Aim To investigate the effect of N6-methy-ladenosine(m6A)demethylase ALKBH5 on the prolif-eration and migration of cardiac fibroblasts(CFs)in-duced by high glucose.Methods Primary CFs were isolated from neonatal mouse hearts and identified u-sing optical and confocal microscopy.Cell activation was induced using a high-glucose medium(33 mmol·L-1 glucose).An ALKBH5 overexpression model was established by transfecting CFs with an ALKBH5 ex-pression vector in a high-glucose medium.The expres-sion of ALKBH5 in CFs was assessed through immuno-fluorescence staining,Western blot and RT-qPCR.Changes in m6A levels were evaluated using Dot blot a-nalysis.Additionally,Alterations in the expression of proliferating cell nuclear antigen(PCNA)and collagenⅠ,a pivotal fibrosis indicator,were measured using Western blot.The proliferation and migration ability of CFs were assessed through EdU staining and Transwell migration assay,respectively.Results Following treatment with high glucose,the expression of ALKBH5 in CFs notably decreased,while m6A level increased.This was accompanied by a significant increase in the expression of the proliferation marker PCNA and the fi-brosis marker collagen Ⅰ.Additionally,there was a sig-nificant improvement in the ability of proliferation and migration.Overexpression of ALKBH5 resulted in a significant decrease in the expressions of PCNA and collagen Ⅰ,leading to the inhibition of both proliferation and migration in CFs.Conclusion Overexpression of ALKBH5 suppresses the expression of PCNA and colla-gen Ⅰ,consequently reducing the proliferation and mi-gration of CFs,potentially through m6A methylation modification.

Objective To explore the role of secreted phosphoprotein 1(SPP1)and myeloid differentiation primary response 88(MYD88)in morphine-induced cardiomyocyte apoptosis.Methods A morphine addiction model was established in Sprague-Dawley(SD)rats.Twelve SD rats were randomly assigned to the normal saline(NS)group or the morphine-dependent(DAM)group.Histopathological analysis was employed to observe and compare myocardial tissue morphology between the two groups.Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling(TUNEL)staining was performed to assess the number of apoptotic cells in each group.The expression levels of SPP1 and MYD88 were evaluated using immunohistochemistry.Quantitative real-time poly merase chain reaction(RT-qPCR)and Western blot were used to detect the mRNA and protein expression of SPP1,MYD88,Bax,Bcl2,Caspase-3,and Caspase-9.Simultaneously,Western blot analysis was used to detected the expression of Cleaved Caspase-3 and Cleaved Caspase-9 proteins.In vitro,SPP1 expression was knocked down in primary neonatal rat cardiomyocytes(NRCMs),and cells were divided into three groups:control(CON),morphine treated(DA),and shSPP1#3+DA.Cell viability was assessed using the CCK-8 assay,and apoptosis rates were determined by flow cytometry.Results HE and TUNEL staining of myocardial tissues from morphine-addicted SD rats revealed that,compared with the NS group,myofibrils in the DAM group exhibited partial disruption and a significant increase in apoptotic cells(P<0.05).Western blot and RT-qPCR analyses demonstrated that,relative to the NS group,the mRNA and protein levels of SPP1,MYD88,Bax,Caspase-3,and Caspase-9 were significantly upregulated in the DAM group(P<0.05),whereas Bcl2 expression was significantly downregulated at both mRNA and protein levels(P<0.05),and the protein expression levels of Cleaved Caspase-3 and Cleaved Caspase-9 were also increased.with all differences being statistically significant.In NRCMs following morphine intervention,cell viability in the DA group was markedly reduced compared to the CON group(P<0.05),accompanied by a signifi-cant increase in apoptosis rate(P<0.05).Consistently,Western blot and RT-qPCR results showed elevated mRNA and protein expression of SPP1,MYD88,Bax,Caspase-3,and Caspase-9 in the DA group(P<0.05),along with decreased Bcl2 expression(P<0.05).The protein expression levels of Cleaved Caspase-3 and Cleaved Caspase-9 were elevated simultaneously.In contrast,the shSPP1#3+DA group exhibited opposing trends compared to the DA group,with statistically sig nificant differences(P<0.05).Conclusion SPP1 and MYD88 play critical roles in mediating morphine-induced cardiomyocyte apoptosis,and silencing SPP1 has been shown to significantly reduce the extent of cardiomyocyte apoptosis following morphine exposure.

