Objective:To investigate the biological role of miR-1224-5p in the proliferation,migration and apoptosis of prostate cancer cells,as well as its regulatory mechanism of expression.Methods:Human prostate cancer cells(PC3,DU145,LNCaP,22Rv1)and normal prostate epithelial RWPE-1 cells were selected for this study.The expression of miR-1224-5p in prostate cancer cells was detected by qPCR.miR-1224-5p mimic,inhibitor and corresponding negative control(NC)plasmids were transfected into PC3 and DU145 cells by liposome transfection technology,respectively.The transfection efficiency was verified by qPCR.The effects of miR-1224-5p mimic and inhibitor on cell proliferation,migration and apoptosis were detected by CCK-8 assay,plate clone formation assay,scratch healing assay and flow cytometry.The potential N6-methyladenosine(m6A)modification sites in the pri-miR-1224-5p sequence were predicted using SRAMP website,and the prediction results were verified using methylated RNA immunoprecipitation(MeRIP).The expression of methyltransferase 3(METTL3),a key methyltransferase involved in m6A modification,in prostate cancer cells was detected by qPCR.CCK-8 assay and Transwell assay were used to detect the effect of METTL3 siRNA transfection on the proliferation and migration of PC3 and DU145 cells.The regulatory effect of METTL3-mediated m6A modification on the expression of miR-1224-5p was detected by qPCR and MeRIP.Results:miR-1224-5p was upregulated in prostate cancer cells(all P<0.01).Transfection with miR-1224-5p mimic promoted the proliferation,migration and inhibited the apoptosis of PC3 and DU145 cells(P<0.05 or P<0.01).Conversely,miR-1224-5p inhibitor suppressed the proliferation,migration and induced apoptosis of PC3 and DU145 cells(P<0.05 or P<0.01).pri-miR-1224-5p contained m6A modification sites.METTL3 was highly expressed in prostate cancer cells(all P<0.01).Transfection of METTL3 siRNA inhibited the proliferation and migration of PC3 and DU145 cells(all P<0.01).METTL3-mediated m6A modification regulated the expression of miR-1224-5p.Conclusion:miR-1224-5p is upregulated in prostate cancer cells through METTL3-mediated m6A modification.Down-regulation of miR-1224-5p can inhibit the proliferation,migration and induce apoptosis of prostate cancer cells.

Objective:To explore the role of circ_0007766 molecule cyclized from the erb-b2 receptor tyrosine kinase 2(ERBB2)gene in prostate cancer(PCa)cells and the impact of hypoxia on the regulation of its expression.Methods:The expression of circ_0007766 molecule in PCa cells and tissues was detected by qRT-PCR.The siRNA of circ0007766 was transfected to PCa cells(DU145 cell and PC3 cell)respectively;Colony formation assay,CCK-8 assay and Transwell assay were performed to detect the effect of circ_0007766 knockdown on the colony formation,proliferation,migration,and invasion abilities of PCa cells.Hypoxia cell model of DU145 and PC3 cells were established by hypoxic chamber method,and WB was performed to detect the protein expression of HIF-1α,a key molecule of the hypoxic pathway.qRT-PCR was performed to detect the expression of circ_0007766 molecule in hypoxic cell model,and the protein expression of RNA-binding protein eukaryotic translation initiation factor 4A3(EIF4A3)was detected by WB.RNA immunoprecipitation(RIP)was performed to detect the binding of EIF4A3 to circ_0007766 under hypoxic conditions.qRT-PCR assay was performed to further detect the effect of EIF4A3 knockdown on the expression of circ_0007766 under hypoxic conditions.Results:circ_0007766 was highly expressed in PCa cells(P<0.01)and PCa tissues(P<0.05).The knockdown of circ_0007766(P<0.05 or P<0.01)could significantly inhibit the proliferation,migration and invasion abilities of PCa cells(all P<0.01).The upregulation of HIF-1α protein under hypoxic conditions confirmed the successful establishment of the hypoxia cell model.qRT-PCR analysis revealed that compared with that in the normoxia group,circ_0007766 expression was markedly elevated in the hypoxia group(P<0.01).WB analysis demonstrated increased EIF4A3 protein expression in cells of the hypoxia group.RIP assays indicated that circ_0007766 was highly concentrated in the EIF4A3-enriched group(P<0.01).Additionally,qRT-PCR showed that hypoxia significantly boosted circ_0007766 expression,whereas EIF4A3 knockdown notably diminished the hypoxia-induced expression of circ_0007766(P<0.05 or P<0.01).Conclusion:Circ-0007766 plays the role of cancer-promoting molecule in PCa cells,and its expression is related to the regulation of EIF4A3 molecule under hypoxic conditions.

