| Objective:1.To systematically search and summarize the clinical studies on the effect of traditional Chinese medicine(TCM)for the prevention and treatment of age-related macular degeneration(AMD)with the method of evidence map.To provide evidence and research direction for the prevention and treatment of AMD with TCM.2.To explore the pathogenesis of AMD and the effect mechanism of Guben Qingmu Decoction(GBQMD)by observing the characteristics of gut microbiota in patients with early AMD,the influence of gut microbiota on wet AMD mice model,and the intervention effect of GBQMD.To provide a new idea and a novel target for the prevention and treatment of AMD.Methods:1.TCM for the prevention and treatment of AMD:a clinical evidence mapAfter searching the publications from January 2000 to November 2021 about the prevention and treatment of AMD with TCM from databases CNKI,WanFang,VIP,PubMed,Web of Science,EMbase and the official website of Clinical Trials.gov,Chinese Clinical Trial Registry Center,we showed distribution features of the articles in form of text description,graphs and tables.2.The characteristics of gut microbiota in patients with early AMDThis was a case-control study.10 patients with early AMD and 14 healthy controls were selected.They were examined by fundus color photography,optical coherence tomography angiography(OCTA)and so on.We compared the microbial composition of the fecal samples between the two groups though 16SrRNA gene sequencing and bioinformatics analysis so that the characteristics of gut microbiota in patients with early AMD were analyzed accordingly.3.Establishment of a mice model for wet AMDMice were fed with foods containing HQ for 4 months,and CNV was induced by Laser Photocoagulation(LP).We observed the general state of the mice.The retinal morphology and choroidal neovascularization(CNV)area of the mice were observed by fundus color photography,retinal tissue hematoxylin-eosin(HE)staining,and choroidal flatmount.4.The intervention and mechanism exploration of GBQMD on wet AMD mice modelWe established a mice model of wet AMD with the method described above.Mice were divided randomly into three groups:prevention group,treatment group and distilled water group.The intervention for the prevention group started 28 days before LP.The intervention for the treatment group or the distilled water group started one day after LP.The normal group were fed with regular foods without HQ.All groups were observed 7 and 14 days after intervention.The effects and the mechanism of GBQMD on the wet AMD mice were observed by the detection of gut microbiota in fecal samples,intestinal permeability in serum samples,HE and choroidal flatmount in retinal tissues.Results:1.TCM for the prevention and treatment of AMD:a clinical evidence mapThe final 240 articles included 230 clinical studies,8 systematic reviews/metaanalysis,and 2 guidelines,expert consensus and clinical pathway.After studying the articles with a focus on the stage of AMD,study type,sample size,intervention,outcome indicators and the quality of the articles,we summarized and analyzed the intervention of TCM in these studies and found that the studies on the prevention and treatment of AMD with TCM were increasing,most of which were wet AMD.It also showed that the main study type was randomized controlled trials and the sample size was mostly less than 100 cases.Different types of intervention of TCM(such as Chinese medicine,Chinese patent medicine or acupuncture)showed the potential benefits of improving the vision and fundus lesions,and alleviating some symptoms.However,the overall quality of the studies was low.2.The characteristics of gut microbiota in patients with early AMD(1)The general state and ocular conditions:the differences in gender and age between the early AMD patients and the healthy control group were not statistically significant in the study(P>0.05).Compared with the healthy control group,the density of superficial and deep retinal capillary plexus in early AMD patients decreased significantly,and the differences between both groups were statistically significant(P<0.05).The central macular thickness of the early AMD patients decreased,but the differences between them were not statistically significant(P>0.05).(2)The detection of gut microbiota:compared with the healthy control group,the relative abundance of gut microbiota in the early AMD patients were different at the taxonomic level of phylum,class,order,family and genus.In early AMD group,the relative abundance of Oscillospirales and Ruminococcaceae increased,and the differences between both groups were statistically significant(P<0.05).To the control group,the relative abundance of Negativicutes,Lactobacillales,Streptococcaceae and Streptococcus increased,and the differences between both groups were statistically significant(P<0.05).Alpha diversity showed that the sampling was sufficient,and it had been widely covered in the literature and therefore the reliability of bioinformatics analysis was high.There was no significant difference between both groups in Chao1 index and Shannon index(P>0.05).Beta diversity PCoA showed significant differences in community structures between both groups(P<0.05).LEfSe showed there were biomarkers with significantly different influence in the two groups at the taxonomic level of order,family,genus and species.3.The mice model of wet AMDFundus color photography showed the retinal edema,the laser spots located between retinal vascular with an expanded range and partial fusion.The retina damaged by LP appeared porcelain white or light white.HE showed the retinal layers were disordered in structure with loose cell arrangement,and the outer nuclear layers were defective with some of which were accompanied with effusion in the photocoagulation area.It also showed that the retinal pigment epithelium and choroid were damaged,and the proliferating cells infiltrated from choroid to the inner layer of retina formed CNV.Choroidal flatmount showed the change of red fluorescence towards a clustered