Aim To investigate the effect of N6-methy-ladenosine(m6A)demethylase ALKBH5 on the prolif-eration and migration of cardiac fibroblasts(CFs)in-duced by high glucose.Methods Primary CFs were isolated from neonatal mouse hearts and identified u-sing optical and confocal microscopy.Cell activation was induced using a high-glucose medium(33 mmol·L-1 glucose).An ALKBH5 overexpression model was established by transfecting CFs with an ALKBH5 ex-pression vector in a high-glucose medium.The expres-sion of ALKBH5 in CFs was assessed through immuno-fluorescence staining,Western blot and RT-qPCR.Changes in m6A levels were evaluated using Dot blot a-nalysis.Additionally,Alterations in the expression of proliferating cell nuclear antigen(PCNA)and collagenⅠ,a pivotal fibrosis indicator,were measured using Western blot.The proliferation and migration ability of CFs were assessed through EdU staining and Transwell migration assay,respectively.Results Following treatment with high glucose,the expression of ALKBH5 in CFs notably decreased,while m6A level increased.This was accompanied by a significant increase in the expression of the proliferation marker PCNA and the fi-brosis marker collagen Ⅰ.Additionally,there was a sig-nificant improvement in the ability of proliferation and migration.Overexpression of ALKBH5 resulted in a significant decrease in the expressions of PCNA and collagen Ⅰ,leading to the inhibition of both proliferation and migration in CFs.Conclusion Overexpression of ALKBH5 suppresses the expression of PCNA and colla-gen Ⅰ,consequently reducing the proliferation and mi-gration of CFs,potentially through m6A methylation modification.

Objective To explore the role of secreted phosphoprotein 1(SPP1)and myeloid differentiation primary response 88(MYD88)in morphine-induced cardiomyocyte apoptosis.Methods A morphine addiction model was established in Sprague-Dawley(SD)rats.Twelve SD rats were randomly assigned to the normal saline(NS)group or the morphine-dependent(DAM)group.Histopathological analysis was employed to observe and compare myocardial tissue morphology between the two groups.Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling(TUNEL)staining was performed to assess the number of apoptotic cells in each group.The expression levels of SPP1 and MYD88 were evaluated using immunohistochemistry.Quantitative real-time poly merase chain reaction(RT-qPCR)and Western blot were used to detect the mRNA and protein expression of SPP1,MYD88,Bax,Bcl2,Caspase-3,and Caspase-9.Simultaneously,Western blot analysis was used to detected the expression of Cleaved Caspase-3 and Cleaved Caspase-9 proteins.In vitro,SPP1 expression was knocked down in primary neonatal rat cardiomyocytes(NRCMs),and cells were divided into three groups:control(CON),morphine treated(DA),and shSPP1#3+DA.Cell viability was assessed using the CCK-8 assay,and apoptosis rates were determined by flow cytometry.Results HE and TUNEL staining of myocardial tissues from morphine-addicted SD rats revealed that,compared with the NS group,myofibrils in the DAM group exhibited partial disruption and a significant increase in apoptotic cells(P<0.05).Western blot and RT-qPCR analyses demonstrated that,relative to the NS group,the mRNA and protein levels of SPP1,MYD88,Bax,Caspase-3,and Caspase-9 were significantly upregulated in the DAM group(P<0.05),whereas Bcl2 expression was significantly downregulated at both mRNA and protein levels(P<0.05),and the protein expression levels of Cleaved Caspase-3 and Cleaved Caspase-9 were also increased.with all differences being statistically significant.In NRCMs following morphine intervention,cell viability in the DA group was markedly reduced compared to the CON group(P<0.05),accompanied by a signifi-cant increase in apoptosis rate(P<0.05).Consistently,Western blot and RT-qPCR results showed elevated mRNA and protein expression of SPP1,MYD88,Bax,Caspase-3,and Caspase-9 in the DA group(P<0.05),along with decreased Bcl2 expression(P<0.05).The protein expression levels of Cleaved Caspase-3 and Cleaved Caspase-9 were elevated simultaneously.In contrast,the shSPP1#3+DA group exhibited opposing trends compared to the DA group,with statistically sig nificant differences(P<0.05).Conclusion SPP1 and MYD88 play critical roles in mediating morphine-induced cardiomyocyte apoptosis,and silencing SPP1 has been shown to significantly reduce the extent of cardiomyocyte apoptosis following morphine exposure.

