Role and mechanism of MiR-542-3p in regulating TLR4 in nonylphenol-induced neuronal cell pyroptosis.

BACKGROUND: This study aimed to investigate the spatial learning/memory and motor abilities of rats and the alteration of miR-542-3p and pyroptosis in the midbrain nigrostriatal area in vivo after nonylphenol (NP) gavage and to explore the mechanism of miR-542-3p regulation of Toll-like receptor 4 (TLR4) in NP-induced pyroptosis in BV2 microglia in vitro. METHODS: In vivo: Thirty-six specific-pathogen-free-grade Sprague-Dawley rats were divided into three equal groups: blank control group (treated with pure corn oil), NP group (treated with NP, 80 mg/kg body weight per day for 90 days), and positive control group [treated with lipopolysaccharide (LPS), 2 mg/kg body weight for 7 days]. In vitro: The first part of the experiment was divided into blank group (control, saline), LPS group [1 µg/ml + 1 mM adenosine triphosphate (ATP)], and NP group (40 µmol/L). The second part was divided into mimics NC (negative control) group, miR-542-3p mimics group, mimics NC + NP group, and miR-542-3p mimics + NP group. RESULTS: In vivo: Behaviorally, the spatial learning/memory and motor abilities of rats after NP exposure declined, as detected via Y-maze, open field, and rotarod tests. Some microglia in the substantia nigra of the NP-treated rats were activated. The downregulation of miR-542-3p was observed in rat brain tissue after NP exposure. The mRNA/protein expression of pyroptosis-related indicators (TLR4), NOD-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein (ASC), gasdermin-D (GSDMD), cysteinyl aspartate-specific proteinase-1 (caspase-1), and interleukin-1ß (IL-1ß) in the substantia nigra of the midbrain increased after NP exposure. In vitro: ASC fluorescence intensity increased in BV2 cells after NP exposure. The mRNA and/or protein expression of pyroptosis-related indicators (TLR4, NLRP3, GSDMD, caspase-1, and IL-1ß) in BV2 cells was upregulated after NP exposure. The transfection of miR-542-3p mimics inhibited NP-induced ASC expression in BV2 cells. The overexpression of miR-542-3p, followed by NP exposure, significantly reduced TLR4, NLRP3, ASC, caspase-1, and IL-1ß gene and/or protein expression. CONCLUSIONS: This study suggested that NP exposure caused a decline in spatial learning memory and whole-body motor ability in rats. Our study was novel in reporting that the upregulation of miR-542-3p targeting and regulating TLR4 could inhibit NLRP3 inflammatory activation and alleviate NP-induced microglia pyroptosis.

Characteristics of Klebsiella pneumoniae pyogenic liver abscess from 2010-2021 in a tertiary teaching hospital of South China.

OBJECTIVES: Pyogenic liver abscess (PLA) is a severe and potentially fatal infectious disease. Klebsiella pneumoniae (K. pneumoniae) is the predominant pathogen responsible for PLA. This study aims to investigate the clinical characteristics and prognostic factors of K. pneumoniae-induced pyogenic liver abscess (KP-PLA), particularly those caused by carbapenem-resistant K. pneumoniae (CRKP). METHODS: Analyses were performed on PLA patients from January 2010 to December 2021, to investigate the differences of K. pneumoniae from other etiologically infected PLA patients. Univariate and multivariate logistic regression analyses were used to compare prognostic factors between patients with carbapenem-resistant K. pneumoniae PLA (CRKP-PLA) and patients with carbapenem-sensitive K. pneumoniae PLA. RESULTS: Univariate analysis demonstrated a significant association between KP-PLA and factors including diabetes mellitus (P < 0.001), cholecystitis and cholelithiasis (P = 0.032), single abscess (P = 0.016), and abscesses with a diameter over 50 mm (P = 0.004). The CRKP group exhibited a higher prevalence of therapeutic interventions before K. pneumoniae infection, including abdominal surgery, mechanical ventilation, sputum suction, tracheal cannula, routine drainage of the abdominal cavity, and peripherally inserted central venous catheters (P < 0.05). Multivariate logistic regression analysis revealed that admission to the intensive care unit was an independent risk factor associated with CRKP-PLA (odds ratio 36; 95% confidence interval 1.77-731.56; P = 0.020). CONCLUSION: The KP-PLA patients were significantly associated with diabetes and were more likely to have single abscesses larger than 50 mm. PLA patients with a history of admission to intensive care unit or invasive therapeutic procedures should be given special consideration if combined with CRKP infection.

