Aim To establish the metho d of precise lung slicing and examine the validity of neutral red(NR) assay for measuring lung slices viability.Method Inflated with 1% low melting agaro se solution, lung lobes were cut into slices by shaking microtome. With the thic kness of 400,500,600,700 ?m,lung slices were cultured in medium pH 6.8,7 .0,7.2, 7.4, respectively. After 1 h pre-incubation, lung slices were cont inuously submerged in 24-well plate, incubated for 0,2,4,6 h,respectively. NR assay, MTT assay, LDH leakage and SOD activity were used to assess the slices viability under different slices thickness, medium pH and culturing time. Result When the slice thickness was 600 ?m and medium pH was 7.0, t he viability of slices maintained best and steady for 6 h. There were positive c orrelations between NR uptake and MTT reduction in slices under different thickn ess(r 1 = 0.91, P

To study the effect and mechanisms of 18 α-glycyrrhizic acid (18 α-GA) on cytochrome P450 (CYP) enzymes, the expression of CYP1A1, CYP2E1 and CYP3A was determined in rat hepatocyte sandwich cultures by using enzyme assay and semi-quantitative reverse transcriptase-polymerase chain reaction(RT-PCR). The results showed that the activities of CYP1A1 (7-ethoxyresorufin O-deethylase, EROD), CYP2E1(aniline hydroxylase, ANH) and CYP3A (erythromycin N-demethylase, ERD) were decreased in concentration-dependent manner after treatment with 18 α-GA(50－400 mg*L-1), and at the concentration of 200 mg*L-1 inhibitory rate reached the maximum (the maximum inhibitory rate was 59.6%, 69.7% and 44.7%, respectively). The time course revealed that the inhibition reached plateau level at d 4 of culure. 18 α-GA Decreased CYP1A1, CYP2E1 and CYP3A1 mRNA expression in dose-dependent manner, the maximum inhibitory rate was 44.5% , 58.1% and 37.1%, respectively. The results suggest that 18 α-GA down-regulate CYP expression at the transcriptive levels.

AIMTo establish a collagen sandwich system (CSS) wit h self-prepared collagen for rat hepatocytes culture[FQ(14*2。46,X-WZ]in vitro, and study the effect of CYP3A substrates on cyt o chrome P450 activities of hepatocyte cultured by CSS. METHODSS terilized type Ⅰ collagen was prepared from rat tail tendons. The morphology of hepatocyte cultured by collagen coated (CC) and CSS method was investigated and compared. Activities of ALT, AST, and LDH in aliquots of extracellular medium w ere measured by biochemical assay. CYP1A, CYP2E1 and CYP3A activities of hepatoc ytes were also determined. RESULTSBy CSS method, the shape of hepatocytes was maintained, and the structure of cells was integrated after cult ivated for 6 d. However, in CC method after 3 d, hepatocytes dropped from the co llagen matrix, and the activities of ALT, AST and LDH in aliquots were increased . As compared with control group, in CSS cultivated hepatocytes, the activities of CYP3A increased 33%, and CYP2E1 decreased 45% by nimodipine, and CYP3A was in duced 1 94-fold (P

Object To investigate the effects of ethanol sediments obtained from the tuber of Angelica sinensis (Oliv). Diels (ESA) and its litmusless component (ESA 1) on the secretion of TNF ? and IL 1 by mice peritoneal macrophages in vitro. Methods L929 cell line cytotoxicity was used for the assay of TNF ?. The proliferation of L929 cell line was used for the assay of IL 1. Results The secretion of TNF ? and IL 1 by mice peritoneal macrophages which were co cultured with ESA or ESA 1 in vitro can be significantly promoted. At the concentrations in range of 5～20 ?g/mL, there is a dose dependence in the action of ESA, while there is not the similar effect of ESA 1, even though it showed the marked effect. Conclusion ESA and ESA 1 can enhance the secreting TNF ? and IL 1 of mice peritoneal macrophages in vitro.

AIM To explore a long-term Kupffer cell culture method, which can best maintain the function and activity of cell and decrease the cellular damage. METHODS Lactic dehydrogenase (LDH) in the supernatants were measured to observe the severity of cellular damage ,the function of Kupffer cell was reflected with the nitrogen monoxide(NO) content and the intracellular acid phosphatases(ACP) activity. In the conditions of common culture(CC),single collagen culture(SCC) and double collagen culture(DCC), with or without serum, study the maintenance of cellular function and the severity of cellular damage. RESULTS Compared with the other two methods, the LDH contents were significantly lower( P

AIM　To study the effect of 18α-glycyrrhizic acid (18α-GL) on hepatic microsomal drug metabolizing enzymes in rats. METHODS　18α-GL (12.5, 50.0 mg*kg-1*d-1) were given ip to male Wistar rats for 3, 6 or 12 consecutive days. The rats were sacrificed 24 h after the last dose and the liver microsomes were prepared for analysis of cytochrome P450 (CYP) isozymes and phase II transferase activites. RESULTS Aniline hydroxylase (CYP2E1) activities in the rats treated with 18α-GL (12.5, 50.0 mg*kg-1) for 6 days decreased dose-dependently by up to 53.2%; For 3, 6 or 12 days 7-ethoxyresorufin O-deethylase (CYP1A1) activities in the rats of 50 mg*kg-1 dose group decreased time-dependently by 17.6%, 38.3% and 47.3%, respectively; Erythromycin N-demethylase (CYP3A) activities was significantly inhibited from 23.1% to 34.3%. UDP-glucuronosyltransferase activities toward 7-hydroxy-4-methylcoumarin significantly increased ranging from 19.3% to 29.9%. UDP-glucuronosyltransferase activities toward 4-phenylphenol in the rats treated with 18α-GL (12.5, 50.0 mg*kg-1) for 6 days increased by 45.9% and 70.3%. Glutathione S-transferase (GST) activities in the rats treated with 18α-GL (12.5，50.0 mg*kg-1) for 6 days increased by 13.7% and 48.3% in dose-dependent manner. CONCLUSION　18α-GL inhibited rat liver microsomal cytochrome P450 while induced phase II transferase.

