Congenital hepatic fibrosis (CHF) is a rare inherited liver disorder caused by mutations in the polycystic kidney and hepatic disease-1 gene (PKHD1), leading to progressive hepatic fibrosis. In our study, single-cell RNA sequencing of a CHF patient carrying compound heterozygous PKHD1 mutations revealed markedly intercellular communication between cholangiocytes and hepatic stellate cells (HSCs) via the secreted phosphoprotein 1 (SPP1)-integrin α8β1 axis. The CRISPR-Cas9-generated Pkhd1 knockout mouse model (Pkhd1del3-4/del3-4) was generated and recapitulated bile duct hyperplasia and fibrosis. We validated that PKHD1 deficiency led to progressive upregulation of SPP1 in cholangiocytes. Co-culture experiments demonstrated that PKHD1-deficient cholangiocytes secrete OPN, activating PI3K/Akt signaling in HSCs and promoting their activation. Mechanistically, phosphorylated STAT3 (p-STAT3) was identified as a direct transcriptional activator of SPP1. Importantly, cholangiocyte-specific knockdown of Spp1 via AAV8 significantly attenuated hepatic fibrosis in vivo. Collectively, our findings demonstrate that reduced PKHD1 expression in cholangiocytes promotes hepatic fibrosis by activating the p-STAT3-SPP1 pathway, and targeting this axis may offer a potential therapeutic strategy.

The quest for a therapy that can precisely seek out and destroy hepatocellular carcinoma (HCC) has long been hindered by poor targeting and off-target toxicity. Therefore, we present an innovative dual-targeting nanotherapeutic strategy that leverages a single-wavelength NIR-II laser to concurrently activate both photodynamic and photothermal therapy for precise and synergistic HCC ablation. The core of this strategy lies in the conjugated polymer nanoparticles (CPNPs), which uniquely synchronize the generation of reactive oxygen species (ROS) with hyperthermia induction under NIR-II laser, enabling deep-tissue penetration and synergistic tumor eradication. By further conjugating a clinically validated targeting antibody, we constructed the smart nanocomplex (BUD31@CPNPs). These nanocomposites achieve a breakthrough in subcellular targeting by coordinately directing their action to both mitochondria and the nucleus. This feat effectively bypasses lysosomal sequestration and overcomes the drug delivery bottleneck of single-targeted systems. Remarkably, BUD31@CPNPs exhibit exceptional biocompatibility and potent tumor suppression in vivo, heralding a transformative approach to HCC treatment with high clinical translation potential.

OBJECTIVE:To develop and validate a model for early risk stratification of secondary hemophagocytic lymphohistiocytosis (HLH) in patients with severe fever with thrombocytopenia syndrome (SFTS). METHODS:This retrospective cohort included adults with laboratory-confirmed SFTS admitted to The First Affiliated Hospital with Nanjing Medical University between January 2019 and July 2024. Predictor variables were derived from clinical and laboratory data obtained within 3 days after virologic confirmation, corresponding to the predefined early-evaluation window of the study. HLH status (binary outcome) was defined using the entire-course HScore (≥170), calculated from the worst available values over the clinical course; HLH-2004 criteria and early-window HScore distributions were summarized descriptively to provide transparent outcome accounting. Twenty-eight candidate predictors entered LASSO with Boruta refinement, and retained variables were used to construct a multivariable logistic model and nomogram. Model performance was evaluated by discrimination, calibration, and decision-curve analysis in the derivation cohort, an internal validation set, and an external cohort of 60 patients from The First Affiliated Hospital with Anhui Medical University. RESULTS:Among 249 patients (152 non-HLH, 97 HLH), HLH was associated with higher peak temperature, longer fever, more lymphadenopathy/splenomegaly and neurological symptoms, and more severe thrombocytopenia, hypertriglyceridemia, hypofibrinogenemia, hyperferritinemia, higher viral load, elevated muscle/liver enzymes and LDH, and coagulopathy (all p < 0.05). LASSO-Boruta identified six routinely available predictors-peak temperature, splenomegaly, fever duration, triglycerides, fibrinogen, and ferritin. The model showed LR χ² = 214.82 (p < 0.0001), R² = 0.784, C-index = 0.962, Dxy = 0.923, with near-perfect calibration in derivation. In internal validation, discrimination remained near-perfect (AUC 0.997, 95% CI 0.989-1.000); mild miscalibration was corrected by intercept-and-slope recalibration, and decision curves showed net benefit across wide thresholds. External validation (n = 60) confirmed excellent discrimination (AUC 0.907, 95% CI 0.835-0.980), slight miscalibration resolved by recalibration, and preserved net benefit across most thresholds. CONCLUSIONS:A simple model based on early clinical and laboratory variables supports risk stratification for HLH in SFTS and may facilitate closer monitoring, repeated HLH assessment, and timely individualized management.