Tenofovir amibufenamide vs tenofovir alafenamide for treating chronic hepatitis B: A real-world study.

BACKGROUND: The efficacy and safety profile of tenofovir amibufenamide (TMF) in chronic hepatitis B (CHB) patients is not well-established. AIM: To compare the efficacy and safety of TMF and tenofovir alafenamide (TAF) over a 48-wk period in patients with CHB. METHODS: A total of 215 subjects meeting the inclusion criteria were enrolled and divided into two groups: TMF group (n = 106) and the TAF group (n = 109). The study included a comparison of virological response (VR): Undetectable hepatitis B virus DNA levels, alanine transaminase (ALT) normalization rates, renal function parameters, and blood lipid profiles. RESULTS: At 24 and 48 wk, VR rates for the TMF group were 53.57% and 78.57%, respectively, compared with 48.31% and 78.65% for the TAF group (P > 0.05). The VR rates were also similar in both groups among patients with low-level viremia, both hepatitis B e antigen (HBeAg)-positive and HBeAg-negative subgroups. The TMF cohort showed ALT normalization and renal safety profiles similar to the TAF group. There was a notable increase in total cholesterol levels in the TAF group (P = 0.045), which was not observed in the TMF group (P > 0.05). In patients with liver cirrhosis, both groups exhibited comparable VR and ALT normalization rates and renal safety profiles. However, the fibrosis 4 score at 48 wk showed a significant reduction in the TAF group as compared to the TMF group within the liver cirrhosis subgroup. CONCLUSION: Our study found TMF is as effective as TAF in treating CHB and has a comparable safety profile. However, TAF may be associated with worsening lipid profiles.

ANXA1 is identified as a key gene associated with high risk and T cell infiltration in primary sclerosing cholangitis.

BACKGROUND: Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease, with unclear pathogenesis. Although immune disorders, especially T cell infiltration, are thought to play a vital role in PSC, the specific pathogenesis mechanisms remain incompletely understood. This study evaluated the potential key gene associated with the PSC pathogenesis and analyzed the associations of the key gene with prognosis and immune cell infiltration by combining bioinformatics analysis and experimental verification. METHODS: Transcriptome data of PSC and normal human liver tissues (GSE159676) were obtained from the gene expression omnibus database. Differentially expressed genes (DEGs) were identified, and differences in biological states were analyzed. A protein-protein interaction (PPI) network was constructed. Hub genes were identified, and their expression was verified using transcriptome data of mice fed 0.1% 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) and Mdr2-/- mice (GSE179993, GSE80776), as well as by immunohistochemistry staining on clinical samples. The correlations between the key gene and other factors were evaluated by Pearson's correlation coefficient. Immune cell infiltration into human liver (GSE159676) was analyzed by xCell and verified by immunofluorescence staining on PSC liver samples. RESULTS: Of the 185 DEGs identified, 113 were upregulated and 72 were downregulated genes in PSC. Genes associated with immune cell infiltration and fibrosis were significantly enriched in PSC. PPI network showed close interactions among DEGs. A module strongly associated with immune infiltration was identified, with annexin A1 (ANXA1) being the core gene. High expression of ANXA1 in PSC was confirmed in two public datasets and by immunohistochemistry staining on clinical samples. High ANXA1 expression was strongly associated with high-risk score for PSC. Also, ANXA1 expression was positively associated with chemokines and chemokine receptors and with the infiltration of immune cells, especially T cells, into liver with PSC. Immune infiltration, fibrosis, and cancer-related processes were markedly enriched in PSC with high expression of ANXA1. CONCLUSION: ANXA1 is a key gene associated with high risk and infiltration of immune cells, especially T cells, in PSC. These findings provide new insight into the key biomarker of PSC and suggest that targeting ANXA1 may be a valuable strategy for the treatment of PSC.

Clinicopathologic characteristics of liver inflammation and fibrosis in 310 patients with chronic hepatitis B. / 310ä¾‹æ ¢æ€§ä¹™åž‹è‚ç‚Žæ‚£è€ è‚è„ç‚Žç—‡åŠçº¤ç»´åŒ–çš„ä¸´åºŠç— ç†ç‰¹å¾.

