Microbiota in Umbilical Dirt and Its Relationship with Odor.

The umbilicus accumulates more dirt than other body surfaces and is difficult to clean. Hygiene in this area is vital, particularly for surgery, because of its proximity to the laparotomy site. Although microorganisms in the umbilicus have been extensively examined, those in umbilical dirt have not due to the lack of an efficient method of collection. We previously established a technique to extract umbilical dirt using the anchor effect of polymers, which are injected into the umbilicus. In the present study, we applied this technique for the first time to investigate umbilical dirt. The results obtained revealed an abundance of Corynebacterium among various bacteria, whereas Cutibacterium and Staphylococcus, which are abundant at other skin sites, were rare. The relationships between the microbiota and issues related to the umbilicus were investigated and some covariates, including the odor score and several bacteria, were identified. A detailed ana-lysis of the genera associated with odor revealed no correlation with Corynebacterium; however, some minor anaerobic bacteria, such as Mobiluncus, Arcanobacterium, and Peptoniphilus, were more abundant in the high odor score group. Therefore, this technique to collect umbilical dirt provided insights into the microbiota in umbilical dirt and suggested functions for minor anaerobes. Furthermore, since various pathogenic microorganisms were detected, their control may contribute to the prevention of both odor production and infectious diseases caused by these microorganisms.

Assessment of Multidimensional Health Care Parameters Among Adults in Japan for Developing a Virtual Human Generative Model: Protocol for a Cross-sectional Study.

BACKGROUND: Human health status can be measured on the basis of many different parameters. Statistical relationships among these different health parameters will enable several possible health care applications and an approximation of the current health status of individuals, which will allow for more personalized and preventive health care by informing the potential risks and developing personalized interventions. Furthermore, a better understanding of the modifiable risk factors related to lifestyle, diet, and physical activity will facilitate the design of optimal treatment approaches for individuals. OBJECTIVE: This study aims to provide a high-dimensional, cross-sectional data set of comprehensive health care information to construct a combined statistical model as a single joint probability distribution and enable further studies on individual relationships among the multidimensional data obtained. METHODS: In this cross-sectional observational study, data were collected from a population of 1000 adult men and women (aged ≥20 years) matching the age ratio of the typical adult Japanese population. Data include biochemical and metabolic profiles from blood, urine, saliva, and oral glucose tolerance tests; bacterial profiles from feces, facial skin, scalp skin, and saliva; messenger RNA, proteome, and metabolite analyses of facial and scalp skin surface lipids; lifestyle surveys and questionnaires; physical, motor, cognitive, and vascular function analyses; alopecia analysis; and comprehensive analyses of body odor components. Statistical analyses will be performed in 2 modes: one to train a joint probability distribution by combining a commercially available health care data set containing large amounts of relatively low-dimensional data with the cross-sectional data set described in this paper and another to individually investigate the relationships among the variables obtained in this study. RESULTS: Recruitment for this study started in October 2021 and ended in February 2022, with a total of 997 participants enrolled. The collected data will be used to build a joint probability distribution called a Virtual Human Generative Model. Both the model and the collected data are expected to provide information on the relationships between various health statuses. CONCLUSIONS: As different degrees of health status correlations are expected to differentially affect individual health status, this study will contribute to the development of empirically justified interventions based on the population. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/47024.

The ceramide [NP]/[NS] ratio in the stratum corneum is a potential marker for skin properties and epidermal differentiation.

BACKGROUND: Specific species of ceramides (Cer), major constituents of lipids in the stratum corneum (SC), are decreased and are correlated with SC barrier and water-holding functions in the skin of patients with atopic dermatitis (AD) or psoriasis (Pso). However, possible correlations between Cer subclass ratios and skin properties in barrier-disrupted skin and in healthy skin remain unclear. The objective of this study was to identify a new marker to evaluate skin properties and epidermal differentiation in SC not only in barrier-disrupted skin but also in healthy skin. METHODS: The Cer subclass ratios in the SC of healthy control subjects and in patients with AD or Pso were evaluated. Correlations with candidate markers and facial skin features of healthy Japanese females (20-74 years old, n = 210) were investigated. Variations of markers during epidermal differentiation were studied in human epidermis and in cultured keratinocytes. RESULTS: The ratios of Cer [NP]/[NS], Cer [NH]/[NS], Cer [NP]/[AS], Cer [NH]/[NS], Cer [NDS]/[AS], Cer [AH]/[AS] and Cer [EOP]/[AS] showed significant differences between non-lesional skin of AD patients and normal skin of healthy control subjects, as well as Pso patients and their healthy control subjects. The Cer [NP]/[NS] ratio was correlated with SC functional parameters (transepidermal water loss and capacitance) and with skin appearance (texture, scaling and color) even in the cheek skin of healthy female subjects. The Cer [NP]/[NS] ratio in the SC was approximately 18-times higher than in living keratinocytes, and it increased as they differentiated. CONCLUSIONS: The Cer [NP]/[NS] ratio in the SC is a potential marker for skin properties and epidermal differentiation in barrier-disrupted skin as well as in healthy skin.

