transition (EMT) by enhancing the expression of the EMT-related factor Snail and decreasing the content and membrane distribution of E-cadherin (19). However, it remains scarce how BAP31 induces the entire EMT process. The current study reported a multi-molecule staining dataset of EMT triplet and BAP31 to estimate tumor heterogeneity and preliminarily explored the relationship between BAP31 and EMT.Conventional immunohistochemistry techniques have several limitations. Multiplex immunohistochemistry(mIHC) technology allows for the simultaneous detection of multiple markers on a single tissue section, providing a new method for comprehensive studies of cell composition, cell function, and intercellular interactions.Moreover, the use of multiplex immunohistochemistry (mIHC) (20).Through the use of mIHC, EMT triplets and BAP31 in liver cancer can be elucidated together.Here, we present our dataset containing numerous images and analyzing data from a digitally scanned high-resolution tissue microarray (TMA) with 138 samples, termed LV138. The TMA was stained for HE and other specific biomarkers, such as BAP31, E-cadherins, N-cadherins, vimentin, and 4,6-diamidino-2-phenylindole (DAPI). Each sample was accompanied by clinical data, pathologist annotations, and staging information. Cell and tissue segmentation, as well as the expression of specific biomarkers, were performed using inForm Advanced image analysis software (inForm 2.6, Akoya). The results of each step were compiled and formed part of the dataset. The utility of our datasets was confirmed by preliminary statistical analysis. Application of dataset can prevent a multitude of repetitive operations and offers an insightful approach for investigating molecular heterogeneity of HCC.This patient group consisted of 138 primary liver cancer patients aged between 21 and 98 years, including 37 women and 101 men. We obtained 138 tissue samples from these individuals and stored them in a TMA-LV138. The study involving human participants was reviewed and approved by the Ethics Committee of the Fourth Military Medical University. Our samples were derived from a commercial tissue microarray (TMA), and the supplier ensured that informed consent was obtained from all patients during the data collection process.The original primary liver cancer tissue microarray (TMA), which was stained with hematoxylin and eosin (HE), was independently evaluated and analyzed by at least two pathologists. This meticulous evaluation led to the identification of 138 viable core samples. Each of these core samples was annotated with essential details such as the diagnosis, grade, TNM classification, stage, and type of cancer. Additionally, each core includes basic patient information and a unique identifier for accurate reference.To enhance visualization, the LV138 was scanned using an Aperio GT 450 scanner from Leica Biosystems (USA). The high-resolution scanning process produced SVS images that were subsequently saved in TIF format, ensuring clear and detailed representation of the tissue samples.The studies involving humans were approved by the institutional review board of the Fourth Military Medical University. The studies were conducted in accordance with the local legislation and institutional requirements. The human samples used in this study were acquired from a by-product of routine care or industry.Multiplexed tyramide signal amplification (TSA) immunofluorescence was performed on the LV138 TMA to simultaneously visualize multiple biomarkers on a single section using Opal 7-plex technology (Akoya).Tyramide Signal Amplification (TSA) is an enzymatic detection method based on horseradish peroxidase (HRP). The principle involves the covalent binding of fluorescently labeled tyramine to the tyrosine residues of the target protein under the catalysis of HRP, thereby labeling the target protein with specific fluorescence (21). The original slides of the LV138 TMA were processed into 5-μm-thick sections. The slides were deparaffinized in xylene and rehydrated in an ethanol gradient.The mIHC staining includes four sequential cycles, each specifically targeting one of the following molecular markers: Vimentin, BAP31, E-cadherin, and N-cadherin. At the start of each staining cycle, the slides are first immersed in either EDTA buffer (pH 9.0) or citrate buffer (pH 6.0). The reaction container with the slides is then placed in a microwave for heat-induced antigen retrieval and stripping. To minimize non-specific binding, we apply a 5% Bovine Serum Albumin (BSA) blocking solution to the slides and allow it to rest for 15 minutes. According to a pre-established and optimized protocol, during each round of staining, the slides are first incubated with primary antibodies-anti-Vimentin (Proteintech; 10366-1-AP), anti-BAP31 (FMU-BAP31-2)(18), anti-E-cadherin (Proteintech; 60335-1-IG), anti-N-cadherin (Proteintech; 66219-1-IG)followed by incubation with HRP-conjugated secondary antibodies (Akoya; Opal Polymer HRP Ms + Rb; ARH1001EA) to detect specific molecular markers. Afterwards,during each round of staining, the slides are incubated with fluorophore-conjugated tyramide-Opal 520, Opal 570, Opal 620, and Opal 690-for 10 minutes to visualize the corresponding molecular markers. Once a TSA staining round is complete, the same method used previously is applied for heat-induced antibody stripping and antigen retrieval. Finally, the cell nuclei are stained with 4,6-diamidino-2-phenylindole (DAPI; Sigma-Aldrich, St. Louis, Missouri, USA; Catalog No. D9542), marking the completion of the entire staining process. The slides were scanned using Vectra 3.0 (Akoya) to acquire multispectral images. All the samples are shown in the resulting images and an example of these images is shown in Figure . 1(A).The multispectral images were unmixed by advanced image analysis software (inForm 2.6.0, Akoya). In this study, we analyzed a total of 138 core samples, comprising 118 hepatocellular carcinoma (HCC) cores, 10 cholangiocarcinoma cores, and 10 normal liver cores. All pertinent data were meticulously documented within the LV138 dataset. The data records were systematically organized into four distinct folders: clinical data, optical acquisition and imaging, Artificial Intelligence(AI) scoring, and colocalization. Each file was methodically named according to the LV138 system, incorporating the file ID and data type to facilitate comprehension of our data structure. To further elucidate our organizational approach, we developed a data result diagram showed in Figure . 2 and appended a file type suffix to each file name (e.g., LV138_ID_coloc_data.txt).The clinical data of the patients are stored in Clinical Data.xlsx, including pathologist annotations, age, sex, diagnosis, grade and unique identifiers for each sample.The images were further analyzed using machine learning for additional interpretation. The inForm 2.6.0 software performed tissue and cell segmentation as well as histochemical scoring of markers, producing outputs of both images and analysis data. To further understand the influence of joint function of biomarks, the colocalizations of biomarkers were analyzed, and the results were divided into two categories, BAP31-EMT markers and EMT markers. The BAP31-EMT marker category includes the colocalization analysis results of each EMT marker with BAP31. The results were divided into four groups: BAP31_E-cadherin, BAP31_Ncadherin, BAP31_Vimentin, and BAP31_E-cad_N-cad_Vimentin. The files, including coloc_data.txt, coloc_quant_data.txt, and quant_data.txt, were stored in the data subfolders, and coloc.tif was stored in the image subfolders.The EMT markers category includes the colocalization analysis results between each pair of EMT markers. The results were divided into the following groups: N-cadherin_E-cadherin, Vimentin_Ecadherin, and Vimentin_N-cadherin. The files, including coloc _data.txt, coloc_quant_data.txt, and quant_data.txt, were stored in the data subfolders, and coloc.tif was stored in the image subfolders.For the preliminary analysis of the data, the datasets were analyzed using GraphPad Prism 10 software (San Diego, CA, United States). The Pearson correlation test was conducted to explore the correlations between four biomarkers, during which the correlation coefficients and P values were calculated, and scatter diagrams were generated. Analysis of variance (ANOVA) was performed to test for the presence of significant differences in different grades, and the H-score was used to represent expression levels. For non-normally distributed rank/ordered variables and data, the Kruskal-Wallis test was used. All P values were two-sided, and a P values less than 0.05 were considered to indicate statistical significance.ANOVA was used to compare the expression levels of BAP31 across different grades of cancer and in normal liver tissue. Notably, BAP31 expression was significantly greater in cancerous tissue than in normal liver tissue, indicating a strong correlation between elevated BAP31 expression and poor prognosis. Pearson correlation analysis revealed a significant relationship between the expression of N-cadherin and E-cadherin in liver cancer and normal liver tissues, with a Pearson's r value of 0.7749 and a P value less than 0.0001. This finding aligns with the results from GEPIA2 (Figure GEPIA 2 -Copyright © 2018) (http://gepia2.cancer-pku.ac.cn), where the Pearson's r value was 0.4 with a P value of 0. To further investigate the correlation between N-cadherin and E-cadherin expression in