[Specific changes of intestinal microflora in children with nonalcoholic fatty liver disease]

Objective: To analyze the composition and richness of intestinal microflora in children with non-alcoholic fatty liver disease (NAFLD) and the role of which in pathogenesis of NAFLD. Methods: This was a prospective case-control study. From November 2015 to June 2017, 19 children diagnosed with NAFLD according to the 2010 edition of diagnostic criteria were enrolled voluntarily in the Second and First Affiliated Hospitals of Zhejiang Chinese Medicine University. Twenty-two healthy children were enrolled in the control group. Among the patients, 10 were males and 9 were females, at the mean age of (11.0±1.0) years; 10 males and 12 females in the control group, at the mean age of (9.0±1.2) years. The body mass index (BMI) and waist circumference were recorded, and the fasting blood glucose, total cholesterol, triglyceride, high-density lipoprotein and low-density lipoprotein were detected. Feces were collected and the fecal microorganisms were extracted with magnetic beads methods; the composition and the richness of intestinal microflora in the two groups were detected with 16S rDNA high throughput sequencing technology. The KO differential gene expression and KEGG signal pathway enrichment were analyzed with PICRUST software. The intestinal flora characteristics between the two groups were compared with t test or Mann-Whitney U test and Willcoxon W test. Results: The BMI, waist circumference and triglyceride were higher in NAFLD group than those in the control group (BMI (25.1±2.7) vs. (18.2±1.5)kg/m(2), t=9.912, P=0.000; waist circumference (88.6±6.6) vs. (71.5±6.3) cm, t=8.520, P=0.000; triglyceride (0.9±0.4) vs.(0.7±0.3)mmol/L, t=2.060, P=0.046). The abundance and diversity index of intestinal microflora were lower in the NAFLD group (Shannon index 3.99 (3.13, 4.54) vs. 4.63 (4.21, 4.81), Z=-2.065, P=0.039; Simpson index 0.85 (0.70, 0.89) vs. 0.90 (0.88, 0.93), Z=-2.431, P=0.015; ACE index 235.76 (205.26, 361.94) vs. 326.96 (275.34, 368.65), Z=-2.092, P=0.036). At the level of phylum, the proportion of Actinomycetes was lower and the proportion of Thermus was higher in NAFLD group (Actinobacteri 29.807 (14.723, 62.080) ×10(-3) vs. 63.212 (46.133, 172.071) ×10(-3), Z=-2.667, P=0.008; Thermus 0.033 (0.000, 0.226) ×10(-3) vs. 0.000 (0.000, 0.031) ×10(-3), Z=-2.729, P=0.006) . At the level of genus, the proportion of Bacteroides and Bifidobacterium in the NAFLD group were significantly lower (Bacteroides 78.757 (11.430, 151.621) ×10(-3) vs. 356.821 (161.049, 403.037) ×10(-3), Z=-2.771, P=0.006; Bifidobacterium 19.680 (6.181, 53.944) ×10(-3) vs. 54.721 (31.911, 146.410) ×10(-3), Z=-2.458, P=0.014); the proportion of Prevotella in NAFLD group was significantly higher (3.089 (0.165, 63.502) ×10(-3) vs. 0.432 (0.029, 2.257) ×10(-3), Z=-2.112, P=0.035). Based on the KEGG database, 78 differentially expressed genes and 26 differential metabolic pathways were found, among which the function genes of K01470, K01961 and K07258 were concentrated in the pathways of arginine and proline metabolism, fatty acid synthesis, and polysaccharides biosynthesis and metabolism. Besides, these three function genes were related to Bacteroides, Prevotella, Bifidobacterium and Ruminococcus. Conclusion: NAFLD children have intestinal flora disturbances in both diversity and abundance, which may alter lipid metabolic pathways through differential gene expressions, contributing to the pathogenesis of NAFLD.