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Objective:To develop a stoichiometric relationship between iohexol injection and its refractive index. Methods: Ru-dolph J257 and Mettler Toledo RE40D were adopted to determine the refractive index of iohexol injection produced by different facto-ries. The relationship between the refractive index and the concentration of iohexol determined by potentiometric titration was described as C=2.828R-3.769. Results:The relative deviation of simulated content calculated by the stoichiometric and titration content was less than 2%. Little contribution by excipients could be ignored. Conclusion:The proposed method shows simplicity,rapidity and ac-curacy,which can be applied in the process control and the market supervision of iohexol.
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Objective@#To observe the change levels of nuclear factor-kappa B (NF-κB) p65 protein in cytoplasm and nuclear, phosphorylation of inhibitor of kappa B (p-IκB) protein and cytochrome C (Cyt-c) , cleaved cysteinyl aspartate specific proteinase-3 (Cleaved caspase-3) , B-cell lymphoma/leukemia-2 (Bcl-2) in cytoplasm in the process of N, N-dimethylformamide (DMF) -induced apoptosis in H9c2 cardiomyocytes, and explore the tentative mechanism of apoptosis.@*Methods@#H9c2 cardiomyocytes were exposed to 200 mmol/L DMF. Western blotting was used to detect the protein expression levels of p65 in cytoplasm and nuclear, p-IκB after exposure for 0, 2, 4, 6, 8, 12 h, and the protein expression levels of Cyt-c, Cleaved caspase-3, Bcl-2 in cytoplasm after exposure for 0, 2, 4, 8, 12, 24 h. Immunofluorescencecytochemistry (IFC) was used to observe the location of Cyt-c after 200 mmol/L DMF exposure for different times.@*Results@#The levels of p65 in cytoplasm and nuclear and p-IκB among groups were statistically significant (F were 7.79, 33.11, 90.25, respectively, all P<0.01) . Compared with the control group, the levels of p65 in cytoplasm of 2, 4, 6 h group were significantly decreased (all P<0.01) ; the levels of p65 in nuclear of 2, 4, 6, 8 h were significantly increased (all P<0.01) ; the levels of p-IκB of 2, 4, 6 h group were significantly increased (all P<0.01) . The levels of Cyt-c, Cleaved caspase-3 and Bcl-2 among groups were statistically significant (F were 51.42, 503.68, 73.37, respectively, all P<0.01) . Compared with the control group, the levels of Cyt-c of 8, 12 h group were significantly increased (both P<0.01) ; the levels of Cleaved caspase-3 of 2, 4, 8, 12, 24 h were significantly increased (all P<0.01) ; the levels of Bcl-2 of 2, 4, 8, 12, 24 h group were significantly decreased (all P<0.01) . IFC showed that Cyt-c was released from the mitochondria to the cytoplasm gradually as the extension of the exposure time.@*Conclusion@#NF-κB signaling pathway and mitochondrial pathway are involved in the mechanism of DMF-induced apoptosis in H9c2 cardiomyocytes.