Racial and ethnic disparities in metabolic dysfunction-associated steatotic liver disease (MASLD) persist in the US population. We studied the impact of social determinants of health (SDOH) on the likelihood of developing MASLD and explored the contribution of SDOH to racial disparities in MASLD. Data on self-reported SDOH domains and sub-items based on Healthy People 2030 were collected from the US National Health and Nutrition Examination Survey (NHANES) across 10 NHANES cycles from 1999 to 2000 to 2017–2018. The participants were divided into four racial and ethnic groups: White, Black, Hispanic, and other. The primary outcome was MASLD, characterized by a US fatty liver index score exceeding 30. Survey-weighted logistic regression models were used to examine the relationship between SDOH and MASLD. The results were further analysed by race and ethnicity. A total of 22,585 participants (10115 White, 4431 Black, 5978 Hispanic, and 2061 other) were included in our analyses, of whom 7798 had MASLD (survey-weighted prevalence 32.0

Epigallocatechin gallate (EGCG) has shown antiviral potential against the hepatitis B virus (HBV) by restoring lysosomal acidification; however, its application is limited by poor stability and low intestinal permeability. Here, a dual-shell chitosan/ferritin nanosystem (CHE) was developed to enhance the EGCG delivery and functionality. CHE achieved an encapsulation efficiency of 13.61% with a uniform nanoscale size distribution. Compared with free EGCG and single-shell HE, CHE improved storage stability and delayed EGCG release under simulated gastrointestinal conditions. In a Caco-2/HepG2.2.15 or HepAD38 coculture model, CHE enhanced transepithelial transport and intracellular delivery while maintaining epithelial barrier integrity. Mechanistically, CHE promoted lysosomal acidification, increased cathepsin B maturation, and restored autophagic flux, accompanied by reductions in HBV DNA, pgRNA, HBsAg, and HBeAg levels. These findings demonstrate that a dual-shell nanocage integrates stability enhancement with functional intracellular modulation, providing a promising strategy to improve the antiviral efficacy of polyphenols.

