Preparation And Evaluation Of AS-IV-NPs And Its Treatment Of Diabetic Nephropathy

Astragaloside Ⅳ is the most effective active ingredient in the Astragalus polysaccharide extract,derived from the dried rhizome of the plant Astragalus membranaceus,known as Super Astragalus Polysaccharide.Astragaloside Ⅳ has anti-oxidation,anti-inflammatory,immune regulation,anti-tumor and other pharmacological effects,and is widely used to treat many diseases.In the history of Chinese medicine development,astragalus is mostly used to treat diabetic nephropathy.Diabetic nephropathy,one of the main microvascular complications of diabetes,is the result of the composite effect of multi-hemodynamics,multi-cytokines and multi-system,which is caused by the disorder of glucose metabolism.In the world list of diseases that endanger human health,chronic diseases rank third,becoming a new disaster in the 21st century and directly leading to end-stage renal disease.In hundreds of years of traditional Chinese medicine clinical treatment,astragalus has a good effect on improving the clinical symptoms of diabetic patients.With the development of modern medicine,more and more studies have shown that Astragalus can significantly reduce the proteinuria level of diabetic nephropathy,and AS-Ⅳ is the main pharmacological ingredient.However,the pharmacological properties of AS-Ⅳ are insoluble in water,which restricts its clinical application.In this paper,AS-Ⅳ was prepared into a nano-suspension to explore its effect and mechanism on the glomerular mesangial cells as a diabetic nephropathy model.Methods used in the experiment 1.Establish the high performance liquid phase-evaporative light scattering chromatography(HPLC-ELSD)analysis method to determine the content of ASⅣ APIs and reference substances.2.Preparation of AS-Ⅳ-NPs by high-speed homogenization combined with high-pressure homogenization,the optimal formula was screened by the central design,and the AS-Ⅳ-NPs prepared by the optimized formula were characterized and evaluated.3.MTT method compared the survival inhibition rate of free AS-Ⅳ and AS-Ⅳ-NPs on HMC;flow cytometry explored the uptake of free AS-Ⅳ and AS-Ⅳ-NPs by normal cells and DN model cells;the internal ROS production of model cells and the elimination effect in vitro.4.Western Blot to detect the effects of free AS-Ⅳ and AS-Ⅳ-NPs on the inflammatory protein high mobility protein(HMGB1),receptor protein for advanced glycation end products(AGEs),nuclear factor(NF)in the DN cell model-κB)expression and the influence of its pathways.Through high performance liquid chromatography analysis,the concentration of AS-Ⅳ API is≥98%,meeting the requirements of this experiment;the aveages particle size of the experimentally prepared AS-Ⅳ-NPs is 96.51 ± 2.8 nm,the PDI is 0.14 ± 0.02,and the zeta potential is-24.5mV;the inhibitory and therapeutic effects of nano AS-Ⅳ on the DN cell model are more obvious,and the expression of HMGB1,AGEs,and NF-κB protein is reduced.It is concluded that HPLC-ELSD is very suitable for determining the content of AS-Ⅳ,and AS-ⅣNPs successfully improves the disadvantage of low bioavailability of free AS-Ⅳ and increases the practicality of clinical medication.At the same time,AS-Ⅳ-NPs can reduce the expression of HMGB1-AGEs-NF-κB pathway proteins to achieve the effect of treating DN cell models,reducing the development of DN to ESRD significantly,and reducing mortality.