BACKGROUND:Hydroxy safflower yellow A has anti-ischemia,anti-oxidation,anti-thrombotic and anti-inflammatory effects.Whether it affects neuronal pyroptosis after glucose-oxygen deprivation/reglucose-reoxygenation is still unclear. OBJECTIVE:To investigate the protective effect of hydroxy safflower yellow A on neuronal pyroptosis and its mechanism. METHODS:HT22 cells in logarithmic growth phase were randomly divided into five groups:normal group,model group,hydroxy safflower yellow A group,colivelin group,and colivelin+hydroxy safflower yellow A group.HT22 cells were treated with glucose-oxygen deprivation/reglucose-reoxygenation to establish neuronal pyroptosis model,and then treated with STAT3 agonist Colivelin and hydroxy safflower yellow A.JC-1 probe was employed to assess changes in mitochondrial membrane potential.Reactive oxygen species kit was used to determine the content of reactive oxygen species in cells.GSDMD/TUNEL staining was conducted to observe cell pyroptosis.Immunofluorescence analysis was performed to detect STAT3 and GSDMD protein expression.RT-PCR was utilized for assessing mRNA expression levels of STAT3,NLRP3,and Caspase-1.Western blot assay was utilized to measure the protein expression levels of p-STAT3,NLRP3,GSDMD,Cleaved-caspase-1,and interleukin-1β. RESULTS AND CONCLUSION:(1)Compared with the normal group,the number of pyroptotic cells increased in HT22 cells in the model group along with a significant increase in protein expression levels of p-STAT3,NLRP3,Cleaved-caspase-1,GSDMD,and interleukin-1β.Compared with the model group,the number of pyroptotic cells reduced,and the expression of pyroptosis-related proteins significantly decreased in the hydroxy safflower yellow A group.(2)In comparison with the model group,pyroptosis worsened in the colivelin group where mitochondrial membrane potential decreased along with elevated reactive oxygen species content and increased mRNA expression levels of STAT3,NLRP3,and Caspase-1,as well as increased protein expression levels of p-STAT3,NLRP3,GSDMD,Cleaved-caspase-1,and interleukin-1β.Compared with the Colivelin group,above indexes were improved in the colivelin+hydroxy safflower yellow A group.These results suggest that hydroxy safflower yellow A plays a neuroprotective role through STAT3 signaling pathway to inhibit HT22 pyroptosis after glucose-oxygen deprivation/reglucose-reoxygenation treatment.

BACKGROUND: The main cause of restenosis after percutaneous transluminal angioplasty (PTA) is the excessive proliferation and migration of vascular smooth muscle cells (VSMCs). Lin28a has been reported to play critical regulatory roles in this process. However, whether CCAAT/enhancer-binding proteins β (C/EBPβ) binds to the Lin28a promoter and drives the progression of restenosis has not been clarified. Therefore, in the present study, we aim to clarify the role of C/EBPβ-Lin28a axis in restenosis. METHODS: Restenosis and atherosclerosis rat models of type 2 diabetes ( n   =  20, for each group) were established by subjecting to PTA. Subsequently, the difference in DNA methylation status and expression of C/EBPβ between the two groups were assessed. EdU, Transwell, and rescue assays were performed to assess the effect of C/EBPβ on the proliferation and migration of VSMCs. DNA methylation status was further assessed using Methyltarget sequencing. The interaction between Lin28a and ten-eleven translocation 1 (TET1) was analysed using co-immunoprecipitation (Co-IP) assay. Student's t -test and one-way analysis of variance were used for statistical analysis. RESULTS: C/EBPβ expression was upregulated and accompanied by hypomethylation of its promoter in restenosis when compared with atherosclerosis. In vitroC/EBPβ overexpression facilitated the proliferation and migration of VSMCs and was associated with increased Lin28a expression. Conversely, C/EBPβ knockdown resulted in the opposite effects. Chromatin immunoprecipitation assays further demonstrated that C/EBPβ could directly bind to Lin28a promoter. Increased C/EBPβ expression and enhanced proliferation and migration of VSMCs were observed after decitabine treatment. Further, mechanical stretch promoted C/EBPβ and Lin28a expression accompanied by C/EBPβ hypomethylation. Additionally, Lin28a overexpression reduced C/EBPβ methylation via recruiting TET1 and enhanced C/EBPβ-mediated proliferation and migration of VSMCs. The opposite was noted in Lin28a knockdown cells. CONCLUSION Our findings suggest that the C/EBPβ-Lin28a axis is a driver of restenosis progression, and presents a promising therapeutic target for restenosis.