OBJECTIVES: Long-term hepatitis B virus (HBV) infection can cause recurrent inflammation in the liver, and then develop into liver fibrosis, cirrhosis, and liver cancer. The hepatic pathological change is one of the important criteria for guiding antiviral therapy in patients with chronic hepatitis B (CHB). Due to the limitations of liver biopsy, it is necessary to find valuable non-invasive indicators to evaluate the hepatic pathological changes in CHB patients and guide the antiviral therapy. This study aims to analyze the clinical characteristics of different pathological changes in CHB patients, and to explore the factors influnencing the degree of liver inflammation and fibrosis in CHB patients with normal alanine aminotransferase (ALT). METHODS: This retrospective study was conducted on 310 CHB patients. Liver biopsy was performed in all these patients. The clinical data of the patients were collected. The liver biopsy pathological results were used as the gold standard to analyze the relationship between clinical indicators and liver pathological changes. Then CHB patients with normal ALT were screened, and the independent factors influencing the degree of liver inflammation and fibrosis were explored. RESULTS: Among the 310 patients with CHB, there were 249 (80.3%) patients with significant liver inflammation [liver inflammation grade (G) ≥2] and 119 (38.4%) patients with significant liver fibrosis [liver fibrosis stage (S) ≥2]. The results of univariate analysis of total samples showed that the ALT, γ-glutamyl transferase, alkaline phosphatase, and HBV DNA were related to the significant liver pathological changes. Among the 132 CHB patients with normal ALT, the patients with liver pathology G/S≥2, G≥2, and S≥2 were 80.3% (106/132), 68.2% (90/132), and 43.2% (57/132), respectively. The results showed that the independent influencing factor of significant liver inflammation was HBV DNA>2 000 U/mL (OR=3.592, 95% CI 1.534 to 8.409), and the independent influencing factors of significant liver fibrosis were elevated alkaline phosphatase level (OR=1.022, 95% CI 1.002 to 1.043), decreased platelet count (OR=0.990, 95% CI 0.982 to 0.998), and positive in hepatitis B e antigen (HBeAg) (OR=14.845, 95% CI 4.898 to 44.995). According to the multivariate analysis, a diagnostic model for significant liver fibrosis in CHB patients with normal ALT was established, and the area under the receiver operating characteristic curve was 0.844 (95% CI 0.779 to 0.910). CONCLUSIONS: The liver pathological changes should be evaluated in combination with different clinical indicators. A considerable number of CHB patients with normal ALT still have significant liver pathological changes, which need to be identified and treated with antiviral therapy in time. Among them, HBV DNA>2 000 U/mL suggests the significant liver inflammation, and the diagnostic model for significant liver fibrosis based on alkaline phosphatase, platelet count, and HBeAg can help to evaluate the degree of liver fibrosis.

IL-27 Gene Therapy Induces Stat3-Mediated Expansion of CD11b+Gr1+ Myeloid Cells and Promotes Accumulation of M1 Macrophages in the Tumor Microenvironment.

IL-27 is a pleiotropic cytokine that exhibits stimulatory/regulatory functions on multiple lineages of immune cells and has a potential to be used as a therapeutic for cancer. We have recently demonstrated that administration of IL-27 producing adeno-associated virus (AAV-IL-27) exhibits potent inhibition of tumor growth in mouse models. In this study, we demonstrate that AAV-IL-27 treatment leads to significant expansion of CD11b+Gr1+ myeloid cells. AAV-IL-27-induced expansion of CD11b+Gr1+ cells is IL-27R-dependent and requires Stat3 signaling, but it is inhibited by Stat1 signaling. AAV-IL-27 treatment does not increase the self-renewal capacity of CD11b+Gr1+ cells but induces significant expansion of Lin-Sca1+c-Kit+ (LSK) and granulocyte-monocyte progenitor cells. Despite exhibiting significant suppression of T cells in vitro, IL-27-induced CD11b+Gr1+ cells lost the tumor-promoting activity in vivo and overall play an antitumor role. In tumors from AAV-IL-27-treated mice, CD11b+Gr1+ cells are largely F4/80+ and express high levels of MHC class I/II and M1 macrophage markers. Thus, IL-27 gene therapy induces Stat3-mediated expansion of CD11b+Gr1+ myeloid cells and promotes accumulation of M1 macrophages in the tumor microenvironment.

CD200R signaling contributes to unfavorable tumor microenvironment through regulating production of chemokines by tumor-associated myeloid cells.

CD200 is overexpressed in many solid tumors and considered as an immune checkpoint molecule dampening cancer immunity. In this study, we found that CD200R-/- mice were significantly more potent in rejecting these CD200+ tumors. scRNA sequencing demonstrated that tumors from CD200R-/- mice had more infiltration of CD4+ and CD8+ T cells, and NK cells but less infiltration of neutrophils. Antibody depletion experiments revealed that immune effector cells are crucial in inhibiting tumor growth in CD200R-/- mice. Mechanistically, we found that CD200R signaling regulates the expression of chemokines in tumor-associated myeloid cells (TAMCs). In the absence of CD200R, TAMCs increased expression of CCL24 and resulted in increased infiltration of eosinophils, which contributes to anti-tumor activity. Overall, we conclude that CD200R signaling contributes to unfavorable TME through chemokine-dependent recruitment of immune suppressive neutrophils and exclusion of anti-cancer immune effectors. Our study has implications in developing CD200-CD200R targeted immunotherapy of solid tumors.

