BackgroundLong-term immunosuppression following transplantation places recipients at a high risk of malignancy. Hepatocellular carcinoma (HCC) recurrence after transplantation poses significant challenges to long-term survival of recipients. Several studies in transplant oncology have established cancer-transplant models to support the development of therapies for reducing the risk of post-transplant cancer recurrence. However, existing models fail to recapitulate the complex immune status of recipients and the authentic tumor microenvironment after transplantation.MethodsC57BL/6 recipient mice were injected intravenously with luciferase-expressing Hepa1-6 cells 7 days before receiving Balb/c cardiac allografts, followed by post-transplant immunosuppression with tacrolimus. On day 7 post-transplantation, the immune state of recipients was assessed by measuring serum inflammatory cytokines, and allograft rejection was evaluated by hematoxylin-eosin staining. Cancer progression was evaluated using in vivo imaging and measurements of serum alpha-fetoprotein levels, with subsequent histological confirmation.ResultsAllogeneic cardiac transplantation resulted in a significant increase in serum levels of pro-inflammatory cytokines accompanied by a marked reduction in cancer burden. Marked lymphocytic infiltration, hemorrhage, and structural disintegration were observed in the grafts of untreated animals. In contrast, tacrolimus treatment effectively attenuated both the inflammatory cytokine response and acute allograft rejection but, conversely, resulted in a significant increase in tumor burden. Histological analysis confirmed that malignancies were exclusively localized to the lungs, mirroring the most common site of clinical HCC recurrence post-transplantation.ConclusionThis model effectively simulates the elevated risk of cancer recurrence under post-transplant immunosuppression and faithfully recapitulates lung metastasis of HCC mediated by circulating cancer cells—the most common site of clinical recurrence following liver transplantation for HCC. It thus provides a reliable tool for basic research aimed at reducing risk of post-transplant cancer recurrence.

Hepatocellular carcinoma (HCC) recurrence after liver transplantation (LT) remains a major cause of mortality, with current selection criteria such as the Milan and UCSF standards relying solely on tumor morphology and failing to fully account for biological aggressiveness. Emerging evidence implicates dysregulated lipid metabolism, particularly sterol regulatory element-binding protein 2 (SREBP2), in HCC progression, yet its prognostic role in post-LT recurrence remains unexplored. To address this gap, this study developed and validated a novel SREBP2-integrated nomogram for improved risk stratification in 206 HCC patients undergoing LT (2015-2022), randomly split into development (n = 144) and validation (n = 62) cohorts. SREBP2 levels were quantified via ELISA (sensitivity: 0.1 ng/mL) and then incorporated into a multivariate Cox model alongside tumor number and AFP to predict recurrence-free survival (RFS). The nomogram demonstrated superior discrimination, with C-indices of 0.778 (development) and 0.796 (validation). High SREBP2 levels (≥ 40 ng/mL) independently predicted recurrence (HR = 1.757, p = 0.011), whereas decision curve analysis confirmed greater clinical net benefit across 1-/3-/5-year RFS predictions. As the first study to integrate SREBP2 into LT candidate selection, this biologically informed nomogram significantly enhances recurrence prediction, offering a refined tool for transplant eligibility assessment and adjuvant therapy guidance in HCC management. By combining molecular biomarkers with clinical parameters, this approach addresses a critical unmet need in optimizing post-LT outcomes.

Ferroptosis is reported to be closely involved in acetaminophen (APAP) overdose induced liver injury. The development of hepatoprotective drugs that target ferroptosis inhibition is highly valuable. In our previous work, we have confirmed the antioxidant properties of syringaresinol (SYR), but its effects on ferroptosis and the potential molecular mechanisms are unclear. This study aimed to investigate the impact of SYR on APAP-induced hepatocyte ferroptosis and elucidate the role of Caveolin-1 (CAV-1) in the protective function of SYR, focusing on its regulation of the Nrf2/HO-1 signaling pathway. In vitro validation was performed in APAP-treated AML12 and HL7702 cells. A mouse model of acute liver injury was established using intraperitoneal APAP injection. SYR was administered at low and high doses. We found that SYR significantly reduced ROS accumulation and Fe2+ overload, thereby alleviating APAP-induced cell death in vitro. Similarly, SYR alleviated APAP-induced liver injury by decreasing the level of Fe2+ and lipid peroxidation in mice. Western blot analysis also demonstrated SYR’s suppression of APAP-induced ferroptosis. Furthermore, SYR treatment dramatically elevated the protein expression of CAV-1 and activated the Nrf2/HO-1 pathway. Most importantly, knockdown of CAV-1 markedly abolished the protective effects of SYR against APAP-induced hepatocyte ferroptosis, as well as its activation effect on Nrf2/HO-1 pathway, indicating its critical role in SYR’s function on ferroptosis. Collectively, our results indicate that SYR protects against APAP-induced hepatocyte ferroptosis through the CAV-1/Nrf2/HO-1 signaling pathway.

