Based On The Theory Of "toxic Damage To Brain Collaterals", The Protective Effect And Mechanism Of Naoshuantong Capsule On Brain And Intestinal Barrier Function In Rats With Cerebral Small Vessel Disease Were Explore

Research purpose and significance:This study aims to explore the medication patterns of cerebral small vessel disease(CSVD)by leveraging data mining techniques and incorporating clinical research prescriptions for CSVD treatment.It also aims to predict the ingredients,targets,and pathways of detoxifying and dredging collaterals therapy through network pharmacology methods.Furthermore,it intends to validate the therapeutic effects and potential mechanisms of detoxify ing and dredging collaterals therapy in a rat model of CSVD through experimental verification.The goal is to provide new evidence for the ’toxic damage to collateral vessels’ theory and contribute insights into clinical medication and research.Research methods:Data Mining:The published literature on the treatment of CSVD by traditional Chinese medicine was included in this study.By using frequency statistics,cluster analysis,association rule analysis,complex network analysis and other methods,the TCM syndromes,medication rules,core prescriptions and treatment methods of CSVD were discussed.Network Pharmacology:Firstly,the effective components of the core Chinese medicine were screened out through the TCMSP and TCMID databases,and the target proteins of the effective components were further obtained.Secondly,the disease targets of cerebral smallvessel disease were searched,and the intersection target data set of drug component targets and disease targets was established.Finally,the protein interaction network,GO enrichment analysis and KEGG pathway analysis were performed.On this basis,through molecular docking technology,the binding ability of the intersection target to Nao Shuan Tong Capsule(representative Chinese patent medicine for the detoxification and promoting collaterals treatment method)was predicted,which provided insights and evidence for further mechanism research.Experimental research:The CSVD rat model was prepared using the BCCAO modeling method,and model evaluation was conducted after 4 weeks to select the successfully modeled rats.Pharmaceutical intervention was performed on the selected rats for 4 weeks.The efficacy and potential mechanism of action of Cerebral Thrombus-Dissolving Capsules in treating CSVD rats were investigated using behavioral tests such as water maze and open field,pathological tests including HE,LFB,and TUNEL,as well as molecular biology tests including Western blotting,qRT-PCR,and IHC.Using the method of 16S rRNA gene amplification sequencing of the gut microbiota,the analysis of species diversity,microbial composition and structure,and their correlation with memory,inflammation,and barrier indicators were conducted.The multidimensional mechanisms and connections were explored to understand the mechanisms and connections at multiple levels.The species diversity,flora composition and structure and their correlation with memory,inflammation and barrier indexes were analyzed to explore the multi-dimensional mechanism and connection by the method of 16S rRNA gene amplification and sequencing of intestinal flora.The research results:Data Mining:A total of 116 prescriptions were included in the study,most of which were randomized controlled trials(77%).The studies were mainly concentrated in 2015-2023(79%).The subtypes of the disease were CSVD with cognitive dysfunction(47%),lacunar cerebral infarction(20%),etc.TCM syndrome differentiation of kidney deficiency and blood stasis syndrome(29%),qi deficiency and blood stasis syndrome(17%),phlegm and blood stasis syndrome(13%).A total of 153 Chinese herbal medicines were used,and the cumulative frequency of Chinese medicines was 1161 times,with an average of 10 medicines per prescription.The top 30 traditional Chinese medicines such as Chuanxiong and Angelica were set as high-frequency traditional Chinese medicines.Most of these drugs are tonic drugs,blood circulation drugs.These drugs mainly enter the liver,heart,spleen and kidney meridians,with’warm,flat,slightly warm ’,’sweet,spicy.bitter’medicinal properties.K-means clustering classification is divided into 5 categories:qi-tonifying and blood-nourishing drugs,bloodactivating and stasis-resolving drugs,liver-calming and liver-nourishing drugs,kidneytonifying and essence-filling drugs,and resuscitation and refreshing drugs.Association rule analysis showed that based on Buyang Huanwu Decoction,Danggui-Chuanxiong,TaorenHonghua,Quanxie-Dilong.Yuanzhi-Shichangpu,Shanzhuyu-Shudihuang and other important compatibility drug pairs.Network Pharmacology:A network pharmacology analysis was conducted on the top 8 herbs in the detoxification treatment for promoting blood circulation and removing blood stasis.A total of 102 active ingredients.211 target proteins,2315 target points associated with cerebrovascular small vessel disease(CSVD),and 113 overlapping targets were identified.GO enrichment analysis found that the intersection targets were mainly on cell components such as membrane vesicles,and involved in biological processes such as hormone response through protein homodimerization activity.KEGG pathway analysis showed that WNT,NF-κB,adhesion connection and other pathways were closely related to CSVD.Molecular docking results revealed that the important target indicators(IL-6,MMP-9),pathway indicators(WNT3A),and blood-brain barrier indicators(Occludin,Caveolin-1)exhibited docking scores≥-5 with the brain-targeting components of Nao Shu Tong capsule.Experimental research:(1)After 4 weeks of modeling,compared with Sham group,the NORT cognitive index of BCCAO group decreased significantly(P<0.001),the expression of MBP decreased significantly(P<0.05),the expression of GFAP increased significantly(P<0.001),and the expression of Iba-1 increased significantly(P<0.01).