Engineered myocardium model to study the roles of HIF-1α and HIF1A-AS1 in paracrine-only signaling under pathological level oxidative stress

Abstract

Studying heart tissue is critical for understanding and developing treatments for cardiovascular diseases. In this work, we fabricated precisely controlled and biomimetic engineered model tissues to study how cell-cell and cell-matrix interactions influence myocardial cell survival upon exposure to pathological level oxidative stress. Specifically, the interactions of endothelial cells (ECs) and cardiomyocytes (CMs), and the role of hypoxia inducible factor-1α (HIF-1α), with its novel alternative regulator, HIF-1α antisense RNA1 (HIF1A-AS1), in these interactions were investigated. We encapsulated CMs in photo-crosslinkable, biomimetic hydrogels with or without ECs, then exposed to oxidative stress followed by normoxia. With precisely controlled microenvironment provided by the model tissues, cell-cell interactions were restricted to be solely through the secreted factors. CM survival after oxidative stress was significantly improved, in the presence of ECs, when cells were in the model tissues that were functionalized with cell attachment motifs. Importantly, the cardioprotective effect of ECs was reduced when HIF-1α expression was knocked down suggesting that HIF-1α is involved in cardioprotection from oxidative damage, provided through secreted factors conferred by the ECs. Using model tissues, we showed that cell survival increased with increased cell-cell communication and enhanced cell-matrix interactions. In addition, whole genome transcriptome analysis showed, for the first time to our knowledge, a possible role for HIF1A-AS1 in oxidative regulation of HIF-1α. We showed that although HIF1A-AS1 knockdown helps CM survival, its effect is overridden by CM-EC bidirectional interactions as we showed that the conditioned media taken from the CM-EC co-cultures improved CM survival, regardless of HIF1A-AS1 expression.

Human iPSC-derived myocardium-on-chip with capillary-like flow for personalized medicine

“…Previous characterizations from our group have shown that cells differentiated using this protocol are very similar to HUVECs in mRNA and protein expression as well as in tube formation ability. 43 In this study, we characterize the cells further under flow. iECs seeded into both the single channel device and the MOC appeared morphologically similar to primary endothelial cells.…”

“…39 In previous studies, we have characterized the phenotypical and functional properties of these iEC under static culture conditions. 43 In this study, we seeded iECs into a microfluidic device with a single channel (0.1 mm high  1 mm wide  10 mm long, Figure 4) to apply physiological-level shear stress and examined the morphology and expression of endothelial cell-specific markers under flow. iECs were seeded into the single-channel microfluidic device (Figure 4(b)) and 24 h after seeding, the device was perfused with culture media up to 72 h. For the first 24 h, the perfusion was kept at a flow rate that corresponded to a wall shear stress of 9.7 dynes/cm 2 which corresponds to average values seen in human capillaries (Figure 4(c)).…”

Transcriptome profiling of 3D co-cultured cardiomyocytes and endothelial cells under oxidative stress using a photocrosslinkable hydrogel system

“…Taken together, these results indicate a protective effect of the co-culture condition on the CMs as we have investigated in detail in our recent publication [33]. In order to evaluate the crosstalk between CMs and iECs in response to oxidative stress, the transcriptome of CMs in single culture or in the co-culture with iECs with or without H 2 O 2 treatment were profiled and screened for significantly changed genes.…”

Technical Feasibility and Physiological Relevance of Hypoxic Cell Culture Models

“…Similarly, 3D cell culture technology has been used to study the effect of hypoxia in the context of ischemia in various cell types, including cardiomyocytes (152)(153)(154), astrocytes (155), endothelial cells (156), and hypoxia related to pulmonary fibrosis in fetal lung fibroblasts (157). Furthermore, the effects of both continuous hypoxia and IH on vascular sprouting has been explored in endothelial cells (158)(159)(160).…”