OBJECTIVE: To investigate the prevention effect of aqueous extract of Epimedii sagittatum (ESE) on ovariectomy-induced (OVX) bone loss in rats. METHODS: Rats were divided into sham-operated and OVX groups. The OVX rats were divided into four groups treated with distilled water, 17beta-estradiol (1 mg/kg, ig) and ESE (0.5 and 1 g/kg, ig) for 11 weeks. Serum calcium, phosphorus, estradiol, bone gla protein concentrations and serum alkaline phosphatase activity were measured. Bone density was assayed by dual-energy X-ray absorptiometry. The undecalcified longitudinal proximal tibial metaphysical sections were cut and stained for the bone histomorphometric analysis. RESULTS: In OVX rats, alkaline phosphatase activity in serum was markedly increased by ESE treatment, which had no obvious influence on the body weight. Meanwhile, atrophy of uterus and descent of bone mineral density were suppressed by ESE treatment. In addition, ESE completely corrected the decreased concentrations of calcium and E2 in serum observed in OVX rats. Histological results also showed ESE prevented the increases in trabecular separation (Tb.Sp) in OVX rats whereas it did not alter trabecular thickness (Tb.Th) and trabecular number (Tb.N) in OVX rats. Moreover, ESE had remarkable effect on bone formation rate with bone volume as referent (BFR/BV) and bone formation rate with bone surface as referent (BFR/BS). CONCLUSION: The findings assessed on the basis of biochemical test, bone mineral density and histomorphometric parameters show that aqueous extract of Epimedii sagittatum has a definite antiosteoporotic effect and can prevent the OVX-induced bone loss in rats.

Objective To investigate the effect of 12-lipoxygenase(12-LO) on the p27Kip1 expression in diabetic glomeruli. Methods Mesangial cells were exposed to 12-LO product 12 (S)-HETE (10-7 mmol/L) with or without p38 MAPK (p38) inhibitor (SB203580, 1 μmol/L) for 24 hours. Rats fed with high fat diet received low dose streptozotoein (ST-Z, 35 mg/kg, IP injection) to develop type 2 diabetes and were divided into 2 groups: low dose STZ, low dose STZ+12-LO inhibitor cinnamyl-3,4-dihydroxy-α-cynanocinnamate (CDC, 8 mg/kg) treatment. Rats fed with regular chow were divided into two groups: controls, CDC treatment. The rats received injection of CDC or vehicle subcutaneously in the hind leg. CDC or vehicle injection was performed three times weekly on alternate days. All the rats were sacrificed after 4 weeks, Wild type and 12-LO knockout C57BL/6 mice were divided into 4 groups: wild type control, 12-LO knockout, STZ-induced wild type type 1 diabetes and STZ-induced 12-LO knockout type 1 diabetes. All the mice were sacrificed after 16 weeks. Urine, blood, kidney cortical tissue and isolated glomeruli by sieving method were collected at the end of study respectively. Western blot and immunohistochemistry for target protein were performed respectively. Results Inhibition of p38 activation could significantly reduce p27Kip1 expression induced by 12 (S)-HETE in mesangial cells (P<0.01). Increased glomerular volume, microalbuminuria, elevated glomeluli p38 activation, p27Kip1 expresssion in type 2 diabetic glomeruli was decreased after CDC treatment (P<0.01). Compared with wild type diabetic mice, glomerular p38 activation, p27Kip1 exprcsssion and extracellular matrix accumulation in the 12-LO knockout diabetic mice were significantly decreased (P <0.01, respectively). Conclusions 12-LO induces p27kipl expression via p38 pathway in diabetic glomeruli.

AIM To investigate the anticonvulsive action of supercritical CO2 ethanol extract from Pinellia Pedatisecta Schott(SEE-CO2PP). METHODS The rat convulsive model was induced by penicillin localized injected in rat cortex. The effects of SEE-CO2PP on the latency of seizure and changes of convulsive behaviors were investigated. The latency of epileptiform discharge, and frequency and amplitude of highest wave in cortex and hippocampus were recorded by using RM6240C multichannel physiological signal collection and analysis recorder. At the same time, the contents of glutamic acid (Glu), aspartic acid (Asp), glycine (Gly) and γ-aminobutyric acid (GABA) in hippocampus were determined with high performance liquid chromatography. RESULTS SEE-CO2PP 15 and 30 g·kg-1, ig, prolonged the latent period of seizure and weakened the extent. SEE-CO2PP also prolonged the latent period of epileptiform discharge, reduced the frequency and decreased amplitude of the highest wave in both cortex and hippocampus. Moreover, SEE-CO2PP increased the content of GABA in hippocampus, but the levels of Gly,Asp and Glu had no obvious changes. CONCLUSION SEE-CO2PP inhibits the epileptiform discharge and convulsive behaviors of convulsive model rats, which suggests SEE-CO2PP has anticonvulsive action.