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OBJECTIVE: To explore the effect of N,N-dimethylformamide( DMF)-induced inflammatory injury in H9c2 cardiomyocytes and its mechanism. METHODS: H9c2 cardiomyocytes were cultured in vitro and randomly divided into 4different groups: control group,50 mmol / L-group,100 mmol / L-group,200 mmol / L-group. These 4 groups of cells were treated with different DMF concentrations( 0,50,100,200 mmol / L) for 12 hours. The cells were also divided into 6groups and treated with 200 mmol / L DMF at different time points( 0,2,4,6,8,12 h) : control group,2 h-group,4 hgroup,6 h-group,8 h-group and 12 h-group. The level of lactate dehydrogenase( LDH) was detected by colorimetry. The levels of creatine kinase( CK) and isoenzyme of creatine kinase( CK-MB) were detected by ultraviolet spectrometry. The levels of tumor necrosis factor-α( TNF-α),interleukin( IL)-1β,IL-6,and IL-8 were detected by enzyme linked immunosorbent assay. The level of reactive oxygen species( ROS) was detected by fluorescence probe. The location of nuclear factor-kappa B( NF-κB) p65 protein was detected by immunofluorescence cytochemistry( IFC) staining. RESULTS: The levels of LDH,CK and CK-MB in the 50 mmol / L-group,100 mmol / L-group and 200 mmol / L-group were higher than that of the control group( P < 0. 05) and showed a significant dose-effect( P < 0. 05). The levels of LDH,CK and CK-MB in the 6 h-group,8 h-group and 12 h-group were higher than that of the control group( P < 0. 01) and showed a significant time-effect( P < 0. 01). The levels of TNF-α,IL-1β,IL-6 and IL-8 of the 200 mmol / L-group were higher than the control group( P < 0. 05). Compared with the control group,the levels of TNF-α of the 4 h-group,12 h-group were higher( P < 0. 05),the levels of IL-1β of the 2 h-group,4 h-group,6 h-group,8 h-group and 12 h-group were higher( P < 0. 05),the levels of IL-6 of the 2 h-group and 4 h-group were higher( P < 0. 05),the level of IL-8 of the 2 h-group was higher( P < 0. 05). In addition,the levels of TNF-α,IL-1β and IL-6 reached a peak at 4 h-group and the level of IL-8 reached a peak at 2 h-group. The ROS levels of the 2 h-group,4 h-group and 6 h-group were higher than the control group( P < 0. 01),and the level of ROS reached a peak at 2 h-group. Furthermore,IFC staining showed that the fluorescence intensity of NF-κB p65 protein in nucleus of the 2h-group and 4 h-group increased after treatment with DMF,comparing with the control group. CONCLUSION: DMF leads to inflammatory injury in H9c2 cardiomyocytes. ROS and NF-κB might be involved in the process.
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Objective To investigate the effects of uncoupling protein 2 (UCP2) on the myocardial cells of mice with type 2 diabetes mellitus combined with hyperuricemia (HUA), and clarify the mechanism thereof. Methods The mouse cardiac myocytes (MCM) cultured with 25mmol/L high glucose (HG) medium were divided into two groups: HG plus 300μmol/L sodium palmitate for 18 hours as high glucose and high fat (HG+HF) group, and HG+HF plus 1500μmol/L uric acid (UA) for 18 hours as HG+HF+HUA group. Then the myocardial cells in HG+HF+HUA group, by use or not use UCP2 inhibitor genipin, were further divided into two groups: vehicle group and genipin group. In order to verify the mechanism of UCP2 in myocardial cells injury caused by high glucose, high lipid and high uric acid, the myocardial cells were divided again into genipin group and genipin+N-acetylcysteine (NAC) group. Accordingly, the apoptosis of myocardial cells were measured by flow cytometry at specific time, the mRNA and protein expressions of UCP2 were determined by q-PCR and Western blotting, and the levels of reactive oxygen species (ROS) were detected by DHE staining and ELISA. Results The apoptosis rate of myocardial cells increased obviously, and the expression levels of UCP2 decreased and of ROS elevated significantly in HG+HF+HUA group than in HG+HF group (P<0.05). As the expression levels of UCP2 decreased by genipin intervention, the apoptosis rate of myocardial cells and ROS level in HG+HF+HUA group increased more obviously (P<0.05). In contrast, such an effect was reversed by the application of antioxidants NAC (P<0.05). Conclusion UCP2 can inhibit oxidative stress and alleviate the apoptosis of myocardial cells induced by high glucose, high fat and high uric acid.
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Objective:To determine the content of ferric oxide in calamine powder. Methods:Flame atomic absorption spectrom-etry ( FAAS) was used to determine the content of ferric oxide. The determination results were compared with those of the titration method. Results:There was a good linear relationship between the absorbance and the concentration of iron within the range of 0. 5-4μg·ml-1 , the relative standard deviation of the repeatability test was 1. 2%, the average recoveries were 100. 4% and 99. 4%, the limit of quantitation and detection was 0. 081 μg·ml-1 and 0. 024 μg·ml-1 , respectively. Conclusion:The method of FAAS is ac-curate and quick with good specificity and high sensitivity, which can be used for the determination of ferric oxide in calamine powder.
