FORMULATING FOR MILDNESS
7000 6000 5000 4000 3000 2000 1000 0
-1000
+852% $$$
-63% ***
-50% ***
-65% ***
Control
NiSO4 10µM
Dexa 1µM+Ni CAP
0.01% dm + Ni
0.005% dm + Ni
Figure 5: Anti-inflammatory effect of CAP extract against cytokine IL8 following Nickel sensitising stress in keratinocytes. ($$$ p<0.001 vs Untreated; ** p< 0.01/ *** p<0.001 vs NiSO4- one-factor ANOVA followed by Tukey test, GraphPad Prism 5 software).
stabilisation of the plasma membrane where it inhibits the intracellular penetration of Ca++, since this ion is essential to mast cell degranulation. Under the experimental conditions of
this study, CAP extract, tested at 0.033% and 0.1% (dm), presented a clear concentration- dependent inhibitor effect on the activation of basophils induced by fMLP (22% and 39% inhibition, respectively) (Fig 6).
Inhibition of chemoattractant factors following a Th2 type stress The gene expression of pro-inflammatory chemokines was analysed in normal human epidermal keratinocytes (NHEK) stimulated by a mixture of cytokines Th2 (IL-4 + IL-13 + IL-22 + TNF-α) reproducing a late phase (chronic inflammation) “atopic dermatitis” type phenotype as described in the introduction. Chlamydomonas Acidophila (CAP) extract 0.01% was compared to the reference JAK inhibitor I.
A
400 350 300 250 200 150 100 50 0
Control B
1600 1400 1200 1000 800 600 400 200 0
Control +245% CAP
50 45 40 35 30 25 20 15 10 5 0
Control
31
$$$
-26% *
-46% **
-22% *
-39% **
fMLP1µM
SB202190 30µM
Cromoglycate 10mM
CAP 0.033% dm
CAP 0.1% dm
Figure 6: Inhibitor effect of CAP extract on basophil activation ($$$ p<0.001 vs Non-stimulated control; * p< 0.05/ ** p<0.01 vs fMLP - Student t test)
Results The stress due to cytokines Th2 strongly induced the gene expression level of CCL5 and CCL27 in keratinocytes (Fig 7). CCL5 (C-C motif chemokine ligand 5 or RANTES) and CCL27 (C-C motif chemokine ligand 27 or CTACK) code for chemokines involved in the amplification of the cutaneous inflammatory and allergic reaction. These chemokines work specifically by inducing the recruitment of immune cells (monocytes, T memory cells, eosinophils), inducing histamine release by basophils and activating eosinophils. In keratinocytes subjected to Th2 stress,
Chlamydomonas acidophila (CAP) extract inhibited CCL5 and CCL27 gene expression and therefore protected keratinocytes from Th2 stress. Therefore, CAP extract could contribute to limiting Th2 type allergic response.
Conclusion By inhibiting on the one hand, the activation of basophils and on the other hand the gene
CCL5
expression of chemoattractant cytokines in keratinocytes subjected to a Th2 stress; Chlamydomonas acidophila (CAP) extract could contribute to the modulation of the processes involved in the initiation of the allergic reaction and therefore be of interest in atopic dermatitis.
Action on barrier function and hydration Activation of barrier and hydration markers gene expression Chlamydomonas acidophila (CAP) extract at 0.05% of dry matter (dm) was added in the culture medium of reconstituted human epidermises. After 6 hours of incubation, the expression
of the markers selected was evaluated by quantitative RT-PCR.
Results CAP extract significantly increased the level of gene expression of GBA (Glucosylceramidase, Beta-glucocerebrosidase) and RAB11A (Ras- related protein Rab-11A) markers, +91% and +114% respectively (Fig 8). The GBA/GBAP1 gene codes for the enzyme
-49% -71%
Glucosylceramidase or β-glucocerebrosidase. The β-glucocerebrosidase secreted by lamellar bodies, is responsible for the transformation of glucosylceramides from lamellar bodies into ceramides composing the lipid leaflets.9 RAB11A codes for a GTPase (Ras-related
Th2 stress
(IL4+IL13+IL22+ TNFa)
JAK Inhibitor 10µM Th2 stress
CAP 0.01% dm + Th2 stress
CCL27 +748% -41%
protein Rab-11A) involved in the biogenesis of lamellar bodies within keratinocytes. Reynier et al. recently demonstrated that silencing the RAB11A gene in a reconstituted epidermis 3D model induced a deficit in the barrier function, a reduction in the quantity of lipids in the stratum corneum as well as a reduction in the density of lamellar bodies and their secretion.10 This highlights the importance of RAB11A in the homeostasis of the epidermal barrier. CAP extract also significantly increases the
Th2 stress
(IL4+IL13+IL22+ TNFa)
JAK Inhibitor 10µM Th2 stress
CAP 0.01% dm + Th2 stress
Figure 7: Inhibition chemokines CCL5 (a) and CCL27 (b) gene expression following a Th2 type stress in keratinocytes.
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level of gene expression of HAS2 and HAS3 markers coding for hyaluronan-synthase-2 and 3, respectively, enzymes which are responsible for the synthesis of hyaluronic acid (Fig 8). In this study, the CAP extract also increased
April 2021 PERSONAL CARE
IL8 (pg/mL/mg proteins Relative quantity CCL27 Relative quantity CCL5
% activated basophilic population (CCR3+/CD63+)
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