Tenofovir alafenamide versus entecavir for treating hepatitis B virus-related acute-on-chronic liver failure: real-world study.

Background and aims: Real-world data regarding hepatitis B virus-related acute-on-chronic liver failure (HBV-ACLF) patients receiving tenofovir alafenamide (TAF) as an antiviral drug are limited. Hence, we evaluated the efficacy and kidney safety of TAF among this population. Methods: A total of 272 HBV-related ACLF patients hospitalized at Xiangya Hospital of Central South University were enrolled in this retrospective research. All patients received antiviral therapy with TAF (n = 100) or ETV (n = 172) and comprehensive medical treatments. Results: Through 1:1 propensity score matching, 100 patients were finally included in each group. At week 48, the survival rates without transplantation of the TAF group and ETV group were 76.00 and 58.00%, separately (P = 0.007). After 4 weeks of treatment, the TAF treatment group exhibited a significantly decline in HBV DNA viral load (P = 0.029). The mean estimated glomerular filtration rate was apparently improved in the TAF group compared with the ETV group (TAF 5.98 ± 14.46 vs. ETV 1.18 ± 18.07 ml/min/1.73 m2) (P < 0.05). There were 6 patients in TAF group and 21 patients in ETV group with chronic kidney disease (CKD) stage progression ≥ 1. By contrast, the ETV treatment group has a greater risk of renal function progression in CKD 1 stage patients (P < 0.05). Conclusion: This real-world clinical study showed that TAF is more effective than ETV in reducing viral load and improving survival rate in HBV-ACLF patients and the risk of renal function decline is lower. Clinical trial registration: https://ClinicalTrials.gov, identifier NCT05453448.

Composition, distribution and enrichment of trace metals in sediments from the muddy area off the southern Shandong Peninsula in the Northwestern South Yellow Sea of China since 10,000 years.

We analyzed ten trace metals (Cu, Pb, Zn, Cr, Cd, Hg, As, Ni, V, Co and Ni) in sediments of the upper 14.98 m of core WHZK01 from the muddy area off the Shandong Peninsula, northwestern South Yellow Sea, to analyze their content, vertical distribution, and the enrichment status. Except for Hg and As, the other metals (Cu, Pb, Zn, Cr, Cd, Ni, V, Co and Ni) were mainly controlled by grain size. When the sediment particle size became smaller, the metal content reached a high level. Moreover, oxides and hydroxides of Al, Ti, Fe, and Mn also contributed to the metal enrichments due to their strong adsorption to the metals. Over the past four stages of 10-7 kyr BP, 7-4.5 kyr BP, 4.5-2.5 kyr BP, and 2.5 kyr BP to date, the metal values have shown a trend of increasing - fluctuating to high values - decreasing - re-increasing, respectively. However, since 4.5 kyr BP, Hg concentrations have been on an increasing trend, associated with the release of large amounts of contaminants into the environment from ancient human metal mining and smelting activities. As concentrations, despite the fluctuating changes, have remained relatively stable at high levels since 5.5 kyr BP, associated with their high background values.

Distribution, contamination and provenance of heavy metals in sediments from the nearshore area of Weihai City, eastern Shandong Peninsula, China.

We analyzed eight heavy metals (Cu, Pb, Zn, Cr, Cd, Hg, As, and Ni) in 85 seabed sediments off the coast of Weihai City, eastern Shandong Peninsula, China, to reveal their distributions, enrichment status, and pollutant sources. Cu, Pb, Zn, Cr, As, and Ni were enriched in all bays in both the inner and outer waters. However, Cd and Hg were more abundant in Weihai Bay, followed by Rongcheng Bay and Chaoyang Port, with denser populations and more developed industries near the coast. Most areas exhibited slight contamination with As and Pb, with relatively severe contamination in localized areas. Moreover, Weihai Bay showed slight contamination with Cd, Zn, and Hg. Heavy metals are largely influenced by the discharge of anthropogenic pollutants along the coast. It is recommended to implement strict control measures on the discharge of waste into the sea, and ensure the sustainable development of the marine ecological environment.

