Corneal defects due to diabetes mellitus (DM) may cause severe vision impairments. Current studies focus on the corneal epithelium and nerve defects neglecting the corneal stroma. The aim of this study was to develop a 3D in vitro model to examine the interactions between corneal stroma and nerves in the context of DM. Primary human corneal stromal fibroblasts isolated from healthy (HCFs), Type 1 (T1DM) and Type 2 (T2DM) patients were stimulated with stable ascorbic acid to secrete and assemble an extracellular matrix (ECM). Human neuronal cells were then seeded on top and differentiated to create the 3D co-cultures. Our data revealed successful co-culture of stromal fibroblasts and neuronal cells with large elongated neuron extensions. T2DM showed significant upregulation of Collagen III and IGF1 when compared to T1DM. Interestingly, upon nerve addition, those markers returned to HCF levels. Neuronal markers were also differentially modulated with T2DM co-cultures expressing high levels of βIII tubulin where T1DM co-cultures expressed Substance P. . Overall, our unique 3D co-culture model provides us with a tool that can be utilized for both molecular and therapeutic studies for diabetic keratopathy.
Keywords: 3D in vitro model; Corneal nerves; Diabetes mellitus; Diabetic corneal stroma.
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