Universidade de São Paulo, Faculdade de Odontologia de Ribeirão Preto, Bone Research Laboratory, Ribeirão Preto, São Paulo, Brasil.
Instituto Militar de Engenharia, Laboratório de Biomateriais, Rio de Janeiro, Rio de Janeiro, Brasil.
J Appl Oral Sci. 2020 Feb 7;28:e20190156. doi: 10.1590/1678-7757-2019-0156. eCollection 2020.
The present study aimed to investigate the participation of focal adhesion kinases (FAK) in interactions between osteoblastic cells and titanium (Ti) surfaces with three different topographies, namely, untreated (US), microstructured (MS), and nanostructured (NS).
Osteoblasts harvested from the calvarial bones of 3-day-old rats were cultured on US, MS and NS discs in the presence of PF-573228 (FAK inhibitor) to evaluate osteoblastic differentiation. After 24 h, we evaluated osteoblast morphology and vinculin expression, and on day 10, the following parameters: gene expression of osteoblastic markers and integrin signaling components, FAK protein expression and alkaline phosphatase (ALP) activity. A smooth surface, porosities at the microscale level, and nanocavities were observed in US, MS, and NS, respectively.
FAK inhibition decreased the number of filopodia in cells grown on US and MS compared with that in NS. FAK inhibition decreased the gene expression of Alp, bone sialoprotein, osteocalcin, and ALP activity in cells grown on all evaluated surfaces. FAK inhibition did not affect the gene expression of Fak, integrin alpha 1 ( Itga1 ) and integrin beta 1 ( Itgb1 ) in cells grown on MS, increased the gene expression of Fak in cells grown on NS, and increased the gene expression of Itga1 and Itgb1 in cells grown on US and NS. Moreover, FAK protein expression decreased in cells cultured on US but increased in cells cultured on MS and NS after FAK inhibition; no difference in the expression of vinculin was observed among cells grown on all surfaces.
Our data demonstrate the relevance of FAK in the interactions between osteoblastic cells and Ti surfaces regardless of surface topography. Nanotopography positively regulated FAK expression and integrin signaling pathway components during osteoblast differentiation. In this context, the development of Ti surfaces with the ability to upregulate FAK activity could positively impact the process of implant osseointegration.
本研究旨在探讨粘着斑激酶（FAK）在成骨细胞与具有三种不同形貌的钛（Ti）表面相互作用中的作用，这三种形貌分别为未经处理（US）、微结构化（MS）和纳米结构化（NS）。
从 3 日龄大鼠的颅盖骨中提取成骨细胞，在存在 PF-573228（FAK 抑制剂）的情况下在 US、MS 和 NS 圆盘上培养，以评估成骨细胞分化。在 24 小时后，我们评估了成骨细胞形态和粘着斑蛋白表达，在第 10 天评估了以下参数：成骨细胞标记物和整合素信号成分的基因表达、FAK 蛋白表达和碱性磷酸酶（ALP）活性。US、MS 和 NS 分别观察到光滑表面、微尺度水平的孔隙和纳米腔。
FAK 抑制减少了在 US 和 MS 上生长的细胞中的丝状伪足数量，与 NS 相比。FAK 抑制降低了在所有评估表面上生长的细胞中 Alp、骨涎蛋白、骨钙素和 ALP 活性的基因表达。FAK 抑制不影响在 MS 上生长的细胞中 Fak、整合素α 1（Itga1）和整合素β 1（Itgb1）的基因表达，增加了在 NS 上生长的细胞中 Fak 的基因表达，并增加了在 US 和 NS 上生长的细胞中 Itga1 和 Itgb1 的基因表达。此外，FAK 蛋白表达在 FAK 抑制后在 US 培养的细胞中减少，但在 MS 和 NS 培养的细胞中增加；在所有表面生长的细胞中均未观察到粘着斑蛋白表达的差异。
我们的数据表明，FAK 在成骨细胞与 Ti 表面的相互作用中很重要，与表面形貌无关。纳米形貌在成骨细胞分化过程中正向调节 FAK 表达和整合素信号通路成分。在这种情况下，开发具有上调 FAK 活性能力的 Ti 表面可能会对植入物骨整合过程产生积极影响。
