Toll-Like Receptor Signalling Pathways Regulate Hypoxic Stress Induced Fibroblast Growth Factor but Not Vascular Endothelial Growth Factor-A in Human Microvascular Endothelial Cells

Akhtar, Rukhsar; Tahir, Husain; Stewart, Elizabeth; Wei, Ruoxin; Mohammed, Imran; Amoaku, Winfried M.

Abstract

Retinal diseases are the leading causes of irreversible blindness worldwide. The role of toll-like receptor (TLR) signalling mechanisms (MyD88 and TRIF) in the production of pro-angiogenic growth factors from human microvascular endothelial cells (HMEC-1) under hypoxic stress remains unexplored. HMEC-1 was incubated under normoxic (5% CO2 at 37 °C) and hypoxic (1% O2, 5% CO2, and 94% N2; at 37 °C) conditions for 2, 6, 24, and 48 h, respectively. For TLR pathway analysis, HMEC-1 was pre-treated with pharmacological inhibitors (Pepinh-MyD88 and Pepinh-TRIF) and subjected to normoxia and hypoxia conditions. Gene and protein expressions of vascular endothelial growth factor-A (VEGF-A), fibroblast growth factor (FGF-2), hypoxia inducible factor 1-alpha (HIF1-α) were performed using quantitative polymerase chain reaction (qPCR), ELISA, and Western blot methodologies. Levels of TLR3 and TLR4 were analysed by flow cytometry. Under hypoxia, levels of VEGF-A and FGF-2 were elevated in a time-dependent fashion. Inhibition of MyD88 and TRIF signalling pathways decreased FGF-2 levels but failed to modulate the secretion of VEGF-A from HMEC-1. Blocking a known regulator, endothelin receptor (ETR), also had no effect on VEGF-A secretion from HMEC-1. Overall, this study provides the proof-of-concept to target TLR signalling pathways for the management of blinding retinal diseases.

Citation

Akhtar, R., Tahir, H., Stewart, E., Wei, R., Mohammed, I., & Amoaku, W. M. (2021). Toll-Like Receptor Signalling Pathways Regulate Hypoxic Stress Induced Fibroblast Growth Factor but Not Vascular Endothelial Growth Factor-A in Human Microvascular Endothelial Cells. International Journal of Translational Medicine, 1(1), 25-38. https://doi.org/10.3390/ijtm1010003

• Journal Article Type: Article
• Acceptance Date: May 12, 2021
• Online Publication Date: May 27, 2021
• Publication Date: 2021-06
• Deposit Date: Nov 12, 2021
• Publicly Available Date: Nov 17, 2021
• Journal: International Journal of Translational Medicine
• Print ISSN: 2673-8937
• Electronic ISSN: 2673-8937
• Publisher: MDPI AG
• Peer Reviewed: Peer Reviewed
• Volume: 1
• Issue: 1
• Pages: 25-38
• DOI: https://doi.org/10.3390/ijtm1010003
• Public URL: https://nottingham-repository.worktribe.com/output/5619771
• Publisher URL: https://www.mdpi.com/2673-8937/1/1/3