|Corneal endothelial autocrine VIP enhances its integrity in stored human donor corneoscleral explant. |
Koh, SW; Gloria, D; Molloy, J
Investigative ophthalmology & visual science
To demonstrate corneal endothelial (CE) integrity enhanced during eye banking by a brief treatment of human donor corneoscleral explant (explant) with CE autocrine trophic factor vasoactive intestinal peptide (VIP).Paired explants were used as control versus VIP (10 nM)-treated before storage in corneal storage medium (4°C). CE ciliary neurotrophic factor receptor (CNTFRα) and CNTF (0.83 nM) responsiveness in connexin 43 upregulation were monitored (Western blot analysis). CE damage in CNTF-modulated explants and corneal buttons from explants was quantified by analysis of panoramic and microscopic images of the alizarin red-stained corneal endothelium. CE cells scraped from the Descemet's membrane were counted. CE VIP receptor was demonstrated (Western blot analysis).CE cells in every VIP-treated, freshly dissected explant demonstrated higher CNTFRα levels than controls (100% vs. 142% ± 15%; P = 0.014; 7 pairs stored for 4 to 25 days). Nine days after VIP treatment of previously preserved explants, CNTF responsiveness was 174% ± 23% (P = 0.023; 4 pairs) of controls. Panoramic images of explants and corneal buttons revealed that VIP treatment reduced CE damage to 75% ± 6% (P = 0.023; 4 pairs) and 71% ± 11% (P = 0.016; 9 pairs) of controls, respectively, whereas CE damage to 39% (2 pairs) and 23% ± 4% (P less than 0.001; 7 pairs), respectively, was revealed in microscopic images. Twenty-one days after VIP treatment of previously preserved explants, CE cell retention was 206% ± 38% (P = 0.008; 14 pairs) of the control. CE cells from human donor corneas expressed VIP receptor VPAC1 (not VPAC2).CE integrity during eye banking was enhanced by a brief treatment of the explant with the CE autocrine VIP.
|VIP down-regulates the inflammatory potential and promotes survival of dying (neural crest-derived) corneal endothelial cells ex vivo: necrosis to apoptosis switch and up-regulation of Bcl-2 and N-cadherin. |
Shay-Whey M Koh, Jason Cheng, Rebecca M Dodson, Chao-Yar T Ku, Cara J Abbondandolo, Shay-Whey M Koh, Jason Cheng, Rebecca M Dodson, Chao-Yar T Ku, Cara J Abbondandolo, Shay-Whey M Koh, Jason Cheng, Rebecca M Dodson, Chao-Yar T Ku, Cara J Abbondandolo, Shay-Whey M Koh, Jason Cheng, Rebecca M Dodson, Chao-Yar T Ku, Cara J Abbondandolo
Journal of neurochemistry
The neuropeptide vasoactive intestinal peptide (VIP) is anti-inflammatory and protective in the immune and nervous systems, respectively. This study demonstrated in corneal endothelial (CE) cells injured by severe oxidative stress (1.4 mM H(2)O(2)) in bovine corneal organ cultures that VIP pre-treatment (0, 10(-10), 10(-8), and 10(-6) M; 15 min), in a VIP concentration-dependent manner, switched the inflammation-causing necrosis to inflammation-neutral apoptosis (showing annexin V-binding, chromatin condensation, and DNA fragmentation) and upheld ATP levels in a VIP antagonist (SN)VIPhyb-sensitive manner, while up-regulated mRNA levels of the anti-apoptotic Bcl-2 and the differentiation marker N-cadherin in a kinase A inhibitor-sensitive manner. As a result, VIP, in a concentration-dependent and VIP antagonist-sensitive manners, promoted long-term CE cell survival. ATP levels, a determining factor in the choice of apoptosis versus necrosis, measured after VIP pre-treatment and 0.5 min post-H(2)O(2) were 39.6 +/- 3.3, 50.8 +/- 6.2, 60.1 +/- 4.8, and 53.6 +/- 5.3 pmoles/microg protein (mean +/- SEM), respectively (p 0.05, anova). VIP treatment alone concentration-dependently increased levels of N-cadherin (Koh et al. 2008), the phosphorylated cAMP-responsive-element binding protein and Bcl-2, while 10(-8) M VIP, in a VIP antagonist (SN)VIPhyb-sensitive manner, increased ATP level by 38% (p 0.02) and decreased glycogen level by 32% (p 0.02). VPAC1 (not VPAC2) receptor was expressed in CE cells. Thus, CE cell VIP/VPAC1 signaling is both anti-inflammatory and protective in the corneal endothelium.Full Text Article