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Comparative Study
. 2011 May;151(5):752-759.e2.
doi: 10.1016/j.ajo.2010.11.008. Epub 2011 Feb 18.

Transforming growth factor-β signaling pathway activation in Keratoconus

Christoph Engler  1 , Shukti Chakravarti, Jefferson Doyle, Charles G Eberhart, Huan Meng, Walter J Stark, Clare Kelliher, Albert S Jun
Affiliations
Comparative Study

Transforming growth factor-β signaling pathway activation in Keratoconus

Christoph Engler et al. Am J Ophthalmol. 2011 May.

Abstract

Purpose: To assess the presence of transforming growth factor-β (TGFβ) pathway markers in the epithelium of keratoconus patient corneas.

Design: Retrospective, comparative case series of laboratory specimens.

Methods: Immunohistochemistry results for TGFβ2, total TGFβ, mothers against decacentaplegic homolog (Smad) 2, and phosphorylated Smad2 was performed on formalin-fixed, paraffin-embedded sections of keratoconus patient corneas and normal corneas from human autopsy eyes. Keratoconus patient corneas were divided in two groups, depending on their severity based on keratometer readings and pachymetry. Autopsy controls were age-matched with the keratoconus cases. Immunohistochemistry signal quantification was performed using automated software. Real-time reverse-transcriptase polymerase chain reaction was performed on total ribonucleic acid of epithelium of keratoconus patient corneas and autopsy control corneas.

Results: Immunohistochemistry quantification showed a significant increase in mean signal in the group of severe keratoconus cases compared with normal corneas for TGFβ2 and phosphorylated Smad2 (P < .05). Immunohistochemistry analysis using antibodies against total TGFβ and Smad2 did not show any significant increase in the keratoconus cases versus the autopsy controls. Reverse-transcriptase polymerase chain reaction exhibited elevated messenger ribonucleic acid levels of Smad2 and TGFβ2 in severe keratoconus corneal epithelium.

Conclusions: This work shows increased TGFβ pathway markers in severe keratoconus cases and provides the rationale for investigating TGFβ signaling further in the pathophysiology of keratoconus.

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Figures

FIGURE 1
FIGURE 1
Immunohistochemistry for transforming growth factor-β (TGFβ) signaling pathway markers. TGFβ2, pSmad2, and TGFβ (total) were assessed in normal human corneas and corneas with keratoconus. Horseradish peroxidase (HRP)-positive signal was quantified as described in Methods. Error bars indicate standard deviation. *P < .05; **P < .01; ***P < .001; +P ≥ .05; Mann–Whitney U test. Original magnification ×200. (A and B): Immunohistochemistry for TGFβ signaling pathway markers (TGFβ2 and phosphorylated Smad2 [pSmad2]) in (Top left) normal human corneas and (Top right) in patients with mild and (Bottom left) severe keratoconus. (Bottom right) Signal quantification. (C): Immunohistochemistry results for TGFβ (total) in (Left) normal human corneas and (Middle) patients with severe keratoconus. (Right) Signal quantification.
FIGURE 2
FIGURE 2
Photomicrograph showing immunohistochemistry results for phosphorylated Smad2 (pSmad2). Enlarged detail of keratoconus epithelium (×400 magnification) stained for pSmad2, showing a (*) perinuclear and (arrow) nuclear staining pattern.
See this image and copyright information in PMC

References

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