A rapid method has been developed based on the sample preparation procedure named as QuEChERS （Quick, Easy, Cheap, Effective, Rugged and Safe）, combined with reversed-phase high performance liquid chromatography with fluorescence detector and C18 column after precolumn derivatization using o-phthalaldehyde and 2-mercaptoethanol to determine dopamine in porcine muscle. Methanol and deionized water （0.1% acetic acid, v/v） with a ratio of 60：40 was used as mobile phase. The flow rate was 0.8 mL/min and dopamine was eluted within 15 min. The linearity range was 0.003-8 μg/mL with r=0.9992. The detection limit for dopamine was 4 μg/kg and the quantification limit was 9 μg/kg. Recovery studies were carried out at 0.1, 0.5 and 1.0 mg/kg fortification levels and the average recoveries obtained ranged from 90.4% to 98.2% with relative standard deviations between 3.5% and 8.1%. The method was found to be suitable for detection of dopamine in animal product tissues at the maximum residue level.

Objective To assess the value of the myocardial segment number of color change in the bull's eyes of three-dimensional speckle tracking echocardiography (3D-STE)in the detection of multi-vessel coronary stenosis at the resting state.Methods A total of 125 consecutive patients were enrolled and divided into the following three groups according to the coronary angiography(CAG):multi-vessel coronary stenosis group (n=48),single-vessel stenosis group(n =34)and control group (n =43).All patients underwent two-dimensional (2D)and three-dimensional echocardiography (3D)and three-dimensional speckle tracking echocardiography (STE).The bull's eyes of left ventricular 17 myocardial segments including longitudinal strain(LS),circumferential strain (CS),area strain(AS)and radial strain(RS)were acquired by 3D-STE.The homogeneous colors were regarded as normal wall motion myocardial segment (LS,CS,AS is a uniform red,RS for uniform blue ),the colors were uneven or shallower,or changes were regarded as abnormal wall motion myocardial segment.The myocardial segment number of color change in the bull's eyes can be calculated.Receiver operating characteristic (ROC ) curves were computed to determine optimal strain cutoff values to predict multi-vessel coronary stenosis.Results The myocardial segments of abnormal wall motion in the bull's eyes (LS,CS,AS,and RS)were significantly increased compared with the control group(P <0.001).In particular,the multi-vessel coronary stenosis group were dramatically increased than the single-vessel coronary stenosis group.Meanwhile,these parameters were higher in patients with multi-vessel coronary stenosis than in those with single-vessel coronary stenosis (P <0.05).ROC analysis demonstrated areas under the curve of 0.84 for 3D GLS,0.89 for 3D GCS,0.95 for 3D GAS,and 0.89 for 3D GRS.An optimal cutoff value of magnitude,sensitivity and specificity were LS,12,89.6%,76.7%;CS,11,89.6%,74.4%;AS,12,91.7%,88.4%;RS,13,81.2%,86.0%,respectively. Conclusions The myocardial segment number of color change in the bull's eyes by 3D-STE is useful to detect multi-vessel coronary stenosis,where in GAS are more valuable indicators with higher sensitivity and specificity.

A rapid method has been developed based on the sample preparation procedure named as QuEChERS (Quick,Easy,Cheap,Effective,Rugged and Safe),combined with reversed-phase high performance liquid chromatography with fluorescence detector and C18 column after precolumn derivatization using o-phthalaldehyde and 2-mercaptoethanol to determine dopamine in porcine muscle.Methanol and deionized water (0.1％ acetic acid,v/v) with a ratio of 60∶40 was used as mobile phase.The flow rate was 0.8 mL/min and dopamine was eluted within 15 min.The linearity range was 0.003-8 μg/mL with r=0.9992.The detection limit for dopamine was 4 μg/kg and the quantification limit was 9 μg/kg.Recovery studies were carried out at 0.1,0.5 and 1.0 mg/kg fortification levels and the average recoveries obtained ranged from 90.4％ to 98.2％ with relative standard deviations between 3.5％ and 8.1％.The method was found to be suitable for detection of dopamine in animal product tissues at the maximum residue level.

