Prediction and association analyses of skin phenotypes in Japanese females using genetic, environmental, and physical features.

BACKGROUND: Skin characteristics show great variation from person to person and are affected by multiple factors, including genetic, environmental, and physical factors, but details of the involvement and contributions of these factors remain unclear. OBJECTIVES: We aimed to characterize genetic, environmental, and physical factors affecting 16 skin features by developing models to predict personal skin characteristics. METHODS: We analyzed the associations of skin phenotypes with genetic, environmental, and physical features in 1472 Japanese females aged 20-80 years. We focused on 16 skin characteristics, including melanin, brightness/lightness, yellowness, pigmented spots, wrinkles, resilience, moisture, barrier function, texture, and sebum amount. As genetic factors, we selected 74 single-nucleotide polymorphisms of genes related to skin color, vitamin level, hormones, circulation, extracellular matrix (ECM) components and ECM-degrading enzymes, inflammation, and antioxidants. Histories of ultraviolet (UV) exposure and smoking as environmental factors and age, height, and weight as physical factors were acquired by means of a questionnaire. RESULTS: A linear association with age was prominent for increase in the area of crow's feet, increase in number of pigmented spots, decrease in forehead sebum, and increase in VISIA wrinkle parameters. Associations were analyzed by constructing linear regression models for skin feature changes and logistic regression models to predict whether subjects show lower or higher skin measurement values in the same age groups. Multiple genetic factors, history of UV exposure and smoking, and body mass index were statistically selected for each skin characteristic. The most important association found for skin spots, such as lentigines and wrinkles, was adolescent sun exposure. CONCLUSION: Genetic, environmental, and physical factors associated with interindividual differences of the selected skin features were identified. The developed models should be useful to predict the skin characteristics of individuals and their age-related changes.

The protective role of DJ-1 in ultraviolet-induced damage of human skin: DJ-1 levels in the stratum corneum as an indicator of antioxidative defense.

DJ-1 is a multifunctional protein associated with Parkinson's disease and plays a significant role in protecting nerve cells from oxidative stress. DJ-1 is expressed in the skin, although its function there is unknown. In this study, we investigated DJ-1 function in keratinocytes. DJ-1 was induced by H2O2 exposure and UV irradiation in keratinocytes. DJ-1 knockdown with small interfering RNA (siRNA) increased reactive oxygen species (ROS) and lactate dehydrogenase (LDH) release after UVB irradiation, suggesting that DJ-1 reduces ROS and might protect skin cells from UV damage in vitro. To investigate the in vivo role of DJ-1 in the skin, we determined DJ-1 levels in human stratum corneum samples obtained by the tape-stripping method. DJ-1 levels in the stratum corneum (scDJ-1) correlated with total antioxidant capacity. We also examined the effect of scDJ-1 on changes in skin after UVB irradiation. DJ-1 was elevated in SC from the upper arm 1 to 2 weeks after UVB irradiation. One day after UVB irradiation, L* (brightness) and a* (redness) values, indicators of skin color, were altered regardless of scDJ-1 expression. However, these values recovered more quickly in subjects with high scDJ-1 expression than in those with low scDJ-1 expression. These data suggest that DJ-1 in skin plays a significant role in protection against UV radiation and oxidative stress, and that DJ-1 levels in the SC might be an indicator of antioxidative defense against UV-induced damage.

Macrophage migration inhibitory factor (MIF) in the stratum corneum: a marker of the local severity of atopic dermatitis.

Different biomarkers are used to evaluate the severity of atopic dermatitis (AD); however, it remains difficult to determine the severity of localized skin lesions. MIF plays an essential role in the pathophysiology of skin inflammation. To establish whether the MIF level in the stratum corneum (SC) serves as a marker of the severity of AD lesions, we examined the SC MIF (scMIF) levels in AD patients. The SC of the cheek, neck and upper arm skin was collected using tape stripping, and the scMIF levels were measured. Consequently, the scMIF levels were found to be significantly higher in the involved skin lesions than the uninvolved areas within the same patient. Moreover, the scMIF levels were significantly correlated with the severity of local skin lesions. These findings suggest that the scMIF level can be used as an effective marker for evaluating the local severity of AD.

An atypical case of lymphoproliferative pulmonary involvement in a patient with Sjögren's syndrome: a case report.

