BACKGROUND AND AIMS:Liver sinusoidal endothelial cells (LSECs) are crucial nonparenchymal cells involved in liver fibrosis progression. Liver fibrosis develops from various chronic liver diseases, with pathological progression associated with the spleen. However, the precise mechanism by which the spleen influences LSECs during fibrosis is not fully understood. This study investigated the spleen's effect on LSEC angiogenesis and its underlying mechanisms using animal and cellular experiments. METHODS:Splenectomy or sham operations were performed on carbon tetrachloride (CCl4)-induced liver fibrosis mice and oil-treated control mice. Intraoperative spleen serum, peripheral serum from hepatitis B cirrhosis patients pre- and post-splenectomy, and normal human peripheral serum were collected and used to treat cultured LSECs. LSEC proliferation, migration, and tubule formation were then assessed. Active serum cytokines were identified by cytokine array analysis, and activated signaling pathways in LSECs were investigated through functional experiments. RESULTS:The spleen promoted LSEC angiogenesis in fibrotic mice. Compared to peripheral serum, spleen serum from cirrhotic patients significantly enhanced LSEC proliferation, migration, and tubule formation. Growth differentiation factor 15 (GDF15) was elevated in spleen serum and promoted LSEC angiogenesis. In vitro, GDF15 induced Smad2/3 phosphorylation in LSECs. Furthermore, inhibition of Smad3 phosphorylation attenuated the pro-angiogenic effects of spleen serum or GDF15 on LSECs. CONCLUSION:The spleen promotes LSEC angiogenesis by activating the TGFβ/Smad2/3 pathway through GDF15 in serum.

Introduction: Icaritin is a bioactive flavonol isolated from the Chinese medicinal herb Epimedium. The comprehensive understanding of antifibrotic effects and associated molecular mechanisms of icaritin remains incomplete. This study aims to explore the protective effects of icaritin against liver fibrosis and to further elucidate the mechanisms involved. Methods: Human hepatic stellate LX-2 cells stimulated with TGF-β1 and a carbon tetrachloride (CCl4)-induced liver fibrosis mouse model were employed. in vitro assays were carried out to evaluate collagen type I (COl I) and α-smooth muscle actin (α-SMA) expression, while in vivo studies assessed fibrosis alleviation. Molecular mechanisms were explored via analysis of TGF-β1, phosphorylated Smad2/3, and HIF-1α protein levels using Western blotting. Results: Icaritin suppressed TGF-β1-induced COl I and α-SMA expression in LX-2 cells and ameliorated liver fibrosis in CCl4-treated mice. Mechanistically, it significantly reduced TGF-β1 levels, inhibited Smad2/3 phosphorylation, and downregulated HIF-1α protein expression in LX-2 cells. Conclusion: Icaritin attenuated experimental liver fibrosis through the inhibition of the TGF-β/Smad and HIF-1α signaling pathways, highlighting its therapeutic potential for fibrotic liver diseases.

