Management of the human hair follicle microbiome by a synthetic odorant.

BACKGROUND: Human scalp hair follicles (HFs) engage in olfactory receptor (OR)-dependent chemosensation. Activation of olfactory receptor family 2 subfamily AT member 4 (OR2AT4) by the synthetic, sandalwood-like odorant Sandalore® up-regulated HF antimicrobial peptide expression of dermcidin (DCD), which had previously been thought to be produced exclusively by sweat and sebaceous glands. OBJECTIVES: To understand if intrafollicular DCD production can be stimulated by a commonly used cosmetic odorant, thus altering human HF microbiome composition in a clinically beneficial manner. METHODS: DCD expression was compared between fresh-frozen scalp biopsies and microdissected, full-length scalp HFs, organ-cultured in the presence/absence of the OR2AT4 agonist, Sandalore® and/or antibiotics and/or the competitive OR2AT4 antagonist, Phenirat®. Amplicon-based sequencing and microbial growth assays were performed to assess how this treatment affected the HF microbiome. RESULTS: Synthetic odorant treatment upregulated epithelial DCD expression and exerted antimicrobial activity in human HFs ex vivo. Combined antibiotic and odorant treatment, during an ex vivo dysbiosis event, prevented HF tissue damage and favoured a more physiological microbiome composition. Sandalore®-conditioned medium, containing higher DCD content, favoured Staphylococcus epidermidis and Malassezia restricta over S. aureus and M. globosa, while exhibiting antimicrobial activity against Cutibacterium acnes. These effects were reversed by co-administration of Phenirat®. CONCLUSIONS: We provide the first proof-of-principle that a cosmetic odorant impacts the human HF microbiome by up-regulating antimicrobial peptide production in an olfactory receptor-dependent manner. Specifically, a synthetic sandalwood-like odorant stimulates intrafollicular DCD production, likely via OR2AT4, and thereby controls microbial overgrowth. Thus, deserving further exploration as an adjuvant therapeutic principle in the management of folliculitis and dysbiosis-associated hair diseases.

Insights into the molecular determinants of thermal stability in halohydrin dehalogenase HheD2.

Halohydrin dehalogenases (HHDHs) are promising enzymes for application in biocatalysis due to their promiscuous epoxide ring-opening activity with various anionic nucleophiles. So far, seven different HHDH subtypes A to G have been reported with subtype D containing the by far largest number of enzymes. Moreover, several characterized members of subtype D have been reported to display outstanding characteristics such as high catalytic activity, broad substrate spectra or remarkable thermal stability. Yet, no structure of a D-type HHDH has been reported to date that could be used to investigate and understand those features on a molecular level. We therefore solved the crystal structure of HheD2 from gamma proteobacterium HTCC2207 at 1.6 Å resolution and used it as a starting point for targeted mutagenesis in combination with molecular dynamics (MD) simulation, in order to study the low thermal stability of HheD2 in comparison with other members of subtype D. This revealed a hydrogen bond between conserved residues Q160 and D198 to be connected with a high catalytic activity of this enzyme. Moreover, a flexible surface region containing two α-helices was identified to impact thermal stability of HheD2. Exchange of this surface region by residues of HheD3 yielded a variant with 10 °C higher melting temperature and reaction temperature optimum. Overall, our results provide important insights into the structure-function relationship of HheD2 and presumably for other D-type HHDHs. DATABASES: Structural data are available in PDB database under the accession number 7B73.

Pregnant adolescent women's perceptions of depression and psychiatric services in the United States.

PROBLEM: Adolescent mothers and their children are at high-risk for depression and the associated negative educational, social, health, and economic outcomes. BACKGROUND: However, few pregnant adolescent women with depression receive psychiatric services, especially low-income or racial/ethnic minority adolescent women. AIM: This qualitative study explores perceptions of depression, psychiatric services, and barriers to accessing services in a sample of low-income, pregnant racial/ethnic minority adolescent women. Our goal was to better understand the experiences of depression during pregnancy for these vulnerable adolescent women, and thereby improve their engagement and retention in services for perinatal depression. METHODS: We recruited 20 pregnant adolescent women who screened positive for depression from 2 public health prenatal clinics in the southeastern United States. Participants were low-income and primarily racial/ethnic minority women between 14 and 20 years old. Data were collected through individual in-depth, ethnographically informed interviews. FINDINGS: Generally, participants lacked experience with psychiatric services and did not recognize their symptoms as depression. However, participants perceived a need for mood improvement and were interested in engaging in services that incorporated their perspective and openly addressed stigma. DISCUSSION: Participants reported practical and psychological barriers to service engagement, but identified few cultural barriers. Family perceptions of psychiatric services served as both a barrier and support. CONCLUSION: Adolescent women are more likely to engage in psychiatric services if those services reduce practical and psychological barriers, promise relief from the symptoms perceived as most meaningful, and address underlying causes of depression. Culture may affect Latina adolescent women's perceptions of depression and services.