various tissues, we examined their relationship in normal liver and cancer tissues using the same methodology. The results indicated a significant correlation between a normal liver (Pearson's r = 0.7204, P = 0.0188) and liver cancer (Pearson's r = 0.7723, P < 0.0001). However, no significant correlation was found between the two biomarkers in cholangiocarcinoma (P = 0.7475). These findings were corroborated by data from GEPIA2 (http://gepia2.cancer-pku.ac.cn).Contrary to the traditional view of decreased E-cadherin and increased N-cadherin, the strong correlation between N-cadherin and E-cadherin suggests that the conventional epithelialmesenchymal transition (EMT) model may not be applicable for characterizing hepatocellular carcinoma, but its applicability to cholangiocarcinoma remains to be explored. Recent multicenter study have shown that the expression of N-and E-cadherin are markers for normal hepatocytes and cholangiocytes, respectively, and that the expression of E-and N-cadherin is retained in HCC and intrahepatic cholangiocarcinoma (iCCA) (22,23). his research aligns with our findings, further validating the credibility and value of our dataset.Additionally, the colocalization of E-cadherin and BAP31 was analyzed using ANOVA across four grades of cancer, yielding a p value of 0.0104. The colocalization analysis showed the extent of the overlapping area between the two biomarkers within the entire tissue core, suggesting a potential relationship between the degree of interaction between the biomarkers and the progression of cancer. Notably, survival data from the TCGA database (https://www.cancer.gov/aboutnci/organization/ccg/research/structural-genomics/tcga) indicate a trend toward diminished overall survival with increased transcription of the corresponding mRNA. This observation supports the validity of our database for survival assessments.Our findings suggest that multispectral analysis can significantly contribute to the diagnosis of liver cancer. Further studies based on a larger patient cohort are warranted. Moreover, incorporating survival outcomes could provide a critical indicator for predicting survival rates.Figure1(A): Four biomarkers of normal cores and HCC cores at different stages stained by mIHC technology; a darker color indicates higher biomarker expression. (B) The stained core was multispectral split, and the merged image was split into five images: Opal520-stained vimentin (green), Opal570-stained BAP31 (brown), Opal620-stained E-cadherin (orange), Opal690-stained Ncadherin (red) and DAPI (blue). (C) Colocalization of E-cadherin and N-cadherin. Colocalization analysis was subsequently performed. The percentage colocalization area (golden) was calculated. (D) All three steps to train the inForm for segmentation and the calculation of the H-score. From left to right are the tissue segmentation, cell segmentation and H-score. Each step is further analyzed based on the previous step. Different tissues were first divided into cancer (red), stromal (green), and background (blue) tissues. Then, the cells were segmented. According to the biomarker expression level, we set thresholds to divide cells into 4 grades (blue indicates no expression, yellow indicates +1 expression, orange indicates +2 and brown indicates +3), and the H-score was calculated. The percentage of the colocalization area (golden) was calculated.Figure2: Detailed structure drawing of the data record.

Tumor metastasis is the main cause of hepatocellular carcinoma (HCC) related death. Loss of cell polarity may lead to weakened cell adhesion, epithelial-mesenchymal transition (EMT), and metastasis of HCC. However, the mechanism involved in HCC cells polarity loss is still less studied. Here, we found that BAP31 expression increased with tumor grade and metastasis. Moreover, BAP31 silencing inhibited invasion and migration and recovered the polarity of HCC cells. RNA-seq identified SPINK6 was a downstream gene of BAP31, and was associated with tumor stage and metastasis in HCC. IP-MS and IF assays showed that BAP31 bound to the RNA binding protein ELAVL1, and promoted its maturation. In addition, RIP, RNA-FISH, RNA stability and luciferase reporter assays confirmed that ELAVL1 could bind to the 3 'UTR region of SPINK6 mRNA to stabilize its expression. Depletion of SPINK6 inhibited the invasion and migration, re-established the cell polarity and suppressed EMT in HCC cells, while overexpression of SPINK6 partially counteracted BAP31/ELAVL1 knockdown caused attenuation of metastasis and recovery of polarity. Finally, in vivo experiments verified that BAP31-ELAVL1-SPINK6 axis induced cell polarity loss and promoted metastasis in HCC. Our study shed new light on the mechanism of cell polarity loss and metastasis in HCC.