Objective To study the correlation between the genetic polymorphism of tumor necrosis factor-α(TNF-α)-308 with benazepril treatment response in the patients with hypertensive renal damage in Ningxia area.Methods Two hundred and eighty-four patients initially diagnosed as hypertension were enrolled and the hypertensive renal damage defined by the measurement of urinary albumin excretion rate(UAER).At the same time 160 individuals undergoing healthy physical examination were selected as the normal blood pressure control group.The plasma samples were obtained from all the subjects,and plasma level of TNF-α and TNF-α-308 gene polymorphism were detected.Then the patients with hypertensive renal damage were interfered with benazepril as one of the antihypertensive drugs,and the treatment response of different TNF-α-308 genotypes to benazepril was observed,and the comparative analysis was performed.Results Among the TNF-α-308 genotypes in the patients with simple hypertension,genotype GA was the most common,followed by GG and AA successively,with the G/A allele frequency of 53 ％/47 ％ (P＜0.05).In the patients with hypertensive renal damage,genotype GG was the most common,followed by GA and AA successively,with the G/A allele frequency of 70％/30％,the genotypes and allele frequency had no statistical difference(P＜0.05).Before and after benazepril treatment,the change range of UAER in the patients with genotype AA was maximal,followed by the genotype GA and GG,the difference among 3 groups was statistically significant(P＜0.05).Conclusion The TNF-α-308 gene is correlated with hypertensive renal damage and its response to benazepril treatment.

Objective To study the relationship between the genetic polymorphism of interleukine-6 (IL-6)-174 and the response to benazepril treatment in patients with hypertensive renal damage. Methods Two hundred and eighty-four patients with hypertension were enrolled in this study. The hypertensive renal damage was defined by the measurement of urinary albumin excretion rate (UAER). One hundred and sixty healthy subjects were enrolled simultaneously as control group. Blood samples were obtained from all the subjects, and plasma levels of IL-6 and the genotype of gene IL-6-174 were detected. The patients with hypertensive renal damage were treated with benazepril for 16 weeks. The responses were evaluated by the changes of UAER level to benazepril in different genotypes. Results Genotype CC was the most common of the gene IL-6-174 in patients with hypertension, followed by GG and GC successively, with the G/C allele frequency of 47%and 53%(P<0.05), while in patients with hypertensive renal damage, GG was the most common genotype of the gene IL-6-174, followed by GC and CC successively, with the G/C allele frequency of 68%and 32%(P<0.05). After benazepril treatment, the UAER was decreased most in patients with genotype CC, followed by GC and GG successively ( P<0.05). Conclusion The G allele frequency of the gene IL-6-174 is related with hypertensive renal damage in patients in Ningxia, with GG as the most common genotype. The patients with CC genotype have the best response to benazepril treatment, with most decreased UAER.

Objective To study the protective effect of total flavonoids of lycium barbarum(TFLB) on vascular endothelial cells in streptozotocin(STZ)-induced diabetic mellitus(DM) rats injury model.Methods The injury model of DM rats was induced by STZ,and the rats were divided into normal control group,DM model group and TFLB interventiongroup(60,90 and 120 mg/mL).Rats in the normal control group and the DM model group were given the same amount of intragastric administration of normal saline every day.The rats in the intervention group were treated with TFLB at different concentrations for 4 weeks.After filling the stomach for 4 weeks,the blood samples were taken from the chest cavity,and the changes of serum vWF,sTM and sEPCR were measured by ELISA.The tissue of the thoracic aorta was removed and stained by HE,and the protective effect of TFLB on the vascular endothelium in DM rats was observed.Results Compared with DM model group,the levels of vascular endothelial function related factors vWF,sTM and sEPCR were down-regulated in the TFLB intervention group,and the difference was statistically significant(P＜0.05,P＜0.01).In the TFLB intervention group,with the increase of TFLB dose,the intima thickening degree of the rats was significantly reduced;the formation of mountain-like protrusions was gradually decreased;the outer membrane showed no significant damage;the vascular morphology arrangement tended to be regular.Conclusion TFLB can improve the hyperglycemiacaused damage of vascular endothelial cells,regulate the content of vascular injury related factors,delay the progression of DM complications,and protect the cardiovascular system.

