Ameliorative Effect And Underlying Mechanisms Of Total Triterpenoids From Psidium Guajava Linn. Leaves On Diabetic Peripheral Neuropathy In Rats

Objectives To investigate the effect of total triterpenoids from Psidium Guajava Linn. leaves(TTPGL) on diabetic peripheral neuropathy(DPN) rats by testing nerve functions, evaluating histopathological and measuring inflammatory expression. And explore mechanism of TTPGL by measuring gene expression of pro-inflammatory molecules(TNF-α, IL-6 and i NOS) and tissue proteins that NF-κB pathway mediated.Methods We used high-fat feeding and STZ(35 mg/kg) injection in the Sprague Dawley(SD) rats. Diabetic rats were randomized into 5 groups: TTPGL-30 group(30 mg/kg/d, i.g.); TTPGL-60 group(60 mg/kg/d, i.g.); TTPGL-120 group(120 mg/kg/d, i.g.); Diabetic mellitus(DM) group; rosiglitazone(RSG) group(3 mg/kg/d, i.g.) used as positive control. Other normal rats were conducted as Control group; After 8 weeks administration(i.g.), we conducted nerve conduction velocity and sensitive tests in vivo. In vitro, we performed histopathological evaluations of pro-inflammatory molecules(TNF- α, IL-6 and i NOS) by Hematoxylin and eosin(H&E) and immunofluorescence(IF) staining; Real Time-PCR was conducted to measuring the m RNA expression of pro-inflammatory molecules(TNF- α, IL-6 and i NOS) and Western Blotting was conducted to assess tissue proteins p-Akt/Akt,p-IKKα/IKKα,p-p65/p65.Results 1 Effects of TTPGL on body weight(BW) and fast blooding glucose(FBG). After 8 weeks of feeding, the body weight of the Control group increased from 254.5±14.8 g to 490.6±11.0 g(Mean±SD) during the entire study and FBG was in a stable level which under 7.0 mmol/L; DM group was significantly difference(P ≤ 0.01) in the FBG than that in the Control group and DM group was in a high blooding glucose which above 15.0 mmol/L. 2 Effects of TTPGL on MNCV, SNCV and thermal response latency.Compare to Control group, DM group was significantly slowed in MNCV and SNCV(P ≤ 0.01); While compare to DM group, TTPGL(120 mg/kg) and RSG groups were significantly fast in MNCV and SNCV(P ≤ 0.05). During the first 4 weeks, compare to the Control, DM group has shown no significantly difference in thermalgesia liminal value. TTPGL and RSG groups also shown no significantly difference to DM group. After 8 weeks, compare to the Control, DM group has shown significantly difference in thermalgesia liminal value(P ≤ 0.01); While compare to the DM group, TTPGL(120 mg/kg) and RSG groups were significantly sensitive in thermalgesia liminal(P ≤ 0.05). 3 Effects of TTPGL on sciatic nerve inflammatory mediator(TNF-α, IL-6 and i NOS) expression. Compare to Control group, DM has shown significantly difference in gene expression and IF staining(P ≤ 0.01); While compare to DM group, TTPGL groups and RSG group have their significantly ameliorative effects, specifically(P ≤ 0.05 or P ≤ 0.01). 4 Effects of TTPGL on sciatic nerve histological alterations. It was observed well organized myelin sheets round axons and absence of mononuclear infiltrate in Control group. Conversely, in DM group, the histological analysis revealed that diabetes significantly increased the degraded myelin sheets, edema as well as mononuclear cells. TTPGL treatment presented less myelin sheets appeared more organized than in DM animals. The above histological evaluation confirmed that TTPGL treatment ameliorates the structural changes in a dose-dependently manner. 5 The effect of TTPGL on NF-κB activation via the Akt/IKKα pathway. Compare to the Control group, DM has shown significantly increased in value of p-Akt/Akt,p-IKKα/IKKα,p-p65/p65(P ≤ 0.01). It means that the activation via the Akt/IKKα pathway was enhanced. After administration, TTPGL groups or RSG group has significantly decreased in expression of phospho-Akt, phospho-IKKα and phospho-NF-κB-p65. There have shown significantly difference compare to that of DM group(P ≤ 0.05 or P ≤ 0.01).Conclusions: These findings uncover a previously unrecognized mechanism for the neuro protective effects of TTPGL and support the concept that TTPGL contribute to the combined modulation of levels of NF-κB and inflammatory mediators may offer a novel therapeutic approach for the treatment of diabetic peripheral neuropathy.