Ursodeoxycholic acid induces apoptosis in hepatocellular carcinoma xenografts in mice

AIM: To evaluate the efficacy of ursodeoxycholic acid(UDCA) as a chemotherapeutic agent for the treatment of hepatocellular carcinoma(HCC).METHODS: BALB/c nude mice were randomized into four groups 24 h before subcutaneous injection of hepatocarcinoma BEL7402 cells suspended in phosphate buffered saline(PBS) into the right flank. The control group(n = 10) was fed a standard diet while treatment groups(n = 10 each) were fed a standard daily diet supplemented with different concentrations of UDCA(30,50 and 70 mg/kg per day) for 21 d. Tumor growth was measured once each week,and tumor volume(V) was calculated with the following equation: V =(L × W2) × 0.52,where L is the length and W is the width of the xenograft. After 21 d,mice were killed under ether anesthesia,and tumors were excised and weighed. Apoptosis was evaluated through detection of DNA fragmentation with gel electrophoresis and the terminaldeoxynucleotidyl transferase-mediated d UTP-biotin nick end labeling(TUNEL) assay. Western blot analysis was performed to determine the expression of apoptosisrelated proteins BAX,BCL2,APAF1,cleaved caspase-9,and cleaved caspase-3.RESULTS: UDCA suppressed tumor growth relative to controls. The mean tumor volumes were the following: control,1090 ± 89 mm3; 30 mg/kg per day,612 ± 46 mm3; 50 mg/kg per day,563 ± 38 mm3; and 70 mg/kg per day,221 ± 26 mm3. Decreased tumor volumes reached statistical significance relative to control xenografts(30 mg/kg per day,P < 0.05; 50 mg/kg per day,P < 0.05; 70 mg/kg per day,P < 0.01). Increasing concentrations of UDCA led to increased DNA fragmentation observed on gel electrophoresis and in the TUNEL assay(control,1.6% ± 0.3%; 30 mg/kg per day,2.9% ± 0.5%; 50 mg/kg per day,3.15% ± 0.7%,and 70 mg/kg per day,4.86% ± 0.9%). Western blot analysis revealed increased expression of BAX,APAF1,cleaved-caspase-9 and cleaved-caspase-3 proteins,which induce apoptosis,but decreased expression of BCL2 protein,which is an inhibitor of apoptosis,following administration of UDCA. CONCLUSION: UDCA suppresses growth of BEL7402 hepatocellular carcinoma cells in vivo,in part through apoptosis induction,and is thus a candidate for therapeutic treatment of HCC.

Correlations of β-catenin,Ki67 and Her-2/neu with gastric cancer

Objective:To study the clinical pathologic characteristics ofβ-catenin,Ki67 and Her-2/neu in gastric cancer and the correlation ofβ-catenin and Ki67 to the protein expression and gene conditions of Her-2/Neu.Methods:The protein expression ofβ-catenin,Ki67 and Her-2/Neu was detected by immunohistochemistry in 101 cases of gastric cancer and the gene conditions of Her-2/Neu by fluorescence in situ hybridization(FISH).Results:The protein expression ofβ-catenin,Ki67 and Her-2/Neu had close relationship with the clinical pathologic characteristics of gastric cancer.Theβ-catenin and Ki67 had obvious correlation to the differentiation,infiltration and lymphatic metastasis of the gastric cancer(P<0.05).The Ki67 had close relationship with the tumor-node-metastasis staging staging of gastric cancer(P<0.05).Her-2/Neu had close relationship with the differentiation and tumor-node-metastasis staging of gastric cancer(P<0.05)but had no relationship with the infiltration and lymphatic metastasis of the gastric cancer(P<0.05).The protein expression of Ki67 had significantly positive correlation to the protein expression and gene amplification conditions of Her-2/Neu(r=0.567,P<0.05 for protein;r=0.304,P<0.05 for gene).Conclusions:Combined detection ofβ-catenin,Ki67 and Her-2/Neu can be used as a reliable method to help the observation of biological behavior,diagnosis and prognosis of gastric cancer,and Ki67 can be used to serve the preliminary screening of Her-2/Neu gene state.

Newly designed “pieced” stent in a rabbit model of benign esophageal stricture

AIM:To investigate a newly designed stent and its dilatation effect in a rabbit model of benign esophageal stricture.METHODS:Thirty-four New Zealand white rabbits underwent a corrosive injury in the middle esophagus for esophageal stricture formation.Thirty rabbits with a successful formation of esophageal strictures were randomly allocated into two groups.The control group(n = 15) was implanted with a conventional stent,and the study group(n= 15) was implanted with a detachable "pieced" stent.The study stent(30 mm in length,10 mm in diameter) was composed of three covered metallic pieces connected by surgical suture lines.The stent was collapsed by pulling the suture lines out of the mesh.Two weeks after stricture formation,endoscopic placement of a conventional stent or the new stent was performed.Endoscopic extraction was carried out four weeks later.The extraction rate,ease of extraction,migration,complications,and survival were evaluated.RESULTS:Stent migration occurred in 3/15(20%)animals in the control group and 2/15(13%) animals in the study group;the difference between the two groups was not statistically significant.At the end of four weeks,the remaining stents were successfully extracted with the endoscope in 100%(11/11) of the animals in the study group,and 60%(6/10)of the animals in the control group;this difference was statistically significant(P < 0.05).There was no difference in the mean number of follow-up days between the control and study groups(25.33 vs25.85).Minor bleeding was reported in five cases in the study group and four in the control group.There were no severe complications directly associated with stent implantation or extraction in either of the two groups.CONCLUSION:In this experimental protocol of benign esophageal strictures,the novel "pieced" stent demonstrated a superior removal rate with a similar migration rate compared to a conventional stent.

