Peroxisome Proliferator-Activated Receptor-γ as a Target and Regulator of Epigenetic Mechanisms in Nonalcoholic Fatty Liver Disease.

Peroxisome proliferator-activated receptor-γ (PPARγ) belongs to the superfamily of nuclear receptors that control the transcription of multiple genes. Although it is found in many cells and tissues, PPARγ is mostly expressed in the liver and adipose tissue. Preclinical and clinical studies show that PPARγ targets several genes implicated in various forms of chronic liver disease, including nonalcoholic fatty liver disease (NAFLD). Clinical trials are currently underway to investigate the beneficial effects of PPARγ agonists on NAFLD/nonalcoholic steatohepatitis. Understanding PPARγ regulators may therefore aid in unraveling the mechanisms governing the development and progression of NAFLD. Recent advances in high-throughput biology and genome sequencing have greatly facilitated the identification of epigenetic modifiers, including DNA methylation, histone modifiers, and non-coding RNAs as key factors that regulate PPARγ in NAFLD. In contrast, little is still known about the particular molecular mechanisms underlying the intricate relationships between these events. The paper that follows outlines our current understanding of the crosstalk between PPARγ and epigenetic regulators in NAFLD. Advances in this field are likely to aid in the development of early noninvasive diagnostics and future NAFLD treatment strategies based on PPARγ epigenetic circuit modification.

Molecular Links between Cancer and Metabolic Diseases: New Perspectives and Therapeutic Strategies for Cancer Prevention and Treatment by Targeting Nutritional Patterns and Metabolic Alterations.

Cancer-related mortality is reported to be elevated in cases with metabolic dysfunction [...].

Noncoding RNAs as additional mediators of epigenetic regulation in nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) has emerged as the most common cause of chronic liver disorder worldwide. It represents a spectrum that includes a continuum of different clinical entities ranging from simple steatosis to nonalcoholic steatohepatitis, which can evolve to cirrhosis and in some cases to hepatocellular carcinoma, ultimately leading to liver failure. The pathogenesis of NAFLD and the mechanisms underlying its progression to more pathological stages are not completely understood. Besides genetic factors, evidence indicates that epigenetic mechanisms occurring in response to environmental stimuli also contribute to the disease risk. Noncoding RNAs (ncRNAs), including microRNAs, long noncoding RNAs, and circular RNAs, are one of the epigenetic factors that play key regulatory roles in the development of NAFLD. As the field of ncRNAs is rapidly evolving, the present review aims to explore the current state of knowledge on the roles of these RNA species in the pathogenesis of NAFLD, highlight relevant mechanisms by which some ncRNAs can modulate regulatory networks implicated in NAFLD, and discuss key challenges and future directions facing current research in the hopes of developing ncRNAs as next-generation non-invasive diagnostics and therapies in NAFLD and subsequent progression to hepatocellular carcinoma.

In Vitro Molecular Study of Titanium-Niobium Alloy Biocompatibility.

Titanium dental implants have common clinical applications due to their biocompatibility, biophysical and biochemical characteristics. Although current titanium is thought to be safe and beneficial for patients, there are several indications that it may release toxic metal ions or metal nanoparticles from its alloys into the surrounding environment, which could lead to clinically relevant complications including toxic reactions as well as immune dysfunctions. Hence, an adequate selection and testing of medical biomaterial with outstanding properties are warranted. This study was designed to explore the biocompatibility of smooth titanium-niobium alloy (S_TiNb) versus smooth titanium commercially pure (S_TiCp)-a reference in implantology. All experiments were performed in vitro using human osteoblast-like SaOs-2 and monocyte THP-1 cell lines as models. Cell adhesion and growth morphology were determined by scanning electron microscopy, while cell viability was evaluated using WST-1 assay. Because niobate anions or niobium nanoparticles can be released from implants during biomaterial-cell interaction, potential immunotoxicity of potassium niobate (KNbO3) salt was evaluated by examining both metabolic activity and transcriptomic profiling of treated THP-1 monocytes. The main findings of this study are that S_TiCp and S_TiNb discs do not show an impact on the proliferation and viability of SaOs-2 cells compared to polystyrene surfaces, whereas a significant decrease in THP-1 cells' viability and metabolic activity was observed in the presence of S_TiNb discs compared to the control group. However, no significant changes were found neither at the metabolic activity nor at the transcriptomic level of THP-1 monocytes exposed to KNbO3 salt, suggesting that niobium has no effect on the immune system. Overall, these data imply a possible toxicity of S_TiNb discs toward THP-1 cells, which may not be directly related to niobium but perhaps to the manufacturing process of titanium-niobium alloy. Thus, this limitation must be overcome to make titanium alloy an excellent material for medical applications.