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Objective: To investigate the influence of intermittent cold stress on collagen content of atherosclerotic plaque in experimental ApoE-/-mice. Methods: A total of 20 male ApoE-/-mice at 8 weeks of age were divided into 2 groups:Experimental group, the mice had intermittent cold exposure at (4 ± 1)°C from 8am to 12noon and Control group, the mice were living at (24 ± 2) °C. All animals were treated for 12 weeks, n=10 in each group. The collagen content of atherosclerotic plaque at the aortic root in ApoE-/-mice was observed by Masson staining, the protein expressions of aortic MMP-2, MMP-9 and TIMP-1 were examined by Western blot analysis. Results: Compared with Control group, the Experimental group presented the lower collagen content of atherosclerotic plaque at the aortic root, higher protein expressions of MMP-2, MMP-9 and lower protein expression of TIMI 1. Conclusion: Intermittent cold stress may disturb the balance of MMP/TIMP and decrease collagen content of atherosclerotic plaque to form vulnerable plaque in experimental ApoE-/-mice which may cause acute coronary syndrome.
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To rapidly identify the compound Yiganling tablets from different manufacturers by near-infrared fibre-opti-cal spectroscopy and detect the content of silymarin. Methods:The ninety-three batches of compound Yiganling tablets from six manu-facturers were used as the analysis objects. Clustering analysis model of compound Yiganling tablets was established. The spectra were pretreated with the second derivative and vector normalization;the number of smoothing point was thirteen; the wavelength range was 9 000-4 100 cm-1;the distance between the spectra was Euclidean distance, the distance between the spectra and the class was the sum of square of the variance(Ward’s method). The quantitative model was established using the same samples. The spectra were pretreated with the first Derivative and vector normalization;the number of smoothing point was seventeen;the wavelength ranges were 9 100-7 300 cm-1 and 7 100-4 100 cm-1;the regression method was partial least squares ( PLS) algorithm. Results:The clustering model could identify the samples of training set, and validate the thirty spectra of test set accurately. For the quantitative model of the content of silymarin, the root mean square error of cross validation (RMSECV) was 0. 082 8 and the determination coefficient R2 was 97. 98%. The root mean square error of prediction (RMSEP) was 0. 044 1 and the correlation coefficient R2 was 98. 95%. Conclu-sion:The clustering analysis and the quantitative model are rapid and simple, accurate and reliable, and can be applied in fast drug a-nalysis.
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Objective To establish an animal model of calpastatin ( CAST) transgenic mice by inserting the full hu-man CAST into the genome of C57BL/6J mice.Methods Recombinant transgenic vector pRP .EX3d-EF1A-CAST-IRES-eGFP was constructed by Gateway technology .It was injected into the fertilized eggs from C 57BL/6J mice.The injected eggs were transplanted into the oviduct of pseudopregnant mice .Tail DNA PCR screening was performed to identify the positive founder mice.The expressions of CAST mRNA and protein in tissues of the transgenic mice were detected by RT -PCR and Western blotting.Results Ninty eggs were transplanted into the oviducts of 3 recipients.The transplantation success rate was 100%.23 viable offsprings were born from the recipients .Tail DNA PCR screening showed that two of the offsprings were positive transgenic mice .The positive rate of transgenic mice was 9%.RT-PCR assay revealed that CAST mRNA ex-pressions were present in the heart , liver, kidney, lung, spleen, brain and skeletal muscle of the transgenic mice .Addition-ally, the CAST protein expression was significantly increased in the transgenic mice .Conclusion CAST transgenic mice have been successfully established and provide a good animal model support for further studies on the CAST function .
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As nanomedicines are developing fast in both academic and market areas, building up suitable methods for nanomedicine analysis with proper techniques is an important subject, requiring further research. The techniques, which could be employed for grain size analysis of nanomedicines, were reviewed. Several key techniques were discussed with their principles, scope of applications, advantages and defects. Their applications to nanomedine analysis were discussed according to the properties of different nanomedicines, with the purpose of providing some suggestions for the control and administration of nanomedicines.