Development of a cognitive function marker based on D-amino acid proportions using new chiral tandem LC-MS/MS systems.

The incidence of dementia, a clinical symptom characterized by severe cognitive decline, is increasing worldwide. Predictive biomarkers are therefore required for early identification and management. D-amino acids in the brain contribute to cognitive function and are suggested as useful biomarkers for diagnosing dementia risk. To clarify their relationship with human cognitive decline, we developed an identification method of chiral metabolomics for detecting slight differences in chiral amino acid amounts. Chiral tandem liquid chromatography-tandem mass spectrometry systems were applied for sensitive and selective amino acid species along with chiral species determination based on anion and zwitterion exchange mechanisms. In a comprehensive health cohort (cross-sectional study), we measured blood chiral amino acid levels from 305 women (65-80 years old) classified into Control, Mild-cognitive-Impairment (MCI), and Dementia groups using the Mini-Mental State Examination. MCI exhibited higher D-Pro (D-Pro/(D-Pro + L-Pro)) proportion vs the Control group, suggesting this proportion as a useful biomarker for MCI. Biomarker accuracy was improved in combination with D-Ser proportion. Receiver operating characteristics analysis of the Control vs. MCI proportion obtained area under the curve (0.80) with 70% sensitivity and 84% specificity at the optimal cutoff value (0.30). Thus, dementia monitoring can be improved by including trace D-amino acids measurements.

Ethnic differences in stratum corneum functions between Chinese and Thai infants residing in Bangkok, Thailand.

BACKGROUND/OBJECTIVES: Ethnic and racial differences in infant skin have not been well characterized. The purpose of this study was to establish whether there are ethnic differences and similarities in the stratum corneum (SC) functions of Thai and Chinese infants. METHODS: Healthy infants 6 to 24 months of age (N = 60; 30 Thai, 30 Chinese) who resided in Bangkok, Thailand, were enrolled. Transepidermal water loss (TEWL) and SC hydration (capacitance) on the thigh, buttock, and upper arm were measured. Ceramide content was determined in the SC on the upper arm. RESULTS: SC hydration was not remarkably different between the two ethnicities at any site measured, but TEWL was significantly higher in Chinese infants than in Thai infants at all sites. Hydration of the SC was not significantly correlated with age in either ethnicity. TEWL had significant but weak correlations with age on the thigh and upper arm in Thai infants. Ceramide content was significantly higher in Chinese SC than in Thai SC. No relationship between ceramide content and TEWL or hydration was observed in either ethnicity. CONCLUSION: The significant differences in TEWL and ceramide contents between Chinese and Thai infant skin could prove useful in designing skin care and diapering products that are best suited for each ethnicity.

Coffee polyphenols extracted from green coffee beans improve skin properties and microcirculatory function.

Coffee polyphenols (CPPs), including chlorogenic acid, exert various physiological activities. The purpose of this study was to investigate the effects of CPPs on skin properties and microcirculatory function in humans. In this double-blind, placebo-controlled study, 49 female subjects with mildly xerotic skin received either a test beverage containing CPPs (270 mg/100 mL/day) or a placebo beverage for 8 weeks. The ingestion of CPPs significantly lowered the clinical scores for skin dryness, decreased transepidermal water loss, skin surface pH, and increased stratum corneum hydration and the responsiveness of skin blood flow during local warming. Moreover, the amounts of free fatty acids and lactic acid in the stratum corneum significantly increased after the ingestion of CPPs. These results suggest that an 8-week intake of CPPs improve skin permeability barrier function and hydration, with a concomitant improvement in microcirculatory function, leading to efficacy in the alleviation of mildly xerotic skin.

Incomplete KLK7 Secretion and Upregulated LEKTI Expression Underlie Hyperkeratotic Stratum Corneum in Atopic Dermatitis.