Transcriptional Factors and Epigenetic Mechanisms in Obesity and Related Metabolic Comorbidities.

Recent advances in the study of chromatin remodeling and transcriptional machinery complex dysfunction, and how they drive metabolic-related gene expression have considerably increased our understanding of several molecular processes underlaying obesity and its complications [...].

Dietary Patterns Influence Target Gene Expression through Emerging Epigenetic Mechanisms in Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD) refers to the pathologic buildup of extra fat in the form of triglycerides in liver cells without excessive alcohol intake. NAFLD became the most common cause of chronic liver disease that is tightly associated with key aspects of metabolic disorders, including insulin resistance, obesity, diabetes, and metabolic syndrome. It is generally accepted that multiple mechanisms and pathways are involved in the pathogenesis of NAFLD. Heredity, sedentary lifestyle, westernized high sugar saturated fat diet, metabolic derangements, and gut microbiota, all may interact on a on genetically susceptible individual to cause the disease initiation and progression. While there is an unquestionable role for gene-diet interaction in the etiopathogenesis of NAFLD, it is increasingly apparent that epigenetic processes can orchestrate many aspects of this interaction and provide additional mechanistic insight. Exciting research demonstrated that epigenetic alterations in chromatin can influence gene expression chiefly at the transcriptional level in response to unbalanced diet, and therefore predispose an individual to NAFLD. Thus, further discoveries into molecular epigenetic mechanisms underlying the link between nutrition and aberrant hepatic gene expression can yield new insights into the pathogenesis of NAFLD, and allow innovative epigenetic-based strategies for its early prevention and targeted therapies. Herein, we outline the current knowledge of the interactive role of a high-fat high-calories diet and gene expression through DNA methylation and histone modifications on the pathogenesis of NAFLD. We also provide perspectives on the advancement of the epigenomics in the field and possible shortcomings and limitations ahead.

Epigenetic Regulation of Peroxisome Proliferator-Activated Receptor Gamma Mediates High-Fat Diet-Induced Non-Alcoholic Fatty Liver Disease.

Non-alcoholic fatty liver disease (NAFLD) is highly prevalent in Western countries and has become a serious public health concern. Although Western-style dietary patterns, characterized by a high intake of saturated fat, is considered a risk factor for NAFLD, the molecular mechanisms leading to hepatic fat accumulation are still unclear. In this study, we assessed epigenetic regulation of peroxisome proliferator-activated receptor γ (PPARγ), modifications of gene expression, and lipid uptake in the liver of mice fed a high-fat diet (HFD), and in hepatocyte culture challenged with palmitic acid. Bisulfate pyrosequencing revealed that HFD reduced the level of cytosine methylation in the pparγ DNA promoter. This was associated with increased expression of the hepatic PPARγ, very low-density lipoprotein receptor (VLDLR) and cluster differentiating 36 (CD36), and enhanced uptake of fatty acids and very low-density lipoprotein, leading to excess hepatic lipid accumulation. Furthermore, palmitic acid overload engendered comparable modifications in hepatocytes, suggesting that dietary fatty acids contribute to the pathogenesis of NAFLD through epigenetic upregulation of PPARγ and its target genes. The significance of epigenetic regulation was further demonstrated in hepatocytes treated with DNA methylation inhibitor, showing marked upregulation of PPARγ and its target genes, leading to enhanced fatty acid uptake and storage. This study demonstrated that HFD-induction of pparγ DNA promoter demethylation increased the expression of PPARγ and its target genes, vldlr and cd36, leading to excess lipid accumulation, an important initiating mechanism by which HFD increased PPARγ and lipid accumulation. These findings provide strong evidence that modification of the pparγ promoter methylation is a crucial mechanism of regulation in NAFLD pathogenesis.

The Emerging Role and Promise of Circular RNAs in Obesity and Related Metabolic Disorders.

