The Study Of Buzhong-Yiqi Decoction On Insomnia Patients With Spleen Deficiency Syndrome:A Perspective Of Gut Microbiota

ObjectiveThe overall objective of this study was to observed the personality,gut microbiota of insomnia patients with spleen deficiency syndrome.Secondly,we evaluated the association among the human subjects with sleep quality,gastrointestinal symptoms and intestinal flora.Furthermore,we observed the efficacy and safety of Buzhong-Yiqi Decoction of insomnia patients with spleen deficiency,as well as its effect and possible mechanism on gut microbiota.MethodsThis is a double-blind,randomized controlled trial,which were conducted in Guangdong Provincial Hospital of Traditional Chinese Medicine.Subjects who were diagnosed insomnia with spleen deficiency syndrome were divided into two groups at random.The control group was treated with placebo（Buzhong-Yiqi Decoction Simulant）and the experimental group was treated with Buzhong-Yiqi Decoction Granules.And they received a one-week start-up period by taking Buzhong-Yiqi Decoction Simulant,followed by a 4-week drug intervention（Buzhong-Yiqi Decoction Granules or Simulant）,and finally followed up for one month.Before treatment we evaluated sleep quality,gastrointestinal symptoms,gut microbiota and the personality of two groups.Simultaneously,before and after treatment,the efficacy and safety of Buzhong-Yiqi Decoction in treating insomnia patients with spleen deficiency were evaluated.The association among sleep quality,gastrointestinal symptoms and gut microbiota were examined.ResultsFrom January 2018 to March 2019,we prospectively and consectively enrolled 62 insomnia patients with spleen deficiency syndrome from The Second Affiliated Hospital of Guangzhou University of Chinese Medicine（Guangdong Provincial Hospital of Traditional Chinese Medicine）.55 cases met the inclusion criteria,which were randomly divided into two groups,including 27 in the test group and 28 in the control group.The following results are described using data that fits the normal distribution,and data that does not fit the normal distribution are described using the median（Q1,Q3）.1.Baseline results of insomniac with spleen-deficiency1.1 Demographic characteristics and personalityDemographic characteristics:the average age of insomniacs with spleen deficiency was 41 years old,and the ratio of male to female was 1:2.9.Personality:the results of MMPI hysteria（Hy）scores was 65.04±12.45,higher than that of Chinese norm（T=60）.1.2 Sleep Quality1.2 Subjective sleep qualityThe Pittsburgh Sleep Quality Index（PSQI）was 13.56±2.22,the Insomnia Severity Index（ISI）was 20.35±0.56,and the degree of insomnia was moderate.1.3 Gastroenteric symptomThe score of gastrointestinal symptom rating scale（GSRS）was 5.21±0.47.The main symptoms of insomnia patients with spleen-deficiency included abdominal pain,abdominal distention,increased defecation,etc.1.4 Gut microbiota1.4.1 Alpha Diversity:Simpson index（richness and evenness）was 0.825.1.4.2 Species composition:1.4.2.1 Distribution of top 10 flora in abundance arrangement（1）Phylum-level Differences:The intestinal flora of patients with spleen deficiency type insomnia mainly consists of the following 6 phylums（the abundances are arranged from high to low）:Firmicutes,Bacteroidetes,Actinobacteria,Proteobacteria,Fusobacteria,Verrucomicrobia.（2）Class-level Differences:Clostridia,Negativicutes,Bacilli,Erysipelotrichia;Bacteroidia;Actinobacteria and Coriobacteriia;Gammaproteobacteria;Fusobacteriia;Verrucomicrobiae.Among them,the most dominant bacteria are Bacteroides and Clostridia.（3）Genus-level Differences:Prevotella₉,Megamonas,Faecalibacterium,Bifidobacterium,Collinsella,Bacteroides,Fusobacterium,Veillonella,Sutterella,Escherichia-Shigella.1.4.2.2 Classification according to inflammationAccording to the relationship between intestinal flora and immune inflammation,the patients with insomnia of spleen deficiency type can be divided into inflammation promoting related flora and anti-inflammatory related flora.See the following table for details:2.1 Sleep qualityAfter intervention,the total scores of PSQI and ISI in the two groups decreased significantly（P<0.05）,and the subjective sleep quality of the patients improved.There was no significant difference between the two groups（P>0.05）2.2 Gastrointestinal symptomsAfter intervention,the total score of GSRS in the experimental group was significantly lower than that before intervention（P<0.05）.That in the control group was lower than that before the intervention（P>0.05）.2.3 Safety evaluation（1）Shedding cases:There were 16 cases were shedding during the intervention period of this study.Among them,there were 4 patients who withdrew consciously while taking placebo,2 patients withdrew due to conscious discomfort while taking Buzhong Yiqi Decoction,and 1 patient withdrew from Buzhong Yiqi Decoction without conscious effect.The remaining 9 patients withdrew from the trial for other reasons.