胎儿小梁网细胞的超微结构与流体流动生理学

Lee On-Tat, Wong Joshua, Liepmann Dorian, Lang Tess, Lin Shan

Department of Ophthalmology, School of Medicine, University of California, San Francisco, California, USA.

Curr Eye Res. 2008 Oct;33(10):849-56. doi: 10.1080/02713680802429210.

PURPOSE

To assess the ultrastructural and fluid flow characteristics of cultured trabecular meshwork (TM) cells derived from fetal sources.

METHODS

Fetal eyes were carefully dissected to isolate the developing TM tissue for culture. Immunostaining was used to assess the expression of the junction-associated proteins zonula occludens-1 (ZO-1) and occludin. Fetal and adult TM cells were grown to confluence on permeable membranes for both flow and ultrastructural studies. Fluid flow resistance was measured by permeation of horseradish peroxidase (hrp) activity and hydraulic conductivity (HC) experiments. The effects of dexamethasone (Dex) on permeability and HC were also evaluated.

RESULTS

ZO-1 and occludin are expressed in the TM region of tissue sections and at cell borders in cultured fetal and adult TM cells. Transmission electron microscopy demonstrated that cultured TM cells possessed numerous mitochondria, electron-dense bodies, surface microvilli, and adherens and gap junctions. The permeation of hrp across fetal TM cell monolayers (0.030 +/- 0.010) and of adult TM cells (0.031 +/- 0.010) had similar values for absorbance at 470 nm (p = 0.83, 95% CI: -0.004, 0.005). Dex treatment significantly reduced the permeability to 0.022 +/- 0.008 (p = 0.002) and 0.018 +/- 0.009 (p = 0.004) for fetal and adult TM cells, respectively. The average HC (microl/min/mmHg/cm(2)) of fetal cells (2.78 +/- 1.03) and of the adult cells (2.15 +/- 1.31) was not significantly different (p = 0.24, 95% CI: -1.01, 0.26). Dex treatment significantly reduced HC in both fetal (1.24 +/- 0.72, p = 0.0004) and adult (1.29 +/- 0.29, p = 0.00001) TM.

CONCLUSIONS

Cultured fetal TM cells exhibited similar expression of junctional proteins and ultrastructural features as their adult counterparts. The permeability and HC of the fetal cells were similar to their older adult counterparts. Dex treatment induced increased fluid flow resistance in both cell types. These cells may serve as a source for in vitro studies of meshwork physiology.

目的

评估源自胎儿的培养小梁网（TM）细胞的超微结构和流体流动特性。

方法

仔细解剖胎儿眼睛以分离发育中的TM组织用于培养。免疫染色用于评估紧密连接相关蛋白闭合蛋白-1（ZO-1）和闭合蛋白的表达。将胎儿和成人TM细胞在可渗透膜上培养至汇合，用于流动和超微结构研究。通过辣根过氧化物酶（hrp）活性渗透和水力传导率（HC）实验测量流体流动阻力。还评估了地塞米松（Dex）对通透性和HC的影响。

结果

ZO-1和闭合蛋白在组织切片的TM区域以及培养的胎儿和成人TM细胞的细胞边界处表达。透射电子显微镜显示，培养的TM细胞具有大量线粒体、电子致密体、表面微绒毛以及黏着连接和缝隙连接。hrp穿过胎儿TM细胞单层（0.030±0.010）和成人TM细胞单层（0.031±0.010）在470nm处的吸光度值相似（p = 0.83，95%可信区间：-0.004，0.005）。Dex处理分别使胎儿和成人TM细胞的通透性显著降低至0.022±0.008（p = 0.002）和0.018±0.009（p = 0.004）。胎儿细胞（2.78±1.03）和成人细胞（2.15±1.31）的平均HC（微升/分钟/毫米汞柱/平方厘米）无显著差异（p = 0.24，95%可信区间：-1.01，0.26）。Dex处理显著降低了胎儿（1.24±0.72，p = 0.0004）和成人（1.29±0.29，p = 0.00001）TM的HC。