Controlling the Growth of the Skin Commensal Staphylococcus epidermidis Using D-Alanine Auxotrophy

ABSTRACTUsing live microbes as therapeutic candidates is a strategy that has gained traction across multiple therapeutic areas. In the skin, commensal microorganisms play a crucial role in maintaining skin barrier function, homeostasis, and cutaneous immunity. Alterations of the homeostatic skin microbiome are associated with a number of skin diseases. Here, we present the design of an engineered commensal organism, Staphylococcus epidermidis, for use as a live biotherapeutic product (LBP) candidate for skin diseases. The development of novel bacterial strains whose growth can be controlled without the use of antibiotics, or genetic elements conferring antibiotic resistance, enables modulation of therapeutic exposure and improves safety. We therefore constructed an auxotrophic strain of S. epidermidis that requires exogenously supplied D-alanine. The S. epidermidis strain, NRRL B-4268 Δalr1Δalr2Δdat (SEΔΔΔ) contains deletions of three biosynthetic genes: two alanine racemase genes, alr1 and alr2 (SE1674 and SE1079), and the D-alanine aminotransferase gene, dat (SE1423). These three deletions restricted growth in D-alanine deficient media, pooled human blood, and skin. In the presence of D-alanine, SEΔΔΔ colonized and increased expression of human β-defensin 2 in cultured human skin models in vitro. SEΔΔΔ, showed a low propensity to revert to D-alanine prototrophy, and did not form biofilms on plastic in vitro. These studies support the potential safety and utility of SEΔΔΔ as a live biotherapeutic strain whose growth can be controlled by D-alanine.

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Controlling the Growth of the Skin Commensal Staphylococcus epidermidis Using D-Alanine Auxotrophy

mSphere ◽

10.1128/msphere.00360-20 ◽

2020 ◽

Vol 5 (3) ◽

Author(s):

David Dodds ◽

Jeffrey L. Bose ◽

Ming-De Deng ◽

Gilles R. Dubé ◽

Trudy H. Grossman ◽

...

Keyword(s):

Antibiotic Resistance ◽

Staphylococcus Epidermidis ◽

Skin Diseases ◽

Skin Barrier ◽

Skin Barrier Function ◽

Skin Microbiome ◽

Bacterial Strains ◽

Content Type ◽

Genetic Elements

ABSTRACT Using live microbes as therapeutic candidates is a strategy that has gained traction across multiple therapeutic areas. In the skin, commensal microorganisms play a crucial role in maintaining skin barrier function, homeostasis, and cutaneous immunity. Alterations of the homeostatic skin microbiome are associated with a number of skin diseases. Here, we present the design of an engineered commensal organism, Staphylococcus epidermidis, for use as a live biotherapeutic product (LBP) candidate for skin diseases. The development of novel bacterial strains whose growth can be controlled without the use of antibiotics or genetic elements conferring antibiotic resistance enables modulation of therapeutic exposure and improves safety. We therefore constructed an auxotrophic strain of S. epidermidis that requires exogenously supplied d-alanine. The S. epidermidis NRRL B-4268 Δalr1 Δalr2 Δdat strain (SEΔΔΔ) contains deletions of three biosynthetic genes: two alanine racemase genes, alr1 and alr2 (SE1674 and SE1079), and the d-alanine aminotransferase gene, dat (SE1423). These three deletions restricted growth in d-alanine-deficient medium, pooled human blood, and skin. In the presence of d-alanine, SEΔΔΔ colonized and increased expression of human β-defensin 2 in cultured human skin models in vitro. SEΔΔΔ showed a low propensity to revert to d-alanine prototrophy and did not form biofilms on plastic in vitro. These studies support the potential safety and utility of SEΔΔΔ as a live biotherapeutic strain whose growth can be controlled by d-alanine. IMPORTANCE The skin microbiome is rich in opportunities for novel therapeutics for skin diseases, and synthetic biology offers the advantage of providing novel functionality or therapeutic benefit to live biotherapeutic products. The development of novel bacterial strains whose growth can be controlled without the use of antibiotics or genetic elements conferring antibiotic resistance enables modulation of therapeutic exposure and improves safety. This study presents the design and in vitro evidence of a skin commensal whose growth can be controlled through d-alanine. The basis of this strain will support future clinical studies of this strain in humans.