I. QUALITY THRESHOLDS, FEATURES, AND DOSAGE IN EARLY CARE AND EDUCATION: INTRODUCTION AND LITERATURE REVIEW.

This monograph addresses the hypotheses that preschool children benefit most strongly when early care and education (ECE) is at or above a threshold of quality, has specific quality features, and/or is of longer duration. These issues are pivotal in recent policies designed to improve the quality of ECE, especially for children from low-income families. Evidence of quality thresholds in which ECE quality has stronger impacts in settings with moderate to high levels of quality than in settings with low quality would inform policy initiatives in which monetary incentives or consequences are allocated to ECE settings based on their level of quality. Evidence that specific features of quality, such as quality of teacher-child interactions and of literacy and mathematics instruction, are predictors of gains in child outcomes could help inform quality improvement efforts. Evidence that more time spent in center-based ECE or in instruction in specific content areas predict larger gains among preschoolers could be useful in designing public preschool programs such as Head Start or prekindergarten. Secondary data analyses of eight large studies of preschool children in center-based ECE were conducted. Analyses focused on quality thresholds and quality features examined the extent to which three types of quality measures predicted gains in children's language, literacy, mathematics, and social skills. The measures comprised (1) global quality measures that provide an overall or global rating of quality, focusing on interactions as well as on physical features of the environment, activities, and routines; (2) interaction-specific measures that focus in depth on the quality of interactions between teachers and children with respect to instructional and emotional support; and (3) domain-specific measures that focus on the quality of instruction and stimulation in specific content areas such as early language and literacy. The goal was to provide replicated analyses with data from several projects in order to address each question. Multilevel analyses that controlled for entry skills were conducted, and results were combined by using meta-analysis, nonlinear and nonparametric analyses, and propensity score analyses. With respect to thresholds, the analyses suggest that increases in the quality of instruction are related to larger gains in language and literacy outcomes, but only in higher quality classrooms. Results point to stronger associations between quality and child outcomes in higher versus lower quality classrooms for measures of the instructional quality of teacher-child interactions and of the quality of specific activities thought to promote early literacy, such as teaching phonemic skills and book reading. In addition, the items focusing on quality of interactions on the global measure also predicted acquisition of language and social skills in higher but not in lower quality classrooms. With respect to quality features, interaction-specific and especially domain-specific measures of quality remained significant predictors of child outcomes, whereas global measures of quality were never significant positive predictors, when both global and more specific measures of quality were included simultaneously in analyses. There is thus consistent evidence that more specific measures of quality are better predictors of child outcomes. With respect to dosage, several approaches were used in operationalizing both the cumulative and current dosage of children's exposure to ECE. Propensity score analyses that included baseline scores on outcomes to control for selection into larger dosages suggested that children with two as opposed to one year of Head Start had stronger vocabulary and literacy skills both immediately upon exit from Head Start and at the end of kindergarten. Fewer absences and more observed time spent on instruction were associated with stronger gains in literacy and mathematics skills. Finally, findings revealed that more time spent on instruction in classrooms with higher overall quality was particularly important to the development of mathematics skills. No other replicated evidence of quality by quantity interactions emerged.

Activity-Fed Translation (AFT) Assay: A New High-Throughput Screening Strategy for Enzymes in Droplets.

There is an increasing demand for the development of sensitive enzymatic assays compatible with droplet-based microfluidics. Here we describe an original strategy, activity-fed translation (AFT), based on the coupling of enzymatic activity to in vitro translation of a fluorescent protein. We show that methionine release upon the hydrolysis of phenylacetylmethionine by penicillin acylase enabled in vitro expression of green fluorescent protein. An autocatalytic setup where both proteins are expressed makes the assay highly sensitive, as fluorescence was detected in droplets containing single PAC genes. Adding a PCR step in the droplets prior to the assay increased the sensitivity further. The strategy is potentially applicable for any activity that can be coupled to the production of an amino acid, and as the microdroplet volume is small the use of costly reagents such as in vitro expression mixtures is not limiting for high-throughput screening projects.

