In vitro cell culture model for anti-cataract drug penetration studies.
Immortalized human corneal epithelial cells (HCECs) and human lens epithelial cells (HLECs) were cultured in vitro. Cells were observed under a phase-contrast microscope and the integrity of cell monolayers was assayed by transepithelial electrical resistance (TEER) determination. The permeability of disulfiram (DSF) through a HCECs monolayer was compared with that of DSF through an excised rabbit cornea. The permeability coefficients of DSF through a HCECs monolayer and excised rabbit cornea were 29.5 +/- 4.8 x 10(-6) cm/s and 34.7 +/- 5.2 x 10(-6) cm/s, respectively. Diethyldithiocarbamate (DDC) had high permeability through HLECs monolayer with a permeability coefficient of 44.6 +/- 7.1 x 10(-6) cm/s. The cytotoxicity of DDC against HLECs was investigated using the trypan blue exclusion test. For a DDC concentration of 5 mmol/l, more than 85% cells were viable. DH3a1 mRNA was expressed in cultured HLECs. The expression of aldehyde dehydrogenase 3a1 (ALDH3a1), which may be be responsible for DSF-DDC conversion, was detected using RT-PCR and agarose gels electrophoresis. These results demonstrate that the permeability of DSF can be detected and intra-ocular drug action may be predicted using the cultured HCEC and HLEC monolayers as model.
Jiang
T Y
TY
School of Pharmacy, Shenyang Pharmaceutical University, China.
Wang
S L
SL
Liu
Y
Y
Nie
S F
SF
Ito
Y
Y
Nagai
N
N
Wu
C F
CF
D000444:Aldehyde Dehydrogenase / Q000235:genetics; D000818:Animals; D002386:Cataract / Q000188:drug therapy* / Q000378:metabolism*; D002460:Cell Line; D002463:Cell Membrane Permeability; D024141:Collagen Type IV / Q000378:metabolism; D004396:Coloring Agents; D003315:Cornea / Q000378:metabolism; D004221:Disulfiram / Q000493:pharmacokinetics; D004050:Ditiocarb / Q000493:pharmacokinetics; D004553:Electric Conductivity; D004847:Epithelial Cells / Q000378:metabolism; D019573:Epithelium, Corneal / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007908:Lens, Crystalline / Q000378:metabolism; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014343:Trypan Blue
D024141:Collagen Type IV; D004396:Coloring Agents; D004050:Ditiocarb; D000444:Aldehyde Dehydrogenase; D014343:Trypan Blue; D004221:Disulfiram