Objective To measure the parameters of mitral valve leaflets and mitral annulus in patientswithmildmitralregurgitation( MR )byreal timethree-dimensionaltransesophagealechocardiography (RT-3D-TEE),and explored the mechanism of MR.MethodsFifty-seven MR subjects were selected and twenty-eight subjects without mitral regurgitation were served as control group,all subjects were examined by RT-3D-TEE and acquired image,mitral valve quantification (MVQ) software was used for post-processing.Mitral annulus parameters (H/DAIPm,E2D,θAv-Mv,mitral annulus θnpa) and mitral valve leaflets parameters(A3DE,L2DAIPm,VA1-3tentVp1-3tentVtentHtentθnpa ) at the end of systolic were measured.The results of two groups were compared,and the most affected parameters to mild mitralregurgitation were selected.Results Compared with control group,VA3tent was decreased,mitral annulus θnpa and L2DAIPm increased,and the mitral valve leaflets θnpa was independently correlation factor of mild mitral regurgitation.ConclusionsThe mitral annulus geometry to flat in subjects with mild MR,the mitral valve local area is increased in subjects with mild MR,the mitral valve leaflets θnpa is independently correlation factor of mild mitral regurgitation.

Objective To assess the left ventricular(LV) regional myocardial systolic function after bone marrow mesenchymal stem cells (BMSCs) injection in rabbits with acute myocardial infarction(AMI) by 2-dimensional ultrasound speckle-tracking imaging.Methods Twenty-four healthy rabbits were randomly divided into three groups:group of sham-operated,group of masculine control (AMI was induced by ligation of left anterior descending coronary artery),and group of cell infusion(after two weeks of AMI,bone marrow mesenchymal stem cells were injected to the region of AMI).Four weeks after cell deliver two-dimensional strain images were acquired from LV short-axis view(at the levels of mitral annulus,muscle papillary and apex),radial strain rate (SrR),circumference strain rate (SrC),rotation rate (RotR) and rotation (Rot) of three levels in short-axis views were measured by speckle-tracking imaging.The conventional echocardiography indices included LV diameter of end diastole(LVEDd),LV ejection fraction(LVEF),LV fractional shortening(LVFS).The catheter indices included LV systolic pressure(LVSP),LV end diastolic pressure (LVEDP)and maximum rate of rise and descend of LV pressure(LVdp/dtmax).ResultsCompared with sham-operated group,rabbits had significantly larger LVEDd,lower LVEF and LVFS,lower LVSP and LVdp/dtmax,higher LVEDP in control group,SrR,SrC,RotR and Rot of LV anterior regional myocardial function and global myocardial function of three levels in short-axis views were lower.Compared with control group,the rabbits had significantly smaller LVEDd,larger LVEF and LVFS,larger LVSP and LVdp/dtmax,lower LVEDP,SrR,SrC,RotR and Rot of LV anterior regional myocardial function and global myocardial function of three levels in short-axis views were larger in group of cell infusion.There was no significant between group of cell infusion and group of sham-operated.Conclusions Speckle tracking imaging can evaluate the regional myocardial systolic function of the area of BMSCs transplantation accurately.

Objective To explore the effect and mechanism of Cyclosporin A (CsA) and cholic acid on reducing the human liver cell damage induced by α-amanitin (AMA).Methods According to the previous research results,the minimum hepatocellular survival concentration against αt-AMA (1.4 g/L),the experiment was conducted in 5 groups:control group,damage group,glycochenodeoxycholic acid group,CsA group,and CsA combined with cholic acid group (CsA+ taurocholic acid,CsA+ chenodeoxycholic acid,CsA+ glycocholic acid,CsA+ glycochenodeoxycholic acid,and CsA+ taurochenodeoxycholic acid).For each group,there were 3 time points for observation (24 h,48 h and 72 h after attacking),CsA and CsA+ glycochenodeoxycholic acid was used to protect hepatocytes,respectively,and morphological changes in cells were observed with inverted phase contrast microscope,and the live cells were counted by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) method,and aspertate aminot ransferase (AST) and alanine aminotransferase (ALT) in the culture supernatant were detected by biochemical method.Results Compared with the control group,hepatocellular growth in the injury group was obviously suppressed,with progressive cellular apoptosis and significantly decreased absorbance value of MTIT (0.345 ± 0.021);the activity of AST and ALT increased gradually,reaching the highest after 72 h [(98.4 ± 6.7) U/L and (116.2 ± 9.5) U/L,respectively].Compared with injury group,broken organelles decreased significantly and absorbance value elevated in glycochenodeoxycholic acid group,CsA group and CsA combined with cholic acid group,and at each time point,the highest absorbance value in the CsA+ taurochenodeoxycholic acid group [the highest was (0.656 ± 0.014),P ＜ 0.05];at the same time,the activity of AST and ALT didn't increase obviously,at each time point,the lowest in CsA+ taurochenodeoxycholic acid group [the lowest was (22.3 ± 6.2) U/L and (20.2 ± 5.4) U/L,P ＜ 0.05,respectively].Conclusions CsA,as well as cholic acid,can protect human liver cells from the attack of α-AMA.The mechanism may be CsA,as an organic anion transfer peptide in humans (OATP1B1 and OATP1B3) suppressant,inhibits the absorption of α-AMA.The joint application of CsA and taurochenodeoxycholic acid is superior to the single OATP substrate or inhibitor.

