ABSTRACT
The bioactive form of vitamin D [1,25(OH)2D3] regulates mineral and bone homeostasis and exerts potent anti-inflammatory and antiproliferative properties through binding to the vitamin D receptor (VDR). The 3D structures of the VDR ligand-binding domain with 1,25(OH)2D3 or gemini analogs unveiled the molecular mechanism underlying ligand recognition. On the basis of structure-function correlations, we generated a point-mutated VDR (VDR(gem)) that is unresponsive to 1,25(OH)2D3, but the activity of which is efficiently induced by the gemini ligands. Moreover, we show that many VDR target genes are repressed by unliganded VDR(gem) and that mineral ion and bone homeostasis are more impaired in VDR(gem) mice than in VDR null mice, demonstrating that mutations abolishing VDR ligand binding result in more severe skeletal defects than VDR null mutations. As gemini ligands induce VDR(gem) transcriptional activity in mice and normalize their serum calcium levels, VDR(gem) is a powerful tool to further unravel both liganded and unliganded VDR signaling.
Subject(s)
Receptors, Calcitriol/chemistry , Receptors, Calcitriol/metabolism , Animals , Chromatin Immunoprecipitation , Crystallography, X-Ray , Fluorescence Polarization , Genotype , HEK293 Cells , Humans , MCF-7 Cells , Mice , Mice, Knockout , Mutation/genetics , Protein Binding/genetics , Receptors, Calcitriol/genetics , Spectrometry, Mass, Electrospray Ionization , Vitamin D/metabolismABSTRACT
Ageing is associated with skeletal muscle impairment. Changes in mitochondrial homeostasis are thought to play a key role in this process. This study examined whether chronic intake of polyphenols (PPs), which are known to be modulators of oxidative stress, might prevent the age-related decline of mitochondrial functions in skeletal muscle. Three groups of 10 Wistar rats were investigated. Rats aged 16 weeks were compared with rats aged 40 weeks that were given 75 mg kg(-1) day(-1) PPs or solvent in the drinking water starting at week 16. Mitochondrial respiratory chain complex activities were measured in saponin-skinned fibres of soleus muscles using glutamate-malate (V(max)), succinate (V(succ)) and N,N,N',N'-tetramethyl-p-phenylenediamine dihydrochloride-ascorbate (V(TMPD)). Production of reactive oxygen species was assessed using dihydroethidium staining. Transcript levels of genes involved in antioxidant defence were determined using RT-PCR. Ageing reduced muscle V(max) (from 8.8 ± 0.45 to 6.17 ± 0.51 µmol O(2) min(-1) g(-1), -30.5%, P < 0.01), V(TMPD) (from 20.67 ± 1.24 to 16.55 ± 1.16 µmol O(2) min(-1) g(-1), -19.9%, P < 0.05), increased production of reactive oxygen species (from 100 ± 9.9 to 351.1 ± 31.7%) and decreased transcripts of mitochondrial superoxide dismutase 2 (-59.3%, P < 0.01), peroxisome proliferator-activated receptor γ coactivator-1ß (PGC-1ß; -61.5%, P < 0.05) and sirtuin 1 (-54.2%, P < 0.05). Chronic PP intake normalized V(max) (8.63 ± 0.63 µmol O(2) min(-1) g(-1)), decreased production of reactive oxygen species (141.7 ± 16.7%, P < 0.001) and enhanced antioxidant defence (superoxide dismutase 2 expression, +151.3%, P < 0.05) and PGC-1ß expression (+185.7%, P < 0.05) in comparison to age-matched untreated rats. The present data indicate that regular intake of PPs starting at a young age prevents age-related mitochondrial respiratory impairment in skeletal muscle, probably through decreased oxidative stress and enhancement of PGC-1ß expression.
