Neocollagenesis after Ellansé™ injection. Research results

Author: Pierre Nicolau, plastic surgeon, teacher, member of the International Society of Aesthetic Plastic Surgery ISAPS (France)

Source: KOSMETIK international journal, No. 3/2013, pp. 62-64

As an advertisement

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Ellanse™ fillers (Ellanse-S, Ellanse-M, Ellanse-L and Ellanse-E), developed and manufactured by AQTIS Medical BV, are biostimulants and, unlike other dermal fillers, have stable effectiveness, a unique predictable duration of action from 1 years (Ellanse™-S) up to 4 years (Ellanse™-E) and complete and predictable biodegradation (STaT™ technology). Ellanse™ consists of perfectly smooth, spherical, non-crosslinked polycaprolactone (PCL) microspheres uniformly suspended in a special water-based carboxymethylcellulose (CMC) gel carrier. Figure 1 shows SEM and optical microscope images of PCL microspheres, demonstrating the smooth surface of the particles and the ideal spherical shape of the microspheres.

Rice. 1. Images of PCL microspheres obtained using SEM and optical microscope

PCL and CMC are successfully used in numerous CE and FDA approved self-absorbable materials used in medicine, cosmetics and pharmacy - for example, in other dermal fillers, plastic and maxillofacial surgery, wound dressings and controlled drug administration.

The physical characteristics of the filler, such as particle size 25–50 microns, concentration, smooth surface, ideally spherical particle shape, viscosity and elasticity of the carrier gel, gel uniformity and injectability, are identical for the entire Ellanse™ family.

The unique distinguishing feature that creates the predicted duration of action is the initial length of the PCL polymer chain within the microspheres. The variable initial length of this polymer chain is the basis for the controlled and predictable duration of action of Ellanse™.

PCL as a self-resorbable medical polymer is very promising due to its controlled and safe biodegradation through hydrolysis of ester bonds, resulting in the formation of non-toxic products that are quickly eliminated from the body through natural metabolic pathways. These data have been obtained from numerous scientific biocompatibility studies and have been published. Complete controlled biodegradation of PCL has been proven in studies of H and C-labeled PCL implants. Extensive safety studies have been conducted on the Ellanse™ implant formulation, including in vitro, preclinical and clinical studies. The results of this work show that all Ellanse™ fillers are safe and biocompatible and demonstrate excellent performance.

Ellanse™ is the first dermal filler to use fully self-absorbable, non-cross-linked PCL microspheres to volumize soft tissue.
All these fillers are CE marked and are on the market in more than 70 countries.

Dermal fillers

Although all dermal fillers can add volume after injection, they can be divided into two categories: those that stimulate collagen production (biostimulants) and those that do not stimulate collagen production (volumizants). The properties of new collagen fibers, such as the type and density around the particles underlying the fillers, depend on several characteristics of the dermal filler - in particular, particle volume, uniformity of microspheres in the gel, viscosity and elasticity of the gel, absorption time of the carrier gel, particle size , microsphere size, microsphere shape, surface smoothness/roughness and biodegradation process.

Collagen is the main protein of connective tissue. There are different types of collagen, but collagen types I and III predominate in the skin. Type III collagen is most often found in rapidly growing tissues. As body growth slows, the content of type III collagen in the skin decreases, while the content of type I collagen, which is coarser and more stable, increases.

Collagen in the skin is formed by fibroblasts. It has been noted that as the body responds to the introduction of foreign bodies with dermal filler, type III collagen first appears, which is gradually replaced by type I collagen.

Ellanse™ filler, designed for implantation into the deep dermis and hypodermis, consists of unique, completely smooth PCL microspheres, specially designed for optimal biocompatibility. Thanks to the optimal viscosity and elasticity of the carrier gel, wrinkles and folds are instantly corrected immediately after the procedure. The carrier gel is taken up by macrophages gradually over several weeks after the procedure, during which time PCL microspheres stimulate neocollagenesis to replace the volume of absorbed vehicle. Microspheres are not absorbed by phagocytes due to their size and surface characteristics. PCL microspheres trigger human skin's natural response and stimulate the natural wound healing process through neocollagenesis, which results in the deposition of new collagen around the microspheres.

Research method

The study was conducted at the Central Animal Facility of Utrecht University (GDL, Holland) with the approval of the Animal Ethical Committee (DEC, Utrecht University, Holland). Histological staining and analysis were performed in the Department of Pathobiology, Faculty of Veterinary Medicine. The study was conducted in accordance with ISO 10993: Evaluation of the biological effects of medical devices; part 6 (2007): Studies of local effects after implantation.
Two New Zealand White rabbits received a bolus injection of 0.1–0.2 ml Ellanse™-S and Ellanse™-M intradermally laterally along the spinal column (Fig. 2) .