Objective To investigate the effect and significance of regulating endoplasmic reticulum stress on the expression of histone methyltransferases SET7/9 in the kidneys of db/db mice.Methods Db/db mice were randomly divided into two groups according to random number table method:diabetic nephropathy model group (DN group,n=18) and betaine treatment group (DN+B group,n =18),db/m mice were defined as normal control group (NC group,n =18).At the end of 4,8 and 12 weeks,the expression of GRP78,SET7/9,H3K4me2,and monocyte chemoattractant protein 1 (MCP-1) was determined by real-time fluorescence PCR and Western blotting.24-hour urinary protein excretion rate (UPER) and urine MCP-1 were measured by enzyme linked immunosorbent assay (ELISA).The dynamic changes of blood glucose(BG),serum creatinine (Scr),blood urea nitrogen (BUN) were tested by completely automatic biochemistry analyzer.The morphology of kidney was estimated by special staining of periodic acid-schiff (PAS).Results The levels of BG,BUN,UAER and MCP-1 were significantly higher in DN group than those in NC group (P ＜ 0.05),and were in time-dependent manner.Glomerular basement membrane thickening and mesangial cells proliferation began to emerge in DN group at the end of week 4 and mesangial matrix expansion was more obvious at the end of week 12.The mRNA and protein expression of GRP78 and SET7/9 were elevated significantly in DN group as compared to NC group.The H3K4me2 protein expression level was also increased in time-dependent manner.Compared with the DN group,in DN+B group glomerular lesions attenuated and the GRP78 and SET7/9 expression levels obviously decreased (P ＜ 0.05).Furthermore,the levels of BG,BUN,UPER,MCP-1,H3K4me2 in DN+B group were also reduced (P ＜ 0.05).Conclusion Endoplasmic reticulum stress may be the upstream mechanism of mediating the expression of SET7/9 in the kidneys of DN mice.

Objective To investigate the application of parametric quantification (PQ) with targeted microbubbles on angiogenesis after stem cell transplantation in rabbits with myocardial infarction.Methods A CD34-targeted microbubble was prepared.Acute myocardial infarction (AMI) model were established by ligating coronary artery in 80 New Zealand white rabbits.Rabbits in common microbubble group (T + C group) and CD34-targeted microbubble group (T + T group) were subjected to myocardial contrast echocardiography(MCE),then compare myocardial perfusion parameters (A,β and A × β values) pre- and 3d post- ligation and 4 weeks after stem cell transplantation.Microvessel density (MVD) were determine by CD34 antibody immunohistochemistry.Results Compared to before stem cell transplantation,the perfusion scores ( P ＜0.05) and A,β and A× β values( P ＜0.01) of T+ C group and T+ T group were increased than before transplantation and control group.MVDs after stem cell transplantation in experimental groups were higher than those of control group ( P ＜0.01),especially the T + T group.A × β value in the T + C group was correlated with MVD ( r =0.658,P ＜ 0.05) ; β and A × β value in the T + T group were correlated with MVD ( r =0.620,0.859,P ＜0.05).A × β value ( X ) and MVD ( Y ) were included to establish the following regression equation:Y =- 130.986 + 34.505 X ( R 2 =0.556,P ＜ 0.05).Conclusions PQ and CD34-targeted microbubble probably provides some value in invasive evaluation of angiogenesis after stem cell transplantation.

