Department of Ophthalmology and Visual Sciences, University of Illinois at Chicago, College of Medicine, Chicago, Illinois 60612, USA.
Invest Ophthalmol Vis Sci. 2011 Oct 11;52(11):8025-32. doi: 10.1167/iovs.11-8332.
To determine the effect of lamellar transection surgery on the nerve fiber density (NFD) and pattern of nerve regeneration in the cornea of thy1-YFP transgenic mice.
Wide-field stereo fluorescence microscopy was used to obtain serial images of nerves in live thy1-YFP mice, which express a fluorescent protein in their axons. NFD (mm/mm(2)) was calculated from maximum intensity projection images as the total length of fibers within the area of the contour in which nerves were traced. Whole-mount confocal microscopy was performed to analyze the arrangement of nerves and the types of regenerating fibers.
NFD in normal corneas was 35.3 ± 1.8 mm/mm(2). Stereo fluorescence microscopy revealed the presence of a subbasal hairpin nerve layer and an intrastromal nerve trunk layer. After surgery, regenerative sprouting was observed from transected distal ends of intrastromal nerve trunks. NFD also increased, with this increase being maximal between 4 and 6 weeks after surgery. NFD approximated baseline values at 6 weeks and did not change any further at 8 weeks. Regenerated nerves did not readopt the normal corneal nerve arrangement. A dense interlacing network of regenerated nerves was present in the corneal bed. Branches from this network traversed the flap to innervate the epithelium. Immunofluorescence staining revealed that regenerating fronds contained peptidergic nociceptive fibers (positive for calcitonin gene-related peptide and substance P) and myelinated non-nociceptive fibers (positive for neurofilament 200).
Although corneal NFD recovers to normal levels by 8 weeks after nerve transection, the arrangement of regenerated nerves is abnormal.
确定板层横断术对 Thy1-YFP 转基因小鼠角膜神经纤维密度（NFD）和神经再生模式的影响。
使用宽场立体荧光显微镜获取表达轴突中荧光蛋白的 Thy1-YFP 活鼠的神经连续图像。从最大强度投影图像中计算 NFD（mm/mm²），作为在追踪神经的轮廓区域内纤维的总长度。进行全角膜共聚焦显微镜检查以分析神经排列和再生纤维类型。
正常角膜的 NFD 为 35.3±1.8 mm/mm²。立体荧光显微镜显示存在亚基底发夹状神经层和基质内神经干层。手术后，从基质内神经干的横断远端观察到再生的发芽。NFD 也增加，术后 4 至 6 周增加最大。NFD 在 6 周时接近基线值，在 8 周时没有进一步变化。再生神经没有重新采用正常的角膜神经排列。在角膜床中存在密集交织的再生神经网络。该网络的分支穿过皮瓣以支配上皮。免疫荧光染色显示再生的羽枝含有肽能伤害性纤维（降钙素基因相关肽和 P 物质阳性）和有髓非伤害性纤维（神经丝 200 阳性）。
尽管在神经横断后 8 周，角膜 NFD 恢复到正常水平，但再生神经的排列是异常的。
