Objective To identify the N6-methyladenosine (m6A)-related characteristic genes analyzed by gene clustering and immune cell infiltration in myocardial ischemia-reperfusion injury (MI/RI) after cardiopulmonary bypass through machine learning. Methods The differential genes associated with m6A methylation were screened by the dataset GSE132176 in GEO, the samples of the dataset were clustered based on the differential gene expression profile, and the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of the differential genes of the m6A cluster after clustering were performed to determine the gene function of the m6A cluster. R software was used to determine the better models in machine learning of support vector machine (SVM) model and random forest (RF) model, which were used to screen m6A-related characteristic genes in MI/RI, and construct characteristic gene nomogram to predict the incidence of disease. R software was used to analyze the correlation between characteristic genes and immune cells, and the online website was used to build a characteristic gene regulatory network. Results In this dataset, a total of 5 m6A-related differential genes were screened, and the gene expression profiles were divided into two clusters for cluster analysis. The enrichment analysis of m6A clusters showed that these genes were mainly involved in regulating monocytes differentiation, response to lipopolysaccharides, response to bacteria-derived molecules, cellular response to decreased oxygen levels, DNA transcription factor binding, DNA-binding transcription activator activity, RNA polymerase Ⅱ specificity, NOD-like receptor signaling pathway, fluid shear stress and atherosclerosis, tumor necrosis factor signaling pathway, interleukin-17 signaling pathway. The RF model was determined by R software as the better model, which determined that METTL3, YTHDF1, RBM15B and METTL14 were characteristic genes of MI/RI, and mast cells, type 1 helper lymphocytes (Th1), type 17 helper lymphocytes (Th17), and macrophages were found to be associated with MI/RI after cardiopulmonary bypass in immune cell infiltration. Conclusion The four characteristic genes METTL3, YTHDF1, RBM15B and METTL14 are obtained by machine learning, while cluster analysis and immune cell infiltration analysis can better reveal the pathophysiological process of MI/RI.

BACKGROUND:At present,open reduction and internal fixation and minimally invasive needle aspiration are commonly used in patients with Sanders types Ⅱ and Ⅲ calcaneal fractures.However,there is little comparison between the clinical efficacy of the two methods and high-level clinical evidence is still available. OBJECTIVE:To compare the curative effect of Sanders types Ⅱ and Ⅲ calcaneal fractures treated by three-dimensional digital model-assisted minimally invasive needle penetration and tarsal sinus incision and manual reduction and internal fixation with steel plate. METHODS:From January 2021 to October 2022,80 patients with Sanders types Ⅱ and Ⅲ calcaneal fractures who were treated in the Department of Orthopedics,Cangzhou Hospital of Integrated Traditional Chinese and Western Medicine in Hebei Province were randomly divided into control group(40 cases)and observation group(40 cases).The control group was treated with manual reduction and internal fixation with steel plate through the traditional tarsal sinus incision,while the observation group was treated with a three-dimensional digital model assisted with minimally invasive needle penetration fixation.The operation time,blood loss,hospitalization time and fracture healing time of the two groups were recorded.The changes in Maryland score,AO-FAS score,pain visual analog scale score,quality of life score(SF-36 score),and imaging parameters(B?hler angle,Gissane angle,calcaneal length,width and height)were observed before and 12 months after operation in the two groups.The complications during the follow-up were recorded. RESULTS AND CONCLUSION:(1)Operation time,blood loss,hospitalization time and fracture healing time in the observation group were lower than those in the control group(all P<0.05).(2)The Maryland score,AO-FAS score,SF-36 score,B?hler angle,Gissane angle,calcaneal length and height of the two groups after treatment were significantly higher than those before treatment(all P<0.05).Visual analog scale score and calcaneal width were significantly lower than those before treatment(all P<0.05).(3)After 12 months of follow-up,the incidence of complications in the observation group was lower than that in the control group(all P<0.05).(4)In conclusion,the treatment of Sanders types Ⅱ and Ⅲ calcaneal fractures with three-dimensional digital model-assisted minimally invasive needle penetration fixation can significantly improve the operation time,bleeding volume and other perioperative indicators,and can reduce the occurrence of multiple complications.The recovery of ankle function,relief of pain symptoms,and improvement of quality of life are equivalent to traditional therapy.

