Elasticity
Firmness
Cellular adhesion
plant-
EGF
The importance of growth factors in the skin
2
Growth factors are naturally occurring proteins that stimulate cellular growth, proliferation and differentiation.
They play an important role in maintaining a healthy skin structure, mediating communication between
epidermal and dermal cells by binding to specific receptors on the surface of the target cell.
Well known to play an important role in reversing the effects of skin aging, topical application of human like
growth factors has been shown to act locally stimulating cellular renewal, reducing wrinkles and boosting
collagen synthesis.
EGF
Epidermal Growth Factor is a 7 kDa protein that stimulates cell growth, proliferation, and differentiation by
binding with high affinity to its receptor on the surface of the target cells. This binding initiates the MAPK/ERK
(protein-tyrosine kinase) pathway and ends with nuclear DNA expressing proteins.
EGF is naturally found in our skin but its presence diminishes notably with age (over 50% after the age of 30)
and UV light exposure and the renewal of epidermal cells noticeably slows down. The deficiency of EGF results
in skin aging, in formation of wrinkles, freckles and age marks.
EGF works very efficiently even at very low concentrations. Topically applied, and has been found to accelerate
the rate of skin renewal and re-epithelialization, to stimulate collagen production and to help preventing photodamage and fine lines and wrinkles. EGF shows a good complementary effect with humectants such as
hyaluronic acid.
plant-EGF
plant-EGF (Nicotiana benthamiana Hexapeptide-40 sh-Oligopeptide-1) is synthetic bioengineered highly purified
Epidermal Growth Factor produced in plants.
It is a biotech ingredient for the prevention of skin aging. Its ability to induce collagen and fibronectin
biosynthesis helps improving the appearance of wrinkles and the elasticity of the skin.
In vitro efficacy tests
All assays were third party performed
Wound Healing Assay
Human keratinocytes (HaCaT cells) serum
starved were seeded in 24-well plates and
grown up to 90% confluence. Then 1 mm
scratch was made with a pipet tip and pictures
were taken immediately. Cells were allowed to
close the damage when kept at 37°C.
Migration of HaCaT cells in a dose-dependent
manner highlights the regenerative properties of
plant-EGF, based on its promoting effect on skin
cell migration, proliferation and survival. Wound
healing assay demonstrated the potential to
significantly stimulate and promote skin cell
regeneration by 100% in 78 hours under proaging conditions. plant-EGF high purity
guarantees activity at very low concentrations.
CN: Untreated cells; (1): 1 ng/ml plant-EGF; (5) 5 ng/ml plantEGF; (10) 10 ng/ml plant-EGF.
1 mm wounds were closed at 78 hours
after
treatment
under
pro-aging
conditions
Human keratinocytes cell proliferation
The aim of this assay is to assess quantitatively
the effects of plant-EGF on the stimulation of the
cell cycle and viability of keratinocytes, the
predominant cell type in the epidermis.
300%
3
% cell viability
250%
Cell viability of HaCaT cell extracts from the wound
healing assay was assessed by MTT. Untreated
cells were used as negative control. plant-EGF
promotes proliferation of HaCaT cells in a dose
dependent manner, supporting the regenerating
and re-epithelializing performance of the wound
healing assay.
200%
150%
100%
50%
0%
0
1
plant-EGF (ng/ml)
5
Proliferation increased by 152% at 78
hours after treatment under pro-aging
conditions
Human fibroblast cell proliferation
Dermal fibroblasts are the cells that synthesize
collagen and the extracellular matrix to maintain
the skin’s structure. As a result of the process of
aging turnover of new fibroblast decreases,
collagen production slows and the skin becomes
thinner and wrinkled.
250%
% cell viability
200%
150%
Primary dermal fibroblasts were seeded in 24-well
plates, grown up to 40% confluence and treated
with different concentrations of plant-EGF for 24
hours. Cell viability was assessed by MTT assay
and showed significant stimulation of fibroblasts
proliferation at 1 ng/ml versus the untreated
control.
100%
50%
0%
0
1
plant-EGF (ng/ml)
5
Fibroblast proliferation boosts the skin repairing
process stimulating collagen synthesis and the
formation of connective tissue.
Proliferation increased by 95% at 24 hours
after treatment
Type I Collagen synthesis
Collagen is one of the main structural proteins of human skin, over 90% of the skin's total proteins, and provides
structural support to cells and tissues.
Collagen I expression profile was performed by indirect immunofluorescence on primary dermal fibroblasts
grown up to 60-70% confluence, showing a remarkable intracellular increment of collagen versus
the untreated fibroblasts 24 hours after treatment with 5 ng/ml of plant-EGF.