Background: This study aimed to evaluate the impact of intraosseous (IO) access on inflammatory mediators, hematopoietic cell function, and coagulation-metabolic disturbances in patients presenting with emergency traumatic hemorrhagic shock (THS), thereby providing clinical evidence to refine IO resuscitation protocols in emergency settings. Methods: We conducted a randomized controlled trial involving 84 THS patients admitted between February 2024 and February 2025. Participants were allocated equally into two groups: the IO group (n= 42), where vascular access was established via humeral or proximal tibial puncture, and the intravenous (IV) group (n= 42), where conventional peripheral or central venous access was prioritized. Serial measurements were performed at baseline (T0), 24 hours (T1), and 72 hours (T2) post-intervention to assess: (1) inflammatory mediators (IL-1 b, IL-6, IL-10, HMGB1, MDA); (2) hematopoietic parameters (CD34+ cell proportion, CFU-GM /BFU-E colony formation, CXCL12, EPO, and TPO ); (3) coagulation profiles (PT, APTT, and D-dimer); and (4) tissue perfusion indicators (blood lactate and lactate clearance rate). Comparative analyses were conducted both between groups and across different time points. Results: The IO group demonstrated significantly elevated levels of IL-1P, HMGB1, and MDA at T1 and T2 compared to the IV group (P< 0.05), coupled with reduced IL-10 expression (P< 0.05), indicating exacerbated inflammatory imbalance and oxidative stress. Hematopoietic evaluation revealed progressive declines in CD34+ cell populations, CFU-GM /BFU-E colony formation, and CXCL12 concentration in the IO group at T1 and T2 (P< 0.05), despite modest compensatory increases in EPO and TPO that remained inferior to the IV group (P< 0.05). Coagulation studies showed prolonged PT/APTT (P< 0.01) and higher D-dimer levels (P< 0.05) in the IO group, along with worse blood lactate levels and lactate clearance rates compared to the IV group (P< 0.05), suggesting increased tissue hypoxia and coagulopathy risk. Conclusions: While IO access enables rapid vascular access for resuscitation and reduces critical intervention time, despite its procedural efficiency in rapid vascular access for resuscitation, IO may inadvertently aggravate systemic inflammatory dysregulation, impair hematopoietic function, and worsen coagulation-metabolic disturbances through mechanisms such as mechanical stimulation, hypothermic fluid infusion, and oxidative stress.

Our study focused on investigating the bioeffects of extremely low-frequency electromagnetic fields (ELF-EMFs) on the immune function of the spleen. We designed an electromagnetic instrument that can locally target on spleen, the spleens of mice were locally exposed to the ELF-EMF (50 Hz, 30 mT) for 14 days (4 h/day). Parallelly, the isolated splenic T cells were exposed to ELF-EMF (50 Hz, 15 mT) for 2 h. After the exposure, the splenocyte showed a reduced apoptosis rate. Among the splenocytes, the CD4+ T cells and natural killer cells accumulated, the percentage of B cells decreased. In vitro study demonstrated that ELF-EMF induced the alteration of T cell subsets, showing an increased percentage of CD4+ T cells and a decreased percentage of CD8+ T cells. Within CD4+ T cells, the population of T helper (Th) 17 cells increased, and the population of regulatory T cells (Treg) cells decreased. The enrichment of the nuclear factor (NF)-κB pathway in the splenic T cells was found to be reduced after exposure to ELF-EMF. Our findings suggest that ELF-EMF regulated the immune function of the spleen by changing the proportion of immune cells in the spleen. Specifically, the differentiation of spleen T cells was induced by ELF-EMF toward Th17 cells and inhibited by ELF-EMF into Treg cells. The NF-κB signaling pathway probably accounts for the effects of ELF-EMF on the spleen T cells.

Background:Splenic immunomodulation triggered by ultrasound shows a significant anti-inflammatory effect against various inflammatory diseases, whose mechanism is mainly attributable to the activation of cholinergic anti-inflammatory pathway (CAP). However, the potential role and underlying mechanism of splenic ultrasound stimulation in cancer management have been rarely reported and superficially defined. Methods:Following optimization of ultrasonic parameters, this study evaluated the anti-tumor efficacy of splenic sonication across multiple tumor models (eg, orthotopic H22 hepatocellular carcinoma (HCC), orthotopic Hepa1-6 HCC, and subcutaneous 4T1 breast cancer), and applied flow cytometry to quantify dynamic alterations in immune cell populations. Furthermore, in orthotopic H22 HCC models, this study employed fluorescence-activated cell sorting, RNA sequencing, splenic nerve blockade via absolute ethanol ablation, and in vitro Ca²⁺ flux assays to delineate the mechanisms underlying ultrasound-mediated splenic anti-tumor immunity. Results:This study first assessed the therapeutic effect of focused ultrasound precisely targeting the spleen (FUS sti. spleen) on various tumors at specific ultrasonic doses. It fully demonstrated that FUS directly stimulated splenic immune cell proliferation and activation (especially NK and CD8 T cells) rather than CAP excitation to modulate splenic immune function. Particularly, NK cells are much more indispensable and important in responding to FUS stimulation for cancer suppression than CD8 T cells. RNA sequencing of NK and CD8 T cells, as well as in vitro experiments revealed that FUS firstly regulated calcium-related signaling pathways to further modulate others, such as PI3K-AKT, Rap1, and Hippo pathways to promote immune cell proliferation, migration and activation to suppress cancer cell deterioration. Particularly, FUS sti. spleen and FUS intervention on the tumor synergistically induced the best tumor suppression than each of the two taken individually. Conclusion:FUS sti. spleen facilitated immunocyte proliferation and activation through altering calcium-dependent signaling rather than CAP excitation to modulate anti-tumor immunity, indicating substantial clinical translation potential.

