In this study, we offer new insights into the contrasting effects of electrospun fiber orientation on microglial polarization under normoxia and hypoxia, and establish for the first time, the intrinsically protective roles of electrospun meshes against hypoxia-induced microglial responses. First, resting microglia were cultured under normoxia on poly(caprolactone) fibers possessing two distinctly different fiber orientations. Matrix-guided differences in cell shape/orientation and differentially expressed Rho GTPases (RhoA, Rac1, Cdc42) were well-correlated with the randomly oriented fibers inducing a pro-inflammatory phenotype and the aligned fibers sustaining a resting phenotype. Upon subsequent hypoxia induction, both sets of meshes offered protection from hypoxia-induced damage by promoting a radical phenotypic switch and beneficially altering the M2/M1 ratio to different extents. Compared to 2D hypoxic controls, meshes significantly suppressed the expression of pro-inflammatory markers (IL-6, TNF-α) and induced drastically higher expression of anti-inflammatory (IL-4, IL-10, VEGF-189) and neuroprotective (Nrf-2) markers. Consistent with this M2 polarization, the expression of Rho GTPases was significantly lower in the mesh groups under hypoxia compared to normoxic culture. Moreover, meshes—particularly with aligned fibers—promoted higher cell viability, suppressed caspase 3/8 and LC-3 expression and promoted LAMP-1 and LAMP-2 expression, which suggested the mitigation of apoptotic/autophagic cell death via a lysosomal membrane-stabilization mechanism. Notably, all protective effects under hypoxia were observed in the absence of additional soluble cues. Our results offer promise for leveraging the intrinsic therapeutic potential of electrospun meshes in degenerative diseases where microglial dysfunction, hypoxia and inflammation are implicated.

在这项研究中，我们对静电纺丝纤维取向对常氧和缺氧条件下小胶质细胞极化的对比影响提供了新的见解，并首次确定了静电纺丝网对缺氧诱导的小胶质细胞反应的内在保护作用。首先，在具有两种明显不同纤维取向的聚（己内酯）纤维上，在常氧条件下培养静止的小胶质细胞。基质引导的细胞形状/方向差异和差异表达的 Rho GTP 酶（RhoA、Rac1、Cdc42）与诱导促炎表型的随机定向纤维和维持静息表型的对齐纤维密切相关。在随后的缺氧诱导后，两组网格通过促进激进的表型转换和不同程度地有益地改变 M2/M1 比率来提供对缺氧引起的损伤的保护。与 2D 缺氧对照相比，网格显着抑制促炎标志物（IL-6、TNF-α) 并诱导抗炎 (IL-4、IL-10、VEGF-189) 和神经保护 (Nrf-2) 标志物的显着升高。与这种 M2 极化一致，与常氧培养相比，缺氧下网状组中 Rho GTPase 的表达显着降低。此外，网格——尤其是排列整齐的纤维——促进了更高的细胞活力，抑制了 caspase 3/8 和 LC-3 的表达，并促进了 LAMP-1 和 LAMP-2 的表达，这表明通过溶酶体膜减轻了凋亡/自噬性细胞死亡。稳定机制。值得注意的是，在没有其他可溶性线索的情况下，观察到了缺氧条件下的所有保护作用。我们的研究结果为利用静电纺丝网在退行性疾病中的内在治疗潜力提供了希望，其中小胶质细胞功能障碍，