（2）Inspection index:in this study,the abnormal indexes of the experimental group mainly reflected in the aspects of uric acid,urine routine and ECG.There were 5 patients,including 1 case of urine occult blood,3 cases of the increase in uric acid,1 case of the electrocardiogram of short P-R interval.The abnormal indexes in the control group were mainly reflected in liver function,uric acid,urine routine,stool routine and ECG.There were 5 cases in total,including 1 case of the slightly increase in glutamic oxaloacetic transaminase（GOT）,1 case of the increase in uric acid,1 case of urine occult blood,1 case of weak positive fecal occult blood test（FOBT）,1 case of abnormal ST-T segment of ECG.（3）Adverse events:there were 9 patients,which had mild adverse events in the experimental group.Among them,2 cases had symptoms in digestive system,6 cases had symptoms in respiratory system and 1 case had symptoms in skin.There were 13 patients,which had mild adverse events in the control group.Among them,6 cases had symptoms in digestive system,6 cases had symptoms in respiratory system and 1 case had symptoms in skin.All the adverse events were improved and cured before the end of the intervention.3.The effect mechanism of Buzhongyiqi Tang on insomnia with spleen-deficiency 3.1 The changes of intestinal flora after intervention3.1.1 α-DiversityAfter intervention,the Simpson index of α-diversity in the experimental group showed a trend of first decreasing and then slightly increasing.There was no significant difference（P>0.05）.3.1.2 The diversity of intestinal floraAt the phylum level,the abundance of Synergistetes increased（P<0.05）;the abundance of Fusobacteria and Spirochaetes decreased（P<0.05）.At the Class level,the intestinal flora with significantly increased abundance（P<0.05）includes:Negativicutes,Deltaproteobacteria,and Synergistia;abundance is significantly reduced The intestinal flora（P<0.05）includes:Fusobacteriia,Actinobacteria,Erysipelotrichia,Melainabacteria,and Spirochaetia.At the gnues level,After intervention,the abundance of bacteria in the experimental group increased significantly（P<0.05）as follows:Phascolarctobacterium and Prevotellaceae,which were belongs to probiotics;The abundance of bacteria in the experimental group decreased significantly（P<0.05）as follows:probiotics such as Faecalibacterium,Bifidobacterium,Anaerostipes and Prevotellaceae-2,and conditioned pathogen such as Fusobacterium,Escherichia-Shigella and Clostridium_sensu_stricto₁.3.1.3 Firmicutes/Bacillus（F/B）ratio,Bacillus/Clostridia（B/C）ratio and Bifidobacterium/Escherichia-Shigella（B/E）After intervention,the F/B ratio,B/C ratio and B/E ratio of the experimental group increased first and then decreased（P>0.05）.3.2 Correlation analysis of bacteria and sleep,Gastrointestinal tract before and after intervention.At the level of the class,PSQI total scores are linear with Actinobacteria,Negativicutes,Deltaproteobact eria,Melainabacteria,and Synergistia.Negative correlation trend,PSQI total score showed a linear positive correlation trend with Fusobacteriia and Erysipelotrichia,but the correlation were not statistically significant（P>0.05）;There were linear negative correlation between GSRS total score and Bifidobacteria（P<0.05）,and there were no correlation between GSRS and other Bacterobacteria.ConclusionIn this study,the patients with chronic moderate insomnia and spleen deficiency syndrome,accompanied with gastrointestinal symptoms and intestinal flora composition disorder.Insomnia may be related to the immune inflammation caused by the flora.Buzhong Yiqi Decoction intervenes in spleen-deficiency insomnia with obvious curative effect and safety,and it can improve the gastrointestinal state of insomnia and regulation disorder.The mechanism of its efficacy may be related to the regulation of intestinal flora,increase the abundance of anti-inflammatory-associated flora,reducing the abundance of proinflammatory bacteria,regulating probiotics and conditionally pathogenic bacteria.Thereby it may health for insomnia patients to improve the immune level and regulate the intestinal flora.