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Darlene A. Dartt, Robin R. Hodges; Activation Of P2X7 Receptors In Lacrimal Gland Acini Increases Protein Secretion, But Not Cellular Permeability Or IL-1beta Secretion. Invest. Ophthalmol. Vis. Sci. 2012;53(14):626.
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© ARVO (1962-2015); The Authors (2016-present)
To determine if activation of P2X7 receptors is beneficial for the lacrimal gland by increasing protein secretion or deleterious by increasing cellular permeability and producing inflammatory mediators
Acini were isolated by collagenase digestion from male rat lacrimal glands. Cellular permeability was determined by measuring pore formation with and without BzATP (P2X7 agonist) using Yo-Pro. Secretion of mature IL-1β was determined by western blotting analysis of supernatant from acini incubated with BzATP or the positive control IL-1α. ATP release was investigated by preincubating acini with the pannexin inhibitor carbenoxolone (CBX) and stimulating with the cholinergic agonist carbachol (Cch) or the α1D-adrenergic agonist phenylephrine (Ph). Intracellular [Ca2+] ([Ca2+]i) was determined using fura 2. Acini were preincubated with CBX or a peptide pannexin inhibitor (PanX1) and stimulated with BzATP or Cch. Acini isolated from P2X7 null and wild type (WT) male mouse lacrimal glands were used to determine the effect of P2X7 on function. Peroxidase secretion (measures protein secretion) and [Ca2+]i were determined after stimulation of null and WT acini with BzATP, Cch, or Ph.
In rat lacrimal gland acini: 1) BzATP did not increase cellular permeability, but the positive control 1% Triton X100 increased permeability compared to basal, 2) BzATP did not increase secretion of mature IL-1β, but IL-1α elevated it, 3) Cch and Ph stimulated ATP release but this release was not altered by CBX, and 4) BzATP increase in [Ca2+]i was blocked 76.3 ± 9% by CBX and 92 ± 5% by PanX1, but Cch [Ca2+]i was unaltered. In mouse P2X7 null compared to WT lacrimal gland acini: 1) BzATP stimulated [Ca2+]i was 26 ± 16 and 440 ± 104 nM, Cch and Ph induced [Ca2+]i was unaltered and 2) BzATP stimulated peroxidase secretion was 1.3 ± 0.2 and 2.6 ± 1.4 fold over basal, Cch induced secretion was 3.8 ± 0.8 and 2.6 ± 0.4 fold, and Ph elevation in secretion was 6.8 ± 0.8and 13.5 + 1.4 fold, respectively.
Activation of P2X7 receptors stimulates protein secretion and their absence does not increase cellular permeability or produce inflammatory mediators. Thus, in the lacrimal gland activation of P2X7 receptors is beneficial, not deleterious.
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