Inhibition of RNase 7 by RNase inhibitor promotes inflammation and Staphylococcus aureus growth: Implications for atopic dermatitis.

BACKGROUND: The antimicrobial ribonuclease RNase 7 is abundantly expressed in the epidermis of lesional skin of atopic dermatitis (AD). Host RNase inhibitor (RI) binds to RNase 7 and blocks its ribonuclease activity. This study aimed to evaluate the impact of RNase 7-RI interactions on AD. METHODS: Cultured human primary keratinocytes, with siRNA-mediated downregulation of RNase 7 and RI, were stimulated with the synthetic RNA polyinosinic-polycytidylic acid (poly I:C). Induction of proinflammatory mediators was analyzed by real-time PCR and ELISA. RI expression in AD non-lesional and lesional skin biopsies and healthy controls was analyzed by real-time PCR and immunostaining. RI protein release in vivo on the AD skin surface was determined by western blot. Antimicrobial and ribonuclease assays were used to investigate the functional role of RI. RESULTS: RNase 7 inhibited the RNA-induced expression of proinflammatory mediators in keratinocytes. Accordingly, downregulation of RNase 7 in keratinocytes enhanced RNA-mediated induction of proinflammatory mediators, whereas downregulation of RI had the opposite effect. RI was released by damaged keratinocytes and epidermis. In vivo expression and release of RI on the skin surface were enhanced in lesional AD skin. Rinsing solution from the surface of lesional AD skin blocked the ribonuclease activity of RNase 7. The anti-Staphylococcus aureus activity of RNase 7 was abrogated by RI. CONCLUSIONS: Our data suggest a novel role of RI as a trigger factor of inflammation in AD by blocking the ribonuclease and antimicrobial activity of RNase 7, thereby enhancing RNA-mediated inflammation and S. aureus growth.

3D-printed wound dressing platform for protein administration based on alginate and zinc oxide tetrapods.

Wound treatment requires a plethora of independent properties. Hydration, anti-bacterial properties, oxygenation and patient-specific drug delivery all contribute to the best possible wound healing. Three-dimensional (3D) printing has emerged as a set of techniques to realize individually adapted wound dressings with open porous structure from biomedically optimized materials. To include all the desired properties into the so-called bioinks is still challenging. In this work, a bioink system based on anti-bacterial zinc oxide tetrapods (t-ZnO) and biocompatible sodium alginate is presented. Additive manufacturing of these hydrogels with high t-ZnO content (up to 15 wt.%) could be realized. Additionally, protein adsorption on the t-ZnO particles was evaluated to test their suitability as carriers for active pharmaceutical ingredients (APIs). Open porous and closed cell printed wound dressings were tested for their cell and skin compatibility and anti-bacterial properties. In these categories, the open porous constructs exhibited protruding t-ZnO arms and proved to be anti-bacterial. Dermatological tests on ex vivo skin showed no negative influence of the alginate wound dressing on the skin, making this bioink an ideal carrier and evaluation platform for APIs in wound treatment and healing.

Antimicrobial skin peptides in premature infants: Comparison with term infants and impact of perinatal factors.

Introduction: Preterm infants have an immature epidermis barrier function that may lead to an increased permeability to pathogens. On the surface of the human skin, antimicrobial peptides (AMPs) are important molecules of the innate immune system, have broad antimicrobial properties, and provide an essential role in integrity of the microbiome. Given the marked susceptibility of preterm infants to infection, we hypothesize a decreased expression of AMPs on the skin of preterm infants. Materials and methods: In a prospective single-center study with 35 preterm and 20 term infants, we analyzed skin rinsing probes for the presence of the AMPs psoriasin (S100A7) and ribonuclease 7 (RNase 7) via enzyme-linked immunosorbent assay. Samples were taken from preterm infants < 34 0/7 weeks gestational age (mean ± SD gestational age, 28.8 ± 2.4 weeks) on days 0, 7, 14, and 28 after birth. Term infants (> 36 6/7 weeks) (controls) were washed on days 0 and 28. Results: Psoriasin and RNase 7 were both expressed on skin of preterm and term infants and increased in concentration significantly over time. RNase 7 was more expressed in term infants on day 0 [preterm = 1.1 (0.7-2.9) vs. term = 2.0 (1.1-3.4) ng/ml, p = 0.017]. On day 28, premature infants showed higher values of psoriasin [preterm = 10.9 (5.6-14.2) vs. term = 6.3 (3.4-9.0) ng/ml, p < 0.001]. Notably, preterm infants with infectious or inflammatory context driven by histological proof of chorioamnionitis and early-onset or late-onset sepsis had higher concentrations of psoriasin as compared with non-affected preterm infants. After exclusion of infants with inflammatory hit, median concentrations of RNase 7 and psoriasin did not differ between preterm and full-term infants on days 0 and 28. Discussion: Psoriasin and RNase 7 concentrations increase over time on the skin of newborn infants and seem to play a role in the first defense against infection. This is of particularly interest as the role of AMPs on a maturing skin microbiome and its possible new prevention strategies is unclear and needs to be determined.