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Spermidine-induced recovery of human dermal structure and barrier function by skin microbiome

Communications Biology ◽

10.1038/s42003-020-01619-4 ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Gihyeon Kim ◽

Misun Kim ◽

Minji Kim ◽

Changho Park ◽

Youngmin Yoon ◽

...

Keyword(s):

Human Skin ◽

Barrier Function ◽

Skin Diseases ◽

Lipid Synthesis ◽

Skin Barrier ◽

Streptococcus Thermophilus ◽

Skin Barrier Function ◽

Skin Microbiome ◽

Skin Structure

AbstractAn unbalanced microbial ecosystem on the human skin is closely related to skin diseases and has been associated with inflammation and immune responses. However, little is known about the role of the skin microbiome on skin aging. Here, we report that the Streptococcus species improved the skin structure and barrier function, thereby contributing to anti-aging. Metagenomic analyses showed the abundance of Streptococcus in younger individuals or those having more elastic skin. Particularly, we isolated Streptococcus pneumoniae, Streptococcus infantis, and Streptococcus thermophilus from face of young individuals. Treatment with secretions of S. pneumoniae and S. infantis induced the expression of genes associated with the formation of skin structure and the skin barrier function in human skin cells. The application of culture supernatant including Streptococcal secretions on human skin showed marked improvements on skin phenotypes such as elasticity, hydration, and desquamation. Gene Ontology analysis revealed overlaps in spermidine biosynthetic and glycogen biosynthetic processes. Streptococcus-secreted spermidine contributed to the recovery of skin structure and barrier function through the upregulation of collagen and lipid synthesis in aged cells. Overall, our data suggest the role of skin microbiome into anti-aging and clinical applications.

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Potential of Tamanu (Calophyllum inophyllum) Oil for Atopic Dermatitis Treatment

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2021/6332867 ◽

2021 ◽

Vol 2021 ◽

pp. 1-9

Author(s):

Amadeus Pribowo ◽

Jyothsna Girish ◽

Marsia Gustiananda ◽

Rakrya Galih Nandhira ◽

Pietradewi Hartrianti

Keyword(s):

Atopic Dermatitis ◽

Treatment Option ◽

Skin Diseases ◽

Treatment Options ◽

In Vivo Studies ◽

Skin Barrier Function ◽

Skin Microbiome ◽

Calophyllum Inophyllum

Tamanu oil, derived from the nut of Calophyllum inophyllum L., has been traditionally used to treat various skin-related ailments. In recent years, this oil is increasingly gaining popularity as researchers continue to search for novel natural alternative therapies for various skin diseases. There have been a number of in vitro and in vivo studies investigating various skin-active properties of tamanu oil, and it has been proven to have potent anti-inflammatory, antioxidant, antimicrobial, analgesic, and even wound-healing abilities. These properties make tamanu oil an especially interesting candidate for the treatment of atopic dermatitis (AD). This multifaceted disease is marked by the disruption of the skin barrier function, chronic inflammation, and skin microbiome dysbiosis with limited treatment options, which is free from adverse events and inexpensive, making it desperate for a new treatment option. In this review, we examine previous in vitro and in vivo studies on AD-relevant pharmacological properties of tamanu oil in order to evaluate the potential of tamanu oil as a novel treatment option for AD.

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Skin care as a way to recover microbiome in patients with atopic dermatitis

Rossiiskii Allergologicheskii ZHurnal ◽

10.36691/rja526 ◽

2014 ◽

Vol 11 (1) ◽

pp. 17-22

Author(s):

K S Pavlova

Keyword(s):

Staphylococcus Aureus ◽

Atopic Dermatitis ◽

Staphylococcus Epidermidis ◽

Barrier Function ◽

Skin Barrier ◽

Skin Barrier Function ◽

Skin Microbiome ◽

Important Goal ◽

Staphylococcus Haemolyticus ◽

Genomic Technologies

Recently microbiome of the skin was characterized using genomic technologies in norm and in pathology. Microbiome of the affected skin in atopic dermatitis is characterized by a lack of the variety of bacteria, decrease of the Actinomycetes and Proteobacteries species and increase of Staphylococci colonization (Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus and others). Restoration of the skin barrier function is the most important goal in the overall concept of the atopic dermatitis treatment. Recent studies demonstrated the possibility of reductionof inflammation, xerosis, itching and restoration of skin microbiome of the affected areas by emollients use (Lipikar Baume AP, La Roche Posay), as a result of the skin barrier function improvement.

