The Mediation/Moderation Effects of Gut Microbiota on Sleep Quality and Primary Liver Cancer: A Mendelian Randomization and Case–Control Study

Figures & data

Figure 1 Flow chart of the case–control study. *By Figdraw.

Figure 2 Clinicopathological features of individuals with and without Primary Liver Cancer. *P < 0.05; **P < 0.01; ***P ≤ 0.001.

Abbreviations: AFP, Alpha Fetoprotein; ALB, Albumin; ALT, Alanine aminotransferase; AST, Aspartate Transaminase; BMI, Body Mass Index; CA199, Carbohydrate Antigen199; CEA, Carcinoembryonic Antigen; HBV, Hepatitis B Virus; PT, Prothrombin Time; TBIL, Total Bilirubin.

Figure 3 Analysis and description of differences in alpha diversity and beta diversity of gut microbiota between the individuals with and without Primary Liver Cancer. (A) Observed_otus; (B) Goods_coverage; (C) Chao Index; (D) Simpson Index; (E) PCoA of weighted Unifrac distance matrix; (F) NMDS Analysis; *P < 0.05; **P < 0.01; ***P < 0.001.

Abbreviations: NMDS, non-metric multidimensional scaling; PCoA, principal coordinates analysis.

Figure 4 Differences of gut microbiota between the individuals with and without Primary Liver Cancer. (A) Predominant taxa distribution in two group were showed in a phylogenetic tree with clad-ogram; (B) Differential flora between two group were showed with the bar plot.

Table 1 The Mediation Effects of Various Levels of Gut Microbiota on the Relationship Between Sleep Quality and Primary Liver Cancer

Level	Class
Gut microbiota	Actinobacteria
Sleep status	PSQI Score	Self-sleep quality assessment
Mediation effect	0.003	0.018
95% CI Lower	0.00003	0.00044
95% CI Upper	0.01	0.05
P-value	0.048*	0.046*
Direct effect	0.05	0.15
95% CI Lower	0.03	0.05
95% CI Upper	0.06	0.24
P-value	<0.001***	<0.001***
Total effect	0.05	0.17
95% CI Lower	0.04	0.07
95% CI Upper	0.06	0.26
P-value	<0.001***	<0.001***
Proportion of Mediation effect	6.09%	10.88%

Notes: *P-value < 0.05; **P-value < 0.01; ***P-value < 0.001. P-values below 0.05 are highlighted in bold typeface.

Abbreviations: PSQI Score, Pittsburgh Sleep Quality Index Score; 95% CI, 95% confidence interval.

Table 2 The Moderation Effects of Various Levels of Gut Microbiota on the Relationship Between Sleep Quality and Primary Liver Cancer

Level	Gut Microbiota	Sleep Status	Effect	Estimate	SE	[95% CI]	P-value
Class	Actinobacteria	Sleep time at night	Main effect	0.597	0.230	[0.146, 1.047]	0.009**
			Moderation effect	−0.050	0.021	[−0.092, −0.007]	0.021*
Order	Erysipelotrichales	Daytime dysfunction	Main effect	0.640	0.172	[0.304, 0.976]	<0.001***
			Moderation effect	0.156	0.073	[0.012, 0.299]	0.033*
Genus	Roseburia	PSQIScore	Main effect	0.357	0.075	[0.210, 0.505]	<0.001***
			Moderation effect	0.168	0.070	[0.031, 0.306]	0.017*
		Time required to go to sleep at night	Main effect	0.574	0.159	[0.262, 0.886]	<0.001***
			Moderation effect	0.369	0.141	[0.093, 0.645]	0.009**
	Bifidobacterium	Sleep time at night	Main effect	0.617	0.228	[0.171, 1.064]	0.007**
			Moderation effect	−0.048	0.021	[−0.088, −0.007]	0.022*

Notes: *P-value < 0.05; **P-value < 0.01; ***P-value < 0.001. P-values below 0.05 are highlighted in bold typeface.

