We have reported previously a new model for the induction, propagation and perpetuation of dry skin

1
P411
Stratum corneum protein and pro-inflammatory
cytokine biomarkers as endpoints for
understanding the effects of a high-efficacy body
moisturizer containing niacinamide
P J Matts PhD1, T Albader PhD1, S W Hendrix MSc2,
P Barton1, K Vanoosthuyze PhD1 and D Mulligan
PhD1, The Procter Gamble Company, Egham, UK1
and Cincinnati, USA2
RESULTS AND DISCUSSION
METHODS
INTRODUCTION
CONCLUSIONS

• A moisturizer vehicle containing high
  concentrations of niacinamide and glycerin was
  able to mitigate the micro-inflammatory status
  of dry leg skin, as evidenced by a decrease in
  IL1-RAIL-1a ratio and expression of keratins 1,
  10 and 11
• A combination of tape-stripping and a novel
  multiple-analyte method has proven a valuable new
  tool in providing rapid and accurate
  high-throughput determinations of important
  stratum corneum components.

For interpretation of the results, all analyte
levels obtained were normalised to total protein
found in each tape strip. Since the levels of the
biomarkers obtained were not normally distributed
over the data set, nonparametric testing
(Wilcoxon Signed Rank test) was conducted.
Two-tailed tests were performed with plt0.05 as
the significance level. Values below represent
mean pg analyte / µg total protein the values
obtained from the first and second tape strip (1
and 2 respectively) are shown below for
comparison
accompanying micro-inflammation and that
treatments effective in barrier improvement can
lead to an improvement in this status5,1.
Furthermore, induction of a micro-inflammatory
state has been proposed as a core propagating
factor in the recently-published Dry Skin Cycle
model of xerosis1. The observed amelioration of
this process by the niacinamide-containing
moisturizer is, therefore, of prime
importance. The decrease in expression of
keratins 1,10 and 11 observed is also
significant, as elevated levels of these proteins
are associated with hyper-keratotic,
barrier-compromised skin1. Although results
from both tape strips 1 and 2 were similar,
future studies will combine these to ensure a
representative result. Finally, it is clear
that the combination of tape-stripping and
multiple-analyte methods is able to provide rapid
and accurate high-throughput determinations of
important stratum corneum components (up to 100
from one sample).
Skin sites within the dry, outer lower legs of 24
female subjects were treated daily with a topical
moisturizer containing a high level of
niacinamide and glycerin, for 3 weeks, according
to a pre-determined randomization. A site was
left untreated as a control. Two sequential
tape-strips were acquired from each site at
baseline and 24h after the last application of
product (at 3 weeks). SC material from the strips
was solubilized and then submitted to two
separate analyses. The pro-inflammatory
cytokine, interleukin 1? (IL-1?) was assayed
using a a standard commercial ELISA methodology,
whereas its receptor antagonist (IL-1RA) was
assayed using a state-of-the-art commercial
multiple-analyte ELISA technique (involving
flow-cytometry of color-coded immuno-indexed
microspheres). The multiple-analyte technique was
also used to detect changes in specific SC
keratins. The multiple-analyte technique uses
novel 5µm polystyrene microspheres dyed
internally with a defined blends of fluorophores
to create up to 100 unique fluorescent signatures
that can be assigned to different analyte targets
(e.g., IL-1?). This signature, along with that of
reporter dyes captured during incubation of
microspheres with the sample, are detected and
quantified by a commercialized instrument
combining flow cytometry and laser excitation
(Fig 1).
We have reported previously a new model for the
induction, propagation and perpetuation of dry
skin which we have termed the Dry Skin
Cycle1,2. An underlying assumption in this model
is the pivotal role played by stratum corneum
(SC) barrier function. Disruption of this
barrier, even to a small extent, can induce a
chronic micro-inflammatory state, leading to the
over-hasty production of a variety of
sub-standard SC structural protein components.
Without appropriate intervention, xerosis ensues
with classic clinical manifestation. We have also
reported previously on the utility of niacinamide
as a unique and effective agent in augmenting SC
barrier function, principally through
up-regulation of endogenous ceramide synthesis3.
Here we report on characterizing further the
effect of topical niacinamide by assaying SC
structural protein and cytokine status, using
high-throughput novel multiple-analyte
methodology, reported by us previously4.
References 1. Stratum corneum moisturisation at
the molecular level an update in relation to the
Dry Skin Cycle, Rawlings, A.V. and Matts, P.J.,
J. Invest. Dermatol., 1241099-1102, 2005 2. The
Dry Skin Cycle Introducing a New Model for
the Induction and Propagation of Xerosis in
Normal Skin, Rawlings, A.V. and Matts, P.J.,
American Academy of Dermatology 63rd Annual
Conference, 2005 3. Topical niacinamide and
barrier enhancement, Bissett D., Cutis. 70(6
Suppl) 8-12, 2002 4. Novel microarray analysis
of skin surface biomarkers, S Hendrix, Barrier
function of mammalian skin, Gordon Conference,
2005 5. Improvement of mild inflammatory changes
of the facial skin induced by winter environment
with daily applications of a moisturizing cream.
A half-side test of biophysical skin parameters,
cytokine expression pattern and the formation of
cornified envelope, Kikuchi K., Kobayashi H.,
Hirao T., Ito A., Takahashi H. and Tagami H.,
Dermatology 207(3) 269-75, 2003
Table 1 cytokine and keratin analysis at week 3
It is clear from the results presented above that
treatment with the moisturizer containing high
levels of glycerin and niacinamide led to
significantly lower values (plt0.05) of the
IL1-RAIL-1a ratio and also significantly lower
levels (plt0.05) of keratins 1, 10 and 11. These
observations fit extremely well with in-going
hypotheses based on previous studies. It has been
reported that increased IL1-RAIL-1a ratios are
associated with winter xerosis with
OBJECTIVE
Evaluate the ability of niacinamide delivered
from a moisturizing vehicle to mitigate the
micro-inflammatory state induced during the Dry
Skin Cycle, via non-invasive tape-stripping and
subsequent multiple-analyte analysis
Microsphere with bound capture molecules
Instrument combining flow cytometry and laser
detection
Dual lasers to both identify and quantify
specific analytes
This work was supported in its entirety by
Procter Gamble.
Figure 1 schematic diagrams of the
multiple-analyte method
diagrams courtesy of the Luminex Corporation