Vasoactive intestinal peptide inhibits TNF-α-induced apoptotic events in acinar cells from nonobese diabetic mice submandibular glands

Abstract

IntroductionThe role of apoptotic secretory epithelium as a pro-inflammatory triggering factor of exocrine dysfunction is currently explored in Sjogren's syndrome patients and in the nonobese diabetic (NOD) mouse model. Vasoactive intestinal peptide (VIP) has anti-inflammatory effects in various models of chronic inflammation. Our goal was to analyse the effect of TNF-α on apoptotic mediators in isolated acinar cells from NOD submandibular gland and their modulation by VIP.MethodsAcinar cells were isolated from submandibular glands of 16-week-old NOD females with salivary flow decline. Age-matched BALB/c females or eight-week-old NOD females were used as controls. Apoptotic mediators and TNF-α receptor expression were assessed by immunoblotting and RT-PCR, caspase 3 activity was assessed by optical density at 405 nm with Ac-DEVD-pNA as a substrate and chromatin condensation by Hoechst stain. They were evaluated in resting conditions and after a 3.5 or 6 hours of culture with TNF-α. VIP effects in acinar cells were assessed at 100 nM in TNF-α-treated cultures and VIP receptor functional assays by radio immunoassay (cAMP) or enzymatic detection (amylase).ResultsNOD acinar cells at 16 weeks present an increased expression of TNF-α receptor1 together with increased Bax, tumour protein 53-induced nuclear protein1α (TP53INP1α), caspase 3 activity and chromatin condensation. Acini from NOD mice were more sensitive to TNF-α-induced increases of apoptotic mediators than control cells. VIP inhibited TNF-α-induced apoptotic events through functional VPAC1 receptors coupled to the protein kinase A (PKA) signalling pathway.ConclusionsOur results indicate that acinar cells isolated from submandibular glands of NOD mice with salivary dysfunction are more sensitive to apoptosis induced by TNF-α which could be prevented by VIP through a PKA-mediated pathway.

Vasoactive intestinal peptide/vasoactive intestinal peptide receptor relative expression in salivary glands as one endogenous modulator of acinar cell apoptosis in a murine model of Sjögren's syndrome

“…Submandibular glands were removed and transferred immediately to ice-cold RPMI-1640, 10% fetal bovine serum (FBS) for acinar cell isolation, as described previously [16]. Acinar cells were washed and seeded on flat-bottomed 24-well microtitre plates (Corning Glass, Corning, NY, USA) and incubated for 2 h at 37°C in a humidified incubator with 5% CO2 to separate immune adherent cells and viability determination [16].…”

“…Among them, a progressive loss of activity of the neural isoform of nitric oxide synthase (NOS 1) in NOD exocrine glands at the Sjögren's syndromelike period has been described [12,15]. The lower levels of NOS activity were found in glands of 16-week-old NOD mice that presented increased apoptosis of acinar cells and increased levels of tumour necrosis factor (TNF)-a, among other T helper type 1 (Th1) cytokines in the serum [15,16]. Vasoactive intestinal peptide (VIP), described initially as a vasodilator and prosecretory neuropeptide, has trophic effects on acini [17,18] and strong anti-inflammatory properties in several models of chronic inflammatory diseases [19][20][21].…”

“…Prediabetic NOD mice treated systemically with VIP showed increased serum interleukin (IL)-10 and reduced Th1 cytokine levels [22] while gene-transfer of VIP onto NOD submandibular glands prevented saliva secretion loss and partly reduced glandular Th1 cytokine expression [23]. Furthermore, VIP showed a clear anti-apoptotic effect on acinar cells isolated from NOD submandibular glands driven to apoptosis through TNF-a/TNF-aR1-mediated pathways [16].…”

Vasoactive Intestinal Peptide (VIP) and VIP Receptors-Elucidation of Structure and Function for Therapeutic Applications

“…VIP administration reportedly inhibited apoptosis in submandibular gland-derived acinar cells and neurinoma cells [43,44].…”

“…10) VIP might inhibit apoptosis by stimulating the expression of the apoptosis-inhibiting gene Bcl2 or by inhibiting the activity of caspase 3, which is a part of the caspase cascade [43]. VIP administration reportedly inhibited apoptosis in submandibular gland-derived acinar cells and neurinoma cells [43,44].…”