The influence of the commensal skin bacterium Staphylococcus epidermidis on the epidermal barrier and inflammation: Implications for atopic dermatitis.

The skin microbiota is a crucial component in maintaining cutaneous barrier function. Staphylococcus epidermidis is considered as a beneficial commensal member of the cutaneous microbiota promoting skin health. However, S. epidermidis is also frequently detectable in the skin of patients with the inflammatory skin disease atopic dermatitis (AD) and some studies reported a significantly higher presence of S. epidermidis in severe AD as compared to mild AD. Therefore, this study aimed to analyse the impact of S. epidermidis on the expression of cutaneous inflammatory mediators and skin barrier molecules. Various S. epidermidis skin-derived isolates activated the proinflammatory transcription factor NF-kappaB and induced expression of AD-associated proinflammatory cytokines in human primary keratinocytes and 3D skin equivalents. Skin barrier molecules such as filaggrin were downregulated by S. epidermidis. In general, AD-derived S. epidermidis strains elicited a higher response than strains derived from the skin of healthy individuals. Taken together, our results provide further evidence that the abundance of S. epidermidis in AD may trigger the inflammatory scenario associated with this disease.

Skin microbiota analysis in patients with anorexia nervosa and healthy-weight controls reveals microbial indicators of healthy weight and associations with the antimicrobial peptide psoriasin.

Anorexia nervosa (AN), a psychiatric condition defined by low body weight for age and height, is associated with numerous dermatological conditions. Yet, clinical observations report that patients with AN do not suffer from infectious skin diseases like those associated with primary malnutrition. Cell-mediated immunity appears to be amplified in AN; however, this proinflammatory state does not sufficiently explain the lower incidence of infections. Antimicrobial peptides (AMPs) are important components of the innate immune system protecting from pathogens and shaping the microbiota. In Drosophila melanogaster starvation precedes increased AMP gene expression. Here, we analyzed skin microbiota in patients with AN and age-matched, healthy-weight controls and investigated the influence of weight gain on microbial community structure. We then correlated features of the skin microbial community with psoriasin and RNase 7, two highly abundant AMPs in human skin, to clarify whether an association between AMPs and skin microbiota exists and whether such a relationship might contribute to the resistance to cutaneous infections observed in AN. We find significant statistical correlations between Shannon diversity and the highly abundant skin AMP psoriasin and bacterial load, respectively. Moreover, we reveal psoriasin significantly associates with Abiotrophia, an indicator for the healthy-weight control group. Additionally, we observe a significant correlation between an individual's body mass index and Lactobacillus, a microbial indicator of health. Future investigation may help clarify physiological mechanisms that link nutritional intake with skin physiology.

Skin Care Product Rich in Antioxidants and Anti-Inflammatory Natural Compounds Reduces Itching and Inflammation in the Skin of Atopic Dermatitis Patients.