Circular RNAs (circRNAs) are genome transcripts that are produced from back-splicing of specific regions of pre-mRNA. These single-stranded RNA molecules are widely expressed across diverse phyla and many of them are stable and evolutionary conserved between species. Growing evidence suggests that many circRNAs function as master regulators of gene expression by influencing both transcription and translation processes. Mechanistically, circRNAs are predicted to act as endogenous microRNA (miRNA) sponges, interact with functional RNA-binding proteins (RBPs), and associate with elements of the transcriptional machinery in the nucleus. Evidence is mounting that dysregulation of circRNAs is closely related to the occurrence of a range of diseases including cancer and metabolic diseases. Indeed, there are several reports implicating circRNAs in cardiovascular diseases (CVD), diabetes, hypertension, and atherosclerosis. However, there is very little research addressing the potential role of these RNA transcripts in the occurrence and development of obesity. Emerging data from in vitro and in vivo studies suggest that circRNAs are novel players in adipogenesis, white adipose browning, obesity, obesity-induced inflammation, and insulin resistance. This study explores the current state of knowledge on circRNAs regulating molecular processes associated with adipogenesis and obesity, highlights some of the challenges encountered while studying circRNAs and suggests some perspectives for future research directions in this exciting field of study.

circRNAs Signature as Potential Diagnostic and Prognostic Biomarker for Diabetes Mellitus and Related Cardiovascular Complications.

Circular RNAs (circRNAs) belong to the ever-growing class of naturally occurring noncoding RNAs (ncRNAs) molecules. Unlike linear RNA, circRNAs are covalently closed transcripts mostly generated from precursor-mRNA by a non-canonical event called back-splicing. They are highly stable, evolutionarily conserved, and widely distributed in eukaryotes. Some circRNAs are believed to fulfill a variety of functions inside the cell mainly by acting as microRNAs (miRNAs) or RNA-binding proteins (RBPs) sponges. Furthermore, mounting evidence suggests that the misregulation of circRNAs is among the first alterations in various metabolic disorders including obesity, hypertension, and cardiovascular diseases. More recent research has revealed that circRNAs also play a substantial role in the pathogenesis of diabetes mellitus (DM) and related vascular complications. These findings have added a new layer of complexity to our understanding of DM and underscored the need to reexamine the molecular pathways that lead to this disorder in the context of epigenetics and circRNA regulatory mechanisms. Here, I review current knowledge about circRNAs dysregulation in diabetes and describe their potential role as innovative biomarkers to predict diabetes-related cardiovascular (CV) events. Finally, I discuss some of the actual limitations to the promise of these RNA transcripts as emerging therapeutics and provide recommendations for future research on circRNA-based medicine.

Circular RNAs in hypertension: challenges and clinical promise.

Hypertension (HT), or high blood pressure (BP), is a chronic disease that is common among populations worldwide. The occurrence of HT is one of the leading causes of cardiovascular morbidity and mortality in adults. Although multiple studies have stressed the multifactorial and multigenic nature of HT, uncertainties about its etiology persist, and current diagnostic biomarkers can explain only a small part of the phenotypic variance of BP. Hence, the search for novel biomarkers that enable early disease prevention and guided therapy is warranted. Regulatory circRNAs have emerged as the newest player in HT-related gene networks and hold promise for improving the accuracy of diagnosis. These RNAs are genome products that are formed through back-splicing of specific regions of pre-mRNAs. Evidence suggests that these RNA species are involved in various metabolic diseases. Recent studies have revealed that aberrant expression of circRNAs is relevant to the occurrence and development of HT. Accordingly, circRNAs are proposed as a new generation of predictive biomarkers and potential therapeutic targets for different forms of HT, including pulmonary hypertension and preeclampsia. This paper presents an overview of the findings from current research focusing on the emerging role of circRNAs in the pathogenesis of hypertension. Furthermore, some of the challenges encountered by circRNA studies are highlighted, and perspectives are provided on the future of research in this area.

Circular RNAs as Potential Biomarkers and Therapeutic Targets for Metabolic Diseases.