Abbreviations: PSQI Score, Pittsburgh Sleep Quality Index Score; 95% CI, 95% confidence interval.

Table 3 The Mediation Effects of Various Levels of Gut Microbiota on the Relationship Between Specific Sleep Disorder and Primary Liver Cancer

Specific Sleep Disorder	Level	Gut Microbiota	OR	[95% CI]	P-value	Proportion of Mediation Effect
Sleep time at night	Phylum	Elusimicrobia	1.052	[1.024, 1.096]	0.012*	34.23%
	Order	Micrococcales	1.030	[1.002, 1.055]	0.022*	34.48%
	Family	Micrococcaceae	1.033	[1.002, 1.055]	0.023*	35.97%
Daytime dysfunction	Order	Micrococcales	1.033	[1.010, 1.058]	0.010*	41.56%
	Family	Enterococcaceae	1.024	[1.013, 1.060]	0.038*	23.33%
	Family	Micrococcaceae	1.022	[1.009, 1.058]	0.011*	28.57%

Notes: *P-value < 0.05. P-values below 0.05 are highlighted in bold typeface.

Abbreviations: OR, odds ratio; 95% CI, 95% confidence interval.

Table 4 The Moderation Effects of Various Levels of Gut Microbiota on the Relationship Between Specific Sleep Disorder and Primary Liver Cancer

Specific Sleep Disorder	Level	Gut Microbiota	OR	[95% CI]	P-value
Self sleep quality assessment	Phylum	Epsilonbacteraeota	0.388	[0.072, 0.773]	0.037*
Time required to go to sleep at night	Class	Coriobacteriia	1.126	[1.008, 1.260]	0.036*
	Order	Coriobacteriales	1.115	[1.006, 1.262]	0.023*
	Family	Coriobacteriaceae	1.149	[1.015, 1.301]	0.028*
	Genus	Collinsella	1.148	[1.015, 1.300]	0.028*
	Genus	Roseburia	1.446	[1.097, 1.906]	0.009**
Sleep time at night	Phylum	Chloroflexi	0.065	[0.005, 0.287]	0.041*
	Class	Actinobacteria	0.951	[0.912, 0.993]	0.021*
	Class	Alphaproteobacteria	0.027	[0.001, 0.252]	0.018*
	Order	Bifidobacteriales	0.953	[0.916, 0.993]	0.022*
	Order	Pseudomonadales	0.311	[0.069, 0.514]	0.012*
	Family	Bifidobacteriaceae	0.965	[0.925, 0.993]	0.016*
	Family	Acidaminococcaceae	2.782	[1.181, 6.554]	0.019*
	Genus	Bifidobacterium	1.196	[1.007, 1.346]	0.012*
Daytime dysfunction	Phylum	Epsilonbacteraeota	0.233	[0.134, 0.407]	0.011*
	Class	Erysipelotrichia	1.143	[1.021, 1.579]	0.033*
	Class	Bacteroidetes_unclassified	0.029	[0.002, 0.190]	0.001**
	Class	Campylobacteria	0.213	[0.049, 0.412]	0.011*
	Order	Erysipelotrichales	1.169	[1.012, 1.349]	0.033*
	Family	Erysipelotrichaceae	1.159	[1.008, 1.432]	0.031*
	Family	Prevotellaceae	0.911	[0.834, 0.995]	0.039*
	Genus	Dialister	1.252	[1.007, 1.556]	0.043*

Notes: *P-value < 0.05; **P-value < 0.01. P-values below 0.05 are highlighted in bold typeface.

Abbreviations: OR, odds ratio; 95% CI, 95% confidence interval.

Data Sharing Statement

All GWAS summary statistics analysed in this study are publicly available for download by qualified researchers. The GWAS for the disorder of the sleep-wake schedule was provided by the Finland Database (https://r9.finngen.fi/pheno/F5_SLEEPWAKE). The GWAS for PLC was provided by the UK Biobank (https://github.com/Nealelab/UK_Biobank_GWAS). All data used to support the conclusions of the case–control study are accessible upon reasonable request from the principal investigator.