BACKGROUND: Sjögren's syndrome is characterized by lymphocytic infiltration of the exocrine glands, together with polyclonal B-cell activation, and lung diseases are well-known complications of the disease. Therefore, in most cases associated with Sjögren's syndrome, infiltrating lymphocytes in the lung specimen exhibit the features of B-cells. We herein report an atypical case of lymphoproliferative pulmonary involvement in a patient with Sjögren's syndrome. CASE PRESENTATION: A 46-year-old female was admitted to our hospital because of an abnormal chest roentgenogram finding on a medical checkup. Chest computed tomography showed randomly-distributed micronodules and patchy ground-glass opacities. A surgical biopsied specimen showed an atypical pattern of interstitial pneumonia with numerous lymphoid follicles. Among the infiltrating lymphocytes in the lung, only the monoclonality of the T-cells was proven by a gene rearrangement analysis, but there was no cytological atypicality or genetic disorder revealed by testing the bone marrow aspirate. A diagnosis of Sjögren's syndrome was made based on the patient's other symptoms and these negative findings. The patient's pulmonary lesions have been successfully treated and remission has been maintained for over three years with corticosteroid treatment alone. CONCLUSION: The present patient was an atypical case of lymphoproliferative pulmonary involvement in a patient with Sjögren's syndrome. Although monoclonality of the infiltrating T-cells was proven, the clinical course and the findings of the imaging and laboratory examinations were inconsistent with the previously-reported cases of primary pulmonary T-cell lymphoma. This suggests that the monoclonality of lymphocytes does not always define malignancy. The diagnosis of malignant lymphoma or lymphoproliferative diseases should be made clinically, pathologically and cytogenetically to rule out other similar diseases.

Antiproliferative effects of galectin-1 from Rana catesbeiana eggs on human leukemia cells and its binding proteins in human cells.

Galectin-1 from American bullfrog, RCG1, was isolated to high purity, and its growth inhibitory properties against human cells were examined. The results demonstrated that highly purified RCG1 induced large cell aggregates and revealed cell-type-specific growth inhibition. It significantly inhibited all human leukemia cell lines tested such as HL-60, U937, and K562 cells but did not inhibit human colon cancer cell line, Colo 201, or mouse mammary tumor cell line FM3A cells. Although most of the galectin-induced growth inhibitions are known to be apoptic, RCG1 induced growth arrest and neither apoptosis nor necrosis. RCG1-mediated growth inhibition was specifically suppressed by the corresponding sugar, lactose, but not by sucrose or even the structurally similar sugar, melibiose. Several studies have reported that galectin-mediated biological functions were modulated by charge modification. Since the high purity of RCG1 was demonstrated but a moderate degree of growth inhibition occurred, it is possible protein charge modification was examined by isoelectric focusing, and it was found to be highly heterogeneous in charge. RCG1 binding proteins in human cells were analyzed by lectin blotting using biotinylated RCG1, and lectin blotting revealed that in human cell extracts the specific proteins at molecular weight 37 and 50 kDa possessed the responsive features of RCG1 binding and lactose competition.

Clinical features of healthcare-associated pneumonia (HCAP) in a Japanese community hospital: comparisons among nursing home-acquired pneumonia (NHAP), HCAP other than NHAP, and community-acquired pneumonia.

BACKGROUND AND OBJECTIVE: More than 100000 Japanese die of pneumonia every year. The number of people residing in nursing homes is increasing with the ageing of the population. In 2005, the American Thoracic Society/Infectious Diseases Society of America (ATS/IDSA) published important guidelines for the management of healthcare-associated pneumonia (HCAP). In Japan, however, the optimum strategy for management of HCAP is still unclear. The purpose of this study was to clarify the clinical features of patients with HCAP. METHODS: Patients (n = 202) who were consecutively admitted with a diagnosis of acute pneumonia between October 2007 and September 2009 were retrospectively evaluated. Using the ATS/IDSA guidelines, patients were divided into three groups: a community-acquired pneumonia (CAP) group (n = 123), a nursing home-acquired pneumonia (NHAP) group (n = 46) and a HCAP other than NHAP (O-HCAP) group (n = 33). These groups were then compared with respect to laboratory data, microbiological findings and mortality. RESULTS: Thirty-day mortality in the NHAP group (10.9%) tended to be higher than that in the CAP group (3.3%) or the O-HCAP group (0%). The pathogens most frequently identified were Streptococcus pneumoniae and Haemophilus influenzae in the CAP group, methicillin-resistant Staphylococcus aureus and Klebsiella pneumoniae in the NHAP group, and S. pneumoniae and K. pneumoniae in the O-HCAP group. CONCLUSIONS: The NHAP group was clinically different from the O-HCAP group, based on bacteriological examination and mortality rates. In order to accurately diagnose, and formulate optimum treatment strategies for Japanese patients, the categories of HCAP, as specified in the ATS/IDSA guidelines, should not be applied directly either to patients with NHAP or those with O-HCAP.