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<p><b>OBJECTIVE</b>To study the effect of polysaccharides in processed Sibiraea on the immunologic function of immunosuppression mice.</p><p><b>METHOD</b>The immunosuppressed mice were induced by cyclophosphamide. After the treatment, the organ weight index and the delayed type hypersensitivity of the mice were investigated. The humoral immune function was determined by serum hemolysin assay. Non-specific immune function was determined by carbon clearance method. Cellular immune function was determined by spleen lymphocyte proliferation test. Two hundred kunming mice were randomly divided into five groups: normal controls, model group, low-dose group (110 mg x kg(-1)), middle-dose group (220 mg x kg(-1)), high-dose group (440 mg x kg(-1)). Drugs were given to the mice by oral gavage every day.</p><p><b>RESULT</b>The immunosuppressed mice treated with Sibiraea polysibcharide at intragastrica dose of 110-440 mg x kg(-1) have increased weight of the immune organs, increased content of DTH and content in serum hemolysin lgG and lgM. Mean while the rate of carbon clearance was enhanced and the proliferation of spleen lymphocyte was increased.</p><p><b>CONCLUSION</b>Polysaccharides in processed Sibiraea can increase the weight of the immune organs. At the same time, non-specific immune, DTH, humoral immune and cellular immune function were enhanced significantly.</p>
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Animals , Female , Humans , Male , Mice , Disease Models, Animal , Immunity , Immunocompromised Host , Organ Size , Polysaccharides , Random Allocation , Rosaceae , Chemistry , Spleen , Allergy and ImmunologyABSTRACT
Objective The purpose of the present study was to investigate the effect of peroxisome proliferator-activated receptors δ（PPAR δ）activation with dietary GW610742X on the expression of tenascin-C in the infarcted and remodeling myocardium.Methods Sixty male Wistar rats were divided into four groups,including control group,sham group,myocardial infarction（MI） group,and MI＋GW610742X（GW）group.The left coronary artery was ligated to establish the MI model.PPAR δ activator GW610742X（100mg/kg/d）was administrated into the rats of GW group.At 3 months after procedure,the expression and distribution of tenascin-C in left ventricular free wall from each group were examined by Western blotting and immunofluorescence,respectively.Results After 3 months following procedure,there were obvious necrosis and fibrosis in left ventricular free wall from MI group.The expression of tenascin-C in MI and GW group was significantly higher than those in control and sham group（P 〈 0.01）.Moreover,tenascin-C expression in GW group was remarkably decreased compared to MI group（P 〈 0.05）.Additionally,tenascin-C expression in sham group was similar to that in control group（P 〉 0.05）.Conclusion The tenascin-C is upregulated in infarcted myocardium during the remodeling process,which can be significantly attenuated by PPAR δ activation.
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Objective To investigate the involvement of mitogen-activated protein kinase(MAPK)and PI3K(phosphatidylinositol 3 kinase)/ Protein kinase B(Akt) signal transduction pathways in the mechanism of myocardium remodeling in patients with congestive heart failure(CHF).Methods Thirty nine patients of mitral valve disease with CHF were randomly selected and 30 cases of healthy persons were included as controls.Cardiac function parameters were measured by echocardiography.Concentration of AngⅡ in plasma and myocardial tissues was determined by radio immunoassay.Activity of PKC was determined by using competive prote in binding method,activity of MAPK was detected by the methods of immunoprecitipation.Immunoprecitipation was used to assay the protein expression and phosphorylation of PI3K and Akt(Protein kinase B),protein expression of C-FOS and ?-skeletal-actin in myocardial tissues.Results Pathological changes of myocardial tissues in CHF with valvular heart diseaseshowed typical myocardial remodeling.The hypertrophy was dominant at early stagy of CHF,while at end stage the characteristics include disordered alignament of the myocytes,the discontinuity and dissolving of cardiomyofibrills,destroyed subcellular organs,and the hyperplasia of interstitial tissue.AngⅡ concentration in plasm and myocardial tissues in patients with CHF was higher than those in the control group(P
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Objective To investigate the involvement of mitogen-activated protein kinase(MAPK)system and hypertrophy related genes in myocardial remodeling mechanism of patients with congestive heart failure(CHF).Methods Thirty-nine patients of mitral valve disease with CHF were randomly selected and 30 cases of healthy persons were included as controls.Cardiac function parameters were measured by echocardiography.Concentration of AngⅡ in plasma and myocardial tissues were determined by radio immunoassay.Immunoprecipitation was used to assay the protein expression and phosphorylation of c-jun NH2-terminal kinase(JNK),extracellular signal regulated kinase(ERK),P 38- mitogen-activated protein kinase(P 38-MAPK)in myocardial tissues;protein expression of c-myc,c-myb,c-jun was also determined by immunoprecipitation.Results Compared to the control group,the phosphorylation of ERK1/2 was obvious in heart function class Ⅱ group(P0.05);phosphorylation of JNK was obvious in CHF groups(P0.05);there was no expression in control group.Conclusion MAPK activated by renin angiotensin system may play an important role by causing the protein expression of c-myc,c-myb,c-jun in myocardial remodeling in patients with CHF.