Type I Collagen expression profile after treatment with plant-EGF at 5ng/ml. Scale bar 20 µm.
4
Fibronectin synthesis
Elastin synthesis
Fibronectin together with collagen and proteoglycans are
integral components of the ECM, and plays an important
role in cell adherence.
Elastin gives the skin with the ability to stretch and
recoil and plays a critical role in supporting and
maintaining strong skin cells. Even being the longest
lasting protein in the body, elastin damage caused by
sun overexposure and aging may be sufficiently
repaired and the skin gradually loses its elasticity.
Fibronectin expression profile was performed by indirect
immunofluorescence on primary dermal fibroblasts grown
up to 60-70% confluence, showing an extracellular
increment of the protein 24 hours after treatment, and a
better organization versus the untreated fibroblast after
treatment with different concentrations of plant-EGF.
Fibronectin expression profile (red fluorescence) after treatment
with plant-EGF at 1ng/ml and 5 ng/ml. Scale bar 50 µm
Elastin expression profile was performed by indirect
immunofluorescence on primary dermal fibroblasts
grown up to 60-70% confluence, showing an
intracellular increment of the protein 24 hours after
treatment. An intense Elastin fluorescence can be
noticed in the plant-EGF treated cells.
Elastin expression profile (green fluorescence) after treatment
with plant-EGF at 1ng/ml and 5 ng/ml. Scale bar 50 µm
Uses
Improving the skin appearance particularly after age 30: anti-aging, anti-wrinkle, anti-sagging, improving aspect
of scars and marks, minimizing pores.
Bibliography
-Heck D.E., Laskin D.L., Gardner C.R., Laskin J.D. 1992. Epidermal growth factor suppresses nitric oxide and hydrogen peroxide production by keratinocytes. Potential role
for nitric oxide in the regulation of wound healing. J Biol Chem 267:21277-80.
-Tsang M.W., Wong W.K.R., Hung C.S., Lai K., Tang W., Cheung E.Y.N., Kam G., Leung L., Chan C.W., Chu C.M., Lam E.K.H. 2003. Human epidermal growth factor
enhances healing of diabetic foot ulcers. Diabetes Care 26:1856-1861.
-Schouest J.M., Luu T.K., Moy R.L. 2012. Improved texture and appearance of human facial skin after daily topical application of barley produced, synthetic, human-like
epidermal growth factor (EGF) serum. Source Moy-Fincher-Chipps Facial Plastics and Dermatology, Beverly Hills, CA, USA. J Drugs Dermatol. 11(5):613-20.
-Atkin D.H., Trookman N.S., Rizer R.L., Schreck L.E., Ho E.T., Gotz V., Ford R.O., Mehta R.C. 2010. Combination of physiologically balanced growth factors with
antioxidants for reversal of facial photodamage. J Cosmet Laser Ther. 12(1):14-20. doi: 10.3109/14764170903449786.
-Kong M. and Hong S.E. 2013. Topical Use of Recombinant Human Epidermal Growth Factor (EGF)-Based Cream to Prevent Radiation Dermatitis in Breast Cancer
Patients: a Single-Blind Randomized Preliminary Study. Asian Pac J Cancer Prev. 14(8):4859-4864.
-Carpenter G.J. 1985. Epidermal growth factor: biology and receptor metabolism. Cell Sci Suppl. 3:1-9.
-Greenhalgh D.G. 1996. The role of growth factors in wound healing. J Trauma 41(1):159-67.
-Alemdaroglu C., Degim Z., Celebi N., Sengezer M., Alomeroglu M., Nacar A. 2008. Investigation of epidermal growth factor containing liposome formulation effects on burn
wound healing. J Biomed Mater Res 85(1):271-83.
-Alemdaroglu C., Degim Z., Celebi N., Zor F., Ozturk S., Erdogan D. 2006. An investigation on burn wound healing in rats with chitosan gel formulation containing epidermal
growth factor. Burns 32(3):319-27.
-Brown G.L., Nanney L.B., Griffen J., Cramer A.B., Yancey J.M., Curtsinger L.J. 1989. Enhancement of wound healing by topical treatment with epidermal growth factor. N
Engl J Med. 321(2):76-9.
-Couch J.H. 1987. Mitotic activity of corneal endothelial cells in organ culture with recombinant human EGF. Ophthalmology 94(1):1-6.
-Kitazawa T. 1990. The mechanism of accelerated corneal ephithelial healing by human EGF. Invest Ophthalmol Vis Sci 31(9):1773-7.