OBJECTIVE:Spinal cord injury (SCI) is a severe secondary injury that often results in impaired motor function, imposing a significant burden on both individuals and society. Therefore, there is an urgent need for new therapeutic targets and strategies to address this challenge. METHODS:To construct a mouse model of SCI, an aneurysm clip was used in vivo to clamp the abdominal aorta below the left renal artery in C57BL/6J mice. After 60 min, the aneurysm clip was removed to restore blood flow. In vitro, primary neuronal cells were subjected to OGD/R to mimic the conditions of SCI. Cell viability was assessed using the CCK-8 assay, and the levels of neuronal death, autophagy, and ferroptosis were determined using a combination of WB, IF, and transmission electron microscopy. IP/MS and Co-IP techniques were employed for the identification and validation of proteins interacting with USP5. RESULTS:Neurons exhibit significant ferroptosis in the SCI mice. USP5 is markedly upregulated in SCI neurons and mediates neural ferroptosis. In vitro experiments demonstrate that overexpression of USP5 promotes neuronal ferroptosis, whereas knockout of USP5 significantly reduces it, with consistent results observed in vivo. Notably, the upregulation of USP5 expression markedly increases the accumulation of autophagosomes and autophagic flux in neurons, which may represent a potential mechanism by which USP5 mediates neuronal ferroptosis. Further investigations utilizing IP/MS and Co-IP confirmed the interaction between USP5 and c-MAF. Additionally, Western blot analysis revealed that USP5, through its deubiquitinating enzyme activity, enhances c-MAF protein stability, thereby activating autophagy and subsequently promoting neuronal ferroptosis. CONCLUSION:In summary, our results indicate a close relationship between ferroptosis and SCI. USP5 regulates c-MAF expression through deubiquitination, thereby activating autophagy-dependent ferroptosis in neurons and mediating the progression of SCI. USP5 may serve as a potential therapeutic target for SCI.

ABSTRACT Objective Capecitabine (CAP) is an orally administered prodrug of fluorouracil, predominantly utilized in the treatment of solid tumors. Triggering tumor ferroptosis is an important mechanism for the treatment of hepatocellular carcinoma (HCC). However, the potential of CAP to induce ferroptosis in HCC, along with the underlying mechanisms, remains unknown. Methods In this study, a subcutaneous HCC model was constructed using the syngeneic Hepa1‐6 cell line in C57BL/6 mice, followed by treatment with metronomic CAP (mCAP). The anti‐tumor effects of mCAP were evaluated by monitoring tumor volume, performing pathological staining, and evaluating tumor oxidative stress levels. In vitro, various thymidylate synthase (TYMS) inhibitors were used to treat both mouse and human HCC cell lines. Furthermore, TYMS‐overexpressing plasmids were transfected into mouse and human HCC cell lines to directly investigate their impact on intracellular oxidative stress. Intracellular oxidative stress and ferroptosis‐related markers were detected using flow cytometry, transmission electron microscopy, and Western blotting. Results 5‐FU, an active metabolite of CAP, as well as TYMS‐specific inhibitors, suppressed the proliferation of Hepa1‐6 and HepG2 cells. These treatments promoted p67phox expression, activated nicotinamide adenine dinucleotide phosphate hydrogen oxidase (NOX), induced reactive oxygen species production, and increased ferrous ion accumulation. Electron microscopy revealed mitochondrial alterations characteristic of ferroptosis. Raltitrexed, another TYMS inhibitor, also induced ferroptosis in hepatocellular carcinoma cells. In vivo, the anti‐tumor effect of mCAP was antagonized by co‐treatment with an NOX inhibitor. Conclusion CAP targets TYMS to induce ferroptosis by activating NOX, thereby inhibiting HCC progression.