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Aim To investigate the contribution of cardiac L-and L/T-type Ca 2+ channels blockers on matrix metalloproteinases (MMPs) protein expression and fibronectin (FN) degradation following myocardial infarction(MI). Methods Rat MI model was established by permanent ligation of left coronary artery. Infarcted rats were orally treated with placebo, amlodipine (L-channel blocker, 4 mg?kg -1?d -1) or mibefradil (L/T-Channel blocker, 10 mg?kg -1?d -1) for 7 days before induction of myocardial infarction. Protein levels of MMP-2,3,9 and FN distribution were assayed by immunoflorescence at 1, 3, 7 days post coronary occlusion in left ventricular free wall (LVFW) and myocardium interventricular septum (IS). Results Pathological changes of myocardial tissues in IS and LVFW showed typical myocardial remodeling. The hypertrophy was dominant in IS, but in LVFW the characteristics including disordered alignament, hypertrophy of the myocytes, the discontinuity, dissolving of carediomyofibrills, and the hyperplasia of interstitial tissue were found 7 days after MI. The protein levels of MMP-2,3,9 increased in IS 1,3,7 days post MI gradually, whereas the protein levels of FN decreased gradually, the protein levels of MMP-2,3,9 increased significantly in LVFW 7 days post MI, and the protein levels of FN decreased significantly compared to control group, respectively(P
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Aim To invesitgate the effect of PTEN(phosphatase and tensin homologue deleted on chromo-some ten) /PI3K(phosphatidylinositol 3 kinase) signaltransduction pathway in hypertrophied myocardium me-diated by overloaded pressure in rats,and the effect ofangiotensionⅡ(AngⅡ) receptors(AT1,AT2) antag-onists on it.Methods Overloaded pressure hypertro-phied myocardiumratmodel was established by abdom-inal arota constriction.Thirty two male wistar rats weredivided randomly into four groups,namely sham-opera-ted group,banding group,valsartan group(bandinggroup and valsartan administation),and PD123319group(banding group and PD123319 administration).AngⅡconcentration in plasma and left ventricular myo-cardium were measured by radioimmunoassays.Cardiacindex was measured and HE staining was performedwith left ventricular myocardium,immunoprecitipationwas used to assay the protein expression and phospho-rylation of PTEN、PI3K and Akt(Protein kinase B),protein expression of?-skeletal-actin in myocardial tis-sues.Results AngⅡ concentration in serum and leftventricular myocardium tissue in banding group washigher than that in sham-operated group,valsartangroup andPD123319 group(P0.05).Conclusions PTEN/PI3Ksignal pathway is involved in myocardium hyper-trophy in overloaded pressure rat model.The up-regu-lations of PI3K/Akt and down regulation of PTEN aremediated via AT1 and can be reduced by AT1 receptorantagonists.