The atopic dermatitis (AD) complex pathogenesis mechanism reveals marked changes of certain signaling factors as well as some morphological alterations in the epidermis. Reduced resilience against environmental factors and oxidative stress often makes the treatment with corticosteroids or tacrolismus ointments indispensable. In view of the correlation between oxidative stress and AD pathological factors, antioxidants can be incorporated into AD management strategies. This study investigates a curly kale, apple and green tea-containing natural extract rich in antioxidants for its effects on signaling inflammatory molecules and skin barrier enhancement in human epidermal keratinocytes- (NHEKs) based cell assays. Furthermore, the skin penetration on porcine ears was measured ex vivo using Raman micro spectroscopy. Finally, in a double-blind half-side, placebo-controlled clinical study, the effects of a formulation containing this extract were analyzed for the influence of lesion severity, epidermal barrier function, and pruritus in mild to moderately AD patients. Summarizing our results: The extract reduces expression of inflammatory cytokines in keratinocytes and increases barrier-related molecules. The verum formulation with a very high antioxidant capacity used in AD patients with mild to moderate lesions reduces itching, local SCORAD, and improves barrier function and the hydration of skin lesions.

Staphylococcus epidermidis-Derived Protease Esp Mediates Proteolytic Activation of Pro‒IL-1ß in Human Keratinocytes.

The Gram-positive bacterium Staphylococcus epidermidis (SE) is an abundant skin commensal. It plays an important role in cutaneous defense by activation of IL-1 signaling. In keratinocytes (KCs), SE induces the release of mature IL-1ß. IL-1ß serves as an important cytokine of host defense. It contains an N-terminal prodomain that has to be cleaved off to generate active mature IL-1ß. Typically, the processing and release of IL-1ß are associated with inflammasome assembly and activation of the protease caspase-1. In this study, we report that the bacterial challenge of KCs with SE induced the release of mature IL-1ß in a caspase-1‒independent manner. Instead, the SE-derived serine protease Esp was identified as a pro‒IL-1ß‒processing factor leading to a proteolytic maturation of active IL-1ß. Esp production and secretion by various SE strains correlated with their capacity to induce the release of mature IL-1ß in human primary KCs. Reconstitution of Esp-lacking SE strains with Esp enhanced their capacity to induce IL-1ß release in KCs and skin. Intracellular abundance of pro‒IL-1ß and cytotoxic effects of SE suggest a release of pro‒IL-1ß during injury, followed by extracellular Esp-mediated processing to mature IL-1ß. These findings provide further insights into how a skin commensal interacts with KCs to activate cutaneous host innate defense.

Staphylococcus aureus Activates the Aryl Hydrocarbon Receptor in Human Keratinocytes.

Staphylococcus aureus is an important pathogen causing various infections, including - as most frequently isolated bacterium - cutaneous infections. Keratinocytes as the first barrier cells of the skin respond to S. aureus by the release of defense molecules such as cytokines and antimicrobial peptides. Although several pattern recognition receptors expressed in keratinocytes such as Toll-like and NOD-like receptors have been reported to detect the presence of S. aureus, the mechanisms underlying the interplay between S. aureus and keratinocytes are still emerging. Here, we report that S. aureus induced gene expression of CYP1A1 and CYP1B1, responsive genes of the aryl hydrocarbon receptor (AhR). AhR activation by S. aureus was further confirmed by AhR gene reporter assays. AhR activation was mediated by factor(s) <2 kDa secreted by S. aureus. Whole transcriptome analyses and real-time PCR analyses identified IL-24, IL-6, and IL-1beta as cytokines induced in an AhR-dependent manner in S. aureus-treated keratinocytes. AhR inhibition in a 3D organotypic skin equivalent confirmed the crucial role of the AhR in mediating the induction of IL-24, IL-6, and IL-1beta upon stimulation with living S. aureus. Taken together, we further highlight the important role of the AhR in cutaneous innate defense and identified the AhR as a novel receptor mediating the sensing of the important skin pathogen S. aureus in keratinocytes.

Bladder management in individuals with spinal cord injury or disease during and after primary rehabilitation: a retrospective cohort study.