BACKGROUND:Tumor-associated macrophages (TAMs) are the dominant myeloid population in hepatocellular carcinoma (HCC) and critically shape the chemokine milieu that governs CD8+ T-cell entry. However, how macrophage-intrinsic regulators of chemokine expression influence antitumor immunity and response to immunotherapy remains unclear. We postulated that the macrophage RNA helicase DEAH-box helicase 34 (DHX34) suppresses production of the chemokine fractalkine (CX3CL1), thereby limiting chemotaxis of CX3CR1+ CD8+ T cells and diminishing the therapeutic efficacy of programmed cell death protein-1 (PD-1) blockade. We therefore examined the impact of macrophage DHX34 on T-cell trafficking and tumor control. METHODS:We profiled DHX34 expression and subcellular distribution across human single-cell RNA-sequencing datasets and confirmed its pattern in murine TAMs. To investigate its function, we implemented myeloid-restricted Dhx34 deletion and evaluated tumor growth, intratumoral CD8+ T-cell abundance, and the efficacy of PD-1 blockade. Mechanistic experiments were performed using bone marrow-derived macrophages treated with tumor culture supernatant. CX3CL1 expression was measured by qPCR, ELISA, and immunoblotting, and CD8+ T-cell migration was assayed using Transwell assays. RESULTS:DHX34 was enriched in TAMs, and higher DHX34 abundance correlated with lower CD8+ T-cell numbers and faster tumor growth. Myeloid-restricted Dhx34 deletion increased CX3CL1 expression and release in TAMs, thereby augmenting the influx of CX3CR1+ CD8+ T cells without altering their proliferation or viability. Consequently, DHX34 deficiency elevated intratumoral CD8+ T cell levels, slowed tumor expansion, and increased susceptibility to PD-1 blockade. CONCLUSIONS:Within tumor-associated macrophages, DHX34 suppressed CX3CL1 output, limiting the tumor entry of CX3CR1-positive CD8+ T cells and, in turn, impairing control of HCC and reducing responsiveness to checkpoint therapy. Targeting DHX34 may potentiate PD-1 blockade in HCC.

Background: Hemorrhagic shock (HS) is a severe condition involving stress proteins, inflammation, and gut microbiota dysbiosis. Understanding whether regulatory proteins influence HS through microbial pathways is crucial for improving therapeutic strategies. Methods: We used Mendelian randomization (MR) combined with animal experiments to investigate the role of regulatory proteins in HS. Two-sample MR was performed to assess the impact of various stress-related proteins. Additionally, 16 S rRNA sequencing was conducted in a rat HS model to analyze gut microbiota diversity and composition at baseline, 24 h, and 72 h after hemorrhage. Results: Two-sample MR identified HSPB1 and HIF1A as protective proteins, while APAF1, F7, and F10 increased susceptibility to HS. In the rat model, microbiota alpha diversity decreased at 24 h but partially recovered by 72 h, with significant shifts in beta diversity. Genus-level analysis revealed transient expansion of Lactobacillus, followed by dominance of Blautia and Romboutsia. Stage-specific predictions from PICRUSt2 suggested enrichment of amino acid metabolism and protein synthesis, particularly at 72 h, implicating microbial regulation in cellular recovery and stress adaptation. Conclusions: Our findings support a "protein-microbiota-HS" regulatory framework, highlighting the gut microbiota as key mediators of host stress responses. This integrative approach provides mechanistic insights into HS pathogenesis and suggests potential microbiome-targeted therapeutic strategies. We propose that targeting specific microbial communities, such as Blautia and Lactobacillus, could enhance recovery from HS.