Severe fever with thrombocytopenia syndrome virus (SFTSV), also known as Dabie bandavirus, is the pathogen causing severe fever with thrombocytopenia syndrome (SFTS). Human cell receptors are the first barriers to viral entry, proliferation, and host immune response. This study explored the association between single nucleotide polymorphisms (SNPs) in genes encoding receptors or related molecules that mediate SFTSV cell entry, such as dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN), DC-SIGN-related protein (DC-SIGNR), and non-muscle myosin heavy chain IIA (NMMHC-IIA), and SFTSV infection susceptibility in the Chinese Han population, as well as the underlying potential biological mechanisms. From September 2020 to December 2022, 454 subjects from the First Affiliated Hospital with Nanjing Medical University were enrolled and divided into SFTSV-infected and uninfected groups based on SFTSV RNA and antibody test results. TaqMan SNP genotyping was performed on DC-SIGN (rs7248637, rs4804800, and rs11465421), DC-SIGNR (rs2277998), and NMMHC-IIA (rs2269529 and rs2269530). Logistic regression was used to assess the association between the candidate SNPs and SFTSV infection susceptibility, and the online bioinformatics tools were employed to explore potential biological functions of positive SNPs. Logistic regression analysis showed that individuals carrying the DC-SIGNR rs2277998 AA genotype were more susceptible to SFTSV infection, and in silico bioinformatics analysis generated the hypothesis that this locus might be involved in influencing gene transcription and expression, which warrants further functional validation. DC-SIGNR rs2277998 is significantly associated with SFTSV infection susceptibility in the Chinese Han population. The exact regulatory mechanisms require further experimental validation.IMPORTANCESevere fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by severe fever with thrombocytopenia syndrome virus (SFTSV). Host genetic variation may modulate both disease susceptibility and severity by altering key functional proteins. Our study was the first to explore the association between single nucleotide polymorphisms (SNPs) in host genes encoding receptors or related molecules that mediate SFTSV cell entry, such as dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN), DC-SIGN-related protein (DC-SIGNR), and non-muscle myosin heavy chain IIA (NMMHC-IIA), and SFTSV infection susceptibility in the Chinese Han population. The results showed that the homozygous mutation of DC-SIGNR rs2277998 AA genotype was associated with a significantly higher risk of SFTSV infection in the Chinese Han population, with in silico predictions pointing to a potential, yet unverified, role in regulating the DC-SIGNR gene transcription and expression, which may provide a new scientific basis for SFTSV prevention and treatment.

The drug resistance to targeted therapy in patients with advanced hepatocellular carcinoma (HCC) is gradually increasing. Thus, it is very important to further examine the molecular signaling pathways related to HCC and the corresponding targeted therapy. The study identified a significant association between low TBX15 expression and sorafenib resistance in HCC. TBX15 increased sorafenib sensitivity in HCC cells by inducing ferroptosis in vivo and vitro experiments. Mechanistically, TBX15 promoted the secretion of mitochondrial DNA (mtDNA) into the cytoplasm through the opening of the mitochondrial permeability transition pore, then stimulated the cGAS/STING pathway due to the increased cytosolic mtDNA. TBX15 binds to the HERC5 promoter region, increasing cGAS ISGylation and enhancing its stability. Furthermore, the activation of cGAS/STING pathway increased ferroptosis in HCC cells. Our research underscores TBX15's potential to counteract sorafenib resistance and establishes the TBX15-cGAS/STING axis as a key regulator in HCC pathobiology.

Autoimmune hepatitis (AIH) is a chronic inflammatory liver disease typically managed with broad-spectrum immunosuppressants that carry significant systemic side effects and often provide incomplete efficacy. While gut-microbiota-derived metabolites are known to influence AIH progression, the specific microbial drivers that maintain hepatic immune homeostasis remain poorly defined. Here, we show that Bacteroides acidifaciens (BA) and its metabolite 1-oleoyl-sn-glycero-3-phosphoethanolamine (O-LysoPE) are enriched in self-healing mouse models of hepatitis but markedly depleted in AIH patients. We demonstrate that O-LysoPE induces a 'hepatocyte-driven active immunosuppression' by targeting the Qa-1b (HLA-E): NKG2A immune checkpoint. Mechanistically, O-LysoPE selectively redirects the transcription factor Creb1 to the H2T23 promoter under inflammatory conditions, thereby upregulating hepatocytic Qa-1b expression. This elevation of Qa-1b engages the inhibitory receptor NKG2A on T cells, suppressing their overactivation and restoring a quiescent phenotype. Genetic disruption of H2T23 abrogates the hepatoprotective effects of O-LysoPE, confirming the central role of this metabolic-immune axis. Our findings reveal that the BA-O-LysoPE axis mobilizes the liver's intrinsic self-rescue mechanisms to restore immune quiescence. This study establishes a robust biological framework for liver-specific, targeted immunotherapy in AIH, offering a precision alternative to current systemic immunosuppression.

