Cosmetic compositions and method of making the same
The present invention generally relates to cosmetic compositions comprising serum derived from cultured keratinocyte and fibroblast cells. The invention also relates to methods of producing serum for use in the cosmetic compositions through novel cell culture techniques.
1. A method of making a cosmetic composition comprising combining water, vitamin C, vitamin A, and a human Keratinocyte Fibroblast Serum (KFS),
wherein the human KFS is a compound mixture secreted by keratinocytes and fibroblasts in culture and substantially excludes other cell types, and wherein the human KFS is produced by the steps comprising:
(1) obtaining a cell culture by culturing epithelial cells derived from human neonatal foreskin in a cell culture container, the cell culture container comprising a first media that contains nutrients and one or more exogenous growth factors selected from L-Glutamine, hydrocortisone hemisuccinate, linoleic acid, Lecithin, HSA, rh FGF β, rh EGF/TGF β-1, rh-insulin, and ascorbic acid, wherein the first media is free from porcine derived components,
(2) culturing the cells in the first media until keratinocyte and fibroblast cells predominate a viable cell population in substantially equal proportions, and cells reach at least about 80 percent confluence,
(3) removing the first media from the cell culture container and washing the cell culture cells with a buffer solution,
(4) adding a volume of a sterile second media to the cell culture container, wherein the second media comprises nutrients of the first media but lacks exogenous growth factors of the first media, porcine-derived components, and bovine-derived components,
(5) culturing the cells in the second media for a sufficient period of time to produce a conditioned media comprising at least one of fibronectin, laminin, collagen, nidogen, vinculin, decorin, moesin, plastin-3 and lumican,
(6) removing and retaining a volume of the conditioned media from the cell culture container,
(7) replacing the removed volume of conditioned media with an equal volume of the sterile second media,
(8) repeating steps (6) and (7) at least once, and
(9) combining the retained volumes of the conditioned media to produce the human KFS.
2. The method of claim 1 , wherein steps (6) and (7) are repeated three times.
3. The method of claim 1 , further comprising filtering the conditioned media and/or the human keratinocyte fibroblast serum with a micro filter.
4. The method of claim 1 , wherein the first media is free from bovine derived components.
5. The method of claim 1 , wherein the conditioned media substantially excludes components from cells other than human keratinocytes and human fibroblasts.