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E Ghinelli, JD Rios, D Zoukhri, J Johansson, DA Dartt; Identification of Neurotrophins and Their Receptors in Adult Rat Conjunctiva and Lacrimal Gland . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3172.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:To determine which of the neurotrophins (NT): nerve growth factor (NGF), brain derived neurotrophic factor (BDNF), neurotrophin 3 (NT3), and neurotrophin 4 (NT4) and their receptors (NTrec): TrkA, TrkB, TrkB truncated form, TrkC and p75 are present in the adult rat conjunctiva and lacrimal gland. Methods:Conjunctival tissue, lacrimal gland and brain (the positive control) were removed from male adult Sprague Dawley rats (n=3). The expression of NGF, BDNF, NT3, NT4, TrkA, TrkB, TrkB truncated form, TrkC and p75 evaluated by Western Blotting technique. Tissues were homogenized in RIPA buffer containing DTT, 4x sample buffer and protease inhibitor cocktail and centrifuged at 4,000 rpm at 4°C. Proteins were separated by SDS-PAGE and transferred to nitrocellulose membranes. Membranes were blocked in 5% dried non-fat milk and incubated for 1 hour with the primary antibody. Immunoreactive bands were detected by the enhanced chemiluminescent method. Results:Western blot analysis showed that all NTs and NTrecs were present in the positive control , the brain. The conjunctiva contained the NTs: BDNF and NT3, a band for NT3 was obtained at ∼60 kD in both brain and conjunctiva rather than 30 kD. The conjunctiva also contained the NTrecs TrkA, TrkC and p75. The lacrimal gland contained the NTs: NGF, BDNF and NT3. A band for NT3 was again obtained at 60kD both in brain and lacrimal gland. The lacrimal gland contained the NTrecs TrkB TrkC and p75. TrkB appeared as light multiple bands perhaps representing multiple glycosylated forms of this receptor. Conclusions:We conclude that members of the NT family growth factors and their receptors are present in both the rat conjunctiva and lacrimal gland. Rat conjunctiva and lacrimal gland do not contain the same patterns of NT and NTrec expression Supported by EY04057 and EY06177.
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