Dendritic cell expression of CD24 contributes to optimal priming of T lymphocytes in lymph nodes.

CD24 is a GPI anchored cell surface glycoprotein whose function as a co-stimulatory molecule has been implicated. However, the function of CD24 on antigen presenting cells during T cell responses is not well understood. Here we show that in the CD24-deficient host, adoptively transferred CD4+ T cells undergo inefficient expansion and have accelerated cell death in lymph nodes, which results in insufficient priming of T cells. Insufficient expansion of T cells in the CD24-deficient host was not due to host anti-CD24 response by NK, T and B lymphocytes. Transgenic expression of CD24 on DC in CD24-/- mice restored T cell accumulation and survival in draining lymph nodes. Consistent with these findings, MHC II tetramer staining also revealed that an antigen-specific polyclonal T cell response was reduced in lymph nodes of CD24-/- mice. Taken together, we have revealed a novel role of CD24 on DC in optimal T cell priming in lymph nodes. These data suggest that CD24 blockade should lower unwanted T cell responses such as those in autoimmune diseases.

Role of miR-219a-5p in regulating NMDAR in nonylphenol-induced synaptic plasticity damage.

Nonylphenol (NP) is a typical environmental endocrine disruptor with estrogenic effects. It serves as an emulsifier and as the main ingredient of detergents and has become an increasingly common pollutant in both fresh and salt water, vegetables, and fruits. This study aimed to clarify whether NP exposure could lead to cognitive dysfunction and synaptic plasticity impairment, and also explore the mechanism of microRNA (miR)- 219a-5p regulation of N-methyl-D-aspartate receptor (NMDAR) in NP-induced synaptic plasticity impairment in vivo and in vitro. In vivo, 30 male Sprague-Dawley rats were randomly divided into 2 groups: blank control group (pure corn oil) and NP-exposed group [NP 80 mg/(kg·d)], with 15 rats in each group. In vitro, the extracted hippocampal neurons were divided into 6 groups: blank control group, mimics NC group, miR-219 mimics group, NP group (70 µmol/L NP), NP + mimics NC group, and NP + miR-219 mimics group. In vivo, the content of NP in hippocampal tissues after 90 days of NP exposure was significantly higher in the NP-stained group than in the blank control group. NP exposure could lead to a decrease in the ability to learn and memory, ability to remember, and space spatial memory ability in rats. The dendrites in the NP-stained group were disordered, with few dendritic spines and significantly decreased dendritic spine density. The postsynaptic densities were loosely arranged, the thickness and length of the postsynaptic densities shortened, and the length and width of the synaptic gap increased. Glutamine (Glu) and γ-aminobutyric acid (GABA) contents in hippocampal tissues decreased in the NP-stained group. The expression of miR-219a-5p mRNA decreased in the NP-stained group after 3 months of NP exposure. The expression of NMDAR1, NMDAR2A, NMDAR2B, nerve growth-associated protein (GAP-43), and Ca/calmodulin-dependent kinase II (CaMKII) mRNA/proteins decreased in the NP-stained group. In vitro, NMDAR protein expression decreased, while GAP-43 and CaMKII protein expression increased in the miR-219 mimics group compared with the control group. The expression levels of NMDAR and GAP-43 and CaMKII proteins were higher in the NP + miR-219 mimics group compared with the NP group. The levels of neurotransmitters Glu and GABA decreased in the NP and NP + mimics NC groups compared with the blank group. Shortened synaptic active band length, decreased thickness of postsynaptic densities, and shortened length of postsynaptic densities were observed in the NP, NP + mimics NC, and NP + miR-219 mimics groups compared with the blank control group. In vivo, NP exposure reduced learning memory capacity and neurotransmitter content in rats and caused a decrease in dendritic spine density and synaptic number density and a decrease in miR-219a-5p expression. In vitro, high expression of miR-219a-5p inhibited the expression of NMDAR, thus reducing the effect of NP on synaptic plasticity impairment in hippocampal neurons. Our study provided a scientific basis for the prevention of cognitive impairment owing to NP exposure and the development of targeted drug treatment strategies.

Demographics and Clinical Outcomes of Culture-Positive versus Culture-Negative Pyogenic Liver Abscess in an Asian Population.