Nasopharyngeal carcinoma (NPC) is the third most common malignancy with a high recurrence and metastasis rate in South China. Natural compounds extracted from traditional Chinese herbal medicines have been developed and utilized for the treatment of a variety of cancers with modest properties and slight side effects. Maackiain (MA) is a type of flavonoid that was first isolated from leguminous plants, and it has been reported to relieve various nervous system disorders and exert anti-allergic as well as anti-inflammatory effects. In this study, we demonstrated that MA inhibited proliferation, arrested cell cycle and induced apoptosis in nasopharyngeal carcinoma CNE1 and CNE2 cells in vitro and in vivo. The expression of the related proteins associated with these processes were consistent with the above effects. Moreover, transcriptome sequencing and subsequent Western blot experiments revealed that inhibition of the MAPK/Ras pathway may be responsible to the anti-tumor effect of MA on NPC cells. Therefore, the effects of MA and an activator of this pathway, tertiary butylhydroquinone (TBHQ), alone or combination, were investigated. The results showed TBHQ neutralized the inhibitory effects of MA. These data suggest that MA exerts its anti-tumor effect by inhibiting the MAPK/Ras signaling pathway and it has the potential to become a treatment for patients with NPC.
Humans ; Nasopharyngeal Carcinoma/pathology* ; Cell Line, Tumor ; Cell Proliferation ; Apoptosis ; Signal Transduction ; Nasopharyngeal Neoplasms/pathology*

Molecular knowledge of human gastric corpus epithelium remains incomplete. Here, by integrated analyses using single-cell RNA sequencing (scRNA-seq), spatial transcriptomics, and single-cell assay for transposase accessible chromatin sequencing (scATAC-seq) techniques, we uncovered the spatially resolved expression landscape and gene-regulatory network of human gastric corpus epithelium. Specifically, we identified a stem/progenitor cell population in the isthmus of human gastric corpus, where EGF and WNT signaling pathways were activated. Meanwhile, LGR4, but not LGR5, was responsible for the activation of WNT signaling pathway. Importantly, FABP5 and NME1 were identified and validated as crucial for both normal gastric stem/progenitor cells and gastric cancer cells. Finally, we explored the epigenetic regulation of critical genes for gastric corpus epithelium at chromatin state level, and identified several important cell-type-specific transcription factors. In summary, our work provides novel insights to systematically understand the cellular diversity and homeostasis of human gastric corpus epithelium in vivo.
Humans ; Epigenesis, Genetic ; Gastric Mucosa/metabolism* ; Chromatin/metabolism* ; Stem Cells ; Epithelium/metabolism* ; Fatty Acid-Binding Proteins/metabolism*

Objective To investigate the effect and mechanism of miR-27a-3p on nerve cell apoptosis induced by oxygen glucose deprivation/reoxygenation(OGD/R)through regulation of Rho family GTPase 3(Rnd3)expression.Methods PC12 neurons were cultured in vitro and reoxygenated for 3,6,9 h and 12 h after 2 h oxygen glucose deprivation.Cell viability,miR-27a-3p expression and Rnd3 mRNA expression were assessed at each time point and the optimal reoxygenation time point was screened.After transfection of miR-27a-3p Mimic,miR-27a-3p Inhibitor and their negative control,transfection of shRnd3 and its negative control,or co-transfection of shRnd3 and miR-27a-3p Inhibitor through lentivirus,CCK-8 assay was used to detect cell activity.The apoptosis rate of the cells was detected using flow cytometry.Expression of miR-27a-3p and Rnd3 mRNA was detected by RT-qPCR.Expression of apoptosis-related protein and Rnd3 protein was detected by Western blot.The dual luciferase reporter assay confirmed the targeting relationship between miR-27a-3p and Rnd3.Results Upregulation of miR-27a-3p increased cell viability,decreased total cell apoptosis rate,suppressed pro-apoptotic proteins Cleaved Caspase-3(C-caspase-3)and Bax,and promoted expression of anti-apoptotic protein Bcl-2(P<0.05);The opposite result was found when down-regulating miR-27a-3p.The double luciferase reporter gene assay showed that Rnd3 was the target gene of miR-27a-3p.Down-regulation of Rnd3 increased cell viability,decreased the total rate of apoptosis,suppressed the pro-apoptotic protein C-caspase-3,Bax,and promoted expression of the anti-apoptotic protein Bcl-2(P<0.05).However,miR-27a-3p Inhibitor reversed the protective effect of shRnd3.Conclusion miR-27a-3p alleviates OGD/R-induced damage to PC12 neurons by targeting Rnd3 to inhibit cell apoptosis.

Objective To investigate the efficacy and safety of lenvatinib in patients with unresectable hepatocellular carcinoma (HCC) previously treated with tyrosine kinase inhibitor (TKI). Methods A retrospective analysis was performed for the clinical data of 76 patients with unresectable HCC who were treated with lenvatinib in Beijing Ditan Hospital, Capital Medical University, from January 2019 to January 2020, and according to the treatment method, they were divided into TKI previously untreated group with 49 patients and TKI treatment-experienced group with 27 patients. The patients were observed till one year after enrollment, adjustment of treatment regimen, tumor progression, or death. The two groups were compared in terms of progression-free survival (PFS) time, objective response rate (ORR), disease control rate (DCR), and incidence rate of adverse events. The t -test or the Wilcoxon rank-sum test was used for comparison of continuous data between two groups, and the chi-square test or the Wilcoxon rank-sum test was used for comparison of categorical data between two groups; the Kaplan-Meier method was used for survival analysis, and the log-rank test was used for comparison between groups. Results There were no significant differences between the TKI previously untreated group and the TKI treatment-experienced group in median PFS time (115 days vs 72 days, P =0.148), ORR (36.7% vs 18.5%, P =0.098), DCR (65.3% vs 55.6%, P =0.402), and incidence rates of grade ≥3 adverse events (24.5% vs 18.5%, P =0.550). Conclusion Patients with unresectable HCC previously treated with TKI can benefit from lenvatinib and have good safety, with similar results to those treated with TKI for the first time.

Objective To investigate the efficacy and safety of lenvatinib combined with sintilimab as the second-line therapy for advanced intrahepatic cholangiocarcinoma (ICC). Methods A retrospective analysis was performed for the clinical data of the patients with advanced ICC who were admitted to Beijing Ditan Hospital from October 31, 2019 to October 31, 2021 and could not undergo surgery or experienced metastasis after surgery. All patients were treated with lenvatinib combined with sintilimab as the second-line therapy. The patients were followed up, and the RECIST1.1 criteria were used to assess treatment outcome. The primary endpoint was time to progression (TTP), and the secondary endpoints were tumor objective response rate (ORR), disease control rate (DCR), overall survival (OS) time, and safety. The Kaplan-Meier method was used to plot survival curves, and the log-rank test was used for comparison between groups. Results A total of 27 patients were enrolled, among whom there were15 male patients (55.6%) and 12 female patients (44.4%), with a median age of 58 years (range 33-73 years). The median TTP for these patients was 5.5 (95% confidence interval [ CI ]: 1.7-9.3) months, and 13 patients (48.1%) died of disease progression, with a median OS time of 11.2 (95% CI : 5.0-17.4) months. The overall ORR and DCR were 40.7% and 70.3%, respectively. Of all patients, 66.7% experienced varying degrees of adverse events, and among these patients, 44.4% had an increase in alanine aminotransferase, 44.4% had an increase in aspartate aminotransferase, 37.0% had hypertension, 29.6% had an increase in bilirubin, 29.6% experienced diarrhea, and 25.9% each experienced proteinuria, anorexia, and weakness. No treatment-related death was observed, and only 1 patient developed grade Ⅳ immune-related hepatotoxicity and was relieved without sequelae after corticosteroid therapy, resulting in permanent withdrawal of sintilimab. The patients with lymph node metastasis had a significantly shorter median TTP than those without lymph node metastasis (4.5 months vs 18.8 months, P =0.035), and the patients who achieved disease remission had a significantly longer median TTP [11.6 months (95% CI : 5.6-17.6) vs 2.8 months (95% CI : 1.8-3.8), P < 0.001]; the patients with lymph node metastasis had a shorter median OS time [9.6 months (95% CI: 7.9-11.3) vs 21.9 months (95% CI : 0-44.9), P =0.053], and the patients who achieved disease remission had a significantly longer median OS time [16.6 months (95% CI : 9.0-24.2) vs 6.9 months (95% CI : 3.6-10.2), P =0.011]. Conclusion Lenvatinib combined with sintilimab has a marked clinical effect and a low incidence rate of serious adverse events as the second-line therapy for advanced ICC, and therefore, it is a safe and effective treatment regimen.

