HIF1α Regulates Early Metabolic Changes due to Activation of Innate Immunity in Nuclear Reprogramming

Abstract

Innate immune signaling has recently been shown to play an important role in nuclear reprogramming, by altering the epigenetic landscape and thereby facilitating transcription. However, the mechanisms that link innate immune activation and metabolic regulation in pluripotent stem cells remain poorly defined, particularly with regard to key molecular components. In this study, we show that hypoxiainducible factor 1a (HIF1a), a central regulator of adaptation to limiting oxygen tension, is an unexpected but crucial regulator of innate immune-mediated nuclear reprogramming. HIF1a is dramatically upregulated as a consequence of Toll-like receptor 3 (TLR3) signaling and is necessary for efficient induction of pluripotency and transdifferentiation. Bioenergetics studies reveal that HIF1a regulates the reconfiguration of innate immune-mediated reprogramming through its well-established role in throwing a glycolytic switch. We believe that results from these studies can help us better understand the influence of immune signaling in tissue regeneration and lead to new therapeutic strategies.

Stemness of Human Pluripotent Cells: Hypoxia-Like Response Induced by Low Nitric Oxide

“…Hydroxylated HIF-α is targeted for ubiquitination by E3 ubiquitin protein ligase containing Von Hippel–Lindau protein (VHL). Hypoxia also enhances the generation of inducible pluripotent stem cells (iPSCs) [ 18 , 19 ], and it has been reported that reprogramming cells to pluripotency requires HIF-1α and HIF-2α at an early state to induce the metabolic switch from an oxidative to a highly glycolytic state, as well as for the acquisition of an undifferentiated state [ 19 , 20 ]. The evidence thus reviewed indicates that changes in metabolism are relevant for transition from the pluripotent to the differentiated state.…”

Consecutive Hypoxia Decreases Expression of NOTCH3, HEY1, CC10, and FOXJ1 via NKX2-1 Downregulation and Intermittent Hypoxia-Reoxygenation Increases Expression of BMP4, NOTCH1, MKI67, OCT4, and MUC5AC via HIF1A Upregulation in Human Bronchial Epithelial Cells

“…It is noteworthy that HIF1α and BMP4 have both been reported not only to promote MUC5AC expression, but also to upregulate expression of OCT4 ( Qi et al, 2004 ; Zhang K. et al, 2010 ; Chen et al, 2011 ; Mathieu et al, 2011 ; Iida et al, 2012 ; Li et al, 2016 ; Liu et al, 2020 ), which is a pluripotency gene and a marker for lung and cancer stem cells ( Ling et al, 2006 ; Chen Y. et al, 2007 ; Chen et al, 2008 ; Gonzalez et al, 2009 ; Karoubi et al, 2009 ; Chiou et al, 2010 ; Zhang X. et al, 2010 ; Khatri et al, 2012 ; Bora-Singhal et al, 2015 ; Jen et al, 2017 ). In fact, our immunostaining and qPCR analyses revealed that, in the submerged cultures, the significant increases in both the protein and mRNA levels of OCT4 by intermittent H/R in both the NHBE and DHBE cells and by consecutive hypoxia in the DHBE cells were concordant with the significant increases in both the protein and mRNA levels of HIF1A and BMP4 ( Figures 9I−K , 10A , and data not shown).…”

A protocol for transdifferentiation of human cardiac fibroblasts into endothelial cells via activation of innate immunity

“…This step sorts the iECs from the transdifferentiation protocol. CD144, also known as VE-Cadherin has been widely used for the isolation of ECs derived from embryonic stem cells (ESCs) and iPSCs ( Liu et al., 2020 ; Sayed et al., 2020 ; Sayed et al., 2015 ). Aspirate the spent medium and dissociate the cells using 1 mL TrypLE express per well for 10 mins at 37°C.…”

“…CD31 has been primarily used to demonstrate the presence of ECs. Similarly, CD144, also known as vascular endothelial cadherin (VE-cadherin) is a marker for ECs ( Liu et al., 2020 ; Sayed et al., 2015 ). RNA extraction Dissociate iECs from 1-well of a 6-well plate using 1 mL TrypLE for 5 min at 37°C.…”

“…Our protocol produces bona fide human ECs with 95%–98% purity by first passage. For complete details on the use and execution of this protocol, please refer to Liu et al. (2020) and Sayed et al.…”