or a wheeled shape.In addition,after a whole body examination,compared with the normal group,the mice in the model group had the syndromes of anorexia,less water input,slow weight gain or reduced weight,hair whitening or partial shedding,listless spirit,pale in lip,claw nail and tail,reduced activity,blue and black stool,thin and soft,and some other manifestations similar to syndrome of spleen deficiency.4.The intervention effect and mechanism of GBQMD on wet AMD mice model(1)The results of gut microbiota detection(analyzed on the phylum and genus level):7 days after LP,compared with the normal group,the relative abundance of Firmicutes,Verrucomicrobiota,Proteobacteria,Cyanobacteria,Dubosiella,Akkermansia,Clostridia_UCG-014 and Turicibacter increased in the distilled water group,while the relative abundance of Bacteroidota,Patescibacteria,Campilobacterota,Desulfobacterota,Muribaculaceae,Lachnospiraceae_NK4A136_group,Helicobacter,Alloprevotella,Candidatus_Saccharimonas decreased in the same group.Meanwhile,the ratio of Firmicutes/Bacteroidota(F/B)increased and the relative abundance of Cyanobacteria increased,that of Campilobacterota decreased.The differences between the groups were statistically significant(P<0.05).Both the prevention group and the treatment group showed decreasing in the relative abundance of Firmicutes and increasing in the relative abundance of Bacteroidota,and the F/B value decreased.14 days after LP,compared with the normal group,the relative abundance of gut microbiota in the distilled water group increased or decreased basically in the same way as that in the distilled water group on the 7th day after LP,and the effect of GBQMD also improved the relative abundance ratio of beneficial bacteria in the prevention group and the treatment group.Alpha diversity showed the sampling was sufficient,and it had been widely covered in the literature and therefore the reliability of bioinformatics analysis was high.The differences in Chao1 index and Shannon index among the groups were not statistically significant(P>0.05)except that Chao1 index of the treatment group was significantly higher than that of the distilled water group on the 14th day after LP(P<0.01).Beta diversity PCoA showed that there were no significant differences in community structure among the groups(P>0.05).LEfSe showed there were biomarkers with significantly different influences at the classification level of kingdom,phylum,class,order,family,genus in the normal group,the 7d treatment group,the 14d prevention group and the 14d treatment group.(2)The results of intestinal permeability test:7 days after LP,compared with the normal group,the concentration of serum Evans blue(EB)increased in the distilled water group,and the difference was statistically significant(P<0.05).The concentration of serum EB in the prevention group and the treatment group was lower than that in the distilled water group,and there was significant difference between the prevention group and the distilled water group(P<0.05).14 days after LP,compared with the normal group,the concentration of serum EB increased in the distilled water group,and the difference was statistically significant(P<0.05).The concentration of serum EB in the prevention group and the treatment group was lower than that in the distilled water group,but the differences between the groups were not statistically significant(P>0.05).(3)The results of HE:7 days after LP,compared with the normal group,the inner and outer nuclear layers of the retina in each model group were disordered.The cells proliferated abnormally and the retinal pigment epithelial cell layer and Bruch membrane were damaged and interrupted.The choroidal capillaries were damaged and disordered.14 days after LP,the number of fibroblasts and collagen fibers increased in the photocoagulation area.7 days after LP,the retinopathy in the prevention group and the treatment group was reduced,and the central thickness of CNV was lower than that in the distilled water group.The differences between the prevention group and the distilled water group were statistically significant(P<0.01).14 days after LP,the retinopathy in the prevention group and the treatment group was reduced,and the central thickness of CNV was lower than that in the distilled water group,but the differences between the groups were not statistically significant(P>0.05).(4)The results of choroidal flatmount:7 days after LP,the cluster red fluorescence could be seen in each model group,which suggested the formation of CNV.The area of CNV in the prevention group and the treatment group was smaller than that in the distilled water group,and the differences between the prevention group and the distilled water group were statistically significant(P<0.05).14 days after LP,the range of each group was expanded,which were characterized by clusters and wheels in shape.The CNV area of the prevention group and the treatment group was smaller than that of the distilled water group,but the differences between the groups were not statistically significant(P>0.05).Conclusions:1.The sample sizes of clinical studies on the prevention and treatment of AMD with TCM were small.The outcome indicators were not unified and the effect on the stage of AMD was unclear.The overall quality of the early studies was lower.As there were very few studies done about the intervention of TCM on the early AMD,the advantages of TCM for the prevention and treatment of AMD remains unclear.Therefore,more standardized clinical research could be carried out with more trials on early AMD so that TCM could play a more important role in the prevention and treatment of such disease.2.Gut microbiome may be involved in the occurrence and development of AMD.3.GBQMD can serve as an intervention for the occurrence and development of wet AMD mice model,and its mechanism of action may be related to increasing the relative abundance of beneficial bacteria in the gut and remodeling the intestinal microecological balance. |
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