ObjectiveHepatocellular carcinoma (HCC) represents 90% of all primary liver cancers. The main risk factors associated with HCC include viral hepatitis (B and/or C), alcohol abuse, and metabolic dysfunction-associated steatotic liver disease (MASLD), which progressively advance to liver fibrosis, cirrhosis, and ultimately evolve into HCC. Surgical resection represents the most effective treatment for HCC, while recent advances in immunotherapy, including immune checkpoint inhibitors and adoptive cell therapies, have provided improved treatment prospects for patients with unresectable HCC. However, the complex metabolic heterogeneity of HCC limits the therapeutic efficacy. Metabolic intermediates acyl-CoA not only provide energy and substrates for numerous biochemical reactions but also serve as donors for protein lysine acylation, a major class of post-translational modification (PTM). Therefore, a deeper understanding of the molecular mechanisms underlying protein lysine acylation and hepatocarcinogenesis is urgently needed. MethodsThe levels of protein lysine acylation and silence information regulator 5 (SIRT5) expression levels in clinical HCC samples were analyzed by Western blot. Quantitative malonylome and succinylome of HCC samples were analyzed by antibody-based affinity enrichment coupled with tandem mass spectrometry. The proliferation of HCC cells was analyzed with Cell Counting Kit-8 (CCK-8) assays, the apoptosis was quantified by Annexin V-FITC/propidium iodide (PI) staining coupled with flow cytometry, and the ability of cells to migrate was assayed by Transwell assays. The enzymatic activity of glutathione S-transferase Mu 1 (GSTM1) was quantified. Transgenic mice with hepatic overexpression of SIRT5 were constructed using CRISPR-Cas9, and primary hepatocarcinogenesis was induced by administration of diethylnitrosamine. ResultsWestern blot analysis indicated that the expression level of SIRT5 was elevated in clinical samples from HCC patients, and the levels of lysine malonylation, glutarylation, and succinylation were significantly reduced in HCC tissues. Knockout of SIRT5 in MHCC-97H and MHCC-97L hepatoma cells suppressed cell proliferation, and increased the percentage of apoptotic cells significantly. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses of the differentially malonylome and succinylome of HCC samples revealed significant enrichment in two major classes of biological processes: core energy metabolism (e.g., glycolysis/gluconeogenesis, tricarboxylic acid metabolic process, fatty acid beta oxidation) and detoxification and oxidative stress response (e.g., response to toxic substance, chemical carcinogenesis, reactive oxygen species (ROS)). SIRT5 removes malonylation from lysine residues in GSTM1 and restores its detoxification activity, which is crucial for the survival of hepatocytes under stressed conditions. More importantly, in vivo experiment indicated that hepatic-specific overexpression of SIRT5 in mice accelerated diethylnitrosamine-induced liver fibrosis and hepatocarcinogenesis, indicating the critical role of SIRT5 in HCC progression. ConclusionThis study highlights the previously unrecognized SIRT5-GSTM1 axis as a key regulator in hepatocarcinogenesis, and suggests a potential target for the treatment of patients with HCC.