PURPOSE: The purpose of this study was to investigate the course of bladder evacuation and the predictors of intermittent self-catheterization (ISC) in individuals with neurogenic lower urinary tract dysfunction (NLUTD) during and after primary rehabilitation. METHODS: The patient database of a single spinal cord injury rehabilitation center was screened for patients with NLUTD admitted for primary rehabilitation. Patient characteristics and bladder evacuation details were collected during and after rehabilitation. Binary logistic regression analysis was used to evaluate predictors of ISC: sex, age > 65 years, injury severity, and bladder capacity ≥ 400 ml. RESULTS: Data of 255 men (74.3%) and 88 women (25.7%) with a mean age of 54 ± 19 years were analyzed. Early in rehabilitation, 21.6% of the evaluated individuals used ISC. In 17.8%, the bladder was evacuated by transurethral catheterization (TUC). The proportion of TUC decreased during rehabilitation and dropped below 2% at the last follow-up. In contrast, the proportion of ISC and suprapubic catheterization (SPC) increased to 28% and 12.8%, respectively, during rehabilitation. These proportions increased further thereafter and reached 37.7% and 18.6% for ISC and SPC, respectively. Age and injury severity were significant (p ≤ 0.041) negative predictors, whereas male sex and above-average bladder capacity were positive predictors of ISC. CONCLUSIONS: There is a shift toward bladder evacuation by ISC and SPC during and after primary rehabilitation. Bladder evacuation by ISC, regarded as the gold standard, is less common in individuals > 65 years or with high-level tetraplegia. The optimal bladder evacuation method needs to be established individually, considering all medical and psychosocial factors rather than simply following a guideline.

Platelet-Released Growth Factors Influence Wound Healing-Associated Genes in Human Keratinocytes and Ex Vivo Skin Explants.

Platelet-released growth factors (PRGFs) or other thrombocyte concentrate products, e.g., Platelet-Rich Fibrin (PRF), have become efficient tools of regenerative medicine in many medical disciplines. In the context of wound healing, it has been demonstrated that treatment of chronic or complicated wounds with PRGF or PRF improves wound healing in the majority of treated patients. Nevertheless, the underlying cellular and molecular mechanism are still poorly understood. Therefore, we aimed to analyze if PRGF-treatment of human keratinocytes caused the induction of genes encoding paracrine factors associated with successful wound healing. The investigated genes were Semaphorin 7A (SEMA7A), Angiopoietin-like 4 (ANGPLT4), Fibroblast Growth Factor-2 (FGF-2), Interleukin-32 (IL-32), the CC-chemokine-ligand 20 (CCL20), the matrix-metalloproteinase-2 (MMP-2), the chemokine C-X-C motif chemokine ligand 10 (CXCL10) and the subunit B of the Platelet-Derived Growth Factor (PDGFB). We observed a significant gene induction of SEMA7A, ANGPLT4, FGF-2, IL-32, MMP-2 and PDGFB in human keratinocytes after PRGF treatment. The CCL20- and CXCL10 gene expressions were significantly inhibited by PRGF therapy. Signal transduction analyses revealed that the PRGF-mediated gene induction of SEMA7A, ANGPLT4, IL-32 and MMP-2 in human keratinocytes was transduced via the IL-6 receptor pathway. In contrast, EGF receptor signaling was not involved in the PRGF-mediated gene expression of analyzed genes in human keratinocytes. Additionally, treatment of ex vivo skin explants with PRGF confirmed a significant gene induction of SEMA7A, ANGPLT4, MMP-2 and PDGFB. Taken together, these results describe a new mechanism that could be responsible for the beneficial wound healing properties of PRGF or related thrombocytes concentrate products such as PRF.

Platelet-Released Growth Factors Induce Genes Involved in Extracellular Matrix Formation in Human Fibroblasts.

Platelet concentrate products are increasingly used in many medical disciplines due to their regenerative properties. As they contain a variety of chemokines, cytokines, and growth factors, they are used to support the healing of chronic or complicated wounds. To date, underlying cellular mechanisms have been insufficiently investigated. Therefore, we analyzed the influence of Platelet-Released Growth Factors (PRGF) on human dermal fibroblasts. Whole transcriptome sequencing and gene ontology (GO) enrichment analysis of PRGF-treated fibroblasts revealed an induction of several genes involved in the formation of the extracellular matrix (ECM). Real-time PCR analyses of PRGF-treated fibroblasts and skin explants confirmed the induction of ECM-related genes, in particular transforming growth factor beta-induced protein (TGFBI), fibronectin 1 (FN1), matrix metalloproteinase-9 (MMP-9), transglutaminase 2 (TGM2), fermitin family member 1 (FERMT1), collagen type I alpha 1 (COL1A1), a disintegrin and metalloproteinase 19 (ADAM19), serpin family E member 1 (SERPINE1) and lysyl oxidase-like 3 (LOXL3). The induction of these genes was time-dependent and in part influenced by the epidermal growth factor receptor (EGFR). Moreover, PRGF induced migration and proliferation of the fibroblasts. Taken together, the observed effects of PRGF on human fibroblasts may contribute to the underlying mechanisms that support the beneficial wound-healing effects of thrombocyte concentrate products.

