Objective:To study the preventive and health effects of Ampelopsis grossedentata on hyperlipidemia. Method:Thirty Wistar rats were divided into three groups:control group, model group and test group. The control group was fed normal diet, while the model and test groups were fed high fat diet.The control and model group were given water while the test group was given water with 20% Ampelopsis grossedentata for 15 w. Serum lipids, hemorrheological indices, myocardial enzymes and IL-6 were measured. Results: At the end of experiment, serum TC, TG, LDL-C, and apoB/apoA, serum and myocardial MDA, myocardial enzymes activities, hemorrheological indices and serum IL-6 were obviously higher in model group while the activities of serum and myocardial SOD and GSH-Px were lower. In test group. serum TC, TG and apoB/apoA levels, serum and myocardial MDA, myocardial enzymes activities, hemorrheological indices, and serum IL-6 were significantly lower than those in model group, but HDL-C, SOD and GSH-Px activities higher. Conclusion:Ampelopsis grossedentata could significantly decrease blood lipids and hemorrheological indices of rats fed with high fat diet, and protect myocardial cells fromoxidation, and as a result to prevent hyperlipidemia and cardiovascular disease.

Aim To investigate the effect of Trillium tschonoskii Maxim ( TTM ) ethanol extract on hypoxia ischemia brain damage ( HIBD ) in neonatal rats and potential mechanisms. Methods Fifty healthy SD rats of 7 day-old were randomly divided into three groups:the sham operation group ( n=10 ) , the model group ( n=20 ) and TTM treatment group ( n=20 ) , which received 3-day intraperitoneal injection of normal saline or ethanol extract of TTM respectively. TTC staining and Nissl staining were performed to detect the cerebral ischemia area and neuronal death. Western blot was used to detect the expression of Bcl-2 and Bax. Re-sults The brain tissue of model group was slightly swollen, and white necrotic zone induced by ischemia occured on the right side of the brain, while the brain morphology of TTM treatment group was good. After TTC staining, ischemia zone was clearly seen on the right side of the brain in model group, while after TTM treatment, the size of ischemic zone was decreased. Compared with the model group , Nissl staining showed the neuronal cells increased in TTM treatment group. Western blot showed the expression of Bcl-2 protein in TTM group increased than that in HIBD model group ( P <0. 01 ) , while the expression of Bax protein de-creased ( P <0. 01 ) . Conclusion TTM therapy is beneficial for HIBD,which may be related to reducing neuronal apoptosis.

AIM:To construct recombinant adenovirus vector carrying human miR-133a and study its expression in human mesenchymal stem cells(hMSCs).METHODS:The PCR product containing miR-133a was amplified from human genomic DNA and inserted into the adenoviral shuttle vector pAdTrack-CMV.Then the recombinant shuttle plasmid linearized by pmeⅠwas cotransformed into competent E.coli.BJ5183 with the adenoviral backbone plasmid pAdEasy-1 to generate the recombinant adenovirus vector rAd-mir-133a.rAd-mir-133a was then packaged and amplified in human embryonic kidney 293(HEK293) cells.The purified rAd-miR-133a was used to infect the hMSCs and the expression of miR-133a was detected by non-quantitative RT-PCR and real-time PCR.RESULTS:The recombinant adenovirus shuttle vector pAdTrack-CMV-miR-133a was constructed and verified by restriction endonuclease analysis and DNA sequence analysis.rAd-miR-133a was successfully packaged and amplified in HEK293 cells.The transcriptions of primary miR-133a and mature miR-133a were over-expressed in the hMSCs infected with rAd-miR-133a.CONCLUSION:The recombinant adenovirus vector carrying human miR-133a is successfully constructed,which lay a foundation for miR-133a function study.

AIM:To investigate the effect of caspase-8 small hairpin RNA ( shRNA) on attenuating apoptosis of human mesenchymal stem cells ( hMSCs ) .METHODS: Two recombinant plasmids for over-expression of caspase-8 shRNA, pAd-Cap8 shRNA1 and pAd-Cap8 shRNA2, were constructed.Caspase-8 mRNA was determined in pAd-Cap8 shRNA-transfected human HEK293 cells by Q-PCR.The screened pAd-Cap8 shRNA was used to construct the recombinant adenovirus plasmid , which was linearized and transfected into HEK 293 cells for packaging and amplification of the recombi-nant adenovirus rAd-Cap8 shRNA.The expression of caspase-8 at mRNA and protein levels was determined by Q-PCR and Western blotting .Annexin V/PI staining and determination of caspase-8 activity were performed to assess apoptosis of hM-SCs under the conditions of serum deprivation and hypoxia .The mRNA expression of vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), insulin-like growth factor 1 (IGF-1), Bcl-2 and Bcl-xL was analyzed by Q-PCR.RESULTS:The pAd-Cap8 shRNA, which efficiently inhibited caspase-8 expression, was screened by Q-PCR.The recombinant adenovirus plasmid for caspase-8 shRNA was constructed and used to package and amplify the recombinant ad-enovirus ( rAd)-Cap8 shRNA successfully .rAd-Cap8 shRNA-mediated caspase-8 shRNA markedly inhibited caspase-8 ex-pression in hMSCs .Over-expression of caspase-8 shRNA by infection of rAd-Cap8 shRNA also efficiently decreased the ap-optotic rate and caspase-8 activity in hMSCs under the conditions of serum deprivation and hypoxia , with up-regulation of the mRNA expression of HGF, IGF-1 and Bcl-2.CONCLUSION:Caspase-8 shRNA attenuates hMSC apoptosis under the conditions of serum deprivation and hypoxia .

