132 J. Cosmet. Sci., 74, 132–142 (March/April 2023) Address all correspondence to Mariana Brito da Cruz, mariana.cruz@campus.ul.pt Hyaluronic Acid Aesthetic Fillers: A Review of Rheological and Physicochemical Properties PATRÍCIA PAULINO, MARIANA BRITO DA CRUZ AND VIRGÍNIA SANTOS Faculdade de Medicina Dentária da Universidade de Lisboa, Lisboa, Portugal (P.P., V.S.) Unidade de Investigação em Ciências Orais e Biomédicas (UICOB), Faculdade de Medicina Dentária da Universidade de Lisboa, Lisboa, Portugal (M.B.D.C.) Accepted for publication July 01, 2023. Synopsis Hyaluronic acid’s (HA) main functions are absorbing water into the tissues and structuring the skin. It is mostly used in dermal fillers, treatments for certain diseases, and wound healing. This study intends to review current literature of HA’s rheology and its physicochemical properties as an injectable filler. Data were acquired from articles concerning HA-based biomaterials published within the last 25 years in PubMed and ScienceDirect. The MeSH terms “hyaluronic acid” and “dermal fillers,” were used either alone or combined with “rheology,” “physicochemical concepts,” “cross-linking reagents,” “viscoelastic substances,” “cohesivity,” and “cosmetic techniques.” Some articles not found during the initial search were chosen from the reference lists of previously selected publications. All articles that fit in the theme were considered valid regardless of study type. Available literature describes intrinsic properties of HA as a glycosaminoglycan. As an injectable filler, its rheology (viscoelasticity and cohesivity) and its physicochemical properties (cross-linking, hydrophilia, particle size, and HA concentration) define its clinical behavior by influencing its longevity, lifting capacity, resistance to external forces, and needle extrusion force. HA is promising as a dermal filler and healing agent. Understanding its properties is essential, as each patient benefits from different products. Future research should continue to explore these properties. INTRODUCTION Hyaluronic acid or hyaluronan (HA) is the most abundant glycosaminoglycan present in the human dermis, with around 50% of total body HA being found in the skin, and a component of all mammalians’ connective tissue. HA constitutes a compound of cell surfaces and extracellular matrix in the skin, eyes, joints and muscles, and umbilical cord. Its main functions are to draw water into the tissues, volumizing and giving structure to the skin by binding collagen and elastin fibres into a supportive matrix.1-4 Additionally, HA intervenes in inflammation regulation, drug delivery, angiogenesis, cell migration and proliferation (caused by HA binding itself specifically to proteins that are responsible for these processes, whether they are in the extracellular matrix, on the cell surface, where they’re called hyaladherins, or in the cellular cytosol), wound healing (based on its

133 HYALURONIC ACID AESTHETIC FILLERS antioxidant properties and ability to eliminate free radicals), and cancer progression (based on its hydrodynamics and ability to interact with tumour cell surfaces and influence the porosity and malleability of extracellular and pericellular matrices, and also based on the fact that an increase in HA links to apoptosis, invasiveness, and drug resistance).5-12 HA was first discovered in 1934 at Colombia University of New York by two American scientists, Karl Meyer and John Palmer, when they isolated it from bovine vitreous humor.1,5 HA was then commercialized for the first time by Endre Balazs, who used it as a substitute for egg whites in bakery products. More recently, HA has been used for numerous purposes, such as wound treatment, ophthalmic surgery, drug delivery, arthritis treatment (serving as an intra-articularly injected lubricant), and aesthetic treatment.1,2,13 The first biocompatible gel (hylan B gel, Hylaform) was created in 1980 by Balaz,5 and the first HA filler (Restylane) was approved in the United States in December 2003 for the correction of deep wrinkles and folds.14-17 In 2006, the American Society of Aesthetic Plastic Surgeons declared HA dermal fillers to be the fastest noninvasive aesthetic procedure in the United States.1 Currently, there is a wide panoply of different HA-based dermal fillers, each of them manufactured in a distinct way and with different characteristics, with none of them being a universally fitting filler for every situation.1,5,15,18 Currently, HA is also used in antiaging treatments, due to its high biocompatibility, volumizing effect, low potential for adverse reactions, reversibility in cases of complications, and possibility of storage without refrigeration for up to two years. Using this polysaccharide as a dermal filler is essential to facial harmonization and can gather many of the desired dermal filler properties. In fact, an ideal filler must be safe, biocompatible, efficient, easy to store, low cost, easily eliminated when necessary, and independent of allergy testing. Thus, clinicians should familiarize themselves with HA’s rheological and physicochemical properties as these properties will influence clinical performance. These fillers usually last from 6 to 18 months, depending on several factors such as the HA itself (cross-linking degree, HA concentration, particle size), skin type, medication, age, injection technique, physical activity, and the presence of free radicals in the tissues (particularly in the skin) that quickly degrade uncross-linked HA polymers (depolymerization), among others.5,14,19-23 Degradation of HA fillers by free radicals happens due to a transient inflammatory reaction derived from their injection into tissues, and it is caused by the cleavage of glycosidic bonds.12,23 Oxidative damage and enzymatic degradation represent the two mechanisms through which HA is separated in the body, and these two mechanisms are responsible for the degradation of 30% of the 15g of HA that is locally present. The other 70% of HA is systematically catabolized by the endothelial cells of lymphatic vessels.12 DATA SOURCES AND METHODS All information used to elaborate the present review was found using the PubMed and ScienceDirect primary databases, where we searched for and selected experimental studies (such as clinical trials and in vitro and in vivo studies, both in humans and animals) and reviews pertaining to the uses of HA in medicine, particularly those reviews related to aesthetic and healing procedures, and to HA’s intrinsic properties. Several research terms were used, such as “hyaluronic acid” and “dermal fillers,” both alone and combined with terms such as “rheology,” “physicochemical concepts,” “water absorption,” “cross-linking reagents,” “viscoelastic substances,” “particle size,” “cohesivity,” “soft tissue augmentation,”