Stoppoloni D, Politi L, Leopizzi M, Gaetani S, Guazzo R, Basciani S, Moreschini O, De Santi M, Scandurra R, Scotto d'Abusco A
Dept. of Biochemical Sciences, Sapienza University of Roma, P.le Aldo Moro, 5, 00185 Roma, Italy.
Dept. of Medico-Surgical Sciences and Biotechnologies, Faculty of Medicine and Pharmacy, Sapienza University, Polo Pontino, Corso Della Repubblica 79, Latina, Italy.
Osteoarthritis Cartilage. 2015 Jan;23(1):103-13. doi: 10.1016/j.joca.2014.09.005. Epub 2014 Sep 16.
Aim of this study is to investigate the effects of Glucosamine (GlcN) and its peptidyl-derivative, 2-(N-Acetyl)-L-phenylalanylamido-2-deoxy-β-D-glucose (NAPA), on extracellular matrix (ECM) synthesis in human primary chondrocytes (HPCs).
Dose-dependent effect of GlcN and NAPA on Glycosaminoglycan (GAG), Collagen type II (Col2) and Small Leucine-Rich Proteoglycans (SLRPs) was examined by incubating HPCs, cultured in micromasses (3D), with various amounts of two molecules, administered as either GlcN alone or NAPA alone or GlcN plus NAPA (G + N). Immunohystochemical and immunofluorescent staining and biochemical analysis were used to determine the impact of the two molecules on ECM production. Gene expression analysis was performed by TaqMan Real-Time Polymerase Chain Reaction (PCR) assays.
The lowest concentration to which GlcN and NAPA were able to affect ECM synthesis was 1 mM. Both molecules administered alone and as G + N stimulated GAGs and SLRPs synthesis at different extent, NAPA and mainly G + N stimulated Col2 production, whereas GlcN was not effective. Both molecules were able to induce Insulin Growth Factor-I (IGF-I) and to stimulate SOX-9, whereas NAPA and G + N were able to up-regulate both Hyaluronic Acid Synthase-2 and Hyaluronic acid. Very interesting is the synergistic effect observed when chondrocyte micromasses were treated with G + N.
The observed anabolic effects and optimal concentrations of GlcN and NAPA, in addition to beneficial effects on other cellular pathways, previously reported, such as the inhibition of IKKα, could be useful to formulate new cartilage repair strategies.
本研究旨在探讨氨基葡萄糖（GlcN）及其肽基衍生物2-(N-乙酰基)-L-苯丙氨酰胺基-2-脱氧-β-D-葡萄糖（NAPA）对人原代软骨细胞（HPCs）细胞外基质（ECM）合成的影响。
通过将以微团形式（3D）培养的HPCs与不同量的两种分子单独孵育，即单独使用GlcN或NAPA或GlcN加NAPA（G + N），来检测GlcN和NAPA对糖胺聚糖（GAG）、II型胶原蛋白（Col2）和富含亮氨酸的小分子蛋白聚糖（SLRPs）的剂量依赖性效应。采用免疫组织化学和免疫荧光染色以及生化分析来确定这两种分子对ECM产生的影响。通过TaqMan实时聚合酶链反应（PCR）测定法进行基因表达分析。
GlcN和NAPA能够影响ECM合成的最低浓度为1 mM。单独使用以及作为G + N使用的这两种分子均在不同程度上刺激了GAG和SLRPs的合成，NAPA以及主要是G + N刺激了Col2的产生，而GlcN则无效。这两种分子均能够诱导胰岛素生长因子-I（IGF-I）并刺激SOX-9，而NAPA和G + N能够上调透明质酸合酶-2和透明质酸。当用G + N处理软骨细胞微团时观察到的协同效应非常有趣。
除了先前报道的对其他细胞途径的有益作用（如抑制IKKα）外，观察到的GlcN和NAPA的合成代谢作用以及最佳浓度可能有助于制定新的软骨修复策略。
