Based On The Intestinal Flora To Explore The Clinical And Mechanism Research Of Jiangtang Tiaozhi Recipe In Regulating Glucose, Lipid And Fat Metabolism Disorders

1 Background and PurposeAccording to the latest epidemiological data,China has 114.4 million diabetic patients in 2017 and this number still ranks first in the world.In addition,as the way of life modernized and the eating habits changed,the number of diabetes patients with obesity and hyperlipidemia is increasing.Type 2 diabetes(T2D),obesity,and hyperlipidemia are all the risk factors for cardiovascular disease.T2D patients with obesity and hyperlipidemia are facing difficult to control blood sugar levels,having higher risk of cardiovascular event and higher mortality.The purpose of the comprehensive treatment of T2D is to balance the hypoglycemic effect,the risk of inducing cardiovascular events,drug tolerance and adverse reactions.To date,a comprehensive and effective treatment for patients with multiple metabolic disorders is uncommon.Current studies have shown that T2D frequently cause from insulin resistance,and T2D and obesity are manifested as chronic low-level inflammation.As a highly complex bacterial community in gastrointestinal tract,gut microbiota has a close relationship with the metabolic diseases.Therefore,this study aims at(1)Evaluating the efficacy and safety of using Jiangtangtiaozhi(JTTZ),a Chinese herbal formula,to treat T2D patients with obesity and lipid disorders;(2)Investigating the comprehensive intervention mechanism of JTTZ formula from the perspectives of insulin resistance,body inflammation,and gut microbiota.2 Methods2.1 The Methods of Randomized Controlled Clinical Trial450 subjects with T2D,obesity,hyperlipidemia,and gastrointestinal heat syndrome were divided into treatment group and control group with a proportion of 1 to 1 by an Interactive Web Response System.The treatment group received JTTZ formula(JTTZ group),while the control group received metformin(MET group).The course of treatment included a 4-week screening period and a 12-week treatment period.Quality control of the traditional Chinese medicine(TCM)compound was conducted using high-performance liquid chromatography.The main efficacy indicators include changes in glycated hemoglobin(HbA1c),triglycerides(TG),body weight,and waist circumference compared with baseline,and the rates of reaching the target of HbAlc and TG.The secondary efficacy indicators were FPG,2h-PG,TC,LDL-C,BMI,hip circumference and TCM syndrome efficacy changed from baseline.The safety indicators were vital signs,routines of blood,urine,and stool samples,functions of liver and kidney,and electrocardiogram.Meanwhile,adverse events and hypoglycemic events were recorded carefully during the treatment.2.2 The Methods of Mechanisms of Insulin Resistance and InflammationBy measuring serum fasting insulin(FINS),adiponectin,leptin,insulin resistance index(HOMA-IR),and ? cell function index(HOMA-?)were calculated.By using the ELISA method,the serum lipopolysaccharide binding protein(LBP),al-acid glycoprotein(al-AGP),serum amyloid A protein(SAA),tumor necrosis factor ?(TNF-?),and interleukin 6(IL-6)expression levels were detected.2.3 The Methods of Gut Microbiota MechanismStool samples were collected with Stool Collection Tubes with DNA Stabilizer,and DNA was extracted using InviMag_Stool_DNA_Kit.The 16S rRNA gene V3-V4 region fragments with index at both ends were obtained by two rounds of PCR,and the sequencing library was constructed.PCR purification was performed by magnetic beads adsorption method.Using the Illumina Hiseq 2500 high-throughput sequencing platform,the qualified library was sequenced by a double-end method.Bioinformatics analyses,including diversity analysis,species structure analysis,variance analysis,and correlation analysis were also performed.3 Results3.1 The Results of Clinical TrialA total of 12252 subjects underwent initial screening.According to the inclusion and exclusion criteria,450 subjects were enrolled.In the JTTZ group,216 subjects completed the 12-week clinical study and 215 were included in the PPS analysis.In the MET group,212 subjects completed the study and 199 were included in the PPS analysis.The values of the baseline characteristics showed no differences between the two groups.3.1.1 Blood Glucose EfficacyHbA1c decreased by 0.75±1.32%and 0.71 ±1.2%in the JTTZ and MET groups,respectively,after the 12-week treatment.Using HbAlc ?7.0%,?6.5%,and ?6.0%as the target respectively,the achievement rates were 45.1%,26.0%,and 11.2%in the JTTZ group,and 42.7%,18.1%,and 7.5%in the MET group.The FPG levels(mmol/L)were reduced by 1.4±2.4 in the JTTZ group and by 1.33±2.33 in the MET group.The 2h-PG levels(mmol/L)were reduced by 2.42±4.53 in the JTTZ group and by 2.18±4.48 in the MET group.Compared with the baseline,both groups had the significant