New horny layer marker proteins for evaluating skin condition in atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) has a complicated pathogenesis and its clinical features vary greatly among patients. Although many clinical parameters have been reported, it remains difficult to evaluate AD skin conditions adequately. OBJECTIVE: To support better evaluation of AD patients, we attempted to develop a new, objective and noninvasive method that assesses skin condition in AD using biochemical markers in the skin's horny layer (HL). METHODS: Thirty-six patients with AD, 8 with psoriasis and 16 healthy volunteers were recruited. HL samples were obtained by tape stripping from involved and uninvolved skin of the forearms. Expression levels of 6 proteins in the HL [fatty acid-binding protein-5 (FABP-5), squamous cell carcinoma antigens 2 (SCCA2), alpha-enolase, annexin II, apolipoprotein A-I and albumin] were analyzed by immunoblotting and compared with clinical data. RESULTS: The 6 proteins were detected at a high level in AD skin lesions, but scarcely in the normal controls. FABP-5 showed correlation with the local severity of the involved skin. Annexin II, apoprotein A-I and albumin showed correlation with the severity of specific eruptions. SCCA2 correlated significantly with total serum IgE level. Albumin levels in the uninvolved skin of AD patients showed significant correlation with the local severity in the involved skin of the same patient and with the trans-epidermal water loss. Albumin levels in psoriatic skin were very low, even with scratch marks, compared to those in AD skin. CONCLUSION: FABP-5, albumin and some other proteins in HL seem to be useful as biomarkers to evaluate inflammation and skin barrier conditions in AD patients.

Localization of laminin alpha3B chain in vascular and epithelial basement membranes of normal human tissues and its down-regulation in skin cancers.

The basement membrane (BM) proteins laminins, which consist of alpha, beta and gamma chains, play critical roles in the maintenance of tissue structures. One of laminin alpha chains, alpha3 has two isoforms, the truncated form alpha3A and the full-sized form alpha3B. In contrast to alpha3A laminins, little is known about alpha3B laminins. To show the histological distribution of the laminin alpha3B chain, we prepared alpha3B-specific monoclonal antibodies. Immunohistochemical analysis showed that the alpha3B chain was colocalized with the alpha3A, beta3 and gamma2 chains in the epithelial BMs of the skin, esophagus, breast and lung, suggesting the presence of laminin-3B32 (laminin-5B) and laminin-3A32 (laminin-5A). In the lung alveoli, laminin-3B32 was dominant over laminin-3A32, but vice versa in other epithelial BMs. In contrast, the BMs of blood vessels including capillaries were strongly positive for alpha3B, but almost or completely negative for alpha3A, beta3 and gamma2. alpha3B was colocalized with beta1 and gamma1 in these BMs. The alpha3B chain was scarcely detected in the vessels of malignant skin cancers, though the gamma2 and beta3 chains were highly expressed in the cancer cells. These results strongly suggest that the laminin alpha3B chain is widely expressed in vascular BMs of normal tissues, probably as laminin-3B11/3B21 (laminin-6B/7B).

Identification of membrane-bound serine proteinase matriptase as processing enzyme of insulin-like growth factor binding protein-related protein-1 (IGFBP-rP1/angiomodulin/mac25).