Capecitabine (CAP) is widely used in cancer treatment for its oral convenience and tumor targeting. However, its effectiveness in hepatocellular carcinoma (HCC) is suboptimal, possibly due to metabolic enzyme expression differences. This study aims to analyze these enzymes’ expression differences and explore their correlation with clinical pathological factors, to inform personalized CAP treatment. This retrospective study used Immunohistochemistry (IHC) to analyze tumor and non-tumorous samples from HCC patients for CAP metabolic enzyme expression. PRM protein quantification was performed on 10

Osteoarthritis (OA) is a multifactorial joint disease that encompasses not just cartilage deterioration but also meniscal degeneration, subchondral bone remodeling, and fibrosis of the infrapatellar fat pad and synovial membrane. OA is characterized by a progressive and irreversible loss of the articular cartilage, due in main part to the cleavage of type II collagen within the matrix by the enzyme matrix metalloproteinases (MMPs). As an important member of MMP family, MMP-3 plays a prominent role in the pathologenesis of OA. The pathological factors and signaling pathways mediating Mmp-3 expression activation during OA development are not well defined and have been a focus of intense research. Mmp-3 promoter is featured by CpG sites. So far there have been no reports concerning the functional role of DNA methylation in Mmp-3 promoter during OA development. In this study, we sought to investigate DNA methylation status in Mmp-3 promoter, the role of DNA methylation in Mmp-3 activation in OA, and the underlying mechanisms. Our results showed that Mmp-3 promoter demethylation played an important role in Mmp-3 expression activation in OA, which facilitated signal transducer and activator of transcription 4 (STAT4) binding on Mmp-3 promoter to activate Mmp-3 expression. More importantly, OA pathological phenotype of mice was alleviated in response to siRNA-mediated STAT4 knockdown. Our study illustrates how subtle changes in DNA methylation/demethylation status can directly influence gene expression and contribute to pathology and takes us one step further in the understanding of the pathogenesis of OA.

Objective: To explore the clinical application effect of a new rehabilitation training device for elderly knee joints, and to provide a basis for its promotion and the optimization of elderly knee joint rehabilitation nursing. Methods: Eighty elderly patients with knee joint diseases were selected to conduct a 4-week clinical trial of the new rehabilitation training device, and targeted nursing interventions were implemented. The knee joint function score (HSS) and visual analog scale (VAS) were used to evaluate the patients' knee joint function and pain degree, and the treatment compliance, satisfaction and incidence of adverse events were counted. Results: After the trial, the patients' HSS score (78.5±7.2) was significantly higher than that before the trial (62.3±8.5), and the VAS score (2.3±1.1) was significantly lower than that before the trial (6.8±1.5). The differences were statistically significant (P＜0.05). The patients' treatment compliance was 95%, the total satisfaction with the device and nursing work was 97.5%, and the incidence of adverse events was only 2.5% (2 cases of mild knee joint soreness and discomfort, which were relieved after intervention). Conclusion: The new rehabilitation training device for elderly knee joints combined with targeted nursing intervention can effectively improve the patients' knee joint function, relieve pain, and has high safety and patient recognition, which is worthy of clinical promotion.