A new siderophore chelate (1) and 8 known compounds were identified from the liquid co-cultures of the marine-derived Streptomyces sp. IMB18-531 and Cladosporium sp. IMB19-099 by a combination of chromatography methods, including C18 reversed-phase medium pressure chromatography, gel column chromatography and HPLC. Their structures were determined by spectroscopic analysis and chemical methods as aluminioxamine E (1), desferrioxamine E (2), ferrioxamine E (3), terragine E (4), capsimicin (5), cyclo(L-prolinyl-L-tyrosine) (6), anthranilic acid (7), (Z)-14-methylpentadec-9-enoic acid (8), and (Z)-hexadec-8-enoic acid (9). Compound 2 showed inhibitory activities against the expression of liver fibrosis related genes COL1A1, MMP2, and TIMP2. Compounds 5, 8, and 9 displayed antibacterial activities against methicillin-resistant Staphylococcus aureus, S. epidermidis and Bacillus subtilis, with MICs of 16-64 μg·mL^-1. Compound 5 showed cytotoxicities against human pancreatic cancer MIA Paca-2 and human colon cancer HT-29 cell lines with IC₅₀ of 2.9 and 6.3 μmol·L^-1, respectively.

Aim To investigate the effect of DNA methyltransferase 3A (DNMT3A) on the proliferation and migration of cardiac fibroblasts (CFs) in C57 mice under high glucose environment. Methods The hearts of C57 mice were taken from 1 to 3 days. After cutting and digesting, CFs were extracted by differential adherance centrifugattion and observed under microscope. After cell attachment, the cells were cultured under low glucose (5.5 mmol • L

Objective: To investigate the clinical features and long-term prognosis of primary biliary cholangitis (PBC) in patients with past hepatitis B virus (HBV) infection. Methods: 353 cases with PBC who visited the Liver Disease Center of Beijing Friendship Hospital Affiliated to Capital Medical University between January 2000 and January 2018 were retrospectively analyzed and were divided into the past HBV infection group (156 cases) and the no HBV infection group (197 cases). The two groups' baseline clinical features were compared. Ursodeoxycholic acid response rate after one year, GLOBE score, UK-PBC score, and long-term liver transplantation-free survival rate were compared through outpatient and telephone follow-up. Results: PBC with past HBV infection had a significantly reduced female proportion compared to the no HBV infection group (91.9% vs. 79.5%, P = 0.001). However, there were no statistically significant differences in age, biochemical indices, immunological indicators, platelet count, cirrhosis proportion, and others. Ursodeoxycholic acid biochemical response rate was reduced in patients with past HBV infection at the end of one year of treatment, but the difference was not statistically significant (65.8% vs. 78.2%, P = 0.068). In addition, there were no statistically significant differences between the GLOBE score (0.57 vs. 0.59, P = 0.26) and UK-PBC 5-year (2.87% vs. 2.87%, P = 0.38), 10-year (9.29% vs. 8.2%, P = 0.39) and 15-year liver transplantation rates (16.6% vs. 14.73%, P = 0.39). Lastly, the overall 5-year liver transplantation-free survival rate had no statistically significant difference between the two groups of patients (86.4% vs. 87.5%, P = 0.796). Conclusion: Primary biliary cholangitis had no discernible effect in terms of age at onset, biochemical indices, immunological indicators, cirrhosis proportion, ursodeoxycholic acid response rate after one year, GLOBE score, UK-PBC score, or overall liver transplantation-free survival rate in patients with past hepatitis B virus infections.