Objective: Despite its high case-fatality risk, pyogenic liver abscess (PLA) lacks clear management guidelines in patients with negative microbial cultures. Our aim was to evaluate differences in clinical characteristics between patients with culture-negative liver abscess (CNLA) and those with culture-positive liver abscess (CPLA), and identify differences in the main causative pathogen. Methods: In this study, we retrospectively collected medical records of PLA patients admitted to a teaching hospital from January 2010 to December 2019. Results: In total, 324 PLA patients were enrolled in this study. Of these, 202 (62.3%) cases were confirmed cultural positive, including 109 patients (54%) and 20 (9.9%) patients infected with Klebsiella pneumoniae (K. pneumoniae) and Escherichia coli (E.coli), respectively. Patients in the CPLA group were older (p=0.029) and had higher prevalence of abscesses larger than 5 cm in diameter (p = 0.003), gas-forming rate (p = 0.016), and percutaneous drainage (p < 0.001) compared with CNLA group. Patients with CPLA had significantly longer hospitalizations than those with CNLA (p = 0.010). Nevertheless, there was no significant difference in in-hospital mortality between the two groups (p = 0.415). Compared with patients with E. coli, those with K. pneumoniae had higher incidence of diabetes mellitus (p = 0.041), solitary abscess (p < 0.001), localization in the right hepatic lobe (p = 0.033), abscess size larger than 5 cm (p < 0.001) and percutaneous drainage (p = 0.002), but mortality was not significantly different (p = 1.000). Conclusion: No significant difference in in-hospital mortality was found between patients with CNLA and those with CPLA group. However, clinical characteristics and management were different between the main causative pathogens, including K. pneumoniae and E. coli.

Identification of AKR1B10 as a key gene in primary biliary cholangitis by integrated bioinformatics analysis and experimental validation.

Background: Primary biliary cholangitis (PBC) is a chronic autoimmune liver disease that eventually progresses to cirrhosis and hepatocellular carcinoma (HCC) in the absence of proper treatment. However, Gene expression and molecular mechanisms involved in the pathogenesis of PBC have not been completely elucidated. Methods: Microarray expression profiling dataset GSE61260 was downloaded from the Gene Expression Omnibus (GEO) database. Data were normalized to screen differentially expressed genes (DEGs) using the limma package in R. Moreover, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) enrichment analyses were performed. A protein-protein interaction (PPI) network was constructed to identify hub genes and an integrative regulatory network of transcriptional factor-DEG-microRNA was established. Gene Set Enrichment Analysis (GSEA) was used to analyze differences in biological states for groups with different expressions of aldo-keto reductase family 1 member B10 (AKR1B10). Immunohistochemistry (IHC) analysis was performed to validate the expression of hepatic AKR1B10 in patients with PBC. The association of hepatic AKR1B10 levels with clinical parameters was evaluated using one-way analysis of variance (ANOVA) and Pearson's correlation analysis. Results: This study identified 22 upregulated and 12 downregulated DEGs between patients with PBC and healthy controls. GO and KEGG analysis revealed that DEGs were mainly enriched in immune reactions. AKR1B10 was identified as a key gene and was further analyzed by screening out hub genes from the PPI network. GSEA analysis indicated that high expression of AKR1B10 might promote PBC to develop into HCC. Immunohistochemistry results verified the increased expression of hepatic AKR1B10 in patients with PBC and demonstrated its positive correlation with the severity of PBC. Conclusion: AKR1B10 was identified as a hub gene in PBC by integrated bioinformatics analysis and clinical validation. The increase of AKR1B10 expression in patients with PBC was associated with disease severity and might promote the progression of PBC to HCC.

Relationships among Problematic Smartphone Use, Mathematics Achievement, Teacher-Student Relationships, and Subjective Well-Being: Results from a Large-Scale Survey in China.

This study examined the mediating role of mathematics performance and the moderating role of teacher-student relationships on the effects of problematic smartphone use on students' subjective well-being. Through probability proportionate to size sampling (PPS), a total of 20,321 fourth graders from a city in central China were invited to complete a paper-based mathematics achievement test and an online questionnaire survey, including demographic information, problematic smartphone use, subjective well-being, and teacher-student relationship scales. The results showed that: after controlling for SES and gender, (1) problematic smartphone use had a direct and negative effect on students' subjective well-being; (2) mathematics performance partially mediated the effects of problematic smartphone use on students' subjective well-being; (3) teacher-student relationships moderated the effects of problematic smartphone use on mathematics performance/students' subjective well-being; (4) with the increase in problematic smartphone use, high teacher-student relationships produced a lower rate of the positive moderating effect than low teacher-student relationships. The implications of this study and suggestions for future research are discussed.

Novel CRISPR/Cas9-mediated knockout of LIG4 increases efficiency of site-specific integration in Chinese hamster ovary cell line.

