Restoring hyaluronic acid (HA) content is important for maintaining the function of photo-aged skin. This study aimed to evaluate the passive delivery into skin of HA nanoparticles formed by the polyion complex method. Nanoparticles were prepared by mixing and stirring anionic HA with a cationic polymer, protamine, at the charge ratio 55:45. The permeation of fluorescently-labelled HA nanoparticles (HANP) or free HA through hairless mouse skin was characterized in vitro. HANP or free HA was applied to ultraviolet (UV)-irradiated mice in vivo, and their transepidermal water loss (TEWL) was measured after 4 days. HA that had been delivered into skin was separated and characterized by molecular sieve chromatography. HANP were able to deliver HA into the dermis both in vitro and in vivo, whereas free HA penetrated no further than the stratum corneum. Following HANP application, HA within the skin was present in the form of free HA rather than nanoparticles. When applied in vivo, HANP significantly reduced the TEWL caused by UV irradiation. Thus, although free HA does not penetrate into the skin by passive diffusion, HA can be effectively delivered by nanoparticles. HA is then released from the nanoparticles and can contribute to barrier recovery following UV irradiation.

The permeation pathway of macromolecules and nanospheres through skin was evaluated using fluorescent isothiocyanate (FITC)-dextran (average MW, 4 kDa) (FD-4) and nanospheres (500 nm in diameter) in hairless rat abdominal skin and porcine ear skin as well as a three-dimensional cultured human skin model (cultured skin model). A low molecular hydrophilic compound, sodium fluorescein (FL) (MW, 376 Da), was used for comparison. FL penetrated the stratum corneum and permeated the viable epidermis of hairless rat skin, whereas less permeation of FL was observed through the cultured skin model, suggesting that the primary permeation pathway for the hydrophilic material may be skin appendages through the rat skin. A macromolecular compound, FD-4, was distributed through the hair follicles of the rat skin. In addition, nanospheres were detected in the hair follicles of porcine skin, although no skin permeation was detected. These findings suggest that appendage routes such as hair follicles can be a penetration pathway of macromolecules and nanospheres through skin.