Background The incidence of metabolic-associated steatotic liver disease in patients with chronic hepatitis B is increasing annually; however, the interaction between hepatitis B virus (HBV) infection and lipid metabolism remains unclear. This study attempted to clarify whether fatty acid metabolism regulation could alleviate mitochondrial dysfunction caused by HBV infection.Methods A public gene set of human livers was analyzed, and a proteomic analysis on mouse livers was conducted to explore metabolic disorders and affected organelles associated with HBV infection. The effect of decanoylcarnitine on fatty acid beta-oxidation and mitochondria was investigated in vivo and in vitro. The pathways involved were shown by proteomic analysis and confirmed by Western blot.Results HBV infection could cause fatty acid beta-oxidation disorder and mitochondrial dysfunction in vivo and in vitro. CPT1A overexpression could improve mitochondrial function in hepatocytes. Furthermore, decanoylcarnitine supplementation could activate CPT1A expression, thus improving fatty acid metabolism and repairing mitochondrial dysfunction. Proteomic analysis of mouse livers suggests that decanoylcarnitine stimulates the peroxisome proliferator-activated receptor (PPAR) signaling pathway, and PPAR alpha was the most important among PPARs.Conclusions Impaired fatty acid metabolism and mitochondrial dysfunction in hepatocytes caused by HBV infection could be partially restored by exogenous supplementation of decanoylcarnitine. This work elucidates the therapeutic potential of decanoylcarnitine in HBV infection and provides a new approach for diseases related to mitochondrial dysfunction. Hepatitis B virus infection could cause fatty acid beta-oxidation disorder and mitochondrial dysfunction in vivo and in vitro, which could be partially restored by exogenous supplementation of decanoylcarnitine, mainly through the PPAR alpha-CPT1A pathway.

BACKGROUND AND AIMS:Metabolic dysfunction-associated steatohepatitis (MASH)-related fibrosis plays an important role in MASH prognosis; however, the underlying mechanism remains unknown. Here, we explored the involvement of miR-210-3p in MASH-associated fibrosis. METHODS:We examined miR-210-3p expression in patients with MASH, with or without fibrosis using microarray analysis. miR-210-3p expression both in vivo and in vitro was validated using real-time quantitative reverse transcription PCR (RT-qPCR). Target genes and mechanisms were explored using western blotting, dual luciferase assay and immunofluorescence staining. Ferroptosis was assessed based on the levels of malondialdehyde (MDA), glutathione (GSH), iron and ferroptosis-related protein. RESULTS:miR-210-3p decreased significantly in mice fed methionine-choline-deficient (MCD)-fed and in those fed a high-fat diet (HFD) + CCL4 diets. Palmitic acid (PA)-stimulated LX2 and primary hepatic stellate cells showed miR-210-3p downregulation, consistent with the microarray results. miR-210-3p overexpression alleviated MASH-related fibrosis by inducing ferroptosis in hepatic stellate cells. Iron-sulfur cluster assembly enzyme (ISCU) was validated as the downstream target of miR-210-3p and its overexpression reversed miR-210-3p-induced ferroptosis. Overall, the ISCU-IRP1-CD71 axis is vital to miR-210-3p-induced ferroptosis. CONCLUSIONS:miR-210-3p expression is decreased in MASH-related fibrosis and is involved in ferroptosis by targeting ISCU.