Gene testing for osteonecrosis of the femoral head in systemic lupus erythematosus using targeted next-generation sequencing:A pilot study

BACKGROUND Previous publications indicated that genetic predisposition might play important roles in the onset of osteonecrosis of the femoral head(ONFH)in systemic lupus erythematosus(SLE).Some gene loci such as complement C3d receptor 2(CR2),nitric oxide synthase 3(NOS3),collagen type II alpha 1 chain(COL2A1),protein tyrosine phosphatase non-receptor type 22(PTPN22),and transient receptor potential cation channel subfamily V member 4(TRPV4)were reported to be involved in this process.AIM To investigate whether the risk of ONFH in SLE is associated with single nucleotide variations(SNVs)in these five genes.METHODS SNVs in the CR2,NOS3,COL2A1,PTPN22,and TRPV4 genes were examined by using FastTarget and Illumina Miseq sequencing technologies in 49 cases of SLE with ONFH.Burrows–wheeler aligner was used to align the sequencing reads to hg19,and GATK and Varscan programs were used to perform SNV calling.PolyPhen-2,SIFT,and MutationTaster were used to assess the functional effects of non-synonymous SNVs.RESULTS Six of the 49 patients were confirmed to have low frequency SNVs,including one patient with SNVs in NOS3(exon 6:c.814G>A:p.E272K and exon 7:c.814G>A:p.E272K.),four in COL2A1(rs41263847:exon 29:c.1913C>T:p.T638I,exon 28:c.1706C>T:p.T569I,and rs371445823:exon 8:c.580G>A:p.A194T,exon 7:c.373G>A:p.A125T),and one in CR2(rs45573035:exon 2:c.200C>G:p.T67S).CONCLUSION The onset of ONFH in SLE might be associated with the identified SNVs in NOS3,COL2A1,and CR2.

The Change of Quantitative of HBeAg Can Predict the Efficacy of Peg-IFN-α 2a in HBeAgpositive CHB Patients

Objective To investigate the quantitation of hepatitis B e antigen (HBeAg) at week 24 in predicting the efifcacy of pegylated-interferon alfa-2a (Peg-IFN-α2a) in HBeAg-positive chronic hepatitis B (CHB) patients at week 48 and to find a useful predictor for treatment efficacy and investigate individualized treatment of antiviral therapy. Methods Ninety-six HBeAg-positive CHB patients with detectable HBeAg who were treated with Peg-IFN-α2a were enrolled in this trial. They were categorized into 3 groups according to the changes of HBeAg in week 24:HBeAg decline>2 log10 group (group A), HBeAg decline between 1 1og10-2 log10 (group B), HBeAg decline<1 log10 group (group C), and group C was randomly distributed into C1 and C2. The patients in group A, group B, and group C1 continued the original therapy and the patients in group C2 were given lamivudine plus Peg-IFN-α2a for 24 weeks. At week 48, the treatment efifcacy and hepatitis B virus covalently closed circular DNA (HBV cccDNA) in liver biopsies were analyzed. Results At week 48, mean reduction of serum HBV DNA:group A:5.8 log10 copies/ml, group B:3.8 log10 copies/ml, group C1:2.8 log10 copies/ml, group C2:5.7 log10 copies/ml, the reduction of HBV DNA in group A was greater than groups B and C1 (P<0.01), that in group C1 was greater than group C2 (P<0.01), the difference between groups B and C1 had no statistical signiifcance (P=0.19). Mean reduction of HBeAg:group A:2.7 log10S/CO, group B:1.9 log10S/CO, group C1:0.9 log10S/CO, group C2:1.5 log10S/CO, the difference among groups A, B and C1 and between groups C1 and C2 were statistically signiifcant (P<0.01). At week 48, HBV DNA undetectable rate in group A, group B, group C1 and group C2 were 87.5%, 34.5%, 17.4%and 81.9%, respectively, the rate in group A was greater than groups B and C1 (P<0.01),that in group C1 was greater than group C2 (P<0.01). HBeAg seroconversion rate were 75.0%, 24.1%, 13.0%and 22.7%, respectively, that in group A was greater than groups B and C1 (P<0.01). Group A had lower cccDNA in liver tissue than group B and group C1 (P<0.01). The difference of HBV cccDNA between groups B and C1 and that between groups C1 and C2 had no statistical signiifcance. Conclusions HBeAg decline > 2 log10 at week 24 in Peg-IFN-α 2a-treated hepatitis B patients suggested a better efficacy at week 48; HBeAg decline < 2 log10 at week 24 suggests a worse efficacy at week 48, the combined therapy of Peg-IFN-α and lamivudine could improve the clinical responses. The change of quantitative of HBeAg at week 24 may be used as a predictor of treatment effects at week 48.