Figure 5.

Autophagy is not activated in TXNIP-KO mice during fasting. Deficiency in autophagy was observed in the liver of TXNIP-KO mice. (a) The morphological differences of mitochondria between WT and TXNIP-KO mice were examined by TEM. In the liver of WT mice, autophagy increases during the fasted state, and in the present study, increased number of autophagosomes (arrow heads) containing mitochondria (arrows) were observed. By contrast, autophagosomes were rarely found in the liver of TXNIP-KO mice by TEM analysis. (b,c) The levels of LC3 mRNA and protein were evaluated by RT-qPCR and western blotting, respectively. The expression LC3 mRNA was upregulated during fasting in both WT and TXNIP-KO mice but there was no obvious difference between WT and KO mice. In WT mice the expression of LC3-II, a key molecule in autophagy, was upregulated during the fasted state, but this change was not observed in TXNIP-KO mice. The expression of LC3-II was statistically lower in TXNIP-KO mice during the fasted state. (c,e) The levels of Pink1 and Parkin, key molecules in mitophagy, were examined by RT-qPCR and western blotting, revealing that the expression patterns during the fed and fasted states were similar between WT and KO mice. (f) Other proteins playing essential roles in autophagy, Atg5, Atg14, Lamp1, Lamp2, and Beclin, were examined, and no statistical significance was found between WT and TXNIP-KO mice in the fed and fasted state in both strains. *p < 0.05, **p < 0.01 (five mice per strain were used for RT-qPCR, and ten were used for western blotting). TXNIP thioredoxin-interacting protein, WT wild-type mice, KO knockout mice, WT wild-type mice, KO knockout mice, TEM transmission electron microscopy, LC3 microtubule-associated protein light chain 3, Pink1 PTEN induced kinase 1, Atg5 autophagy related 5, Atg14 autophagy related 14, Lamp1 lysosomal-associated membrane protein 1, Lamp2 lysosomal-associated membrane protein 2.