Intrabody against prolyl hydroxylase 2 promotes angiogenesis by stabilizing hypoxia-inducible factor-1?.

ABSTRACT: Hypoxia-inducible factor (HIF)-1? is a crucial transcription factor that regulates the expression of target genes involved in angiogenesis. Prolyl hydroxylase 2 (PHD2) dominantly hydroxylates two highly conserved proline residues of HIF-1? to promote its degradation. This study was designed to construct an intrabody against PHD2 that can inhibit PHD2 activity and promote angiogenesis. Single-chain variable fragment (scFv) against PHD2, INP, was isolated by phage display technique and was modified with an endoplasmic reticulum (ER) sequence to obtain ER-retained intrabody against PHD2 (ER-INP). ER-INP was efficiently expressed and bound to PHD2 in cells, significantly increased the levels of HIF-1?, and decreased hydroxylated HIF-1? in human embryonic kidney cell line (HEK293) cells and mouse mononuclear macrophage leukaemia cell line (RAW264.7) cells. ER-INP has shown distinct angiogenic activity both in vitro and in vivo, as ER-INP expression significantly promoted the migration and tube formation of human umbilical vein endothelial cells (HUVECs) and enhanced angiogenesis of chick chorioallantoic membranes (CAMs). Furthermore, ER-INP promoted distinct expression and secretion of a range of angiogenic factors. To the best of our knowledge, this is the first study to report an ER-INP intrabody enhancing angiogenesis by blocking PHD2 activity to increase HIF-1? abundance and activity. These results indicate that ER-INP may play a role in the clinical treatment of tissue injury and ischemic diseases in the future.

SUBMITTER: Zhao L 

PROVIDER: S-EPMC6694103 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature