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Objective To observe the effect of different concentrations of realgar on the apoptosis of gastric adenocarcinoma cell Line SGC-7901. Methods MTT assay was conducted to detect the growth inhibition ratio of SGC-7901 cells treated by different concentrations of realgar (0,10,20,50 and 80 μg·mL-1 ). AnnexinⅤ/ Propidium iodide double staining method together with flow cytometry were used to detect the effect of realgar at 0,20,50 and 80 μg·mL-1 on the apoptosis of SGC-7901 cells. Results The growth inhibition ratio of SGC-7901 cells by various concentrations of high-purity realgar (10,20,50 and 80 μg·mL-1 ) was 6. 15% ,7. 54% ,42. 31% and 63. 54% ,respectively,24 h after realgar treatment. At 48 h after the treatment,the growth inhibition ratio rose to 32. 56% ,46. 14% ,64. 51% and 87. 52% ,respectively (P0. 05). Conclusion High purity realgar can inhibit the proliferation and induce early apoptosis of gastric adenocarcinoma SGC-7901 cells.
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BACKGROUND: Preactivation of peripheral blood mononuclear cells (PBMCS) is one of the most important eerly events and facilitating factor for the formation of atherosclerosis. Tanshinone is a lipolytic component extracted from traditional Chinese medicine of denshen, it has definite anti-atherosclerotic effect.OBJECTTVE: To analyze whether PBMCS preactivation existed at early essential hypertension, and investigate the effects of tanshinone on inhibiting the PBMCS activation cultured in vitro by detecting the adhesion and excretory activities of PBMCS.DESTGN: A case-controlled analysis.SETTING: Department of Emergency and Research Room of Traditional Chinese Medicine, Tongji Hospital affiliated to Tongji Medical College, Huazhong University of Science and Technology.PARTTCTPANTS: Thirty patients with untreated essential hypertension or with withdrawal from antihypertensives for at least 2 weeks were selected from the Department of Cardiology, Tongji Hospital affiliated to Tongji Medical College,Huazhong University of Science and Technology from January 2003 to October 2004, including 16 males and 14 females, aged (44.6±7.4) years, body mass index of (26.2±4.5) kg/m2, average disease course of (38.5±16.9) months.Informed contents were obtained from all the subjects. Their hypertension was grade Ⅰ-Ⅱ according to the diagnostic standards for hypertension by WHO/ISH in 1999. Secondary hypertension, organic heart disease, hyperglyceridemia,diabetes mellitus, liver and kidney dysfunction, heart, brain, kidney, vessel and other target damaged induced by infection and other clinical conditions and hypertension were excluded by history, physical examination and assistant examination.Another 30 healthy physical examinees with normal blood pressure were enrolled as the normal control group. Human umbilical vein endothelial cells (Species Reserving Center of Wuhan University); Tanshinone injection (Yaan Sanjiu Pharmaceutical, Co., Ltd., batch number: 020724);METHODS: ① Venous blood samples (4.0-5.0 mL) were drawn from all the subjects, and mononuclear cells were separated by means of Ficoll-Hypaque density gradient centrifugation and plastic adhesion, then incubated at 37 ℃ for 40-60 minutes, and the adherent cells were the PBMCS. These cells (viability ＞ 95%, Trypan blue staining) had the characteristics of mononuclear cells (Wright staining). The newly separated adherent PBMCS were resuspended, and then inoculated to the 24-well plate (4×107 L-1). There were 3 wells for each sample: the first was for basic excretion, the second for angiotensin Ⅱ stimulation, and the third for tanshinone pretreatment. The PBMCS were co-incubated with tanshinone for 30 minutes before angiotensin Ⅱ stimulation. The terminal concentration was 1×10-8 mol/L and 1×10-8 g/L for angiotensin Ⅱ andtanshinone respectively, and that of PBMCS was 2×107 L-1. The cells were cultured in the incubator (CO2 of 0.05 in volume fraction) at 37 ℃ for 24 hours, then the supernatant and cell ingredients were collected respectively. ② The PBMCS suspension was preparedl, and the cellular