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S.–W.M. Koh, X.–H. Liu, A.J. Symes, J.A. Waschek; Ciliary Neurotrophic Factor (CNTF)Induction of VasoactiveIntestinal Peptide (VIP)Expression In the Corneal Endothelium (CE) In Preserved Donor Human Corneas . Invest. Ophthalmol. Vis. Sci. 2004;45(13):3764.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To demonstrate that CNTF, a cytokine released by CE cells surviving oxidative stress, induces in CE cells the expression of VIP, a factor that has previously been shown to protect CE cells against lethal oxidative stress. Methods: Human donor corneas that had been stored in Optisol–GS (cornea storage medium used in eye banking) at 4oC and determined not suitable for transplantation due to less than optimal CE cell numbers were obtained from the Central Florida Eye Bank and stored at 4oC until experimentation. The presence of CNTF receptor (CNTFRα) in CE was demonstrated by Western blot analysis. For CNTF induction of VIP, corneas were preconditioned in DMEM (24 h, 37oC) before CNTF (1 nM) treatment (24 h, 37oC) in DMEM plus 5% fetal calf serum. CE total RNA was isolated from 10 paired corneas (control vs CNTF–treated) and subjected to Northern analysis using a human VIP cDNA. In other experiments, two halves bisected from the same cornea were used as control vs CNTF–treated in reverse transcription/real time PCR quantification (real–time RT–qPCR) of CE cell VIP mRNA. Results: CE cells expressed a 50 kDa CNTFRα. In Northern blots, a single hybridizing band appeared corresponding to the predicited 1.8 kb VIP mRNA, and was induced in the CE of CNTF–treated corneas. Real–time RT–qPCR showed that VIP mRNA level in the CE of CNTF–treated cornea halves was 2.25+0.38 (mean+sem) times that of the control cornea halves in 24 corneas. In corneas that had been stored for four weeks or longer, CNTFRα was undetectable, and CNTF treatment did not increase VIP mRNA levels (i. e. 0.91+0.09 [mean+sem]–fold induction in 8 corneas). Conclusions: CE cells in human donor corneas in storage for up to four weeks express CNTFRα and VIP mRNA, the latter of which can be induced by exogenous CNTF. Although CE in stored cornea appeared to have lost its CNTF (unpublished data), CNTF is enriched in the ciliary body and iris. While CNTF is an injury factor released only after injury, CNTF upregulation of VIP in CE may take place in transplanted cornea in the recipient eye.
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