Epidemiological studies provide evidence of a continuous rise in metabolic diseases throughout industrialized countries. Metabolic diseases are commonly associated with different abnormalities that hold a key role in the emergence and progression of frequent disorders including diabetes mellitus (DM), non-alcoholic fatty liver disease (NAFLD), obesity, metabolic syndrome and cardiovascular diseases. The burden of metabolic diseases is believed to arise through complex interaction between genetic and epigenetic factors, lifestyle changes and environmental exposure to triggering stimuli. The diagnosis and treatment of metabolic disorders continue to be an overwhelming challenge. Thus, the development of novel biomarkers may enhance the accuracy of the diagnosis at an early stage of the disease and allow effective intervention. Over the past decade, progress has been made in exploring the potential role of noncoding RNAs (ncRNAs) in the regulation of gene networks involved in metabolic diseases. A growing body of evidence now suggests that aberrant expression of circular RNAs (circRNAs) is relevant to the occurrence and development of metabolic diseases. Accordingly, circRNAs are proposed as predictive biomarkers and potential therapeutic targets for these diseases. As the field of circRNAs is rapidly evolving and knowledge is increasing, the present paper provides current understanding of the regulatory roles of these RNA species mainly in the pathogenesis of DM, NAFLD and obesity. Furthermore, some of the limitations to the promise of circRNAs and perspectives on their future research are discussed.

Epigenetic Regulation of ATP-Binding Cassette Protein A1 (ABCA1) Gene Expression: A New Era to Alleviate Atherosclerotic Cardiovascular Disease.

The most important function of high density lipoprotein (HDL) is its ability to remove cholesterol from cells and tissues involved in the early stages of atherosclerosis back to the liver for excretion. The ATP-binding cassette transporters ABCA1 and ABCG1 are responsible for the major part of cholesterol efflux to HDL in macrophage foam cells. Thus, promoting the process of reverse cholesterol transport (RCT) by upregulating mainly ABCA1 remains one of the potential targets for the development of new therapeutic agents against atherosclerosis. Growing evidence suggests that posttranscriptional regulation of HDL biogenesis as well as modulation of ABCA1 expression are under the control of several genetic and epigenetic factors such as transcription factor (TFs), microRNAs (miRNAs) and RNA-binding proteins (RBPs).These factors may act either individually or in combination to orchestrate ABCA1 expression. Complementary to our recent work, we propose an exploratory model for the potential molecular mechanism(s) underlying epigenetic signature of ABCA1 gene regulation. Such a model may hopefully provide the basic framework for understanding the epigenetic regulation of RCT and contribute to the development of novel therapeutic strategies to alleviate the burden of cardiovascular diseases (CVD).

Epigenetic regulation of genes involved in the reverse cholesterol transport through interaction with miRNAs.

microRNAs (miRNAs) are a group of small non-coding RNA molecules known to regulate target genes at the post-transcriptional level. miRNAs are implicated in the regulation of multiple pathophysiological processes including dyslipidemia, a major risk factor for atherosclerosis. Emerging evidence suggests that miRNAs act as a novel class of epigenetic regulators of high-density lipoproteins cholesterol (HDL-C) from synthesis to clearance contributing remarkably to the pathogenesis of atherosclerosis. Accumulating studies have revealed that miRNAs such as miR-33, miR-27, miR-144, miR-758 and miR-20 are involved in the post-transcriptional control of ABCA1, ABCG1 and SCARB1 genes regulatory network of the reverse cholesterol transport (RCT). These miRNAs have been shown to be central players in the impairment of RCT pathway leading to the development of atherosclerosis. In this article, we present most recent understanding of involvement of relevant miRNAs in different steps of HDL metabolism and RCT pathway. We also discuss some of the actual limitations to the promise of these miRNAs and perspectives on their translation to clinical settings.

The "forgotten" modified lipoprotein subspecies.

Insights from preclinical and clinical studies have attempted to highlight the importance of modified lipoprotein particles in the pathogenesis of cardiovascular diseases (CVD). However, evidence is not conclusive. Since there is a relative dearth of clinical research in collecting useful information from traditional advanced lipoproteins testing, this present editorial introduces the aim of a special issue on modified lipoproteins as potential biomarkers for CVD. This issue aims at gathering a selection of insightful articles that address major challenges related to potential clinical use of modified lipoproteins as new CVD biomarkers. The editors seek to promote better mobilization of lipoproteins measurement for the pursuit of sustainable CVD clinical outcome and development of potential biomarkers. Knowledge and progress in this particular field will certainly help answering questions about clinical relevance of circulating modified lipoprotein subspecies and their potential use for better patient care and disease prevention. We hope that, when taken together, the focus on modified lipoproteins will stimulate new vision and reveal study key aspects for better clinical data outcome and more effective therapeutic strategies.