-Ryu S.H., Kim Y.H., Lee S.W., Hong J.P. 2010. The preventive effect of recombinant human growth factor (rhEGF) on the recurrence of radiodermatitis. -J Radiat Res.
51(5):511-7.
-Ryu S.H., Moon S.Y., Yang Y.J., Moon S.R., Hong J.P., Choi J., Lee S.W. 2009. Recombinant human epidermal growth factor accelerates the proliferation of irradiated
human fibroblasts and keratinocytes in vitro and in vivo. J Radiat Res. 50(6):545-52.
-Kim Y.S., Lew D.H., Tark K.C., Rah D.K., Hong J.P. 2010. Effect of recombinant human epidermal growth factor against cutaneous scar formation in murine full-thickness
wound healing. J Korean Med Sci 25(4):589-96. doi: 10.3346/jkms.2010.25.4.589.
-Levi-Montalcini R. and Calissano P. 1979. The Nerve-Growth Factor. Scientific American 240: 44-53
5
DESCRIPTION: plant-EGF is a single chain
recombinant human peptide produced by
transient expression in Nicotiana benthamiana
plants. The starting gene is a synthesized copy of
the human gene which codes for Epidermal
Growth Factor, used as such or adapted to the
production host. It contains a maximum of 54
amino acids which may contain disulfide bonds
and/or glycosylation. The protein consists of the
proper sequence of the 20 standard amino acids.
SUGGESTED INCI: Nicotiana benthamiana
Hexapeptide-40 sh-Oligopeptide-1
IECIC 2014: OLIGOPEPTIDE-1
CAS NUMBER: 62229-50-9
ORIGIN: Biotechnology.
CONCENTRATION: ≤5%
PURITY: 97%, purified by sequential
chromatography (Affinity and Anionic exchangeFPLC).
PROTEIN
APPEARANCE: Lyophilized white fine powder.
RECOMMENDED DOSE: 0.01 ppm - 0.02ppm.
SOLUBILITY: Soluble in water.
RECONSTITUTION: Reconstitute with deionized
water and prevent from microbial contamination.
Trace dilution method is suggested to dissolve
the highly pure, frozen dried product. Add to the
cosmetic formula during the final phase,
preferably below 40°C.
•
100 µg vials: add 5 ml of water and use it in
5 - 10 liters/kg of final formula.
•
250 µg vials: add 5 ml of water and use it in
12 - 25 liters/kg of final formula.
•
500 ug vials: add 5 ml of water and use it in
25 - 50 liters/kg of final formula.
STERILE FILTRATION: 0.22 µm
ENDOTOXIN LEVEL: Less than 0.04EU/µg as
measured by LAL method.
RECOMMENDED PH: 5-8.
PROTEASE ACTIVITY: Very low to zero protease
activity.
STORAGE: This lyophilized powder is stable at
room temperature. We recommend being
stored at 2-8°C. Use immediately after
reconstitution, or keep aliquots at -20°C. Keep
away from direct sunlight.
TOXICOLOGY: The toxicological profile of the
product for cosmetic purposes was assessed in
vitro.
Cytotoxicity on human dermal fibroblasts:
No cytotoxic.
Cytotoxicity
on
human
keratinocytes: No cytotoxic.
epidermal
Genotoxicity: No genotoxic. Ames test (OECD
471).
Ocular Irritation: Not irritating for the eyes
(NRU - Neutral Red Uptake test).
Skin irritation:
(OECD 439).
Not irritating for the skin
Pro-sensitizing potential: Potentially not
sensitizing for the skin. (Monocytes cell line
THP-1).
.
The information in this publication is given in good faith based on our present knowledge and experience. The only guaranteed analytical specifications are
those appearing in the certificate of analysis sent with each delivery. It is good practice to determinate suitability for a particular purpose prior to use and to
conduct safety tests on all final formulations prior to marketing. The recipient is solely responsible for ensuring that products marketed to consumers comply with
all relevant laws and regulations. We will not assume any expressed or implied liability in connection with any use of this information, or other legal responsibility
on our part, including with regard to existing third party intellectual property rights. PLANTADERMA, S.L. c/ Santiago Grisolía 2, Madrid Scientific Park. D132
28760 Tres Cantos. Madrid- Spain. Tel.: +34 916 331 340 Fax: +34 916 332 197. www.plantaderma.es.
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regeneration
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biotechnology
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performance
technology
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laboratory
proteins biology
green
The development of this product had finantial support
under R+D Programs of
Plantaderma. Madrid Scientific Park
c/ Santiago Grisolía 2, D132
28760 Tres Cantos. SPAIN
T +34 916 331 340 F +34 916 333 197
[email protected]
www.plantaderma.es
V: 4/2014