OBJECTIVETo explore the effects of glucose concentration fluctuation on function of cultured bovine arterial endothelial cells and underlying mechanism.

METHODSThe thoracic aorta of newborn calf was used for primary endothelial cells culture. Cells were divided into 3 groups and cultured for 48 h: control group (C, 5.5 mmol/L), constant high glucose group (HG, 30 mmol/L) and glucose fluctuation (GF, three circles of 2 h 30 mmol/L followed by 3 h 5.5 mmol/L, 30 mmol/L overnight, repeat the whole procedure on the following day) groups. The membranes fluidity of endothelial cells was detected by fluorescence polarization method. The contents of sorbierite, aldose reductase (AR), sorbitol dehydrogenase (SDH) and advanced glycation end products (AGEs) were measured. RAGE, eNOS and ET-1 mRNA expressions were detected by semi-quantitative RT-PCR.

RESULTSThe membranes fluidity of endothelial cells in HG or GF group were significantly decreased compared with the control group (all P < 0.01) and significantly lower in GF group than those in HG group (all P < 0.01). Sorbierite, AR and AGEs concentrations were significantly higher in HG and GF groups than those in control group (all P < 0.01) and AR and AGEs concentrations were significantly higher in GF group than that in HG group (all P < 0.01). SDH of endothelial cells in HG or GF group were decreased compared with the control group and lower in GF group than in HG group (all P < 0.05). In addition, the mRNA levels of RAGE, eNOS and ET-1 were significantly upregulated compared with the control group (all P < 0.01).

CONCLUSIONSGlucose concentration fluctuation can result in more severe bovine arterial endothelial cells dysfunction than high glucose via activating polyols metabolic pathways, upregulating the expression of AGEs, eNOS and ET-1. Therefore, glucose concentration fluctuation might play a crucial role on macrovascular complications of diabetes.

Aldehyde Reductase ; analysis ; Animals ; Aorta, Thoracic ; cytology ; Cattle ; Cells, Cultured ; Endothelial Cells ; metabolism ; pathology ; Endothelin-1 ; analysis ; Endothelium, Vascular ; cytology ; metabolism ; Glucose ; metabolism ; Glycation End Products, Advanced ; analysis ; L-Iditol 2-Dehydrogenase ; analysis ; Membrane Fluidity ; Nitric Oxide Synthase Type III ; analysis

Carbon source plays an important role in the constitutive expression of foreign proteins in Pichia pastoris. In present study, glucose , glycerol , methanol and oil acid, was used respectively as the only carbon source to constitutively express hAS in Pichia pastoris GS115 (pGAP9K-AS)in shaking flask. The result shows that oleic acid is the best (163 mg/L) compared with glycerol (83mg/L), glucose (76 mg/L)and methanol (57 mg/L). Since oleic acid is insoluble in water, glycerol was used as the carbon source in the high-density cell culture of GS115 (pGAP9K-AS) in a 30 liter bioreactor and 169 mg/L of angiostatin was obtained after 48h of culture. The expressed angiostatin is immunologically active as shown by Western blotting. The recombinant hAS inhibits bFGF induced CAM angiogenesis and suppresses the growth of B16 melanoma in C57BL/6J mice. The tumor inhibition rate is 90% after 12 days of treatment. Statistics analysis revealed that the tumor volume difference of mice between the hAS group and PBS group is prominent (P < 0.01).
Angiogenesis Inhibitors ; biosynthesis ; genetics ; therapeutic use ; Angiostatins ; biosynthesis ; genetics ; therapeutic use ; Animals ; Bioreactors ; microbiology ; Culture Media ; pharmacology ; Fermentation ; Glycerol ; pharmacology ; Humans ; Melanoma, Experimental ; drug therapy ; Mice ; Mice, Inbred C57BL ; Pichia ; genetics ; growth & development ; metabolism ; Recombinant Proteins ; biosynthesis ; genetics ; therapeutic use

OBJECTIVEGua Sha and Blood-letting at the acupoints were Chinese traditional therapies for heatstroke. The purpose of present study was to assess the therapeutic effect of Gua Sha on the DU Meridian and Bladder Meridian combined with Blood-letting acupoints at Shixuan (EX-UE 11) and Weizhong (BL 40) on heatstroke.

METHODSAnesthetized rats, immediately after the onset of heatstroke, were divided into four major groups: Gua Sha group, Blood-letting group, Gua Sha combined with Blood-letting group and model group. They were exposed to ambient temperature of 43 °C to induce heatstroke. Another group of rats were exposed to room temperature (26 °C) and used as normal control group. Their survival times were measured. In addition, their physiological and biochemical parameters were continuously monitored.