Objective:Surgical intervention and fixation is the recognized measurement to treat pubic symphysis diastasis caused by high-energy trauma. The purpose of this retrospective study was to assess the clinical application of modified pedicle screw-rod fixation (modified PSRF) and open reduction plate fixation (ORPF) for treating pubic symphysis diastasis. Methods:The data of this retrospective analysis were collected from 32 patients with pubic symphysis diastasis managed with modified PSRF or ORPF from January 2012 to December 2017, with or without posterior fixation. Indicators of clinical assessments including operating time, intraoperative blood loss, relevant surgical complications as well as follow-up were recorded. Majeed scores were performed for functional evaluation, as well as Matta criteria were applied to evaluate the quality of reduction. Results:The average time from injury to operation was 2.9 days in modified PSRF group and 3.2 days in ORPF group. Significant differences regarding average operation time (41.8 min versus 64.3 min) and average intraoperative blood loss (46.6 ml versus 304.6 ml) were presented between modified PSRF groups and ORPF group. Neither Majeed scores nor Matta evaluation showed a significant difference between two groups. In ORPF group, the incision infection occurred in one patient and two patients developed loosening of screws. In modified PSRF group, loosening of screws was found in one patient during the operative procedure and one patient experienced femoral nerve palsy. Irritation to the lateral femoral cutaneous nerve (LFCN) was detected in two patients in modified PSRF group. Conclusions:Satisfactory clinical outcomes were provided with applications of both fixation methods for treating pubic symphysis diastasis. Modified PSRF, as a minimal invasive technique, could serve as an effective and reasonable option for treating pubic symphysis diastasis.Level of evidence: III: retrospective cohort study.Trial registration: researchregistry3906.

BACKGROUNDS:The decision to perform a stoma during surgical resection of penetrating Crohn's disease (CD) is a critical consideration. The objective of this study was to identify factors that influence stoma creation and develop a predictive nomogram model to assist surgeons in making optimal surgical decisions. METHODS:A retrospective study was conducted at a tertiary center from December 1, 2012, to December 1, 2021, involving consecutive patients with penetrating CD who underwent elective surgical resection. The LASSO regression method was used to select preoperative predictors, and a nomogram was constructed using multivariate logistic regression. The performance of the nomogram was validated in an internal cohort by assessing its discrimination, calibration, and clinical usefulness. RESULTS:The study included 405 cases in the training group and 135 cases in the validation group. Nine risk factors for stoma formation were identified, including disease location, fistula resulted from previous anastomosis, absence of preoperative exclusive enteral nutrition support, albumin levels below 35 g/L, C-reactive protein levels above 10 mg/L, hemoglobin levels below 100 g/L, perianal disease, internal fistula, and surgical approach. These risk factors were selected using the LASSO regression method, and a nomogram was developed based on them. The area under the curve and the coefficient of determination (R2) of the nomogram were 0.821 and 0.394, respectively. And the nomogram demonstrated good performance in the validation cohort. CONCLUSIONS:The nomogram exhibited good predictive ability for stoma formation during elective surgical resection for penetrating CD, which can assist surgeons in making informed clinical decisions.

目的 研究胆道闭锁患儿肝外门静脉管壁增厚的病理形态学特征,同时探讨肝外门静脉管壁增厚程度的临床意义及危险因素.方法 分析2022年6月至2022年12月天津市第一中心医院儿童器官移植科60例行肝移植治疗的胆道闭锁患儿的临床病理资料,观察胆道闭锁患儿肝外门静脉管壁增厚的病理形态学改变.根据肝外门静脉总管壁厚度的中位数,将胆道闭锁患儿分为门静脉轻度增厚组及门静脉重度增厚组.比较两组间Kasai术后自体肝生存时间.单因素和多因素Logistic回归分析肝外门静脉管壁增厚程度的影响因素.结果 胆道闭锁患儿肝外门静脉管壁出现不同程度增厚,以血管内皮细胞下间质水肿、纤维和成纤维细胞增生及少量炎细胞浸润为主要病理学改变.肝外门静脉管壁内膜厚度为110(30～640)μm,总管壁厚度为373(160～1320)μm,门静脉内膜厚度/门静脉总管壁厚度的比值0.341(0.105～0.636).对胆道闭锁进行分组,总管壁厚度≤373 μm定义为门静脉轻度增厚组,总管壁厚度＞373 μm定义为门静脉重度增厚组.门静脉重度增厚组自体肝生存时间显著低于门静脉轻度增厚组,差异有统计学意义(P＜0.05).单因素分析结果显示:Kasai手术史和胆管炎病史是影响胆道闭锁肝外门静脉管壁增厚程度的相关因素(P＜0.05).多因素分析结果显示:胆管炎病史是影响胆道闭锁肝外门静脉管壁增厚程度的独立危险因素(优势比=4.000,95%可信区间=1.272～12.578,P＜0.05).结论 胆道闭锁患儿肝外门静脉管壁增厚以血管内皮细胞下间质水肿、纤维和成纤维细胞增生及少量炎细胞浸润为主要特征.胆道闭锁肝外门静脉管壁增厚程度可能影响Kasai术后自体肝生存时间.预防并治疗胆管炎,有助于减轻门静脉管壁增厚的程度.