Abstract The spleen, as the largest immune organ, plays a pivotal role in modulating immune responses, particularly in the context of carcinogenesis and tumor progression. Non-pharmacological manipulation, particularly splenic ultrasound stimulation (SUS), has demonstrated significant immunomodulatory efficacy in alleviating chronic inflammatory diseases, suggesting its potential to revitalize splenic immunocompetence suppressing tumor proliferation, yet remains underexplored. This study applied low-frequency pulsed focused ultrasound (FUS) noninvasively stimulating the spleen (FUS sti. spleen) to investigate the efficacy in enhancing antitumor immunity and suppressing hepatocellular carcinoma (HCC). The results showed that FUS sti. spleen significantly suppressed tumor proliferation, achieving a suppression rate of >70% for H22-HCC and >83% for Hepa1-6-HCC, along with significantly prolonged survival. Comprehensive flow cytometry, single-cell RNA sequencing (scRNA-seq) and cytokine analyses demonstrated that SUS profoundly reshaped the splenic and intratumoral immune landscape, specifically activating cytotoxic CD8+ T cells and NK cells while suppressing immunosuppressive cell populations. Mechanistically, FUS facilitated calcium influx in splenic immunocytes, activating multiple signaling pathways, such as TNF, NFκB, MAPK, HIF-1, and ErbB, thereby counteracting tumor-driven immunosuppressive polarization while potentiating robust immune activation that impedes malignant progression and neoplastic proliferation. Leveraging above insights, we developed spleen-targeted nanodroplets encapsulating bioavailable calcium ions (STNDs@Ca²⁺), which, upon FUS stimulation, undergo cavitation-mediated controlled release of Ca²⁺, further amplifying immunocyte activation and tumor suppression, achieving a remarkable H22-HCC suppression rate of over 90%. This study highlights the therapeutic potential of ultrasound-mediated splenic immunomodulation, both as a standalone intervention and in synergy with STNDs@Ca²⁺, as a novel and noninvasive strategy for cancer immunotherapy.

Tumors often evade immune surveillance by crippling their immunogenicity in the microenvironment. DHX34, an RNA helicase involved in nonsense-mediated mRNA decay pathway, is critical for aberrant RNA degradation. However, the effect of DHX34 in regulating the immunogenicity in hepatocellular carcinoma (HCC) is still unclear. Here, a surprising function of DHX34 in inhibited HCC immunogenicity is identified. DHX34-deficient tumors were infiltrated by activated T cells that impaired tumor growth and enhanced survival in mice. Mechanistically, DHX34 depletion triggered dsRNA accumulation which may activate cytosolic RNA-sensing pathway effectors such as MAVS, p-IKK, p-IRF3, and the subsequent type-I interferon response, evoking tumor-intrinsic immunity and leading to CD8 T activation. Collectively, DHX34 is implicated as a regulator that orchestrates a barrier in HCC by suppressing dsRNA-driven innate immune activation. Targeting DHX34 may enhance tumor immunogenicity and synergize with immunotherapies, offering a novel therapeutic strategy for HCC.