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Impact of pH on growth of Staphylococcus epidermidis and Staphylococcus aureus in vitro

Journal of Medical Microbiology ◽

10.1099/jmm.0.001421 ◽

2021 ◽

Vol 70 (9) ◽

Author(s):

Vidula Iyer ◽

Janhavi Raut ◽

Anindya Dasgupta

Keyword(s):

Staphylococcus Aureus ◽

Growth Kinetics ◽

Staphylococcus Epidermidis ◽

Type Species ◽

Ph Dependence ◽

Skin Microbiome ◽

Content Type ◽

Link Type ◽

Kinetics Of

The pH of skin is critical for skin health and resilience and plays a key role in controlling the skin microbiome. It has been well reported that under dysbiotic conditions such as atopic dermatitis (AD), eczema, etc. there are significant aberrations of skin pH, along with a higher level of Staphylococcus aureus compared to the commensal Staphylococcus epidermidis on skin. To understand the effect of pH on the relative growth of S. epidermidis and S. aureus , we carried out simple in vitro growth kinetic studies of the individual microbes under varying pH conditions. We demonstrated that the growth kinetics of S. epidermidis is relatively insensitive to pH within the range of 5–7, while S. aureus shows a stronger pH dependence in that range. Gompertz’s model was used to fit the pH dependence of the growth kinetics of the two bacteria and showed that the equilibrium bacterial count of S. aureus was the more sensitive parameter. The switch in growth rate happens at a pH of 6.5–7. Our studies are in line with the general hypothesis that keeping the skin pH within an acidic range is advantageous in terms of keeping the skin microbiome in balance and maintaining healthy skin.

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An integrated multiscale, multicellular skin model

10.1101/830711 ◽

2019 ◽

Author(s):

Ryan Tasseff ◽

Boris Aguilar ◽

Simon Kahan ◽

Seunghwa Kang ◽

Charles C. Bascom ◽

...

Keyword(s):

Differential Equations ◽

Cell Biology ◽

High Performance ◽

Skin Barrier ◽

Skin Barrier Function ◽

Emergent Properties ◽

Skin Model ◽

Environmental Chemical ◽

Software Models

ABSTRACTSkin is our primary barrier to the outside world, protecting us from physical, biological and chemical threats. Developing innovative products that preserve and improve skin barrier function requires a thorough understanding of the mechanisms underlying barrier response to topical applications. In many fields, computer simulations already facilitate understanding, thus accelerating innovation. Simulations of software models allow scientists to test hypothesized mechanisms by comparing predicted results to physical observations. They also enable virtual product optimization, without physical experiments, once mechanisms have been validated. The physical accessibility and abundant knowledge of skin structure makes it a prime candidate for computational modeling. In this article, we describe a computational multiscale multicellular skin model used to simulate growth and response of the epidermal barrier. The model integrates several modeling styles and mathematical frameworks including ordinary differential equations, partial differential equations, discrete agent-based modeling and discrete element methods. Specifically, to capture cell biology and physical transport, we combined four distinct sub-models from existing literature. We also implemented methods for elastic biomechanics. Our software implementation of the model is compatible with the high-performance computing simulation platform Biocellion. The integrated model recapitulates barrier formation, homeostasis and response to environmental, chemical and mechanical perturbation. This work exemplifies methodology for integrating models of vastly different styles. The methodology enables us to effectively build on existing knowledge and produce “whole-system” tissue models capable of displaying emergent properties. It also illustrates the inherent technical difficulties associated with the mounting complexity of describing biological systems at high fidelity. Among the challenges are validation of the science, the mathematical representations approximating the science and the software implementing these representations. Responsibility for a discrepancy observed between in silico and in vitro results may as easily lie at one of these three levels as at another, demanding that any sustainable modeling endeavor engage expertise from biology, mathematics and computing.

