Genome-wide association studies in the Japanese population identified genetic loci and target gene associated with epidermal turnover.

The epidermis is an essential organ for life by retaining water and as a protective barrier. The epidermis is maintained through metabolism, in which basal cells produced from epidermal stem cells differentiate into spinous cells, granular cells and corneocytes, and are finally shed from the epidermal surface. This is epidermal turnover, and with aging, there is a decline in epidermis function. Other factors that may affect epidermal turnover include ultraviolet damage and genetic factors. These genetic factors are of particular interest as little is known. Although recent skin-focused genome-wide association studies (GWAS) have been conducted, the genetic regions associated with epidermal turnover are almost uninvestigated. Therefore, we conducted a GWAS on epidermal turnover in the Japanese population, using the corneocyte area, which correlates to the rate of epidermal turnover, as an indicator. As a result, rs2278431 (p = 1.29 × 10-7 ) in 19q13.2 was associated with corneocyte size. Furthermore, eQTL analysis suggested that rs2278431 was related to the SPINT2 gene. In addition, SPINT2 knockdown studies using epidermal keratinocytes revealed that SPINT2 is involved in keratinocyte proliferation and in corneocyte size regulation in reconstructed epidermis. These results suggest that rs2278431 is involved in the expression of SPINT2 and affects epidermal turnover.

Identification of mitochondrial genetic variants associated with human corneocyte size in Japanese women.

Mitochondria have their own DNA (mtDNA). Genetic variants are likely to accumulate in mtDNA, and its base substitution rate is known to be very fast, 10-20 times faster than that of nuclear DNA. For this reason, mtSNPs (mitochondrial genome single nucleotide polymorphisms) are frequently detected in mtDNA. Several thousands of copies of mtDNA are considered to be present in a cell, and variants that have occurred in mtDNA are expected to markedly affect the intracellular energy production system and ROS (reactive oxygen species) kinetics. Therefore, recently, mtSNPs have come to be considered very important as a determinant of the individual constitution such as the life-span and disease susceptibility. In this study, we searched for mtSNPs that affect the individual corneocyte size using samples from 358 Japanese women. As a result, mtSNPs 10609C and 12406A were found to be significantly related to the corneocyte size in the outermost layer of the epidermis. There have been a large number of reports concerning the association between mtSNPs and individual constitution, but little evaluation of their relationships with epidermal properties has been made. The results of the present study first suggested that mtSNPs may affect the epidermal properties in Japanese women.

Analysis of the effect of daily stress on the skin and search for genetic loci involved in the perceived stress of an individual.

Background: Stress may have various effects on our bodies. In particular, the skin may be readily influenced by stress. In addition, there are individual differences in the way we feel stress, suggesting the involvement of genetic factors in such individual differences. Objectives: In this study, we analysed the influence of stress on skin condition and ageing involving Japanese females, and investigated single nucleotide polymorphisms (SNPs) that influence perceived stress of an individual. Methods: We collected genotype data from 1200 Japanese females. At the same time, a questionnaire was conducted on the degree of stress that each subject feels on a daily basis and the current skin condition. We analysed the effects of stress on skin condition and searched for SNPs related to individual stress susceptibility by genome-wide association studies. Results: Our data suggested that stress influences skin condition and ageing, as previously reported. And, we found rs74548608 as a SNP that affects perceived stress of an individual. This SNP is located on the upstream of Patched-1, which is a gene that functions as a sonic hedgehog receptor. Conclusions: Our study has identified new genetic factors for perceived stress of an individual in the Japanese female. The SNP found in this study may be a candidate factor important for understanding the perceived stress of an individual of Japanese.

Increase in inhibin beta A/Activin-A expression in the human epidermis and the suppression of epidermal stem/progenitor cell proliferation with aging.

BACKGROUND: Age-related thinning and reduced cell proliferation in the human epidermis are associated with the accumulation of senescent cells and decreases in the number and function of epidermal stem cells. OBJECTIVE: This study examined the expression of INHBA/Activin-A in human epidermis and expression differences with age, and the effect of Activin-A on epidermal stem/progenitor cells. METHODS: Immunohistochemical staining was used to analyze age-related changes in the expression of INHBA/Activin-A in the epidermal tissue of young and old subjects. Epidermal INHBA/Activin-A expression levels, epidermal morphology, and the number of epidermal stem/progenitor cells or proliferating cells were investigated using older abdominal skin samples. The effects of Activin-A on the development of a three-dimensional (3D) reconstructed epidermis and cell proliferation were also assessed. RESULTS: INHBA/Activin-A expression levels in the human epidermis increased with age, although they varied among individuals. In the epidermis of older abdominal skin samples, INHBA/Activin-A expression levels negatively correlated with epidermal thickness, the rete ridge depth and the interdigitation index. The proportion of epidermal stem/progenitor cells and proliferating cells decreased with increases in INHBA/Activin-A expression levels. Activin-A had no effect on the differentiation of keratinocytes in the 3D-reconstructed epidermis; however, thinning of the 3D epidermis was noted. Moreover, the addition of Activin-A inhibited the proliferation of epidermal stem/progenitor cells in a concentration-dependent manner. CONCLUSIONS: Age-related increased in INHBA/Activin-A expression levels were observed in the human epidermis, and may contribute to epidermal thinning and decreases in the number of epidermal stem/progenitor cells and proliferative activity.