Insulin-like growth factor (IGF) binding protein-related protein-1 (IGFBP-rP1) modulates cellular adhesion and growth in an IGF/insulin-dependent or independent manner. It also shows tumor-suppressive activity in vivo. We recently found that a single-chain IGFB-rP1 is proteolytically cleaved to a two-chain form by a trypsin-like, endogenous serine proteinase, changing its biological activities. In this study, we attempted to identify the IGFBP-rP1-processing enzyme. Of nine human cell lines tested, seven cell lines secreted IGFBP-rP1 at high levels, and two of them, ovarian clear cell adenocarcinoma (OVISE) and gastric carcinoma (MKN-45), highly produced the cleaved IGFBP-rP1. Serine proteinase inhibitors effectively blocked the IGFBP-rP1 cleavage in the OVISE cell culture. The conditioned medium of OVISE cells did not cleave purified IGFBP-rP1, but their membrane fraction had an IGFBP-rP1-cleaving activity. The membrane fraction contained an 80-kDa gelatinolytic enzyme, which was identified as the membrane-type serine proteinase matriptase (MT-SP1) by immunoblotting. When the membrane fraction was separated by SDS/PAGE, the IGFBP-rP1-cleaving activity comigrated with matriptase. A soluble form of matriptase purified in an inhibitor-free form efficiently cleaved IGFBP-rP1 at the same site as that found in a naturally cleaved IGFBP-rP1. Furthermore, small interfering RNAs for matriptase efficiently blocked both the matriptase expression and the cleavage of IGBP-rP1 in OVISE cells. These results demonstrate that IGFBP-rP1 is processed to the two-chain form by matriptase on the cell surface.

Regulation of cell adhesion and type VII collagen binding by the beta3 chain short arm of laminin-5: effect of its proteolytic cleavage.

The basement membrane protein laminin-5 (Lm5), a heterotrimer of alpha3 (or alpha3A), beta3, and gamma2 chains, regulates cellular adhesion and motility. Here we examined the proteolysis and biological function of the laminin beta3 chain. First, we found that the beta3 chain of Lm5 is cleaved at its N-terminal, short arm by an endogenous proteinase(s) in normal human keratinocytes and some other cell lines. To examine the effect of beta3 chain cleavage, we expressed a wild-type Lm5 and two Lm5 mutants with partially deleted beta3 chains in HEK293 cells. Experiments with the purified Lm5 forms demonstrated that the deletion of the beta3 short arm or its N-terminal domain LN decreases the cell adhesion activity of Lm5, but does not significantly affect the motility activity. A recombinant beta3 short arm protein enhanced integrin-mediated cell adhesion to Lm5 by binding to an unidentified cell receptor. It was also found that the laminin EGF-like domain of the beta3 short arm is a binding site for type VII collagen. These results suggest that the beta3 short arm is involved not only in the matrix assembly of Lm5, but also in its cell adhesion activity. The proteolytic cleavage of the beta3 chain may modulate these functions of Lm5 in vivo.

Regulation of biological activity and matrix assembly of laminin-5 by COOH-terminal, LG4-5 domain of alpha3 chain.

The basement membrane protein laminin-5 (LN5; alpha3beta3gamma2) undergoes specific proteolytic processing of the 190-kDa alpha3 chain to the 160-kDa form after the secretion, releasing its COOH-terminal, LG4-5 domain. To clarify the biological significance of this processing, we tried to express a recombinant precursor LN5 with a 190-kDa alpha3 chain (pre-LN5), in which the cleavage sequence Gln-Asp was changed to Ala-Ala by point mutation. When the wild-type and mutated LN5 heterotrimers were expressed in HEK293 cells, the wild-type alpha3 chain was completely cleaved, whereas the mutated alpha3 chain was partially cleaved at the same cleavage site (Ala-Ala). pre-LN5 was preferentially deposited on the extracellular matrix, but this deposition was effectively blocked by exogenous heparin. This suggests that interaction between the LG4-5 domain and heparan sulfate proteoglycans on the cell surface and/or extracellular matrix is important in the matrix assembly of LN5. Next, we purified both pre-LN5 and the mature LN5 with the processed, 160-kDa alpha3 chain (mat-LN5) from the conditioned medium of the HEK293 cells and compared their biological activities. mat-LN5 showed higher activities to promote cell adhesion, cell scattering, cell migration, and neurite outgrowth than pre-LN5. These results indicate that the proteolytic removal of LG4-5 from the 190-kDa alpha3 chain converts the precursor LN5 from a less active form to a fully active form. Furthermore, the released LG4-5 fragment stimulated the neurite outgrowth in the presence of mat-LN5, suggesting that LG4-5 synergistically enhances integrin signaling as it is released from the precursor LN5.