Graft adhesion in descemet membrane endothelial keratoplasty dependent on size of removal of host's descemet membrane.

IMPORTANCE: It is essential to devise strategies that improve graft adhesion after Descemet membrane endothelial keratoplasty (DMEK) to reduce the rebubbling rate. OBJECTIVE: To evaluate the influence of the extent of descemetorhexis on graft adhesion properties after DMEK. DESIGN, SETTING, AND PARTICIPANTS: Single-surgeon, retrospective, observational case series conducted in the Department of Ophthalmology, University of Erlangen-Nuremberg, Germany, that reviewed the medical records of 200 consecutive patients undergoing DMEK. Fifty-three eyes of 51 patients undergoing DMEK for Fuchs endothelial dystrophy fulfilling the inclusion criteria were enrolled in this study. Based on intraoperative drawings, postoperative slitlamp examination, and photographs, eyes were divided into 2 groups. The diameter of the descemetorhexis was approximately 10 mm in group A (30 eyes), resulting in a peripheral 1-mm zone of denuded stroma between the graft and the host's Descemet membrane, and approximately 6 mm in group B (23 eyes), resulting in a peripheral 1-mm zone of overlapping between the graft and the host's Descemet membrane. MAIN OUTCOMES AND MEASURES: Graft detachment rate, extent of graft detachment (in clock hours of graft's circumference), and rebubbling rate. RESULTS: Four days after DMEK, the graft detachment rate was 33.3% (10 of 30) in group A and 78.3% (18 of 23) in group B (P = .002). The mean (SD) extent of graft detachment was 0.6 (0.9) and 2.8 (2.5) clock hours in groups A and B, respectively (P < .001), 4 days after surgery. The rebubbling rate was 6.7% (2 of 30) and 30.4% (7 of 23) for groups A and B, respectively (P = .03). CONCLUSIONS AND RELEVANCE: A larger descemetorhexis in DMEK is correlated with better graft adhesion and lower rebubbling rates. Therefore, patients with a larger descemetorhexis require less intensive follow-up.

Longitudinal analysis of progression in glaucoma using spectral-domain optical coherence tomography.

PURPOSE: To compare the longitudinal loss of RNFL thickness measurements by SD-OCT in healthy individuals and glaucoma patients with or without progression concerning optic disc morphology. METHODS: A total of 62 eyes, comprising 38 glaucomatous eyes with open angle glaucoma and 24 healthy controls, were included in the study (Erlangen Glaucoma Registry, NTC00494923). All patients were investigated annually over a period of 3 years by Spectralis SD-OCT measuring peripapillary RNFL thickness. By masked comparative analysis of photographs, the eyes were classified into nonprogressive and progressive glaucoma cases. Longitudinal loss of RNFL thickness was compared with morphological changes of optic disc morphology. RESULTS: Mixed model analysis of annual OCT scans revealed an estimated annual decrease of the RNFL thickness by 2.12 µm in glaucoma eyes with progression, whereas glaucoma eyes without progression in optic disc morphology lost 1.18 µm per year in RNFL thickness (P = 0.002). The rate of change in healthy eyes was 0.60 µm and thereby also significantly lower than in glaucoma eyes with progression (P < 0.001). The intrasession variability of three successive measurements without head repositioning was 1.5 ± 0.7 µm. The loss of mean RNFL thickness exceeded the intrasession variability in 60% of nonprogressive eyes, and in 85% of progressive eyes after 3 years. CONCLUSIONS: LONGITUDINAL MEASUREMENTS OF RNFL THICKNESS USING SD-OCT SHOW A MORE PRONOUNCED REDUCTION OF RNFL THICKNESS IN PATIENTS WITH PROGRESSION COMPARED WITH PATIENTS WITHOUT PROGRESSION IN GLAUCOMATOUS OPTIC DISC CHANGES. (www.clinicaltrials.gov number, NTC00494923.).

Fungal interface keratitis by Candida orthopsilosis following deep anterior lamellar keratoplasty.