density was adjusted to 2.5×109 L-1. The human umbilical vein endothelial cells were cultured on the 24-well plate with M199 medium containing fetal bovine serum (0.1 in volume fraction), and spread to monolayer after the cells entered the logarithm phase. Each well was added with PBMCS suspension (100 μL), incubated at 37 ℃ for 2 and 4 hours respectively. The unadherent cells were removed, and the adherent ones were counted after fixed with 20 g/L glutaral, 40 visual sights were counted for each well under high power microscope, and the average value was used. ③ The double-antibody sandwich enzyme-linked immunoabsorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR) were used to determine the concentrations of tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β) and interleukin-6 (IL-6) in supernatant of PBMCS, and the expressions of their mRNA.MATN OUTCOME MEASURES: ① Changes of PBMCS adhesion activity; ② Concentrations of cytokines and their mRNA expressions in supernatant of PBMCS.RESULTS: ① At 2 and 4 hours of inoculation, the numbers of PBMCS adhered to endothelial cells under basic conditions were similar between the hypertension group and normal control group (t =1.153-1.577, P ＞ 0.05); After angiotensin Ⅱ stimulation, the adherent cells were obviously more in the hypertension group than in the normal control group (t =3.842-4.536, P ＜ 0.01); The numbers of the adherent cells were decreased to the same levels after tanshinone pretreatment (t =0.855-1.702, P ＞ 0.05). ②Under basic state, the concentrations of TNF-α, IL-1β and IL-6 in supernatant were all lower in both groups (t =0.981-1.829, P ＞ 0.05); The concentrations of the cytokines after angiotensin Ⅱ stimulation were obviously higher in the hypertension group than in the normal control group (t = 2.442, 5.075, P ＜ 0.01,0.01, 0.05); The concentrations of the cytokines after tanshinone pretreatment were all decreased to different extent, and there were no significant differences (t =1.227-1.940, P ＞ 0.05). Similar changes were observed in the mRNA expressions of the cytokines in PBMCS in the two groups.CONCLUSTON: ① The number of PBMCS adhered to endothelial cells, the concentrations and mRNA expressions of the cytokines under basic state in patients with essential hypertension were at the levels of normal subjects, and they were significantly increased after angiotensin Ⅱ stimulation, suggesting that the PBMCS were at preactivation at early essential hypertension. ② Tanshinone could decrease the adhesion and excretory activities of PBMCS in patients with essential hypertension to the normal levels, it is proved that tanshinone can inhibit the further activation of the preactivated PBMCS, and can prevent the occurrence of atherosclerosis to some extent.
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BACKGROUND: As a reliable marker of inflammation, intercellular adhesion molecule-1 (ICAM-1) plays an important role in atherogenesis. Recently, it is assumed in researches of recent years that chronic inflammation mediated by ICAM-1 is involved in hypertensive left ventricular hypertrophy probably, but there are still some objections reported. Tanshinone ⅡA is a kind of liposoluble extract from danshen(Radix Salviae Mitiorrhizae) . It is verified in animal experiment that it can inhibit hypertensive left ventricular hypertrophy.OBJECTIVE: To investigate the relationship between ICAM-1 and hypertensive left ventricular hypertrophy and the effect of tanshinone ⅡA on expression of ICAM-1.DESIGN:A randomized controlled observation was designed.SETTING: Tongji Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology.MATERIALS: The experiment was performed in the Laboratory of Emergency Department of Tongji Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology from October 2002 to January 2004. Ten male WKY rats(10 WKY rats) of 12-week-old and 20 rats with spontaneous hypertension(20 SHR rats) were employed and divided into the control group(10 WKY rats), hypertension group(10 SHR rats) and tanshinone ⅡA group(10 SHR rats).injected from caudal vein for treatment and distilled water at the same