Acta Biomaterialia ( IF 9.4 ) Pub Date : 2023-04-10 , DOI: 10.1016/j.actbio.2023.04.004 Yan Zhou 1 , Timmy Fyrner 2 , Charlotte H Chen 2 , Nicholas A Sather 2 , Erin L Hsu 3 , Samuel I Stupp 4 , Malcolm L Snead 1
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Various peptide amphiphile (PA) molecules have been developed to promote bone regeneration. Previously we discovered that a peptide amphiphile with a palmitic acid tail (C16) attenuates the signaling threshold of leucine-rich amelogenin peptide (LRAP)-mediated Wnt activation by increasing membrane lipid raft mobility. In the current study, we found that treatment of murine ST2 cells with an inhibitor (Nystatin) or Caveolin-1-specific siRNA abolishes the effect of C16 PA, indicating that Caveolin-mediated endocytosis is required. To determine whether hydrophobicity of the PA tail plays a role in its signaling effect, we modified the length of the tail (C12, C16 and C22) or composition (cholesterol). While shortening the tail (C12) decreased the signaling effect, lengthening the tail (C22) had no prominent effect. On the other hand, the cholesterol PA displayed a similar function as the C16 PA at the same concentration of 0.001% w/v. Interestingly, a higher concentration of C16 PA (0.005%) is cytotoxic while cholesterol PA at the higher concentration (0.005%) is well-tolerated by cells. Use of the cholesterol PA at 0.005% enabled a further reduction of the signaling threshold of LRAP to 0.20 nM, compared to 0.25 nM at 0.001%. Caveolin-mediated endocytosis is also required for cholesterol PA, as evidenced by Caveolin-1 siRNA knockdown experiments. We further demonstrated that the noted effects of cholesterol PA are also observed in human bone marrow mesenchymal stem cells (BMMSCs). Taken together, these results indicate that the cholesterol PA modulates lipid raft/caveolar dynamics, thereby increasing receptor sensitivity for activation of canonical Wnt signaling.
Statement of significance
Cell signaling involves not only the binding of growth factors (or other cytokines) and cognate receptors, but also their clustering on the cell membrane. However, little or no work has been directed thus far toward investigating how biomaterials can serve to enhance growth factor or peptide signaling by increasing diffusion of cell surface receptors within membrane lipid rafts. Therefore, a better understanding of the cellular and molecular mechanism(s) operating at the material-cell membrane interface during cell signaling has the potential to change the paradigm in designing future biomaterials and regenerative medicine therapeutics. In this study, we designed a peptide amphiphile (PA) with a cholesterol tail to enhance canonical Wnt signaling by modulating lipid raft/caveolar dynamics.
中文翻译：
通过改变肽两亲亲脂性优化肽两亲分子-脂筏相互作用
已经开发了多种肽两亲物 (PA) 分子来促进骨再生。之前我们发现，具有棕榈酸尾部 (C16 ) 的肽两亲物通过增加膜脂筏的流动性来减弱富含亮氨酸牙釉蛋白肽 (LRAP) 介导的 Wnt 激活的信号阈值在当前的研究中，我们发现用抑制剂（制霉菌素）或 Caveolin-1 特异性 siRNA 处理鼠类 ST2 细胞会消除 C 16 PA的作用，表明需要 Caveolin 介导的内吞作用。为了确定 PA 尾部的疏水性是否对其信号效应起作用，我们修改了尾部的长度（C 12、C16和 C22) 或成分（胆固醇）。虽然缩短尾巴 (C 12 ) 降低了信号效应，但加长尾巴 (C 22 ) 没有显着影响。另一方面，胆固醇 PA 在 0.001% w/v 的相同浓度下表现出与 C 16 PA 相似的功能。有趣的是，更高浓度的 C 16PA (0.005%) 具有细胞毒性，而较高浓度 (0.005%) 的胆固醇 PA 可被细胞很好地耐受。与 0.001% 的 0.25 nM 相比，使用 0.005% 的胆固醇 PA 能够将 LRAP 的信号阈值进一步降低至 0.20 nM。正如 Caveolin-1 siRNA 敲低实验所证明的，胆固醇 PA 也需要 Caveolin 介导的内吞作用。我们进一步证明，在人骨髓间充质干细胞 (BMMSC) 中也观察到了胆固醇 PA 的显着影响。总之，这些结果表明胆固醇 PA 调节脂筏/小窝动力学，从而增加受体对激活典型 Wnt 信号传导的敏感性。
重要性声明
细胞信号传导不仅涉及生长因子（或其他细胞因子）和同源受体的结合，还涉及它们在细胞膜上的聚集。然而，到目前为止，很少或根本没有研究生物材料如何通过增加细胞表面受体在膜脂筏内的扩散来增强生长因子或肽信号传导。因此，更好地了解细胞信号传导过程中材料-细胞膜界面的细胞和分子机制，有可能改变设计未来生物材料和再生医学疗法的范式。在这项研究中，我们设计了一种带有胆固醇尾巴的肽两亲物 (PA)，以通过调节脂筏/小窝动力学来增强典型的 Wnt 信号传导。
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