Role of conventional protein kinase Cγ and its interacting protein 14-3-3γ in oxygen\|glucose deprivation-induced ischemic injury of primary cultured mouse cortical neurons

doi:10.16098/j.issn.0529-1356.2015.02.001

Abstract

Abstract:

Objective To explore the role of conventional protein kinase Cγ (cPKCγ) and its possible interacting protein 14-3-3γ in 1hour oxygen-glucose deprivation (OGD)/24hours reoxygenation and glucose restoration (R)-induced ischemic injury in primary cultured cortical neurons of mice.Methods Using cPKCγ gene knockout (cPKCγ^-/-) mice, 1hour OGD/24hours R-induced ischemic injury model of primary cultured cortical neurons, and the techniques of co-immunoprecipitation (Co-IP), Western blotting and 3-(4,5-dimethyl thiazole-2)-2,5-diphenyl four azole nitrogen bromine salt (MTT) assay, the interaction of cPKCγ and 14-3-3γ and their effects on ischemic injury were determined. Results The Co-IP results confirmed the existence of cPKCγ interaction with 14-3-3γ in the cerebral cortex of mice; the decreased protein expression of cPKCγ was accompanied with the increase of 14-3-3γ protein levels in primary cultured cortical neurons from cPKCγ+/+mice after 1hour OGD/24hours R treatment (P<0.05, n=6 per group). Although the 14-3-3γ protein level in cortical neurons from cPKCγ-/-mice was higher than that from cPKCγ+/+ mice, 1hour OGD/24hours R treatment caused a significant decrease of 14-3-3γ protein level in cortical neurons from cPKCγ-/-mice (P<0.05, n=6 per group), suggesting cPKCγ may affect 14-3-3γ protein expression level in cortical neurons. In addition, the MTT results demonstrated that the 14-3-3γ inhibitor R18 (100μmol/L) significantly enhanced 1hour OGD/24hours R-induced ischemic injury of primary cultured cortical neurons from both cPKCγ^+/+and cPKCγ^-/-mice (P<0.05, n=6 per group).Conclusion cPKCγ may interact with 14-3-3γ and affect 14-3-3γ protein expression level in cortical neurons of mice. cPKCγ and 14-3-3γ may protect primary cultured cortical neurons against 1hour OGD/24hours R-induced ischemic injury.

Key words: Primary cultured cortical neuron, Oxygen-glucose deprivation, Ischemic injury, Conventional protein kinase Cγ, 14-3-3γ, Western blotting, Mouse

ZHANG Xin-xin YANG Xuan MA Wei CHEN Lu-mian HAN Song LI Jun-fa*. Role of conventional protein kinase Cγ and its interacting protein 14-3-3γ in oxygen\|glucose deprivation-induced ischemic injury of primary cultured mouse cortical neurons[J]. AAS, 2015, 46(2): 145-150.

References

［1］Zhang N, Yin Y, Han S, et al. Hypoxic preconditioning induced neuroprotection against cerebral ischemic injuries and its cPKCgamma-mediated molecular mechanism ［J］. Neurochem Int, 2011,58（6）: 684-692.
［2］Meller R, Schindler CK, Chu XP, et al. Seizure-like activity leads to the release of BAD from 14-3-3 protein and cell death in hippocampal neurons in vitro ［J］. Cell Death Differe, 2003, 10（5）: 539-547.
［3］Ye SQ, Zhou XY, Lai XJ, et al. Silencing neuroglobin enhances neuronal vulnerability to oxidative injury by down-regulating 14-3-3γ ［J］. Acta Pharmacol Sin, 2009, 30(7): 913-918.
［4］Luo H, Su JE, Han S, et al. cPKCγ in HPC protects N2a cells against OGD-induced injury and its molecular mechanism［J］. Basic and Clinical Medicine,2013,33(3):308-313. (in Chinese)
罗宏，苏吉儿，韩松，等. cPKCγ在HPC抗N2a细胞OGD损伤中作用及分子机制［J］. 基础医学与临床，2013，33（3）：308-313.
［5］Chen XQ, Yu AC. The association of 14-3-3gamma and actin plays a role in cell division and apoptosis in astrocytes ［J］. Biochem Biophys Res Commun, 2002, 296(3): 657-663.
［6］Chen XQ, Chen JG, Zhang Y, et al. 14-3-3γ is upregulated by in vitro ischemia and binds to protein kinase raf in primary cultures of astrocytes ［J］. Glia, 2003, 42(4): 315-324.
［7］Gonelli A, Milani D, Rimondi E, et al. Activation of PKCε counteracts maturation and apoptosis of HL-60 myeloid leukemic cells in response to TNF family members ［J］. Eur J Histochem, 2009,53(3):177-182.
［8］Fujii K, Tanabe Y, Kobayashi K,et al. Detection of 14-3-3 protein in the cerebrospinal fluid in mitochondrial encephalopathy with lactic acidosis and stroke-like episodes ［J］. J Neurol Sci, 2005, 239(1): 115-118.
［9］Dong Y, Liu HD, Zhao R, et al. Ischemic activates JNK/c-Jun/AP-1 pathway to up-regulate 14-3-3γ in astrocyte ［J］. J Neurochem, 2009, 109(Sulpp 1): 182-188.
［10］Jang SE, Ryu KR, Park SH, et al. Nobiletin and tangeretin ameliorate scratching behavior in mice by inhibiting the action of histamine and the activation of NF-κB, AP-1 and p38 ［J］. Int Immunopharmacol, 2013,17(3):502-507.
［11］Su QJ, Chen XW, Chen ZB, et al. Involvement of ERK1/2 and p38 MAPK in up-regulation of 14-3-3 protein induced by hydrogen peroxide preconditioning in PC12 cells ［J］. Neurosci Bull, 2008,24(4):244-250.
［12］Fujii K, Tanabe Y, Kobayashi K, et al. Detection of 14-3-3 protein in the cerebrospinal fluid in mitochondrialencephalopathy with lactic acidosis and stroke-like episodes ［J］. J Neurol Sci, 2005,239(1):115-118.
［13］Wang B, Yang H, Liu YC, et al. Isolation of high-affinity peptide antagonists of 14-3-3 proteins by phage display ［J］. Biochemistry, 1999, 38(38): 12499-12504.