dose was injected in the other two groups. Twelve weeks later, the rats were sacrificed and myocardium was collected for specimen preparation. Haematoxylin and eosin(HE) staining, VG staining, immunocytochemical staining and myocardial ED1 labeling were applied to determine myocardial macrophage infiltration degree. The reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay(ELISA) were used to determine the expressions of ICAM-1 mRNA and protein.MAIN OUTCOME MEASURES: Degree of macrophages infiltration and the expressions of ICAM-1 mRNA and protein in myocardium in each group.RESULTS: Twelve SHR and 10 WKY rats were employed in the experiment pared with the control group, ICAM-1 mRNA and protein expressions in hypertrophic myocardium were increased significantly in hypertension group (0.176±0.087,0.537±0.195;0.104±0.011,0.173±0.027, P ＜0.01or P ＜ 0.05) . Infiltration of macrophage was significant(0. 62 ±0.07,non Ⅱ A group, the expressions of ICAM-1 mRNA and protein were decreased significantly(0. 537 ±0. 195, 0.291 ±0. 106; 0. 173 ±0.027, 0. 125± 0. 014, P ＜ 0.01 or P ＜ 0. 05); the amount of macrophages infiltration was decreased(1.85 ±0. 23, 1.16 ±0. 17, P ＜ 0.05) and the degree of cardiomyocytic hypertrophy and interstitial fibrosis was remarkably relieved.CONCLUSION: Excessive expression of myocardial ICAM-1 and its mediated inflammatory cellular infiltration plays an important role in hypertensive left ventricular hypertrophy. The inhibition of tanshinone ⅡA on left ventricular hypertrophy may be contributed to decreased expression of ICAM-1 and alleviated inflammatory cellular infiltration in myocardium.
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BACKGROUND:Aldosterone is main pathogenic factor of left ventricular hypertrophy, for which, to inhibit its biosynthesis effectively may prevent and treat hypertension induced by left ventricular hypertrophy.OBJECTIVE: To investigate the effects of Tanshinone Ⅱ A on myocardiac aldosterone synthesis and expression of its relevant genic CYP11B1and CYP1 1B2 in spontaneously hypertensive rats(SHR) and explore the possi ble nechanism of Tanshinone Ⅱ A on inhibiting hypertension induced by left ventricular hypertrophy.DESIGN:SHR were employed as the objects in the experiment and WKY rats with normal blood pressure (WKY rats) were employed in the controlgroup. Complete group division and randomized control experiment was designed. Analysis of variance was used for the means comparison among groups.SETTING :Department of Emergency and Institute of Integrated Traditional Chinese and Western Medicine in Tongji Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology.MATERIALS :The experiment was performed in the laboratory of Department of Emergency in Tongji Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology from January 2003 to February 2004, in which, 10 male WKY rats with normal blood pressure of 12-week-old were employed in the control group and 20 male SHR of 12-week-old were randomized into two groups, named hypertension group and Tanshinone ⅡA group, 10 rats each group.METHODS:In Tanshinone Ⅱ A group, Tanshinone Ⅱ A 1.5 mg/kg was injected abdominally everyday. The distilled water of same volume was injected abdominally in hypertension group and the control group. Twelve weeks after experiment, myocardial specimens were collected after rats sacrificed. Radioimmunoassay was used to determine the contents of aldosteron and angiotensin Ⅱ in myocardial tissue and reverse transcriptase polymerase chain reaction (RT-PCR) were used to measure mRNA level of CYP11B1 and CYP11B2 genes relevant to aldosterone synthesis.MAIN OUTCOME MEASURES: Contents of aldosterone and angiotensin Ⅱ in myocardial tissue and mRNA expression of CYP11B1 and CYP1 1B2 genes.RESULTS:All of 20 SHR and 10 WKY rats entered result analysis. ① Myocardial aldosterone content of rats in 12 weeks after experiment: that in hypertension group and Tanshinone Ⅱ A group was higher remarkably than that in the control group [(0.056±0.014), (0.031±0.010), (0.018±0.009) ng/g,P ＜ 0.01, 0.05]; that in Tanshione Ⅱ A group was lower remarkably than that in