Hypoxia-induced autophagy of stellate cells inhibits expression and secretion of lumican into microenvironment of pancreatic ductal adenocarcinoma.

Abstract

Lumican is secreted by pancreatic stellate cells and inhibits cancer progression. Extracellular lumican inhibits cancer cell replication and restrains growth of early-stage pancreatic adenocarcinoma (PDAC) such that patients with tumors containing stromal lumican experience a three-fold longer survival after treatment. In the present study, patient tumor tissues, ex-vivo cultures of patient-derived xenografts (PDX), PDAC stellate and tumor cells were used to investigate whether hypoxia (1% O2) within the tumor microenvironment influences stromal lumican expression and secretion. We observed that hypoxia significantly reduced lumican expression and secretion from pancreatic stellate cells, but not cancer cells. Although hypoxia enhanced lactate dehydrogenase A (LDHA) expression and lactate secretion from all cells, neither hypoxia-induced nor exogenous lactate influenced lumican expression. Autophagy was induced by hypoxia in ex vivo cultures of PDX and pancreatic stellate cells, but not cancer cells cultured in 2D. Autophagic flux inhibitors, bafilomycin A1, chloroquine diphosphate salt, and ammonium chloride prevented hypoxia-mediated reduction in lumican expression in stellate cells. Furthermore, inhibition of AMP-regulated protein kinase (AMPK) phosphorylation or hypoxia-inducible factor (HIF)-1α expression within hypoxic stellate cells restored lumican expression levels. Hypoxia did not affect lumican mRNA expression, indicating that hypoxia-induced reduction of lumican occurs post-transcriptionally; in addition, AMPK inhibition prevented hypoxia-reduced phosphorylation of the mTOR/p70S6K/4EBP signaling pathway, a key contributor to protein synthesis. Taken together, these findings demonstrate that hypoxia reduces stromal lumican in PDAC through autophagy-mediated degradation and reduction in protein synthesis within pancreatic cancer stellate cells.