| Objective:To investigate the contents of orthosiphon stamineus aqueous extract(OSE);to observe the effects of OSE on acute hyperuricemic in mice;to observe the effects of OSE on acute urarthritis in rats;to observe the effects of OSE in uric acid nephropathy rats and investigate the mechanism of renal protection and uric acid descending.Methods:OSE was extracted with traditional water extracting technology and the may contents of it was determined by High-performance liquid chromatography(HPLC).Acute hyperuricemic in mice was induced by hypoxanthine and potassium oxonate respectively,the levels of serum uric acid(SUA)was determined.Acute urarthritis in rats was induced by urate crystal to observe the effects of OSE on whole blood leukocyte,neutrophil and lymphocyte levels,the levels of NO in ankle joint and articular swelling.Uric acid nephropathy in rats was induced by yeast extract power and adenine to observe the effects of OSE on SUA,Serum Creatinine(SCr),Blood urea nitrogen(BUN),Urine Uric acid(UUA),Urine Creatinine(UCr),ranctional excretion of urine(FEUA),Renal index(RI),Serum Xanthine oxidase(SXO),Liver Xanthine oxidase(LXO),and 24hVurine.Rat kidneys were stained with hematoxylin and eosin(HE)for conventional morphological evaluation.The URAT1 and OUAT1 protein expression were investgated by immunohistochemistry.The URAT1 and OUAT1 mRNA expression were investgated by Polymerase chain reaction(PCR).The URAT1,OUAT1,bFGF and NLRP3 protein expression were investgated by Western blot(WB).Results:1.May contents of OSE was determined by HPLC,the results show that caffeic acid and rosmarinic acid were separated well with other contens.caffeic acid contents of crude drug in the sample was 8.4245mg/g,rosmarinic acid contents of crude drug in the sample was8.6457mg/g.2.For the acute hyperuricemic in mice,preventive therapy of OSE with 3g/kg,1.5g/kg and 0.75g/kg doses respectively for 7 days.Rats model made by hypoxanthine intraperitoneal injection in the 7th day and OSE markedly decrease the levels of SUA(P<0.01).3.Rats model made by intragastric administration of potassium oxonate for 7 days,at the same time by intragastric administration of OSE with 3g/kg,1.5g/kg and 0.75g/kg doses.The results show that the dose of 3g/kg markedly decrease the levels of SUA(P<0.05),but dose of 1.5g/kg and 0.75 g/kg had no effects on SUA levels.4.For the acute gouty arthritis in rats,preventive therapy of OSE with 6g/kg,3g/kgand 1.5g/kg doses respectively.Rats model made by injecting 0.1 ml of monosodium urate solution into the right ankle.The results show that OSE with 3g/kg effectively on the joint swelling at 6 to 48 hour and OSE with 6g/kg effectively on the joint swelling at 12 to 24 hour.Dose of 6g/kg and 3g/kg markedly decrease the levels of WBC,neutrophil and lymphocyte.Dose of 6g/kg and 3g/kg markedly increase the level of serum NO and decrease the level of tissue NO.5.yeast extract power and adenine(100mg/kg)has been used to induce model hyperuricemic in rats,at the same time by intragastric administration of OSE with 6g/kg,3g/kgand 1.5g/kg doses.(1)Serum biochemical indicator detection showed:the levels of SUA,SCr and BUN in model group were higher than in normal group(P<0.01),which in high dose and middle dose of OSE groups were lower than in model group(P<0.05 或 P<0.01).(2)Urine analysis in rats with uric acid nephropathy showed:OSE had no effects on 24hVurine during all groups.The levels of 24h UUA,24h UCr and FEUA in model group were higher than in normal group(P<0.01),which in high dose and middle dose of OSE groups were lower than in model group(P<0.05 或<0.01).(3)The levels of SXO and LXO in model group were higher than in normal group(P<0.01),which in high dose and middle dose of OSE were inhibit by high dose of OSE remakely(P<0.05 或 P<0.01).(4)The levels of RI in model group were higher than in normal group(P<0.01),which in all doses of OSE were lower than in model group(P<0.05 或P<0.01).(5)Kidney pathological morphological changes:Inflammatory cell infiltration and expansive interstitial fibrosis,simultaneous with large urate deposition in renal tubular when compared with the normal control rats.However,allopurinol have no obvious effects on renal trauma.As expect,these kidney pathological changes were alleviated by the treatment of OSE,especially OSE at 6g/kg ameliorated the development of renal tubular dilatation and inflammatory cell infiltration in a good way.(6)Immunohistochemical detection showed that IOD of URAT1 in model group were higher than normal group,IOD of URAT1 in high dose and middle dose of OSE groups were lower than model group;IOD of OAT1 in model group were lower than normal group,IOD of OAT1 in high dose and middle dose of OSE groups were higher than model group.(7)Real-Time PCR assay showed:The increased renal levels of URAT1 mRNA were observed in hyperuricemic rats compared to normal group(P<0.001),OS at6 and3 g/kg effectively down-regulated URAT1 mRNA(P<0.001,P<0.01 或 P<0.05);the decreased renal levels of URAT1 mRNA were observed in hyperuricemic rats compared to normal group(P<0.001),OS at6 and3 g/kg effectively up-regulated URAT1 mRNA(P<0.001 或 P<0.01).(8)Western blot assay showed:The increased renal levels of URAT1,bFGF and NLRP3 protein were observed in hyperuricemic rats compared to normal group(P<0.001或P<0.01),OS at6 and3 g/kg effectively down-regulated URAT1,bFGF and NLRP3 protein(P<0.001 或 P<0.01);the decreased renal levels of URAT1 protein were observed in hyperuricemic rats compared to normal group(P<0.01),OS at6 and3 g/kg effectively up-regulated URAT1 protein(P<0.001).Conclusions:1.OSE can reduce the levels of SUA,Scr and BUN of hyperuricemia rats and mice,improving the kidney function.2.OSE have a good curative effect on acute gouty arthritis rats.3.OSE can inhibit the levels of SXO and LXO,reducing the level of uric acid.4.OSE can improve kidney pathological morphology,down-regulated URAT1,bFGF and NLRP3 protein,protect and repair the nephron,ameliorated the development of renal tubular dilatation and inflammatory cell infiltration in a good way.5.OSE can down-regulate the expression of URAT1 and up-regulate the expression of OAT3 in order to reduce the level of serum uric acid by promoting excretion of uric acid. |