A 39-year-old male patient underwent uncomplicated deep anterior lamellar keratoplasty due to keratoconus. On day 5 after surgery, small whitish infiltrates developed in the corneal interface. The diagnosis of fungal keratitis was made when the culture medium of the graft grew Candida after the surgical intervention. Despite intensive antimycotic treatment and irrigation of the interface, the infiltrates persisted and eventually enlarged. Therefore, revision surgery with penetrating keratoplasty was performed. Microbiological analysis showed Candida orthopsilosis in the culture of the excised graft button. Histopathological staining of the excised graft showed periodic acid-Schiff-positive and Grocott methenamine silver-positive clusters of yeast between Descemet's membrane and the deep corneal stroma with focal perforations through Descemet's membrane. The treatment of mycotic keratitis caused by C orthopsilosis is challenging. Antimycotic treatment was unsuccessful in this case. Progression of the keratitis and perforation of Descemet's membrane suggest that early surgical intervention by penetrating keratoplasty is required.

A prospective study of perinatal depression and trauma history in pregnant minority adolescents.

OBJECTIVE: Adolescent pregnancy is common and minority adolescents are at high risk. We sought the following: (1) to prospectively assess prevalence of antenatal depression (AND) and postpartum depression (PPD) in minority adolescents and (2) to examine the association of social support and adjustment, trauma, and stress on depression status. STUDY DESIGN: A total of 212 pregnant adolescents were recruited from public prenatal clinics and administered a prospective research survey during pregnancy and 6 weeks' postpartum. Depression was measured using the Edinburgh Postnatal Depression Scale. Univariate, bivariate, and multivariable analyses were performed using logistic regression to assess predictors of AND and PPD. RESULTS: In our cohort, 20% screened positive for AND and 10% for PPD. The strongest predictor of PPD was AND (odds ratio [OR], 4.89; P < .001). Among adolescents with trauma history, there was a 5-fold increase (OR, 5.01) in the odds of AND and a 4-fold increase (OR, 3.76) in the odds of PPD. AND was associated with the adolescent's poor social adjustment (P < .001), perceived maternal stress (P < .001), less social support (P < .001), and a less positive view of pregnancy (P < .001). PPD was significantly associated with primiparity (P = .002), poor social adjustment (P < .001), less social support and involvement of the baby's father (P < .001), and less positive view of pregnancy (P < .001). CONCLUSION: Significant independent risk factors for PPD include AND, view of pregnancy, and social support. Trauma history was highly prevalent and strongly predicted AND and PPD. Point prevalence decreased postpartum, and this may be due to transient increased social support following the birth, warranting longer follow-up and development of appropriate interventions in future work.

Invasion of lymphatic vessels into the eye after open globe injuries.

PURPOSE: We analyzed whether lymphatic vessels can be detected in eyes enucleated after an open globe injury. METHODS: The presence of lymphatic vessels was analyzed immunohistochemically using podoplanin as a specific lymphatic endothelial marker in 21 globes that had been enucleated after open globe injury. The localization of pathologic lymphatic vessels (within the eye wall or inside the eye) was correlated with the mechanism of trauma, anatomic site of perforation or rupture, and time interval between trauma and enucleation. RESULTS: Pathologic lymphatic vessels were detected in 15 of 21 eyes (71%) enucleated after an open globe injury. In 5 globes (24%) they were found within the eye, located in retrocorneal membranes, underneath the sclera, and adjacent to uveal tissue (ciliary body, iris). No significant association was observed between the presence of pathologic lymphatic vessels and the mechanism of trauma (P = 0.598), anatomic site of perforation or rupture (P = 0.303), and time interval between trauma and enucleation (P = 0.145). CONCLUSIONS: The human eye can be invaded secondarily by lymphatic vessels if the eye wall is opened by trauma. This mechanism could be important for wound healing, immunologic defense against intruding microorganisms, and autoimmune reactions against intraocular antigens.

FoxO proteins mediate hypoxic induction of connective tissue growth factor in endothelial cells.

