促肾上腺皮质激素对N-甲基-D-天冬氨酸诱发幼鼠痉挛模型应激蛋白表达的影响
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摘要
目的分析促肾上腺皮质激素(ACTH)对婴儿痉挛模型应激蛋白表达的影响,为婴儿痉挛症的治疗提供理论依据。方法将60只新生幼鼠随机分为对照组、N-甲基-D-天冬氨酸(NMDA)组和ACTH组,每组20只。幼鼠出生第11天,ACTH组腹腔注射10mg/(kg·d)ACTH,其他两组注射等量的生理盐水。幼鼠出生的第15天,NMDA组和ACTH组腹腔注射15mg/(kg·d)NMDA,对照组注射等量生理盐水。对三组幼鼠痉挛症状进行评分,采用q-PCR法测海马白介素1β(IL-1β)、白介素6(IL-6)和ACTH的mRNA转录水平,Western Blot法测海马促肾上腺皮质激素释放激素(CRH)和促肾上腺皮质激素释放激素1型受体(CRHR1)蛋白表达。结果 NMDA组中幼鼠的痉挛症状评分高于对照组(t=11.236,P<0.001);ACTH组幼鼠的痉挛症状评分低于NMDA组(t=-6.347,P<0.001)。与对照组相比,NMDA组海马IL-1β和IL-6的mRNA转录水平升高(IL-1β:t=6.237,P<0.001;IL-6:t=6.553,P<0.001),ACTH组海马中IL-1β和IL-6的mRNA转录水平低于NMDA组(IL-1β:t=-7.669,P<0.001;IL-6:t=-8.125,P<0.001)。ACTH组海马中ACTH mRNA转录水平高于NMDA组(t=7.758,P<0.001)。与对照组相比,NMDA组海马体CRH和CRHR1蛋白表达上调(CRH:t=7.517,P<0.001;CRHR1:t=7.745,P<0.001),ACTH组海马CRH和CRHR1蛋白表达低于NMDA组(CRH:t=-6.120,P=0.003;CRHR1:t=-6.050,P=0.005)。结论 ACTH可缓解幼鼠的痉挛症状,推测与降低海马体中IL-1β和IL-6的mRNA转录水平,上调ACTH的mRNA转录水平,抑制CRH和CRHR1蛋白表达有关。
Objective To analyze the effect of adrenocorticotropic hormone(ACTH)on the expression of stress proteins in infantile spasm model induced by N-methyl-D-aspartate(NMDA),in order to provide theoretical evidence for the treatment of infantile spasm.Methods Totally 60 neonatal rats were randomly divided into control group,NMDA group and ACTH group,with 20 rats in each group.On the 11 th day of birth,ACTH group was injected with 10 mg/(kg·d)ACTH intraperitoneally,and the other two groups were injected with the same amount of saline.On the 15 th day of birth,15 mg/(kg·d)NMDA was intraperitoneally injected into NMDA group and ACTH group,and the same amount of saline was injected into control group.Spasticity score was used to score the spasticity symptoms of three groups.The mRNA transcription levels of interleukin 1 beta(IL-1β),interleukin 6(IL-6)and ACTH in hippocampus were measured by qPCR.The expressions of corticotropin releasing hormone(CRH)and corticotropin releasing hormone type 1 receptor(CRHR1)in hippocampus were measured by Western Blot method.Results The score of spasticity in NMDA group was significantly higher than that in control group(t=11.236,P<0.001).The score of spasmodic symptoms in ACTH group was lower than that in NMDA group(t=-6.347,P<0.001).Compared with the control group,the transcription levels of IL-1βand IL-6 in the hippocampus of NMDA group increased(IL-1β:t=6.237,P<0.001;IL-6:t=6.553,P<0.001).The transcriptional levels of IL-1βand IL-6 in the hippocampus of ACTH group were lower than those of NMDA group(IL-1β:t=-7.669,P<0.001;IL-6:t=-8.125,P<0.001).The transcription level of ACTH mRNA in hippocampus of group ACTH was higher than that of NMDA group(t=7.758,P<0.001).Compared with the control group,the expression levels of CRH and CRHR1 protein in hippocampus of NMDA group were up-regulated(CRH:t=7.517,P<0.001;CRHR1:t=7.745,P<0.001).While the expression of CRH and CRHR1 protein in ACTH group was lower than that in NMDA group(CRH:t=-6.120,P=0.003;CRHR 1:t=-6.050,P=0.005). Conclusions ACTH could relieve spasm symptom induced by NMDA in neonatal rats.It is speculated that ACTH may help decrease the mRNA expressions of IL-1βand IL-6 in the hippocampal tissues of neonatal rats,up-regulate the mRNA expression of ACTH,and inhibit the protein expressions of CRH and CRHR1.