Antimicrobial peptides and proteins: Interaction with the skin microbiota.

The cutaneous microbiota comprises all living skin microorganisms. There is increasing evidence that the microbiota plays a crucial role in skin homeostasis. Accordingly, a dysbiosis of the microbiota may trigger cutaneous inflammation. The need for a balanced microbiota requires specific regulatory mechanisms that control and shape the microbiota. In this review, we highlight the present knowledge suggesting that antimicrobial peptides (AMPs) may exert a substantial influence on the microbiota by controlling their growth. This is supported by own data showing the differential influence of principal skin-derived AMPs on commensal staphylococci. Vice versa, we also illuminate how the cutaneous microbiota interacts with skin-derived AMPs by modulating AMP expression and how microbiota members protect themselves from the antimicrobial activity of AMPs. Taken together, the current picture suggests that a fine-tuned and well-balanced AMP-microbiota interplay on the skin surface may be crucial for skin health.

Free human DNA attenuates the activity of antimicrobial peptides in atopic dermatitis.

BACKGROUND: The high susceptibility of AD patients to microbial skin infections has been attributed to a deficient antimicrobial peptide (AMP) expression, which is contradicted by a growing amount of recent studies clearly demonstrating that AMP expression is not impaired in lesional skin of AD patients. The reasons for the high susceptibility of AD patients to microbial infections are still unknown. METHODS: The influence of self-DNA on the antimicrobial activity of RNase 7, LL-37, and hBD2 has been investigated using antibacterial and antiviral assays. The amount of self-DNA on skin has been analyzed by skin rinsings and subsequent quantification using dsDNA assays. DNA source was identified by qPCR. RESULTS: Complex formation of the AMPs with self-DNA significantly impaired their antibacterial activity against Staphylococcus aureus and their antiviral activity against HSV-1. The inhibition of the antibacterial activity was dependent on the DNA concentration but not on the length of the DNA molecules. Of note, we detected significant higher amounts of cell-free self-DNA in skin rinses taken from lesional AD skin compared to skin rinses from non-lesional skin and from normal skin of healthy donors. Consequently, rinse solution from AD lesional skin prevented antibacterial activity of LL-37. CONCLUSION: Our study indicates that extracellular self-DNA is released in considerable amounts in AD skin lesions and AMP-self-DNA-complex formation leads to a significant loss of antibacterial and antiviral activity in atopic dermatitis. Studies on strategies to reduce the amount of extracellular DNA in AD are needed to identify possible methods relevant in clinical settings.

Expression of epidermal antimicrobial peptides is increased in tinea pedis.

BACKGROUND: Tinea pedis is often chronic or recurrent, but not all individuals are equally susceptible to this infection. Dermatophytes are able to induce the expression of antimicrobial peptides and proteins (AMPs) in human keratinocytes and certain AMPs can inhibit the growth of dermatophytes. OBJECTIVE: The focus of this study was to analyse the secretion of relevant AMPs, especially RNase 7, human beta-defensin-2 (hBD-2) and the S-100 protein psoriasin (S100A7), in patients with confirmed tinea pedis. METHODS: To verify the diagnosis, skin scales were obtained from all patients (n = 13) and the dermatophytes were identified by potassium hydroxide mount, culture and molecular analysis. To determine the AMP concentrations, the lesional skin area of the foot was rinsed with a buffer that was subsequently analysed by ELISA. The corresponding area of the other unaffected foot as well as defined healthy skin areas of the forearm and forehead and samples from age and gender-matched healthy volunteers served as controls. RESULTS: In tinea pedis patients the AMP concentrations were higher in lesional skin than in non-lesional skin and in healthy skin of controls. In particular, concentrations of hBD-2 and psoriasin were significantly elevated. CONCLUSIONS: The induction of AMPs in tinea pedis might be triggered directly by the dermatophytes; furthermore, attendant inflammation and/or differentiation processes may play a role. Our results indicate that there is no defect in the constitutive expression and induction of the analysed AMPs by dermatophytes in the epidermis of affected patients. However, it is not known why the elevated AMP concentrations fail to efficiently combat dermatophyte growth.