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Objective To evaluate the role of transfected angiotensinⅡ(Ang Ⅱ) receptor AT1 anti-sense nucleotide (AT1A) in the expression of subtypes of AngⅡ receptor mRNA, synthesis of protein and nucleic acid in cardiomyocytes. Methods AT1 cDNA sequence (476 bp) was cloned with RT-PCR and reversely inserted into PcDNA3.1 (5.4 kb) to construct an intact plasmid containing AT1A (PAT1A). The plasmid was then transfected into the cultured cardiomyocytes and identified with RT-PCR and Western blot. The synthesis of protein and nucleic acid identified by 3H-Leu and 3H-TdR incorporation, and expressions of AT1 and AT2 mRNA by RT-PCR, were compared between transfected and nontransfected cardiomyocytes after being stimulated with 10-7 mol/L AngⅡ for 24 h. Results The plasmid PAT1A were successfully constructed. The AT1 mRNA and its protein were expressed significantly less in the transfected cardiomyocytes than in the control (P<0.01). In the transfected cardiomyocytes, AT1 mRNA expression was markedly decreased, but that of AT2 mRNA obviously increased (P<0.01) when compared with the nontransfected cardiomyocytes after stimulation for 24 h with AngⅡ 10-7 mol/L; no significant difference was found in 3H-Leu and 3H-TdR incorporation between them. Conclusion After the cardiomyocytes was tranfected with AT1A, the expression of AT1 mRNA was markedly suppressed，while AT2 mRNA up-regulated at the same time. Our results indicate that AT1A blocking can not effectively interrupt the Ang Ⅱ-induced synthesis of the protein and nucleic acid in cardiomyocytes.
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AIM: To investigate the effects of intracellular free calcium ([Ca 2+ ]i) from different resources on the proliferation mediated by mitogen activated protein kinase (MAPK) in vascular smooth muscle cells (VSMCs). METHODS: Cultured VSMCs were used in all experiments. Calcium influx was stimulated by angiotension Ⅱ(AngⅡ). The release of intracellular calcium stores was induced by inositol trisphosphate(IP 3) and ryanodine(RY). MAPK activity was measured by [?- 32 P]-ATP incorporation , MAPK protein expression by western blot, VSMCs proliferation by -Leucine(-Leu) and -Thymidine(-TdR) incorporation. RESULTS: Compared to the control VSMCs, AngⅡ, IP 3 and RY significantly increased [Ca 2+ ]i concentration , activity of MAPK and its protein content in VSMCs. The promotion of -Leu and -TdR incorporation in VSMCs was also observed ( P
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AIM: To investigate the protective effect of calcium antagonists on anoxia/reoxygenation (A/R) injury of cardiomyocytes. METHODS: Primary-cultured cardiomyocytes were divided into four groups, namely A/R, A/R+nifedipine(Nif), A/R+ruthenium red(Ru)+heparin (Hep) and control groups. The following parameters were measured in all groups: intracellular calcium concentration ([Ca 2+ ]i), cardiac cell viability, ATP content, lactate dehydrogenase (LDH) activity in the medium, PKC and MAPK activity and -Leucine(-Leu) incorporation. RESULTS: In comparison with A/R group, A/R+nifedipine(Nif) and A/R+ruthenium red(Ru)+heparin (Hep) groups showed a marked decrease in [Ca 2+ ]i and LDH content, and a significant increase in cell viability ,ATP content, activity of PKC and MAPK and -Leu incorporation(P
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Object To gain a clear idea on the resources of medicinal plants of Cynanchum L (Asclepiadaceae) in Gansu Province Methods Field investigation, specimen collection, taxonomic study and literature retrieval were carried out Results The distribution, ecological environment and medicinal parts of 18 species and 1 variety of 6 sections, including 17 medicinal plants, 2 Gansu newly recorded species have been clarified Conclusion The investigation may prove to be useful for the utility and further study of Cynanchum L in Gansu Province
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Ecological charactcristics and biological specific feature of Helleborus thibetanus Fran ch. were briefly described and cultivation technique to transform the wild plant into itscultivated variety was studied to provide a basis for the protection and expliotation ofthe wild resource of H. thibetanus.