Protein tyrosine phosphatase mitochondrial 1 (PTPMT1), is a member of the protein tyrosine phosphatase superfamily localized on the mitochondrial inner membrane, and regulates the biosynthesis of cardiolipin. Given the important position of PTPMT1 in mitochondrial function and metabolism, pharmacological targeting of PTPMT1 is considered a promising manner in disease treatments. In this study, we mainly investigated the role of PTPMT1 in hepatocellular carcinoma (HCC) ferroptosis, a new type of cell death accompanied by significant iron accumulation and lipid peroxidation. Herein, the pharmacological inhibition of PTPMT1 was induced by alexidine dihydrochloride (AD, a dibiguanide compound). Human HCC cell lines with PTPMT1 knockout and PTPMT1 overexpression were established using CRISPR/Cas9 and lentiviral transduction methods, respectively. The position of PTPMT1 in regulating HCC ferroptosis was evaluated in vitro and in vivo. Our results indicated that pharmacological inhibition of PTPMT1, facilitated by AD treatment, heightens the susceptibility of HCC to cystine deprivation-ferroptosis, and AD treatment promoted the conversion from ferritin-bound Fe3+ to free Fe2+, which contributed to the labile iron pool in cytoplasm. Meanwhile, pharmacological inhibition of PTPMT1 also induced the formation of both swollen mitochondria and donut mitochondria, and enhanced the metabolism process form succinate to fumarate in mitochondrial tricarboxylic acid (TCA) cycle, which increased the sensitivity of HCC cells to cystine deprivation-induced ferroptosis. In total, our work reveals the close association of PTPMT1 with cysteine deprivation-induced ferroptosis, providing a novel insight into chemotherapy strategies against human HCC.

Purpose:DDX11 antisense RNA 1 (DDX11-AS1) has been recognized for its strong correlation with hepatocellular carcinoma (HCC). Nevertheless, the exact biological functions and fundamental molecular processes of DDX11-AS1 in HCC require further in-depth investigation. Methods:A comprehensive bioinformatics analysis was carried out to explore the expression of DDX11-AS1 and its clinical implication in HCC utilizing the TCGA data. qRT-PCR was employed to validate the expression of DDX11-AS1 in HCC tissues/cell lines. RNA fluorescence in situ hybridization (RNA-FISH) was used to observe the subcellular localization of DDX11-AS1 in HCC cells. Loss-of-function experiments, both in vitro and in vivo, were executed to elucidate the biological functions of DDX11-AS1 in HCC. RNA sequencing (RNA-seq) was employed to identify genes and signaling pathways potentially regulated by DDX11-AS1. Rescue experiments were conducted to validate that carbonic anhydrase IX (CA9) mediates DDX11-AS1 promoting HCC progression. The influence of nuclear respiratory factor 1 (NRF1) on the transcription of DDX11-AS1 was investigated through dual-luciferase reporter assays and ChIP-qPCR. Results:The increased expression of DDX11-AS1 is positively associated with several aggressive clinical characteristics (pathologic T stage, histologic grade, AFP level, and vascular invasion), and is closely linked to unfavorable outcomes in HCC patients, acting as a separate hazardous factor for overall survival. DDX11-AS1 is predominantly situated in the nucleus of HCC cells. DDX11-AS1 knockdown impeded the growth, migration, and invasion capabilities of HCC cells in vitro, and reduced the tumor enlargement in a subcutaneous mouse model. RNA-Seq unveiled that silencing DDX11-AS1 lessened the expression of CA9 and suppressed the activity of the MEK/ERK signaling cascade in HCC cells. Rescue experiments uncovered that CA9 acts as a downstream target facilitating the cancer-causing roles of DDX11-AS1 in HCC. Furthermore, DDX11-AS1 was revealed to be transcriptionally regulated by NRF1. Conclusion:DDX11-AS1, a NRF1-induced lncRNA, facilitates HCC development by upregulating CA9 expression and activating the MEK/ERK signaling cascade.