[1]	HONG Xiao-min LI San-qiang CUI Qin-yi ZHENG Run-yue YANG Meng-li LUO Ren-li LI Qian-hui. Nuclear factor-κB signaling pathway and gender differences in alcoholic liver fibrosis [J]. Acta Anatomica Sinica, 2024, 55(1): 55-61.
[2]	WEI Xi-xi LI Chao-hong ZHAO Chen-lu TANG Jia-ping LIU Yu-zhen. Characterization of group Ⅰ metabotropic glutamate receptors in rat superior cervical ganglion and their changes following chronic intermittent hypoxia [J]. Acta Anatomica Sinica, 2024, 55(1): 3-9.
[3]	YUAN Lei YANG Zhi-wei YANG Hui LIU Zheng-hai HE Jie WAN Wei. Panax notoginseng saponin relieving the inflammatory pain caused by complete Freund’s adjuvant by inhibiting the activation of astrocytes in mice [J]. Acta Anatomica Sinica, 2024, 55(1): 25-31.
[4]	MA Ting-ting CHEN Jian-hong LIU Ai-cui LI Hai-ning. Role of inhibiting lncRNA TUG1 to down-regulate nucleotide binding oligomerization domain like receptor protein 1 inflammasome in delaying the progression of Alzheimer’s disease [J]. Acta Anatomica Sinica, 2024, 55(1): 32-42.
[5]	ZOU Cheng-gong FENG Hao CHEN Bing TANG Hui SHAO Chuan SUN Mou YANG Rong HE Jia-quan . Research on the dynamic changes of neurological dysfunction and cognitive function impairment in traumatic brain injury [J]. Acta Anatomica Sinica, 2024, 55(1): 43-48.
[6]	LÜ Hai-yan SHEN Xi-ya ZHAO Fu-sheng ZHU Mei . Effects of tricholoma matsutake polysaccharides on 1-methy-4-pehnyl-pyridine ion-induced PC12 cell damage [J]. Acta Anatomica Sinica, 2024, 55(1): 49-54.
[7]	ZHENG Li-ping LIU Xia DU Xiao-hua WU Ya-juan LIU Shan-shan . Expression and distribution of brain-derived neurotrophic factor in different cerebrum regions of yak and cattle [J]. Acta Anatomica Sinica, 2024, 55(1): 10-16.
[8]	HAO Yong-ming GUO Lei ZHOU Cheng-hui PEI Han-jun LI Li ZHAO Zhe . Establishment of a rat model of myocardial hypertrophy by a modified abdominal aortic coarctation method [J]. Acta Anatomica Sinica, 2024, 55(1): 120-124.
[9]	XU Ya-ping YU Yue-xin CHEN Jin-fu WANG Ke-ke GUO Zhikun . Structural distribution and mechanism of elastic fibers and collagen fibers in ventricular interstitium of aged rats [J]. Acta Anatomica Sinica, 2023, 54(6): 716-721.
[10]	WANG Wen-juan DING Yu-tong SU Hao REN Jie SUN Yan-rong WANG Han-fei LU Jia-li ZHANG Lin-qian BAI Yu QIN Li-hua. Changes of Nogo-A expression in hippocampus and striatum of rats under low estrogen condition [J]. Acta Anatomica Sinica, 2023, 54(6): 620-627.
[11]	ZHANG Qin YANG Jun-xia JIANG Qing-song. Effect of fosinopril on angiotensin converting enzyme 2 and vascular endothelial growth factor in diabetic retinopathy mice [J]. Acta Anatomica Sinica, 2023, 54(6): 628-634.
[12]	CHEN Zi-xuan SI Li-na SHU Wei-han ZHANG Xin WEI Chen-yang CHENG Lu-yang YANG Song-he QIAO Yue-bing. Mechanism of melatonin regulation of PI3K/Akt/mTOR signaling in hypothalamus delaying the initiation of puberty in female mice mice [J]. Acta Anatomica Sinica, 2023, 54(6): 644-651.
[13]	LIANG Pan-pan YU Ai-mei DU Jing KOU Wen-hui WANG Huan-huan SONG Ai-xia. Inhibitory effect of sodium ferulate on inflammatory response in migraine rats based on c-Jun N-terminal kinase/p38 mitogen-activated protein kinase signaling pathway [J]. Acta Anatomica Sinica, 2023, 54(6): 652-659.
[14]	XU Ying LIU Bin ZHENG Xing HE Zong-zhao. Role and mechanism of complement C3a receptor in the cognitive impairment of septic rats induced by cecal ligation and perforation [J]. Acta Anatomica Sinica, 2023, 54(6): 668-675.
[15]	LIU Zhe-chuan LI Kun MA Shuai-nan MENG Jia-qi WANG Yan-qin. Effects of liraglutide on inflammation and mitochondrial fusion/division in Parkinson’s disease model of mice induced by paraquat [J]. Acta Anatomica Sinica, 2023, 54(6): 676-681.