Classification Of MAFLD Subtypes Based On TCM Clinical Phenotype And Oral-intestinal Flora Characteristics Of Obese-MAFLD

ObjectiveThe etiology,pathogenesis,syndrome typing,subgroup analysis and typing of metabolic-associated fatty liver disease（MAFLD）were discussed theoretically.To provide new insights into the heterogeneity of MAFLD,the subtypes of MAFLD were classified using electronic medical records of Traditional Chinese medicine as data sources.Combined with subtype classification of obesity MAFLD disease and sequencing analysis of oral（tongue coating）-intestinal flora,the biological differences of metabolic disorders of complex"disease syndrome cluster" bacteria were identified.Methods1.The ancient and modern definitions,etiology and pathogenesis,disease location,syndrome classification and syndrome differentiation and treatment of MAFLD were summarized based on ancient and modern medical literatures.The current status of the disease-syndrome classification of MAFLD at home and abroad was analyzed based on factors such as body constitution,body mass index（BMI）,microbial metabolism,comorbidiosis,imaging indicators,pathological types and omics information.2.The electronic TCM medical records in accordance with MAFLD diagnosis from 2016 to 2020 were included for entity labeling and standardized treatment of each clinical phenotype.Conduct disease-gene set and drug-target matching.A heterogeneous medical record network（HEMnet）based on MAFLD population was constructed with gender,age,disease diagnosis,symptoms and western medicine prescription as nodes and the co-occurrence times of each phenotype as edge,and clustering was divided by k-means clustering algorithm.Relative risk was used to assess the specificity of each phenotype.Statistical methods were used to compare the differences of laboratory indicators among subtypes.The traditional Chinese medicine prescriptions involved in subtypes were analyzed by using Liquorice platform.Based on functional annotation data of molecular pathway and Gene Ontology,the functional analysis of corresponding molecular network modules of each disease group was carried out.4.Subtypes of obese MAFLD patients were classified by the above classification methods.Collect tongue coating and fecal biological samples of patients according to standard operating procedures and extract DNA;Bioinformatics analysis of 16S rDNA full-length assembly and sequencing was performed.ResultsChapter 1The pathogenesis of MAFLD mostly involves "dampness","phlegm","blood stasis" and other tangible evil,which can be classified into"hypochondriac","accumulation","ruffia","liver disease","fat qi" and other categories.Its clinical types are mainly treated by the syndromes of liver stagnation and spleen deficiency,internal obstruction of phlegm and dampness,interaction of phlegm and blood stasis,obstruction of dampness and heat and deficiency of liver and kidney Yin.Chapter 212 626 patients were enrolled,with a mean age of（55.02±14.21）years old.The top 3 TCM combined diagnosis were diabetes,vertigo and chest paralysis.The top 3 TCM syndromes were qi stagnation and blood stasis syndrome,damp-heat accumulation syndrome and phlegm and blood stasis syndrome.There are 1 117 kinds of concomitant diseases in western medicine,and the top 3 are hypertension,hyperlipidemia and type 2 diabetes.A total of 7 954 patients with MAFLD received oral Chinese medicine compound,involving 455 kinds of Chinese medicine,the top 5 Chinese medicine were Poria cocos,glycyrrhiza,Atractylodes atractylodes,Astragalus membranaceus and tangerine peel.A total of 8 469 patients were treated with 312 kinds of Chinese patent medicines,the frequency of which was in order of removing blood stasis,strengthening and clearing heat.A total of 12 000 patients with MAFLD were treated with western drugs that were metabolized in vivo,involving 506 kinds of drugs.The top three drugs in frequency were hormones and endocrine affecting drugs,drugs for cardiovascular system,and drugs for digestive system.The results of cluster analysis showed that when the cluster number was 5,the effect was the best,and the differentiation between subtypes was obvious.There were 8 351 cases of CO subtype with complicated digestive system diseases,and the subtype population accounted for the largest proportion.Chinese medicine prescription drugs are mainly to buzhong Yiqi decoction,Erchen decoction and so on as the core of the main prescription.The biological enrichment involved is mainly associated with inflammation and is heavily enriched in tumor-related disease signaling pathways.There were 1 694 cases of C1 subtype with mental disorders and gynecological diseases,which were mainly manifested as abnormal mood,vaginal bleeding,irregular menstruation and menstrual tendons.The matching degree of core prescription was the highest in Liujunzi Decoction and Bazhen Decoction.The biological enrichment involved is mainly related to the dysfunction of NMDA receptor,the abnormality of GABA system and the disturbance of hormone regulation in vivo,and is involved in the neuroactive ligand-receptor interaction,y-aminobutyric acid neurotransmission and estrogen signaling pathway.There were 384 patients in C2 subtype with chronic liver disease and