Metabolic pathway alterations in microvascular endothelial cells in response to hypoxia
Fig 1
Hypoxia alters the metabolite profile of microvascular cells.
(A) Lysates were prepared from HMEC-1 cells exposed to a time course of hypoxia and immunoblotted for HIF1α expression. Relative HIF1α expression was calculated using LI-COR Image Studio and is expressed as fold change relative to time = 0 sample. Representative of N = 1. (B) mRNA abundance of HIF1α target genes in HMEC-1 cells at 48 hrs of normoxia or hypoxia exposure was measured by quantitative real-time polymerase chain reaction (qRT-PCR) and is expressed as log(2) fold change relative to HMEC-1 cells cultured in normoxia, N = 3 biological replicates. Significance by two-way ANOVA. (C) Fold change metabolite abundance over normoxia control for 178 metabolites detected in all samples as measured by LC-MS/MS in HMEC-1 cells grown in hypoxia for 48 hrs. Number indicates biological replicate, letter represents technical replicate. (D) Effect size and significance by t-test of metabolite changes measured in C. Colored circles indicate p < 0.05 and fold change > 1.5. Top five most significant metabolites over 1.5-fold change are labeled. Colors indicate pathways: green, amino acid metabolism; orange, nucleotide metabolism; pink, sugar and energy metabolism; blue, other. (E) Fold change relative to normoxia of top five most significantly altered metabolites with over 1.5-fold change after 48 hrs in hypoxia. Different symbols represent biological replicates. Significance by unpaired t-test. All error bars represent SEM. *, p < 0.05; ****, p < 0.0001.