136 JOURNAL OF COSMETIC SCIENCE argue that all fillers should be considered monophasic as they all have the same composition throughout and believe that there is not a real phase separation.16,17 Some fillers also have lidocaine in their composition to reduce the pain that can accompany this treatment, both during the injection and after it. Lidocaine appears to alter some of HA’s properties, such as G’.4,30 Cohesivity. Cohesivity is an essential property for determining the filler’s integrity, contributing to it maintaining its microscopic shape after being injected into the patient’s tissues.24 It corresponds to the internal adhesion forces that bind the different cross-linking units together within the gel and translates the resistance to vertical compression/stretching forces. In this way, it defines the initial vertical projection of the dermal filler.17,19,25 It also has a role in defining the filler’s modeling capacity since a less cohesive gel is a more malleable one. However, this property’s clinical relevance decreases over time. As the filler is integrated into the tissues, it naturally becomes less malleable and more stuck in place. Cohesivity depends on HA concentration and cross-linking technique, but it is not influenced by the cross-linking degree unlike a lot of other HA properties.17,25 It has been suggested that lower cohesivity values contribute to a more uniform distribution of the dermal filler in the tissues, reducing lump formation and allowing for a more superficial placement without inducing the Tyndall effect (when the skin gains a bluish tone due to superficial dermal filler placement),27 and that the more cohesive a product is, the bigger its tissue integration and lifting capacity are. Still, this property’s clinical relevance remains widely debated, due to the lack of standardized measurement techniques and conflicting opinions on cohesivity’s true effects on fillers.16,26 There are currently four known methods for determining cohesivity, even though none of them tend to provide consistent data: the linear compression test, the average drop-weight, the dye diffusion test, and the Gavard-Sundaram Cohesivity Scale.16 It has also been noted that measuring a gel’s cohesivity before injection is irrelevant, since there is still uncross- linked HA, which will be quickly degraded postinjection making the filler more cohesive.31 PHYSICOCHEMICAL PROPERTIES Cross-Linking. Other deeply important characteristics of HA fillers are their clinical persistence and durability, which are also influenced by the gel’s viscous and elastic components in synergy with other important properties. For example, a higher cross-linking degree (which is the percentage of HA disaccharide monomer units bound to a cross-linker molecule), HA concentration, particle size, and molecular weight tend to increase HA’s biostability and resistance to degradation over time. Due to these qualities, adding 1,4-butanediol diglycidyl ether (BDDE) or other cross-linking agents to the filler’s formula is important as HA’s natural duration is one of its main limitations. These cross-linking agents modify the HA by creating “bridges” (ether bonds) between its molecules, which increase the filler’s biostability by transforming the filler from a viscous liquid into a gel and making it harder for the filler to be degraded by hyaluronidase and increasing resistance to oxidative stress.2,3,14,18-21,32 Cross-linking degree is reported by many as the most influential factor for rheological properties, particularly when it comes to gel stiffness.31 However, even though they are rare, hypersensitivity reactions tend to happen because of the cross-linking process that is caused by the epoxide groups in the residual BDDE

137 HYALURONIC ACID AESTHETIC FILLERS (a BDDE molecule that hasn’t reacted with any other molecules). Because of this, the amount of BDDE used in each filler is maintained at trace amounts (2 ppm), so that its clinical use can be considered safe. Besides residual BDDE, this agent can be present in dermal fillers in three other states: a fully reacted cross-linker (a BDDE molecule that reacted with HA on both ends), a pendant cross-linker (a molecule that reacted with only one end of HA), and a deactivated cross-linker (hydrolyzed BDDE).3,5,23 Degrees of cross- linking that are considered too high can lead to problems with biocompatibility because the HA is furthest away from its natural form. As a result, the organism may perceive the HA as more foreign.4 In recent years, a different cross-linking agent from BDDE has been studied and more frequently used showing lower levels of cytotoxicity. This agent is part of Neauvia Stimulate, a HA filler of bacterial origin cross-linked with PEG (poly-ethylene-glycol) and 1% of micromolecules of calcium hydroxyapatite. These micromolecules grant the neauvia acid its colagenesis activity, in addition to the filler’s volumizing effect. In an in vitro study published in 2018, this filler showed no cytotoxicity until at least 24 hours postapplication and caused no alterations to cell viability, morphology, or structure.33 Cross-Linking Technologies. As has been previously stated, dermal fillers from distinct brands are produced using different cross-linking technologies, which will shape the filler’s properties for the target tissue and desired postinjection effect.31,34 There is a large variety of cross-linking technologies including Vycross, nonanimal stabilized hyaluronic acid (NASHA), and Tri-Hyal. Vycross, which is used in Juvéderm fillers, is composed of a mixture of high-molecular- weight HA and a significantly higher ratio of low-molecular-weight HA (1 MDa). These fillers are cross-linked with BDDE at both ends (a fully reacted cross-linker) and show a narrower range of available G’ values. They tend to be harder gels and can be noncohesive or partially cohesive.31,34 Their homogeneous matrix is smoother rather than granular, making them highly malleable with a more even distribution in the tissues. The higher amount of low-molecular-weight HA and a lower overall amount of HA reduces the water absorption of these fillers, thus reducing their swelling. As previously described, the small percentage of noncross-linked HA present helps lower the overall extrusion force.30 Juvéderm fillers are monophasic and monodensified, which means they are produced by mixing the HAs and cross-linking them in one single moment. In comparison with most other FDA approved fillers, they also present a higher cross-linking degree and lower G’ values. Because of these characteristics, they tend to show greater longevity.35 Tri-Hyal is a technology applied by Fillmed in their ART FILLER gamma, which is advertised by the manufacturer as consisting of monophasic fillers, and it’s characterized by a combination of long chain, very-long chain, and free noncross-linked HA (which facilitates injection extrusion and creates more natural results). The triple cross-linked HAs, all with different molecular weights, provide a more suitable environment for dermal fibroblasts to produce extracellular matrix components, which contributes to skin self-renewal. This makes the filler a good choice as it derives its effect not only from its volumizing action, but also from being a rejuvenating agent. This technology also shows a natural entanglement of HA, which may allow a reduction in the amount of cross-linking agent used (such as BDDE) and a higher sculpting ease, cohesivity, and malleability. This triple cross-linking technique can provide a sustained release of free HA.36 It also has 0.3% of lidocaine hydrochloride in its composition for anaesthetic properties and a phosphate buffer at pH =7.2.37