Objective To determine the content of emodin in Qinhuang Houzhen Capsule and to provide evidence for its quality criteria.Method A HPLC method was established as follows: Dionex AcclaimTM120 C18 cloumn(4.6? 150 mm,5 ? m),methnol-0.4 % phosphoric acid aqueous solution(85∶ 15) as mobile phase,flow rate at 1.0 mL/min and detection wavelength at 254 nm.Results The average content of emodin in Qinhuang Houzhen Capsule was 0.2057 mg/g,and RSD=4.11 %.Conclusion The content of rhubarb calculated by emodin in 1 g Qinhuang Houzhen Capsule could be affirmed at 0.150 mg/g tentatively.The method is simple,acurate,repeatible,and can be used for the quality control of Qinhuang Houzhen Capsule.

Objective To determine the concents of palmatine hydrochloride and berberine hydrochloride in the preparation of Fu Yan Xiao Lotion(FYXL).Methods The HPLC method was established for the determination of palmatine hydrochloride and berberine hydrochloride in the preparation of FYXL.The chromatographic column is Agilent Extend C18(4.6mm? 150 mm,5 ? m),mobile phase consists of water-acetonitrile(40 ∶ 60,3.5 g potassium dihydrogen phosphate and 1.5 g sodium dodecyl sufate per 1 000 mL),column tempreture was 30 ℃,and the flow rate was 1.0 mL/min.The detection wavelength was 345 nm.Results A good linearity is obtained in the range of 0.062 5 ? g to 1.250 ? g of berberine hydrochloride,and 0.068 6～ 1.372 ? g of palmatine hydrochloride.The average recovery of berberine was 98.63 %,RSD being 2.01 %.The average recovery of palmatine was 102.34 %,RSD being 1.67 %.Conclusion This method is simple and quick,and can be used for the quality control of FYXL.

AIM: To study the effects of Zhongjingweiling Capsule (Fructus Amomi, Cortex Cinnamemi, Rhizoma Alpiniae Galangae, Fructus Foeniculi, Radix Glycyrrhizae, etc.) on analgesia, anti inflammation and gastrointestinal peristalsis inhibition. METHODS: The effect of analgesia was tested by the method of writhes and hot plate on mice. The effect of anti inflammation was tested by injecting acetic acid and implanting cotton pellet and xylene into mice. RESULT: Zhongjingweiling Capsule effectively inhibited the swelling of mouse pinna induced by xylene and the increase of peritoneal capillary permeability caused by the acetic acid injected into mice. The capsule also decreased the proliferation of granuloma caused by implantation cotton pellet significantly and had an abirritant effect on the algesia induced by acetic acid and hot plate. Besides, it effectively inhibited the gastrointestinal peristalsis. CONCLUSION: Zhongjingweiling Capsule is an effective medicine in anti inflammation, analgesia, and gastrointestinal peristalsis inhibition.

Objective To observe the effects of cyclovirobuxinum D(Cvb-D)on the myocardium cell apoptosis of myocardial ischemia rats.Methods The rats were given Cvb-D 0.55,1.1 and 2.2 mg/kg orally per day for 21 days respectively.The myocardial ischemia model was induced by isoprenaline intraperitoneal injection.Myocardium tissue was observed under light microscope and end labelling TUNEL method was used to determine the cell apoptosis.Then the average percentage of apoptosis cell was figured out to evaluate the effect of Cyb-D.Results Cvb-D obviously lightened isoprenaline-induced myocardial pathological changes such as turbulence of myocardial fiber,plasmolysis of necrotic cell,inflammatory cell infiltration of necrotic interstitial region.Cvb-D remarkably inhibited myocardium cell apoptosis of myocardial ischemia rats caused by isoprenaline.Conclusion Cvb-D has the action of ameliorating myocardium cell apoptosis caused by myocardial ischemia.

AIM　To investigate the relationship between sinomenine distribution and its organic toxicology in rats so as to give some pharmacological data for clinical application of sinomenine. METHODS　Three kinds of administration plans were designed in the experiment, ie sinomenine was ip administered at the dosage of l50 mg*kg-1 per day, repreat-dosed for 6 wk and suspended the drug for 1 wk after 6wk repeat-doses．At the end of the each administration plan，the animals were sacrificed and their blood and their main internal organs were collected for the purpose of measurement of sinomenine concentration in each sample by HPLC. Meanwhile，the histopathological and serological examinations were also done in the experiments. RESULTS　The sinomenine concentration in rats internal organs were in order of liver, heart, lung and brain either in single-dosed treated animals or in repeat-dosed treated animals for 6 wk. However，the concentration of sinomenine could not be detected by HPLC after l wk drug-suspension，the histopathological examination showed that sinomenine at the dosage of l50 mg*kg-l per day for 6wk treatment could slightly damage liver ce11s, dominant1y caused the cell edema，but no any influence on the sero1ogy of liver and kidney. Sinomenine ip could also cause a mild hyperaemia of the rats heart tissues but no any histopathological changes had been observed. In testis tissues no sinomenine had beed detected although the animals were treated by repeat treatment for 6 wk and no any histopathological changes had been found yet. However, Sinomenine could partialy inhibit the sperm vitalities and amount of the dead sperms were a1so augmented. It was similar to in vitro eperiments. These influences of sinomenine on testis could be quickly recovered by drug suspension. CONCLUSION　Sinomenine concentration were in order of liver, heart, kidney, lung and brain either in treatment by single dose or by repeat-dose administration. The histopathological changes were only abserved in liver cells of the animals which indicates that it should be in consideration of the liver functions during treatment course of the drug.

AIM To investigate the relationship between sinomenine distribution and its organic toxicology in rats so as to give some pharmacological data for clinical application of sinomenine. METHODS Three kinds of administration plans were designed in the experiment, ie sinomenine was ip administered at the dosage of l50 mg?kg -1 per day, repreat dosed for 6 wk and suspended the drug for 1 wk after 6wk repeat doses.At the end of the each administration plan,the animals were sacrificed and their blood and their main internal organs were collected for the purpose of measurement of sinomenine concentration in each sample by HPLC. Meanwhile,the histopathological and serological examinations were also done in the experiments. RESULTS The sinomenine concentration in rats internal organs were in order of liver, heart, lung and brain either in single dosed treated animals or in repeat dosed treated animals for 6 wk. However,the concentration of sinomenine could not be detected by HPLC after l wk drug suspension,the histopathological examination showed that sinomenine at the dosage of l50 mg?kg -l per day for 6wk treatment could slightly damage liver ce11s, dominant1y caused the cell edema,but no any influence on the sero1ogy of liver and kidney. Sinomenine ip could also cause a mild hyperaemia of the rats heart tissues but no any histopathological changes had been observed. In testis tissues no sinomenine had beed detected although the animals were treated by repeat treatment for 6 wk and no any histopathological changes had been found yet. However, Sinomenine could partialy inhibit the sperm vitalities and amount of the dead sperms were a1so augmented. It was similar to in vitro eperiments. These influences of sinomenine on testis could be quickly recovered by drug suspension. CONCLUSION Sinomenine concentration were in order of liver, heart, kidney, lung and brain either in treatment by single dose or by repeat dose administration. The histopathological changes were only abserved in liver cells of the animals which indicates that it should be in consideration of the liver functions during treatment course of the drug.

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 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 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 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 .