Atopic Dermatitis (AD) is a frequent inflammatory skin disease characterized by recurrent eczema. It associates genetic/epigenetic-induced alterations of epidermal barrier and type-2 inflammation/hypersensitivity, which may be triggered by different antigens that pass through the altered skin . Some studies have reported that environmental pathogens such as house dust mites are able to induce type-2 inflammation through particular activation of innate immunity . Multiple staphylococcal strains are commonly found on the skin of AD patients. Interestingly, recent findings suggest that S. aureus may be a key factor of AD inflammation: (i) 90% of AD patients have S. aureus skin colonization on lesional skin , (ii) AD patients with S. aureus skin colonization have more increased type-2 inflammatory markers in comparison with AD patients without SA skin colonization , (iii) skin colonization by monoclonal S. aureus strains correlate with severe flares and (iv) S. aureus is detected in both epidermis and dermis during AD flares; In this study, our hypothesis is that S. aureus induces AD flares through a type 2 T cell-mediated hypersensitivity against S. aureus, involving innate and adaptive responses. Conversely, S. epidermidis, a commensal strain, has a protective effect against S. aureus dysbiosis. To this end, we will characterize, in the skin and the blood, the immune response induced by cutaneous application of : i) S. aureus isolated from patients with moderate-to-severe AD which will mimic the cutaneous dysbiosis occurring in the natural course of AD; ii) S. aureus toxins without bacteria to evaluate the skin response against those particular proteins; iii) a laboratory strain of S. epidermidis, a common well-tolerated skin commensal bacteria; iv) a mix of S. aureus and S. epidermidis to evaluate the regulatory effect of S. epidermidis on the S. aureus-induced AD inflammation. Importantly, this characterization will be led in AD patients (with alterations of skin barrier), compared to healthy volunteers (without alterations of skin barrier), as controls.
Study Type
INTERVENTIONAL
Allocation
NON_RANDOMIZED
Purpose
BASIC_SCIENCE
Masking
NONE
Enrollment
7
A 50 mL blood sample will be collected in Lithium Heparin tubes (45 mL) and dry tube (5 mL), by venipuncture, at screening and Day 3. Bacteriological samples from AD patients will be performed by swabbing the skin at screening visit (Day -42 to Day -28). Each sample will be cultured in a RPMI/human serum AB medium and methi.R (methicillin Resistant) strains will be eliminated. Thus, only S. aureus methi.S (methicillin Sensitive) will be isolated to be re-applied (via patchtest) to AD patient. A well-characterized S. epidermidis lab strain will be also applied to AD patients. Patch tests containing S. aureus, S. epidermidis or a mix S. aureus/S. epidermidis will be applied on healed or improved area as defined by a lesional score ≤ 1 or a 2-point change from the baseline lesional score. Patch tests will be applied 48h and reading of the patch tests results and biopsies will be performed 72h after patch test application.
A 50 mL blood sample will be collected in Lithium Heparin tubes (45 mL) and dry tube (5 mL), by venipuncture, at Day 0 and Day 3. Each bacteriological sample from AD patients performed by swabbing the skin at screening visit will be cultured in a RPMI/human serum AB medium and methi.R (methicillin Resistant) strains will be eliminated. Thus, only S. aureus methi.S (methicillin Sensitive) will be isolated to be applied (via patchtest) to a paired (age/sex) healthy volunteer. A well-characterized S. epidermidis lab strain will be also applied to healthy volunteers. Patch tests containing S. aureus, S. epidermidis or a mix S. aureus/S. epidermidis will be applied on healthy skin. Patch tests will be applied 48h and reading of the patch tests results and biopsies will be performed 72h after patch test application
Centre Hospitalier Lyon Sud
Pierre-Bénite, France
Occurrence of a clinical inflammatory skin lesion after application of patch test containing a solution of methi-sensible (methi-S) S.aureus, in AD patients or healthy volunteers.
A difference of the local AD severity will be searched between baseline and after S. aureus patch test application (in AD patients and healthy volunteers) according to the lesional scoring and the patch test scoring. A logistic mixed effects model will be performed to model the occurrence of clinical inflammatory skin lesion. For each sub-population (AD patients or healthy volunteers), a linear mixed effect model will be used to model the lesional scoring.
Time frame: Day 3
Molecular analysis of cytokine skin expression induced by S.aureus patch test.
Results will be expressed as fold change compared to control skin from the same individual (qPCR analysis). * Type 2 inflammation: interleukins (IL)-4, 5, 13, 31 * Type 1 inflammation: TNF, IFNγ * Type 17-22 inflammation: IL-12, IL-23, IL-17a, IL-22 * Innate inflammation: TSLP, IL-25, IL-33, IL-1b, IL-1a, IL-18 Then, a significant difference of each parameter cited above will also be searched between baseline and after S. aureus patch test application (in AD patients and healthy volunteers).
Time frame: Day 3
Type 2 specific cellular and molecular regulatory and inflammatory response in blood - OPTIONAL (only in case of additional funding/grant)
A significant difference of parameters will be searched between baseline and after S. aureus patch test application (in AD patients and healthy volunteers)
Time frame: Day 3
Presence/identification of S. aureus-specific virulence factors inducing a sensitization (immunoblotting) - OPTIONAL (only in case of additional funding/grant)
A significant difference of parameters will be searched between baseline and after S. aureus patch test application (in AD patients and healthy volunteers).
Time frame: Day 3
Differences in clinical skin response against S. epidermidis and mix S. aureus/S. epidermidis.
Results will be expressed as fold change compared to control skin from the same individual (qPCR analysis). * Type 2 inflammation: interleukins (IL)-4, 5, 13, 31) * Type 1 inflammation: TNF, IFNγ * Type 17-22 inflammation: IL-12, IL-23, IL-17a, IL-22 * Innate inflammation: TSLP, IL-25, IL-33, IL-1b, IL-1a, IL-18 Then, a significant difference of each parameter cited above will also be searched between S. epidermidis alone, mix S. epidermidis/S. aureus and S. aureus alone patch tests.
Time frame: Day 3
Molecular analysis of cytokine skin expression induced by S. epidermidis and mix S. aureus/S. epidermidis.
Results will be expressed as fold change compared to control skin from the same individual (qPCR analysis). * Type 2 inflammation: interleukins (IL)-4, 5, 13, 31) * Type 1 inflammation: TNF, IFNγ * Type 17-22 inflammation: IL-12, IL-23, IL-17a, IL-22 * Innate inflammation: TSLP, IL-25, IL-33, IL-1b, IL-1a, IL-18 Then, a significant difference of each parameter cited above will also be searched between S. epidermidis alone, mix S. epidermidis/S. aureus and S. aureus alone patch tests. A linear mixed effects model will be used to model each parameter of the molecular analysis of cytokine skin expression.
Time frame: Day 3
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