Zhang Huan, Li Zhuoya, Lin Min
（ 710049） The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China.
（ 710049） Bioinspired Engineering and Biomechanics Center (BEBC), School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China.
Sichuan Da Xue Xue Bao Yi Xue Ban. 2024 Mar 20;55(2):321-329. doi: 10.12182/20240360104.
To investigate the synergistic regulation of the polarization of mesenchymal stem cells by integrin and N-cadherin-mediated mechanical adhesion and the underlying mechanobiological mechanisms.
Bilayer polyethylene glyeol (PEG) hydrogels were formulated and modified with RGD and HAVDI peptides, respectively, to achieve mechanical adhesion to integrin and N-cadherin and to replicate the integrin-mediated mechanical interaction between cells and the extracellular matrix and the N-cadherin-mediated cell-cell mechanical interaction. The polar proteins, phosphatidylinositol 3-kinase (PI3K) and phosphorylated myosin light chain (pMLC), were characterized through immunofluorescence staining in individual cells with or without contact with HAVDI peptides under integrin-mediated adhesion, N-cadherin-mediated adhesion, and different intracellular forces. Their expression levels and polar distribution were analyzed using Image J.
Integrin-mediated adhesion induced significantly higher polar strengths of PI3K and pMLC in the contact group than in those in the no contact group, resulting in the concentration of the polar angle of PI3K to β-catenin in the range of 135° to 180° and the concentration of the polar angle of pMLC to β-catenin in the range of 0° to 45° in the contact group. Inhibition of integrin function led to inhibition of the polarity distribution of PI3K in the contact group, but did not change the polarity distribution of pMLC protein. The effect of N-cadherin on the polarity distributions of PI3K and pMLC was similar to that of integrin. However, inhibition of the mechanical adhesion of N-cadherin led to inhibition of the polarity intensity and polarity angle distribution of PI3K and pMLC proteins in the contact group. Furthermore, inhibition of the mechanical adhesion of N-cadherin caused weakened polarity intensity of integrin β1, reducing the proportion of cells with polarity angles between integrin β1 and β-catenin concentrating in the range of 135° to 180°. Additionally, intracellular forces influenced the polar distribution of PI3K and pMLC proteins. Reducing intracellular forces weakened the polarity intensity of PI3K and pMLC proteins and their polarity distribution, while increasing intracellular forces enhanced the polarity intensity of PI3K and pMLC proteins and their polarity distribution.
Integrin and N-cadherin co-regulate the polarity distribution of cell proteins and N-cadherin can play an important role in the polarity regulation of stem cells through local inhibition of integrin.
研究整合素和N-钙黏蛋白介导的机械黏附对间充质干细胞极化的协同调节作用及其潜在的机械生物学机制。
分别用RGD和HAVDI肽对双层聚乙二醇（PEG）水凝胶进行配制和修饰，以实现对整合素和N-钙黏蛋白的机械黏附，并复制整合素介导的细胞与细胞外基质之间的机械相互作用以及N-钙黏蛋白介导的细胞间机械相互作用。通过免疫荧光染色对在整合素介导的黏附、N-钙黏蛋白介导的黏附以及不同细胞内力作用下，与HAVDI肽接触或不接触的单个细胞中的极性蛋白磷脂酰肌醇3激酶（PI3K）和磷酸化肌球蛋白轻链（pMLC）进行表征。使用Image J分析它们的表达水平和极性分布。
整合素介导的黏附使接触组中PI3K和pMLC的极性强度显著高于非接触组，导致接触组中PI3K与β-连环蛋白的极角集中在135°至180°范围内，pMLC与β-连环蛋白的极角集中在0°至45°范围内。抑制整合素功能导致接触组中PI3K的极性分布受到抑制，但未改变pMLC蛋白的极性分布。N-钙黏蛋白对PI3K和pMLC极性分布的影响与整合素相似。然而，抑制N-钙黏蛋白的机械黏附导致接触组中PI3K和pMLC蛋白的极性强度和极性角分布受到抑制。此外，抑制N-钙黏蛋白的机械黏附导致整合素β1的极性强度减弱，降低了整合素β1与β-连环蛋白之间极角在135°至180°范围内集中的细胞比例。另外，细胞内力影响PI3K和pMLC蛋白的极性分布。降低细胞内力会减弱PI3K和pMLC蛋白的极性强度及其极性分布，而增加细胞内力则会增强PI3K和pMLC蛋白的极性强度及其极性分布。
整合素和N-钙黏蛋白共同调节细胞蛋白的极性分布，且N-钙黏蛋白可通过局部抑制整合素在干细胞的极性调节中发挥重要作用。
