The Study Of Shuangdan Oral Liquid And Its Main Effective Composition On The Treatment Of Diabetic Nephropathy

Backgroud and ObjectiveDiabetic nephropathy(DN) is characterized by hypertension, edema, proteinuria and renal insufficiency of kidney in the course of diabetes mellitus(DM). DN is the result of multiple factors, of which the role of free radicals is very important. The increasing free radicals can cause oxidative damage in DM. Free radicals may also participate in the change process of DM early glomerular hemodynamics, lipid peroxide can directly damage the basement membrane, leading to renal dysfunction. The main active ingredient of Shuangdan oral liquid(SDO) are danshensu and paeonol. The mechanism of Danshensu on DN may be to remove the oxygen free radical organization, inhibit platelet aggregation, reduce blood viscosity, significantly improve the microcirculation, inhibit endogenous cholesterol synthesis and inhibition of lipid peroxidation. At the same time paeonol can reduce serum total cholesterol(TC) in animal models and low density lipoprotein(LDL) level, improve blood rheology, decrease lipid, and inhibit lipid peroxidation. Thus, the pharmacological effects of principal components in SDO was consistent with the pathological mechanism of DN, then, whether the SDO of DN is valid, and what is the mechanism? Based on this question, we first design and study the role of SDO and its main effective composition on DN rats, and to explore the mechanism of SDO and its main effective composition for the treatment of early DN.MethodsThe experiment with SD male rats is to establish the cell model of mesangial cells in animal models of DN and high glucose environment to evaluate the effect and mechanism of SDO and its main effective composition for the treatment of DN. The experiment us ing MTT, flow cytometry, RT-PCR, Western Blotting methods to observe the following four aspects: 1. Therapeutic effect of SDO in DN rats; 2. The treatment effect of Danshensu combined with paeonol in DN rats; 3. The inhibitory effect of SDO and mechanism of mesangial cell proliferation; 4. Inhibitory effect of Danshensu combined with paeonol effect and mechanism of mesangial cell proliferation. Result and discussion1. The animal experiment of SDO on treatment of DN(1) Compared with model group, SDO can significantly improve renal function in DN rats, the 24 h urine volume, the levels of SCr and BUN of SDO treatment groups significantly decreased in DN rat(P<0.01).(2) Compared with model group, SDO can significantly improve the disorder of lipid metabolism in DN rats, the contents of T-CHO, TG and LDL-C in DN rats significantly decreased and the level of HDL-C increased(P<0.05).(3) Compared with model group, SDO can significantly improve the antioxidative ability of DN rats, the levels of T-AOC and T- SDO in the treatment group significantly increased and the levels of NO and MDA decreased(P<0.05).(4) SDO significantly improved DN renal pathological injury in rats, under the light microscope, the degree of glomerular hypertrophy of renal tubular epithelial vacuolar degeneration degree are significantly reduced.2. The animal experiment of danshensu combined with paeonol on treatment of DN(1) DSS + Pae had the best effect on significantly improving renal function in DN rats reducing 24 h urine volume, the levels of SCr and BUN(P < 0.01) among the other treatment groups.(2) DSS + Pae had the best effect on improving blood lipid metabolic disorder, decreasing the contents of T-CHO, TG and LDL-C, increasing the level of HDL-C(P < 0.05).(3) DSS + Pae had the best effect on improving the antioxida nt capacity, increasing T-AOC, T-SOD activity and reducing the content of no and MDA(P < 0.01)(4) Each treatment group can improve renal pathological injury in DN rats. Under the light microscope, the degree of glomerular hypertrophy of renal tubular epithelial vacuolar degeneration were significantly reduced, among which the DSS + Pae treatment group had the best effect.(5) The glomeruli and renal tubular epithelial hypertrophy and vacuolar degeneration degree are significantly reduced, the DSS + Pae treatment group had the best effect.3. The cell experimentalof SDO on treatment of DN(1) SDO significantly decreased glomerular mesangial cells in experimental MTT activity.(2) SDO significantly increased mesangial cell percentage of G1 phase.(3) SDO significantly reduced glomerular mesangial cells by flow cytometry in the proportion of B3.(4) Under the transmission electron microscope observed cells appeared vacuolar and mesangial cel s in mitochondria obvious nucleolus pyknosis.4. The cell experiment of danshensu combined with paeonol on treatment of DN(1) DSS + Pae treatment group had the best effect on reducing glomerular mesangial cel s in experimental MTT activity.(2) DSS + Pae treatment group had the best effect on increasing mesangial cell percentage of G1 phase.(3) DSS + Pae treatment group had the best effect on reducing glomerular mesangial cel s by flow cytometry in the proportion of B3.(4) The each treatment group can significantly promote the apoptosis of glomerular mesangial cells, and model group compared to DSS + Pae in each treatment group can significantly up-regulated cytochrome c, Bax, PARP-1 and caspase-3 m RNA expression levels(P < 0.01) and protein expression(P < 0.05), and significantly down-regulated the m RNA of Bcl-2 expression level(P < 0.05) and protein expression(P < 0.05); and significantly reduced the mitochondrial membrane potential(P < 0.05), of which DSS + Pae group treatment had the best effect.ConclusionsThe study confirmed that the shuangdan oral liquid and its main effective composition had the therapeutic effect on treatment of DN in vitro and in vivo. The mechanism may be related to lipid metabolism, improve the antioxidative ability and mitochondria mediated apoptosis pathway. The results of the study provided strong support for the clinical research of shuangdan oral liquid and its main effective component on the treatment of DN, and it also provided effective theoretical basis for the development and application of molecular medicine in shuangdan oral liquid med icine for basic research and development in the future.