[Effects of triterpenoid from Psidium guajava leaves ursolic acid on proliferation, differentiation of 3T3-L1 preadipocyte and insulin resistance].

OBJECTIVE: To investigate the influences of triterpenoid from Psidium guajava Leaves (ursolic acid) on the proliferation, differentiation of 3T3-L1 preadipocyte, and its possible mechanism treat for insulin resistance. METHODS: 3T3-L1 preadipocyte was cultured in vitro. After adding ursolic acid to the culture medium for 48h, the cell viability was tested by MTT assay. Induced for 6 days, the lipid accumulation of adipocyte was measured by Oil Red O staining. The insulin resistant cell model was established with Dexamethasone. Cellular glucose uptake was determined with GOD-POD assays and FFA concentration was determined at the time of 48h. Secreted adiponectin were measured by ELISA. The protein levels of PPARgamma and PTP1B in insulin resistant adipocyte were measured by Western Blotting. RESULTS: Compared with medium control group, 30, 100 micromol/L ursolic acid could increase its proliferation and differentiation significantly (P < 0.05 or P < 0.01). Compared with the model group, ursolic acid at 100 micromol/L could enhance cellular glucose uptake of insulin resistant adipocyte significantly both in basic and insulin stimulation state (P < 0.01), while ursolic acid at 30 micromol/L could already enhance its glucose uptake significantly (P < 0.05), and could already decrease its FFA production significantly (P < 0.05). Ursolic acid at 30 micromol/L could increase the secretion of adiponectin on insulin resistant adipocyte significantly (P < 0.05), up-regulate the expression of PPARgamma protein (P < 0.05), but showed no effect on the PTP1B protein expression (P > 0.05). CONCLUSION: Ursolic acid can improve the proliferation and differentiation of 3T3-L1 preadipocyte, enhance cellular glucose uptake, inhibit the production of FFA, promote the secretion of adiponectin insulin resistant adipocyte, its mechanism may be related to upregulating the expression of PPARgamma protein.

[Nephro-protective effects of total triterpenoids from Psidium guajava leaves on type 2 diabetic rats].

OBJECTIVE: To investigate the nephro-protective effects of total triterpenoids from Psidium guajava leaves (TTPGL) on type 2 diabetic rats. METHODS: Diabetic rats were induced by intraperitoneal injection of streptozotocin (STZ, 35 mg/kg) and a high-fat diet. Diabetic rats were divided into five groups: diabetic model control, low-dose TTPGL-treated (60 mg/kg, L-TTPGL), medium-dose TTPGL-treated (120 mg/kg, M-TTPGL), high-dose TTPGL-treated (240 mg/kg, H-TTPGL) and rosiglitazone-treated (3 mg/kg, RSG). The rats received daily treatment for six weeks. At the end of the period,the levels of fasting blood glucose (FPG), fasting insulin (FINS), creatinine (Cr) and blood urea nitrogen (BUN) in serum were measured. Kidneys for histopathological evaluation were stained with Hematoxylin and Eosin (HE). RESULTS: Compared with normal control group, the level of FPG was increased, the insulin and insulin sensitivity index were decreased in the model group; The levels of BUN and Cr were increased with histopathological changes related to diabetic nephropathy in the kidney, which were the glomerular endothelium and mesangial cell proliferation, capillary narrowed, the base-membrane incrassation, glomerular swelling, cysts narrowed and tubules edema. Compared with the model group, the levels of FPG were decreased, serum insulin and insulin sensitivity index were increased significantly in M-TTPGL and H-TTPGL groups (P<0.01 or P<0.05); The levels of BUN and Cr were decreased significantly (P<0.01 or P<0.05) and the renal structural damages were improved significantly. CONCLUSION: TTPGL could decrease the level of blood glucose of diabetic rat effectively, increase the insulin sensitivity index and protect renal lesions in diabetic rats.

Doxorubicin-induced vasomotion and [Ca(2+)](i) elevation in vascular smooth muscle cells from C57BL/6 mice.

