How to improve skin elasticity: COSSMA

Because improving skin elasticity is a major objective in cosmetics, the elastin precursor TE could play a tremendous role in the formulation of body care products, as Prof Dr Dr Reinhard H.H. Neubert from the University of Halle-Wittenberg explains.

Lately, intensive developments have been realised in research, marketing and commerce in the field of cosmetics to generate new and effective actives for improving the elasticity of the human skin. Identifying relevant macromolecules is a major concern.

Macromolecules have a short history in cosmetics; one of the first macromolecules in cosmetics was hyaluronic acid.

The latest cosmetically relevant macromolecule in the pipeline is tropoelastin (TE). Elasticity is one of the requirements as organisms transform into complex multicellular systems.

Elastin is the main component of elastic fibres and provides tissues with this exceptional property.

The protein of elastic fibres is responsible for the elasticity and resilience of organs: skin (2–8%)1, lung (7%), major vascular vessels (32%), aorta (57%) and biomaterial2. Mature elastin is the most stable protein in the extracellular matrix (ECM) located in the human dermis. However, age and sun exposure have a damaging effect on dermal elastin. Fig. 1 shows the difference between a sun-protected 6-year-old (A) and a sun-exposed 90-year-old (B)¹.

No wonder that improving skin elasticity is a major objective in cosmetics. The elastin precursor TE could play a tremendous role in the formulation of body care products.

fig. 1: The impact of age and sun on elastin fibres: A: sun-protected 6-year and sun-exposed 90-year old1

How does tropoelastin work in the biosynthesis of elastin?

In elastin biosynthesis, the monomer TE, (see fig. 2), is the key protein. The protein with a size of 60–72 kDa is water-soluble, in contrast to elastin. TE is secreted, then rapidly matures into a complex polymer through a process of extensive cross-linking between TE units (see fig. 3). Cross-linking is a multistep process that begins with the alignment of TE units so that lysin residues come to proximity.

Coacervation appears to facilitate the alignment process and thus help the oxidative deamination of certain lysin residues to form α-aminoadipic-δ-semialdehyde (allysine). Deamination is catalysed with the help of a family of enzymes known as lysyl oxidases (LOXs). These are the key enzymes in crosslinking and present in a lot of tissues such as skin and cartilage.

fig. 2: Idealised model of tropoelastin structure.Source: adoption of 3

How is tropoelastin produced?

Biotechnological protein production in microorganisms such as bacteria or yeast is of enormous importance in many economic fields. The best studied model organism is Escherichia coli (E. coli), named after the German doctor Theodor Escherich. Genetically modified strains of this bacterium were used to produce TE. E. coli can grow in defined cultivation media and can be cultivated in a large-scale bioreactors (see fig. 4), here with a maximum working volume of 200 litres.

It is not possible to isolate native TE from relevant tissues. Therefore, TE has to be produced recombinantly in E. coli. Separating the host cell proteins from the desired product, e.g. by precipitation is crucial. Unlike the E. coli proteins, TE is soluble in a special solvent mixture. TE is freeze-dried after further downstream processing and can be stored until its application. This means that recombinant human TE can be obtained in sufficient quantities in a relatively short time.

How can tropoelastin be characterised analytically?

Analytical characterisation of intact TE is a challenging task. With the help of HPLC and gel-based separation techniques such as two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), the entire TE molecule can be analytically determined (see fig. 5).

The quality of TE can be characterised after digestion using suitable enzymes such as matrix metalloproteinases (MMPs) by effective mass spectrometric (MS) techniques such as tandem MS (also known as MS/MS or MS2).

fig. 5: Gel electropherogram of tropoelastin in production medium after 0, 0.5, 1.5, and 2 hours of production

How can the ingredient be used in cosmetics?

Using TE in cosmetics makes sense because the LOXs are present in the human dermis. As shown above, the LOXs are responsible for the crosslinking of TE to create elastin, resulting in greater elasticity of the skin. The most efficient means of delivery is applying TE into the dermis with the help of suitable syringes; however, non-invasive delivery can be helpful, too. As mentioned before, it would be extremely helpful to apply TE in suitable formulations for the following cosmetic applications:

to aged skin for recovery of skin elasticity
repair the defective elastic fibres (improvement of skin repair mechanisms)
artificial skin in combination with hyaluronic acid to improve skin elasticity and hydration to sustain skin moisture
wrinkle reduction, etc.

The way forward

In cosmetics, searching for new and effective active ingredients is a challenge. The new and effective macromolecule tropoelastin for formulations and other cosmetic issues improves the elasticity of the skin. Future uses of TE include uses in cartilage because the LOXs are present in human cartilage, in other tissues (e.g. human vessels), in wound care, and in the support of artificial skin.

The non-invasive delivery of macromolecules such as TE to the skin is a tremendously challenging issue, but it would be extremely helpful for cosmetic use of TE.

These procedures are in the focus of cosmetic delivery of TE⁴:

physical methods such as microneedles, microjets, laser beams, electroporation, sonophoresis, and iontophoresis
chemical methods such as penetration enhancer molecules
effective formulations such as colloidal carrier systems (CCDs)

From the procedures mentioned, the application of CCDs appears to be the most effective way for the cosmetic use of TE.

The Investitionsbank Sachsen-Anhalt financially supported the Joint Tropoelastin Project realised by Prof Markus Pietzsch (MLU), Prof Johannes Wohlrab (MLU) and Prof Reinhard Neubert (IADP)

References

¹ Mora Huertas, A.C. et al. (2016) Degradation of tropoelelastin and elastin by neprilysin. Biochimie, 128-129, 163-173.

² Mithieux, S.H., Rasko, J. & Weiss, A. (2004). Synthetic Elastin Hydrogels Derived from Massive Elastic Assemblies of Self-Organized Human Protein Monomers. Biomaterials, 25, 4921-4927.

³ Tamburro, A.M., Pepe, A. and Bochicchio, B. (2006) Localizing alpha-helices in human tropo-elastin: assembly of the elastin “puzzle”. Biochemistry, 45, 9518-9530.

⁴ Münch, S., Wohlrab, J. and Neubert, R.H.H. (2017) Dermal and transdermal delivery of relevant macromolecules. European Journal of Pharmaceutics and Biopharmaceutics, 19, 235-242.

⁵ Farjanel, J., Se`ve, S., Borel, A., Sommer, P. & Hulmes, D. J. S. (2005) Inhibition of lysyl oxidase activity can delay phenotypic modulation of chondrocytes. OsteoArthritis and Cartilage 13, 120-128.