Objective:To investigate the alterations in gut microbiota diversity and inflammatory cytokine levels among patients with varying severities of insomnia, and to explore their interrelationships, in order to provide a theoretical basis for understanding the pathophysiology of insomnia.Methods:A total of 42 patients with chronic insomnia who visited the First Hospital of Hebei Medical University between March and December 2023 were enrolled in the insomnia group, and 22 age-and sex-matched healthy volunteers were recruited from the same hospital as the control group. General demographic data were collected, and Mini-International Neuropsychiatric Interview (MINI) was used to screen for comorbid psychiatric disorders. The Self-Rating Depression Scale (SDS) and the Self-Rating Anxiety Scale (SAS) were employed to evaluate individual′s depressive and anxiety symptoms. Sleep quality and insomnia severity were assessed using the Pittsburgh Sleep Quality Index (PSQI) and the Insomnia Severity Index (ISI), Participants′ gastrointestinal function and symptoms over the past week were evaluated using the Gastrointestinal Symptom Rating Scale (GSRS). Fecal and blood samples were collected from all participants. Gut microbiota diversity was analyzed using 16S rRNA sequencing. Differential taxa were identified using linear discriminant analysis effect size (LEfSe) and random forest analysis. Serum levels of inflammatory cytokines were measured by enzyme-linked immunosorbent assay (ELISA). Spearman correlation analysis was used to explore the relationships between insomnia symptoms, microbial diversity indices, key microbial taxa, and inflammatory markers. Multiple linear regression analysis was conducted to identify factors associated with insomnia severity.Results:Compared to the control group, both the mild insomnia group and the moderate-to-severe insomnia group showed significantly higher GSRS scores ( Z=-3.51, -2.72, both P<0.05). The Chao1 index was significantly lower in the mild and moderate-to-severe insomnia groups than in controls ( Z=-3.53, -3.87, both P<0.05). Similarly, the Observed species index was lower in both the mild and moderate-to-severe groups ( Z=-3.33, -3.74, both P<0.05). The Shannon index was significantly reduced in the moderate-to-severe group compared to both the mild group and controls ( Z=-2.81, -2.23, both P<0.05). The Simpson index in the moderate-to-severe group also tended to be lower than in the mild group ( Z=-1.95, P=0.051). Beta diversity differed significantly among the mild insomnia group, the moderate-to-severe insomnia group ( P<0.05), and the control group ( F=2.96, 3.12, both P<0.05). Random forest analysis identified Ruminococcus_D and Klebsiella as key microbial genera distinguishing between mild and moderate-to-severe insomnia. Inflammatory cytokine levels were significantly elevated in both insomnia groups compared to controls ( P<0.05). PSQI scores were negatively correlated with the Shannon index, the Observed species index, and the relative abundance of Ruminococcus_D ( r=-0.34, -0.30, and -0.25, respectively; all P<0.05). Multiple linear regression revealed that serum IL-1β (β=0.339, 95% CI=0.014-0.716, P=0.042) and Ruminococcus_D (β=-0.309, 95% CI=-194.591--8.318, P=0.034) were independent predictors of insomnia severity. Conclusion:Elevated inflammatory cytokine levels and reduced gut microbial richness may be closely associated with increased insomnia severity. Additionally, Ruminococcus_D and IL-1β may be important factors contributing to the severity of insomnia in affected individuals.