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A Trifecta of New Insights into Ovine Footrot for Infection Drivers, Immune Response and Host Pathogen Interactions.

Infection and Immunity ◽

10.1128/iai.00270-21 ◽

2021 ◽

Author(s):

Adam M. Blanchard ◽

Ceri E. Staley ◽

Laurence Shaw ◽

Sean R Wattegedera ◽

Christina-Marie Baumbach ◽

...

Keyword(s):

Immune Response ◽

Type I Collagen ◽

Global Analysis ◽

Bacterial Species ◽

Skin Barrier ◽

Control Measures ◽

Skin Barrier Function ◽

Type I ◽

Bacterial Populations

Footrot is a polymicrobial infectious disease in sheep causing severe lameness, leading to one of the industry’s biggest welfare problems. The complex aetiology of footrot makes in-situ or in-vitro investigations difficult. Computational methods offer a solution to understanding the bacteria involved, how they may interact with the host and ultimately providing a way to identify targets for future hypotheses driven investigative work. Here we present the first combined global analysis of the bacterial community transcripts together with the host immune response in healthy and diseased ovine feet during a natural polymicrobial infection state using metatranscriptomics. The intra tissue and surface bacterial populations and the most abundant bacterial transcriptome were analysed, demonstrating footrot affected skin has a reduced diversity and increased abundances of, not only the causative bacteria Dichelobacter nodosus , but other species such as Mycoplasma fermentans and Porphyromonas asaccharolytica . Host transcriptomics reveals a suppression of biological processes relating to skin barrier function, vascular functions, and immunosurveillance in unhealthy interdigital skin, supported by histological findings that type I collagen (associated with scar tissue formation) is significantly increased in footrot affected interdigital skin comparted to outwardly healthy skin. Finally, we provide some interesting indications of host and pathogen interactions associated with virulence genes and the host spliceosome which could lead to the identification of future therapeutic targets. Impact Statement Lameness in sheep is a global welfare and economic concern and footrot is the leading cause of lameness, affecting up to 70% of flocks in the U.K. Current methods for control of this disease are labour intensive and account for approximately 65% of antibiotic use in sheep farming, whilst preventative vaccines suffer from poor efficacy due to antigen competition. Our limited understanding of cofounders, such as strain variation and polymicrobial nature of infection mean new efficacious, affordable and scalable control measures are not receiving much attention. Here we examine the surface and intracellular bacterial populations and propose potential interactions with the host. Identification of these key bacterial species involved in the initiation and progression of disease and the host immune mechanisms could help form the basis of new therapies.

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Efficacy of Phototherapy With 308-nm Excimer Light for Skin Microbiome Dysbiosis and Skin Barrier Dysfunction in Canine Atopic Dermatitis

Frontiers in Veterinary Science ◽

10.3389/fvets.2021.762961 ◽

2021 ◽

Vol 8 ◽

Author(s):

Ju-Yong Park ◽

Seon-Myeong Kim ◽

Jung-Hyun Kim

Keyword(s):

Atopic Dermatitis ◽

Relative Abundance ◽

Barrier Function ◽

Therapeutic Option ◽

Clinical Signs ◽

Skin Barrier ◽

Skin Barrier Function ◽

Skin Microbiome ◽

Canine Atopic Dermatitis ◽

Allergic Skin

The management of canine atopic dermatitis, an allergic skin disorder, is challenging. To investigate the effect of phototherapy using a 308-nm excimer light as a topical treatment for canine atopic dermatitis, 10 dogs with canine atopic dermatitis and 10 with non-allergic skin were enrolled in this study. Phototherapy was applied every 7 days for a total of 2 months. The skin microbiome, skin barrier function, and clinical outcomes were evaluated after phototherapy. Phototherapy significantly changed the composition of the skin microbiome of dogs with atopic dermatitis and significantly increased the relative abundance of the phyla Actinobacteria and Cyanobacteria. It significantly alleviated the clinical signs of canine atopic dermatitis without serious adverse effects. Transepidermal water loss, as a measure of skin barrier function, significantly decreased after phototherapy. In addition, phototherapy increased microbial diversity and decreased the relative abundance of Staphylococcus pseudintermedius associated with the severity of canine atopic dermatitis. These results suggest that the excimer light therapy is a suitable and safe therapeutic option for canine atopic dermatitis, which is also a spontaneous animal model of atopic dermatitis.

