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G.W. Laurie, N. Wang, R.W. Raab, R.L. McKown; NFAT/mTOR Signaling and Downstream Promotion of MUC16 Expression by Lacrimal Prosecretory Mitogen Lacritin . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1606.
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© ARVO (1962-2015); The Authors (2016-present)
Lacritin is released from human lacrimal acinar cells during reflex tearing then flows past ductal cells onto the corneal surface. Lacritin is also expressed by human corneal and conjunctival cells. Addition to: (1) lacrimal acinar cells promotes enhanced secretion, (2) subconfluent salivary ductal cells triggers mitogenesis, and (3) subconfluent human corneal epithelial cells promotes proliferation, protection from TNFalpha and expression of corneal lacritin. Here we probe lacritin mitogenesis in more detail, and reveal a new secretory effect on the corneal surface.
Recombinant human lacritin was generated from the pLACm vector in E.coli and purified. Mitogenic signaling using 10 nM lacritin was studied in human salivary gland ducted cells (HSG) via 3H–thymidine incorporation. For siRNA depletion, SMARTpool siRNA were transfected with Lipofectamin 2000 in Opti–MEMI reduced serum medium. Transfected cells were monitored by Western Blotting and 3H–thymidine incorporation. Secretion studies utilized the human corneal epithelial cell line (HCE). MUC16 mRNA and protein were respectively monitored by RT–PCR and with monoclonal antibody CA125 in Western blotting and sandwich ELISA. Each experiment was done in a dose– and time– dependent manner.
(1) In confirmation of pharmacological studies with cyclosporin and rapamycin, siRNA depletion of mTOR and the transcription factor NFATc1 abrogates lacritin–dependent mitogenesis. (2) Lacritin–dependent NFATc1 nuclear translocation is dependent on calcium entry into cells. siRNA reveals that this is not via TRPC1. However STIM1 appears to be involved. (3) COX–2, but not IL–6 or OSCAR expression is downstream of NFAT. (4) In HCE cells, lacritin promotes mRNA and protein expression of membrane mucin MUC16 and possibly could be one mechanism by which lacritin is protective against TNFalpha.
Lacritin is a multipurpose growth and secretory factor of the human lacrimal–corneal axis that may play an important role in the renewal and secretory physiology of these epithelia.
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