differences(P<0.001).3.1.2 Blood Lipid EfficacyAfter the 12-week treatment,the TG value in 27.4%of the subjects in JTTZ group and 27.1%of the subjects in the MET group achieved the target,when using TG ?1.7 mmol/L as the target achievement.The TG levels(mmol/L)were significantly reduced by 0.64±2.37(P<0.001)in the JTTZ group and by 0.37±2.18(P=0.018)in the MET group,compared to baseline.The TC levels(mmol/L)were reduced by 0.39±0.99 in the JTTZ group and by 0.36±1.16 in the MET group.The LDL-C levels(mmol/L)were reduced by 0.29±0.79 in the JTTZ group and by 0.31±0.71 in the MET group.Compared with the baseline,both groups had the significant differences(P<0.05).3.1.3 Obesity EfficacyAfter the 12-week treatment,the weight levels(kg)were reduced by 2.47±2.71 in the JTTZ group and by 2.03±2.36in the MET group.The BMI levels(kg/m2)were reduced by 0.9±0.99 in the JTTZ group and by 0.74±0.86 in the MET group.The waist circumference levels(cm)were reduced by 2.52±4.29 in the JTTZ group and by 2.49±3.86 in the MET group.The hip circumference levels(cm)were reduced by 3.04±3.86 in the JTTZ group and by 2.65±3.51 in the MET group.Compared with the baseline,both groups had the significant differences(P<0.001).3.1.4 TCM Syndromes EfficacyIn response to the syndrome of gastrointestinal heat syndrome,the JTTZ formula significantly improved participants' total symptom scores and single symptom score of abdominal fullness,thoracoabdominal distension,congestion,dry mouth,bitter mouth,Halitosis,thirst and drinking cold water,increased eating with rapid hungering,red tongue and yellow tongue coating,and power and rapid pulse after the 12-week treatment(P<0.05).The level decreased significantly more in the JTTZ group compared to the MET group(P<0.001).3.1.5 Safety EvaluationOne case of non-severe hypoglycemia was reported in the MET group,and no severe adverse events were reported during the trial.3.2 The Results of Mechanisms of Insulin Resistance and InflammationAfter the 12-week treatment,the HOMA-IR levels were significantly reduced by 0.19±0.91 in the JTTZ group(P<0.01)and by 0.16±0.82(P<0.05)in the MET group.The HOMA-? levels were significantly increased by 0.26±0.82 in the JTTZ group(P<0.001)and by 0.27±0.87(P<0.001)in the MET group,compared to baseline.The leptin levels(pg/mL)in the JTTZ group and MET groups were increased 55.04±216.78 and 16.41±128.5,respectively.Compared with the baseline,only the JTTZ group had a significant difference(P=0.005).There was no statistical difference between the two groups of the changes of adiponectin.The LBP levels(ng/mL)in the JTTZ and MET groups were decreased 1.98±4.36 and 1.85±4.81,respectively.The ?1-AGP levels(ng/mL)in the JTTZ and MET groups were decreased 3.25±10.87 and 3.73±9.96,respectively.The SAA levels(?g/mL)were decreased 0.47±1.04 in the JTTZ group and 0.47±1.22 in MET group,after the 12-week treatment.Compared with the baseline,both groups had the significant differences(P<0.01).There was no significant difference in the changes of TNF-? and IL-6 between the two groups.3.3 The Results of Gut Microbiota MechanismThe intervention of JTTZ formula for 12 weeks were able to reduce the species numbers,richness and diversity of gut microbiota,and improve the gut microbiota structure.However,MET group did not affect the diversity and structure of the gut microbiota.Boths groups could significantly reduce the abundance of Romboutsia.The MET group could also significantly reduce the abundance of Fusicatenibacter.The JTTZ group could also significantly reduce the abundance of Ruminococcus,Clostridium_sensu_stricto,and unclassified_Ruminococcaceae;and increase the abundance of Anaerostipes and unclassified_Lachnospiraceae.Anaerostipes and unclassified_Lachnospiraceae were able to be used as the specific biomarkers of the JTTZ formula.In addition,the changed abundance of unclassified_Lachnospiraceae was correlated with FPG and 2h-PG before and after the intervention of the JTTZ formula.4 ConclusionsThe JTTZ formula led to safe and significant improvements in the blood glucose,blood lipids,and weight levels;relieved gastrointestinal heat syndrome for T2D patients with concomitant obesity and hyperlipidemia.Improing insulin resistance,enhancing ? cell function;reducing chronic inflammation levels and changing gut microbiota structure were its possible intervention mechanisms.The JTTZ formula has shown positive results as a comprehensive treatment for T2D and could potentially be developed as an alternative medicine for patients with type-2 diabetes,obesity and hyperlipidemia,particularly those who cannot tolerate metformin or other hypoglycemic and hypolipidemic drugs.This study provided an evidence-based evidence of large national population and possible biological mechanisms evidence for comprehensive treatment of T2D with obesity and hyperlipidemia using Chinese herbal medicine.