Are 200 units of onabotulinumtoxin A sufficient for the suppression of neurogenic detrusor overactivity in individuals with established 300-unit botulinum toxin treatment? A retrospective cohort study.

STUDY DESIGN: Retrospective cohort study. OBJECTIVES: To investigate the effects of 200-unit onabotulinumtoxin A detrusor injections on neurogenic detrusor overactivity (NDO) in patients who have previously been treated with 300-unit injections. SETTING: Tertiary urologic referral center in Switzerland. METHODS: The patient database was screened for patients with NDO as a result of chronic (≥ 12 months) spinal cord injury (SCI), who had been treated with 300- followed by 200-unit onabotulinumtoxin A detrusor injections. Patient characteristics, bladder management data and concurrent bladder medication as well as urodynamic data were collected. The percent changes in the urodynamic parameters from the 300- to the 200-unit treatment phase were calculated to test for non-inferiority of the 200-unit treatment. RESULTS: The data of 61 individuals with a mean age of 44 ± 15 years (range 18-73 years) and a mean 13.2 ± 9.5 years (range 2-43 years) since SCI were analyzed. The 200-unit treatment was not inferior regarding the urodynamic parameters compared to the 300-unit treatment. Furthermore, the proportion of patients with urinary incontinence was similar for both doses. There was no significant difference in the number of daily bladder evacuations (p = 0.13) or used incontinence pads (p = 0.43) between the two dosage phases. Moreover, there was no significant (p = 0.19) increase in the use of concurrent NDO medication (antimuscarinics or mirabegron) during the 200-unit treatment. CONCLUSIONS: The treatment of NDO with 200 units of onabotulinumtoxin A was not inferior to a 300-unit treatment regarding urodynamic parameters in patients with chronic SCI.

Skin microbiota analysis in human 3D skin models-"Free your mice".

In the May issue of Experimental Dermatology 2018, we published a review article focusing on human 3D skin models in the context of microbiota research. The principal intention was to provide an overview of present and future concepts to use skin models in microbiota analyses. With the present viewpoint, we would like to draw the reader's attention again to the use of human skin models in microbiota research with the aim to highlight the benefits and necessity of human skin models to analyse the human skin-microbiota interaction. This is accompanied by a critical view on mice models that often are not suitable to analyse the functional impact of the human skin microbiota. In addition, we present novel and future concepts highlighting the benefits of human 3D skin models in microbiota research.

Real-World Effects of Mirabegron in Patients with Chronic Neurogenic Detrusor Overactivity - A Retrospective Cohort Study.

PURPOSE: To investigate the tolerability and the effects of the ß-3-adrenoceptor-agonist mirabegron on urinary incontinence and urodynamic parameters in patients with chronic neurogenic detrusor overactivity (NDO). PATIENTS AND METHODS: The patient database of a spinal cord injury rehabilitation center in Switzerland was screened for patients with chronic (>12 months) NDO, who had been prescribed mirabegron. Patient characteristics, data regarding bladder management, urinary incontinence and concurrent medication for NDO as well as urodynamic parameters were collected retrospectively. The changes in the urodynamic parameters and the occurrence of urinary incontinence over time were investigated. RESULTS: The data of 63 patients with a median age of 48 years and a median NDO duration of 8.9 years at the initiation of the mirabegron treatment were analyzed. A median 3.0 and 12.7 months had elapsed from the initiation of the mirabegron therapy to the first and second follow-up evaluation, respectively. The majority of patients (73%) received mirabegron in combination with an established antimuscarinic or onabotulinum toxin therapy. The number of patients suffering from urinary incontinence decreased significantly (p≤0.005) from 60.3% (95% CI 47.2/72.4%) to 38.1% (95% CI 23.6/54.4%). Furthermore, the maximum detrusor pressure during the storage phase was significantly (p≤0.04) lower at the second follow-up evaluation (29.5cmH2O, 95% CI 22/40cmH2O) compared to before the mirabegron treatment (35cmH2O, 95% CI 29/41cmH2O). The bladder capacity and detrusor compliance were significantly (p≤0.005) increased during the mirabegron treatment. No patient had discontinued the mirabegron treatment as a result of side effects. CONCLUSION: Mirabegron demonstrated a clinically relevant effect and a good safety profile. Concomitant treatment of NDO with mirabegron may allow reduction in the dose of antimuscarinic medication and thus, improve the long-term persistence of NDO treatment.

