The Improvement Effect And Mechanism Of Total Saponins From The Fruits Of Panax Ginseng And Ginsenoside Rg3 On AMD Retinopathy

According to ancient records,Panax ginseng C.A.Meyer has the function of improving vision and promoting intelligence.Modern pharmacological research has shown that the active ingredient ginsenosides in ginseng can be used to prevent or improve eye diseases that threaten vision,such as glaucoma,diabetic retinopathy,and cataracts.However,the non-medicinal part of ginseng,the ginseng fruit,is rarely used.Ginseng fruit contains richer saponin compounds than ginseng and has protective effects on the cardiovascular system,endocrine system,immune system,and anti-aging properties,but there are no reported studies on the relationship between ginseng fruit saponins and age-related macular degeneration（AMD）.Age-related macular degeneration（AMD）,a disease that mainly occurs in people over 50 years old,increases in incidence with age.Sodium iodate（NaIO₃）,an inorganic oxidant,is often used as a modeling agent for AMD.Therefore,this study used ginseng fruit as the research object to explore the protective effect and molecular mechanism of ginsenosides against retinal degeneration induced by NaIO₃ to investigate its potential for treating AMD.Combining techniques such as H&E staining,optical coherence tomography（OCT）,fundus fluorescein angiography（FFA）,immunofluorescence,and Western Blot,the protective effect and mechanism of ginseng fruit total saponins on NaIO₃-induced AMD in mice were studied.To further explore its active ingredients,the protective effect of ginsenoside Rg3 on retinal injury was investigated using a model of NaIO₃-induced retinal pigment epithelial cell（RPE）damage,and its potential molecular mechanism of action was examined.This study will provide scientific evidence for the treatment of AMD with ginseng active ingredients.The main research work is divided into the following three aspects:1.The protective effect and mechanism of total saponins from the fruits of Panax ginseng （SFPG）on sodium iodate-induced age-related macular degeneration in mice.After adapting to the environment for 7 days,C57BL/6J mice were intravenously injected with NaIO₃ at a dose of 20 mg/kg to establish an age-related macular degeneration（AMD）model.After 7 days,the morphology of the mice’s retina was observed through fundus photography to confirm the successful construction of the AMD model.The protective effect of total ginsenosides from Panax notoginseng was observed by pre-administering it to the mice.The experimental results showed that the expression of tight junction protein（ZO-1）and（Occludin）in the model group decreased as analyzed by Western blot and immunofluorescence staining.H&E staining and optical coherence tomography（OCT）were used to detect changes in the morphology of the mice’s retina,which showed that sodium iodate resulted in retinal thinning,significant disorder of the outer nuclear layer,and large amounts of yellow-white deposits in the retina of the model group.Fluorescein angiography（FFA）showed obvious shadowing and dark spots in the fundus of the model group mice.However,pre-treatment with SFPG at doses of 150 mg/kg and 300 mg/kg for 30 days significantly relieved the above symptoms.The expression of tight junction proteins ZO-1 and occludin in the mice’s retina increased,and the relative thickness of the retina increased.The accumulation of papillary structures in the mice’s fundus decreased,and the disorder of the inner and outer nuclear layers of the retina decreased.Western blot analysis preliminarily suggested that SFPG alleviate AMD-induced retinal damage by increasing retinal tight junction proteins.2.The protective effect and molecular mechanism of ginsenoside Rg3 on sodium iodate-induced damage of RPE.In order to further clarify which monomeric component of total saponins from the fruits of Panax ginseng plays the main therapeutic role,this study established a NaIO₃-induced retinal pigment epithelial cell（RPE）injury model and screened the optimal dose of NaIO₃ through MTT assay.Finally,the concentration of NaIO₃ was determined to be 5 m M as the optimal modeling dose,and the major saponin including Re,Rd,Rb1,Rc,S-Rg3,and R-Rg3 in the fruits of Panax ginseng were screened for activity.The study found that S-Rg3 exhibited excellent pharmacological activity.Therefore,S-Rg3 was selected as the research object for subsequent experiments.MTT results showed that S-Rg3 at doses of 1μM,2μM,and 4μM could restore the viability of RPE cells in a dose-dependent manner.S-Rg3 could partially inhibit the excessive production of ROS induced by NaIO₃.Western Blot results further elucidated that S-Rg3 inhibited the apoptosis of NaIO₃-induced RPE cells by reducing Cytochrome c,Cl-Caspase 9,Cl-Caspase 3,and increasing the expression of Bcl-2 protein.S-Rg3 could inhibit JNK over-phosphorylation to regulate autophagy and apoptosis.Cell apoptosis analysis by flow cytometry and JC-1 staining techniques showed that S-Rg3 pretreatment could effectively improve cell apoptosis.3.Protective effect and mechanism of ginsenoside Rg3 on sodium iodate-induced age-related macular degeneration in mice.Through establishing a NaIO₃-induced mouse model of age-related macular degeneration（AMD）,this study aimed to further investigate the in vivo molecular mechanisms and protective effects of ginsenoside Rg3 on NaIO₃-induced retinal injury.The results showed that H&E staining and optical coherence tomography detected significant thinning of the retinas in NaIO₃-treated mice,with obvious disorganization of the inner and outer nuclear layers.Fundus photography revealed extensive yellow-white deposits in the retinas of model mice.Fluorescein angiography showed poor masking of the mouse retina and prominent dark spots.Treatment with S-Rg3 at doses of 20 and 40 mg/kg via oral administration for 30 consecutive days significantly alleviated the above symptoms,with an increase in relative retinal thickness and a decrease in the degree of disorganization of the inner and outer nuclear layers.Its improvement effect on age-related macular degeneration was similar to that of total ginsenosides from ginseng fruit.To further investigate the mechanism of action of S-Rg3 in treating AMD retinal damage,Western blot and immunofluorescence staining were performed,revealing that S-Rg3 could inhibit the over-phosphorylation of JNK and down-regulate the level of P62 protein,promoting autophagy circulation.In the study of the apoptotic signaling pathway,it was found that ginsenoside Rg3 could inhibit the apoptosis of retinal pigment epithelial cells by regulating the expression of apoptotic-related proteins,thereby exerting a protective effect.In summary,this study verified through in vivo and in vitro experiments that total ginsenosides from ginseng fruit and S-Rg3 both have a certain improvement effect on NaIO₃-induced AMD.Its mechanism of action may be through the inhibition of JNK over-phosphorylation,the reduction of autophagy protein accumulation,and the inhibition of apoptosis factor expression to exert a protective effect.This study provides some scientific basis for the treatment of AMD disease with total ginsenosides from ginseng fruit and ginsenoside Rg3,as well as further development and utilization.