AIM: To explore the action of doxorubicin on vascular smooth muscle cells. METHODS: Isometric tension of denuded or intact thoracic aortic vessels was recorded and [Ca(2+)](i) in isolated aortic smooth muscle cells was measured by using Fluo-3. RESULTS: Doxorubicin induced phasic and tonic contractions in denuded vessels and increased levels of [Ca(2+)](i) in single muscle cells. Treatment with 10 micromol/L ryanodine had no effect on basal tension, but it did abolish doxorubicin-induced phasic contraction. Treatment with 10 mmol/L caffeine induced a transient phasic contraction only, and the effect was not significantly altered by ryanodine, the omission of extracellular Ca(2+) or both. Phenylephrine induced rhythmic contraction (RC) in intact vessels. Treatment with 100 micromol/L doxorubicin enhanced RC amplitude, but 1 mmol/L doxorubicin abolished RC, with an increase in maximal tension. Caffeine at 100 micromol/L increased the frequency of the RC only. In the presence of 100 micromol/L caffeine, however, 100 micromol/L doxorubicin abolished the RC and decreased its maximal tension. Treatment with 10 micromol/L ryanodine abolished the RC, with an increase in the maximal tension. In Ca(2+)-free solution, doxorubicin induced a transient [Ca(2+)](i) increase that could be abolished by ryanodine pretreatment in single muscle cells. The doxorubicin-induced increase in [Ca(2+)](i) was suppressed by nifedipine and potentiated by ryanodine and charybdotoxin. CONCLUSION: Doxorubicin not only releases Ca(2+) from the sarcoplasmic reticulum but also promotes the entry of extracellular Ca(2+) into vascular smooth muscle cells.

[Effects of extracts of Ginkgo biloba leaves on apelin concentration and expressions in plasma and myocardium of rats with myocardial ischemia injury].

OBJECTIVE: To investigate the changes of apelin in plasma and myocardium of model rats and the protective mechanisms of Extracts of Ginkgo biloba leaves (EGb) on myocardial ischemia injury induced by isoproterenol. METHODS: The model of myocardial ischemia injury was induced by subcutaneous injection of high dose isoproterenol. ELISA (enzyme linked immunosorbent assay) was used to measure the apelin concentration in plasma and myocardium. Semi-Quantitative RT-PCR was used to measure the apelin mRNA level in myocardium. The pathomorphology changes of myocardium was observed with light microscope. EGb was administered for 7 weeks. RESULTS: Compared with the normal control group, the apelin concentration in plasma and myocardium and the apelin mRNA level in myocardium significantly decreased in the model group (P < 0.01). Compared with the model group, the apelin concentration in plasma and myocardium and the apelin mRNA level in myocardium obviously increased in the EGb group (P < 0.01). Meanwhile, the NO content in serum also obviously increased and the pathological damage of myocardium was obviously improved. CONCLUSION: The protective mechanisms of EGb on myocardial ischemia injury may be related to the elevation of apelin contents and apelin mRNA level.

[Effects of EGb 761 on the cell apoptosis induced by H2O2 in RIN-m beta cells].

OBJECTIVE: To investigate the effects of ginkgo biloba extract (EGb 761) on apoptosis induced by hydrogen peroxide (H2O2) in RIN-m beta-cells. METHODS: The apoptotic model was made by H2O2 exposed for six hours with a concentration of 500 micromol/L The cytotoxicity was measured by MTT. Hoechst 33258 fluorescent staining were used to detect the protective effect of EGb 761 on the apoptosis of RIN-m beta-cells induced by H2O2. Annexin V-PI double staining of Flow cytometry were used to detect apoptosis quantitively. RESULTS: Compare to control group, after exposed to 500 micromol/L H2O2 for 6 hours, the apoptosis rate incereased and cell survival rate were decreased considerably (P < 0.01). Pretreated for 10 hours with EGb 761, the flow cytometry results showed that the apoptosis rate decreased and cell survival rate were increased considerably (P < 0.01, compared to H2O2 control group). CONCLUSION: EGb 761 can decrease RIN-m beta-cells damage and apoptosis induced by H2O2.

Serum from rabbit orally administered cobra venom inhibits growth of implanted hepatocellular carcinoma cells in mice.