Animals ; Cell Proliferation/genetics* ; Cell Movement/genetics* ; Muscle, Smooth, Vascular/metabolism* ; Rats ; DNA Methylation/physiology* ; CCAAT-Enhancer-Binding Protein-beta/genetics* ; Male ; Myocytes, Smooth Muscle/cytology* ; Rats, Sprague-Dawley ; RNA-Binding Proteins/genetics* ; Cells, Cultured ; Coronary Restenosis/metabolism*

ObjectiveTo investigate the incidence rate of primary liver cancer （PLC） and the progression of liver fibrosis in chronic hepatitis B （CHB） patients with low-level viremia （LLV） （HBV DNA<2 000 IU/mL but ≥20 IU/mL） after treatment adjustment， and to provide more robust evidence for clinical practice. MethodsA retrospective analysis was performed for the clinical data of LLV patients who initially received nucleos（t）ide analogue （NAs） for at least 48 weeks at the Fifth Medical Center of PLA General Hospital from August 2007 to April 2017 and subsequently underwent NAs adjustment due to LLV， and according to the virologic response after 48 weeks of treatment adjustment， the patients were divided into LLV group and complete virological response （CVR） group （HBV DNA<20 IU/mL）. The patients were followed up once every 3‍ ‍—‍ ‍6 months till the primary endpoint event of PLC or October 2024. The incidence rate of PLC and the progression of liver fibrosis were observed， and the progression of liver fibrosis was defined as an increase of ≥1 grade in fibrosis-4 （FIB-4） index. The independent-samples t test was used for comparison of normally distributed continuous data between two groups， and the Mann-Whitney U test was used for comparison of continuous data with skewed distribution between two groups； the chi-square test was used for comparison of categorical data between groups. The Kaplan-Meier method was used to calculate the cumulative incidence rate of PLC， and the Log-rank test was used for comparison between groups； the Cox regression analysis was used to investigate the risk factors for PLC， and the Logistic regression analysis was used to investigate the influencing factors for the progression of liver fibrosis. ResultsA total of 307 patients were enrolled， with a mean age of 50.0 years， and the male patients accounted for 80.5%. After 48 weeks of treatment with the adjusted NAs regimen， 254 patients （82.7%） achieved CVR， and 53 patients （17.3%） still had LLV. For the LLV group， the incidence rate of PLC was 30.2% and the rate of liver fibrosis progression was 22.6%， while for the CVR group， the incidence rate of PLC was only 13.4%， and the rate of liver fibrosis progression was 7.5%. The multivariate regression analyses showed that LLV was an independent risk factor for the onset of PLC （hazard ratio=2.623， 95% confidence interval ［CI］： 1.315‍ ‍—‍ ‍5.234， P=0.006） and the progression of liver fibrosis （odds ratio=3.213， 95%CI： 1.385‍ ‍—‍ ‍7.455， P=0.007）. ConclusionActive adjustment of treatment is needed immediately after the diagnosis of LLV to improve CVR， and if LLV persists after treatment adjustment， it is necessary to enhance the monitoring of liver fibrosis progression and PLC， so as to facilitate early diagnosis and treatment.

