Abstract
Background:
Lymphatic endothelial cells (LECs) derived from lymphatic malformations (LMs) bear activated PIK3CA alleles yet display an inflammatory gene expression profile. A basis for the inflammatory phenotype was sought by screening for coexisting somatic mutations.
Methods and results:
Fourteen independent LEC populations bearing activated PIK3CA alleles were isolated from LM. These were characterized by the expression of growth and inflammatory genes (VEGFC, IL-6, COX-2, IL-8, HO-1, E-SEL) by qRT-PCR. Most commonly upregulated gene products were VEGFC, COX2, HO-1, and ANGPTL4. The specific inhibition of PI3K reduced VEGFC expression without resolving inflammation. Whole exome sequencing of six LM-LEC populations identified five novel somatically acquired alleles coexisting with activated PIK3CA alleles. Two affected genes regulate lipid droplet metabolism (FITM2 and ATG2A), two are gene regulators (MTA1 and TAF1L), and the fifth is an isoform of ANK3 (an endosomal/lysosomal protein). Inhibition of AMPK implicated its involvement in regulating COX-2 and HO-1 overexpression. ANGPTL4 expression was independent of AMPK and PI3K activity and reflected lipid stress demonstrated in normal LECs. AMPK activation with AICAR had a selective growth-limiting effect in a subset of LM-LEC isolates.
Conclusions:
Inflammatory stress displayed by LM-LECs is consistent with errors in lipid metabolism that may be linked to acquired mutations. The acquisition of PIK3CA alleles may be a permissive event that antagonizes inflammation and metabolic defect.
Keywords:
PIK3CA; endothelial cells; lymphatic malformations.
MeSH terms
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AMP-Activated Protein Kinases / genetics
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AMP-Activated Protein Kinases / metabolism
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Alleles*
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Aminoimidazole Carboxamide / analogs & derivatives
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Aminoimidazole Carboxamide / pharmacology
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Angiopoietin-Like Protein 4 / genetics
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Angiopoietin-Like Protein 4 / metabolism
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Ankyrins / genetics
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Ankyrins / metabolism
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Autophagy-Related Proteins / genetics
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Autophagy-Related Proteins / metabolism
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Cell Line
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Class I Phosphatidylinositol 3-Kinases / genetics*
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Class I Phosphatidylinositol 3-Kinases / metabolism
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Cyclooxygenase 2 / genetics
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Cyclooxygenase 2 / metabolism
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E-Selectin / genetics
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E-Selectin / metabolism
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Endothelial Cells / drug effects
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Endothelial Cells / metabolism*
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Endothelial Cells / pathology
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Gene Expression Regulation
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Heme Oxygenase-1 / genetics
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Heme Oxygenase-1 / metabolism
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Histone Deacetylases / genetics
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Histone Deacetylases / metabolism
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Humans
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Interleukin-6 / genetics
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Interleukin-6 / metabolism
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Interleukin-8 / genetics
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Interleukin-8 / metabolism
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Lipid Metabolism / genetics
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Lymphatic Abnormalities / genetics*
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Lymphatic Abnormalities / metabolism
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Lymphatic Abnormalities / pathology
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Lymphoid Tissue / metabolism*
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Lymphoid Tissue / pathology
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Membrane Proteins / genetics
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Membrane Proteins / metabolism
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Mutation
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Primary Cell Culture
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Repressor Proteins / genetics
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Repressor Proteins / metabolism
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Ribonucleotides / pharmacology
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Signal Transduction
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Trans-Activators
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Transcription Factors / genetics
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Transcription Factors / metabolism
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Vascular Endothelial Growth Factor C / genetics
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Vascular Endothelial Growth Factor C / metabolism
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Vesicular Transport Proteins / genetics
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Vesicular Transport Proteins / metabolism
Substances
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ANGPTL4 protein, human
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ANK3 protein, human
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ATG2B protein, human
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Angiopoietin-Like Protein 4
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Ankyrins
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Autophagy-Related Proteins
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E-Selectin
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FITM2 protein, human
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IL6 protein, human
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Interleukin-6
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Interleukin-8
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Membrane Proteins
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MTA1 protein, human
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Repressor Proteins
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Ribonucleotides
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SELE protein, human
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TAF1L protein, human
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Trans-Activators
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Transcription Factors
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VEGFC protein, human
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Vascular Endothelial Growth Factor C
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Vesicular Transport Proteins
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Aminoimidazole Carboxamide
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HMOX1 protein, human
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Heme Oxygenase-1
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Cyclooxygenase 2
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PTGS2 protein, human
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Class I Phosphatidylinositol 3-Kinases
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PIK3CA protein, human
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AMP-Activated Protein Kinases
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Histone Deacetylases
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AICA ribonucleotide