Objective To screen and identify the main pro-angiogenic compounds of Shexiang Baoxin pill ( SBP) presenting in the plasma.Methods The pro-angiogenic effects of SBP and its compounds absorbed into blood were measured by the cell prolif-eration and cell migration assays by xCELLigence .And the cell tube formation and rat aortic ring models were established to evaluate their pro-angiogenic effect.Results SBP(10 -4 ~10 -2 μg/ml), ginsenoside Rg3(1 ~10 μmol/L) and ginsenoside Rh2(1 ~10μmol/L)significantly stimulated human umbilical vein endothelial cells (HUVECs) proliferation, migration and tube-like structures formation at different concentrations (P<0.05).In addition, compared to the control group, only the high concentration group of SBP (10 -2 μg/ml), Rg3(10 μmol/L) and Rh2(10 μmol/L)could induce endothelial cell sprouting from the aortic ring(P<0.05) .Conclusion SBP, ginsenosideRg3 and Rh2 exhibited significantly pro-angiogenic effect in vitro.

ObjectiveTo investigate the difference in protein expression between hepatocellular carcinoma (HCC) patients with recurrence and those with good prognosis, the differential expression and regulatory mechanism of miR-152-3p target proteins, and the role of miR-152-3p in the recurrence of HCC. MethodsTMT-labeled proteomic sequencing and RT-PCR were used to measure the expression of proteins and the expression of miR-152-3p in the HCC tissue of six patients with recurrence at 2 years after HCC resection and six patients with good prognosis at 5 years. Six databases were used to analyze the target genes of miR-152-3p, and Gene Ontology, DAVID, and REACTOME databases were used to perform target gene screening, enrichment annotation, and signal transduction pathway enrichment analysis. Gene mutation frequency and survival curve analysis were performed for the target genes of miR-152-3p to verify the role of miR-152-3p target genes in patients with HCC recurrence. The independent samples t-test was used for comparison of continuous data between two groups, and a Kaplan-Meier analysis was performed to investigate the survival rates of liver-related genes. ResultsCompared with the patients with HCC recurrence, the patients with good prognosis after HCC resection had a significantly higher transcriptional expression level of miR-152-3p in HCC tissue (P＜0.05). The results of protein sequencing showed that there were 365 differentially expressed proteins in HCC tissue between the patients with good prognosis and the patients with recurrence, and the analysis of HCC recurrence databases showed that 17 proteins were regulated by miR-152-3p. Further analysis of the signaling pathways showed that the function of the 17 target genes regulated by miR-152-3p was enriched in the translation and regulation of mitochondria and ribosome, and multiple enrichment revealed that six target genes were closely associated with mitochondrial respiratory chain complex, i.e., AKAP1, FOXRED1, MRPL28, MRPL50, SHC1, and STAU1. Gene mutation frequency and survival curve analysis showed that the loss or weakening of the function of mitochondrial respiratory chain-related target proteins seriously affected the prognosis and survival rate of patients. ConclusionThere is a significant difference in the expression of miR-152-3p in HCC tissue between patients with good prognosis and those with recurrence after HCC resection, and miR-152-3p may lead to the recurrence of HCC by regulating the target genes AKAP1, FOXRED1, MRPL28, MRPL50, SHC1, and STAU1, acting on the mitochondrial respiratory chain, and affecting the oxidative respiratory function of cells.

Objective To build hypoxia/reoxygenation injury model in cultured neonatal rat cardiomyocyte and screen active components from Shexiang Baoxin Pill ( SBP) absorbed in blood against hypoxia/reoxygenation injury .Methods Cardiomyocytes were isolated and purified from hearts of neonatal Sprague Dawley rats (1~3 days old) and were used to build hypoxia/reoxygenation injury model.The components of SBP absorbed in blood were screened by methyl thiazolil tetracolium (MTT) colorimetic method.Results SBP showed significant protective effect against cardiomyocytes hypoxia /reoxygenation injury atthe concentration of 50 μg/ml.Ginsen-oside Rb1, Rb2, bufalin and muscone of twenty components from SBP absorbed in blood also possessed significant protective effect a -gainst cardiomyocytes hypoxia/reoxygenation injury .Conclusion SBP have the protective activity against cardiomyocytes hypoxia /reoxygenation injury , and ginsenoside Rb1, Rb2, bufalin, muscone are the main active components of SBP .This experiment offered basis for further pharmacodynamics and mechanism study of SBP .

Objective : To investigate the inhibitory effect and molecular mechanism of ribosomal translation factor inhibitor Avilamycin on hepatitis B virus replication. Methods: Liver cancer Hep3B cells were treated with different concentrations of Avilamycin. Cell activity was detected by CCK⁃8 ; the apoptosis was detected by flow cytometry , and HBV⁃DNA、pgRNA、MTIF2、RPL10 gene expression level was detected by qPCR method. The HBsAg and HBeAg was detected by ELISA. The AFP was detected by chemiluminescence. Aspartate aminotransferase (AST) , alanine aminotransferase(ALT) , and alkaline phosphatase (ALP) proteins was detected by Biochemistry method. Results : Avilamycin had no inhibitory effect on Hep3B cell proliferation and apoptosis. However, it could promote cellular AST secretion , reduce AFP levels , and have less effect on ALP secretion. In Hep3B cells , Avilamycin promotes accumulation of pgRNA expression by intervening with MTIF2 and feedback upregulates mRNA expression of host RPL10 and MTIF2 genes. It can effectively reduce the HBsAg , HBeAg , and HBV - DNA levels. Conclusion Avilamycin can inhibit MTIF2 translation initiation , regulate the translation process of viral assembly protein by affecting translation initiation , and then inhibit hepatitis B virus replication.

