Effects Of N-acetylglucosamine On Hyaluronic Acid Synthesis In Human Epidermal Keratinocytes

Backgrounds and objectives: Hyaluronic acid (HA) is the linear chain giant molecular polycose composed of glucuronic Acid and N-Acetyl glucosamine (NAG), the polymer of keeping water which exists in the human skin. It is situated in extracellular matrix(ECM) as a water-absorbing macromolecule, and generally exists in vertebrate connective tissue and some bacteria. The strong extension and random curling of HA molecule result in tremendous domain, the hydrodynamics volume is ten times larger than the possessing space of its un-hydrated form. Further more, molecule chains twist between each other to form a consecutive reticular structure, which reacts with water molecule via polar bond and hydrogen bond. HA can absorb and keep water weighing up to thousand fold more than itself because of its structure like"molecule sponge", and is one of the best internationally accepted moisturizers , keeping extracellular space and promoting ion disolving and transportation of nutrient substances. Besides, proteoglycan composed of HA and protein , has the large molecular weight. Proteoglycan binding to water through glucidamin, exists in connective tissue. The cells are binded by the gelatum composed of HA-protein-water, to run normally the functions of cell metabolism and tissue keeping water, to protect themselves from pathogenic bacteria and prevent infection, and to keep tenacity and elasticity of the skin. Youthful and healthy skin is closely related to its high moisture capacity, which decreased as the age increased. And then the skin elasticity decreases, the gloss fades away and the skin become rough and aging. The previous study indicated that the moisture capacity of corium is related to HA content. The content of skin HA, comprised 50% of the total content, is the highest in the body. With in-depth study on the physiologic function of HA, HA applyingin skin care has received wide attention as a cosmetic ingredient.HA is mainly produced by human epidermal keratinocytes and dermal fibroblasts in skin, and NAG is one of the materials for HA synthesis.The present experiment is focused on adult human epidermal keratinocytes (HEKa), in order to study the effects of exogenous NAG on HA production in HEKa cells and investigate the molecular mechanism of the HA synthesis in HEKa cells affected by NAG.Methods: The cell proliferation of HEKa cells affected by NAG were measured by MTT method. Effects of NAG at different concentrations on HA production in HEKa cells were measured by Enzyme linked immunosorbent assay (ELISA). Effects of NAG on Hyaluronic acid synthetase(HAS)mRNA in HEKa cells , including HAS1, HAS2 and HAS3 mRNA, were measured by Reverse transcription-polymerase chain raction (RT-PCR), in order to study the molecular mechanism of NAG.Results: The result of MTT indicated that NAG at 1mM/L, 3mM/L and 5mM/L had no effect on cell proliferation of HEKa cells(P>0.05), but NAG at 10 mM/L significantly inhibited the cell proliferation of HEKa cells (P<0.05). The result of ELISA showed that NAG promoting HA production in HEKa cells in dose dependence(P<0.01),and the best concentration was 5mM/L. The result of RT-PCR showed that NAG had no effect on the expression of HAS1 and HAS2 mRNA in HEKa cells, however, NAG at 1mM/L and 5mM/L upregulated the expression of HAS3 mRNA. NAG at 10mM/L had no effect on the expression of HAS3mRNA.Conclusions:(1)NAG at different concentrations can promoting HA production and secretion.(2)NAG at high concentration inhibits the cell proliferation of HEKa cells, but there are no effect at lower concentrations, which show that the effect of promoting HA production and secretion is not through cell proliferation. However, the inhibitive effect needs further study.(3)NAG may up-regulate the level of HAS3 mRNA and increase the activity, and may act as material to promoting HA production in HEKa cells. (4)There is good development prospect of NAG as a cosmetic ingredient.