Low-level viremia (LLV) in chronic hepatitis B (CHB) represents a significant challenge and area of interest in current clinical management. While nucleos (t)ide analogs (NAs) have demonstrated substantial efficacy in suppressing hepatitis B virus (HBV) replication, the application of highly sensitive detection methods has revealed that some treated patients continue to exhibit persistent or intermittent low-level viremia (HBV DNA: 12–2000 IU/mL). The mechanisms underlying LLV involve a synergistic interplay between host immune response deficiencies and HBV covalently closed circular DNA (cccDNA) persistence. Furthermore, the complex regulation of LLV is influenced by metabolic-associated steatotic liver disease (MASLD). Limitations in the clearance of cccDNA by current antiviral regimens also contribute to this phenomenon. LLV may elevate the risk of liver fibrosis progression, hepatocellular carcinoma (HCC), and end-stage liver disease. Current management strategies emphasize optimizing antiviral regimens, such as switching to tenofovir alafenamide (TAF) or combining therapies with pegylated interferon-alpha (Peg-IFN-α). Enhanced dynamic monitoring, including high-sensitivity HBV DNA assays and quantitative hepatitis B surface antigen (HBsAg) measurements, is also crucial. Moreover, exploring combination therapies involving immunomodulation and hepatocyte regeneration is warranted. Further research should integrate multi-omics technologies with prospective cohort studies to elucidate the host-virus interaction network in LLV. This will allow for the validation of synergistic effects between metabolic interventionsand immunotherapy, thereby advancing personalized precision medicine. This review systematically synthesizes the epidemiological characteristics, pathogenesis, influencing factors, prognosis, and clinical management advancements of LLV, aiming to provide novel perspectives for optimizing therapeutic strategies and translational research.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a prevalent, multifactorial systemic metabolic disorder, now recognized as the most common chronic liver disease globally. Female susceptibility to MASLD varies across menstrual states, influenced by genetic factors, age, menopausal status, and physical activity. Postmenopausal women, experiencing a significant reduction in estrogen, are particularly vulnerable to metabolic imbalances, increasing their risk of MASLD, disease progression, liver fibrosis, insulin resistance, and adverse cardiovascular events compared to premenopausal women and age-matched men. This review systematically synthesizes current research on extrahepatic abnormalities associated with MASLD in postmenopausal women. This review identifies key extrahepatic markers associated with MASLD in postmenopausal women, highlighting gaps in current research and proposing targeted screening and management strategies. (Graphical Abstract).

Congenital hepatic fibrosis (CHF) caused by mutations in the polycystic kidney and hepatic disease 1 (PKHD1) gene is a rare genetic disorder with poorly understood pathogenesis. We hypothesized that integrating gut microbiome and metabolomic analyses could uncover distinct host-microbiome interactions in CHF mice compared to wild-type controls. Pkhd1del3–4/del3–4 mice were generated using CRISPR/Cas9 technology. Fecal samples were collected from 11 Pkhd1del3–4/del3–4 mice and 10 littermate wild-type controls. We conducted a combined study using 16 S rDNA sequencing for microbiome analysis and untargeted metabolomics. The gut microbiome and metabolome data were integrated using Data Integration Analysis for Biomarker discovery using Latent cOmponents (DIABLO), which helped identify key microbial and metabolic features associated with CHF. CHF mouse model was successfully established. Our analysis revealed that the genera Mucispirillum, Eisenbergiella, and Oscillibacter were core microbiota in CHF, exhibiting significantly higher abundance in Pkhd1del3–4/del3–4 mice and strong positive correlations among them. Network analysis demonstrated robust associations between the gut microbiome and metabolome. Multi-omics dimension reduction analysis demonstrated that both the microbiome and metabolome could effectively distinguish CHF mice from controls, with area under the curve of 0.883 and 0.982, respectively. A significant positive correlation was observed between the gut microbiome and metabolome, highlighting the intricate relationship between these two components. This study identifies distinct metabolic and microbiome profiles in Pkhd1del3–4/del3–4 mice. Multi-omics analysis effectively differentiates CHF mice from controls and identified potential biomarkers. These findings indicate that gut microbiota and metabolites are integral to the pathogenesis of CHF, offering novel insights into the disease mechanism.