During antiviral immunity, MHC‑I molecules display endogenous peptides to CD8+ T‑cell receptors, prompting cytotoxic elimination of infected cells. The present study focused on dominant epitopes derived from the nucleocapsid protein (NP) of Hantaan virus (HTNV) and revealed their high affinity for the HLA‑I and H‑2 superfamilies. Through immunogenicity and conservation analyses, four selective epitopes were precisely identified. Molecular docking validated the binding characteristics of selective epitopes with MHC‑I molecules. Bidirectional hierarchical clustering analysis uncovered complex interaction patterns between NP 9‑mer peptides and MHC‑I haplotypes. Moreover, in‑depth investigation of 11 HTNV variants revealed three amino acid substitutions (I241S, E242A and F384I) within the four selective epitopes; however, these substitutions did not significantly affect the pan‑HLA‑I immunoreactivity of these epitopes. Safety assessments highlighted the potential of four selective epitopes for practical applications. Utilizing ELISpot, ELISA and flow cytometry, the immunogenicity of these selective epitopes was comprehensively confirmed. In summary, the present study thoroughly evaluated the pan‑MHC‑I immunoreactivity of HTNV NP, providing a robust foundation for developing effective epitope vaccines for population immunity.

Background: The tumor microenvironment (TME) plays a crucial role in the progression of lung adenocarcinoma (LUAD). However, understanding its dynamic immune and stromal modulation remains a complex challenge. Methods: We utilized the ESTIMATE algorithm to evaluate the immune and stromal components of the LUAD TME from the TCGA database. Correlations between these components and clinical characteristics and patient prognosis were analyzed. Toll-like receptor 7 (TLR7) was identified as a key prognostic biomarker through PPI network and COX regression analysis. Validation of TLR7 expression was conducted using GEO data, qPCR, WB, and IHC. A prognostic model was developed using a nomogram, incorporating TLR7 expression. Enrichment analysis, the Tumor Immune Estimation Resource database, and single-sample gene set enrichment analysis were used to explore TLR7’s potential function. The response of the TLR7 subgroup to immunotherapy and drug sensitivity was observed. Results: We found significant associations between the immune and stromal components of LUAD TME and clinical features and prognosis. Specifically, TLR7 was identified as a prognostic biomarker, where lower expression in tumor tissues was linked to worse outcomes. This finding was further confirmed by comparing TLR7 expression in LUAD cells to normal bronchial epithelial cells, revealing lower expression in the tumor cells. Incorporating TLR7 into a nomogram prognostic model resulted in a good predictor of patient survival. Additionally, TLR7 was associated with immune function and positively correlated with various immune cells. Importantly, patients with high TLR7 expression were more likely to benefit from anti-PD-1 checkpoint blockade therapy. We also identified four treatment candidates for patients with high TLR7 expression. Conclusion: TLR7 is a powerful clinical feature that predicts patient prognosis, immunotherapeutic response, and drug candidates, providing additional insights for the treatment of LUAD.

Medical immunology is an important link between basic experiments and clinical applications, and it is also the intersection of many cutting-edge disciplines. At present, there are still limitations in immunology teaching such as single teaching methods and insufficient scientific research training for students. Department of Immunology has implemented practice and exploration of "scientific research back-feeding teaching" in medical immunology course. By incorporating scientific research into teaching activities, the team has carried out teaching practice through scientific research platforms, enriched teaching content with scientific research progress, conducted scientific research activities to stimulate students' interest, which not only increase students' inner drive, improves teaching efficiency, and leads to a "win-win" situation of teaching and scientific research, but also contributes to training medical innovative talents who can shoulder the heavy responsibility of the country.

Vaccines has long been the focus of antiviral immunotherapy research. Viral epitopes are thought to be useful biomarkers for immunotherapy (both antibody-based and cellular). In this study, we designed a novel vaccine molecule, the Hantaan virus (HTNV) glycoprotein (GP) tandem Th epitope molecule (named the Gnc molecule), in silico. Subsequently, computer analysis was used to conduct a comprehensive and in-depth study of the various properties of the molecule and its effects as a vaccine molecule in the body. The Gnc molecule was designed for DNA vaccines and optimized with a lysosomal-targeting membrane protein (LAMP) strategy. The effects of GP-derived Th epitopes and multiepitope vaccines were initially verified in animals. Our research has resulted in the design of two vaccines based on effective antiviral immune targets. The effectiveness of molecular therapies has also been preliminarily demonstrated in silico and in laboratory animals, which lays a foundation for the application of a vaccines strategy in the field of antivirals.

