Mtor-inhibitor-containing medicine for treating or preventing ophthalmic symptoms, disorders, or diseases, and application thereof
a technology of mtor and inhibitor, which is applied in the field of medicine for treating or preventing eye conditions, disorders, or diseases, can solve the problems of corneal edema, reduced density, and limited regeneration ability of human corneal endothelial cells, and achieve the effects of preserving and/or growing or promoting the growth of corneal endothelial cells
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example 4
on Effect of Rapamycin on Phosphorylation Activity of Smad2 / 3 Induced by TGF-β2
[0207]This Example demonstrated suppression effect of rapamycin, which is a typical example of an mTOR inhibitor, on phosphorylation activity of Smad2 / 3 induced by TGF-β2.
[0208](Materials and Methods)
[0209]The medium was removed from a culture dish in which iFECDs were being cultured, and the cells were supplemented with 1×PBS (−) that was preheated to 37° C., and were washed. This was repeated twice. The cells were supplemented again with 1×PBS (−) and incubated for 3 minutes at 37° C. (5% CO2). After removing the PBS (−), the cells were supplemented with 0.05% Trypsin-EDTA (Nacalai Tesque, 32778-34) and incubated for 5 minutes at 37° C. (5% CO2). The cells were then suspended in a medium, and collected by centrifugation at 1500 rpm for 3 minutes. DMEM (Nacalai Tesque, 08456-36)+10% FBS (Biowest, S1820-500)+1% P / S (Nacalai Tesque, 26252-94) was used as the medium.
[0210]iFECDs were seeded on a 12-well pla...
example 5
on Effect of Rapamycin on Production of Fibronectin Induced by TGF-β2
[0217]This Example demonstrated suppression effect of rapamycin, which is a typical example of an mTOR inhibitor, on the production of fibronectin induced by TGF-β2.
[0218](Materials and Methods)
[0219]The medium was removed from a culture dish in which iFECDs were being cultured, and the cells were supplemented with 1×PBS (−) that was preheated to 37° C., and were washed. This was repeated twice. The cells were supplemented again with 1×PBS (−) and incubated for 3 minutes at 37° C. (5% CO2). After removing the PBS (−), the cells were supplemented with 0.05% Trypsin-EDTA (Nacalai Tesque, 32778-34) and incubated for 5 minutes at 37° C. (5% CO2). The cells were then suspended in a medium, and collected by centrifugation at 1500 rpm for 3 minutes. DMEM (Nacalai Tesque, 08456-36)+10% FBS (Biowest, S1820-500)+1% P / S (Nacalai Tesque, 26252-94) was used as the medium.
[0220]iFECDs were seeded on a 6-well plate at a ratio of ...
example 6
A Suppression Effect on mTOR Expression and Phosphorylation of S6K
[0228]This Example demonstrated the suppression effect of mTOR siRNA, which is another typical example of an mTOR inhibitor, on mTOR expression and phosphorylation of S6K.
[0229](Materials and Methods)
[0230]The medium was removed from a culture dish in which iFECDs were being cultured, and the cells were supplemented with 1×PBS (−) that was preheated to 37° C., and were washed. This was repeated twice. The cells were supplemented again with 1×PBS (−) and incubated for 3 minutes at 37° C. (5% CO2). After removing the PBS (−), the cells were supplemented with 0.05% Trypsin-EDTA (Nacalai Tesque, 32778-34) and incubated for 5 minutes at 37° C. (5% CO2). The cells were then suspended in a medium, and collected by centrifugation at 1500 rpm for 3 minutes. DMEM (Nacalai Tesque, 08456-36)+10% FBS (Biowest, S1820-500)+1% P / S (Nacalai Tesque, 26252-94) was used as the medium.
[0231]iFECDs were seeded on a 12-well plate at a ratio...
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