Purpose:Hyperactive immune responses in severe fever with thrombocytopenia syndrome (SFTS) is considered to associated with disease severity, prognosis and complications. This article aims to evaluate the validity of complement C3a, C5a, and sC5b-9 in predicting the severity and clinical outcomes in SFTS. Patients and Methods:Patients diagnosed with SFTS at the First Affiliated Hospital with Nanjing Medical University from March to November 2021 were enrolled in this retrospective analysis. The study evaluated C3a, C5a, and sC5b-9 levels between SFTS patients and healthy controls. The diagnostic and prognostic efficiency of C3a, C5a, and sC5b-9 for SFTS was assessed utilizing receiver operating characteristic (ROC) curve analysis. Correlation analysis was performed to examine the relationships between these complement components and clinical laboratory parameters in SFTS patients. Results:A total of 67 hospitalized SFTS patients were enrolled. SFTS patients exhibited significantly higher concentrations of C3a, C5a, and sC5b-9 compared to healthy controls. Non-survival and severe SFTS patients had notably higher C3a and sC5b-9 levels than survival and mild, respectively. ROC curve analysis revealed that C3a and sC5b-9 demonstrated effective performance for distinguishing severity in SFTS patients, with the area under the curve (AUC) of 0.784 (95% CI: 0.671-0.896, p < 0.001) and 0.703 (95% CI: 0.573-0.832, p = 0.005), respectively. The correlation analysis indicated that C3a and sC5b-9 positively correlated with SFTS RNA, CRP, PCT, ALT, AST, ALP, LDH, CK, HBDH, APPT, TT and D-dimer, while C3a negatively correlated with PLT. Conclusion:This study revealed abnormalities in complement components among patients with SFTS. C3a and sC5b-9 levels show promise as biomarkers for linking with disease severity and prognosis, potentially providing therapeutic targets for the management of SFTS patients and guide future mechanistic research.

Viral infections disrupt glucose metabolism; however, their impact on disease prognosis in highly pathogenic viruses remains largely unknown. There is an additional need to investigate the antiviral mechanisms of glucose-lowering therapeutics. Here, our multicenter clinical study shows that hyperglycemia and pre-existing diabetes are independent risk factors for mortality in patients infected with severe fever with thrombocytopenia syndrome virus (SFTSV), an emerging and highly pathogenic bunyavirus. SFTSV infection triggers gluconeogenesis, which, in turn, inhibits AMPK activity and subsequent interferon I (IFN-I) responses, thereby facilitating viral replication. In vitro and animal studies further reveal that metformin inhibits SFTSV replication by suppressing autophagy through the AMPK-mTOR pathway, contributing to protection against lethal SFTSV infection in mice. Importantly, our large cohort study demonstrates that metformin reduces viremia and SFTSV-related mortality in patients with hyperglycemia or pre-existing diabetes, contrasting with the disadvantageous effect of insulin. These findings highlight the promising therapeutic potential of metformin in treating viral infections, particularly among individuals with hyperglycemia or diabetes. IMPORTANCE Severe fever with thrombocytopenia syndrome virus (SFTSV), an emerging tick-borne bunyavirus, causes severe hemorrhagic fever with a high mortality rate. Previous studies have shown metabolic disturbances, particularly hyperglycemia, in SFTSV-infected individuals. However, the mechanism underlying this metabolic derangement remains unclear, and further investigation is needed to determine whether glucose-lowering therapeutics could be beneficial for SFTSV-infected patients. In this study, our multicenter clinical data show that hyperglycemia and pre-existing diabetes are independent risk factors for mortality in patients with SFTSV infection. Furthermore, we observed that SFTSV infection triggers gluconeogenesis, which promotes viral replication through the regulation of the AMPK-IFN-I signaling pathway. Notably, metformin significantly reduces viremia and SFTSV-related mortality in patients with hyperglycemia or pre-existing diabetes, attributed to its inhibitory effect on autophagy through the AMPK-mTOR pathway. Therefore, our study uncovers the interaction between SFTSV infection and glucose metabolic disorder and highlights the promising therapeutic potential of metformin for treating SFTSV infection.

