【目的】基质力学微环境异常是导致骨性关节炎(osteoarthritis,OA)的关键因素之一。然而,软骨细胞在基质力学微环境发生变化过程中的炎症响应目前尚不清楚。【方法】利用聚二甲基硅氧烷(polydimethylsiloxane,PDMS)模拟正常和OA软骨细胞周基质(pericellularmatrix,PCM)硬度,制备不同硬度的细胞培养基底,定量分析基底硬度和炎症因子白细胞介素-1β(interleukin-1β,IL-1β)协同作用对软骨细胞形态、炎症介质和基质重塑蛋白表达的影响。首先,定量分析了不同基底硬度和IL-1β对软骨细胞前列腺素E2(prostaglandin,PGE2)和一氧化氮(NO)分泌水平的影响;其次,采用免疫荧光技术分析了IL-1β对不同硬度基底上软骨细胞铺展面积和核面积的影响;最后,采用Westernblot分析了不同硬度基底上软骨细胞Ⅱ型胶原(typeIIcollagen,COLII)和基质金属蛋白酶13(matrixmetalloproteinase-13,MMP13)蛋白水平的表达。【结果】实验结果表明,基底硬度介导软骨细胞对炎症信号IL-1β的响应。软基底显著增强PGE2和NO释放量(P<0.0001)及MMP-13的蛋白表达水平(P<0.05),而IL-1β能进一步增强这一调控作用;硬基底能显著促进软骨细胞铺展面积、核面积(P<0.0001)和Ⅱ型胶原蛋白表达水平(P<0.01),IL-1β加入后同样能进一步增强这种调控作用。【结论】有助于揭示软骨细胞感受基质力学微环境的力学生物学调控机制,同时也为优化细胞诱导性生物材料的设计提供了参考。

【Purposes】Osteoarthritis (OA) is a degenerative joint disease that commonly occurs in middle-aged and elderly people.  Mechanical microenvironment is one of the most significant factors in the development of OA.  However, when the mechanical microenvironment changes, the inflam-matory response of chondrocyte is elusive.  【Methods】By adopting polydimethylsiloxane (PDMS) substrates with varying stiffness which can mimic the physiological stiffness of chondrocyte pericellu-lar matrix (PCM), influences of co-regulated substrate stiffness and inflammatory factors interleu-kin-1β (IL-1β) on chondrocyte morphology, inflammatory mediators, and PCM remodeling pro-tein expression are quantitatively analyzed.  First, prostaglandin E2 (PGE2) and nitric oxide (NO) released in different stiffness substrates and IL-1β stimulated substrates with chondrocytes are de-tected.  Second, the changes in different stiffness substrates and IL-1β stimulated substrates through immunofluorescence technique are observed and recorded.  Third, the protein expressions of type Ⅱ collagen (COLII) and matrix metalloproteinase-13 (MMP13) are measured by Western blot assay.【Findings】The experimental results identify that substrate stiffness regulates the response of chon-drocyte to inflammatory signals.  Soft substrate dramatically enhances the release of PGE2 and NO (P<0. 000 1), and MMP13 (P<0. 05) expression, with  IL-1β further enhances this regulation.  In addition, stiff substrate significantly increases chondrocyte spreading area (P<0. 000 1) and COLII expression level (P<0. 01), IL-1β will further enhance this regulation.  【Conclusions】This study will shed light into the mechanobiological mechanism of the chondrocyte sensing matrix me-chanical microenvironment, and provide a reference for optimizing cell-inductive biomaterials.