Extensive epigenetic reprogramming involves in memory CD8+ T-cell differentiation. The elaborate epigenetic rewiring underlying the heterogeneous functional states of CD8+ T cells remains hidden. Here, we profile single-cell chromatin accessibility and map enhancer-promoter interactomes to characterize the differentiation trajectory of memory CD8+ T cells. We reveal that under distinct epigenetic regulations, the early activated CD8+ T cells divergently originated for short-lived effector and memory precursor effector cells. We also uncover a defined epigenetic rewiring leading to the conversion from effector memory to central memory cells during memory formation. Additionally, we illustrate chromatin regulatory mechanisms underlying long-lasting versus transient transcription regulation during memory differentiation. Finally, we confirm the essential roles of Sox4 and Nrf2 in developing memory precursor effector and effector memory cells, respectively, and validate cell state-specific enhancers in regulating Il7r using CRISPR-Cas9. Our data pave the way for understanding the mechanism underlying epigenetic memory formation in CD8+ T-cell differentiation.
CD8-Positive T-Lymphocytes/metabolism* ; Cell Differentiation ; Chromatin/immunology* ; Animals ; Mice ; Immunologic Memory ; Epigenesis, Genetic ; SOXC Transcription Factors/immunology* ; NF-E2-Related Factor 2/immunology* ; Mice, Inbred C57BL ; Gene Regulatory Networks ; Enhancer Elements, Genetic

Background Chrysotile is widely used in construction and industry. Research has shown that it is associated with lung fibrosis in occupational groups, but the involvement of microRNAs (miRNAs) in chrysotile-induced lung fibrosis has been less well studied, and the specific mechanism is still unclear. Objective Using next-generation sequencing technology to analyze the effects of chrysotile exposure on the miRNAs expression profiles of human lung epithelial cells (BEAS-2B cells), to explore the variations of differentially expressed miRNAs and related signaling pathways, and to identify potential targets and molecular mechanisms of chrysotile-induced lung fibrosis. Methods Chrysotile was analyzed with a laser particle size analyzer and an X-ray diffractometer for particle size and physical phase. BEAS-2B cells were exposed to chrysotile for designed time sessions (12, 24, and 48 h) and doses (0, 50, 100, and 200 μg·mL⁻¹). Cell viability was detected with a cell viability assay kit (CCK8); expression levels of Fibronectin, Collagen-Ⅰ, and α-smooth muscle actin (α-SMA) were detected by Western blot after exposure to 200 μg·mL⁻¹ chrysotile for 24 h. Sample correlation and changes in miRNAs expression profiles between the chrysotile-exposed and the control groups were analyzed by next-generation sequencing technology. The target genes of differentially expressed miRNAs were predicted and subjected to Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. Results The average particle size of the chrysotile dust sample used in this study was 3.58 μm, and the results of X-ray diffraction analysis confirmed the characteristic peaks of chrysotile. Compared with the control group, the chrysotile gradually inhibited the survival rate of BEAS-2B cells with increasing concentration and exposure time (P<0.01). The survival rates of the 50, 100, and 200 μg·mL⁻¹ chrysotile-exposed cells after 12 h exposure were 83.88%±1.86%, 78.07%±3.97%, and 71.95%±2.99%, respectively; the survival rates after 24 h exposure were 77.41%±1.58%, 