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Modulation of Aquaporins to Deliver Consumer Benefits: Applications

E. A. Jewell-Motz, Ph.D., J. R. Kaczvinsky, Skin K. MCare
. Lammers, M.S., S. Xie, Ph.D., R. M. Osborne, Ph.D
P&GBeauty, Cincinnati, OhioUSA

Caffeine Stimulates AQP3

Background Protein Expression in vitro Conclusions
AQP3 levels decrease with 1.6 • AQP3 is expressed in human skin

Fold Change in AQP Protein

Aquaporins (AQP) are an important class of age 1.5
1.4 Product Formulated with • An age-associated decrease in AQP3
proteins that regulate the transport of water 1.3
Caffeine correlates with an age-associated
10000 1.2
and other small solutes across plasma r = -0.446 1.1
Delivers Skin Moisturization increase in dry skin.
p = 0.063
membranes. First discovered in the early 1
• Caffeine increases both the protein
Benefit in Human Clinical

AQP3 Signal
1990’s (1), there are now 13 distinct AQP 0.8 expression and glycerol transport
family members that have been identified in
5000 Control 100uM
Studies activity of AQP3 as determined in
a variety of mammalian tissues, many of * vitro.

In vivo Dry Skin Grade Improvements

which are relevant to the consumer products • Moisturizer formulated with caffeine
ELISA for In Vitro AQP Expression 0.4

(vs. moisturizer alone)

industry. In particular, two aquaporins 20 30 40 50 60 70 • Cultured human neonatal keratinocytes were cultured in * delivers a moisturization benefit to dry
Age (years) 0.3 skin in clinical testing.
(AQP3 and AQP9) are expressed in human 96 well plates and treated at 37°C for 48-96 hrs
• Fixed cells were incubated with anti-AQP3 antibody.
skin cells (2, 3). AQP3 has been implicated 0.2 • Modulation of aquaporins may be
• A peroxidase coupled secondary antibody was added,
as playing a key role in the transport and In vivo Expression of AQP 3 and AQP3 protein content was determined after 0.1 beneficial for a variety of consumer
• 20 Caucasian female subjects, 18-55 years
distribution of epidermal water and glycerol treatment with Horseradish Peroxidase Chromogen
products applications including, but
old TMB.
(3,4) and the regulation of keratinocyte • RNA isolated from hip biopsies & 1 leg not limited to, anti-aging, dry mouth




differentiation (5), both critical processes for biopsy relief, urinary incontinence, or relief for

Caffeine Stimulates AQP3

• AQP RNA expression quantitated via

maintaining skin hydration, barrier function diarrhea.

GeneChip™ assay Activity in vitro
and overall skin health.
Objective 130

1. Preston G. M., Carroll T.P., Guggino W. P. and
Agre P., Science 256, 385-387 (1992).

Glycerol Transport
Clinical Design

% Increase in
110 ●40 female subjects 2. Sougrat R., Morand M., Gondran C., Barre P.,
● Identify compounds that stimulate 100
●4 test sites per leg (8 total per subject), 5x4 cm2 area, Gobin R., Bonte F., Dumas M., and Verbavatz
aquaporin-3 expression and activity in skin 2 ul/cm2, total 40 ul treatment twice/day except
J., J. Invest. Derm., 118(4), 678-685 (2002).
cells. weekends- 1x per day
3. Sugiyama Y., Ota Y., Hara M., and Inoue S.,

●Assessments performed at baseline, 1, 2, and 3 weeks

then 2, 7, and 11 day regression assessments Biochim. Biophys. Acta 1522, 82-88 (2001).
● Determine whether identified materials Control 100uM
●Technical measures via expert visual grading for 4. Hara-Chikuma M. and Verkman A. S., Biol Cell,
promote a skin moisturizing benefit in humans dryness and redness, 97(7), 479-486 (2005).
Glycerol Transport Assay
Stratum basale
Stratum corneum when formulated into product. ●Panelists were asked not to consume caffeine or 5. Zheng X. and Bollinger Bollag W., J. Invest.
● MatTek’s Epiderm cultures were treated with test
herbal drinks
compound for 48 hours before adding glycerol to the Derm., 121(6), 1487-1495 (2003).
medium in the lower chamber.
Aquaporin channel AQP3 is localized to ● Glycerol levels within the upper chamber medium were
human skin determined 24 hours later (Free Glycerol Reagent, Sigma,
(Science 17 October 2003: St. Louis, MO).
Vol. 302. no. 5644, pp. 383 –
● Measurement in the glycerol assay was converted into a
percentage increase over the control (where the control is
equal to 100%). Each mean and SEM were derived from at
least three separate experiments, performed in duplicate.