Study On Expression Of Endoplasmic Reticulum Stress Related Genes In Aorta Of Type 2 Diabetes Mice

Atherosclerosis is the common complication of diabetes, 80% of adults with diabetes die from atherosclerosis. Among the diabetes, type 2 diabetes (T2DM) is 90%. A vast amount of research has gone into understanding the mechanisms by which diabetes promotes atherosclerosis, Several cellular stress mechanisms have been proposed to explain the explain the acceleration of atherosclerosis in diabetes mellitus (DM), including advanced glycation end product, oxidative stress and protein kinase C. Recent findings have shed light on endoplasmic reticulum stress (ERS) in the pathogenesis of atherosclerosis. However, the cellular mechanisms underlying the accelerated progression of atherosclerotic lesions in diabetic arteries are still largely unknown.Therefore, in order to investigate the role of ERS in development and progression of type 2 diabetic atherosclerosis, We therefore used an apolipoprotein E/low-density lipoprotein receptor–deficient (AL) mice model in which T2DM was induced by treatment of a single dose of streptozotocin (STZ) at 6 wk of age combined with high-fat/high-sucrose diet feeding, ND group were normal diet fed mice injected with a same dosage of citrate buffer served as control after normal diet for 3 wk. At the age of 10 wk, T2DM was confirmed by measuring weight, the fasting blood glucose (FBG), insulin, HOMA-insulin resistant (HOMA-IR) and glucose tolerance test (GTT). The two experimental groups mice were sacrificed at 10 and 18 wk of age (n≥10 for each time point), we analysis the atherosclerotic lesion in the Aortic arch, aortic root and aorta enface. Aortic root sections from T2DM group mice and ND group mice at 10 weeks of age or 18 weeks of age were examined for markers of ERS.During the experimental period, T2DM group mice exhibited weight loss compared with ND group mice(P < 0.05). Hyperglycemia developed in T2DM group, plasma glucose levels at 4 weeks after STZ administration were elevated in T2DM group and hyperglycemia persisted until end of the experiment (P < 0.01). Moreover, T2DM group showed significantly higher insulin levels compared with ND group (P < 0.05). HOMA-IR inedx of T2DM group was significantly higher than ND group (P < 0.01) indicating development of insulin resistant (IR). GTT performed at 10 and 18 wk of age showed that T2DM group mice exhibited a significantly higher glucose intolerance compared with the ND group indicating impaired glucose tolerance. T2DM group mice showed significantly elevated TC levels compared with ND group mice at 18-week-old (P < 0.01). TG levels were also higher in T2DM group (P < 0.01), We also observed that LDL-C levels were markedly elevated between 10 and 18 weeks from the start of the study (P < 0.01). Moreover, average HDL-C levels displayed a significant increase in T2DM group mice compared with the ND groups mice at 10 wk of age (P < 0.01). Thus, these dates indicate that we have established a T2DM mouse model exhibiting hyperglycemia, hyperlipidemia, IR and impaired glucose tolerance in AL mice for further research.To investigate the role of T2DM on the development of lesions, we evaluated lesion development in aortic sinus and en face aorta, atherosclerotic lesion formation was considerably increased in the aortic sinus and en face aorta from T2DM group mice in comparison to ND group mice (P < 0.05).Aorta from T2DM group mice were characterized by significantly increased (glucose-regulated protein78, GRP78) and (C/EBP homologous protein, CHOP) mRNA levels at age of 10 weeks and persisted until end of the experiment(P < 0.01).. To understand the contribution of ERS induced apoptosis on the progression of atherosclerosis, we measured the expression of genes about ERS related to apoptosis such as CHOP, JNK2 and Caspase12, the expressions were higher in T2DM group mice at 10 and 18 weeks (P < 0.01).Aortic root sections from T2DM and ND group mice at 10 and 18 weeks of age were examined for markers of ERS (GRP78, CHOP), apoptotic cell death (Actived-Caspase3) and plaque necrosis core. Expression of ERS markers (GRP78) dramatically increased in T2DM group mice. CHOP/GADD153 is an established marker of ERS and a member of the C/EBP gene family of transcription factors with proapoptotic characteristics. We observed weak nuclear CHOP immunostaining in ND group mice at 10and 18 wk of age that showed no morphological signs of apoptosis. In advanced lesions, strong cytoplasmic and nuclear staining was observed in intact cells within the lesion in T2DM group mice at the age of 18 weeks. Lesion necrosis is a consequence of lesion cell apoptosis, cell death in advanced atherosclerotic lesion leads to formation of necrotic cores and promotes lesion instability. We observed Actived-Caspase3-positive staining within the developing necrotic core in T2DM group mice, the area of lesion necrotic core is also lager in T2DM group mice than ND group mice at at the age of 18 weeks (P < 0.01). These data suggest that T2DM-induced ERS plays an important role in the development of atherosclerosis, cell death and formation of necrotic core.