SKIN MICROBIOME 15


response. Moreover, this commensal- induced TLR signaling may be necessary for cell survival and repair during infection. Lipoteichoic acid (LTA) produced by S. epidermidis can inhibit skin inflammation through TLR2 and TLR3-mediated crosstalk mechanism.6 Interestingly, it has been reported that


rosacea affects patients at a certain age. It is also possible that age-specific modulations in TLR expression might play an essential role in the course of this disease. Besides the possible suppression of


inflammatory responses via TLR-TLR cross talk, TLR2 activation signaling enhances tight junction barrier in epidermal keratinocytes that prevents the passage of small molecules through the space between cells.7-10


Furthermore, the


dynamically controlled tight junctions (TJ), essential anchoring systems between differentiating keratinocytes in the stratum corneum in skin, are fundamental in maintaining skin homeostasis and preventing further invasion of pathogens. TJ is essential for anchoring systems


between the differentiating keratinocytes in the stratum granulosum and is crucial for skin barrier function regulating water flux, solutes and pathogens.


Epidermal response Keratinocytes continuously sample the microbiota colonizing the skin surface through pattern recognition receptors (PPRs) such as TLRs. These receptors recognize pathogen-associated molecular patterns (PAMPs). The activation of keratinocytes PPRs by PAMPs immediately initiates the innate immune response, resulting in the secretion of antimicrobial peptides (AMPs), cytokines and chemokines. Beyond affecting an adaptive immune response, AMPs also directly kill bacteria, fungi and enveloped viruses. Therefore, there is a constant interplay among keratinocytes, immune cells and microorganisms that is modulated by AMPs, cytokines, chemokines and microbial peptides The activation of TLRs induces a signaling


cascade which results in the release of AMPs by the keratinocytes. The latter have a direct action to regulate bacterial growth but also an indirect action by activating the immune system. It has been shown that the commensal


bacterium S.epidermidismodulates TLR3- dependent inflammation by initiating a TLR2- mediated cross-talk mechanism to suppress inflammation. This bacterium also induces keratinocytes to express endogenous AMPs through TLR2-dependant mechanism. In order to evaluate this effect, levels of


Elafin mRNAs, RNase7, Psoriasin, Calprotectin and Lysozyme, were measured in vitro.


October 2020 4 n Control n Kalmethic®


+216% **


3


+161% **


+111% *


2


+78% *


+66% ***


+55% *


0.04%


1


0 Elafin RNASE7 Figure 4: Evaluation of Kalmethic® Psoriasin S100A8 S100A9 Calprotectine


40-year-old normal Adult Human Primary Epidermal Keratinocyte (nHEK) (control in light grey profile) and treated with Kalmethic®


effect on antimicrobial peptides mRNA levels in keratinocytes. 0.04% (gold profile), were maintained in an incubator equilibrated with


5±1.5% CO2 at 37±2°C in a specific medium «Dermal Cell Basal Medium» supplemented with Keratinocyte Growth kit components. Target mRNA were purified and then quantified using RT-qPCR. mRNA levels are expressed as induced arbitrary unit compared to control. The SEM bars are represented in black lines on each histogram. The statistical analysis was performed using T-test (* p- value<0.05; ** p-value<0.01; *** p-value<0.001 vs control, n=3).


a 1.6


+43% *


b 1.5 n Control n Kalmethic® 0.04% Lysozyme


1.2 1.2 0.9 0.8 0.6 0.4 0.3


+33% *


+30% *


0 miR-210 Figure 5: Effect of Kalmethic®


0 HIF1AN RHOB on (a) miR-210 and (b) its target genes level in keratinocytes. Expression


ratios were calculated against untreated keratinocytes (NT). qPCR experiments were performed in triplicate for each biological sample. Gene expression was normalized using RNU1A1 house-keeping gene. Each experiment was performed in biological duplicate. The third sample correspond to RT-PCR from the pool of total RNA of the two biological samples. The SEM bars are represented in black lines on each histogram. The statistical analysis was performed using T-test (* p-value<0.05 vs control, n=3).


Compared to control, Elafin, RNASE7,


Psoriasin, Calprotectine and Lysozyme expression were significantly upregulated in nHEK (78%, 66%, 161%, 216%, 111% and 55% respectively)(Fig 4). These results highlighted the epidermal response induced by Kalmethic®


and its ability to boost keratinocytes antimicrobial


peptides production. As result, it reinforces natural skin own defenses. Elderly individuals have an increased


susceptibility to infections, partly because of a suppressed antimicrobial peptide landscape, which suggests a dampened ability of aging skin to control growth of specific bacteria as corynebacterial.11


A link PERSONAL CARE NORTH AMERICA


miR-210 expression level


Antimicrobials peptides expression level


Expression level of miR-210 targets


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