This study aimed to investigate the effects of fish oil-derived omega-3 polyunsaturated fatty acids (omega-3 PUFAs) on gut microbiota and serum lipid metabolites in T2DM. In a three-month, randomized, double-blind, placebo-controlled study, 110 T2DM patients received either fish oil (n = 55) or corn oil (n = 55) capsules daily. Serum lipids, glycemic parameters, gut microbiota diversity, and lipidomics were assessed. This study found that fish oil-derived omega-3 PUFAs intervention did not significantly lower the fasting plasma glucose levels when compared with the baseline level (p > 0.05). However, serum fasting blood glucose (p = 0.039), glycosylated hemoglobin levels (p = 0.048), HOMA-IR (p = 0.022), total cholesterol (p < 0.001), triglyceride (p = 0.034), LDL cholesterol (p = 0.048), and non-HDL levels (p = 0.046) were significantly lower in the fish oil group compared with the corn oil group after three months of intervention. Also, it altered glycerophospholipid metabolism and gut microbiota. After three months, the fish oil group showed a significantly lower abundance of Desulfobacterota compared with the corn oil control group (p = 0.003), with reduced levels of Colidextribacter (p = 0.002), Ralstonia (p = 0.021), and Klebsiella (p = 0.013). Conversely, the abundance of Limosilactobacillus (p = 0.017), Lactobacillus (p = 0.011), and Haemophilus (p = 0.018) increased significantly. In addition, relevant glycolipid metabolism indicators showed significant correlations with the altered profiles of serum lipid metabolites, intestinal bacteria, and fungi. This study highlights the impact of fish oil-derived omega-3 PUFAs on intestinal microbiota structure and function in patients with type 2 diabetes. The observed decrease in pathogenic bacterial species and the enhancement of beneficial species may have significant implications for gut health and systemic inflammation, both of which are pivotal in managing diabetes. Further research is warranted to comprehensively elucidate the long-term benefits and underlying mechanisms of these microbiota alterations.
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