Intracellular Insulin and Impaired Autophagy in a Zebrafish model and a Cell Model of Type 2 diabetes

Int J Biol Sci. 2017 Jul 18;13(8):985-995. doi: 10.7150/ijbs.19249. eCollection 2017.

Abstract

Type 2 diabetes mellitus is characterized by insulin resistance. However, the complete molecular mechanism remains unclear. In this study, zebrafish were fed a long-term high-fat diet to induce type 2 diabetes, which resulted in a higher body weight, body mass index, more lipid vacuoles in liver, increased insulin transcription level in liver, brain and muscle, and high fasting blood glucose in the high-fat diet zebrafish. Oppositely, the transcription levels of insulin substrate-2 and glucose transporter 2 were significantly decreased, indicating insulin signaling pathway and glucose transport impaired in the insulin-targeting tissues. Transcription of the autophagy-related genes, ATG3, ATG4B, ATG5, ATG7, ATG12, and FOXO3, were decreased but autophagy inhibitor gene m-TOR increased, and autophagy-flux was inhibited in liver of the high-fat diet zebrafish. Main of these changes were confirmed in palmitic acid-treated HepG2 cells. Further, in co-immunoprecipitation and subcellular co-localization experiments, the conjunction of preproinsulin with cargo-recognition protein p62 increased, but conjuncts of autophagosome with p62-cargo, lysosomes with p62-cargo, and autolysosomes decreased apparently. Interestingly, lysosomes, autolysosomes and conjuncts of p62-insulin localized at the periphery of palmitic acid-treated cells, the margination of lysosomes may mediate deactivation of proteases activity. These findings suggest that intracellular high-lipid may trigger defective autophagy, defective downstream signaling of insulin and accumulated intracellular preproinsulin, leading to dysregulation of cell homeostasis mechanism, which may be one of reasons involved in insulin-resistance in type 2 diabetes.

Keywords: animal models; defective autophagy; insulin-targeting tissues; preproinsulin; subcellular localization; type 2 diabetes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Autophagy / physiology*
  • Autophagy-Related Proteins / metabolism
  • Blotting, Western
  • Diabetes Mellitus, Type 2 / metabolism*
  • Glucose Transporter Type 2 / metabolism
  • Hep G2 Cells
  • Humans
  • Immunoprecipitation
  • Insulin / metabolism*
  • Protein Precursors / metabolism
  • Real-Time Polymerase Chain Reaction
  • Zebrafish

Substances

  • Autophagy-Related Proteins
  • Glucose Transporter Type 2
  • Insulin
  • Protein Precursors
  • preproinsulin