Jester J V, Rodrigues M M, Herman I M
Am J Pathol. 1987 Apr;127(1):140-8.
The characteristics and derivation of corneal wound healing fibroblasts (myofibroblasts) were evaluated by studying the temporal changes in the cellular actin distribution of corneal fibrocytes following full thickness 3-mm diameter central corneal wounds in the rabbit. Under certain conditions these wounds heal without neovascularization, allowing for the detailed analysis of invading fibroblasts with minimal contamination by other cell types. The authors employed transmission electron microscopy to localize microfilaments, fluorescent microscopy using NBD-phallacidin, a mushroom toxin which binds specifically to f-actin and oligomeres of g-actin, to localize actin filaments, and isoelectric focusing gels to characterize actin isotypes. During the early stages of wound healing (1-7 days) there is a gradual change in the corneal fibrocytes adjacent to the wound margin characterized by the development of extensive rough endoplasmic reticulum, microtubules, a prominent Golgi apparatus, and a cortical microfilament network. This is in contrast to the normal fibrocyte, which, for the most part, lacks these structures. The development of microfilaments correlated with increased NBD-phallacidin fluorescence of fibrocytes adjacent to the wound as compared with fibrocytes farther removed from the site of injury. Fibroblasts appearing within the wound from 7 days to 2 months after injury had ultrastructural characteristics similar to those of myofibroblasts, including parallel arrays of microfilaments, stress fibers and cell-cell, cell-matrix attachments. Furthermore, these cells stained intensely with NBD-phallacidin, supporting the ultrastructural findings. At 1 month after injury, cells contained within the wound possessed predominantly nonmuscle isoactins (gamma) as seen by silver staining of isoelectric focusing gels, but little or no (smooth muscle) isoactins could be detected. Moreover, no significant differences could be detected between electrophoretic profiles obtained from wounded versus normal corneas. These morphologic and biochemical data suggest that the corneal fibrocyte may develop into a fibroblastlike cell similar to the myofibroblast, and is characterized by a marked increase in filamentous actin.
通过研究兔角膜中央3毫米直径全层伤口后角膜纤维细胞的细胞肌动蛋白分布的时间变化，评估角膜伤口愈合成纤维细胞（肌成纤维细胞）的特征和来源。在某些情况下，这些伤口愈合时无新生血管形成，从而能够在极少受到其他细胞类型污染的情况下对侵入的成纤维细胞进行详细分析。作者采用透射电子显微镜定位微丝，使用NBD-鬼笔环肽（一种特异性结合f-肌动蛋白和g-肌动蛋白寡聚体的蘑菇毒素）的荧光显微镜定位肌动蛋白丝，以及等电聚焦凝胶来鉴定肌动蛋白同工型。在伤口愈合的早期阶段（1 - 7天），伤口边缘附近的角膜纤维细胞逐渐发生变化，其特征为广泛的粗面内质网、微管、显著的高尔基体和皮质微丝网络的形成。这与正常纤维细胞形成对比，正常纤维细胞在很大程度上缺乏这些结构。微丝的形成与伤口附近纤维细胞的NBD-鬼笔环肽荧光增强相关，与远离损伤部位的纤维细胞相比荧光更强。损伤后7天至2个月内在伤口处出现的成纤维细胞具有与肌成纤维细胞相似的超微结构特征，包括微丝的平行排列、应力纤维以及细胞 - 细胞、细胞 - 基质连接。此外，这些细胞用NBD-鬼笔环肽染色强烈，支持了超微结构的发现。损伤后1个月，伤口内的细胞在等电聚焦凝胶银染中主要含有非肌肉同工肌动蛋白（γ），但几乎检测不到（平滑肌）同工肌动蛋白。此外，从受伤角膜与正常角膜获得的电泳图谱之间未检测到显著差异。这些形态学和生化数据表明，角膜纤维细胞可能发育成类似于肌成纤维细胞的成纤维样细胞，其特征是丝状肌动蛋白显著增加。