Background: Since 2019, the SARS-CoV-2 virus has been responsible for the global spread of respiratory illness. As of 1 September 2024, the cumulative number of infections worldwide exceeded 776 million. There are many structural proteins of the virus, among which the SARS-CoV-2 nucleocapsid (N) protein plays a pivotal role in the viral life cycle, participating in a multitude of essential activities following viral invasion. An important antiviral immune response is the major histocompatibility complex (MHC)-restricted differentiation cluster 8 (CD8+) T cell cytotoxicity. Therefore, understanding the immunogenicity of SARS-CoV-2 NP-specific MHC-I-restricted epitopes is highly important. Methods: MHC-I molecules from 11 human leukocyte antigen I (HLA-I) superfamilies with 98% population coverage and 6 mouse H2 alleles were selected. The affinity were screened by IEDB, NetMHCpan, SYFPEITHI, SMMPMBEC and Rankpep. Further immunogenicity and conservative analyses were performed using VaxiJen and BLASTp, respectively. EpiDock was used to simulate molecular docking. Cluster analysis was performed. Selective epitopes were validated by enzyme-linked immunospot (ELISpot) assay and flow cytometry in the mice with pVAX-NPSARS-CoV-2 immunization. Enzyme-Linked Immunosorbent Assay (ELISA) was used to detect whether the preferred epitope induced humoral immunity. Results: There were 64 dominant epitopes for the H-2 haplotype and 238 dominant epitopes for the HLA-I haplotype. Further analysis of immunogenicity and conservation yielded 8 preferred epitopes, and docking simulations were conducted with corresponding MHC-I alleles. The relationships between the NP peptides and MHC-I haplotypes were then determined via two-way hierarchical clustering. ELISA, ELISpot assay, and flow cytometry revealed that the preferred epitope stimulated both humoral and cellular immunity and enhanced cytokine secretion in mice. Conclusions: our study revealed the general patterns among multiple haplotypes within the humans and mice superfamily, providing a comprehensive assessment of the pan-MHC-I immunoreactivity of SARS-CoV-2 NP. Our findings would render prospects for the development and application of epitope-based immunotherapy in lasting viral epidemics.

Staphylococcal enterotoxins (SEs) serve as the primary cause of staphylococcal food poisoning and other foodborne intoxications. Among them, staphylococcal enterotoxin C (SEC) has the highest prevalence in dairy products, leading to multiple outbreaks all around the world. Thus, it is of great significance to develop a highly sensitive, highly specific and easy to operate chemiluminescent sandwich enzyme immunoassay (CLEIA) for detecting staphylococcal enterotoxin C (SEC1). We selected two pairs of anti-SEC1 monoclonal antibodies (mAbs) (SEC1-G8 and SEC1-C4), and a chemiluminescent sandwich enzyme immunoassay (CLEIA) was constructed. This approach can detect SEC1 within a concentration spectrum of 3.2-4000 pg/mL, with the detection limit being 2.1 pg/mL. At three concentrations (3.2, 20, and 400 pg/mL), both the intra- and inter-assay coefficient variations were coming in at 6.31 % and 11.2 % respectively. No cross-reaction was noticed in the SEA, SEB, and SED tests. SEC1 was successfully detected by employing the CLEIA method in spiked matrices and commercial samples, and the average recovery rate ranges from 81.6 % to 108.1 %. Therefore, the highly sensitive, SEC1- specific, and easy-to-operate CLEIA could be a useful tool in the near future for quantifying SEC1 in public health and food safety.

Marburg hemorrhagic fever (MHF) is a fatal infectious disease caused by Marburg virus (MARV) infection, and MARV has been identified as a priority pathogen for vaccine development by the WHO. The glycoprotein (GP) of MARV mediates viral adhesion and invasion of host cells and therefore can be used as an effective target for vaccine development. Moreover, DNA vaccines have unique advantages, such as simple construction processes, low production costs, and few adverse reactions, but their immunogenicity may decrease due to the poor absorption rate of plasmids. Lysosome-associated membrane protein 1 (LAMP1) can direct antigens to lysosomes and endosomes and has great potential for improving the immunogenicity of nucleic acid vaccines. Therefore, we constructed a DNA vaccine based on a codon-optimized MARV GP (ID MF939097.1) fused with LAMP1 and explored the effect of a LAMP targeting strategy on improving the immunogenicity of the MARV DNA vaccine. ELISA, ELISpot, and flow cytometry revealed that the introduction of LAMP1 into the MARV DNA candidate vaccine improved the humoral and cellular immune response, enhanced the secretion of cytokines, and established long-term immune protection. Transcriptome analysis revealed that the LAMP targeting strategy significantly enriched antigen processing and presentation-related pathways, especially the MHC class II-related pathway, in the candidate vaccine. Our study broadens the strategic vision for enhanced DNA vaccine design and provides a promising candidate vaccine for MHF prevention.