Background and Aims Macrophages play plastic roles during fibrogenesis and fibrosis regression. Phagocytosis is considered a trigger for shifting macrophages from a profibrotic phenotype to a restorative phenotype. However, the underlying mechanism by which macrophages enhance phagocytosis remains unclear. Our present study investigated the role of reactive oxygen species (ROS)-modulated TRPM2 activation in this process.Methods The changes of TRPM2 expression, ROS intensity, and macrophage phagocytosis were assessed in fibrogenesis and fibrosis regression models. RNA sequencing was utilised to reveal pathway enrichment caused by TRPM2, and the role of TRPM2 in enhancing phagocytosis was verified. The coordinate regulation of ROS-TRPM2 in different functions of macrophages was demonstrated by modulating ROS intensity and TRPM2 expression. Mitochondrial dynamics changes induced by ROS-stimulated TRPM2 activation were evaluated by analysing the expression of dynamics-related molecules and mitochondrial imaging, and intervention in mitochondrial dynamics confirmed their impact on macrophage phagocytosis.Results Low-intensity ROS stimulation up-regulated the expression of TRPM2 and coordinately enhanced macrophage phagocytosis and the expression of matrix degradation-related proteins (MMPs), a process akin to fibrosis regression. However, high-intensity ROS inclined macrophages to produce more profibrotic cytokines, associating with oxidative stress caused by liver injury. ROS-mediated TRPM2 activation mobilised Ca2+ and promoted mitochondrial fission; either inhibiting mitochondrial fission or chelating Ca2+ counteracted phagocytosis, as well as decreasing MMPs.Conclusions ROS-TRPM2 coordinately regulate macrophage functions. During the liver fibrosis regression period, ROS-induced activation of TRPM2 helps enhance macrophage phagocytosis and switches them to a restorative phenotype. Modulating this process may provide means for developing effective therapeutic strategies.

Gaudichaudione H (GH) is a naturally occurring small molecular compound derived from Garcinia oligantha Merr. (Clusiaceae), but the full pharmacological functions remain unclear. Herein, the potential of GH in disulfidptosis regulation, a novel form of programmed cell death induced by disulfide stress is explored. The omics results indicated that NRF2 signaling could be significantly activated by GH. The potential targets are associated with hepatocarcinogenesis and cell death. Moreover, both glutathione (GSH) metabolism and NADP+-NADPH metabolism are affected by GH, indicating the potential in disulfidptosis regulation. It is also observed that GH enhanced the sensitivity of hepatocellular carcinoma (HCC) cells to disulfidptosis, which is dependent on the activation of NRF2-SLC7A11 pathway. GH significantly increased the levels of NRF2 and promoted the transcription of NRF2 target gene, SLC7A11, through autophagy-mediated non-canonical mechanism. Under the condition of glucose starvation, GH-induced upregulation of SLC7A11 aggravated uptake of cysteine, disturbance of GSH synthesis, depletion of NADPH, and accumulation of disulfide molecules, ultimately leading to the formation of disulfide bonds between different cytoskeleton proteins and disulfidptosis eventually. Collectively, the findings underscore the potential role of GH in promoting cancer cell disulfidptosis, thereby offering a promising avenue for the treatment of drug-resistant HCC in clinical settings.

The spleen, the largest lymphatic organ, comprises a diverse array of immunocytes in approximately one quarter of the body, including T cells, B cells, natural killer cells, and myeloid cells (such as dendritic cells, neutrophils, myeloid-derived suppressor cells, and macrophages). These immune cells undergo dynamic transitions and mobilization, enabling the spleen to execute a wide range of immunological functions. The spleen's structural organization and multicellular composition, along with its reservoir of lymphocytes, facilitate the capture and clearance of blood-borne antigens while also orchestrating both innate and adaptive immune responses. Additionally, the spleen plays critical roles in hematopoiesis and the removal of aged or damaged red blood cells. Despite being innervated by sympathetic (catecholaminergic) nerve fibers, the spleen lacks parasympathetic (vagal or cholinergic) innervation. The neuroimmune axis, particularly the interplay between sympathetic and parasympathetic nervous system immune circuits, significantly influences disease onset and progression. Extensive research employing physical, genetic, and pharmacological approaches has sought to directly modulate splenic immunocytes and activate neuroimmune interactions to restore immune homeostasis and counteract disease. Two primary mechanisms underlie these immunomodulatory interventions: (1) the cholinergic anti-inflammatory pathway, wherein norepinephrine released by splenic catecholaminergic fibers binds to β2-adrenergic receptors on CD4⁺ T cells, triggering acetylcholine secretion, which in turn suppresses inflammatory cytokine production in macrophages via α7 nicotinic acetylcholine receptor signaling, and (2) direct immunomodulation of splenic immunocytes, which regulates key genes and signaling pathways, alters cytokine secretion, and modulates ion flux to influence cellular functions. Among various therapeutic strategies, physical methods, particularly electrical stimulation and splenic ultrasound stimulation, have demonstrated the greatest promise for clinical applications in splenic immunomodulation and disease management.

