Streptozotocin (STZ) mediates acute upregulation of serum and pancreatic osteopontin (OPN): a novel islet-protective effect of OPN through inhibition of STZ-induced nitric oxide production

Abstract

Osteopontin (OPN) is a secreted acidic phosphoprotein that binds to a cell-surface integrin-binding motif and is involved in many inflammatory and immune-modulating disorders. There is compelling evidence that soluble OPN can in a variety of situations help cells survive an otherwise lethal insult. In this study we show that OPN is localized in the rat pancreatic islets and ducts. Staining of pancreatic serial sections with islet hormone antibodies showed that all islet cells express OPN. Rats treated with a single dose of streptozotocin (STZ; 50 mg/kg) showed acute upregulation of serum OPN levels and pancreatic OPN mRNA and protein. Serum OPN dropped by the end of day 7 but was still higher than prediabetic levels. Pancreatic mRNA and protein showed a similar pattern. Twenty-four hours after STZ injection, the intensified OPN expression was localized towards the periphery of the islets and surrounded the remaining insulin-positive cells. To explore the significance of OPN acute upregulation, freshly isolated islets were pretreated with OPN (0·15-15 nM) before addition of STZ. OPN significantly reduced the STZ-induced NO levels in the islets through an Arg-GlyAsp (RGD)-dependent reduction of inducible NO synthase (iNOS) mRNA levels. Addition of OPN to freshly isolated mildly diabetic islets (blood glucose <300 mg/dl) significantly improved their glucose-stimulated insulin secretion and reduced their NO levels. Next we investigated the regulation of OPN in -cells. When STZ (5 mM) was added to the -cell line RINm5F it significantly increased OPN mRNA levels within 6 h. To distinguish between the effect of STZ and high glucose on OPN transcription, RINm5F cells were transfected with luciferase-labeled rat OPN promoter and treated with STZ (0·05-5 mM) or with glucose (5-25 mM). STZ induced upregulation of OPN promoter activity within 3 h, while high glucose induced upregulation of OPN promoter activity after 48 h. Our data introduce OPN as a novel islet protein that is differentially regulated by STZ and glucose in the islets. OPN initial upregulation after diabetes induction was probably due to STZ-induced toxicity, while maintenance of the high OPN levels might be due to hyperglycemia. The acute induction of OPN after STZ-induced diabetes might represent an endogenous mechanism to protect the islets against STZinduced cytotoxicity, partly via an RGD-dependent NO regulatory mechanism.

Pancreatic islet-specific overexpression of Reg3β protein induced the expression of pro-islet genes and protected the mice against streptozotocin-induced diabetes mellitus

“…5, A and B); upon Reg3␤ overexpression, there was a clear induction of the protein expression in almost all islet cells. Similarly, in wild-type islets, osteopontin was hardly detected on Western blots, although it has been reported to be detectable (24). Islets from RIP-I/Reg3␤ mice, on the other hand, exhibited a clear induction of the protein (Fig.…”

The Effects of Palmitate on Hepatic Insulin Resistance Are Mediated by NADPH Oxidase 3-derived Reactive Oxygen Species through JNK and p38MAPK Pathways

“…Although the current in vitro study may not reflect the in vivo situation, there is strong evidence for oxidative stress-dependent changes in intracellular signaling, resulting in insulin resistance in vivo (35). From a mechanistic perspective, increased reactive molecules can trigger activation of stress-sensitive serine/ threonine kinase signaling pathways, such as JNK and p38…”

Androgen and estrogen receptor–mediated mechanisms of testosterone action in male rat pelvic autonomic ganglia

“…PCR product was run on 2.2% agarose gels alongside a 100 bp ladder (Trackit 100bp DNA ladder, Invitrogen). GAPDH primers (Katakam et al, 2005) were used to amplify GAPDH cDNA to serve as a positive PCR control using the same PCR protocol (Forward "AGA CAG CCG CAT CTT CTT GT"; reverse "CTT GCC GTG GGT AGA GTC AT"; product 207 bp). ERα (Forward: "AAT TCT GAC AAT CGA CGC CAG"; reverse "GTG CTT CAA CAT TCT CCC TCC TC") and ERβ (Forward: "TTC CCG GCA GCA CCA GTA ACC"; reverse "TCC CTC TTT GCG TTT GGA CTA") cDNAs (345 bp and 262 bp respectively) were detected using the primers of (Kuiper et al, 1997) and the same PCR protocol as described above.…”