Aim To investigate Jaridonin′s selective killing of cancer cells and explore the related molecular mechanism. Methods After treatment by Jaridonin for 24 h, the effect of Jaridonin on the cell viability was examined using MTT assay. The effect of Jaridonin on cytomorphology and mitochondrial membrane poten-tial (Δψm) was observed by a fluorescence microsco-py. The apoptosis of cell lines treated with Jaridonin, as well as the level of reactive oxygen species ( ROS ) was analyzed by flow cytometry. Expression of the pro-teins related with mitochondria apoptosis pathways was detected by Western blot. Results Jaridonin caused strong antiproliferative and apoptotic effects on MGC-803 cells, but there were not remarkable effects on GES-1 cells. Furthermore, the expression of Bax was up-regulated, and the release of cytochrome c from mi-tochondria to cytosol was also promoted in MGC-803 cells treated by Jaridonin. The cleavage of caspase-3 in MGC-803 cells was also observed. Jaridonin increased persistently intracellular levels of ROS in MGC-803 cells, whereas the level of ROS in GES-1 rose in the first stage, and then decreased, and dropped to the basic level after 6 h. More interestingly, Jaridonin-in-duced ROS accumulation and the inhibition of MGC-803 cell proliferation were almost completely attenuated in the presence of GSH. Conclusions Jaridonin se-lectively kills cancer cells and induces apoptosis in MGC-803 through ROS-mediated mitochondrial dam-age.

OBJECTIVE To establish and apply a method for the determination of plasma concentration of delamanid （DLM）. METHODS After plasma samples were treated with methanol precipitation， LC-MS/MS was adopted to determine the plasma concentration of DLM. The chromatographic column was Phenomenex SynergiTM Fusion-RP with mobile phase of methanol-0.1% formic acid solution （gradient elution）. The column temperature was 40 ℃ ， the flow rate was 0.3 mL/min， and the sample size was 1 μL. The ion source was electrospray ion source， and positive ion scanning was carried out in multi-reaction monitoring mode. The DLM ion pair used for quantitative analysis was m/z 535.0→352.0. The plasma concentration of DLM in 6 multidrug resistant tuberculosis （MDR-TB） patients were determined by the LC-MS/MS method. RESULTS The linear range of DLM was 0.05-8 μg/mL （r＝0.999 5）， and the lowest limit of quantitation was 0.05 μg/mL. RSDs of intra-batch and inter-batch precision were all less than 10%. The accuracy ranged 92.7%-104.9%. Average matrix effect was 94.3%-107.5%. Average recoveries were 93.2%-98.1%. The plasma concentration of DLM in 6 MDR-TB patients ranged from 0.61-2.76 μg/mL， with an average of （1.67± 0.74） μg/mL. CONCLUSIONS The established LC-MS/MS analysis method has good specificity， high sensitivity， accuracy and precision， and can be used to determine DLM plasma concentration in MDR-TB patients.

Objective To conduct in vitro transdermal test on triamcinolone acetonide spray solution, and investigate the effects of ethanol and propylene glycol alone or in combination on the in vitro transdermal function of triamcinolone acetonide spray solution. Methods Rabbit abdominal skin was selected, and the in vitro penetration test of triamcinolone acetonide spray solution was carried out by Franz diffusion cell method, and the content of triamcinolone acetonide was determined by HPLC. The rate of transdermal absorption was compared. Results The transdermal absorption rate of the combined use of ethanol and propylene glycol was significantly higher than that of the single use (P<0.05), and the order of promoting the penetration of triamcinolone acetonide spray solution when ethanol and propylene glycol were combined by 10% ethanol + 25% propylene glycol >10% ethanol + 20% propylene glycol >15% ethanol + 25% propylene glycol >15% ethanol + 20% propylene glycol. Conclusion The combination of 10% ethanol and 25% propylene glycol could optimize the transdermal function of triamcinolone acetonide spray solution.

OBJECTIVEThe aim of this study was to explore the molecular mechanism of apoptosis in esophageal cancer cells induced by Isodon rubescens.

METHODSThe DNA-damage effect of Jaridonin was detected by single cell gel electrophoresis (SCGE). The p53 protein was determined by Western blot. GSH assay kit was employed to determine the GSH content in human esophageal cancer EC-1 cells. Intracellular levels of hydrogen peroxide (H2O2) or superoxide (O(2).-) were determined using the redox-sensitive probes 2', 7'-dichlorodihydrofluorescein diacetate (DCF) or dihydroethidium (DHE), and the fluorescence signal was assayed by fluorescence microscopy and by flow cytometry.

RESULTSJaridonin induced DNA damage in EC-1 cells remarkably. The olive tail moments (OTM) of control and 20, 40 µmol/L Jaridonin were 3.2, 45.2 and 89.0, respectively. Compared with the control, the differences were significant (P < 0.01 for both). Jaridonin resulted in extensive p53 up-regulation in the EC-1 cells. More importantly, the p53 up-regulation occurred as early as 2 h after Jaridonin incubation, and in a time-dependent manner (P < 0.05). p53 siRNA transfection inhibited apoptosis in the EC-1 cells, and the Jaridonin-induced apoptosis rate was reduced from 38.5% to 8.8%. Intracellular level of H2O2 was increased by Jaridonin, whereas the level of O(2).- was barely changed. The GSH content in EC-1 cells was reduced from (10.3 ± 1.6) nmol/mg protein to (4.6 ± 2.1) nmol/mg protein after 20 µmol/L Jaridonin incubation for 8 h, and it was further reduced with the increase of Jaridonin concentration. Jaridonin induced DNA damage, H2O2 accumulation and apoptosis were significantly attenuated in the presence of GSH, but Jaridonin showed little effect on normal human liver L-02 cells.

CONCLUSIONSJaridonin selectively induces apoptosis in esophageal cancer EC-1 cells through H2O2-mediated DNA damage by depleting GSH.

Antineoplastic Agents ; pharmacology ; Apoptosis ; DNA Damage ; Diterpenes, Kaurane ; pharmacology ; Esophageal Neoplasms ; metabolism ; Humans ; Hydrogen Peroxide ; metabolism ; Up-Regulation