Background Portal vein system thrombosis (PVST) is a common and potentially life-threatening complication following splenectomy plus pericardial devascularisation (SPDV) in patients with cirrhosis and portal hypertension. Early prediction of PVST is critical for timely intervention. This study aimed to develop a machine learning-based prediction model for PVST occurrence within 3 months after splenectomy.Methods 392 patients with cirrhosis who underwent splenectomy at the Second Affiliated Hospital of Xi’an Jiaotong University between 1 July 2016 and 31 December 2022 were enrolled in this study and followed up for 3 months. The predictive model integrated 37 candidate predictors based on accessible clinical data, including demographic characteristics, disease features, imaging results, laboratory values, perioperative details and postoperative prophylactic therapies, and finally, eight predictors were selected for model construction. The five machine learning algorithms (logistic regression, Gaussian Naive Bayes, decision tree, random forest and AdaBoost) were employed to train the predictive models for assessing risks of PVST, which were validated using five fold cross-validation. Model discrimination and calibration were estimated using receiver operating characteristic curves(ROC), accuracy, sensitivity, specificity, positive predictive value, negative predictive value and Brier scores. The outcome of the predictive model was interpreted using SHapley Additive exPlanations (SHAP), which provided insights into the factors influencing PVST risk prediction.Results During the 3-month follow-up, a total of 144 (36.73%) patients developed PVST. The AdaBoost model demonstrated the highest discriminative ability, with a mean area under the receiver operating characteristic curve (AUROC) of 0.72 (95% CI 0.60 to 0.84). Important features for predicting PVST included albumin, platelet addition, the diameter of the portal vein, γ-glutamyl transferase, length of stay, activated partial thromboplastin time, D-dimer level and history of preoperative gastrointestinal bleeding, as revealed by SHAP analysis.Conclusions The machine learning-based prediction models can provide an initial assessment of 3-month PVST risk after SPDV in patients with cirrhosis and portal hypertension. The AdaBoost model demonstrates moderate discriminative ability in distinguishing between high-risk and low-risk patients, with an AUROC of 0.72 (95% CI 0.60 to 0.84). By incorporating SHAP analysis, the model can offer transparent explanations for personalised risk predictions, facilitating targeted preventive interventions and reducing excessive interventions across the entire patient population.

Macrophage plasticity, critical for immune response, is often dysregulated in various infectious and inflammatory diseases. While ion channels have been implicated in immune cell modulation, how they influence macrophage polarization remains poorly understood. Here, it is demonstrated that ectosomes carrying the ion channel Calhm6 effectively suppress severe inflammation triggered by LPS. These Calhm6-bearing ectosomes, secreted by macrophages, facilitate M2-like polarization, elicit an anti-inflammatory response, and foster immune tolerance. Conversely, Calhm6 deficiency leads to suppressed Creb1 activity, which in turn augments M1-like macrophage polarization, enhancing bactericidal activity and the secretion of pro-inflammatory cytokines. Mechanistically, Chp1 serves as a scaffold protein and undergoes phosphorylation by CaMK4. This phosphorylation enhances the localization of the Calhm6-Chp1-CaMK4 complex to the cell membrane, promoting Creb1 activation and M2-like macrophage polarization calcium-dependently. Moreover, the M1-like polarization inducers LPS and IFNγ enhance the binding of Irf1 to the Calhm6 promoter, upregulating its expression and stimulating ectosome formation. Conversely, Stat6, activated by IL-4, competes with Irf1 for binding to the Calhm6 promoter, thereby suppressing its expression. In summary, our findings unravel the intricate interplay between ion channels, ectosomes, and macrophage polarization, revealing that ectosomal-Calhm6 can serve as a novel therapeutic agent to modulate inflammatory responses and facilitate tissue repair.