Differential Expression of miRNAs in Hypoxia (“HypoxamiRs”) in Three Canine High-Grade Glioma Cell Lines

Abstract

Dogs with spontaneous high-grade gliomas increasingly are being proposed as useful large animal pre-clinical models for the human disease. Hypoxia is a critical microenvironmental condition that is common in both canine and human high-grade gliomas and drives increased angiogenesis, chemo-and radioresistance, and acquisition of a stem-like phenotype. Some of this effect is mediated by the hypoxia-induced expression of microRNAs, small (∼22 nucleotides long), non-coding RNAs that can modulate gene expression through interference with mRNA translation. Using an in vitro model with three canine high-grade glioma cell lines (J3T, SDT3G, and G06A) exposed to 72 h of 1.5% oxygen vs. standard 20% oxygen, we examined the global "hypoxamiR" profile using small RNA-Seq and performed pathway analysis for targeted genes using both Panther and NetworkAnalyst. Important pathways include many that are well-established as being important in glioma biology, general cancer biology, hypoxia, angiogenesis, immunology, and stem-ness, among others. This work provides the first examination of the effect of hypoxia on miRNA expression in the context of canine glioma, and highlights important similarities with the human disease.

Establishment and characterization of two novel patient‐derived lines from canine high‐grade glioma

“…In order to assess the growth of our newly established patient derived lines, we sought to compare them to the canine SDT‐3G, G06‐A, (both high‐grade astrocytoma) and J3T‐Bg (anaplastic astrocytoma) cell lines which are commonly used by the veterinary community 16,19,20 . We assessed in vitro cell growth using the IncuCyte live cell imaging system to capture images of cells over time and calculate growth as a measure of confluence, normalized to the first time point (Figures 4A and S1c).…”

“…In order to assess the growth of our newly established patient derived lines, we sought to compare them to the canine SDT-3G, G06-A, (both high-grade astrocytoma) and J3T-Bg (anaplastic astrocytoma) cell lines which are commonly used by the veterinary community. 16,19,20 We assessed in vitro cell growth using the IncuCyte live cell imaging system to capture images of cells over time and calculate growth as a measure of confluence, normalized to the first time point (Figures 4A and S1c). The novel patient-derived lines Mo-HO and Boo-HA exhibit the slowest growth, with doubling times of 128 and 42 h respectively, whereas SDT-3G, J3T-Bg and G06-A are faster growing with doubling times of 36, 22, and 18 h, respectively.…”

Hypoxia inducible factors inhibit respiratory syncytial virus infection by modulation of nucleolin expression

“…An earlier study reported that NCL transcripts were regulated by microRNA-494 (miR-494) 45 . Notably, miR-494 has a HRE in its promoter ( Supplementary Fig.9 ) and was previously reported to be induced under hypoxic conditions 46,47 . To examine this hypothesis, we measured miR-494 levels in Calu-3 cells cultured at 1% oxygen or treated with Daprodustat and found a robust induction ( Fig.4e ).…”

Mitochondrial Genetic and Epigenetic Regulations in Cancer: Therapeutic Potential

“…Overexpression of miR-210 was documented in pancreatic, head and neck, breast and lung cancers [ 266 ]. Similar to miR-210, miR-323 is upregulated in hypoxia, and its low levels are connected to longer survival in human glioblastoma patients [ 267 ]. Although miR-137 functions as a tumour suppressor [ 268 ], it also plays a role in hypoxia and inhibition of ROS accumulation; miR-137 directly downregulates the expression of the hypoxia-mediated mitophagy receptors FUNDC1 and NIX, leading to suppression of apoptosis and mitophagy [ 253 ].…”

“…MiR-216b inhibits proliferation and cell invasion in several types of cancer and is downregulated in hypoxia. Low levels of this miRNA were associated with poor prognosis [ 267 ].…”