Abstract 18576: Hypoxia Induces Phenotypic Switching of Human Pericytes Into Smooth Muscle Cells That is Associated With Changes in Pericyte-specific Non-coding RNA Expression
Background: Pericytes are pivotal for endothelial barrier integrity, angiogenesis, organ function and wound healing. While endothelial hypoxic reponses are well explored, knowledge on the effects of hypoxia on human pericytes (hPC) is sparse. Non-coding RNAs (ncRNAs) control gene expression by various mechanisms, but their functions in hPC remain unknown. We hypothesized that hypoxia influences the hPC-specific transcriptome and hPC phenotype.
Methods: hPC were validated by analyzing established pericyte expression markers via qRT-PCR and immunofluorescence (IF). Subsequently, hPC and HUVEC were subjected towards hypoxia (24h 1%O2), and next generation RNA deep sequencing (RNAseq) was performed. hPC viability following hypoxia was analyzed by CalceinRed assay.
Results: hPC expressed specific pericyte markers (Desmin, NG2, alphaSMA, PDGFRb) on mRNA and protein level. Cell viability of hPC was not affected by hypoxia. RNAseq revealed that 109 coding genes are significantly regulated in both HUVEC and hPC, whereas 520 coding genes were significantly regulated specifically in hPC (P<0.05, n=3). Interestingly, we identified a number of smooth muscle cell (SMC)-related genes that were increased in hypoxic hPC. The increase of the prototypic SMC marker alphaSMA was confirmed by qRT-PCR (2.23±1.18 fold induction) and IF. We observed a significant regulation of 6 ncRNAs, which were commonly regulated in HUVEC and in hPC, including the known hypoxia-induced miR-210 cluster (HUVEC: 7.86±0.09; hPC: 7.1±0.72 fold induction). Further 36 hPC-specific ncRNAs were identified, among them 25 long intergenic ncRNAs (lincRNAs) and 10 antisense RNAs (P<0.05,n=3). Notably, lincRNA H19, that is known to be important for muscle cell differentiation, was strongly upregulated in hypoxic hPC (4.24±1.1 fold induction, P<0.001,n=3).
Conclusion: Hypoxia triggers hPC differentiation towards a SMC phenotype that is associated with a significant change in hPC-specific ncRNA expression without affecting hPC viability. The functions of the newly identified ncRNAs are currently investigated. Our findings may be relevant to understand the role of pericytes in persistent vascular constriction and vessel destabilization in stroke or myocardial infarction.
Author Disclosures: C.M. Zehendner: None. S. Demolli: None. K. Michalik: None. N. Jaé: None. J. Wagner: None. D. John: None. S. Uchida: None. A.M. Zeiher: None. S. Dimmeler: None.
- © 2014 by American Heart Association, Inc.