ObjectiveTo investigate the role and mechanism of DNA repair regulation in the process of hepatocellular carcinoma （HCC） recurrence. MethodsHCC tissue samples were collected from the patients with recurrence within two years or the patients with a good prognosis after 5 years， and the Tandem Mass Tag-labeled quantification proteomic study was used to analyze the differentially expressed proteins enriched in the four pathways of DNA replication， mismatch repair， base excision repair， and nucleotide excision repair， and the regulatory pathways and targets that play a key role in the process of HCC recurrence were analyzed to predict the possible regulatory mechanisms. The independent samples t-test was used for comparison of continuous data between two groups； a one-way analysis of variance was used for comparison between multiple groups， and the least significant difference t-test was used for further comparison between two groups. ResultsFor the eukaryotic replication complex pathway， there were significant reductions in the protein expression levels of MCM2 （P=0.018）， MCM3 （P=0.047）， MCM4 （P=0.014）， MCM5 （P=0.008）， MCM6 （P=0.006）， MCM7 （P=0.007）， PCNA （P=0.019）， RFC4 （P=0.002）， RFC5 （P<0.001）， and LIG1 （P=0.042）； for the nucleotide excision repair pathway， there were significant reductions in the protein expression levels of PCNA （P=0.019）， RFC4 （P=0.002）， RFC5 （P<0.001）， and LIG1 （P=0.042）； for the base excision repair pathway， there were significant reductions in the protein expression levels of PCNA （P=0.019） and LIG1 （P=0.042） in the HCC recurrence group； for the mismatch repair pathway， there were significant reductions in the protein expression levels of MSH2 （P=0.026）， MSH6 （P=0.006）， RFC4 （P=0.002）， RFC5 （P<0.001）， PCNA （P=0.019）， and LIG1 （P=0.042） in recurrent HCC tissue. The differentially expressed proteins were involved in the important components of MCM complex， DNA polymerase complex， ligase LIG1， long patch base shear repair complex （long patch BER）， and DNA mismatch repair protein complex. The clinical sample validation analysis of important differentially expressed proteins regulated by DNA repair showed that except for MCM6 with a trend of reduction， the recurrence group also had significant reductions in the relative protein expression levels of MCM5 （P=0.008）， MCM7 （P=0.007）， RCF4 （P=0.002）， RCF5 （P<0.001）， and MSH6 （P=0.006）. ConclusionThere are significant reductions or deletions of multiple complex protein components in the process of DNA repair during HCC recurrence.

ObjectiveTo investigate the characteristics of mitochondrial translational initiation factor 2 （MTIF2） gene methylation and its association with the development and progression of hepatocellular carcinoma （HCC）. MethodsMethSurv and EWAS Data Hub were used to perform the standardized analysis and the cluster analysis of MTIF2 methylation samples， including survival curve analysis， methylation signature analysis， the association of tumor signaling pathways， and a comparative analysis based on pan-cancer database. The independent-samples t test was used for comparison between two groups； a one-way analysis of variance was used for comparison between multiple groups， and the least significant difference t-test was used for further comparison between two groups. The Cox proportional hazards model was used to perform the univariate and multivariate survival analyses of methylation level at the CpG site. The Kaplan-Meier method was used to investigate the survival differences between the patients with low methylation level and those with high methylation level， and the Log-likelihood ratio method was used for survival difference analysis. ResultsGlobal clustering of MTIF2 methylation showed that there was no significant difference in MTIF2 gene methylation level between different races， ethnicities， BMI levels， and ages. The Kaplan-Meier survival curve analysis showed that the patients with N-Shore hypermethylation of the MTIF2 gene had a significantly better prognosis than those with hypomethylation （hazard ratio ［HR］=0.492， P<0.001）， while there was no significant difference in survival rate between the patients with different CpG island and S-Shore methylation levels （P>0.05）. The methylation profile of the MTIF2 gene based on different ages， sexes， BMI levels， races， ethnicities， and clinical stages showed that the N-Shore and CpG island methylation levels of the MTIF2 gene decreased with the increase in age， and the Caucasian population had significantly lower N-Shore methylation levels of the MTIF2 gene than the Asian population （P<0.05）； the patients with clinical stage Ⅳ had significantly lower N-Shore and CpG island methylation levels of the MTIF2 gene than those with stage Ⅰ/Ⅱ （P<0.05）. Clinical validation showed that the patients with stage Ⅲ/Ⅳ HCC had a significantly lower methylation level of the MTIF2 gene than those with stage Ⅰ/Ⅱ HCC and the normal population （P<0.05）. ConclusionN-Shore hypomethylation of the MTIF2 gene is a risk factor for the development and progression of HCC.