CELL CYCLE OF THE CORNEAL ENDOTHELIUM

内皮是单层细胞，位于血管的后部。 角膜，保持角膜的透明度。在人类中，角膜 内皮细胞密度随着年龄的增长而降低，这表明， 有丝分裂引起的细胞替换跟不上细胞损失的速度。 其他因素，如糖尿病、炎症、眼外伤或手术， 导致细胞损失并可导致内皮代偿失调， 基质水肿和视力丧失。大泡性角膜病变 内皮功能障碍目前在医学上是不可治疗的， 只有通过角膜移植才能实现。我们长久以来- 长期目标是发展医学治疗，以促进愈合的压力 角膜内皮为了达到这一目标，我们必须发现角膜 调节内皮修复。 通常有丝分裂和细胞运动 有助于单层修复;然而，人角膜内皮细胞 损伤后不易分裂，因此修复主要通过细胞运动进行。 这种有丝分裂能力的相对缺乏并不普遍，因为角膜 诸如兔的物种中的内皮细胞容易分裂。研究 在最后一个资助周期，重点是细胞运动的调节。 拟议中的研究将重点转移到监管上， 角膜内皮细胞周期。这种方法更直接， 更有利于实现长期目标。工作假设 尽管在人类中， 衰老的非分裂细胞随着年龄的增长而增加，在所有年龄段， 一群有丝分裂静止的G1期阻滞细胞， 能够刺激有丝分裂。在人类和兔子体内， 条件维持内皮处于静止、分化状态 以保持其重要的生理功能。因素可能 有助于静止包括接触抑制，相对较高 TGF-β（一种潜在的生长抑制剂）在水溶液中的浓度 体液和PGE 2的自分泌有丝分裂抑制。生长停滞恢复 在生长因子的存在下，如EGF，应该促进有丝分裂， 角膜内皮受压。 这些研究将使用细胞 生物学、药理学和分子生物学方法， 具体目的如下：1）比较人和兔角膜的 内皮细胞：活跃周期细胞和静止细胞的相对百分比 以及静止细胞在细胞周期中的相对位置 2）确定在胎儿发育早期， 是人类角膜内皮从活跃的循环到 与形成稳定的接触相关的静止状态 抑制和/或分化的单层，3）确定是否释放 - 接触抑制，特别是在EGF和/或 吲哚美辛可刺激角膜内皮细胞重新进入细胞 4）确定TGF-β是否抑制角膜内皮细胞 分裂以及这种抑制是否可逆。这些研究应 帮助确定人类角膜的相对有丝分裂潜能 内皮细胞，研究体内抑制角膜内皮细胞的原因 帮助发现逆转有丝分裂抑制的方法 以临床相关的方式。

The endothelium is the monolayer of cells, located at the posterior of the cornea, which maintains corneal transparency. In humans, corneal endothelial cell density decreases with age, suggesting that the rate of cell replacement by mitosis does not keep pace with the rate of cell loss. Other factors, such as diabetes, inflammation, ocular trauma or surgery, contribute to cell loss and can lead to endothelial decompensation, stromal edema, and loss of visual acuity. Bullous keratopathy caused by endothelial dysfunction is currently medically untreatable and restoration of vision can only be accomplished by corneal transplantation. Our long- term goal is to develop medical treatments to promote healing of stressed corneal endothelium. To reach this goal, we must discover how corneal endothelial repair is regulated. Normally, both mitosis and cell movement contribute to monolayer repair; however, human corneal endothelium does not readily divide upon injury, so repair occurs mainly by cell movement. This relative lack of mitotic capability is not universal, since corneal endothelial cells in species, such as rabbits, readily divide. Studies during the last grant cycle focussed on the regulation of cell movements. The proposed studies will change focus to concentrate on the regulation of the corneal endothelial cell cycle. This approach is more direct and may be more helpful in achieving the long-term goal. The working hypothesis for these studies is that, although in humans the relative number of senescent, non-dividing cells increases with age, there is, at all ages, a population of mitotically quiescent, G1-phase arrested cells which are capable of mitogenic stimulation. In both humans and rabbits, in vivo conditions maintain the endothelium in a quiescent, differentiated state to preserve its important physiologic functions. Factors which may contribute to quiescence include contact inhibition, the relatively high concentration of TGF-beta (a potential growth inhibitor) in the aqueous humor, and autocrine mitotic inhibition by PGE2. Reversal of growth arrest in the presence of growth factors, such as EGF, should promote mitosis in stressed corneal endothelium. The proposed studies will use cell biological, pharmacological and molecular biological methods to achieve the following Specific Aims: 1) Compare in human and rabbit corneal endothelium the relative percent of actively cycling and quiescent cells and the relative position within the cell cycle in which quiescent cells are arrested, 2) Determine whether, during early fetal development, there is a change in human corneal endothelium from an actively cycling to a quiescent state which correlates with the formation of a stable, contact inhibited and/or differentiated monolayer, 3) Determine whether release from contact inhibition, particularly in the presence of EGF and/or indomethacin can stimulate corneal endothelial cells to re-enter the cell cycle, and 4) Determine whether TGF-beta inhibits corneal endothelial cell division and whether such inhibition is reversible. These studies should help determine the relative mitotic potential of human corneal endothelium, investigate causes for in vivo inhibition of corneal endothelial mitosis and help discover means to reverse mitotic inhibition in a clinically relevant manner.