Beverly A Karpinski, Sonali Pal-Ghosh, Himani Datta-Majumdar, Shelly Dimri, Soneha Datta, Mary Ann Stepp
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引用次数: 0
Abstract
Purpose: The intraepithelial corneal nerves are essential to corneal health. Rho kinase or ROCK inhibitors (RIs) have been reported to play a role in neuron survival after injury. Here we assess integrin and extracellular matrix expression in primary mouse neurons and determine whether treating cells with RI impacts neurite outgrowth in vitro and reinnervation after trephine and debridement injury in mice in vivo.
Methods: Cocultures of human corneal limbal epithelial cells and E11.5 mouse trigeminal neurons and neurons alone were grown on glass coverslips. High-resolution imaging was performed to localize integrins and laminin on neurons and to determine whether RI impacts neurite outgrowth in vitro and in vivo after both 1.5-mm trephine and 1.5-mm debridement injuries.
Results: Several integrin α (α3, α6, αv) chains as well as β4 integrin are expressed on neuron axons and growth cones in cocultures. RI treatment of isolated neurons, cocultures, and in conditioned media increases neurite outgrowth. In vivo, RI positively impacts sensory nerve reinnervation after trephine and debridement injury.
Conclusions: These studies are the first to demonstrate expression of β4 integrin on trigeminal sensory neurons and preferential adhesion of neurons to the laminin-enriched matrices found in footprints deposited by human corneal limbal epithelial cells. In addition, we also document for the first time the positive impact of RI on neurite outgrowth in vitro and reinnervation in vivo.
目的:角膜上皮内神经对角膜健康至关重要。据报道,Rho 激酶或 ROCK 抑制剂(RIs)在神经元损伤后的存活中发挥作用。在此,我们评估了原代小鼠神经元中整合素和细胞外基质的表达情况,并确定用 RI 处理细胞是否会影响体外神经元的生长以及小鼠体内穿刺和剥离损伤后的神经支配:方法: 在玻璃盖玻片上培养人角膜缘上皮细胞和 E11.5 小鼠三叉神经元的共培养物,以及单独培养神经元。通过高分辨率成像定位神经元上的整合素和层粘连蛋白,并确定 RI 是否会影响体外和体内 1.5 毫米穿刺损伤和 1.5 毫米清创损伤后的神经元生长:结果:多种整合素α(α3、α6、αv)链以及β4整合素在神经元轴突和共培养生长锥上表达。对离体神经元、共培养物和条件培养基进行 RI 处理可增加神经元的生长。在体内,RI 对穿刺和清创损伤后的感觉神经再支配有积极影响:这些研究首次证明了β4整合素在三叉神经感觉神经元上的表达,以及神经元优先粘附在人类角膜缘上皮细胞沉积的足印中发现的富含层粘连蛋白的基质上。此外,我们还首次记录了 RI 对体外神经元生长和体内神经支配的积极影响。
期刊介绍:
Investigative Ophthalmology & Visual Science (IOVS), published as ready online, is a peer-reviewed academic journal of the Association for Research in Vision and Ophthalmology (ARVO). IOVS features original research, mostly pertaining to clinical and laboratory ophthalmology and vision research in general.