Lack of TrkA (Nerve growth factor receptor) expression in keratoconus

Abstract

Abstract: : Purpose:Nerve growth factor (NGF) is a neurotrophin affecting sensory nerves, cell survival, healing and fibroblast function in several tissues. Keratoconus is characterized by changes of keratocyte function, an increase of cell apoptosis and a decrease of corneal nerve density. The aim of this study is to investigate the pathway of NGF production and utilization in patients affected by keratoconus Methods:Twenty–one corneas obtained at the time of penetrating keratoplasty (10 keratoconus, 2 corneal ectasia secondary to refractive surgery, 3 bullous keratopathy, 3 Fuchs dystrophy, 3 lattice dystrophy) and 10 normal corneas obtained by the local eye bank were evaluated. All patients affected by keratoconus showed a stage 4 of the disease. Control corneas were obtained from sex and age matched cadavers with negative history for ocular diseases. RT–PCR was performed to evaluate change in mRNA levels and ISH was used to identify which corneal cells produced NGF, TrkA and p75. To evaluate expression of NGF, TrkA and p75, immunohistochemestry, ELISA and Western Blot evaluation was performed on the same corneas Results:Normal cornea produces both NGF and NGF receptors (TrkA and p75) as demonstrated by RT–PCR. In particular, mRNAs for NGF and NGF receptors were expressed by all corneal cells (endothelium, keratocytes and epithelium). In keratoconus mRNAs levels of NGF and p75 were significantly decreased (p<0.05) when compared to normal corneas, while mRNA of TrkA was no detectable in all the evaluated corneas by RT–PCR as well as by ISH. To investigate if changes in mRNA production corresponded to variation of the respectively proteins, IIC, ELISA and Western Blot were performed on the same samples. In kearatoconus corneas NGF and p75 levels were significantly reduced (p<0.05) as demonstrated by ELISA and Western Blot, while no expression of Trka was detectable in keratoconus corneas by IIC and Western Blot. The corneas of secondary keratoconus, as well as of bullous keratopathy, Fuchs’ and lattice dystrophy, express all the mRNAs (NGF, TrkA, and p75) as well as the corresponding proteins, even if at a decreased concentration Conclusions:NGF plays a role in corneal trophism and regulates cell survival and fibroblast function. The absence of TrkA expression in keratoconus suggests a potential involvement of nerve growth factor in the pathogenesis of this disease

Keywords: keratoconus • growth factors/growth factor receptors • degenerations/dystrophies