Goblet cell differentiation from conjunctival keratinocytes

The conjunctival epithelium of the eye comprises goblet cells, which are unicellular mucin- secreting glands. Both the conjunctival keratinocytes as well as the goblet cells derive from a common bipotent epithelial progenitor cell. Mucin deficiency is present in several ocular pathologies such as chemical and thermal burns, Stevens-Johnson syndrome, Dry Eye syndrome, Sjögren syndrome, atopic keratoconjunctivitis, ocular cicatricial pemphigoid or Vitamin A deficiency. Patients with Sjögren syndrome (SS), a systemic autoimmune disease, show a significant reduction of the number of goblet cells, reduced levels of the goblet cell mucin MUC5AC in tears and reduced MUC5AC mRNA expression in impression cytology samples of the conjunctiva. Advances in knowledge of the molecular mechanisms involved in the pathogenesis of SS may allow the development of more selective therapies. This project aims to identify the mechanisms, which direct a proliferating conjunctival epithelial cell to generate a more differentiated mucin secretory goblet cell. Prerequisite to study the differentiation pathways in conjunctival epithelial cells is to gain know-how in culturing human keratinocytes. Only then, enrichment of human goblet cells with EGF to obtain a sufficient number of goblet cells in cell culture for subsequent experiments and assays can be carried out as step one. In a second step the ERK pathway activated by EGF in cultured conjunctival epithelial cells of healthy donors and SS patients will be analyzed and the effect of inhibition of the ERK signaling cascade on the differentiation of conjunctival epithelial cells will be evaluated. In addition, the expression of Sox9 and Math1 in human conjunctival epithelial cells of healthy donors as well as of SS patients and the effect of gene knockdown by siRNA will be investigated. The results should provide new insights into the interaction between extracellular and intracellular regulatory factors in controlling lineage determination of conjunctival stem cells and will shed light on the molecular mechanisms responsible for the conjunctival cell fate decision. Moreover, understanding how signaling pathways are pathologically altered in Sjögren syndrome will be important in improving the therapeutic utility of adult stem cells.

The conjunctival epithelium of the eye comprises goblet cells, which are unicellular mucin- secreting glands. Both the conjunctival keratinocytes as well as the goblet cells derive from a common bipotent epithelial progenitor cell. Mucin deficiency is present in several ocular pathologies such as chemical and thermal burns, Stevens-Johnson syndrome, Dry Eye syndrome, Sjögren syndrome, atopic keratoconjunctivitis, ocular cicatricial pemphigoid or Vitamin A deficiency. Patients with Sjögren syndrome (SS), a systemic autoimmune disease, show a significant reduction of the number of goblet cells, reduced levels of the goblet cell mucin MUC5AC in tears and reduced MUC5AC mRNA expression in impression cytology samples of the conjunctiva. Advances in knowledge of the molecular mechanisms involved in the pathogenesis of SS may allow the development of more selective therapies. This project aims to identify the mechanisms, which direct a proliferating conjunctival epithelial cell to generate a more differentiated mucin secretory goblet cell. Prerequisite to study the differentiation pathways in conjunctival epithelial cells is to gain know-how in culturing human keratinocytes. Only then, enrichment of human goblet cells with EGF to obtain a sufficient number of goblet cells in cell culture for subsequent experiments and assays can be carried out as step one. In a second step the ERK pathway activated by EGF in cultured conjunctival epithelial cells of healthy donors and SS patients will be analyzed and the effect of inhibition of the ERK signaling cascade on the differentiation of conjunctival epithelial cells will be evaluated. In addition, the expression of Sox9 and Math1 in human conjunctival epithelial cells of healthy donors as well as of SS patients and the effect of gene knockdown by siRNA will be investigated. The results should provide new insights into the interaction between extracellular and intracellular regulatory factors in controlling lineage determination of conjunctival stem cells and will shed light on the molecular mechanisms responsible for the conjunctival cell fate decision. Moreover, understanding how signaling pathways are pathologically altered in Sjögren syndrome will be important in improving the therapeutic utility of adult stem cells.