AIM: To investigate the impact of deficiency of LIG4 gene on site-specific integration in CHO cells. RESULTS: CHO cells are considered the most valuable mammalian cells in the manufacture of biological medicines, and genetic engineering of CHO cells can improve product yield and stability. The traditional method of inserting foreign genes by random integration (RI) requires multiple rounds of screening and selection, which may lead to location effects and gene silencing, making it difficult to obtain stable, high-yielding cell lines. Although site-specific integration (SSI) techniques may overcome the challenges with RI, its feasibility is limited by the very low efficiency of the technique. Recently, SSI efficiency has been enhanced in other mammalian cell types by inhibiting DNA ligase IV (Lig4) activity, which is indispensable in DNA double-strand break repair by NHEJ. However, this approach has not been evaluated in CHO cells. In this study, the LIG4 gene was knocked out of CHO cells using CRISPR/Cas9-mediated genome editing. Efficiency of gene targeting in LIG4-/--CHO cell lines was estimated by a green fluorescence protein promoterless reporter system. Notably, the RI efficiency, most likely mediated by NHEJ in CHO, was inhibited by LIG4 knockout, whereas SSI efficiency strongly increased 9.2-fold under the precise control of the promoter in the ROSA26 site in LIG4-/--CHO cells. Moreover, deletion of LIG4 had no obvious side effects on CHO cell proliferation. CONCLUSIONS: Deficiency of LIG4 represents a feasible strategy to improve SSI efficiency and suggests it can be applied to develop and engineer CHO cell lines in the future.

Comparison of tenofovir alafenamide and entecavir for hepatitis B virus-related acute-on-chronic liver failure. / å¯Œé©¬é ¸ä¸™é šæ›¿è¯ºç¦éŸ¦ä¸Žæ©æ›¿å¡éŸ¦æ²»ç–—ä¹™åž‹è‚ç‚Žç— æ¯’ç›¸å ³æ ¢åŠ æ€¥æ€§è‚è¡°ç«­çš„æ•ˆæžœå¯¹æ¯”.

OBJECTIVES: Hepatitis B virus related acute-on-chronic liver failure (HBV-ACLF) is the most common type of liver failure in China, with a high mortality. Early rapid reduction of HBV-DNA load can improve the survival rate of HBV-ACLF patients. At present, the commonly used drugs are nucleoside (acid) analogues, such as entecavir (ETV), tenofovir, and so on. The newly listed tenofovir alafenamide fumarate (TAF) has attracted great attention of clinicians because of its stronger antiviral effect, higher transaminase normalization rate, better bone and kidney safety, and zero drug resistance. However, there are few clinical research data on the efficacy and safety of TAF in the treatment of Chinese HBV-ACLF patients, and there is a lack of pharmacoeconomic evaluation. This study aims to compare the efficacy, safety, and cost-effectiveness between TAF and ETV in patients with HBV-ACLF. METHODS: The data were collected from 196 HBV-ACLF patients (80 patients in the TAF group and 116 patients in the ETV group) who were hospitalized in Xiangya Hospital, Central South University from May 2020 to March 2021. Biochemistry and virology were detected before and after treatment (at baseline, Week 2, 4, and 12). Clinical features, disease prognosis, and cost-effectiveness were compared between the 2 groups. According to the baseline, HBV-ACLF patients were divided into 4 stages including pre-liver failure stage, early stage, medium stage, and end stage. And the liver transplantation rate and mortality was also compared. Pharmacoeconomic evaluation was taken using cost-effectiveness analysis and cost minimization analysis．. RESULTS: After 4 weeks of treatment, there were no significant differences in the efficacy (liver function, viral load) between the 2 groups (all P>0.05). The TAF group showed lower creatinine [(80.35±18.77) µmol/L vs (105.59±82.32) µmol/L, P<0.05] and higher estimated glomerular filtration rate (eGFR) levels [(95.65±23.21) mL/(min·1.73 m2) vs (82.68±26.32) mL/(min·1.73 m2), P<0.05] than the ETV group. After 12 weeks of treatment, the analysis of overall the liver transplantation rate and mortality between the 2 groups showed similar conclusion. However, the TAF group had a lower the liver transplantation rate and mortality than the ETV group in patients with pre-liver failure (0vs13.89%, P<0.05). No evident distinction was found in the liver transplantation rate and mortality during the early, medium, or end stages of liver failure (13.04% vs 17.65%, 37.50% vs 37.04%, and 54.55% vs 68.42%, respectively). Ratio of cost to effectiveness in the ETV group was higher than that in the TAF group. CONCLUSIONS: TAF is not more efficient than ETV group in improving liver function and reducing viral load for HBV-ACLF patients and they also show similar safety. However, TAF has a greater advantage over ETV not only in preserving renal function, but also in reducing the liver transplantation rate and mortality in patients with pre-liver failure. TAF can provide economic benefit to patients with HBV-ACLF.

