Multiple Low-dose Streptozotocin Model Research Articles

MKK3 signalling plays an essential role in leukocyte-mediated pancreatic injury in the multiple low-dose streptozotocin model

In vitro studies have implicated activation of the p38 mitogen-activated protein kinase (MAPK) signalling pathway in cytokine-mediated pancreatic β-cell injury. Activation of the p38 MAPK occurs through two different upstream kinases, mitogen-activated protein kinase kinase 3 (MKK3) and MKK6. This study examined the role of MKK3 signalling in an in vivo model of cytokine-dependent pancreatic injury induced by multiple low doses of streptozotocin (MLD-STZ). Groups of wild-type (WT) or Mkk3−/− C57BL/6J mice received 5 daily injections of STZ (40 mg/kg) and were killed on day 5, week 2 or week 4. MLD-STZ in WT mice exhibited two distinct phases of pancreatic damage: islet cell apoptosis (immunostaining for cleaved caspase-3) on day 5 in the absence of leukocyte infiltration, and this was followed by islet inflammation (leukocyte infiltration and cytokine production) and further islet cell apoptosis on day 14 resulting in a loss of insulin-producing β-cells and an 80% incidence of hyperglycaemia. Mkk3−/− mice were not protected from the initial phase of STZ-induced islet cell apoptosis day 5. However, Mkk3−/− mice were completely protected from the induction of hyperglycaemia. This was attributed to inhibition of leukocyte infiltration, production of pro-inflammatory cytokines and islet cell apoptosis at day 14 of MLD-STZ. In vitro studies showed that cultured islets from Mkk3−/− and WT mice are equally susceptible to STZ and cytokine-induced apoptosis. In conclusion, MKK3 signalling plays an essential role in the development of islet inflammation leading to destruction of β-cells and hyperglycaemia in MLD-STZ-induced pancreatic injury.

C-Jun amino terminal kinase 1 deficient mice are protected from streptozotocin-induced islet injury

In vitro studies have implicated the c-Jun amino terminal kinase (JNK) in cytokine-induced pancreatic injury leading to a loss of insulin production and hyperglycemia. We examined the role of JNK1 in the multiple low dose streptozotocin (MLD-STZ) model in which islet injury and hyperglycemia are dependent upon T cell immunity and pro-inflammatory cytokines. MLD-STZ in wild type mice induced islet leukocyte infiltration, cytokine production, β-cell apoptosis, and hyperglycemia. In contrast, Jnk1−/− mice were substantially protected from a loss of insulin producing cells and hyperglycemia in the MLD-STZ model despite a marked islet T cell and macrophage infiltrate. Based upon several lines of evidence, this protection was attributed to a reduction in TNF-α production by infiltrating Jnk1−/− macrophages leading to reduced β-cell apoptosis. In conclusion, JNK1 signaling plays an essential role in macrophage induced β-cell apoptosis and the development of hyperglycemia in MLD-STZ induced pancreatic injury.

Examination of the immunosuppressive effect of Δ 9-tetrahydrocannabinol in streptozotocin-induced autoimmune diabetes

Δ 9-Tetrahydrocannabinol (Δ 9-THC) is capable of modulating a variety of immune responses, but has not been evaluated in models of immune-based diabetes. The objectives of the present study were: (a) to investigate the effect of Δ 9-THC in an established model of multiple low dose streptozotocin (MLDSTZ)-induced autoimmune diabetes; and (b) to determine the contribution of the immune response in the MLDSTZ model. CD-1 mice were treated with 40 mg/kg STZ for 5 days in the presence or absence of Δ 9-THC treatment. Δ 9-THC administered orally in corn oil at 150 mg/kg for 11 days attenuated, in a transient manner, the MLDSTZ-induced elevation in serum glucose and loss of pancreatic insulin. MLDSTZ-induced insulitis and increases in IFN-γ, TNFα and IL-12 mRNA expression were all reduced on Day 11 by co-administration of Δ 9-THC. In separate studies, six doses of Δ 9-THC, given after completion of STZ treatment, was found equally effective in attenuating mice from MLDSTZ-induced diabetes. Studies performed using B6C3F1 mice showed moderate hyperglycemia and a significant reduction in pancreatic insulin by MLDSTZ in the absence of insulitis. In addition, MLDSTZ produced a less pronounced hyperglycemia compared to CD-1 mice that was not attenuated by Δ 9-THC. These results suggest that MLDSTZ can initiate direct β-cell damage, thereby augmenting the destruction of β-cells by the immune system. Moreover, these results indicate that Δ 9-THC is capable of attenuating the severity of the autoimmune response in this experimental model of autoimmune diabetes.

