The Study Of The Effects Of FFNT-5 On Renal Fibroblasts In UUO Model Rats

Part 1 The study of the effects of different concentrations of FFNT-5 on renal fibroblasts in ratsBackground:Renal interstitial fibrosis is caused by the excessive deposition of a variety of extracellular matrix components in the renal interstitium. Interstitial fibroblasts are the principal intrinsic renal cells and are the most important extracellular matrix secreting cells in renal interstitial fibrosis. Thus, preventing the process of renal fibrosis lies in blocking the excessive fibroblast proliferation and secretion in the kidney. The more complex mechanism of occurrence of renal interstitial fibrosis is not yet fully understood and may be related to the following:interstitial fibroblast proliferation, activation and phenotype;transformation of renal tubular epithelial cells to myofibroblasts; vasoactive substances and cytokines acting on the cell biology,the process of extracellular matrix production and degradation; extracellular matrix production increase and /or reduction and eventually leading to renal interstitial extracellular matrix accumulation and fibrosis. Based on the pathogenesis of renal interstitial fibrosis, and characteristics of targets and multiple linkages, research and development of an effective anti-fibrosis drugs has become a hot topic in kidney disease, and a daunting challenge. AKF-PD independently designed by Central South University,is a new synthetic pyridine ketone, which has obtained national patent. Preliminary experiments in GLP laboratories have proven their low toxicity in cellular and animal experiments.Our data shows that AKF-PD exerts a strong antifibrotic effect, as shown in experimental renal fibrosis in vivo and in vitro.; FFNT-5 is a new pyridine ketone that we constituted. in earlier renal antifibrosis drug experiments, we preliminary found that FFNT-5 can inhibit the growth of renal fibroblasts and proposed to investigate the effect of different concentrations of FFNT-5 on renal fibroblasts proliferation. Objective:Observation of effect of FFNT-5 on renal fibroblast proliferationMethods:MTT assay of different concentrations of FFNT-5 in rat kidney fibroblast (BHK-21) proliferates. Normal control group, FFNT-5 (100μg /ml) group, FFNT-5 (200μg/ml) group, FFNT-5 (400μg/ml) group, AKF-PD (100μg/ml) group, AKF-PD (200μg/ml) group and the AKF-PD (400μg/ml) group were established with monitoring at different time points (24,48,72 hours) Enzyme OD test values and observations under inverted microscope were used to assess cell morphology and growth.Results:The effect of different concentrations of FFNT-5 on rat kidney fibroblasts at 24h,48h,72h time could significantly inhibit cell proliferation in a dose-dependent mannerConclusion:FFNT-5 can inhibit rat kidney fibroblasts proliferation.Part 2 The study of the effects of different doses of FFNT-5 on renal fibroblasts in UUO model ratsBackground:Renal interstitial fibrosis is the outcome and common pathway of development of conditions ranging from chronic kidney disease to end stage renal failure; Are involved a variety of pathogenic factors such as injury, inflammation, medication, genetic factors, inter-interstitial cells and interstitial cells increase, notably matrix protein synthesis,and inhibition of substrate degradation caused by large accumulation of extracellular matrix leading to glomerular sclerosis and tubulointerstitial fibrosis. Accumulation of extracellular matrix in the renal interstitium is complicated.As per the normal mechanism, renal interstitial protein components (collagen typeⅠ,Ⅲ, fibronectin and tendon protein) are increased, but there is also deposition of normal tubular base membrane matrix proteins (collagen type IV and fibronectin, etc.) deposits.The Unilateral Ureteral Obstruction (UUO) model is of the classic renal interstitial fibrosis animal model, induced by ligation of unilateral ureteral obstruction of the kidney drainage system, leading to acute renal functional changes and chronic kidney damage to simulate the common clinical ureteral obstruction caused by renal interstitial injury, typified by simple and easily identifiable lesions. FFNT-5, a new pyridine ketone, was synthesized by our group, previous in vitro stupes having demonstrated that it significantly inhibited the proliferation of fibroblasts in a dose-dependent manner, and had no impact on normal cells. It has cleared acute toxicity testing preliminary experiments in GLP laboratories have proven their low toxicity. so that the influence of FFNT-5 on collagen deposition in renal tissue and anti-kidney fibrosis deserves further study.Objective:1.Observe the effect of different doses of FFNT-5 on the preventive treatment of renal interstitial fibrosis.Methods:25 SD rats were randomly divided into six groups:sham operation group, model group, FFNT-5 62.5mg/kg group,FFNT-5 125mg/kg group and AKF-PD 125mg/kg treatment groups(n=5). Apart from the sham operation group, other groups consisted of the unilateral ureteral ligation model of ureteral obstruction and only ureteral dissection was performed, not ligation and transection. Rats from drug treatment groups were administered 0.5% CMCNa as solvent,1 day before UUO modeling to 13 days after surgery.2 N-5 treatment groups were given N-5 solution using different dosages of 62.5mg/kg/d,125mg/ kg/d respectively;The 1AKF-PD treatment groups were given AKF-PD solution of 125mg/kg/d, dosage respectively,through intra-gastric delivery;The model group and sham operation group were given equal volumes of 0.5% CMCNa gavage.14 days after surgery, the animals were killed, HE and Masson staining performed on sections of the obstructed kidney, and detection of the renal expression of collagen typeⅠ,Ⅲachieved by immunohistochemistry.Results:14 days after UUO, the renal interstitial damage index and interstitial fibrosis was significantly higher in the model group than the sham group (P<0.0001); Excepting the FFNT-5 62.5 mg/kg treatment group, compared with the model group,the damage index and fibrosis were significantly decreased (P<0.0001), but still higher than the sham operation group (P<0.05); The effect in the FFNT-5 125mg/kg group was weaker than in the AKF-PD125mg/kg group (P<0.05); Increased doses in the FFNT-5 groups resulted in gradual reduction of renal interstitial damage index and interstitial fibrosis gradually decreased in a dose-dependent manner.In the model group, renal Collagen typeⅠ,Ⅲexpression was significantly increased; the treatment group compared with the model group showed expression of Collagen typeⅠ,Ⅲreduced by varying degrees.Conclusion:FFNT-5 can treat UUO renal interstitial fibrosis...