Tumor-associated macrophages (TAMs) have been implicated in fostering various hallmarks of cancer progression in gastric cancer (GC). However, the intricate molecular mechanisms underlying TAM-induced chemoresistance remain incompletely understood. Exosomes emerge as key players, mediating TAM-induced resistance to cisplatin (DDP) by regulating ferroptosis. Our investigation reveals that exo-miR-1911-5p, delivered to GC cells from TAMs, significantly contributes to cisplatin resistance. Specifically, direct modulation of MYB by MiR-1911-5p leads to decreased expression of AKR1B10, a crucial factor in preventing ferroptosis. Further exploration confirms the regulation of ACC by AKR1B10. Through targeting the MYB/AKR1B10/ACC axis, exo-miR-1911-5p inhibits ferroptosis to enhances cisplatin resistance. Additionally, exo-miR-1911-5p promotes M2 polarization of TAMs by targeting ARHGEF3. Collectively, our findings highlight the critical role of exo-miR-1911-5p in mediating cisplatin resistance through modulating the cross-talk between TAMs and GC. Targeting exo-miR-1911-5p could represent a promising strategy for overcoming DDP resistance in GC.

BACKGROUND:Severe fever with thrombocytopenia syndrome (SFTS), caused by Dabie bandavirus (DBV) infection, is characterized by early cytokine storm as a primary pathological feature, although the precise mechanisms remain unclear. Low-density neutrophils (LDNs) are elevated in the peripheral blood of patients with autoimmune or infectious diseases and are closely associated with inflammatory damage and disease severity. However, the pathogenic contribution of LDNs to the progression of SFTS is largely unexplored. This study employed single-cell RNA sequencing (scRNA-seq) to profile the transcriptomic characteristics of LDNs during the acute phase of SFTS, aiming to reveal their compositional and functional heterogeneity following DBV infection, explore their role in the cytokine storm, and further understand their impact on disease progression. METHODS:Cells were isolated from 13 acute-phase SFTS patients with varying disease severity and 3 healthy controls using density gradient centrifugation, followed by preparation of single-cell suspensions for 3'-end scRNA-seq. Sequencing data were processed using the Seurat pipeline, including dimensionality reduction, clustering, cell-type annotation, and visualization with Uniform Manifold Approximation and Projection (UMAP). Low-density granulocytes (LDGs) and their subclusters were identified using canonical gene markers. Functional enrichment of differentially expressed genes (DEGs) was analyzed by high-dimensional Weighted Gene Co-expression Network Analysis (hdWGCNA), Gene Ontology (GO), AddModuleScore, single-sample Gene Set Enrichment Analysis (ssGSEA), and immune-related Gene Set Enrichment Analysis (irGSEA), while cellular interactions were explored using CellCall. RESULTS:1. Compositional heterogeneity: The proportion of LDNs in peripheral blood increased in SFTS patients with greater disease severity during the acute phase. 2. Functional heterogeneity: (1) LDN subclusters showed functional diversity but consistently displayed pro-inflammatory or anti-infective properties. (2) With intensification of the systemic inflammatory response, the expression of multiple cytokine genes (e.g., IL6, IL8, TNFA) and gene sets of the inflammatory pathway (e.g., TNFA-SIGNALING-VIA-NFKB, INFLAMMATORY-RESPONSE) were significantly upregulated in LDNs. Concurrently, the expression of gene sets of type I interferon response pathway (e.g., INTERFERON-ALPHA-RESPONSE, INTERFERON-GAMMA-RESPONSE) and genes of interferon-induced antiviral proteins (e.g., EIF2AK2, OAS1, MX1) were also elevated. (3) In severe cases, glucocorticoid therapy downregulated expression of these inflammatory genes, demonstrating anti-inflammatory effects but potentially increasing infection risk. CONCLUSIONS:This study revealed an increased proportion and heightened pro-inflammatory activity of LDNs during the acute phase of SFTS, closely correlating with disease severity. These findings suggest that LDNs may serve as potential early-warning biomarkers for predicting severe progression in patients with SFTS.