Atopic dermatitis (AD) is a common inflammatory skin disorder. Chronic AD lesions present hyperkeratosis, indicating a disturbed desquamation process. KLK7 is a serine protease involved in the proteolysis of extracellular corneodesmosome components, including desmocollin 1 and corneodesmosin, which leads to desquamation. KLK7 is secreted by lamellar granules and upregulated in AD lesional skin. However, despite increased KLK7 protein levels, immunostaining and electron microscopy indicated numerous corneodesmosomes remaining in the uppermost layer of the stratum corneum from AD lesions. We aimed to clarify the discrepancy between KLK7 overexpression and retention of corneodesmosomes on AD corneocytes. Western blot analysis indicated abnormal corneodesmosin degradation patterns in stratum corneum from AD lesions. The KLK activity of tape-stripped corneocytes from AD lesions was not significantly elevated in in situ zymography, which was our new attempt to detect the protease activity more precisely than conventional assays. This ineffective KLK activation was associated with impaired KLK7 secretion from lamellar granules and increased expression of LEKTI in AD. Such imbalances in protease-protease inhibitor interactions could lead to abnormal proteolysis of corneodesmosomes and compact hyperkeratosis. Upregulated expression of LEKTI might be a compensatory mechanism to prevent further barrier dysfunction in AD.

Essential role of the cytochrome P450 CYP4F22 in the production of acylceramide, the key lipid for skin permeability barrier formation.

A skin permeability barrier is essential for terrestrial animals, and its impairment causes several cutaneous disorders such as ichthyosis and atopic dermatitis. Although acylceramide is an important lipid for the skin permeability barrier, details of its production have yet to be determined, leaving the molecular mechanism of skin permeability barrier formation unclear. Here we identified the cytochrome P450 gene CYP4F22 (cytochrome P450, family 4, subfamily F, polypeptide 22) as the long-sought fatty acid ω-hydroxylase gene required for acylceramide production. CYP4F22 has been identified as one of the autosomal recessive congenital ichthyosis-causative genes. Ichthyosis-mutant proteins exhibited reduced enzyme activity, indicating correlation between activity and pathology. Furthermore, lipid analysis of a patient with ichthyosis showed a drastic decrease in acylceramide production. We determined that CYP4F22 was a type I membrane protein that locates in the endoplasmic reticulum (ER), suggesting that the ω-hydroxylation occurs on the cytoplasmic side of the ER. The preferred substrate of the CYP4F22 was fatty acids with a carbon chain length of 28 or more (≥C28). In conclusion, our findings demonstrate that CYP4F22 is an ultra-long-chain fatty acid ω-hydroxylase responsible for acylceramide production and provide important insights into the molecular mechanisms of skin permeability barrier formation. Furthermore, based on the results obtained here, we proposed a detailed reaction series for acylceramide production.

A role of the anteiso branch of 18-MEA in 18-MEA/SPDA to form a persistent hydrophobicity to alkaline-color-treated weathered hair.

The effect of the anteiso-branch moiety of 18-MEA (18-methyleicosanic acid) to create a persistent hydrophobicity of alkaline-color-treated weathered hair treated with 18-MEA/SPDA (stearoxypropyldimethylamine) was investigated by comparing a straight-chain fatty acid (n-heneicosanoic acid, n-HEA) and an iso-branch fatty acid (19-methyleicosanic acid, 19-MEA) with the anteiso-branch fatty acid (18-MEA), using dynamic contact angle measurements, quantification of 18-MEA by LC/MS, and temperature controlled atomic force microscopy (AFM). The dynamic contact angle measurements indicated that the anteiso-branch moiety of 18-MEA is critical for the creation of a persistent hydrophobicity to alkaline-color-treated weathered hair. The temperature-controlled AFM investigations revealed that the anteiso-branch moiety of 18-MEA in the 18-MEA/SPDA system produces a persistent hydrophobicity to alkaline-color-treated weathered hair by providing higher fluidity to the upper region of the 18-MEA/SPDA layer.

Deposition of 18-MEA onto alkaline-color-treated weathered hair to form a persistent hydrophobicity.

