Protective Effect of Tanshinone IIA during the Acute Stage of Focal Cerebral Ischemia in Rats

Abstract

Abstract:

Objective To investigate the effect of tanshinone IIA (TSA) on the protein expression of phosphorylated cyclic adenosine monophosphate response element binding protein (p-CREB) and transducers of regulated CREB1 (TORC1) and volume of cerebral infarction in rats' parietal cortex during focal cerebral ischemia. Methods Male Sprague-Dawley rats were randomly divided into sham group, control group, TSA low dose group and TSA high dose group, twelve in each group. Focal cerebral ischemia model was induced by occlusion of the right middle cerebral artery using the intraluminal suture method. The rats were scored after waking up. 2, 3, 4 or 5 scores were brought into this study. TSA solution (10 mg/kg, TSA-L; or 20 mg/kg, TSA-H) was injected intraperitoneally immediately after the rats waked up. The rats were scored and sacrified at 24 h after waking up. Infarct size was analyzed with 2, 3, 5-triphenyltetrazolium chloride (TTC). Western blotting was used to analyze the protein expression of TORC1 and p-CREB of each group in the nerve cell, and the positive products were analyzed by image analysis system. Results Compared with control group, the infarct size of TSA-H group was significantly decreased (P =0.004). And nuclear accumulation of TORC1 was only significantly increased in TSA-H group (P <0.001). Compared with control group, the decrease of the infarct size in TSA-L group and the increase of nuclear protein expression of TORC1 have no remarkable differences (P =0.148, P =0.083, respectively), the expression of TORC1 (Total) and p-CREB (Nucleus) protein level in TSA-H (P <0.001) and TSA-L ((P =0.002, P =0.001) group was significantly increased, the enhanced TORC1 is localized in the nucleus and cytoplasm of neurons. Conclusion TSA could protect rat brain from ischemic damage and upregulate the expression of TORC1-CREB pathway in rats' parietal cortex during focal cerebral ischemia.

Key words: Transducers of regulated CREB1; cAMP response element binding protein; Tanshinone IIA; Middle cerebral artery occlusion

摘要：

目的探讨丹参酮ⅡA（TashinoneⅡA，TSA）对局灶性脑缺血急性期大鼠顶叶皮质磷酸化环腺苷酸反应元件结合蛋白（phosphorylated cyclic adenosine monophosphate response element binding protein，p-CREB）和CREB1活性调节转导子（transducers of regulated CREB1，TORC1）的蛋白表达和脑梗死体积的影响。方法采用成年健康雄性SD大鼠，随机分为假手术组、对照组、小剂量TSA组和大剂量TSA组，每组各12只。用线栓法制作右侧大脑中动脉闭塞模型，采用Longa评分判断动物模型成功与否。小剂量TSA 组和大剂量TSA组在造模后立刻腹腔注射TSA溶液，剂量分别为：10 mg/kg及20 mg/kg，假手术组及对照组分别腹腔注射等量的生理盐水。造模成功后24 h将大鼠处死，采用2% 2，3，5-三苯基四唑氮红染色计算梗死体积，应用Western blotting法检测各组手术侧顶叶皮质梗死灶神经细胞核TORC1和细胞核p-CREB以及细胞TORC1的蛋白表达。结果与对照组相比，大剂量TSA组大鼠脑组织梗死体积明显降低（P =0.004），细胞核TORC1 表达显著增加（P <0.001）；与对照组相比，小剂量TSA组大鼠脑组织梗死体积减小，无显著差异（P =0.148），细胞核TORC1蛋白表达升高，也无显著差异（P =0.083），细胞核p-CREB和细胞TORC1的蛋白表达水平无论在大剂量组（P均＜0.001），还是小剂量组（P分别为0.002和0.001）均显著升高。结论 TSA可以上调脑梗死大鼠病灶侧顶叶皮质神经细胞TORC1-CREB信号通路的表达。

关键词: 环腺苷酸反应元件结合蛋白活性调节转导子; 环腺苷酸反应元件结合蛋白; 丹参酮ⅡA; 大脑中动脉闭塞

LIU Ling-Ling*, LIU Hong, YIN Jun-Ling, CHEN Hui-Ying, LIU Xiao-Lei, JIAO Qing-Hai, WU Yi-Ping.. Protective Effect of Tanshinone IIA during the Acute Stage of Focal Cerebral Ischemia in Rats[J]. Chinese Journal of Stroke, 2014, 9(01): 20-.

刘玲玲1，刘红1，尹君玲1，陈惠英1，刘晓蕾1，焦清海1，武一平2. 丹参酮ⅡA对局灶性脑缺血大鼠急性期脑保护作用的相关研究[J]. 中国卒中杂志, 2014, 9(01): 20-.

References

左玮, 陈乃宏. 脑缺血后细胞死亡/存活信号的传递及防治策略[J]. 生理科学进展, 2013, 44:200-205.