Klebsiella pneumoniae is a globally recognized microbial pathogen with significant clinical impact. The bacterium harbors the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas systems, which provide adaptive immunity against invading foreign nucleic acids. Recent studies suggest that certain CRISPR-Cas systems can regulate endogenous genes, influencing bacterial virulence. However, their role in regulating pathogenicity in K. pneumoniae remains poorly understood. This study investigates the regulatory role of the type I-E* CRISPR-Cas system in a hypervirulent K. pneumoniae strain, focusing on its impact on histidine metabolism and pathogenicity. Transcriptome analyses identified differentially expressed genes (DEGs) between the casABECD-deletion and wild-type strains, including significant upregulation of the histidine utilization (Hut) operon and downregulation of biofilm-related genes. These molecular changes resulted in enhanced histidine metabolic activity, reduced biofilm formation, attenuated virulence in A549 lung epithelial cells, and improved survival of Galleria mellonella, as validated through phenotypic and virulence assays. Our bioinformatic analysis indicated that the CRISPR-Cas system in K. pneumoniae targets the hutT sequence, which is part of the Hut operon. Furthermore, the overexpression of hutT mitigated CRISPR-Cas-mediated repression of the Hut operon, as observed in virulence assays, while simultaneous deletion of hutH and casABECD restored the reduced virulence in the ΔcasABECD strain. Additionally, deletion of casABECD significantly enhances the growth of the strain in medium with histidine as the sole carbon source, highlighting the intricate regulatory role of the CRISPR-Cas system in metabolic adaptation. Collectively, these findings uncover a novel role for the CRISPR-Cas system in regulating metabolic pathways and virulence in hypervirulent K. pneumoniae.IMPORTANCEClustered regularly interspaced short palindromic repeats (CRISPR)-Cas systems are primarily recognized for their roles in adaptive immunity against foreign genetic elements in bacteria. However, emerging evidence indicates that these systems can also regulate endogenous genes, thereby influencing bacterial physiology and virulence. In this study, we demonstrate that the type I-E* CRISPR-Cas system in Klebsiella pneumoniae targets the hutT gene, a critical component of the histidine utilization (Hut) pathway. This targeting potentially impacts hutT transcription and alters the expression of other hut genes, ultimately enhancing bacterial virulence. Our findings reveal a previously unrecognized regulatory mechanism through which CRISPR-Cas systems facilitate metabolic adaptation and pathogenicity in K. pneumoniae. This study broadens our understanding of the multifaceted roles of CRISPR-Cas systems in bacterial physiology and pathobiology, with implications for clinically relevant pathogens.

ABSTRACT Ultrasound‐derived fat fraction (UDFF) is designed to assess the hepatic fat content quantitatively. A multicenter study that verifies the diagnostic performance of UDFF for detecting hepatic steatosis has not yet been reported. This study aimed to evaluate the performance of UDFF for diagnosing and grading hepatic steatosis. Participants referred for assessment of hepatic steatosis were prospectively recruited from eight hospitals. All participants underwent UDFF and magnetic resonance imaging proton density fat fraction (MRI‐PDFF) examinations. MRI‐PDFF was used as the reference for diagnosing hepatic steatosis. From January 2023 to July 2023, a total of 300 participants were included. The median body mass index was 25.4 kg/m2 (interquartile range: 22.7–28.1). UDFF values were positively correlated with MRI‐PDFF (R = 0.80, p < 0.001). Using MRI‐PDFF ≥ 5%, ≥ 15%, and ≥ 25% as the reference standard for detecting mild, moderate, and severe hepatic steatosis, the best cutoff values of UDFF were 7.6% (area under the receiver operating characteristic curves [AUC] = 0.90), 15.9% (AUC = 0.90), and 22.3% (AUC = 0.91), respectively. Thus, UDFF has excellent diagnostic performance in detecting and grading hepatic steatosis.