Objective To analyse the secondary structure of glutathions S-transferase of Echinoccocus granulosus(EgGST)and predict the B cell and T cell epitopes with informatics tools,in order to provide basic data and references for the following design of epitope vaccine.Methods The B cell and T cell epitopes of EgGST were predicted by DNAstar,Biosun software and Propred MHC class-Ⅱ Binding Peptide Prediction Server software.Results Many distinct antigenic epitopes of EgGST were identified by comput-er,there existed 8 B cell epitopes and 7 T cell epitopes.Conclusion Analysis of predicted epitopes of EgGST antigenic might be sig-nificant for future research of epitope vaccine.

Objective To analyze the effect of smoking on blood lipid before percutaneous coronary intervention(PCI)in male coronary heart disease(CHD).Methods The clinical data in 124 cases of CHD with PCI were randomly extracted and divided into the smoking group (n = 88 )and the non-smoking group (n = 36 ).The age,past disease history,triglycerides (TG),cholesterol (CHOL),low density lipoprotein(LDL)and high density lipoprotein(HDL)were retrospectively investigated.Results The LDL level of the smoking group was(3.1±1.34)mmol/L,which was significantly higher than(2.5 ±0.95)mmol/L in the non-smoking group with statistical difference between them(P <0.05);the level of TG,HDL and LDL in the CHOL≥4.5 mmol/L group were (2.5±1.49)mmol/L,(0.99±0.29)mmol/L and(3.6 ±1.13)mmol/L respectively,which were hither than(1.8 ±1.23)mmol/L, (0.88±0.20)mmol/L and(2.2±0.69)mmol/L in the CHOL≤4.5 mmol/L group respectively,the differences among them had statistical significance(P <0.05 );the Sperman analysis found that there was a positive correlation between the level of vascular change with CHOL and LDL,at the same time the smoking level was negatively correlated with LDLA.Conclusion Smoking, CHOL and LDL have a close relationship with the male coronary heart disease.Moreover,smoking can affect the lipid metabolism and probably increase the risk of coronary heart disease.

In order to construct the recombinant retrovirus vector of human ngn3 gene and its packaging cell line, we successfully amplified the open reading frame (ORF) of ngn3 gene from human fetal pancreatic tissue by RT-PCR. The PCR products of human ngn3 gene was subcloned into pMD18-T vectors and sequenced. Results showed that its sequence was fully consistent with the ngn3 gene published in GenBank(GenBank Accession No. BC126468). The correct fragment was digested by EcoR I and Hpa I from recombinant pMD18-T vector and inserted into the same restriction enzyme sites of retroviral vector pMSCV-neo. We got recombinant retrovirus vector pMSCV-ngn3, which was identified by double restriction enzyme digestion and then transfected into PT67 cells by lipofectamine 2000. We established the PT67-ngn3 packaging cell line by G418 selection, which was detected by RT-PCR and immunohistochemistry staining. The detection results showed that the Ngn3 expressed at the mRNA and protein level in the packaging cell line. RT-PCR detection and electronic microscope analysis showed that the recombinant retroviral vector pMSCV-ngn3 was packaged into infectious virus particles and released into the supernatant of the cells. These results demonstrated that a PT67-ngn3 packaging cell line was successfully established, and this could facilitate the study of differentiation of the human fetal pancreatic progenitor cells into insulin-producing cells by using the ngn3 gene.
Basic Helix-Loop-Helix Transcription Factors ; biosynthesis ; genetics ; Cell Differentiation ; drug effects ; Cell Line ; Cloning, Molecular ; Fetus ; Genetic Vectors ; genetics ; Humans ; Insulin-Secreting Cells ; cytology ; Molecular Sequence Data ; Nerve Tissue Proteins ; biosynthesis ; genetics ; Open Reading Frames ; genetics ; Pancreas ; cytology ; RNA, Messenger ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; pharmacology ; Retroviridae ; genetics ; metabolism ; Stem Cells ; cytology ; Transfection