Membrane proteins are integral components of cellular membranes, accounting for approximately 30% of the mammalian proteome and serving as targets for 60% of FDA-approved drugs. They are critical to both physiological functions and disease mechanisms. Their functional protein-protein interactions form the basis for many physiological processes, such as signal transduction, material transport, and cell communication. Membrane protein interactions are characterized by membrane environment dependence, spatial asymmetry, weak interaction strength, high dynamics, and a variety of interaction sites. Therefore, in situ analysis is essential for revealing the structural basis and kinetics of these proteins. This paper introduces currently available in situ analytical techniques for studying membrane protein interactions and evaluates the characteristics of each. These techniques are divided into two categories: label-based techniques (e.g., co-immunoprecipitation, proximity ligation assay, bimolecular fluorescence complementation, resonance energy transfer, and proximity labeling) and label-free techniques (e.g., cryo-electron tomography, in situ cross-linking mass spectrometry, Raman spectroscopy, electron paramagnetic resonance, nuclear magnetic resonance, and structure prediction tools). Each technique is critically assessed in terms of its historical development, strengths, and limitations. Based on the authors’ relevant research, the paper further discusses the key issues and trends in the application of these techniques, providing valuable references for the field of membrane protein research. Label-based techniques rely on molecular tags or antibodies to detect proximity or interactions, offering high specificity and adaptability for dynamic studies. For instance, proximity ligation assay combines the specificity of antibodies with the sensitivity of PCR amplification, while proximity labeling enables spatial mapping of interactomes. Conversely, label-free techniques, such as cryo-electron tomography, provide near-native structural insights, and Raman spectroscopy directly probes molecular interactions without perturbing the membrane environment. Despite advancements, these methods face several universal challenges: (1) indirect detection, relying on proximity or tagged proxies rather than direct interaction measurement; (2) limited capacity for continuous dynamic monitoring in live cells; and (3) potential artificial influences introduced by labeling or sample preparation, which may alter native conformations. Emerging trends emphasize the multimodal integration of complementary techniques to overcome individual limitations. For example, combining in situ cross-linking mass spectrometry with proximity labeling enhances both spatial resolution and interaction coverage, enabling high-throughput subcellular interactome mapping. Similarly, coupling fluorescence resonance energy transfer with nuclear magnetic resonance and artificial intelligence (AI) simulations integrates dynamic structural data, atomic-level details, and predictive modeling for holistic insights. Advances in AI, exemplified by AlphaFold’s ability to predict interaction interfaces, further augment experimental data, accelerating structure-function analyses. Future developments in cryo-electron microscopy, super-resolution imaging, and machine learning are poised to refine spatiotemporal resolution and scalability. In conclusion, in situ analysis of membrane protein interactions remains indispensable for deciphering their roles in health and disease. While current technologies have significantly advanced our understanding, persistent gaps highlight the need for innovative, integrative approaches. By synergizing experimental and computational tools, researchers can achieve multiscale, real-time, and perturbation-free analyses, ultimately unraveling the dynamic complexity of membrane protein networks and driving therapeutic discovery.

Enzyme-powered micro/nanomotors(MNMs)(EMNMs)use natural enzymes to facilitate the decom-position of fuels,including hydrogen peroxide(H2O2),glucose,triglycerides,and urea to provide power.EMNMs can achieve self-propulsion through the in situ utilization of biofuels without additional fuels,exhibiting excellent biocompatibility and significant potential for application in the biomedical field.Compared with H2O2,which may cause oxidative damage to the body,urea exhibits superior biosafety characteristics.Presently,urease-powered MNMs(UMNMs)have made notable progress in their appli-cations in the biomedical field and have garnered considerable attention from researchers.In this review,we present the latest advancements in the biomedical field of UMNMs,primarily focusing on:1)diverse materials used for constructing the fundamental framework of motors;2)control of motor movement through the regulation of enzymatic reaction rates;and 3)research directions for the clinical application of motors,including in vivo imaging,biomarker detection,cancer treatment,optical therapy,overcoming biological barriers,antibacterial interventions,antithrombotic strategies,and gastric disease manage-ment.Despite showing immense potential in biomedical applications,there are still several challenges impeding its practical implementation,such as maintaining activity in the in vivo environment while accurately targeting specific sites to achieve the desired clinical therapeutic effects.

PURPOSE: The rate of complications among patients undergoing surgery has increased due to infection with SARS-CoV-2 and other variants of concern. However, Omicron has shown decreased pathogenicity, raising questions about the risk of postoperative complications among patients who are infected with this variant. This study aimed to investigate complications and related factors among patients with recent Omicron infection prior to undergoing orthopedic surgery. METHODS: A historical control study was conducted. Data were collected from all patients who underwent surgery during 2 distinct periods: (1) between Dec 12, 2022 and Jan 31, 2023 (COVID-19 positive group), (2) between Dec 12, 2021 and Jan 31, 2022 (COVID-19 negative control group). The patients were at least 18 years old. Patients who received conservative treatment after admission or had high-risk diseases or special circumstances (use of anticoagulants before surgery) were excluded from the study. The study outcomes were the total complication rate and related factors. Binary logistic regression analysis was used to identify related factors, and odds ratio (OR) and 95% confidence interval (CI) were calculated to assess the impact of COVID-19 infection on complications. RESULTS: In the analysis, a total of 847 patients who underwent surgery were included, with 275 of these patients testing positive for COVID-19 and 572 testing negative. The COVID-19-positive group had a significantly higher rate of total complications (11.27%) than the control group (4.90%, p < 0.001). After adjusting for relevant factors, the OR was 3.08 (95% CI: 1.45-6.53). Patients who were diagnosed with COVID-19 at 3-4 weeks (OR = 0.20 (95% CI: 0.06-0.59), p = 0.005), 5-6 weeks (OR = 0.16 (95% CI: 0.04-0.59), p = 0.010), or ≥7 weeks (OR = 0.26 (95% CI: 0.06-1.02), p = 0.069) prior to surgery had a lower risk of complications than those who were diagnosed at 0-2 weeks prior to surgery. Seven factors (age, indications for surgery, time of operation, time of COVID-19 diagnosis prior to surgery, C-reactive protein levels, alanine transaminase levels, and aspartate aminotransferase levels) were found to be associated with complications; thus, these factors were used to create a nomogram. CONCLUSION Omicron continues to be a significant factor in the incidence of postoperative complications among patients undergoing orthopedic surgery. By identifying the factors associated with these complications, we can determine the optimal surgical timing, provide more accurate prognostic information, and offer appropriate consultation for orthopedic surgery patients who have been infected with Omicron.
Humans ; COVID-19/complications* ; Male ; Female ; Middle Aged ; Postoperative Complications/epidemiology* ; SARS-CoV-2 ; Orthopedic Procedures/adverse effects* ; Aged ; Nomograms ; Adult ; Retrospective Studies ; Risk Factors

