Chang Yijin, Wang Yaguang, Liu Jiansheng, Chen Xu, Ma Xuejing, Hu Yu, Tian He, Wang Xiaomei, Mu Changzheng
School of Basic Medicine, Jinzhou Medical University, Liaoning 121000, China.
First Affiliated Hospital of Jinzhou Medical University, Liaoning 121000, China.
Heliyon. 2023 Sep 7;9(9):e19879. doi: 10.1016/j.heliyon.2023.e19879. eCollection 2023 Sep.
The ability of cartilage to regenerate and repair is limited. N-acetyl- d-glucosamine (GlcNAc) is a nutritional supplement commonly used to activate chondrocytes. To prolong the duration of action of GlcNAc and improve its curative effect after cartilage injury, a GlcNAc thermosensitive hydrogel is prepared based on Pluronic F127 (PF127). The physicochemical properties results indicate that this hydrogel is injectable and retards the release of GlcNAc. Further, the therapeutic benefits of GlcNAc hydrogel are detected through intra-articular injection in rat specimens with cartilage injury. Behavioral experiments results indicate that the rats treated with GlcNAc hydrogel had longer step lengths, smaller foot angles and slower fall times. Compared with the sham group, the expression of Sox9 was 1.5 times and the level of collagen II was 2.4 times in the hydrogel treated group. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining result confirmed that the GlcNAc hydrogel reduce apoptosis by about 50%. Our results of immunohistochemical staining, Western blotting assays and enzyme activity detection all suggested that GlcNAc hydrogel reduce the expression of cleaved-caspase3 and caspase8 (Compared to the sham group, the protein contents were reduced by about 50% in the GlcNAc hydrogel group). We also found that GlcNAc hydrogel activates autophagy through ERK signal pathway. The results of Western blotting indicated that GlcNAc hydrogel increase the levels of LC3B and Becline1 (hydrogel group & sham group, LC3B: 1.56 ± 0.07 & 1.00 ± 0.14; Becline1: 1.98 ± 0.07 & 1.00 ± 0.13). Whereas, the content of P62 reduced after GlcNAc hydrogel treatment, the relative level in sham group and hydrogel group are 1.00 ± 0.02 and 0.73 ± 0.06. Our results revealed that the number of P-ERK positive cells in the hydrogel group (57.36 ± 3.56%) was higher when compared with the sham (24.82 ± 2.72%). And, the ratio of P-ERK and ERK was higher than that in the sham group (1.48 ± 0.07 & 1.00 ± 0.08). The GlcNAc thermosensitive hydrogel is a promising and sustainable drug delivery system for intra-articular injection in the treatment of cartilage injury.
软骨的再生和修复能力有限。N-乙酰-D-葡萄糖胺（GlcNAc）是一种常用于激活软骨细胞的营养补充剂。为了延长GlcNAc的作用时间并提高其在软骨损伤后的治疗效果，基于泊洛沙姆F127（PF127）制备了一种GlcNAc热敏水凝胶。物理化学性质结果表明，这种水凝胶可注射且能延缓GlcNAc的释放。此外，通过对软骨损伤大鼠标本进行关节内注射来检测GlcNAc水凝胶的治疗效果。行为学实验结果表明，用GlcNAc水凝胶治疗的大鼠步长更长、足角更小且跌倒时间更慢。与假手术组相比，水凝胶治疗组中Sox9的表达是假手术组的1.5倍，胶原蛋白II的水平是假手术组的2.4倍。末端脱氧核苷酸转移酶dUTP缺口末端标记（TUNEL）染色结果证实，GlcNAc水凝胶使细胞凋亡减少了约50%。我们的免疫组织化学染色、蛋白质免疫印迹分析和酶活性检测结果均表明，GlcNAc水凝胶降低了裂解型半胱天冬酶3和半胱天冬酶8的表达（与假手术组相比，GlcNAc水凝胶组的蛋白质含量降低了约50%）。我们还发现，GlcNAc水凝胶通过ERK信号通路激活自噬。蛋白质免疫印迹结果表明，GlcNAc水凝胶增加了LC3B和Beclin1的水平（水凝胶组和假手术组，LC3B：1.56±0.07 & 1.00±0.14；Beclin1：1.98±0.07 & 1.00±0.13）。然而，GlcNAc水凝胶处理后P62的含量降低，假手术组和水凝胶组的相对水平分别为1.00±0.02和0.73±0.06。我们的结果显示，与假手术组（24.82±2.72%）相比，水凝胶组中P-ERK阳性细胞的数量更高（57.36±3.56%）。并且，水凝胶组中P-ERK与ERK的比值高于假手术组（1.48±0.07 & 1.00±0.08）。GlcNAc热敏水凝胶是一种有前景的、可持续的关节内注射给药系统，可用于治疗软骨损伤。