Objective: To explore the efficacy of chemotherapy re-challenge in the third-line setting for patients with metastatic colorectal cancer (mCRC) in the real world. Methods: The clinicopathological data, treatment information, recent treatment efficacy, adverse events and survival data of mCRC patients who had disease progression after treatment with oxaliplatin-based and/or irinotecan-based chemotherapy and received third-line chemotherapy re-challenge from January 2013 to December 2020 at Tianjin Medical University Cancer Institute and Hospital were retrospectively collected. Survival curves were plotted with the Kaplan-Meier method, and the Cox proportional hazard model was used to analyze the prognostic factors. Results: A total of 95 mCRC patients were included. Among them, 32 patients (33.7%) received chemotherapy alone and 63 patients (66.3%) received chemotherapy combined with targeted drugs. Eighty-three patients were treated with dual-drug chemotherapy (87.4%), including oxaliplatin re-challenge in 35 patients and irinotecan re-challenge in 48 patients. The remaining 12 patients were treated with triplet chemotherapy regimens (12.6%). Among them, as 5 patients had sequential application of oxaliplatin and irinotecan in front-line treatments, their third-line therapy re-challenged both oxaliplatin and irinotecan; 7 patients only had oxaliplatin prescription before, and these patients re-challenged oxaliplatin in the third-line treatment. The overall response rate (ORR) and disease control rate (DCR) reached 8.6% (8/93) and 61.3% (57/93), respectively. The median progression free survival (mPFS) and median overall survival (mOS) were 4.9 months and 13.0 months, respectively. The most common adverse events were leukopenia (34.7%) and neutropenia (34.7%), followed by gastrointestinal adverse reactions such as nausea (32.6%) and vomiting (31.6%). Grade 3-4 adverse events were mostly hematological toxicity. Cox multivariate analysis showed that gender (HR=1.609, 95% CI: 1.016-2.548) and the PFS of front-line treatments (HR=0.598, 95% CI: 0.378-0.947) were independent prognostic factors. Conclusion: The results suggested that it is safe and effective for mCRC patients to choose third-line chemotherapy re-challenge, especially for patients with a PFS of more than one year in front-line treatments.
Humans ; Irinotecan/therapeutic use* ; Oxaliplatin/therapeutic use* ; Colorectal Neoplasms/pathology* ; Retrospective Studies ; Fluorouracil ; Colonic Neoplasms/chemically induced* ; Rectal Neoplasms/drug therapy* ; Antineoplastic Combined Chemotherapy Protocols/adverse effects* ; Camptothecin/adverse effects*

