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D.J. Brown, B. Holguin, A. Lee, C. Nguyen, J.V. Jester; The Transition From Keratocyte to Myofibroblast Is Accompanied by the Increased Expression of Neuregulin–1 . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2324.
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© ARVO (1962-2015); The Authors (2016-present)
The neuregulin family of growth factors plays an important role in mammalian development and regulates cell–cell interactions in a variety of organ systems in the adult. Our recent data demonstrated that multiple splice variants of neuregulin 1 (Nrg–1) are expressed in the normal adult human cornea. This study was designed to assess how changes in corneal stromal cell phenotypes affect neuregulin expression.
Freshly isolated rabbit keratocytes, human corneal fibroblasts, and a telomerized human keratocyte cell line were cultured under serum free conditions. Transforming growth factor beta (TGFß) was added to the cultures and the cells harvested for protein and total RNA with time. To assess the transformation of cells from the keratocyte to fibroblast to myofibroblast phenotype, the appearance of alpha smooth muscle actin was monitored. Quantitative real time polymerase chain reactions (qPCR) along with a combination of fluorescent primers, restriction endonucleases and image analyses were utilized to determine the amount and proportion of various splice variants of neuregulin. Western analyses were performed with specific neuregulin antibodies to verify the RNA results. Finally, the expression of Nrg–1 receptor was determined in these cell phenotypes.
RT–PCR and Western blot analyses demonstrate that Nrg–1 is expressed in these cells cultured in serum free conditions. As expected, exposing these cells to TGFß led to morphologic changes and the induction of alpha smooth muscle actin (SMA), a hallmark of the transition to a myofibroblastic phenotype. Along with this increase in SMA, the mRNA for neuregulin increased within 6 hours of TGFß treatment and protein levels accumulated in the cells. Interestingly, most of the increase in neuregulin was accounted for by an increase in specific isoforms (alpha variants) that were shown to stimulate corneal epithelial cell migration in vitro. However, stromal cells themselves do not produce neuregulin receptor even with TGFß stimulation.
The transition of corneal stromal cells to myofibroblasts induced by TGFß is accompanied by a rapid increase in specific isoforms of neuregulin. The induced forms of neuregulin represent variants affecting epithelial cell migration but have no affect on stromal cells. As such, this system may represent another example of stromal to epithelial interaction. As both the influence of TGFß and the presence of myofibroblasts are closely associated with the normal wound response in the cornea, we suggest that neuregulin may play a role in wound healing.
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