Human formyl peptide receptor 1 (FPR1) c.32C>T SNP is associated with decreased soluble E-selectin levels.

AIMS: The human formyl peptide receptor (FPR) is a G protein-coupled chemoattractant receptor that is thought to mediate inflammatory responses. The FPR1 gene is highly polymorphic. In a recent study, the FPR1 c.32C>T SNP, resulting in the amino-acid substitution I11T, was reported to be significantly associated with C-reactive protein levels. Therefore, this study sought to determine if the impact of such a genetic variation extends to other clinical parameters associated with inflammation, including cytokines, adhesion molecules and inflammatory markers. MATERIALS & METHODS: This study was carried out on a subsample of 325 adults selected from the STANISLAS cohort study. The FPR1 c.32C>T SNP was genotyped using PCR amplification followed by restriction enzyme digestion. Anthropometric measurements and biochemical profiles were assessed for each individual. RESULTS: The allele frequencies of FPR1 c.32C>T were 0.74 for the 32C allele and 0.26 for the 32T allele. Genotype frequencies were 0.55 for C/C, 0.38 for C/T and 0.07 for T/T. After adjusting for age, sex, BMI, alcohol and cigarette consumption, oral contraceptive, antibiotics and anti-inflammatory drug use, statistical analysis (under a recessive model of inheritance) demonstrated that serum E-selectin levels were 68% lower in individuals homozygous for T/T than in those with C/T or C/C genotypes (p = 0.001). However, no significant correlations were found for C-reactive protein or the other 18 tested clinical parameters that were analyzed in this study. CONCLUSION: The FPR1 c.32C>T SNP may be associated with E-selectin levels in the French population. Although of importance, these findings need confirmation in larger studies.

Genetic profiling of human cell lines used as in vitro model to study cardiovascular pathophysiology and pharmacotoxicology.

BACKGROUND: Cell lines are widely used to monitor drug pharmacokinetics and pharmacodynamics and to investigate a number of biochemical mechanisms. However, little is known about the genetic profile of these in vitro models. OBJECTIVES: To analyze genetic profile of Thp1, U937, HL60, K562, HepG2, Kyn2, and Caco2 human cell lines with a focus on genetic variations within genes involved in the development of cardiovascular pathologies and drug treatment response. METHODS: Multiplex polymerase chain reaction (PCR), PCR-restriction fragment length polymorphism and TaqMan assays were used to genotype 120 polymorphisms within 68 genes previously shown to be involved in various processes such as inflammation, lipid metabolism, and blood pressure. RESULTS: We provide here a list of potential polymorphisms known to be associated with cardiovascular disease. Our results show that the seven cell lines examined carry several of these mutations within genes of interest. Due to the abundance of these variations, only two examples will be given in this abstract. For instance, U937 cells are homozygous for APOE varepsilon4, a mutant associated with higher susceptibility to cardiovascular diseases and lower response to statins. Our study also showed that deletion in intron 16 of the ACE gene, which is associated with susceptibility to hypertension and variation of response to ACE inhibitors, can be found in all considered cells but Kyn2 cells. CONCLUSION: We provide here a data bank of different cell lines genetic profile. In our opinion, this useful information may bring insights into the design and choice of an adequate in vitro model and may help to explain mysterious discrepancies in data from different laboratories.

Macrophage culture as a suitable paradigm for evaluation of synthetic vitreous fibers.