the hypertension group (P ＜ 0.05). ② Myocardial angiotension Ⅱ content of rats in 12 weeks after experiment: that in the hypertension group and Tanshinone ⅡA group was higher remarkably than that in the control group [(0.093±0.016), (0.088±0.024), (0.043±0.012) ng/L, P ＜ 0.01]. ③ Expression of CYP11B1 gene in myocardial tissue of rats in 12 weeks after experiment: that in the hypertension group and Tanshinone Ⅱ A group was higher remarkably than that in the control group (2.774±0.138, 2.533 ±0.127, 1.973±0.102, P ＜ 0.05). ④Expression of CYP11B2 gene in myocardial tissue of rats in 12 weeks after experiment; that in the hypertension group and Tanshinone ⅡA group was higher remarkably than that in the control group (1.573±0.106, 1.024±0.113, 0.786±0.121, P ＜ 0.01,0.05); that in the Tanshinone ⅡA group was lower remarkably than that in the hypertension group (P ＜ 0.05). ⑤The electrophoresis band positions of myocardial CYP11B1 and CYP11B2 as well as RT-PCR products of glyceraldehydes-3-phosphate dehydrogenase (GAPDH), the consulting gene,were in conformity with the theoretic values.CONCLUSION:The inhibition of Tanshinone Ⅱ A on hypertension induced by left ventricular hypertrophy is probably contributed to its downregulating effect on myocardiac CYP11B2 gene expression relevant to aldosterone synthesis and to its reducing action on local biosynthesis of aldosterone in heart.
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To investigate the effect of Jianpiyiqi prescription on the expression of heat shock proteins (HSPs) and the changes of HSPs in gastric-ulcerated rats, the rat model of chronic gastric ulcer was induced by acetic acid. The SABC immunohistochemical method was used to observe HSP70 of mucosa around the gastric ulcer. Imaging analysis was performed. Western dot blot was used to detect HSPs contents in the plasma and gastric mucosal homogenate in each group. The results showed that HSP70 contents of the mucosa around the gastric ulcer in the model group and ranitidine-treated group were increased as compared with control group (P<0.01). Jianpiyiqi could increase the expression of HSP70 of the mucosa around the gastric ulcer further as compared with that in the model group and ranitidine-treated group (P<0.01). The HSP70 contents in the serum and mucosa in the model group and ranitidine-treated group were increased as compared with control group (P<0.01, P<0.05 respectively). HSP70 of serum and mucosa in the Jianpiyiqi-treated group was higher than in the model group and ranitidine-treated group (P<0.05, P<0.01 respectively).It was concluded that HSP might play a role in the process of pathophysiology of gastric ulcer. Jianpiyiqi could enhance gastric ulcer-healing through the protective mechanism of HSPs.
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To investigate the effect of Jianpiyiqi prescription on the expression of heat shock proteins (HSPs) and the changes of HSPs in gastric-ulcerated rats, the rat model of chronic gastric ulcer was induced by acetic acid. The SABC immunohistochemical method was used to observe HSP70 of mucosa around the gastric ulcer. Imaging analysis was performed. Western dot blot was used to detect HSPs contents in the plasma and gastric mucosal homogenate in each group. The results showed that HSP70 contents of the mucosa around the gastric ulcer in the model group and ranitidine-treated group were increased as compared with control group (P<0.01). Jianpiyiqi could increase the expression of HSP70 of the mucosa around the gastric ulcer further as compared with that in the model group and ranitidine-treated group (P<0.01). The HSP70 contents in the serum and mucosa in the model group and ranitidine-treated group were increased as compared with control group (P<0.01, P<0.05 respectively). HSP70 of serum and mucosa in the Jianpiyiqi-treated group was higher than in the model group and ranitidine-treated group (P<0.05, P<0.01 respectively).It was concluded that HSP might play a role in the process of pathophysiology of gastric ulcer. Jianpiyiqi could enhance gastric ulcer-healing through the protective mechanism of HSPs.