Studies On The Anti-aging Activity Of A Glycoprotein(GP3) Isolated From Fupenzi(rubus Chingii Hu.) And Its Regulation Of Klotho Gene Expression

The nearly mature fruits of Fupenzi?Rubus chingii Hu.?can benefit the kidney,reduce urination frequency and semen leakage.Four glycoprotein molecules are isolated and purified by the method of water extract-alcohol precipitation in the earlier research group.The experimental study in vitro shows that No.3 glycoprotein molecule?GP3?has the strongest antioxidant activity.Combining with the important role of anti-aging Klotho gene,we systematically study the anti-aging activity of GP3and its expression and regulation of Klotho gene in the mouse kidney tissue.The main research content is divided into the following two aspects:1.Study on anti-aging activity of GP3 in vitro.Testing the total reducing capacity and the hydroxyl radical scavenging rate of GP3,the content of SOD,GSH-Px,CAT,MDA after the action of TCMK-1 cells are assayed.By CCK-8 and?-galactosidase staining assays,evaluating the survival rate of TCMK-1,NRK-52E cells under H₂O₂ injury and the anti-aging activity of GP3.The result shows that GP3 can increase the survival rate of TCMK-1,NRK-52E cells under oxidative damage,and can significantly improve the physiological indexes of SOD,GSH-Px,CAT,MDA in TCMK-1 cells,and there is a certain dose-effect relationship.2.Study on anti-aging activity of GP3 in vivo and its expression and regulation of Klotho gene.The model of aging is established by injection of D-galactose,after the model is successfully established,the mice are dosed by mouth for 30 days and are sacrificed.The viscera coefficient and physiological indexes of serum,heart,liver,spleen,and kidney about mouse are assayed,and the liver and kidney tissues are pathologically sectioned.Meanwhile,total RNA and protein in mouse kidney are extracted and detected the amount of gene expression by RT-PCR and Western-blot.The result shows that GP3 can effectively improve the pathological state of kidney tissues and organs in aging mice and play a role in tonifying kidney,and the ability of kidney reserve can be repaired.GP3 can significantly improve the physiological indexes of SOD,GSH-Px,CAT,MDA in aging mice,it has obvious anti-aging activity.At the same time,it can significantly increase the expression of anti-aging Klotho gene in the mouse kidney tissue.In conclusion,GP3 has significant anti-oxidation and anti-aging activity,and plays an important role in regulating the anti-aging Klotho gene.We infer that the glycoprotein may be the material basis to delay aging of Fupenzi.The mechanism of anti-aging may be the following two aspects.On the one hand,eliminating harmful free radicals in the body to improve the physiological state of the organs of the body,so that the functions of various tissues and organs are repaired,especially the repair of the kidney tissue may be directly related to the anti-aging effects.On the other hand,regulating the expression level of anti-aging Klotho gene in vivo to achieve its anti-aging effect.The result provides scientific basis for Fupenzi as an important member in the ancient prescription of anti-aging and prolonging life,and also provides an experimental basis for the traditional Chinese medicine viewpoint that"Tonifying kidney is the basic approach to delay aging."...