AIM: To investigate the inhibitory effect of serum preparation from rabbits orally administered cobra venom (SRCV) on implanted hepatocellular carcinoma (HCC) cells in mice. METHODS: An HCC cell line, HepA, was injected into mice to prepare implanted tumors. The animals (n=30) were divided randomly into SRCV, 5-fluorouracil (5-FU), and distilled water (control) groups. From the second day after transplantation, 20 mg/kg 5-FU was administered intraperitoneally once a day for 9 days. SRCV (1,000 mg/kg) or distilled water (0.2 mL) was given by gastrogavage. Tumor growth inhibition was described by the inhibitory rate (IR). Apoptosis was detected by transmission electron microscopy (TEM), flow cytometry (FCM), and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL). Student's t-test was performed for statistical analysis. RESULTS: The tumor growth was inhibited markedly by SRCV treatment compared to that in the control group (P<0.01). The treatment resulted in a significant increase in the apoptotic rate of cancer cells by the factors of 10.5+/-2.4 % and 20.65+/-3.2 % as demonstrated through TUNEL and FCM assays, respectively (P<0.01). The apoptotic cells were also identified by characteristic ultrastructural features. CONCLUSION: SRCV can inhibit the growth of implanted HepA cells in mice, and the apoptosis rate appears to elevate during the process.

Mechanism underlying enhanced endothelium-dependent vasodilatation in thoracic aorta of early stage streptozotocin-induced diabetic mice.

AIM: To investigate the mechanism of the enhanced endothelium-dependent vasodilatation in thoracic aorta of the early stage streptozotocin (STZ)-induced diabetic C57BL/6J mice. METHODS: Radioimmunity was used to detect the metabolite of prostaglandin I2 (PGI2), 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha), in the blood serum. Vascular muscle tension and phenylephrine (PE)-induced rhythmic activity in the isolated thoracic aorta of mice were also compared. RESULTS: 6-Keto-PGF1 alpha in the serum was significantly higher in STZ-induced diabetic mice than age-matched controls [(1.8+/-1.0) microg./L vs (0.5+/-0.3) microg/L, P<0.01]. PE induced rhythmic activity in both diabetic and control mouse aorta but the amplitude was markedly higher in diabetic mice than in controls [(4.9+/-1.7) % vs (12+/-5) %, P<0.01]. PE, high K+ solution-induced contraction, and acetylcholine (ACh)-induced relaxation [(56+/-10) % vs (81+/-8) %, P<0.01] were notably enhanced in diabetic mice than those in controls. Alone NG-nitro-L-arginine methyl ester (L-NAME) or 6-(phenylamino)-5,8-quinolinedione (LY-83583) abolished the rhythmic activity and ACh-induced relaxation in controls but only partially inhibited them in diabetic mice. Indomethacin did not affect rhythmic activity but depressed ACh-induced relaxation. L-NAME plus indomethacin significantly depressed the rhythmic activity and ACh-induced relaxation than L-NAME alone (P<0.01). Furthermore tetraethylammonium plus L-NAME abolished them in diabetic mice. CONCLUSION: The mechanism that enhanced endothelium-dependent vasodilatation in STZ-induced diabetic mice is due to enhanced production of PGI2 and endothelium-derived hyperpolarizing factor (EDHF). The phenomena maybe only take place in early stage of diabetic mice.

Total saponins of Panax notoginseng protected rabbit iliac artery against balloon endothelial denudation injury.

AIM: To investigate whether total Panax notoginseng saponins (PNS) could protect endothelium of rabbit iliac artery against balloon endothelial denudation (BED) injury. METHODS: The morphology changes of the endothelium were observed with scanning electron microscope (SEM) and hematoxylin and eosin stain after BED of rabbit iliac artery at 0, 4, 6, and 8 week respectively. Vascular endothelial growth factor (VEGF) and matrix metalloproteinase-2 (MMP-2) was also determined by immunohistochemistry. PNS 10, 30, and 50 mg/kg were administered iv per day from 2 d before to 4 weeks after operation. RESULTS: The endothelium was denudated completely after BED. At the 4th week the endothelium was repaired in some degree, then recovered gradually at 6 and 8 week. The degree of intimal thickening at 4 week was significantly greater than that at 0, 6, or 8 week. The sequence of VEGF or MMP-2 staining from strong to weak was 4, 6, 0, 8 week, and normal control. However at 4 week, endothelial regeneration in PNS 30 and 50 mg/kg groups was significantly faster than that in saline group. The intimal thickness was significantly decreased and expressions of VEGF and MMP-2 were both down-regulated in PNS 30 or 50 mg/kg groups compared with saline control group. CONCLUSION: PNS promoted the endothelial regeneration and reduced ECM thickening, which was related to regulation of the expression of VEGF and MMP-2. PNS may have sustained antirestenotic effect after BED.