A technology for the deposition of a persistent hydrophobicity to alkaline-color-treated weathered hair surfaces using 18-MEA (18-methyleicosanoic acid) is presented. Two approaches were examined in order to make 18-MEA bind tightly to the alkaline-color-treated weathered hair surface. One was to apply 18-MEA as an acid form and the other was to apply 18-MEA as a salt or complex. It was found that the combination of 18-MEA with specific cationic surfactants [stearoxypropyldimethylamine (SPDA) and docosyldimethylamine (DSDA)] makes the alkaline-color-treated weathered hair surface hydrophobic and that its hydrophobicity is maintained even after shampooing. Characterization of adsorbed layers of 18-MEA/SPDA on a mica surface, as a possible hydrophilic surface model, was performed using atomic force microscopy (AFM) and angle-resolved X-ray photoelectron spectroscopy (AR-XPS). The results revealed that 18-MEA/SPDA formed a layer with high wear resistance, with an alkyl chain, the hydrophobic moiety, oriented at an angle of around 25 degrees to the air interface.

Characterization of curved hair of Japanese women with reference to internal structures and amino acid composition.

The variation of hair curvature in Japanese women was quantitatively investigated and the structure of curved hair was characterized with transmission electron microscopy (TEM) and amino acid analysis. Two hundred and thirty Japanese women volunteers, aged from 10 to 70 years, were randomly selected. The evaluation of the volunteers' natural hair shape showed that 53% of Japanese women have straight hair, while the remaining 47% have curved hair (varying from a slightly wavy shape to a frizzy style). The average curl radius of the volunteers' hair was determined to be 4.4 +/- 2.3 cm, and ranged widely from 0.6 to 16 cm. The TEM observation of curved hair fiber revealed an inhomogenous internal structure between the outer and inner regions of the curved shape. In relation to the inhomogeneous structure of the curved hair, different amino acid composition of the hair keratin was observed between the outer and inner regions. Interestingly, these results of the TEM observation and the amino acid analysis are analogous to the difference between the ortho- and paracortical cells in wool fibers, suggesting the universal structure of curved mammalian hair.

Highly sensitive determination of diverse ceramides in human hair using reversed-phase high-performance liquid chromatography-electrospray ionization mass spectrometry.

Since ceramides (CERs) play roles in signal transduction and cell regulation, CERs of human hair might be responsible for apoptosis during keratinization, in addition to their structural barrier and water-holding functions. Although, we previously developed a method for comprehensive profiling of the CERs in hair, that method was too insensitive to quantitatively characterize the CERs in a small amount of hair samples. The aim of this study was to develop a novel method for the highly sensitive determination of the diverse CERs. The method developed is negative ion electrospray ionization mass spectrometry (ESI-MS) coupled to reversed-phase high-performance liquid chromatography (RP-HPLC) using methanol containing 10 mM ammonium acetate as a mobile phase. By this method, 48 peaks derived from 73 kinds of CERs were simultaneously determined in selected ion monitoring measurement using one calibration line of the standard N-palmitoyl dihydrosphigosine, based on extremely small differences in the molar responses among different species of CERs, followed by the calculation of the actual levels using corrections for (13)C and (2)H effects. This method had extremely high sensitivity as indicated in the limit of quantification being in the femtomolar range. Other quantitative validation data, such as reproducibility, linearity and recoveries, were all sufficient. The quantitative levels of CERs determined by RP-HPLC-ESI-MS were comparable with those determined by thin-layer chromatography. This method was successfully applied to the characterization of levels of CERs in only 1-mm pieces derived from a single hair fiber and revealed the presence of interindividual and intraindividual variations of the CER composition. This RP-HPLC-ESI-MS method can be a powerful tool for future research on physicochemical and physiological roles of CERs in hair.

Highly sensitive determination of N-acyl dihydrosphingosine using liquid chromatography-electrospray ionization mass spectrometry. Application to human hair lipids.

An analytical method for highly sensitive determination of four N-acyl dihydrosphingosines (NDSs) of all ceramides (CERs) in human hair, such as N-palmitoyl dihydrosphingosine (N16DS18), N-stearoyl dihydrosphingosine (N18DS18), N-lignocerol dihydrosphingosine (N24DS18) and N-nervonoyl dihydrosphingosine (N24:1DS18), has been developed using electrospray ionization (ESI) MS connected to reversed-phase LC with selected ion monitoring (SIM). The selection of negative ESI under optimal conditions of in-source collision-induced dissociation was determined based on the simplicity of molecular-related ions and their intensities. Of all ESI-MS parameters tested, the flow of dry nitrogen gas strongly affected the sensitivity of molecular-related ions, particularly in N24DS18 and N24:1DS18, while the capillary voltage elicited significantly different effects on the signal-to-noise ratio between N16DS18/N18DS18 and N24DS18/N24:1DS18. This newly developed method to determine the NDSs is the most sensitive of all existing methods, as shown in the limits of detection and quantification being in the range of 0.06-0.29 and 0.18-0.98fmol, respectively. The linearity, precision and accuracy were all sufficient to determine the NDSs in ca. 0.1mg of a hair fiber ( approximately 1cm in length). This method has been used to characterize levels of the NDSs from the proximal root end to the distal tip of each of six hair fibers obtained from two different females. Characteristic changes were observed between both females as well as among fibers derived from each female. This method will be useful not only for clarifying the roles of the CERs in human hair but also for investigating the physiology of CERs relevant to signal transduction and cell regulation in human cells/tissues.

