Tangerine Peel-Derived Exosome-Like Nanovesicles Alleviate Hepatic Steatosis Induced by Type 2 Diabetes: Evidenced by Regulating Lipid Metabolism and Intestinal Microflora

Junju Zou,1– 3,* Qianbo Song,2,* Pang Chui Shaw,4 Yongjun Wu,1,5 Zhong Zuo,2 Rong Yu1,3 1School of Traditional Chinese Medicine, Hunan University of Chinese Medicine, Changsha, Hunan, 410208, People’s Republic of China; 2School of Pharmacy, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong SAR, People’s Republic of China; 3Hunan Key Laboratory of Traditional Chinese Medicine Prescription and Syndromes Translational Medicine, School of Pharmacy, Hunan University of Chinese Medicine, Changsha, Hunan, 410208, People’s Republic of China; 4School of Life Sciences, The Chinese University of Hong Kong, Hong Kong SAR, People’s Republic of China; 5School of Pharmacy, Hunan University of Chinese Medicine, Changsha, Hunan, 410208, People’s Republic of China*These authors contributed equally to this workCorrespondence: Rong Yu, School of Traditional Chinese Medicine, Hunan University of Chinese Medicine, Changsha, 410208, People’s Republic of China, Email [email protected] Zhong Zuo, School of Pharmacy, the Chinese University of Hong Kong, Hong Kong, SAR, 999077, People’s Republic of China, Email [email protected]: Non-alcoholic fatty liver disease (NAFLD) represents a significant global health burden, exhibiting a strong correlation with insulin resistance, obesity, and type 2 diabetes (T2DM). Despite the severity of hepatic steatosis in T2DM patients, no specific drugs have been approved for clinical treatment of the disease. Tangerine peel is one kind of popular functional food and reported to possess hypoglycemic and lipid-lowering potential. In this study, we investigated the effects of Tangerine-peel-derived exosome-like nanovesicles (TNVs) on hepatic lipotoxicity associated with T2DM.Methods: The TNVs was prepared by differential centrifugation of the aqueous extract of Tangerine and chemical properties were characterized using transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA) and LC-MS/MS. The hypoglycemic and lipid-lowering potential of TNVs were possessed by biochemical measurement, RT-PCR, 16S rRNA sequencing, GC/MS, UHPLC-MS/MS, in vivo small animal imaging assay and HE staining. Subsequently, effects of TNVs on lipid accumulation and glycolysis were investigated on 3T3-L1 and AML-12 cells.Results: TNVs significantly inhibited insulin resistance, reduced hepatic lipid accumulation, facilitate intestinal mucosal repair, rescued gut microbiota dysbiosis, regulated colonic SCFA and liver bile acid metabolism in db/db mice. Furthermore, TNVs restored the expression of key genes in glucose and lipid metabolism (ACC, AMPK, CD36, LXRα, PPAR-γ, SREBP-1) while activating the expression of genes related to glycolysis (G6Pase, GLUT2, PCK1, PEPCK) in db/db mice. Further cell-based mechanistic studies revealed that TNVs reduced lipid accumulation in 3T3-L1 and AML-12 cells via regulation of glucose and lipid metabolism-related genes (UCP1, FGFR4, PRDM16, PGC-1α, Tmem26, Cpt1, Cpt2 and PPAR-α).Conclusion: We for the first time demonstrated that TNVs could significantly improve glucose and lipid metabolism via activating the expression of genes related to fatty acid β-oxidation and glycolysis. Keywords: tangerine peel, plant-derived exosome-like nanoparticles, diabetes mellitus, metabolomics, gut microbiota, fat accumulation