T cell exhaustion plays an immunosuppressive role in malignant tumors. Continuous tumor antigen stimulation, the presence of suppressive immune cells and cytokines in the tumor microenvironment, the up regulation of inhibitory receptor expression on the surface of T cells, changes in T cell related transcription factors, and metabolites in the tumor microenvironment may lead to T cell exhaustion. Reversing the exhaustion of T cells in tumor patients is a promising strategy for tumor immunotherapy. This article will review the latest research progress on T cell exhaustion status, pathogenesis, reversal methods, and clinical applications in hematological tumors.
Humans ; Hematologic Neoplasms/immunology* ; T-Lymphocytes/immunology* ; Tumor Microenvironment ; Immunotherapy ; T-Cell Exhaustion

OBJECTIVE: To explore the mechanism of colon dialysis with Yishen Decoction (YS) in improving the autophagy disorder of intestinal epithelial cells in chronic renal failure (CRF) in vivo and in vitro. METHODS: Thirty male SD rats were randomly divided into normal, CRF, and colonic dialysis with YS groups by a random number table method (n=10). The CRF model was established by orally gavage of adenine 200 mg/(kg•d) for 4 weeks. CRF rats in the YS group were treated with colonic dialysis using YS 20 g/(kg•d) for 14 consecutive days. The serum creatinine (SCr) and urea nitrogen (BUN) levels were detected by enzyme-linked immunosorbent assay. Pathological changes of kidney and colon tissues were observed by hematoxylin and eosin staining. Autophagosome changes in colonic epithelial cells was observed with electron microscopy. In vitro experiments, human colon cancer epithelial cells (T84) were cultured and divided into normal, urea model (74U), YS colon dialysis, autophagy activator rapamycin (Ra), autophagy inhibitor 3-methyladenine (3-MA), and SIRT1 activator resveratrol (Re) groups. RT-PCR and Western blot were used to detect the mRNA and protein expressions of zonula occludens-1 (ZO-1), Claudin-1, silent information regulator sirtuin 1 (SIRT1), LC3, and Beclin-1 both in vitro and in vivo. RESULTS: Colonic dialysis with YS decreased SCr and BUN levels in CRF rats (P<0.05), and alleviated the pathological changes of renal and colon tissues. Expressions of SIRT1, ZO-1, Claudin-1, Beclin-1, and LC3II/I were increased in the YS group compared with the CRF group in vivo (P<0.05). In in vitro study, compared with normal group, the expressions of SIRT1, ZO-1, and Claudin-1 were decreased, and expressions of Beclin-1, and LC3II/I were increased in the 74U group (P<0.05). Compared with the 74U group, expressions of SIRT1, ZO-1, and Claudin-1 were increased, whereas Beclin-1, and LC3II/I were decreased in the YS group (P<0.05). The treatment of 3-MA and rapamycin regulated autophagy and the expression of SIRT1. SIRT1 activator intervention up-regulated autophagy as well as the expressions of ZO-1 and Claudin-1 compared with the 74U group (P<0.05). CONCLUSION Colonic dialysis with YS could improve autophagy disorder and repair CRF intestinal mucosal barrier injury by regulating SIRT1 expression in intestinal epithelial cells.
Animals ; Sirtuin 1/metabolism* ; Drugs, Chinese Herbal/therapeutic use* ; Autophagy/drug effects* ; Male ; Intestinal Mucosa/drug effects* ; Rats, Sprague-Dawley ; Epithelial Cells/metabolism* ; Colon/drug effects* ; Humans ; Kidney Failure, Chronic/drug therapy* ; Signal Transduction/drug effects* ; Renal Dialysis ; Rats ; Kidney/drug effects*