69.57%±2.23%, and 62.79%±3.65%, respectively; the survival rates after 48 h exposure were 74.31%±4.93%, 65.84%±2.71%, and 52.74%±6.31%, respectively. The Fibronectin, Collagen-Ⅰ, and α-SMA protein expression levels were elevated in the 200 μg·mL⁻¹ chrysotile-exposed BEAS-2B cells (P <0.05). The results of principal component analysis showed that there were differences in the composition of the samples between the chrysotile exposure group and the control group, and a total of 163 differential miRNAs were screened, of which 79 were up-regulated and 84 were down-regulated. The results of GO analysis showed that the differential miRNAs were mainly associated with biological processes such as regulation of transcription by RNA polymerase II, regulation of DNA templated transcription, cellular differentiation, protein phosphorylation, lipid metabolism, and cell cycle, cellular components such as nucleus, cytomembrane, cytoskeleton, mitochondria, and endoplasmic reticulum, as well as molecular functions such as protein binding, metal ion binding, transferase activity, and DNA binding. The results of KEGG analysis revealed that the differential miRNAs were mainly enriched in cancer pathway, phosphatidylinositol 3-kinase/ protein kinase B (PI3K/AKT) pathway, Ras-associated protein 1 (Rap1) pathway, calcium pathway, cyclic guanosine monophosphate/ protein kinase G (cGMP-PKG) pathway, Hippo pathway, cyclic adenosine monophosphate (cAMP) pathway, and Ras pathway. Conclusion Chrysotile exposure could significantly inhibit BEAS-2B cell survival, elevate the expression of lung fibrosis-associated proteins, and induce differential miRNAs expression, affecting biological processes (such as lipid metabolism, protein phosphorylation, and cell cycle) and cell components (such as mitochondria and endoplasmic reticulum), and interfering with PI3K/AKT pathway, Hippo pathway, cAMP pathway, Rap1 pathway, and Ras pathway.

OBJECTIVE To investigate the protective effect proanthocyanidin B2(PC-B2) on oxidative damage and apoptosis of mouse astrocytes(AS) induced by hydrogen peroxide(H2O2) and its mechanism. METHODS AS were isolated and cultured from neonatal C57BL/6 mice(1−3 d). The optimal concentration of H2O2 and PC-B2 was divided into four groups: normal group, normal+PC-B2 group(100 μg·mL‒1 PC-B2 treated for 24 h), H2O2 model group(200 μmol·L‒1 H2O2 treated for 24 h), PC-B2 group(200 μmol·L‒1 H2O2 and 100 μg·mL‒1 PC-B2 treated for 24 h). The cell viability of each group was detected by CCK-8 method. Cytotoxicity was detected by LDH method. The antioxidant capacity was detected by ABTS and DPPH. The content of MDA and the activity of SOD, CAT and GSH-Px were detected by ELISA kit. Detection of apoptosis in each group was done by TUNEL staining. The mRNA and protein expression levels of Bax, Bcl-2, Caspase-3, Akt/Stat3, p-Akt, p-Stat3 and Nrf2/HO-1 in AS were detected by RT-PCR and Western blotting, respectively. RESULTS PC-B2 could significantly enhance cell viability and inhibit AS apoptosis. Compared with the H2O2 model group, PC-B2 intervention could significantly reduce the content of LDH and MDA in AS, and increase the activity of SOD, CAT and GSH-Px. PC-B2 intervention could inhibit the mRNA and protein expression of Bax and Caspase-3, and up-regulate the mRNA and protein expression of Akt/Stat3, Bcl-2, Nrf2/HO-1. CONCLUSION PC-B2 can enhance the antioxidant capacity of AS through Akt/Stat3 and Nrf2/HO-1 pathways, therefore reduce H2O2-induced AS oxidative damage and apoptosis.