人肝类器官由于具有与靶器官类似的结构与功能,且可在体外稳定的长期培养、扩增与冻存,因而在多个领域发挥着越来越重要的作用.基于类器官技术的肝类器官生物样本库也应运而生.为了全面概述肝类器官生物样本库的构建情况,本文首先综述了常见的肝类器官构建技术,进一步阐述了肝类器官生物样本库建设需考虑的关键要素,最后展望了其未来的发展方向,以期推动我国肝类器官生物样本库的建设与发展.

Liver transplantation is one of the most effective treatments for hepatocellular carcinoma (HCC). The balance between inhibiting immune rejection and preventing tumor recurrence after liver transplantation is the key to determining the long-term prognosis of patients with HCC after liver transplantation. In our previous study, we found that capecitabine (CAP), an effective drug for the treatment of HCC, could exert an immunosuppressive effect after liver transplantation by inducing T cell ferroptosis. Recent studies have shown that ferroptosis is highly associated with autophagy. In this study, we confirmed that the autophagy inducer rapamycin (RAPA) combined with metronomic capecitabine (mCAP) inhibits glutathione peroxidase 4 (GPX4) and promotes ferroptosis in CD4+ T cells to exert immunosuppressive effects after rat liver transplantation. Compared with RAPA or mCAP alone, the combination of RAPA and mCAP could adequately reduce liver injury in rats with acute rejection after transplantation. The CD4+ T cell counts in peripheral blood, spleen, and transplanted liver of recipient rats significantly decreased, and the oxidative stress level and ferrous ion concentration of CD4+ T cells significantly increased in the combination group. In vitro, the combination of drugs significantly promoted autophagy, decreased GPX4 protein expression, and induced ferroptosis in CD4+ T cells. In conclusion, the autophagy inducer RAPA improved the mCAP-induced ferroptosis in CD4+ T cells. Our results support the concept of ferroptosis as an autophagy-dependent cell death and suggest that the combination of ferroptosis inducers and autophagy inducers is a new research direction for improving immunosuppressive regimens after liver transplantation.

Supplementary Figure S3. The effects of extracellular OPN on cytokine production in macrophages.

BACKGROUND:Capecitabine (CAP) is a classic antimetabolic drug and has shown potential antirejection effects after liver transplantation (LT) in clinical studies. Our previous study showed that metronomic CAP can cause the programmed death of T cells by inducing oxidative stress in healthy mice. Ferroptosis, a newly defined non-apoptotic cell death that occurs in response to iron overload and lethal levels of lipid peroxidation, is an important mechanism by which CAP induces cell death. Therefore, ferroptosis may also play an important role in CAP-induced T cell death and play an immunosuppressive role in acute rejection after trans-plantation.AIM:To investigate the functions and underlying mechanisms of antirejection effects of metronomic CAP.METHODS:A rat LT model of acute rejection was established, and the effect of metronomic CAP on splenic hematopoietic function and acute graft rejection was evaluated 7 d after LT. In vitro, primary CD3+ T cells were sorted from rat spleens and human peripheral blood, and co-cultured with or without 5-fluorouracil (5-FU) (active agent of CAP). The levels of ferroptosis-related proteins, ferrous ion concentration, and oxidative stress-related indicators were observed. The changes in mito-chondrial structure were observed using electron microscopy.RESULTS:With no significant myelotoxicity, metronomic CAP alleviated graft injury (Banff score 9 vs 7.333, P < 0.001), prolonged the survival time of the recipient rats (11.5 d vs 16 d, P < 0.01), and reduced the infiltration rate of CD3+ T cells in peripheral blood (6.859 vs 3.735, P < 0.001), liver graft (7.459 vs 3.432, P < 0.001), and spleen (26.92 vs 12.9, P < 0.001), thereby inhibiting acute rejection after LT. In vitro, 5-FU, an end product of CAP metabolism, induced the degradation of the ferritin heavy chain by upregulating nuclear receptor coactivator 4, which caused the accumulation of ferrous ions. It also inhibited nuclear erythroid 2 p45-related factor 2, heme oxygenase-1, and glutathione peroxidase 4, eventually leading to oxidative damage and ferroptosis of T cells.CONCLUSION:Metronomic CAP can suppress acute allograft rejection in rats by triggering CD3+ T cell ferroptosis, which makes it an effective immunosuppressive agent after LT.

