Role of Neurotrophins and Neurotrophin Receptors in Rat Conjunctival Goblet Cell Secretion and Proliferation

Abstract

To determine which neurotrophins (NTs)-nerve growth factor (NGF), brain-derived neurotrophin (BDNF), neurotrophin-3 (NT3), and neurotrophin-4 (NT4)-and their receptors (NTrs) TrkA, TrkB, TrkC, and p75 are present in rat conjunctiva and cultured rat goblet cells (CGCs) and whether NTs stimulate glycoconjugate secretion or cell proliferation. Western blot analysis and immunofluorescence microscopy determined presence and location of NTs and NTrs. CGCs were incubated with NTs (10(-12)-10(-8) M) for 2 or 24 hours to measure secretion or proliferation, respectively. An enzyme-linked lectin assay analyzed glycoconjugate secretion. WST-8 determined cell proliferation. Western blot analysis showed all NTs and NTrs in both conjunctiva and CGCs. The cytoplasm of conjunctival stratified squamous cells and goblet cell lateral membranes contained NGF, BDNF, and NT4. Stratified squamous cell membranes contained NT3. In CGCs, NGF and BDNF had punctuate perinuclear staining. The nucleus contained NT3 and cytosol contained NT4. TrkA, TrkB, and p75 immunoreactivity was on conjunctival goblet cell lateral membranes. Plasma membranes of the basal layer of stratified squamous cells contained TrkA. Stratified squamous cell and goblet cell nuclei contained TrkC. In CGCs, all NTrs were present in the nucleus. NGF and BDNF, but not NT3 and NT4, induced a concentration-dependent stimulation of secretion from CGCs with a maximum increase of 10(-9) M each. No effect on cell proliferation was detected with any NTs. Rat conjunctival goblet cells and CGCs contain all NTs and NTrs. Only NGF and BDNF stimulated goblet cell glycoconjugate secretion, and none induced CGC proliferation.