Bh a t t a ch a r y a P, Pa n d e y AK, Pa u l S, e t a l .Aquaporin-4 inhibition mediates piroxicam-induced neuroprotection against focal cerebral ischemia/reperfusion injury in rodents[J]. PLoS One, 2013,8:e73481.

Meng C, He Z, Xing D. Low-level laser therapy rescues dendrite atrophy via upregulating BDNF expression:implications for Alzheimer's disease[J]. J Neurosci, 2013, 33:13505-13517.

Io u r ge n ko V, Zh a n g W, Mi ck a n i n C, e t a l .Identification of a family of cAMP response elementbinding protein coactivators by genome-scale functional analysis in mammalian cells[J]. Proc Natl Acad Sci USA, 2003, 100:12147-12152.

Deng J, Zhang XL, Wang JW, et al. Expression and regulated nuclear transpor t of transducers of regulation CREB1 in retinal ganglion cells[J].Neuroscience, 2009, 159:1023-1031.

Li S, Zhang C, Takemori H, et al. TORC1 regulates activity dependent CREB-target gene transcription and dendritic growth of developing cortical neurons[J].J Neurosci, 2009, 29:2334-2343.

Longa EZ, Weinstein PR, Carlson S, et al. Reversible middle cerebral artery occlusion without craniectomy in rats[J]. Stroke, 1989, 20:84-91.

Liu L, Zhang X, Wang L, et al. The neuroprotective effects of Tanshinone IIA are associated with induced nuclear translocation of TORC1 and upregulated expression of TORC1, pCREB and BDNF in the acute stage of ischemic stroke[J]. Brain Res Bull, 2010,82:228-233.

Wang L, Zhang X, Liu L, et al. Atorvastatin protects rat brains against permanent focal ischemia and downregulates HMGB1, HMGB1 receptors (RAGE and TLR4), NF-kappaB expression[J]. Neurosci Lett,2010, 471:152-156.

Tatlisumak T, Carano RA, Takano K, et al. A novel endothelin antagonist, A-127722, attenuates ischemic lesion size in rats with temporary middle cerebral artery occlusion:a diffusion and perfusion MRI study[J]. Stroke, 1998, 29:850-857.

Di r nagl U, I a de col a C, Moskowit z M, e t a l.Pathobiology of ischaemic stroke:an integrated view[J]. Trends Neurosci, 1999, 22:391-397.

Lo EH, Dalkara T, Moskowitz MA. Mechanisms,challenges and opportunities in stroke[J]. Nat Rev Neurosci, 2003, 4:399-415.

Yi JH, Park SW, Kapadia R, et al. Role of transcription f a c tor s i n med ia t i ng pos t-i s chemic c e r ebr a l inflammation and brain damage[J]. Neurochem Int,2007, 50:1014-1027.

McTague J, Amyotte N, Kanyo R, et al. Different s ig n a l i n g me c h a n i sms a r e i nvol ve d i n t h e norepinephrine-stimulated TORC1 and TORC2 nucle a r t r a nsloc at ion i n r at pi ne a locy t e s[J ].Endocrinology, 2012, 153:3839-3849.

Martin F, Nunez C, Marin MT, et al. Involvement of noradrenergic transmission in the PVN on CREB activation, TORC1 levels, and pituitary-adrenal axis activity during morphine withdrawal[J]. PLoS One,2012, 7:e31119.

Chat ur vedi RK, Hen nessey T, Joh r i A, et al.Transducer of regulated CREB-binding proteins (TORCs) transcription and function is impaired in Huntington's disease[J]. Hum Mol Genet, 2012,21:3474-3488.

Sasaki T, Takemori H, Yagita Y, et al. SIK2 is a key regulator for neuronal survival after ischemia via TORC1-CREB[J]. Neuron, 2011, 69:106-119.

夏阳阳, 蒋春明, 孙琤, 等. 丹参酮ⅡA对腹膜透析液诱导的氧化应激及腹膜间皮细胞凋亡因子表达的影响[J]. 东南大学学报, 2013, 32:36-41.

孟庆春, 王玲玲, 孔小燕, 等. 丹参酮ⅡA磺酸钠对大鼠肢体缺血再灌注损伤时IL-8、TNF-α的影响[J]. 西部医学, 2013, 25:353-356.

Gao J, Yang G, Pi R, et al. Tanshinone IIA protects neonatal rat cardiomyocytes from adriamycin-induced apoptosis[J]. Transl Res, 2008, 151:79-87.

牛雪花, 华海涌, 郭文建, 等. 丹参酮ⅡA磺酸钠治疗晚期血吸虫病肝纤维化的效果观察[J]. 中国血吸虫病防治杂志, 2013, 25:137-140.

Wei B, You MG, Ling JJ, et al. Regulation of antioxidant system, lipids and fatty acid β-oxidation contributes to the cardioprotective effect of sodium tanshinone ⅡA sulphonate in isoproterenol-induced myocardial infarction in rats[J]. Atherosclerosis, 2013,230:148-156.

徐兰举, 杨卓. 在脑缺血过程中CREB和cAMP反应元件介导的基因表达[J]. 天津医药, 2007, 35:625.

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