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Topical Pioglitazone Nanoformulation for the Treatment of Atopic Dermatitis: Design, Characterization and Efficacy in Hairless Mouse Model

Pharmaceutics ◽

10.3390/pharmaceutics12030255 ◽

2020 ◽

Vol 12 (3) ◽

pp. 255 ◽

Cited By ~ 3

Author(s):

Lupe Carolina Espinoza ◽

Rodrigo Vera-García ◽

Marcelle Silva-Abreu ◽

Òscar Domènech ◽

Josefa Badia ◽

...

Keyword(s):

Atopic Dermatitis ◽

Ex Vivo ◽

In Vitro Release ◽

Skin Barrier ◽

Inflammatory Cells ◽

Hairless Mouse ◽

Skin Barrier Function ◽

Inflammatory Parameters ◽

Therapeutic Benefits

Pioglitazone (PGZ) is a drug used to treat type 2 diabetes mellitus that has been reported to show additional therapeutic activities on diverse inflammatory parameters. The aim of this study was to optimize a topical PGZ-loaded nanoemulsion (PGZ-NE) in order to evaluate its effectiveness for treating atopic dermatitis (AD). The composition of the nanoformulation was established by pseudo-ternary diagram. Parameters such as physical properties, stability, in vitro release profile, and ex vivo permeation were determined. The efficacy study was carried out using oxazolone-induced AD model in hairless mice. PGZ-NE released the drug following a hyperbolic kinetic. Additionally, its properties provided high retention potential of drug inside the skin. Therapeutic benefits of PGZ-NE were confirmed on diverse events of the inflammatory process, such as reduction of lesions, enhancement of skin barrier function, diminished infiltration of inflammatory cells, and expression of pro-inflammatory cytokines. These results were reinforced by atomic force microscope (AFM), which demonstrated the ability of the formulation to revert the rigidification caused by oxazolone and consequently improve the elasticity of the skin. These results suggest that PGZ-NE may be a promising treatment for inflammatory dermatological conditions such as AD.

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Skin Barriers in Dermal Drug Delivery: Which Barriers Have to Be Overcome and How Can We Measure Them?

Pharmaceutics ◽

10.3390/pharmaceutics12070684 ◽

2020 ◽

Vol 12 (7) ◽

pp. 684 ◽

Cited By ~ 3

Author(s):

Christian Gorzelanny ◽

Christian Mess ◽

Stefan W. Schneider ◽

Volker Huck ◽

Johanna M. Brandner

Keyword(s):

Drug Delivery ◽

Skin Diseases ◽

Current Knowledge ◽

Multiphoton Microscopy ◽

Skin Barrier ◽

Barrier Properties ◽

Transepidermal Water Loss ◽

Hair Follicles

Although, drugs are required in the various skin compartments such as viable epidermis, dermis, or hair follicles, to efficiently treat skin diseases, drug delivery into and across the skin is still challenging. An improved understanding of skin barrier physiology is mandatory to optimize drug penetration and permeation. The various barriers of the skin have to be known in detail, which means methods are needed to measure their functionality and outside-in or inside-out passage of molecules through the various barriers. In this review, we summarize our current knowledge about mechanical barriers, i.e., stratum corneum and tight junctions, in interfollicular epidermis, hair follicles and glands. Furthermore, we discuss the barrier properties of the basement membrane and dermal blood vessels. Barrier alterations found in skin of patients with atopic dermatitis are described. Finally, we critically compare the up-to-date applicability of several physical, biochemical and microscopic methods such as transepidermal water loss, impedance spectroscopy, Raman spectroscopy, immunohistochemical stainings, optical coherence microscopy and multiphoton microscopy to distinctly address the different barriers and to measure permeation through these barriers in vitro and in vivo.

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