Objective:To explore the value of a new modified T3 sub-staging for the prognosis evaluation in gallbladder cancer patients.Methods:This is a retrospective case-series study. The clinical data of patients with pathologically confirmed stage T3 gallbladder cancer who were admitted to the Department of Hepatobiliary Surgery,the First Affiliated Hospital of Xi′an Jiaotong University from January 2011 to December 2021 were retrospectively analyzed. A total of 190 patients were enrolled in this study, 67 males and 123 females, with an age ( M(IQR)) of 63(14) years (range:17 to 88 years). The stage T3 was divided into four sub-stages according to the site of tumor invasion: (1) T3a:tumor perforates the serosa,but not invading the liver and one other adjacent structure; (2) T3b:tumor perforates the serosa and invades one other adjacent structure,but not the liver; (3) T3c:tumor perforates the serosa and invades the liver,but not one other adjacent structure; (4) T3d:tumor perforates the serosa,invades the liver and one other adjacent structure. To evaluate the application value of this modified sub-staging,the Kaplan-Meier method was used to draw the survival curve,univariate analysis and multivariate analysis were done using the Log-rank test and Cox proportional hazard model respectively. Results:According to the modified T3 sub-staging method,34 patients (17.9%) were in stage T3a,24 cases(12.6%) were in stage T3b, 97 cases (51.1%) were in stage T3c, and 35 cases (18.4%) were in stage T3d. The median survival time of patients in stages T3a,T3b,T3c and T3d after radical resection was 72.0 months, 32.0 months, 12.0 months and 10.0 months, respectively. The 1-, 3-, and 5-year survival rates of patients in stage T3a, T3b, T3c and T3d were 79.4%, 53.3%, and 53.3%; 79.2%, 44.6%, and 26.0%;49.5%,27.5%,and 18.1%;42.9%,15.9%, and 15.9% ( χ2=18.349, P<0.01),respectively. Univariate analysis showed that gallbladder stones,pathological differentiation,perineural invasion, N stage,postoperative adjuvant therapy and modified T3 substage were factors affecting patient prognosis(all P<0.05). Cox multivariate analysis showed that modified sub-stages with T3c ( HR=2.043, 95% CI:1.176 to 3.549) and T3d( HR=2.419, 95% CI:1.284 to 4.555), accompanied by gallbladder stones ( HR=1.661,95% CI:1.150 to 2.398),pathological differentiation with poorly differentiated( HR=1.709,95% CI:1.198 to 2.438), and the N stage with N1 and N2( HR=1.602, 95% CI:1.090 to 2.355, 2.714, 95% CI: 1.621 to 4.544) were independent prognostic risk factors for patients in stage T3,while postoperative adjuvant chemotherapy( HR=0.351) was a protective factor for prognosis. There was no statistically significant difference in survival between patients with stage T3a and T3b who underwent hepatic wedge resection and liver segment or major resection ( P=0.402). For patients with stage T3c and T3d with liver invasion,the survival difference after hepatic wedge resection and segmental or major resection was statistically significant ( P=0.008). Conclusion:The modified T3 sub-staging system based on the depth and direction of tumor invasion maybe helpful to further stratify the prognosis of patients with gallbladder cancer.

Objective:To explore the value of a new modified T3 sub-staging for the prognosis evaluation in gallbladder cancer patients.Methods:This is a retrospective case-series study. The clinical data of patients with pathologically confirmed stage T3 gallbladder cancer who were admitted to the Department of Hepatobiliary Surgery,the First Affiliated Hospital of Xi′an Jiaotong University from January 2011 to December 2021 were retrospectively analyzed. A total of 190 patients were enrolled in this study, 67 males and 123 females, with an age ( M(IQR)) of 63(14) years (range:17 to 88 years). The stage T3 was divided into four sub-stages according to the site of tumor invasion: (1) T3a:tumor perforates the serosa,but not invading the liver and one other adjacent structure; (2) T3b:tumor perforates the serosa and invades one other adjacent structure,but not the liver; (3) T3c:tumor perforates the serosa and invades the liver,but not one other adjacent structure; (4) T3d:tumor perforates the serosa,invades the liver and one other adjacent structure. To evaluate the application value of this modified sub-staging,the Kaplan-Meier method was used to draw the survival curve,univariate analysis and multivariate analysis were done using the Log-rank test and Cox proportional