Objective To investigate the effects of MG132 on diabetic nephropathy (DN) rats induced with streptozocin. Methods Seventy-two male SD rats were randomly divided into three groups: normal control group (NC, n=24), DN group (n=24) and DN treated with MG132 group (DN+MG132, n=24). At the end of 4, 8 and 12 weeks, 24 hour urinary protein excretion rate (UPER) was detected. Morphology of kidney was examined by special staining of periodic acid-schiff (PAS). Renal 26S proteasome activity was determined by quantifying the hydrolysis of S-LLVY-AMC in a fluorescence reader. Urinary malondialdehyde (MDA) level and renal SOD and GSH-PX activity were detected by commercial kits. Renal SOD, GSH-PX and p47phox mRNA expressions were determined by real-time fluorescence PCR. Renal p47phox protein expression wasdetermined by Western blotting. Results Compared with NC group, the DN group showed a significant increased of UPER at week 4, 8, 12 (all P<0.05), of mesangium proliferation and mesangial matrix expansion at week 12. In DN+MG132 group, UPER was significantly decreased compared with DN group at the end of 4, 8 and 12 weeks (P<0.05, respectively), and the glomeruler pathological alteration induced by diabetes was attenuated. Increased renal 26S proteasome activity in DN rats was significantly inhibited after MC132 administration (P<0.05). Moreover, renal p47phox mRNA expression in DN group was 155%, 149% and 120% more than those in NC group at 3 time points (all P<0.05), and so was the renal p47phox protein expression, 139%, 152% and 186% more (all P<0.05). Urinary MDA levels in DN group were 1.95-, 2.04-and 2.62-folds more than those in NC group (all P<0.05). In addition, compared with NC group at 3 time points, in DN group, renal SOD activity was decreased by 23.09%, 33.59% and 53.31% (all P<0.05); renal GSH-PX activity was decreased by 28.57%, 33.06% and 48.76% (all P< 0.05); renal SOD mRNA was decreased by 38.09%, 61.44% and 76.53% (all P<0.05); renal GSH-PX mRNA group was decreased by 29.16%, 37.26% and 62.40% (all P<0.05). Compared with DN group, renal p47phox mRNA and protein expression, and urinary MDA levels were significantly lower in DN+MG132 group (all P<0.05); renal SOD and GSH-PX activity as well as mRNA expression were significantly increased in DN+MG132 group (all P<0.05). Conclusions MG132 treatment can provide renoprotection for DN rats effectively maybe through enhancing renal anti-oxidative ability.

Objective To investigate the effect of 4-phenylbutyric acid(4-PBA)on the renal pathogenesis of rats with streptozotocin-induced diabetes and its mechanism. Methods Fifty-four male SD rats were randomly divided into three groups:normal control group(NC group,n=18),diabetic nephropathy group(DN group,n=18),diabetic nephropathy plus 4-PBA treatment group(4-PBA group,n=18).At the end of 4,8 and 12 weeks,index of kidney weight/body weight ratio(KI)were measured and calculated.Serum creatinine (Scr),blood urea nitrogen(BUN),urinary MDA levels,urinary SOD activity,and 24 hour urinary protein excretion ram(UAER)were detected by HITACHI automatically.Morphology of kidney wag examined by special staining of periodic acid-schitt (PAS).The p47phox and nitrotyrosine (NT) expression in kidney were determined by real-time fluorescence PCR and Western blotting. Results Compared with the NC group, the DN group rats showed a significant increase of KI(P＜0.05), UAER(mg/24 h) (4.92±0.70 vs 0.26±0.07, 5.29±0.83 vs 0.28±0.08, 5.54±0.81 vs 0.29±0.04,respectively, P＜0.05]for indicated time, mesangial cells proliferation and mesangial matrix expansion at 12 week. However,4-PBA treatment could significantly inhibit the increase of KI (P＜0.05), decrease UAER (mg/24 h) (3.71±0.37, 3.47±0.36, 3.28±0.40, respectively, P＜0.05]for indicated time, and prevent the glomeruler pathological alteration induced by diabetes. Moreover, the mRNA expression of p47phox in the kidney of DN group was 154.72%, 148.60% and 91.95% more than that of NC group (all P＜0.05) for indicated time. The protein expression of p47phox was 118.00%, 140.10% and 177.82% more than that of NC group (all P＜0.05), and the protein expression of NT was 45.29%,59.13% and 89.28% more than that of NC group (all P＜0.05). In addition, urinary MDA levels in DN group were 2.05-, 2.26- and 2.43- folds of NC group, and urinary SOD activities were decreased by 64.78%, 71.29% and 79.32% of NC group. Compared with the DN group, the mRNA and protein expression of p47phox, and protein expression of NT in 4-PBA group were decreased markedly (all P＜0.05) at the end of 8 and 12 weeks. The urinary MDA level was decreased, and the urinary SOD activity was increased significantly in rats with diabetes after 4-PBA treatment for indicated time (all P＜0.05). Conclusion 4-PBA treatment can significantly inhibit the renal pathogenesis of rats with diabetes through inhibition of oxidative stress.