Objective: To explore the efficacy of chemotherapy re-challenge in the third-line setting for patients with metastatic colorectal cancer (mCRC) in the real world. Methods: The clinicopathological data, treatment information, recent treatment efficacy, adverse events and survival data of mCRC patients who had disease progression after treatment with oxaliplatin-based and/or irinotecan-based chemotherapy and received third-line chemotherapy re-challenge from January 2013 to December 2020 at Tianjin Medical University Cancer Institute and Hospital were retrospectively collected. Survival curves were plotted with the Kaplan-Meier method, and the Cox proportional hazard model was used to analyze the prognostic factors. Results: A total of 95 mCRC patients were included. Among them, 32 patients (33.7%) received chemotherapy alone and 63 patients (66.3%) received chemotherapy combined with targeted drugs. Eighty-three patients were treated with dual-drug chemotherapy (87.4%), including oxaliplatin re-challenge in 35 patients and irinotecan re-challenge in 48 patients. The remaining 12 patients were treated with triplet chemotherapy regimens (12.6%). Among them, as 5 patients had sequential application of oxaliplatin and irinotecan in front-line treatments, their third-line therapy re-challenged both oxaliplatin and irinotecan; 7 patients only had oxaliplatin prescription before, and these patients re-challenged oxaliplatin in the third-line treatment. The overall response rate (ORR) and disease control rate (DCR) reached 8.6% (8/93) and 61.3% (57/93), respectively. The median progression free survival (mPFS) and median overall survival (mOS) were 4.9 months and 13.0 months, respectively. The most common adverse events were leukopenia (34.7%) and neutropenia (34.7%), followed by gastrointestinal adverse reactions such as nausea (32.6%) and vomiting (31.6%). Grade 3-4 adverse events were mostly hematological toxicity. Cox multivariate analysis showed that gender (HR=1.609, 95% CI: 1.016-2.548) and the PFS of front-line treatments (HR=0.598, 95% CI: 0.378-0.947) were independent prognostic factors. Conclusion: The results suggested that it is safe and effective for mCRC patients to choose third-line chemotherapy re-challenge, especially for patients with a PFS of more than one year in front-line treatments.
Humans ; Irinotecan/therapeutic use* ; Oxaliplatin/therapeutic use* ; Colorectal Neoplasms/pathology* ; Retrospective Studies ; Fluorouracil ; Colonic Neoplasms/chemically induced* ; Rectal Neoplasms/drug therapy* ; Antineoplastic Combined Chemotherapy Protocols/adverse effects* ; Camptothecin/adverse effects*

OBJECTIVE: To investigate the serum microRNA (miRNA) expression and examine the impact of miRNA expression profiles on T helper type 17 (Th17)/regulatory T cells (Treg) imbalance among patients with cystic echinococcosis, so as to provide insights into the illustration of the mechanisms underlying chronic Echinococcus granulosus infections, and long-term pathogenesis. METHODS: Total RNA was extracted from the sera of cystic echinococcosis patients and healthy controls, and subjected to high-throughput sequencing with the Illumina sequencing platform. Known miRNAs were annotated and new miRNAs were predicted using the miRBase database and the miRDeep2 tool, and differentially expressed miRNAs were identified. The target genes of differentially expressed miRNAs were predicted using the software miRanda and TargetScan, and the intersection was selected for Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Among the differentially expressed miRNAs with the 20 highest fold changes, miRNAs that targeted genes relating to key transcription factors RORC and FOXP3 that determine the production of Th17 and Treg cells or their important regulatory pathways (PI3K-Akt and mTOR pathways) were matched. RESULTS: A total of 53 differentially expressed miRNAs were screened in sera of cystic echinococcosis patients and healthy controls, including 47 up-regulated miRNAs and 6 down-regulated miRNAs. GO enrichment analysis showed that these differentially expressed miRNA were involved DNA transcription and translation, cell components, cell morphology, neurodevelopment and metabolic decomposition, and KEGG pathway analysis showed that the differentially expressed miRNA were mainly involved in MAPK, PI3K-Akt and mTOR signaling pathways. Among the differentially expressed miRNAs with the 20 highest fold changes, there were 3 miRNAs that had a potential for target regulation of RORC, and 15 miRNAs that had a potential to target the PI3K-Akt and mTOR signaling pathways. CONCLUSIONS Significant changes are found in serum miRNA expression profiles among patients with E. granulosus infections, and differentially expressed miRNAs may lead to Th17/Treg imbalance through targeting the key transcription factors of Th17/Treg or PI3K-Akt and mTOR pathways, which facilitates the long-term parasitism of E. granulosus in hosts and causes a chronic disease.
Echinococcosis/genetics* ; Gene Expression Profiling ; Humans ; MicroRNAs/metabolism* ; Phosphatidylinositol 3-Kinases/genetics* ; Proto-Oncogene Proteins c-akt/genetics* ; T-Lymphocytes, Regulatory ; TOR Serine-Threonine Kinases/genetics* ; Th17 Cells ; Transcription Factors/genetics*