hazard model respectively. Results:According to the modified T3 sub-staging method,34 patients (17.9%) were in stage T3a,24 cases(12.6%) were in stage T3b, 97 cases (51.1%) were in stage T3c, and 35 cases (18.4%) were in stage T3d. The median survival time of patients in stages T3a,T3b,T3c and T3d after radical resection was 72.0 months, 32.0 months, 12.0 months and 10.0 months, respectively. The 1-, 3-, and 5-year survival rates of patients in stage T3a, T3b, T3c and T3d were 79.4%, 53.3%, and 53.3%; 79.2%, 44.6%, and 26.0%;49.5%,27.5%,and 18.1%;42.9%,15.9%, and 15.9% ( χ2=18.349, P<0.01),respectively. Univariate analysis showed that gallbladder stones,pathological differentiation,perineural invasion, N stage,postoperative adjuvant therapy and modified T3 substage were factors affecting patient prognosis(all P<0.05). Cox multivariate analysis showed that modified sub-stages with T3c ( HR=2.043, 95% CI:1.176 to 3.549) and T3d( HR=2.419, 95% CI:1.284 to 4.555), accompanied by gallbladder stones ( HR=1.661,95% CI:1.150 to 2.398),pathological differentiation with poorly differentiated( HR=1.709,95% CI:1.198 to 2.438), and the N stage with N1 and N2( HR=1.602, 95% CI:1.090 to 2.355, 2.714, 95% CI: 1.621 to 4.544) were independent prognostic risk factors for patients in stage T3,while postoperative adjuvant chemotherapy( HR=0.351) was a protective factor for prognosis. There was no statistically significant difference in survival between patients with stage T3a and T3b who underwent hepatic wedge resection and liver segment or major resection ( P=0.402). For patients with stage T3c and T3d with liver invasion,the survival difference after hepatic wedge resection and segmental or major resection was statistically significant ( P=0.008). Conclusion:The modified T3 sub-staging system based on the depth and direction of tumor invasion maybe helpful to further stratify the prognosis of patients with gallbladder cancer.

The technology of three-dimensional(3D)printing emerged in the late 1970s and has since undergone considerable development to find numerous applications in mechanical engineering,industrial design,and biomedicine.In biomedical science,several studies have initially found that 3D printing technology can play an important role in the treatment of diseases in hepatopancreatobiliary surgery.For example,3D printing technology has been applied to create detailed anatomical models of disease organs for preoperative personalized surgical strategies,surgical simulation,intraoperative navigation,medical training,and patient education.Moreover,cancer models have been created using 3D printing technology for the research and selection of chemotherapy drugs.With the aim to clarify the development and application of 3D printing technology in hepatopancreatobiliary surgery,we introduce seven common types of 3D printing technology and review the status of research and application of 3D printing technology in the field of hepatopancreatobiliary surgery.

Objective To explore the CT manifestations and clinical features of checkpoint inhibitor-related pneumonitis(CIP).Methods Chest CT images and clinical data of 34 patients with CIP in Ningbo No.2 Hospitael were collected to retrospectively analysis.According to the site of tumor occurrence,22 patients were divided into lung cancer group and 12 patients in other malignant tumor group,and the differences in CT manifestations between two groups were compared.Results Cough(70.59％)and dyspnea(52.94％)were the common clinical symptoms.CIP occurred earlier and over a greater time span in lung cancer group 114.5(41.50,281.50)d than in other maligment tumor group 144(55.75,226.25)d.Eosinophil count was significantly higher only in other maligment tumor group(P=0.009).After hormonal therapy 18 patients improved,8 were stable and 8 progressed or even died.CT signs were prevalent in ground glass shadow(70.59％)and solid shadow(76.47％),and the imaging pattern was dominated by organic pneumonia pattern(47.6％),which was not related to type of primary tumor,and some of them could show nodular granulomatous reaction.Compared to lung cancer group,the other maligment tumor group was more likely to exhibit symmetrical infiltration(58.33％)distribution.Conclusion The clinical features of CIP are nonspecific,compared with other patients with primary malignancies,lung cancer patients are more likely to develop CIP,which is difficult to relieve after treatment,and are easy to develop severe disease.