OBJECTIVE：To study the inhibi tory effects of genistein on the growth of human nasopharyngeal carcinoma. CNE 1 cells and predict its potential target. METHODS ：CCK-8 method was used to test the effects of 0（blank control ），12.5，25，50， 100，150 µmol/L genistein on the proliferation of CNE 1 cells after treated for 24，48，72 h. Flow cytometry was carried out to detect the effects of 0（blank control ），15，30，60 µmol/L genistein on the cell cycle and ap optosis of CNE 1 cells after treated for 24 h. Scratch test was used to investigate the effects of 0（blank control ）， 10， 20， 30 µmol/L genistein on themigration ability of CNE 1 cells after treated for 24 h. High （No.18210156） throughput sequencing was conducted to discover the differential genes in CNE 1 cells after treated with 0（blankcontrol），30 µmol/L genistein for 24 h. RT-qPCR assay was adopted to verify the mRNA expression of related differential genes in above trials. RESULTS ： Compared with blank control，12.5，25，50，100，150 µmol/L genistein sho wed significant inhibitory effect on the proliferation of CNE 1 cells（P＜ 0.01），in a concentration- time-effect manner ；15，30 µmol/L genistein could arrest CNE 1 cell cycle at G 0/G1 stage（P＜0.05 or P＜ 0.01）；30，60 µmol/L could arrest CNE 1 cell cycle at G 2/M stage and promoted cell apoptosis （P＜0.05 or P＜0.01）. 10，20，30 µmol/L genistein could significantly inhibit the migration ability of CNE 1 cells（padj＜0.01）. High throughput sequencing revealed a total of 2 271 differentialgenes（P＜0.05），1 154 of which were up-regulated while 1 117 of which were down-regulated ；8 potential target genes ，including p53，p21，STC2，FGF2，CDK6，CYCLIN D ，PI3K，AKT，were screened by cell experiment. After validated by RT-qPCR assay ，mRNA expression of p53，p21，STC2，FGF2，CDK6，CYCLIN D and AKT were significantly down-regulated（P＜0.05），which consistent with the sequencing results. CONCLUSIONS ：Genistein can effectively inhibit the growth of human nasopharyngeal carcinoma CNE 1 cells，the mechanism of which may associated with inhibiting the expression of mutant gene p53，restoring the function of wild-type P 53 protein and inhibiting the activity of PI 3K/Akt pathway.

Objective To investigate the effects and mechanisms of valproic acid on brain edema,neurobehavioral outcome and inflammatory response after traumatic brain injury (TBI) in rats.Methods TBI animal models were established using Feeney's method.Fifty-four SD male rats,weighting 220-250 g,were randomly divided into 3 groups (n =18):sham operation group (group sham),traumatic brain injury group (group TBI) and valproic acid treatment group (group TBI + VPA).Experimental rats were treated with valproic acid (300 mg/kg,twice daily) by intraperitoneal injection.Rat behavioral outcomes were measured by modified neurologic severity score (mNSS) tests at day 1,3,and 7 after TBI.Brain water content was measured with wet-dry weight method.The blood cells infiltration into cerebral cortex were tested with immunohistochemistry staining against ED-1 for macrophage.Inflammatory cytokines (INF-γ,tumor necrosis factor-α,interleukin-6) were measured by Western blotting.The statistical analysis were performed by ANOVA and chi-square tests using the statistical software program SPSS 13.0.Results Compared with the Sham group,the levels of brain edema,mNSS and macrophage cell infiltration were significantly increased after TBI (all P =0.00).The expressions of inflammatory cytokines were also increased significantly (all P =0.00).Compared with the TBI group,TBI + VAP group had significantly lower brain water content[3day:(80.12 ±0.59)％ vs.(82.14 ±0.67)％,P=0.04;7day:(74.74 ±0.72)％ vs.(77.93 ±0.48)％,P=0.01],and mNSS scores [3 day:(10.53 ±0.32) vs.(11.74 ±0.48),P =0.02;7 day:(7.97 ± 0.32) vs.(10.73 ± 0.42),P =0.01].VPA suppressed macrophage cell infiltration into cerebral cortex [(36.44 ± 0.72) ％ vs.(25.93 ± 0.48) ％ P =0.00].Meanwhile,VPA inhibited the expressions of inflammatory cytokines (INF-γ,TNF-α,IL-6) (P ＜ 0.05).Conclusions Treatment with VPA markedly reduced brain edema and improved neurological outcomes after TBI,possibly mediated by inhibited TBI-induced cerebral inflammatory responses and macrophage cell infiltrating into cerebral cortex.