Liquid chromatography-mass spectrometry for comprehensive profiling of ceramide molecules in human hair.

Ceramides (CERs) play key roles in signal transduction and cell regulation, probably during the keratinization of human hair. Current methods using mass spectrometry (MS), however, are not sufficient to allow the comprehensive analysis of CER molecules, including isobaric and isomeric CERs. Therefore, a method for the comprehensive profiling of CERs was developed. The method developed is based on reversed-phase liquid chromatography (RPLC) coupled to atmospheric pressure chemical ionization (APCI)-MS. Comprehensive identification and profiling of CERs is achieved using two sets of multimass chromatograms obtained from two channel detections that monitor both molecular-related and sphingoid-related ions under two different in-source collision-induced dissociation conditions and using retention times obtained from RPLC. The application of this method revealed that human hair contains 73 species of CER molecules, which were all corroborated by structural analysis using tandem mass spectrometry. The results further revealed that the composition is characterized by predominant molecules consisting of even carbon atom-containing saturated/unsaturated nonhydroxy or alpha-hydroxy fatty acids and C(18) dihydrosphingosine, a minor but distinct content of isobaric/isomeric and odd chain-containing CERs. This successfully developed RPLC-APCI-MS technique allows the comprehensive profiling of CER molecules in hair for the investigation of their physicochemical and physiological roles.

Characterization of hair lipid images by argon sputter etching-scanning electron microscopy.

Hair lipid images, as visualized by argon sputter etching-scanning electron microscopy (ASE-SEM), reveal convex structures with a stitch pattern (SP) at the cell membrane complex (CMC) in the transverse hair plane. Based on interindividual variation, different features of the convex SP were classified into Types 0 to 4 with the corresponding scores 0 to 4. Observations using hair fibers collected from 27 Japanese females revealed significant positive correlations between the scores and the levels of exogenous lipids, which suggests that exogenous lipids internalized at the CMC predominantly constitute the convex SP. Intraindividual variation with different levels of exogenous lipids among hair fibers derived from individual females may be relevant to the uneven physicochemical properties of hair fibers on the scalp. Observations of 380 hair fibers collected from Japanese (Mongoloid), German and American (Caucasoid) females aged 3 to 77 yr demonstrated similar age-related changes in the lipid images, which represent an increase and then a decrease in levels of exogenous lipids with increasing age. This suggests that age-related changes in exogenous lipids are attributable to alterations in sebum excreted during aging and that this elicits age-related changes in physical parameters, which affect human hair texture.

A systematic method for the sensitive and specific determination of hair lipids in combination with chromatography.

A systematic method for the sensitive, precise and accurate determination of hair lipids, including trace amounts of intrinsic endogenous cholesterol (CH), ceramide/N-palmitoyl-DL-dihydrosphingosine (CER/PDS), cholesterol sulfate (CS) and chemically bound 18-methyl eicosanoic acid (18-MEA), has been developed in combination with TLC/FID (flame ionization detection), LC/MS and GC/MS. TLC/FID was used for the simultaneous determination of squalene (SQ), wax esters (WEs), triglycerides (TGs) and free fatty acids (FFAs). Optimal conditions for LC/MS to determine CS and 18-MEA were developed using selected ion monitoring (SIM) under the negative ion mode of electrospray ionization. An alternative procedure for the determination of 18-MEA was also established using commercially available heneicosanoic acid (HEA). In GC/MS, the optimal selection of ions for SIM of trimethylsilylated CH and CER/PDS, and the use of on-column injection has enabled their simultaneous detection. This newly developed method has been used to characterize the hair lipid composition from the proximal root end to the distal tip of chemically untreated hair fibers from two different females, and specific changes of hair lipids probably due to its origin and individuals have been demonstrated for the first time. This method may be useful for clarifying the important roles of intrinsic endogenous 18-MEA, CS, CH and CERs in the function of the cell membrane complex of hair fibers.