Skin rejuvenation with type I collagen

Andrea Corbo, MD, professor at the University of Camerino (MC) (Italy)

Cecilia Luchi, medical director of the private clinic MedicalSpa (Italy)

Giovanni Merone, medical director of the private clinic MedicalSpa (Italy)

Biorejuvenation treatments rely on repeating multiple treatment sessions according to different protocols that aim to provide intermittent anabolic stimulation to dermal fibroblasts. As a result, the biosynthesis of dermal matrix components and hydration are enhanced, the effect of skin tightening and the restoration of the fibrillar network are observed. The results of biorejuvenation are usually satisfactory in the initial and middle stages of photo- and chronoaging of the skin, but periodic examinations are necessary. In the following study, the authors tested a new class III biorejuvenation drug based on type I equine collagen , which has a beneficial effect on the skin at all stages of aging. To do this, the authors of this article determined the optimal dosage of the drug, talked about the need for patient compliance with the treatment regimen and regimen, determined the degree of client satisfaction, and also proposed the best protocol for preserving and consolidating the results of treatment.

For 30 years now, horse collagen type I has been used in dermatology, along with plastic and vascular surgery, to heal wounds and treat ulcers. The most popular drug of this kind is made on the basis of lyophilized collagen tablets; it is usually injected directly into the wound and, by stimulating tissue regeneration, accelerates healing. It has been proven that horse collagen affects fibroblast mitosis, tissue regeneration and stimulation of the biosynthesis of new collagen.

Equine collagen is obtained from the Achilles tendon of the horse. During the manufacturing process, the structure of the collagen molecule does not undergo chemical changes such as protein denaturation or cross-linking. The only chemical manipulation involves separating the collagen triple helix sequences from the N- and C-terminal amino acids - thus reducing the risk of developing allergies.

In this study, equine collagen was micronized, the resulting powder was placed in vials and sterilized. Immediately before use, the crushed powder was dissolved in an aqueous solution of sodium chloride 0.9% or in water for injection with the addition of a small amount of anesthetic. The result was a suspension intended for intradermal administration.

The proposed mechanism of action is based mainly on the presence of tripeptides, in particular Pro-Hyp-Gly, obtained from the hydrolysis of collagen. In fact, hydrolysis of peptide bonds can occur both in vivo after injection of the drug into the receiving tissue, and in vitro - in the same vial after reconstitution of the product. Once in solution, horse collagen undergoes spontaneous hydrolysis, forming tripeptides and collagen amino acids. This guarantees the complete absence of any risk of immunogenicity, since the mass of molecules capable of causing an immune response in the body must exceed 10 kDa, while in solution the mass of molecules does not exceed 3⎼6 kDa (tripeptides)8.

Tripeptides are biological messengers that act similarly to growth factors. By interacting with transmembrane receptors, they activate anabolic pathways and stimulate the fibroblast cell cycle. Pro-Hyp-Gly tripeptide works as a competitive inhibitor of matrix metalloproteinases (MMPs), responsible for the enzymatic degradation of dermal collagen (collagenase).

Finally, free amino acids present in the solution, according to Michaelis ⎼ Menten kinetics, stimulate the work of fibroblasts and the synthesis of new collagen. Some in vitro studies have shown that young fibroblasts, that is, fibroblasts formed as a result of cell division, synthesize predominantly type III collagen, while older fibroblasts synthesize mainly type I collagen. It is known that the collagen I/III ratio increases as the skin ages.

Research and results

Three different dosages of equine collagen were used to determine the optimal dose. 120 patients aged 35 to 65 years were divided into three groups of 40 people each. Each group was prescribed different doses: 50, 100 and 200 mg of fine sterile powder diluted in 4.5 ml of saline with the addition of 0.5 ml of 2% lidocaine solution.

The resulting suspension was administered intradermally to the subjects using the acupressure mesotherapy method (about 0.1 ml per injection). Skin papules were concentrated on the face, neck, décolleté and arms at a distance of 2 cm. Each patient received 4 treatment sessions - one every 2 weeks for 8 weeks. The condition of the skin before and after treatment is documented in the photo. All side effects are recorded.

Of the side effects, the subjects noted mainly redness, increased temperature and discomfort, and transient pain. In the 50 mg dose group, one subject complained of side effects lasting from several hours to a day; in the group with a collagen dosage of 100 mg, one of the subjects noted side effects lasting from several hours to a day; in the group receiving 200 mg of the drug, eight patients reported side effects lasting from several hours to a day.

The dynamics of improvement in skin condition were assessed by the subjects themselves and the doctor. Patients were asked to complete a questionnaire to obtain their opinion of their own skin's texture, brightness, hydration and firmness. The results of the groups prescribed a dosage of 100 and 200 mg were similar, while the dynamics in the group with a dosage of 50 mg were worse.