Genome-wide association studies in Japanese women identified genetic loci associated with wrinkles and sagging.

Wrinkles and sagging are caused by various factors, such as ultraviolet rays; however, recent findings demonstrated that some individuals are genetically predisposed to these phenotypes of skin aging. The contribution of single nucleotide polymorphisms (SNPs) to the development of wrinkles and sagging has been demonstrated in genome-wide association studies (GWAS). However, these findings were mainly obtained from European and Chinese populations. Limited information is currently available on the involvement of SNPs in the development of wrinkles and sagging in a Japanese population. Therefore, we herein performed GWAS on wrinkles at the outer corners of the eyes and nasolabial folds in 1041 Japanese women. The results obtained revealed that 5 SNPs (19p13.2: rs2303098 (p = 3.39 × 10-8 ), rs56391955 (p = 3.39 × 10-8 ), rs67560822 (p = 3.50 × 10-8 ), rs889126 (p = 3.78 × 10-8 ), rs57490083 (p = 3.99 × 10-8 )) located within the COL5A3 gene associated with wrinkles at the outer corners of the eyes. Regarding nasolabial folds, 8q24.11 (rs4876369; p = 1.05 × 10-7 , rs6980503; p = 1.25 × 10-7 , rs61027543; p = 1.25 × 10-7 , rs16889363; p = 1.38 × 10-7 ) was suggested to be associated with RAD21 gene expression. These SNPs have not been reported in other populations, and were first found in Japanese women population. These SNPs may be used as markers to examine the genetic predisposition of individuals to wrinkles and sagging.

Establishment of Three Types of Immortalized Human Skin Stem Cell Lines Derived from the Single Donor.

Currently, human-skin derived cell culture is a basic technique essential for dermatological research, cellular engineering research, drug development, and cosmetic development. But the number of donors is limited, and primary cell function reduces through cell passage. In particular, since adult stem cells are present in a small amount in living tissues, it has been difficult to obtain a large amount of stem cells and to stably culture them. In this study, skin derived cells were isolated from the epidermis, dermis, and adipose tissue collected from single donor, and immortalization was induced through gene transfer. Subsequently, cell lines that could be used as stem cell models were selected using the differentiation potential and the expression of stem cell markers as indices, and it was confirmed that these could be stably cultured. The immortalized cell lines established in this study have the potential to be applied not only to basic dermatological research but also to a wide range of fields such as drug screening and cell engineering.

Search for genetic loci involved in the constitution and skin type of a Japanese women using a genome-wide association study.

The constitution and skin type of individuals are influenced by various factors. Recently, the influence of genetic predispositions on these has been emphasized. To date, genome-wide association studies (GWAS) have shown several single nucleotide polymorphisms (SNPs) that affect individual's constitution and skin type. However, these studies have mainly focused on the Caucasian population, and only a few association analyses with the constitution and skin type of individuals involving a Japanese population have been conducted. In this study, we conducted a GWAS analysis of 9 phenotypes regarding the constitution or skin type of 1108 Japanese women based on a questionnaire. As a result, in addition to SNPs known to be involved in phenotypes in the past, we discovered new SNPs and genetic regions related to darkness of pigmented spots, skin flushing, frequency of rough skin and responsiveness to cosmetics.

Titanium(IV) chloride-mediated ring cleavage and Michael addition of 3,3-dialkylcyclobutanones and 3-[(trimethylsilyl)methyl]-cyclobutanones.

ß'-Chloro and ß',γ'-unsaturated trichlorotitanium enolates, which were formed in situ by titanium(IV) chloride-mediated ring cleavage of 3,3-dialkylcyclobutanones and 3-[(trimethylsilyl)methyl]cyclobutanones, reacted with enones to give Michael adducts with keeping a labile ß'-chloro or ß',γ'-unsaturated group.

Ring cleavage and successive aldol reaction of 3-[(trialkylsilyl)methyl]cyclobutanones.

3-[(Trialkylsilyl)methyl]cyclobutanones reacted with aldehydes by activation with titanium(IV) chloride to give acyclic ß,γ-unsaturated ß'-hydroxyketones.

Titanium(IV) chloride-mediated carbon-carbon bond-forming reaction between 3,3-dialkylcyclobutanones and aldehydes.

Treatment of 3,3-dialkylcyclobutanones with titanium(IV) chloride in the presence of aldehyde gave beta'-chloro-beta-hydroxy ketones in high yields. It was speculated that ring cleavage of the cyclobutanone ring with titanium(IV) chloride gave trichlorotitanium enolate having a tertiary alkyl chloride moiety and then aldol reaction of the titanium enolate proceeded. A trialkylsilylmethyl group at the 2-position of cyclobutanone facilitated the ring cleavage. Synthesis of substituted cyclopentenone from an obtained product is also described.