BACKGROUND: In valerian-ligusticum extract (VLE), valeriana offici nalis extract (VOE) is γ aminobutyric acid (GABA) receptor kinetin, which can relax cerebral vascular spasm; ligusticum wallichii Fr. Extraxt (LWE)can pass through blood-brain barrier, enhance microcirculation of tissue and inhibit blood platelet aggregation and 5-Hydroxytryptamine (5-HT) release.OBJECTIVE: To probe into the effects of VLE prepared with effective components on prevention and treatment of cerebral ischemic injury.DESIGN:Complete randomized, negative and positive control experiment.SETTING: Institute of Senile Medicine and Pharmacology of Tongji Medical College of Huazhong University of Science and Technology.MATERIALS: The experiment was performed in Institute of Senile Medicine Pharmacology of Tongji Medical College of Huazhong University of ent blood perfusion in brain tissue: Fifty Kunming mice were employed,which was randomized into normal group, solvent control (model) group,ligustrazine 50 mg/kg group, VLE 170 mg/kg group and VLE 85 mg/kg Fifty Wistar rats were employed, which was randomized into solvent control (model) group, compound danshen (Radix Salviae Miltiorrhizae) 5 g/kg group,VLE 156 mg/kg group, VLE 94 mg/kg group and VLE 31.3 mg/kg group,Sixty Wistar rats were employed, which was randomized into sham-operation group, solvent control (model) group, ligustrazine 10 mg/kg group, VLE 156 mg/kg group, VLE 95 mg/kg group and VLE 31.3 mg/kg, 10 mice in each were employed, which was randomized into normal group, solvent control (model) group, ligustrazine 10 mg/kg group, VLE 200 mg/kg group and VLE 40 mg/kg, 10 mice in each one.sue, in advance, VLE (85, 170 mg/kg), ligustrazine (50 mg/kg) or solvent enhancer of equal volume (0.2 mL) were injected abdominally in each group. Twenty minutes later, pituitrin (2.5 u/kg) was injected intravenously; and 10 minutes later, isotope 99Tcm+ L, L-EthylCysteinate Dimer and Stannous Chloride (ECD) 3.7×1010Bq/ L(0.1 mL/per mouse) was injected in coccygeal nerve. Fifteen minutes later, radio-immunity counter was used periment of arteral-ovenous bypass method for thrombosis, before the opercal saline successively, continuously for 7 days, once per day. After 24 hours of medication pause, with abdominal anesthesia with pentobarbitol sodium, a catheter (with surgical thread inside) was used in vitro to connect common cervical vein and carotid artery. Thrombus mass was scaled 15 dominal anesthesia of chloral hydrate, intraluminal thread approach (ITA)was used to block unilateral MCA. Except that ITA was not used, the other management in sham-operation group was same as experimental groups.Gastric perfusion was done with VLE(156, 94, 31.3 mg/kg), ligustrazine operation and 3 hours and 12 hours after operation. 24 hours after modeling, the assessment was done for behavioral neurological damage and brain sive cerebral ischemia experiment, the model was prepared by coccygeal injection of collagen + adrenalin (AD). Respectively, 30 minutes before modeling injection and 1 hour after injection, gastric perfusion was done with VLE (200, 40 mg/kg), ligustrazine (10 mg/kg) or solvent enhancer of equal volume successively to observe the numbers of dead mice in 5 minutes after modeling and the numbers of hemiplegia mice in 15 minutes;and to determine brain mass index 8 hours later after sacrificed and lactic acid level of brain tissue homogenate with ultraviolet spectrophotometry.group.RESULTS: In the experiment of acute extensive brain ischemia in mice, in solvent control, during modeling, 3 mice were died and the rest 207 mice brain tissue in mice, the ratios of brain with and blood γ ray pulsating intensity in VLE 85 mg/kg group and VLE 170 mg/kg were higher than model group (0.53±0.09, 0.55±0.08, 0.45±0.08, t=2.234 6, 2.793 3, P method in rats, the thrombus masses in VLE 156 mg/kg group, 94 mg/kg group and 31.3 g/kg group were lower remarkably than the model group [(12.66±4.79), (13.31 ±3.97), (13.49±4.09), (19.21±5.76) g, (t=2.667 0,31.3 mg/kg group, 94 mg/kg group and 156 mg/kg group was lower remarkably than model group successively [(5.9±1.9), (6.0±2.0), (5.8±2.2),(8.7±0.9) score], and cerebral infarction index was lower than model group [(16.52±5.78)%,(16.54±3.00)%, (14.18±6.13)%, (24.03±4.85)%, (t=3.118 9-chemia in mice, brain mass indexes of VLE 40 mg/kg and 200 mg/kg groups were lower remarkably than model group [(0.91 ±0.20) and (0.82±0.24)%, (1.40±0.32)%], and lactic acid in brain tissue was lower than model group [(17.44±6.71),(14.43±2.81), (29.07±7.33) μmol/g (t=3.388 5-5.800 5, P＜ 0.01)].CONCLUSION: Valerian-liqusticum extract improves significantly cerebral ischemia in mice induced by pituitrin and the damage by medium cerebral artery embolism in rats, and it inhibits significantly blood platelet aggregation and thrombosis induced by AD+ collagen mixture or foreign objects. It is suggested that valerian-ligustrazine extract prevents and treats significantly the perfusion disturbance of cerebral microcirculation.