AIM: To establish human umbilical vein endothelial cells (HUVECs) to express green fluorescent protein (GFP), and to study the suppression of GFP by siRNA in HUVECs. METHODS: Using lipofectamine 2000 to transform plasmid pN_3-EGFP encoding GFP into HUVECs. The HUVEC containing pN_3-EGFP, named HUVEC-GFP, was screened and selected by antibiotic G418. Using in vitro transcription T7 kit, GFPsiRNA targeting GFP mRNA and control-siRNA used as control were synthesized. The siRNAs were transfected into HUVEC-GFP with oligofectamine. 48 h later, the expression levels of GFP protein and mRNA in HUVEC-GFP were determined. RESULTS: The HUVEC-GFP was screened to express GFP in the presence of G418. The agarose gel electrophoresis analysis showed that the siRNAs prepared were integrated. 48 h after transfection with siRNAs, compared to control group, the level of GFP fluorescence was obviously decreased in the HUVEC-GFP transfected with GFPsiRNA. The results of RT-PCR detection showed that GFP mRNA expression was obviously suppressed by GFPsiRNA at the rate of 40%, and no obvious suppression of GFP mRNA expression was found in the HUVEC-GFP transfected with control siRNA. CONCLUSION: The siRNA targeting GFP mRNA, synthesized in vitro, efficiently suppresses the GFP expression in HUVECs.

AIM:To investigate the effect of microRNA-214 ( miR-214) on cardiomyocyte hypertrophy and the expression of the potential target genes .METHODS:A cell model of hypertrophy was established based on angiotensin-Ⅱ( Ang-Ⅱ)-induced neonatal mouse ventricular cardiomyocytes ( NMVCs) .Dual luciferase reporter assay was performed to verify the interaction between miR-214 and the 3’ UTR of MEF2C.The expression of MEF2C and hypertrophy-related genes at mRNA and protein levels was determined by RT-qPCR and Western blot , respectively .RESULTS:The expression of ANP, ACTA1,β-MHC and miR-214 was markedly increased in Ang-Ⅱ-induced hypertrophic cardiomyocytes .Dual lu-ciferase reporter assay revealed that miR-214 interacted with the 3’ UTR of MEF2C, and miR-214 was verified to inhibit MEF2C expression at the transcriptional level .The protein expression of MEF2C was markedly increased in the hypertro-phic cardiomyocytes .Moreover, miR-214 mimic, in parallel to MEF2C siRNA, inhibited the expression of hypertrophy-re-lated genes in Ang-Ⅱ-induced NMVCs.CONCLUSION:MEF2C is a target gene of miR-214, which mediates the effect of miR-214 on attenuating cardiomyocyte hypertrophy .

OBJECTIVE To establish the HPLC fingerprint of Jianpi huayu decoction， and to determine the contents of 8 components. METHODS Thermo Hypersil Gold C18 column was used with mobile phase consisted of methanol-0.05% phosphoric acid aqueous solution （gradient elution） at the flow rate of 1.0 mL/min. The column temperature was 30 ℃， the injection volume was 5 μL. The detection wavelength of matrine was 211 nm， and the other components’ detection wavelength was 283 nm. The similarity evaluation of HPLC fingerprints for 10 batches of Jianpi huayu decoction was performed by using the Similarity Evaluation System of Chromatographic Fingerprint of Traditional Chinese Medicine （2012 edition）. The contents of chlorogenic acid， vanillic acid， p-coumaric acid， ferulic acid， hesperidin， quercetin， bergapten and matrine in the samples were determined by HPLC. RESULTS HPLC fingerprint of Jianpi huayu decoction was established. A total of 27 common peaks were identified， and 8 components were identified. The similarity between 10 batches of samples and the control map ranged from 0.942-0.999. RSDs of precision， repeatability and stability tests were less than 3% （n＝6）. The average recoveries of chlorogenic acid， vanillic acid， p-coumaric acid， ferulic acid， hesperidin， quercetin， bergapten and matrine were 99.48%， 101.32%， 101.18%， 100.79%， 101.12%， 99.19%， 99.81% and 102.46%， respectively； RSDs were 1.34%， 0.93%， 1.90%， 1.84%， 0.54%， 1.53%， 1.33% and 1.01%， respectively （n＝6）. The contents were 0.021-0.061， 0.025-0.034， 0.116-0.295， 0.006- 0.062， 0.014-0.053， 0.017-0.026， 0.014-0.027 and 14.05-24.11 mg/g， respectively. CONCLUSIONS The established fingerprint and content determination method can provide a reference for the quality control and subsequent preparation development for Jianpi huayu decoction.