Rice. 2. Photo of a rabbit after a bolus injection of PCL dermal filler

For histological analysis, a tissue sample was taken from each injection site and preserved in neutral buffered formalin 10%. The slides were routinely stained with hematoxylin and eosin (HE) and covered with picrosirius red (PSR). The PSR method is a selective histochemical reaction for the detection of collagen in paraffin-embedded tissue and is used as a specific stain for collagen types I and III. This dye is known to improve the birefringence of collagen fibers. PSR binds to collagen and then exhibits a bright refractive appearance when illuminated with a plane of polarized light when observed using crossed polarizers. Unpolarized PSR is specific for collagen, which is red in color. When polarized, large type I collagen fibers exhibit yellow or orange birefringence, whereas smaller type III collagen fibers exhibit green birefringence.

Research results

9 months after injection

Figure 3 shows a micrograph of an H&E-stained histological specimen of injected tissue 9 months after injection.

Rice. 3. Photomicrograph of a histological specimen stained using the H&E technique, 9 months after intradermal injection of Ellanse™-S: with hair follicles at the top, mature dermal collagen in the middle, dermal muscles at the bottom

Rice. 4. Photomicrograph of PSR-stained tissue under non-polarized light 9 months after Ellanse™-M administration. Noticeably homogeneous distribution of PCL microspheres and their fixation in the collagen structure (*)

The photomicrograph shows mature collagen without any trace of CMC carrier gel or PCL microspheres, indicating that the carrier gel and PCL microspheres have self-resorbed and been cleared by the body from the injection site. A micrograph of tissue under unpolarized light 9 months after Ellanse™-M administration is shown in Figure 4.

Red staining confirms the formation of new collagen around the PCL microspheres, which results in the fixation of the collagen structure by the PCL microspheres. The PCL microspheres were still homogeneously distributed, and no migration or encapsulation was detected around the implant.

Figure 5 shows a more detailed view of the same injected tissue. 9 months after injection, Ellanse™-M microspheres are still spherical in shape and are not affected by this stage of the self-resorption process. A photomicrograph of a non-polarized PSR-stained histological specimen confirms collagen deposits (Fig. 5a) . Polarized (Fig. 5b) PSR staining revealed the presence of both red-orange and green birefringence, indicating the deposition of type I and III collagen fibers, hence confirming neocollagenesis caused by completely smooth and spherical PCL microparticles. 9 months after injection, PCL microspheres are anchored in new collagen with a high-quality structure.

Rice. 5a. Photomicrograph of PSR-stained tissue under non-polarized light 9 months after Ellanse™-M administration. The picture shows the newly formed collagen structure

Rice. 5b. Photomicrograph of PSR-stained tissue under polarized light 9 months after Ellanse™-M administration. Figure demonstrates typical orange-red and green birefringence with red (arrows) type I collagen and green (arrowheads) type III collagen

21 months after injection

Figure 6 shows a photomicrograph of tissue 21 months after administration of Ellanse™-M (under (a) non-polarized light and (b) polarized light). Ellanse™-M microspheres are still present, embedded in the collagen structure. Polarized PSR staining (Fig. 6b) demonstrates the presence of red-orange birefringence, revealing the presence of type I collagen fibers.

Rice. 6a. Photomicrograph of a PSR-stained histological specimen under unpolarized light 21 months after Ellanse™-M administration. Red coloration confirms the formation of new collagen

Rice. 6b. Photomicrograph of a PSR-stained histological specimen under polarized light 21 months after Ellanse™-M administration. Red coloration indicates the presence of type I collagen fibers

conclusions

The data presented demonstrate the biocompatibility and self-absorbability of PCL dermal filler. Histological analysis confirmed that PCL microspheres of both product versions induce the formation of new collagen. Due to the smoothness and spherical shape of PCL microparticles, a high-quality structure of new collagen is formed.

After nine months, Ellanse™-S PCL microspheres were completely self-absorbed. This is consistent with data from a clinical study following treatment of the nasolabial fold, where clinical effectiveness decreased after 12 months.

Ellanse™-M PCL microspheres were still present 9 months after administration, were still spherical in shape, and were uniformly distributed throughout the tissue, with no evidence of migration or encapsulation. Ellanse™-M PCL microspheres were still present 21 months after injection, embedded in a collagen structure consisting predominantly of type I collagen fibers. Thus, the data obtained demonstrate the robust and long-lasting presence of collagen and the replacement of type III collagen by larger type I collagen fibers I, which is consistent with the normal physiological process of wound healing.