(2)After 4 weeks of intervention,compared with the Sham group,the escape latency of the water maze in the BCCAO group was significantly prolonged,the residence time in the quadrant where the platform was located was significantly reduced(P<0.001),the total distance of the open field activity was significantly reduced,and the number of crossings was significantly reduced(P<0.05).Serum IL-6 and IL-1β were significantly increased(P<0.05),and TUNEL apoptosis was significantly increased(P<0.001).The arrangement of hippocampal pyramidal cells was relatively loose,and the myelin sheath connection in the central area of corpus callosum was loose and vacuolated.The expression of Caveolin-1 and MMP-9 was significantly increased(P<0.05),and the expression of Occludin was significantly decreased(P<0.05).The expressions of Wnt3 a,β-Catenin and p-GSK-3β were significantly decreased(P<0.05).(3)Compared with the BCCAO group,the NST group significantly shortened the escape latency of the model water maze,increased the residence time in the quadrant where the platform was located(P<0.001),and increased the total distance of open field activities and the central area activity time(P<0.05).The levels of serum IL-6 and IL-1β were significantly decreased(P<0.05),and the apoptosis of nerve cells was decreased(P<0.001).Restoring the tight arrangement of hippocampal pyramidal cells and the tight connection of myelin sheath;the expression of Caveolin-1 and MMP-9 was decreased(P<0.05),and the expression of Occludin was increased(P<0.05).The expression of Wnt3 a,β-Catenin and p-GSK-3β was significantly increased(P<0.05).(4)The results of intestinal flora showed that:1 Compared with the Sham group,the abundance of intestinal flora species in the BCCAO model was poor,the Shannon index of a diversity was significantly decreased(P<0.05),the proportion of F/B at the gate level was increased,the intestinal villi were shrunk and deformed,and the expression of Occludin and ZO-1 was significantly decreased(P<0.01).The Shannon index of α diversity increased significantly(P<0.01),and the Simpson index decreased(P<0.05).NST could change the distribution distance of β diversity PCoA(P<0.01),improve the F/B ratio at the gate level(P<0.01),increase the abundance of Lachnospirales(P<0.05),and increase the trend of Lachnospirales_NK4A136 at the genus level.It improved the shrinkage and deformation of intestinal villi and in creased the expression of Occludin and ZO-1(P<0.01).3 Cognitive behavioral indicators and intestinal barrier proteins were positively correlated with Lachnospirales(P<0.05),and inflammatory indicators were positively correlated with Enterobacterales(P<0.05).Research conclusion:Data Mining:The common clinical syndrome type of CSVD is blood stasis syndrome.The treatment is based on Buyang Huanwu Decoction.The traditional Chinese medicine with the effects of tonifying qi and nourishing blood,promoting blood circulation and removing blood stasis,calming liver and nourishing liver,tonifying kidney and filling essence,resolving phlegm and opening orifices is selected for treatment.The top 8 core Chinese medicines in data mining are:Chuanxiong.Danggui,Shichangpu.Huangqi,Dilong,Chishao,Danshen,Honghua,mainly for promoting blood circulation and removing blood stasis,resolving phlegm and dredging collaterals.To a certain extent,it reflects that ’phlegm and blood stasis’ is the most critical pathological factor of CSVD.The theory of ’phlegm and blood stasis’ has been transformed into ’phlegm toxin’ and ’blood stasis toxin’ for a long time,which is mutually confirmed with the theory of ’toxin damaging brain collaterals’.Network Pharmacology:The active ingredients of Tong Luo Jie Du Formula have multiple pharmacological effects,such as inhibiting inflammatory responses and anti-neuronal apoptosis.Potential targets may be involved in pathways such as inflammation,cell apoptosis.signal transduction,and angiogenesis.They may participate in signaling pathways such as Wnt.VEGF,and AMPK to improve blood-brain barrier and endothelial cell function,reduce inflammation,inhibit cell apoptosis,nourish nerves,and exert therapeutic effects.Naoshuantong capsule may have a good relationship with inflammatory pathway,blood-brain barrier pathway and Wnt3a signaling pathway.Experimental research:(1)Naoshuantong capsule can improve the spatial memory,depression and other cognitive behavioral functions of BCCAO rats.It may activate the Wnt3a/β-Catenin/GSK-3β signaling pathway,improve inflammatory indicators,restore BBB function and Caveolin-1 endocytosis transport process,reduce the expression of MMP-9,and then treat CSVD.(2)Naoshuantong capsule can increase the diversity and abundance of intestinal flora in rats,improve the composition of intestinal flora in rats,and protect the structure and function of intestinal barrier.