Decreased mitochondrial function in UVA-irradiated dermal fibroblasts causes the insufficient formation of type I collagen and fibrillin-1 fibers.

BACKGROUND: Decreases of collagen fibers and the disappearance of oxytalan fibers are typical symptoms of photoaged skin. Although a low quality of mitochondria (MT) in photoaged skin cells has been observed, it is unknown whether the decreased quality of MT is responsible for the insufficient formation of dermal fibers. OBJECTIVE: To identify the role of mitochondrial quality in skin photoaging focusing on the formation of dermal fibers. METHODS: Type I collagen and fibrillin-1 fibers in normal human dermal fibroblasts (NHDFs) were observed by immunostaining. Type I collagen and fibrillin-1 proteins in NHDFs were quantified by ELISA. Mitochondrial quality was evaluated by measuring levels of intracellular ATP and MITOL, which regulate mitochondrial quality. RESULTS: UVA-irradiated NHDFs formed insufficient type I collagen and fibrillin-1 fibers and had a decreased ratio of extracellular versus intracellular levels of those proteins. Although expression levels of motor proteins that transport those proteins intracellularly were not affected by UVA, intracellular ATP levels, which is the driving force of motor proteins, were decreased by UVA along with decreased MITOL protein. Knockdown of MITOL in NHDFs decreased the level of intracellular ATP and caused the insufficient formation of type I collagen and fibrillin-1 fibers due to interfering with the secretion of those proteins. CONCLUSION: These results indicate that a low quality of MT with ATP depletion in dermal fibroblasts caused by irradiation with UVA induces the insufficient formation of type I collagen and fibrillin-1 fibers due to the decreased extracellular secretion of those proteins.

Pyridoxine (VB6 ) restores the down-regulation of serine palmitoyltransferase mRNA expression in keratinocytes cultured in highly oxidative conditions through enhancement of the intracellular antioxidant system.

BACKGROUND: Pyridoxine (VB6 ), which acts as a coenzyme in the biosynthesis of niacin, is formulated in pharmaceuticals to treat skin roughness. However, the mechanism of action of VB6 is not known precisely. OBJECTIVE: This study was conducted to clarify the influence of highly oxidative conditions on the expression of skin moisture-related mRNAs and to evaluate the preventive effects of VB6 focusing on antioxidant behaviour. METHODS: Intracellular levels of reactive oxygen species (ROS) in normal human epidermal keratinocytes (NHEKs) were determined using the 2',7'-dichlorofluorescein diacetate assay. Real-time PCR was employed to investigate the influence of higher oxidative conditions on the expression of mRNAs encoding serine palmitoyl transferase (SPT) and filaggrin, and to characterize the mechanism of the antioxidant effect of VB6 . Intracellular glutathione was quantified using an assay based on the glutathione recycling system with 5,5'-dithiobis (2-nitrobenzoic acid) reagent and glutathione reductase. Carbonylated proteins (CPs) were semi-quantified by detecting aldehyde residues. RESULTS: Treatment of NHEKs with BSO increased the level of intracellular CPs by interfering with intracellular glutathione synthesis. Further, treatment with BSO down-regulated the expression level of SPT mRNA, but VB6 restored SPT mRNA expression in BSO-treated NHEKs. VB6 decreased the level of intracellular CPs with or without BSO treatment in a dose-dependent manner. In addition, VB6 increased levels of intracellular NADH/NADPH and glutathione through the activation of nuclear factor E2-related factor 2 (Nrf2) signalling. CONCLUSION: These results suggest that highly oxidative conditions cause an impaired skin barrier function due to the down-regulation of SPT that results in skin roughness. VB6 improved the down-regulation of SPT mRNA expression initiated by highly oxidative conditions by enhancing the intracellular antioxidant system.