The effectiveness of treatment was assessed by monitoring changes in the patient's skin condition after four treatment sessions. 20 patients in the 50 mg group reported improvement in skin appearance after 4 treatment sessions, while 38 patients in the 100 mg group reported improvement in skin appearance and 39 patients in the 200 mg group.

As a result, a dose of 100 mg was determined to be optimal for further treatment, since the ratio between the effectiveness of treatment and the risk of side effects in this case is better.

For complex treatment of the skin of the face, neck, décolleté and hands, 321 patients were injected with 100 mg of equine collagen (0.1 ml every 2 cm, 1 session every 2 weeks, 4 sessions in total) in accordance with the protocol described above (100 mg powder dissolved in 4.5 ml of physiological solution with the addition of 0.5 ml of lidocaine 2%, the total volume of the solution is 5 ml). As in the case of the previous group, a photo report was prepared and a survey was conducted.

Among the main side effects lasting from several hours to a day, noted by less than 1% of patients, are redness of the skin, itching, and the occurrence of minor pain or discomfort in the treated areas of the skin. Data collection did not take into account temporary discomfort at the end of the treatment session, as this occurs due to any injection, regardless of the material used.

As a result of treatment, the texture, structure, brightness and hydration of patients' skin improved in more than 95% of cases. Almost all subjects noted restoration of skin texture, 3 people did not notice changes in its structure, 46 patients noted that the brightness of the skin did not change, and another 27 did not observe any moisturizing effect after treatment.

The split face test was performed on 64 patients, who were divided into two groups of 32 people. The drug was used at a dose of 100 mg.

In the first group of patients, half of the face was treated with a freshly prepared preparation, and the other half with a 2-day-old solution, which was stored at a temperature of 4°C. In the second group, half of the face was treated with a freshly prepared solution, the other half with a solution prepared 4 days ago. Processing was carried out in accordance with the protocol described above. In all cases, the dosage was determined in accordance with the standard protocol, the solution was prepared using the specified volume of saline and anesthetic and was administered pointwise in equal volumes (0.1 ml every 2 cm).

During the test, the authors found a significant decrease in efficiency when using a stale solution: the response was weak. Skin areas where the authors injected the solution prepared 4 days earlier showed a very weak response, while the 2-day preparation showed better results. According to patient assessments, only those areas of the skin that were treated with the freshly prepared suspension showed the expected results. The data obtained were confirmed by clinical evaluation.

At the last stage of the study, the authors tested a protocol to maintain and consolidate treatment results. 80 patients were divided into two subgroups (A and B) of 40 people each. For each group, the dosage of the drug was different. Group A was treated with 70 mg of collagen per session and patients were given 3 sessions every 2 weeks for a total dose of 210 mg, while in Group B patients were given 100 mg in each of the 4 sessions 2 weeks apart. collagen, a total of 400 mg. Results were monitored over a period of 6 months and included clinical examination and questionnaires.

According to the study results, the authors observed a more satisfactory result 6 months after 4 sessions of collagen treatment at a dosage of 100 mg once every 2 weeks.

During monthly examinations of patients, it turned out that the treatment results reported by them during the survey declined faster in the group in which the subjects were prescribed three sessions of administering the drug at a dose of 70 mg per session.

Summary

In the field of cosmetic medicine, there is a growing demand for procedures that improve the appearance of the skin and have primarily a rejuvenating therapeutic effect, and not just aimed at correction or increase in volume.

Therefore, today the market is ready to offer a wide range of injection products with anti-aging effects. In particular, non-crosslinked hyaluronic acid has become widely used. Anti-aging injections of hyaluronic acid mainly affect the hydration of skin cells, correlated with the effect of cleaning reactive oxygen species, but there remains room for doubt.

On the other hand, it essentially activates the expression of the metalloproteinase gene, which leads to increased degradation of the dermal matrix. For the same reason, it is recommended to combine collagen treatment with the use of other biostimulants that will help increase collagen production and reduce the activity of metalloproteinases. The main limiting factor for classical biorejuvenating/biostimulating injection procedures (polynucleotides, vitamins) is the residual anabolic effect of old fibroblasts. Therefore, it is necessary to explore the possibility of using equine collagen. It has been proven to stimulate fibroblast mitosis and tissue regeneration. Young fibroblasts, formed as a result of mitosis, have a greater anabolic effect and the ability to produce more type III collagen than old cells. Finally, more than 30 years of history of using horse collagen suggests the absence of an allergic reaction to it and the high safety of the drug, which is very important when it comes to cosmetic injections.

Read the full version of the article in the magazine Les Nouvelles Esthetiques Ukraine 6 (106)/2017