OBJECTIVES: Metastasis is the primary cause of death in osteosarcoma, and current clinical treatments remain limited. BRD4, a key epigenetic regulator, has shown therapeutic promise in various cancers through its inhibition. However, the mechanistic role of BRD4 in osteosarcoma remains poorly understood. This study aims to elucidate the molecular mechanisms by which BRD4 regulate osteosarcoma progression and to explore novel therapeutic strategies. METHODS: Immunofluorescence was used to assess BRD4 expression levels in a tissue microarray containing 80 osteosarcoma samples from different patients. The Gene Expression Omnibus (GEO) dataset (GSE42352, containing survival data from 88 osteosarcoma patients) was downloaded to perform Kaplan-Meier survival analysis based on BRD4 gene expression levels. In vivo, an orthotopic intramedullary osteosarcoma model was established using HOS cells in C57 mice, followed by treatment with varying doses of the BRD4 inhibitor (+)-JQ1. Micro-CT, 3D reconstruction of bone tissue, and HE staining were employed to evaluate pathological changes in bone and intestinal lymph nodes. In vitro, cell viability was measured using the methyl thiazolyl tetrazolium (MTT) assay, while colony formation and Transwell assays assessed proliferative and invasive capacities. Chromatin-bound BRD4 was analyzed via co-immunoprecipitation combined with mass spectrometry (Co-IP/MS), and O-GlcNAc glycosylation sites and glycan chains of BRD4 were identified using Co-IP with Nano-LC MS/MS. Real-time PCR and Western blotting were used to analyze the relative mRNA and protein expression levels of target genes, respectively. RESULTS: BRD4 was positively expressed in 61.25% (49/80) of osteosarcoma tissues. Patients with high BRD4 expression exhibited significantly shorter survival times (P<0.05). In the orthotopic mouse model, intervention with (+)-JQ1, a potent and commonly used BETi, significantly inhibited tumor growth in vivo and reduced bone destruction (P<0.05). (+)-JQ1 treatment significantly suppressed the proliferation (P<0.001), invasion (P<0.001), and migration (P<0.05) of HOS cells. In osteosarcoma cells, BRD4 exhibited O-GlcNAc modifications at both N- and C- C-termini, particularly at Thr73, which is essential for protein stability. This modification also contributed to the activation of the EGFR tyrosine kinase inhibitor resistance pathway (KEGG Pathway: hsa01521). (+)-JQ1 treatment displaced BRD4 from enhancers and downregulated the transcription of pathway-related genes, such as EGFR and PDGFC, thereby suppressing the malignant behavior of osteosarcoma cells. CONCLUSIONS BRD4 promotes osteosarcoma progression via O-GlcNAc modification at Thr73 and plays a crucial role in tumor growth and metastasis.
Osteosarcoma/drug therapy* ; Humans ; Transcription Factors/metabolism* ; Animals ; Cell Cycle Proteins ; Mice ; Bone Neoplasms/drug therapy* ; Azepines/pharmacology* ; Cell Line, Tumor ; Cell Proliferation/drug effects* ; Triazoles/pharmacology* ; Mice, Inbred C57BL ; Nuclear Proteins/metabolism* ; Gene Expression Regulation, Neoplastic ; Male ; Bromodomain Containing Proteins

Research on immune checkpoints such as programmed death protein-1(PD-1)and cytotoxic T lymphocyte antigen-4(CTLA-4)has provided new directions for tumor treatment.V-domain immunoglobulin suppressor of T-cell activation(VISTA)is an emerging immune checkpoint within the B7 family.Functioning as both a ligand and a receptor,VISTA is an inhibitory immune checkpoint molecule expressed in tumor cells,myeloid cells and T lymphocytes.It plays a crucial role in regulating autoimmunity,inflammatory response and tumor immunity.The non-redundant interactions between VISTA and other immune checkpoints,such as PD-1,may offer new therapeutic strategies and serve as a new target for overcoming immunotherapy resistance.This review summarizes the recent research progress on VISTA in hematological tumors,aiming to provide new insights into its application in the treatment of these malignancies.

