Fig 6.

Oligodendrocytes (OLs)-specific Dicer ablation triggers lipid accumulation and peroxisomal β-oxidation malfunction in the brain. (A) ELOVL7 (elongation of very long chain fatty acids protein 7) overexpression resulted in lipid accumulation in the absence of miR-219. Addition of miR-219 triggered a dramatic reduction of lipid droplets in the cells transfected with wild-type (WT) but not mutated 3-prime untranslated region (3′UTR). Scale bar represents 1cm. (B) Quantification of the accumulated lipid in WT and mutant (MT) ELOVL7 constructs in adipocyte cells. The dye was extracted, and absorbance was measured at 492nm. p Values were calculated by an unpaired Student t test. Data are expressed as means ± standard deviation. (C) Brain sections were stained with Oil Red-O and showed much more lipid accumulation in MT than WT littermate mice. Scale bar represents 500μm. (D) Western blotting showed reduced acylcoenzyme A oxidase 1 (ACOX1) levels in older mutant mice indicating that ACOX1 is not a direct target of miR-219. (E) An assay for peroxisomal β-oxidation showed less activity in brain extracts from MT mice versus WT. p Values were calculated using an unpaired Student t test. Data are expressed as means ± standard error of mean. (F) A model of downstream effects caused by postnatal OL-specific Dicer ablation. Mature OL-specific Dicer ablation suppresses the generation of miR-32, miR-144, and miR-219. Myelin-enriched ELOVL7 synthesizes more lipids such as very long chain of fatty acids (VLCFAs), resulting in demyelination and oxidative damage. In parallel, other unknown proteins targeted by these micro-RNAS (miRNAs) may regulate peroxisomal function, also contributing to lipid accumulation, oxidative damage, and ultimately, inflammatory gliosis and neurodegeneration in OL-specific Dicer mutant mice. PLP = proteolipid protein; GAPDH = glyceraldehyde phosphate dehydrogenase.