Supplementary Figure S2. Analysis of activation of MAPKs in macrophages treated with cellular debris.

Objective This study was conducted to examine the association between the A118G polymorphism of the OPRM1 gene and the risk of increased VAS scores in patients with colorectal cancer who underwent laparoscopic radical resection for which fentanyl was used. Methods The OPRM1 A118G genotype in subjects were detected. The relationship between the A118G polymorphism of the OPRM1 gene and increased Visual Analogue Scale (VAS) scores throughout the perioperative period was explored. A total of 101 patients receiving fentanyl anesthesia undergoing laparoscopic radical resection of colon tumors at Zhongshan Hospital, Fudan University between July 2018 and December 2020 were investigated in the present study. The relative risk between the A118G polymorphism of the OPRM1 gene and VAS ≥ 4 in the PACU was estimated using the adjusted effect relationship diagram, baseline characteristic analysis, and multiple logistic regression analysis. The relationship between the A118G polymorphism of the OPRM1 gene and VAS in the PACU, as well as perioperative fentanyl usage, was examined after confounders were adjusted. Results Subjects with OPRM1 A118G wild gene A were less sensitive to fentanyl, which was a risk factor for PACU VAS ≥ 4. Before the model was adjusted, the odds ratio (OR) was 14.73 ( P = 0.001). After adjusting for age, sex, weight, height, and the duration of surgery, the OR increased to 16.55 ( P = 0.001). When adjusting for age, sex, weight, height, surgery duration, COMT Val158Met gene polymorphism, CYP3A4 *1G gene polymorphism, and CYP3A5 *3gene polymorphism, the OR was 19.94 ( P = 0.002). Moreover, OPRM1 A118G wild type gene A was found to be a risk factor for increased dosage of fentanyl in the PACU. Before the model was adjusted, the OR reached 16.90 ( P = 0.0132). After adjusting for age, sex, body weight, intraoperative fentanyl dosage, surgery duration, and height, the OR was 13.81, ( P = 0.0438). When adjusting for age, sex, weight, height, intraoperative fentanyl dosage, surgery duration, COMT Val158Met gene polymorphism, CYP3A4 *1G gene polymorphism, and CYP3A5 *3 gene polymorphism, the OR reached 15.23, ( P = 0.0205). Conclusion The A118G polymorphism of the OPRM1 gene carrying wild gene A was a risk factor for VAS ≥ 4 in the PACU. Moreover, it is a risk factor for increased dosage of fentanyl in the PACU.

BACKGROUND:This study aims to construct and verify a nomogram model for microvascular invasion (MVI) based on hepatocellular carcinoma (HCC) tumor characteristics and differential protein expressions, and explore the clinical application value of the prediction model.METHODS:The clinicopathological data of 200 HCC patients were collected and randomly divided into training set and validation set according to the ratio of 7:3. The correlation between MVI occurrence and primary disease, age, gender, tumor size, tumor stage, and immunohistochemical characteristics of 13 proteins, including GPC3, CK19 and vimentin, were statistically analyzed. Univariate and multivariate analyzes identified risk factors and independent risk factors, respectively. A nomogram model that can be used to predict the presence of MVI was subsequently constructed. Then, receiver operating characteristic (ROC) curve, calibration curve, and decision curve analysis (DCA) were conducted to assess the performance of the model.RESULTS:Multivariate logistic regression analysis indicated that tumor size, GPC3, P53, RRM1, BRCA1, and ARG were independent risk factors for MVI. A nomogram was constructed based on the above six predictors. ROC curve, calibration, and DCA analysis demonstrated the good performance and the clinical application potential of the nomogram model.CONCLUSIONS:The predictive model constructed based on the clinical characteristics of HCC tumors and differential protein expression patterns could be helpful to improve the accuracy of MVI diagnosis in HCC patients.

Supplementary Methods. Description of additional methods and procedures used in the study.