Objective To investigate the clinical and pathological characteristics of adrenal cortical carcinoma(ACC),compare differences between hypercortisolism and non-functional ACC,and assess the diagnostic value of indicators such as Ki-67 index.Methods The clinical data of 57 ACC patients admitted to the First Medical Center of Chinese PLA General Hospital from January 2015 to March 2025 were retrospectively analyzed.According to the results of endocrine function assessment,47 of these patients were divided into hypercortisolism group(n=19)and non-functional group(n=28).The differences in clinical and pathological characteristics between the two groups were compared,and non-parametric tests and Spearman correlation analysis were used to explore the relationship between Ki-67 index and tumor stage as well as imaging features.Results Among the 57 patients,there were 20 males and 37 females,with a male-to-female ratio of 1:1.85.The age ranged from 16 to 76 years,and the age at diagnosis was(48.7±13.3)years.The tumor diameter was(10.53±4.14)cm.The tumors were located on the right side in 12 cases(21.1%),on the left side in 34 cases(59.6%),and bilaterally in 11 cases(19.3%).Among them,16 cases(28.1%)were complicated with glucose metabolism disorders,31 cases(54.3%)had hypertension,and 20 cases(35.1%)had hypokalemia.According to ENSAT staging,there were 0 cases in stage Ⅰ,15 cases(26.3%)in stage Ⅱ,24 cases(42.1%)in stage Ⅲ,and 18 cases(31.6%)in stage Ⅳ.Endocrine function assessment was completed in 47 of the 57 patients,including 28 cases(59.6%)of non-functional ACC and 19 cases(40.4%)of hypercortisolism(including 1 case of hypercortisolism combined with increased sex hormone secretion).Compared with non-functional group,hypercortisolism group had a significantly higher prevalence of hypertension(P=0.014),later ENSAT stage(P=0.010),and a higher proportion of hypervascularization(P=0.048).The median Ki-67 index was 20%(10%-40%),showing no significant correlation with either the maximum tumor diameter or SUVmax value,but it was related to ENSAT staging,with Ki-67 index in stageⅣ patients being significantly higher than that in stage Ⅱ(P=0.032).Immunohistochemistry results showed that the positive rate of Inhibin-α was 84.8%,and the positive rate of Melan-A was 40.9%.Conclusions ACC is a rare malignant endocrine tumor.ACC patients with hypercortisolism are more likely to be complicated with hypertension,have later staging,and more common hypervascular manifestations.Clinically,their endocrine function should be prioritized for assessment,and more active treatment strategies should be adopted.Diagnosis should be combined with imaging characteristics(such as hypervascularization)and immunohistochemical indicators(Ki-67,Inhibin-α,Melan-A).The significant increase in Ki-67 is in the advanced stage can serve as an important prognostic indicator to guide individualized treatment.

Objective To investigate the effects of estrogen signaling on T-cell recruitment and polarization in acutely injured skeletal muscle.Methods One hundred C57BL/6 male mice,one hundred and eighty C57BL/6 female mice were selected.Twenty-five female mice were ovariectomized(OVX)and 10 male mice were taken as the sham-operated(sham).Then,cardiotoxin(CTX)induced tibialis anterior(TA)injury for preparing mice myoinjury model.Subcutaneous injection of 17β-estradiol(E2)or estrogen receptor antagonist 4-hydroxytamoxifen(4-OHT)was performed.A total of 140 mice(70 males and 70 females)were divided into four group including:PBS-male,CTX-male,PBS-female,and CTX-female.Serum estradiol(E2)levels were measured by ELISA,and muscle injury models were validated via HE staining.Subsequently,20 male and 20 female mice were selected for immunofluorescence(IF)and Real-time PCR to assess estrogen receptors(ER)expression in injured muscle tissue.Further,10 male and 40 female mice were allocated into five experimental groups,including CTX,CTX+E2,CTX+4-OHT,CTX+OVX,CTX+sham.HE staining and IF were performed to evaluate inflammatory infiltration in the injured muscle.Additionally,50 female mice were divided into CTX and CTX+OVX groups,and IF combined with flow cytometry were used to analyze T-cell phenotypes and muscle fiber regeneration in the injured muscle.Results In vivo,serum E2 and myofiber ERβ increased post-injury in mice of both sexes,significantly higher in females.Compared to the control group,E2 alleviated inflammation,OVX exacerbated inflammation,increased CD4+T-cell infiltration,elevated T helper 1 cell(Th1)response,decreased regulatory T cells(Tregs),impaired regeneration.In vitro,IFN-γ/LPS significantly upregulated ERβ in myotubes.Conclusion Estrogen signaling critically regulates muscle inflammation.Estrogen deficiency(OVX)delays repair of skeletal muscle by promoting Th1 response and suppressing Tregs function.