CD24 is expressed on FoxP3+ regulatory T cells and regulates their function.

CD24 is a glycosyl-phosphatidylinositol (GPI) anchored cell surface glycoprotein with a variety of immunomodulatory functions such as inhibition of thymic generation of autoreactive T cells, regulation of antigen presenting cell functions, and mediation of autoimmunity. Given the autoimmune nature of FoxP3+ regulatory T cells and their importance in autoimmune diseases, we hypothesize that CD24 regulates the generation and functions of Treg cells. Through the analysis of the Treg repertoire in two strains of CD24-deficient mice, we found that CD24 does not globally affect the thymic generation of Treg cells. However, CD24 is abundantly expressed on Treg cells, and CD24 antibody treatment of Treg cells enhances their suppressive functions. Concurrently, we observed CD24-deficient Treg cells exhibit increased suppressive functions and produce more IL-10 compared to their wild type counterparts. In addition, CD24-deficient Treg cells exhibited more potent suppressive capacity in inhibiting the development of experimental autoimmune encephalomyelitis (EAE) in mice. Thus, CD24 on Treg cells regulates their suppressive functions. Our findings can partially explain the resistance of EAE development in CD24-deficient mice and CD24 polymorphism-associated susceptibility of human autoimmune diseases. Further investigations regarding mechanisms of CD24 regulation of Treg function may lead to a new approach for the immunotherapy of human autoimmune diseases.

IL-27 Induces CCL5 Production by T Lymphocytes, Which Contributes to Antitumor Activity.

IL-27 is a pleiotropic cytokine that exhibits stimulatory/regulatory functions on multiple lineages of immune cells including T lymphocytes. In this study, we demonstrate that IL-27 directly induces CCL5 production by T lymphocytes, particularly CD8+ T cells in vitro and in vivo. IL-27-induced CCL5 production is IL-27R-dependent. In CD4+ T cells, IL-27-induced CCL5 production was primarily dependent on Stat1 activation, whereas in CD8+ T cells, Stat1 deficiency does not abrogate CCL5 induction. A chromatin immunoprecipitation assay revealed that in the CCL5 promoter region, both putative Stat3 binding sites exhibit significant binding to Stat3, whereas only one out of four Stat1 binding sites displays moderate binding to Stat1. In tumor-bearing mice, IL-27 induced dramatic production of CCL5 in tumor-infiltrating T cells. IL-27-induced CCL5 appears to contribute to an IL-27-mediated antitumor effect. This is signified by diminished tumor inhibition in anti-CCL5- and IL-27-treated mice. Additionally, intratumor delivery of CCL5 mRNA using lipid nanoparticles significantly inhibited tumor growth. Thus, IL-27 induces robust CCL5 production by T cells, which contributes to antitumor activity.

Intratumoral delivery of IL-12 and IL-27 mRNA using lipid nanoparticles for cancer immunotherapy.

Cytokines are important immunotherapeutics with approved drugs for the treatment of human cancers. However, systemic administration of cytokines often fails to achieve adequate concentrations to immune cells in tumors due to dose-limiting toxicity. Thus, developing localized therapy that directly delivers immune-stimulatory cytokines to tumors may improve the therapeutic efficacy. In this study, we generated novel lipid nanoparticles (LNPs) encapsulated with mRNAs encoding cytokines including IL-12, IL-27 and GM-CSF, and tested their anti-tumor activity. We first synthesized ionizable lipid materials containing di-amino groups with various head groups (DALs). The novel DAL4-LNP effectively delivered different mRNAs in vitro to tumor cells and in vivo to tumors. Intratumoral injection of DAL4-LNP loaded with IL-12 mRNA was most potent in inhibiting B16F10 melanoma tumor growth compared to IL-27 or GM-CSF mRNAs in monotherapy. Furthermore, intratumoral injection of dual DAL4-LNP-IL-12 mRNA and IL-27 mRNA showed a synergistic effect in suppressing tumor growth without causing systematic toxicity. Most importantly, intratumoral delivery of IL-12 and IL-27 mRNAs induced robust infiltration of immune effector cells, including IFN-γ and TNF-α producing NK and CD8+ T cells into tumors. Thus, intratumoral administration of DAL-LNP loaded with IL-12 and IL-27 mRNA provides a new treatment strategy for cancer.