VIP/VPAC1 expression did not vary in BALB/c glands with mouse age and, in contrast with NOD glands, freshly isolated acinar cells seemed not to be prone to apoptosis. Acinar cells from NOD mice could be further induced to Enzalutamide apoptosis with a concentration
of TNF-α (10 ng/ml) that was almost ineffective in normal acinar cells. VIP inhibited TNF-α-induced apoptosis in NOD acinar cells through a VPAC1/cAMP/PKA pathway, while neither VPAC2 receptors nor the neuropeptide could be detected in acini, indicating that their expression in whole glands would not correspond to acinar cells. Finally, we found a reduced phagocytic index of NOD macrophages to engulf apoptotic acinar cells compared to normal macrophages, but their basal inflammatory phenotype was suppressed during phagocytosis and VIP stabilized this suppressor regulatory phenotype. It is noteworthy that the time–course of VIP/VPAC1 relative expression decline is similar to the kinetics of nNOS activity loss shown previously and parallels NVP-LDE225 ic50 the reduction in the secretory response to muscarinic acetylcholine receptor stimulation . It also coincided with the loss of acinar cell homogeneous structure of the glands and a higher ductal to acinar cell ratio in the glands at 16 weeks of age . The localization of this enzyme is normally confined
to neural fibres in close proximity to gland epithelial cells where NO contributes to salivary flow. Consistent with this, NOD mice submandibular glands showed a
reduced NOS activation through VIP receptors that coincided with the reduction in salivary flow . While VIP can induce NOS in peripheral and central neurones, VIP expression is regulated by neural NOS activity and knock-out mice for neural NOS isoform express lower neuronal VIP levels . In rat salivary glands VIP is localized in nerve fibres rather than in acinar cells, being mainly released from nerves surrounding acini where it displays trophic effects on epithelial cells [17,18]. In fact, the release of trophic and anti-apoptotic stimuli from nerve terminals with long-term effects on salivary gland parenchyma is the rationale of a newly designed device to restore salivary flow in patients with SS and other sicca-associated pathologies . Acinar cells from both normal and NOD isometheptene submandibular glands express only VPAC1 receptors, as reported previously . In these cells, VIP was able to reduce apoptosis via cAMP/PKA pathway, as derived from the fact that H89 reversed VIP effect on bax expression  and Bad phosphorylation, a step previous to the loss of its apoptotic effect through binding to 14–3–3 in cytosol . Evidence shown here indicates that acinar epithelium of NOD but not BALB/c glands present increased apoptosis along with a dysregulated NF-κB basal activation consistent with a predominant apoptosis-to-survival intracellular set-point.