Characterization of laminin 5B and NH2-terminal proteolytic fragment of its alpha3B chain: promotion of cellular adhesion, migration, and proliferation.

Various laminin isoforms have specific biological functions depending on their structures. Laminin 5A, which consists of the three truncated chains alpha3A, beta3, and gamma2, is known to have strong activity to promote cell adhesion and migration, whereas a laminin 5 variant consisting of a full-sized alpha3 chain (alpha3Beta) and the beta3 and gamma2 chains, laminin 5B, has not been characterized yet. In the present study, we for the first time cloned a full-length human laminin alpha3B cDNA and isolated the human laminin 5B protein. The molecular size of the mature alpha3B chain (335 kDa) was approximately twice as large as the mature alpha3A chain in laminin 5A. Laminin 5B had significantly higher cell adhesion and cell migration activities than laminin 5A. In addition, laminin 5B potently stimulated cell proliferation when added into the culture medium directly. Furthermore, we found that the alpha3B chain undergoes proteolytic cleavage releasing a 190-kDa NH(2)-terminal fragment. The 190-kDa fragment had activities to promote cellular adhesion, migration, and proliferation through its interaction with integrin alpha(3)beta(1). These activities of the NH(2)-terminal structure of the alpha3B chain seem to contribute to the prominent biological activities and the physiological functions of laminin 5B.

Deficiency in a mitochondrial aldehyde dehydrogenase increases vulnerability to oxidative stress in PC12 cells.

Mitochondrial aldehyde dehydrogenase 2 (ALDH2) plays a major role in acetaldehyde detoxification. The alcohol sensitivity is associated with a genetic deficiency of ALDH2. We have previously reported that this deficiency influences the risk for late-onset Alzheimer's disease. However, the biological effects of the deficiency on neuronal cells are poorly understood. Thus, we obtained ALDH2-deficient cell lines by introducing mouse mutant Aldh2 cDNA into PC12 cells. The mutant ALDH2 repressed mitochondrial ALDH activity in a dominant negative fashion, but not cytosolic activity. The resultant ALDH2-deficient transfectants were highly vulnerable to exogenous 4-hydroxy-2-nonenal, an aldehyde derivative generated by the reaction of superoxide with unsaturated fatty acid. In addition, the ALDH2-deficient transfectants were sensitive to oxidative insult induced by antimycin A, accompanied by an accumulation of proteins modified with 4-hydroxy-2-nonenal. Thus, these findings suggest that mitochondrial ALDH2 functions as a protector against oxidative stress.

[Comparative evaluation of serum HCV RNA by Roche Monitor Assay. Versions 1.0 and 2.0; as a predictive marker of subsequent response to IFN therapy].

Quantitative measurement of serum hepatitis C virus(HCV) RNA is important in predicting and monitoring interferon(IFN) therapy. We compared the sensitivity of HCV RNA measurement of different HCV genotypes between two available assays, Roche Monitor 1.0(v1.0) and Roche Monitor 2.0(v2.0). We also evaluated serum level of HCV RNA as the predictors of a long-term response to IFN therapy by distinguishing the complete responders(CR), partial responders(PR) and non-responders(NR) for IFN therapy. We quantified the serum HCV RNA levels in 151 patients and determined the genotypes; 96(64%) with genotype 1b(1b), 42(28%) with genotype 2a(2a), and 6(4%) was not identified. The relationship between the genotype and effects of IFN treatment was as follows: 23CR(1b:11, 2a:9), 15PR(1b:8, 2a:7), 20NR(1b:14, 2a:6). The RNA levels of 2a measured by v2.0 were significantly higher than those by v1.0(p < 0.05), although no significant difference was found in 1b between two assays. By using v2.0, when the cut-off level was set at 200 x 10(3) IU/ml before IFN therapy, CR was discriminated from PR, NR with a predicting efficiency of 88%. HCV RNA levels before IFN therapy were significantly lower in patients who became HCV RNA negative within 2 weeks than in patients who did at 4 weeks or longer. These results suggest that v2.0 is more sensitive and accurate than v1.0 for the quantification of 2a. Using v2.0 assay, it was shown that low viral titre at pretreatment and loss of viraemia within 2 weeks after treatment might be important markers for a long-term response to IFN therapy, irrespective of viral genotype. The v2.0 assay was found to be more useful in predicting effectiveness of IFN therapy.