Hypoxia, a driving force in neovascularization, promotes alterations in gene expression mediated by hypoxia-inducible factor (HIF)-1alpha. Connective tissue growth factor (CTGF, CCN2) is a modulator of endothelial cell growth and migration, but its regulation by hypoxia is poorly understood. Therefore, we analyzed signaling pathways involved in the regulation of CTGF by hypoxia in endothelial cells. Exposure to low oxygen tension or treatment with the hypoxia-mimetic dimethyloxalyl glycine (DMOG) stabilized HIF-1alpha and up-regulated CTGF in human umbilical vein endothelial cells and in a murine microvascular endothelial cell line. Induction of CTGF correlated with a HIF-dependent increase in protein and mRNA levels, and nuclear accumulation of the transcription factor FoxO3a. By contrast, gene expression and cellular localization of FoxO1 were not significantly altered by hypoxia. Expression of CTGF was strongly reduced by siRNA silencing of FoxO1 or FoxO3a. Furthermore, nuclear exclusion of FoxO1/3a transcription factors by inhibition of serine/threonine protein phosphatases by okadaic acid inhibited CTGF expression, providing evidence for both FoxO proteins as regulators of CTGF expression. The DMOG-stimulated induction of CTGF was further increased when endothelial cells were co-incubated with transforming growth factor-beta, an activator of Smad signaling. Activation of RhoA-Rho kinase signaling by the microtubule-disrupting drug combretastatin A4 also enhanced the DMOG-induced CTGF expression, thus placing CTGF induction by hypoxia in a network of interacting signaling pathways. Our findings provide evidence that FoxO1, hypoxia-stimulated expression of FoxO3a and its nuclear accumulation are required for the induction of CTGF by hypoxia in endothelial cells.

Hypoxia interferes with connective tissue growth factor (CTGF) gene expression in human proximal tubular cell lines.

BACKGROUND: Hypoxia plays an important role in kidney injury. By the stabilization of the transcription factor HIF-1, hypoxia affects gene expression also in tubular epithelial cells. Increased expression of connective tissue growth factor (CTGF) is observed in different kidney diseases and is associated with deteriorating renal function. Therefore, we hypothesized that the expression of CTGF might be modulated under hypoxic conditions. METHODS: The human proximal tubular epithelial cell lines HK-2 and HKC-8 were treated with reduced oxygen tension (1% O(2)) or the hypoxia mimetic dimethyloxalyl glycine (DMOG). CTGF was analysed by Western blotting, real-time RT-PCR and luciferase gene expression assays. RESULTS: Exposure of HK-2 or HKC-8 cells to hypoxia or treatment with DMOG for up to 24 h reduced cellular as well as secreted CTGF protein synthesis. Downregulation was also detectable at the mRNA level and was confirmed by reporter gene assays. Hypoxic repression of CTGF synthesis was dependent on HIF-1, as shown by HIF-1alpha knockdown by siRNA. Furthermore, exposure to hypoxia reduced CTGF synthesis in response to TGF-beta. A negative correlation between HIF-1alpha accumulation and CTGF synthesis was also observed in renal cell carcinoma cells (RCC4 and RCC10). Reexpression of von Hippel-Lindau protein reduced HIF-1alpha and increased CTGF synthesis. CONCLUSIONS: We provide evidence that hypoxia inhibits CTGF synthesis in human proximal tubular epithelial cells, involving HIF-1alpha. Under hypoxic conditions, induction of CTGF by TGF-beta was repressed. The reduced synthesis of the profibrotic factor CTGF may contribute to a potential protective effect of hypoxic preconditioning in acute renal injury.

Characterization of oxalic acid derivatives as new metabolites of metamizol (dipyrone) in incubated hen's egg and human.

Metamizol (dipyrone, 1), a widely used drug with effective analgesic and antispasmodic properties, shows severe side effects like agranulocytosis and anaphylactic shock reactions, the reasons of which are not known until today. After oral administration 1 is completely metabolized. All hitherto known metabolites have an intact pyrazolinone ring structure like the parent compound and are completely extractable from urine with polar organic solvents. However, only a fractional amount of the applied dosage can be recovered by this procedure. To clarify the reason of this deficit of unknown metabolites we followed the hypothesis of oxidative rupture of the heterocyclic ring during metabolism of 1. On the basis of former in vitro results we now were able to identify in quality three oxalic acid derivatives and one acetic acid phenylhydrazide as new metabolites of metamizol in the allantoic fluid (AF) of incubated hen's eggs as well as in human urine by means of GC-MS analysis and comparison with unequivocally synthesized authentic reference compounds. Whereas the oxamazide 7, the phenylhydrazide 8 and N-methyloxamic acid 9 are only present in trace concentrations and therefore cannot account for the deficit in the balance of metabolites, the oxalic acid monohydrazide 11 seems to be excreted in higher amount. But quantitative determination of this new metabolite would be required to answer the open questions concerning the biotransformation of metamizol and thereby to detect new facts about mode of action and side effects of this drug.