The ultimate goal of toxicologic investigation of synthetic vitreous fibers (SVFs) is to provide essential input for the assessment of human risk to their exposure. Toxicity of mineral fibers is usually evaluated by testing biopersistence in rodent model. However, a cellular model would be much appreciated in order to reduce, refine, and replace animal models. Pulmonary disorders triggered by inhalation of occupational or environmental mineral particulates can be the endpoints of a chronic inflammatory process in which alveolar macrophages (AMs) play a crucial role. Depending on the type of SVF involved, phagocytosis of fiber leads to activation of macrophages, resulting in release of fiber components and potent mediators, such as reactive oxygen or nitrogen species and cytokines. As a matter of fact, macrophages should be the cells of choice since SVF toxicity is the consequence of fibers and alveolar macrophages interaction. Today, monocytes and macrophages culture are firmly established as a paradigm in toxicology when several endpoints are assayed in macrophages: (1) fiber durability, (2) fiber surface changes, (3) oxidative stress and genotoxicity in macrophage, and (4) macrophage cell viability and apoptosis. This article is a review of up-to-date knowledge of in vitro studies involving macrophages, and assesses endpoints of macrophage toxicity with an emphasis on (1) dissolution, (2) scanning electron microscopy analysis, (3) cytotoxicity, and (4) gene expression.

Multifunctional antimicrobial peptides: therapeutic targets in several human diseases.

Antimicrobial peptides have emerged as promising agents against antibiotic-resistant pathogens. They represent essential components of the innate immunity and permit humans to resist infection by microbes. These gene-encoded peptides are found mainly in phagocytes and epithelial cells, showing a direct activity against a wide range of microorganisms. Their role has now broadened from that of simply endogenous antibiotics to multifunctional mediators, and their antimicrobial activity is probably not the only primary function. Although antimicrobial peptide deficiency, dysregulation, or overproduction is not known to be a direct cause of any single human disease, numerous studies have now provided compelling evidence for their involvement in the complex network of immune responses and inflammatory diseases, thereby influencing diverse processes including cytokine release, chemotaxis, angiogenesis, wound repair, and adaptive immune induction. The purpose of this review is to highlight recent literature, showing that antimicrobial peptides are associated with several human conditions including infectious and inflammatory diseases, and to discuss current clinical development of peptide-based therapeutics for future use.

Genetic determinants of blood pressure regulation.

Hypertension is a multifactorial disorder that probably results from the inheritance of a number of susceptibility genes and involves multiple environmental determinants. Existing evidence suggests that the genetic contribution to blood pressure variation is about 30-50%. Although a number of candidate genes have been studied in different ethnic populations, results from genetic analysis are still inconsistent and specific causes of hypertension remain unclear. Furthermore, the abundance of data in the literature makes it difficult to piece together the puzzle of hypertension and to define candidate genes involved in the dynamic of blood pressure regulation. In this review, we attempt to highlight the genetic basis of hypertension pathogenesis, focusing on the most important existing genetic variations of candidate genes and their potential role in the development of this disease. Our objective is to review current knowledge and discuss limitations to clinical applications of genotypic information in the diagnosis, evaluation and treatment of hypertension. Finally, some principles of pharmacogenomics are presented here along with future perspectives of hypertension.

Keratinocyte production of cathelicidin provides direct activity against bacterial skin pathogens.

Immune defense at an interface with the external environment reflects the functions of physical and chemical barriers provided by epithelial and immune cells. Resident epithelial cells, such as keratinocytes, produce numerous peptides with direct antimicrobial activity but also provide a physical barrier against invading pathogens and signal the recruitment of circulating immune cells, such as neutrophils. Antimicrobial peptides such as cathelicidin are produced constitutively by neutrophils and are inducible in keratinocytes in response to infection. The multiplicity of antimicrobial peptides and their cellular sources has resulted in an incomplete understanding of the role of cathelicidin production by epithelial cells in cutaneous immune defense. Therefore, this study sought to evaluate keratinocyte antimicrobial activity and the potential contribution of keratinocyte cathelicidin to host protection against two leading human skin pathogens. Wild-type mice and those with a targeted deletion of the cathelicidin gene, Cnlp, were rendered neutropenic prior to cutaneous infection. Interestingly, Cnlp-deficient mice remained more susceptible to group A streptococcus infection than mice with Cnlp intact, suggesting the involvement of epithelial cell-derived cathelicidin in host immune defense. Keratinocytes were then isolated in culture and found to inhibit the growth of Staphylococcus aureus, an effect that was partially dependent on their ability to synthesize and activate cathelicidin. Further, lentivirus-mediated delivery of activated human cathelicidin enhanced keratinocyte antimicrobial activity. Combined, these data illustrate the potential contribution of keratinocyte cathelicidin to the innate immune defense of skin against bacterial pathogens and highlight the need to consider epithelial antimicrobial function in the diagnosis and therapy of skin infection.