decompensated complications.The subtype patients were older and had the longest hospital stay.In biochemical tests,they showed the characteristics of’three series’ reduction,abnormal liver function and decreased protein level,as well as low lipid level.The core prescriptions were xiaoyao SAN and Chaihu Shugan SAN.The biofunctional enrichment involved is mainly related to bile acid transport,nitric oxide metabolism,T cell differentiation regulation and microbial symbiotic growth.A total of 1 862 patients with type 2 diabetes mellitus and its complications were subtype C3,presenting obvious manifestations of so-called typical clinical polydipsia,polyuria,and more food and weight loss,acral numbness and blurred vision.The levels of fasting blood glucose,glycosylated hemoglobin,insulin resistance index and blood lipid in the population were significantly higher than those in other subtypes.The core prescriptions mainly include Angelica Liuhuang Decoction,liquid enhancement soup and Qiongyu Ointment.The biological enrichment involved is mainly related to the regulation of glucose metabolism,lipid metabolism,protein-containing complex remodeling and T cell-mediated immune regulation.There were 335 cases of C4 subtype with rheumatic immune system diseases,which had the least number of patients and showed higher PLT count,WBC count and CRP level.The core prescriptions were mainly Duhuizi Decoction and Angelica Shaoyao Powder with high matching degree.The bioenrichment involved is mainly related to bone development such as the positive and negative regulation of ossification,bone remodeling and resorption,and the regulation of a large number of JAK/STAT protein tyrosine phosphorylation.Chapter 3Obesity is the main feature of MAFLD in young patients,associated with glucose and lipid metabolism disorders.The three subtypes of obese MAFLD were liver inflammation,abnormal glucose metabolism and abnormal blood pressure.The OM-CO subtype of obese MAFLD patients were mainly complicated with NASH,gout and lumbar spine disease,and mainly focused on loose stools,liver discomfort,abdominal distension,poor appetite,hypochondriac pain and abdominal pain.ALT and CAP value were the highest.The oral microbial diversity of OM-CO subtype was relatively high.The abundance of Subdoligranulum and Eubacterium were significantly lower.Oral Alloprevotella is associated with FPG.OM-C1 subtype mainly characterized by abnormal glucose metabolism.Patients with subtype OM-C1 had insulin resistance,metabolic syndrome,polycystic ovary syndrome and irregular menstruation.Fatigue,weight gain,head and body trapped heavy,irregular menstruation,lazy and other symptoms of spleen deficiency phlegm dampness.In the laboratory,they had higher levels of insulin resistance,uric acid,BMI,body fat percentage,and visceral fat area.The F/B ratio increased,Alloprevotella and Haemophilus were the dominant oral microorganisms of this subtype.At the same time,the abundance of intestinal Eubacterium in OM-C1 subtype patients was significantly negatively correlated with oral Alloprevotella,and the abundance of intestinal Eubacterium was negatively correlated with BMI.Meanwhile,the abundance of Haemophilus of OM-CO subtype was negatively correlated with CAP value.The abundance of Oral Haemophilus in OM-C1 subtype patients was positively correlated with ALB level,but negatively correlated with BMI,body fat percentage and visceral fat area.The OM-C2 subtype population with abnormal blood pressure as the main feature was characterized by hypertension,cervical spondylosis and gastritis,with dizziness,headache,head weight,poor sleep,nausea and other symptoms of hyperactivity of liver Yang.Patients with subtype OM-C2 were admitted to hospital at a relatively high age and presented hyperlipidemia.Due to the small number of oral and intestinal samples collected from obese MAFLD OM-C2 subtype in this study,they were not analyzed.Conclusion1.MAFLD patients have diverse symptoms and complex comorbidities.The disease classification method based on clinical-molecular network feature representation can divide the MAFLD population into spleen and stomach diseases（combined with digestive system diseases）,emotional diseases（mental disorders,gynecological diseases）,hepatobiliary syndrome（chronic liver disease and decompensated disease）,diabetes mellitus and its syndrome（diabetes and its complications）,joint meridian and tendon disease（immune joint system disease）5 categories,and in the "disease-disease-There is consistency in the "symptom-treat-target".2.The obese MAFLD population can be divided into three subtypes:inflammatory state,abnormal glucose metabolism and abnormal blood pressure,and the characteristics of oral-intestinal flora are distinct.3.The abundance of Eubacterium in the intestinal tract of the obese MAFLD subtype with abnormal glucose metabolism was significantly negatively correlated with the oral Alloprevotella genus,while the inflammatory state of the obese MAFLD subtype was opposite.4.Compared with the obese MAFLD subtype in the inflammatory state,the microbiota function of the obese MAFLD subtype with abnormal glucose metabolism is mainly manifested in the abnormal expression of the pentose phosphate pathway（PPP）and its related metabolic pathways.