RESULTSWhen rats underwent heatstroke, their survival time values were found to be 21-25 min. Treatment of Gua Sha combined with Bloodletting greatly improved the survival time (230±22 min) during heatstroke. All heatstoke animals displayed and activated coagulation evidenced by increased prothrombin time (PT), activated partial thromboplastin time (aPTT), D-dimer, and decreased platelet count, protein C. Furthermore, the animals displayed systemic inflammation evidenced by increased the serum levels of cytokines interleukin-1ß (IL-1ß), tumor necrosis factor α (TNF-α) and malondialdehyde (MDA). Biochemical markers evidenced by cellular ischemia and injury/dysfunction included increased plasma levels of blood urea nitrogen (BUN), creatinine, serum glutamic oxaloacetic transaminase (SGOT), serum glutamic pyruvic transaminase (SGPT), and alkaline phosphatase (ALP) were all elevated during heatstroke. Core temperatures (Tco) were also increased during heatstroke. In contrast, the values of mean arterial pressure were signifificantly lower during heatstroke. These heatstroke reactions were all signifificantly suppressed by treatment of Gua Sha and Blood-letting, especially the combination therapy.

CONCLUSIONGua Sha combined with Blood-letting after heatstroke may improve survival by ameliorating systemic inflflammation, hypercoagulable state, and tissue ischemia and injury in multiple organs.

Animals ; Blood Coagulation Disorders ; drug therapy ; therapy ; Bloodletting ; Combined Modality Therapy ; Complementary Therapies ; methods ; Cytokines ; blood ; Heat Stroke ; physiopathology ; Inflammation ; drug therapy ; therapy ; Ischemia ; drug therapy ; therapy ; Male ; Malondialdehyde ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Survival Rate

Objective To learn the status of wild freshwater fish and shrimp infected with metacercaria of clonorchis sinensis in Jinhua city.Methods Wild freshwater fish and shrimp were randomly captured in river channel,reservoir and pond from 3 counties according to the distribution characteristics of main river system in Jinhua city.Direct tabletting microscopic examination was used to detect metacercaria of clonorchis sinensis in the muscle of wild freshwater fish and shrimp.Results A total of 1 1 kinds of wild freshwater fish and shrimp were infected with metacercariae,accounting for 61 .1 1 %(1 1 /1 8),and the total infection rate was 5.63% among 2 326 wild freshwater fish and shrimp.The infection rate of fish(8.24%)was significantly higher than that of the shrimp(2.96%)(P <0.01 ).There were significant differences in the infection rate among different counties (P <0.01 ),and the infection rate in the downstream of the water system in Wu water area (1 2.90%)was the highest.Also,significant differences were observed in infection rate among different water environments (P <0.01 ),and the infection rate of pond (1 0.1 8%)was the highest.Significant differences were observed in the infection rate among different kinds of wild freshwater fish (P =0.00),and the infection rate of side skin fish(1 7.65%)and psendorasbora parve(1 7.65%)were the highest.Conclusion There were metacercaria of clonorchis sinensis infection in wild freshwater fish and shrimp with different degrees in Jinhua city.People who ate raw or undercooked freshwater fish and shrimp may be at the risk of infection.

This study is to investigate the chemical constituents from the whole plant Corydalis edulis. The chemical constituents were separated and purified by macroporous resin D101, silica gel, Sephadex LH-20, ODS, and semi-preparative HPLC. Their structures were determined by physicochemical properties and spectroscopic data. Four compounds were isolated from the dichloromethane and water extracts of the whole plant C. edulis, and identified as 6'-β-D-xylosylicariside B2(1),(3S,5R,6S,7E)-5,6-epoxy-3-hydroxy-7-megastigmen-9-one(2), loliolide(3), and 5,5'-dimethoxybiphenyl-2,2'-diol(4), respectively. Compound 1 is a new compound, of which the absolute configuration was established by electronic circular dichroism(ECD) calculations. Compound 4 is obtained from the plants of Papaveraceae family for the first time. Compounds 2 and 3 are firstly isolated from the Corydalis genus.
Chromatography, High Pressure Liquid ; Corydalis/chemistry* ; Glycosides/isolation & purification* ; Molecular Structure ; Phytochemicals/isolation & purification* ; Sesquiterpenes/isolation & purification*