Ultrastructure and fluid flow physiology of fetal trabecular meshwork cells.
OBJECTIVE
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.
Department of Ophthalmology, School of Medicine, University of California, San Francisco, California, USA.
On-Tat
OT
Lee
Joshua
J
Wong
Dorian
D
Liepmann
Tess
T
Lang
Shan
S
Lin
D000368:Aged; D000369:Aged, 80 and over; D001834:Body Water / Q000378:metabolism*; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D003907:Dexamethasone / Q000494:pharmacology; D005333:Fetus / Q000166:cytology*; D019084:Fluorescent Antibody Technique, Indirect; D005865:Gestational Age; D005938:Glucocorticoids / Q000494:pharmacology; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D062793:Occludin; D010750:Phosphoproteins / Q000378:metabolism; D014129:Trabecular Meshwork / Q000196:embryology / Q000378:metabolism* / Q000648:ultrastructure*; D062826:Zonula Occludens-1 Protein
D005938:Glucocorticoids; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; D010750:Phosphoproteins; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein; D003907:Dexamethasone