Objective:To investigate the alterations in gut microbiota diversity and inflammatory cytokine levels among patients with varying severities of insomnia, and to explore their interrelationships, in order to provide a theoretical basis for understanding the pathophysiology of insomnia.Methods:A total of 42 patients with chronic insomnia who visited the First Hospital of Hebei Medical University between March and December 2023 were enrolled in the insomnia group, and 22 age-and sex-matched healthy volunteers were recruited from the same hospital as the control group. General demographic data were collected, and Mini-International Neuropsychiatric Interview (MINI) was used to screen for comorbid psychiatric disorders. The Self-Rating Depression Scale (SDS) and the Self-Rating Anxiety Scale (SAS) were employed to evaluate individual′s depressive and anxiety symptoms. Sleep quality and insomnia severity were assessed using the Pittsburgh Sleep Quality Index (PSQI) and the Insomnia Severity Index (ISI), Participants′ gastrointestinal function and symptoms over the past week were evaluated using the Gastrointestinal Symptom Rating Scale (GSRS). Fecal and blood samples were collected from all participants. Gut microbiota diversity was analyzed using 16S rRNA sequencing. Differential taxa were identified using linear discriminant analysis effect size (LEfSe) and random forest analysis. Serum levels of inflammatory cytokines were measured by enzyme-linked immunosorbent assay (ELISA). Spearman correlation analysis was used to explore the relationships between insomnia symptoms, microbial diversity indices, key microbial taxa, and inflammatory markers. Multiple linear regression analysis was conducted to identify factors associated with insomnia severity.Results:Compared to the control group, both the mild insomnia group and the moderate-to-severe insomnia group showed significantly higher GSRS scores ( Z=-3.51, -2.72, both P<0.05). The Chao1 index was significantly lower in the mild and moderate-to-severe insomnia groups than in controls ( Z=-3.53, -3.87, both P<0.05). Similarly, the Observed species index was lower in both the mild and moderate-to-severe groups ( Z=-3.33, -3.74, both P<0.05). The Shannon index was significantly reduced in the moderate-to-severe group compared to both the mild group and controls ( Z=-2.81, -2.23, both P<0.05). The Simpson index in the moderate-to-severe group also tended to be lower than in the mild group ( Z=-1.95, P=0.051). Beta diversity differed significantly among the mild insomnia group, the moderate-to-severe insomnia group ( P<0.05), and the control group ( F=2.96, 3.12, both P<0.05). Random forest analysis identified Ruminococcus_D and Klebsiella as key microbial genera distinguishing between mild and moderate-to-severe insomnia. Inflammatory cytokine levels were significantly elevated in both insomnia groups compared to controls ( P<0.05). PSQI scores were negatively correlated with the Shannon index, the Observed species index, and the relative abundance of Ruminococcus_D ( r=-0.34, -0.30, and -0.25, respectively; all P<0.05). Multiple linear regression revealed that serum IL-1β (β=0.339, 95% CI=0.014-0.716, P=0.042) and Ruminococcus_D (β=-0.309, 95% CI=-194.591--8.318, P=0.034) were independent predictors of insomnia severity. Conclusion:Elevated inflammatory cytokine levels and reduced gut microbial richness may be closely associated with increased insomnia severity. Additionally, Ruminococcus_D and IL-1β may be important factors contributing to the severity of insomnia in affected individuals.

OBJECTIVE To study the effects of the Mongolian medicine Sugemule-4 on the metabolism of insomnia rats， and to preliminarily explore its possible mechanisms for improving insomnia. METHODS The rat model of chronic stress insomnia was established by tail clipping stimulation and intraperitoneal injection of p-chlorophenyl alanine solution. Twenty-four male rats were randomly divided into the normal group， model group， diazepam group （positive control， 0.92 mg/kg）， and Sugemule-4 group （5.2 g/kg）， with 6 rats in each group. Since the 7th day of tail clipping stimulation， the Sugemule-4 group and diazepam group began to be intragastrically administered with relevant medicine； the normal group and model group were intragastrically administered with an equal volume of distilled water， once a day， for 14 consecutive days. The learning and memory abilities of rats were tested using a water maze experiment， and the non-invasive sleep activity monitoring system was used to monitor the 24- hour sleep time of rats. A metabolomics study was conducted on rat serum and hippocampal tissue by using ultra-high-performance liquid chromatography-tandem mass spectrometry. The multivariate statistical analysis method was adopted to analyze the differential metabolites in serum and hippocampal tissue of rats， and screen for differential metabolites and metabolic pathways among those groups. RESULTS Compared with the normal group， the escape latency of rats in the model group was significantly increased， the times of crossing platforms were significantly reduced， and the percentage of average 24-hour sleep time was significantly reduced （P＜0.05）. Compared with the model group， the levels of the above indicators were significantly reversed in the diazepam group and Sugemule-4 group （P＜0.05）. Metabolomics studies found that a total of 9 differential metabolites were identified in rat serum and hippocampal tissue， including 5-hydroxyindoleacetic acid， canine urate， canine urinary quinolinic acid， 5-hydroxytryptamine， phenol sulfate， 1-carboxyethyltyrosine， 3-（4-hydroxyphenyl） lactate， N-acetyl tyrosine， tyrosine and phenol sulfate， mainly involving 2 metabolic pathways of tryptophan and tyrosine.CONCLUSIONS Sugemule-4 can improve the sleep time and behavioral performance of insomnia rats， and its mechanism may be associated with affecting amino acid metabolic pathways such as tryptophan and tyrosine.