Department of Ophthalmology, University of California, Davis, CA 95616, USA.
Department of Cell Biology and Human Anatomy, University of California, Davis, CA 95817, USA.
Int J Mol Sci. 2023 Sep 20;24(18):14327. doi: 10.3390/ijms241814327.
Corneal wound healing is a complex biological process that integrates a host of different signals to coordinate cell behavior. Upon wounding, there is the generation of an endogenous wound electric field that serves as a powerful cue to guide cell migration. Concurrently, the corneal epithelium reduces sialylated glycoforms, suggesting that sialylation plays an important role during electrotaxis. Here, we show that pretreating human telomerase-immortalized corneal epithelial (hTCEpi) cells with a sialyltransferase inhibitor, P-3FAX-Neu5Ac (3F-Neu5Ac), improves electrotaxis by enhancing directionality, but not speed. This was recapitulated using Kifunensine, which inhibits cleavage of mannoses and therefore precludes sialylation on N-glycans. We also identified that 3F-Neu5Ac enhanced the responsiveness of the hTCEpi cell population to the electric field and that pretreated hTCEpi cells showed increased directionality even at low voltages. Furthermore, when we increased sialylation using N-azidoacetylmannosamine-tetraacylated (Ac4ManNAz), hTCEpi cells showed a decrease in both speed and directionality. Importantly, pretreating enucleated eyes with 3F-Neu5Ac significantly improved re-epithelialization in an ex vivo model of a corneal injury. Finally, we show that in hTCEpi cells, sialylation is increased by growth factor deprivation and reduced by PDGF-BB. Taken together, our results suggest that during corneal wound healing, reduced sialylated glycoforms enhance electrotaxis and re-epithelialization, potentially opening new avenues to promote corneal wound healing.
角膜伤口愈合是一个复杂的生物学过程，它整合了多种不同的信号来协调细胞行为。在受伤时，会产生一个内源性的伤口电场，作为引导细胞迁移的有力线索。同时，角膜上皮减少了唾液酸化的糖型，表明唾液酸化在电趋性中起着重要作用。在这里，我们表明，用唾液酸转移酶抑制剂 P-3FAX-Neu5Ac（3F-Neu5Ac）预处理人端粒酶永生化角膜上皮（hTCEpi）细胞，可以通过增强方向性而不是速度来改善电趋性。这一点可以通过 Kifunensine 得到重现，它可以抑制甘露糖的切割，从而阻止 N-糖链上的唾液酸化。我们还发现，3F-Neu5Ac 增强了 hTCEpi 细胞群体对电场的反应性，并且预处理的 hTCEpi 细胞即使在低电压下也显示出更高的方向性。此外，当我们使用 N-叠氮乙酰甘露糖四酰化（Ac4ManNAz）增加唾液酸化时，hTCEpi 细胞的速度和方向性都降低了。重要的是，用 3F-Neu5Ac 预处理去眼可以显著改善角膜损伤的体外模型中的再上皮化。最后，我们表明，在 hTCEpi 细胞中，生长因子剥夺会增加唾液酸化，而 PDGF-BB 会减少唾液酸化。综上所述，我们的研究结果表明，在角膜伤口愈合过程中，减少的唾液酸化糖型增强了电趋性和再上皮化，这可能为促进角膜伤口愈合开辟了新途径。