Platelet-Released Growth Factors and Platelet-Rich Fibrin Induce Expression of Factors Involved in Extracellular Matrix Organization in Human Keratinocytes

Platelet-released growth factor (PRGF) is a thrombocyte concentrate lysate which, like its clinically equivalent variations (e.g., Vivostat PRF® (platelet-rich fibrin)), is known to support the healing of chronic and hard-to-heal wounds. However, studies on the effect of PRGF on keratinocytes remain scarce. This study aims to identify genes in keratinocytes that are significantly influenced by PRGF. Therefore, we performed a whole transcriptome and gene ontology (GO) enrichment analysis of PRGF-stimulated human primary keratinocytes. This revealed an increased expression of genes involved in extracellular matrix (ECM) organization. Real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) analysis confirmed the PRGF-mediated induction of selected ECM-related factors such as transforming growth factor beta-induced protein, fibronectin 1, matrix metalloproteinase-9, transglutaminase 2, fermitin family member 1, collagen type I alpha 1 and collagen type XXII alpha 1. PRGF-induced expression of the above factors was influenced by blockade of the epidermal growth factor receptor (EGFR), a receptor playing a crucial role in wound healing. A differential induction of the investigated factors was also detected in skin explants exposed to PRGF and in experimentally generated in vivo wounds treated with Vivostat PRF®. Together, our study indicates that the induction of ECM-related factors may contribute to the beneficial wound-healing effects of PRGF-based formulations.

The Antimicrobial and Immunomodulatory Function of RNase 7 in Skin.

The human ribonuclease RNase 7 has been originally isolated from human skin and is a member of the human RNase A superfamily. RNase 7 is constantly released by keratinocytes and accumulates on the skin surface. The expression of RNase 7 in keratinocytes can be induced by diverse stimuli such as cytokines, growth factors, and microbial factors. RNase 7 exhibits a potent broad spectrum of antimicrobial activity against various microorganisms and contributes to control bacterial growth on the skin surface. The ribonuclease and antimicrobial activity of RNase 7 can be blocked by the endogenous ribonuclease inhibitor. There is also increasing evidence that RNase 7 exerts immunomodulatory activities and may participate in antiviral defense. In this review, we discuss how these characteristics of RNase 7 contribute to innate cutaneous defense and highlight its role in skin infection and inflammation. We also speculate how a potential dysregulation of RNase 7 promotes inflammatory skin diseases and if RNase 7 may have therapeutic potential.

Staphylococcus epidermidis-induced Interleukin-1 Beta and Human Beta-defensin-2 Expression in Human Keratinocytes is Regulated by the Host Molecule A20 (TNFAIP3).

Staphylococcus epidermidis is an abundant skin commensal capable of activating cutaneous defense responses, such as induction of cytokines and antimicrobial peptides. To permanently colonize human skin and prevent inflammation S. epidermidis needs to control the induction of host defense mediators. We report here that S. epidermidis induces expression of the host regulator protein A20 in human keratinocytes, thereby controlling expression and release of interleukin-1 beta. siRNA-mediated knockdown of A20 expression strongly enhanced the induction of interleukin-1 beta gene expression and protein release in keratinocytes stimulated with S. epidermidis. Furthermore, siRNA-mediated knockdown of A20 resulted in enhanced gene expression and secretion of the antimicrobial peptide human beta-defensin-2 in keratinocytes facing S. epidermidis. Mechanistically, A20 negatively controlled S. epidermidis-induced activation of the transcription factor NF-kappaB. Together, these data indicate that S. epidermidis exploits A20 to attenuate cutaneous defense responses, which may help S. epidermidis to persist on human skin.