Fig. 6.

GPx4 enters the mitochondria through TOM/TIM complex. E47 cells were transferred with TOM20, TOM22 or TOM70 siRNA for 48 hrs prior to CCl4 exposure for 12 hrs. (a) Representative fluorescent images of co-localization of GPx4 (red) with HSP60 (green). White arrowheads point to the co-localization of GPx4 with HSP60 (yellow surrounded by green); (b-f) Fluorescence intensity curve of GPx4 and HSP60 in co-localization images of Scramble-DMSO, Scramble-CCl4, siTOM20-CCl4, siTOM22-CCl4 and siTOM70-CCl4 groups; (g) Representative immunoblots of GPx4 level in total (Vinculin as the loading control), mitochondrial (HSP60 as the loading control) and cytoplasmic (Vinculin as the loading control) compartments; (h) Quantitated GPx4 level in total, mitochondria and cytoplasm. (n = 5–6/group); and (i) Schematic diagram depicting proposed mechanism where FUNDC1 promoted hepatocyte injury in liver fibrosis via direct binding of GPx4 to facilitate its mitochondrial translocation through TOM/TIM complex, where GPx4 was degraded by mitophagy, leading to hepatic ferroptosis. Mean ± SEM (detailed statistical results shown in Table S4); Statistical significance was set at p < 0.05. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)