UBC Theses and Dissertations
Requirement of clusterin expression for prosurvival autophagy in hypoxic kidney tubular epithelial cells Alnasser, Hatem
Cellular autophagy is a prosurvival mechanism in the kidney against ischemia reperfusion injury (IRI), but the molecular pathways to activating autophagy in ischemic kidneys are not fully understood. Clusterin (CLU) is a chaperone-like protein and its expression is associated with kidney resistance to IRI. This study investigated the role of CLU in the prosurvival autophagy in the kidney. Renal IRI was induced in mice by clamping renal pedicles at 32°C for 45 min. Hypoxia in renal tubular epithelial cell (TEC) cultures was induced by exposure to 1% O₂ atmosphere. Autophagy was determined by either LC3-BII expression in Western blot or LC3-GFP aggregation in confocal microscopy. Cell apoptosis was determined by flow cytometric analysis. Unfolded protein response (UPR) was determined by PCR array. Here, we showed that autophagy was significantly activated by IRI in wild type (WT) but not CLU deficient kidneys. Similarly, the autophagy was activated by hypoxia in human proximal TECs (HKC-8) and WT mouse primary TECs but was impaired in CLU null TECs. Hypoxia activated autophagy was CLU dependent and was positively correlated with cell survival, and inhibition of autophagy significantly promoted cell death both in HKC-8 and mouse WT/CLU expressing TECs, but not in CLU null TECs. Further studies showed that CLU-dependent prosurvival autophagy was associated with UPR activation in hypoxic kidney cells. In conclusion, these data suggest that activation of prosurvival autophagy by hypoxia in kidney cells is required CLU expression, and may be a cytoprotective mechanism of CLU in the protection of the kidney from hypoxia/ischemia-mediated injury.
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