Troglitazone can prevent development of type 1 diabetes induced by multiple low-dose streptozotocin in mice

Recent investigations suggest that cytotoxic cytokines such as tumor necrosis factor (TNF)α and interleukin (IL)-1β or free radicals play an essential role in destruction of pancreatic β cells in Type 1 diabetes and that, therefore, anti-oxidant or anti-TNFα and IL-1β therapy could prevent the development of Type I diabetes. Troglitazone belongs to a novel class of antidiabetic agent possessing the ability to enhance insulin action provably through activating PPAR γ and to scavenge free radicals. In the present study, we examined whether troglitazone can prevent the development of Type 1 diabetes in multiple, low-dose streptozotocin (MLDSTZ)-injected mice, hi addition, effects of troglitazone on cytokine-induced pancreatic β cell damage were examined in vitro. Type 1 diabetes was induced by MLDSTZ injection to DBA 2 mice (40 mg/kg/day for 5 days). Troglitazone was administered as a 0.2% food admixture (240 mg/kg/day) for 4 weeks from the start of or immediately after STZ injection. MLDSTZ injection elevated plasma glucose to 615 ± 8 mg dl 4 weeks after final STZ injection and was accompanied by infiltration of leukocytes to pancreatic islets (insulitis). Troglitazone treatment with MLDSTZ injection prevented hyperglycemia (230 ± 30 mg dl ) and, suppressed insulitis and TNFα production from intraperitoneal exudate cells. TNFα (10 pg ml ) and IL-1β (1 pg ml ) addition to hamster insulinoma cell line HIT-T15 for 7 days in vitro decreased insulin secretion and cell viability. Simultaneous troglitazone addition (0.03 ~ 3 μM) significantly improved cytokine-induced decrease in insulin secretion and in cell viability. These findings suggest that troglitazone prevents the development of Type 1 diabetes in the MLDSTZ model by suppressing insulitis associated with decreasing TNFa production from intraperitoneal exudate cells and the subsequent TNFα and IL-1β-induced β cell damage.

Beta-cell apoptosis is responsible for the development of IDDM in the multiple low-dose streptozotocin model.

Although insulin-dependent diabetes mellitus (IDDM) results from irreversible loss of beta cells, the mode of cell death responsible for this loss has not previously been categorized. In this study, the multiple low-dose streptozotocin (stz) model (intraperitoneal injection of stz at a concentration of 40 mg/kg body weight per day for five consecutive days) was used to investigate beta-cell death during the development of IDDM in male C57B1/6 mice. Apoptotic cells were evident by light microscopy within the islets of Langerhans of treated animals from day 2 (the day of the second stz injection) until day 17. Immunohistochemical localization of insulin to the dying cells confirmed the beta-cell origin of the apoptosis. Two peaks in the incidence of beta-cell apoptosis occurred: the first at day 5, which corresponded to an increase in blood glucose concentration, and the second at day 11, when lymphocytic infiltration of the islets (insulitis) was maximal. Insulitis did not begin until day 9, by which time treated animals had developed overt diabetes as revealed by blood glucose and pancreatic immunoreactive insulin (IRI) measurements. Beta-cell apoptosis preceded the appearance of T-cells in the islets and continued throughout the period of insulitis. Thus, whether induced by stz or a subsequent immune response, apoptosis is the mode of cell death responsible for beta-cell loss in the multiple low-dose stz model of IDDM.