Objective To analyze the effect of adrenocorticotropic hormone(ACTH)on the expression of stress proteins in infantile spasm model induced by N-methyl-D-aspartate(NMDA),in order to provide theoretical evidence for the treatment of infantile spasm.Methods Totally 60 neonatal rats were randomly divided into control group,NMDA group and ACTH group,with 20 rats in each group.On the 11 th day of birth,ACTH group was injected with 10 mg/(kg·d)ACTH intraperitoneally,and the other two groups were injected with the same amount of saline.On the 15 th day of birth,15 mg/(kg·d)NMDA was intraperitoneally injected into NMDA group and ACTH group,and the same amount of saline was injected into control group.Spasticity score was used to score the spasticity symptoms of three groups.The mRNA transcription levels of interleukin 1 beta(IL-1β),interleukin 6(IL-6)and ACTH in hippocampus were measured by qPCR.The expressions of corticotropin releasing hormone(CRH)and corticotropin releasing hormone type 1 receptor(CRHR1)in hippocampus were measured by Western Blot method.Results The score of spasticity in NMDA group was significantly higher than that in control group(t=11.236,P<0.001).The score of spasmodic symptoms in ACTH group was lower than that in NMDA group(t=-6.347,P<0.001).Compared with the control group,the transcription levels of IL-1βand IL-6 in the hippocampus of NMDA group increased(IL-1β:t=6.237,P<0.001;IL-6:t=6.553,P<0.001).The transcriptional levels of IL-1βand IL-6 in the hippocampus of ACTH group were lower than those of NMDA group(IL-1β:t=-7.669,P<0.001;IL-6:t=-8.125,P<0.001).The transcription level of ACTH mRNA in hippocampus of group ACTH was higher than that of NMDA group(t=7.758,P<0.001).Compared with the control group,the expression levels of CRH and CRHR1 protein in hippocampus of NMDA group were up-regulated(CRH:t=7.517,P<0.001;CRHR1:t=7.745,P<0.001).While the expression of CRH and CRHR1 protein in ACTH group was lower than that in NMDA group(CRH:t=-6.120,P=0.003;CRHR 1:t=-6.050,P=0.005). Conclusions ACTH could relieve spasm symptom induced by NMDA in neonatal rats.It is speculated that ACTH may help decrease the mRNA expressions of IL-1βand IL-6 in the hippocampal tissues of neonatal rats,up-regulate the mRNA expression of ACTH,and inhibit the protein expressions of CRH and CRHR1.
引文
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[16] Oztas B,Sahin D,Kir H,et al.The effect of leptin,ghrelin,and neuropeptide-Y on serum Tnf-Α,IL-1β,IL-6,Fgf-2,galanin levels and oxidative stress in an experimental generalized convulsive seizure model[J].Neuropeptides,2017,61(2):31-37.
[17] Garcia I,Bhullar PK,Tepe B,et al.Local corticotropin releasing hormone(CRH)signals to its receptor CRHR1during postnatal development of the mouse olfactory bulb[J].Brain Struct Funct,2016,221(1):1-20.
[18] Donner NC,Siebler PH,Johnson DT,et al.Serotonergic systems in the balance:CRHR1and CRHR2differentially control stress-induced serotonin synthesis[J].Psychoneuroendocrinology,2016,63:178-190.
[19] Jing Z,Chen Z,Tian J,et al.miR-34battenuates traumainduced anxiety-like behavior by targeting CRHR1[J].Int J Mol Med,2017,40(1):90-100.
[20] Inda C,Pa DSC,Bonfiglio JJ,et al.Different cAMP sources are critically involved in G protein-coupled receptor CRHR1signaling[J].J Cell Biol,2016,214(2):181-195.
[2] Su DJ,Lu JF,Lin LJ,et al.SCN2A mutation in an infant presenting with migrating focal seizures and infantile spasm responsive to a ketogenic diet[J].Brain Dev,2018,40(8):724-727.
[3] Depondt C,Heinzen EL,Goldstein DB.Reply to two patients with TNK2 mutations and late-onset infantile spasm[J].Ann Neurol,2017,81(1):161.
[4] Scantlebury MH,Chun KC,Ma SC,et al.Adrenocorticotropic hormone protects learning and memory function in epileptic kcna1-null mice[J].Neurosci Lett,2017,645:14-18.
[5] Shachor-Meyouhas Y,Ravid S,Hanna S,et al.Legionella pneumophila pneumonia in two infants treated with adrenocorticotropic hormone[J].J Pediatr,2017,186(7):186-188.
[6] Hayashi Y,Yoshinaga H,Akiyama T,et al.Predictive factors for relapse of epileptic spasms after adrenocorticotropic hormone therapy in West syndrome[J].Brain Dev,2016,38(1):32-39.
[7] Chen W,Sheng J,Peng G,et al.Early stage alterations of catecholamine and adrenocorticotropic hormone levels in posttraumatic acute diffuse brain swelling[J].Brain Res Bull,2017,130(8):47-52.
[8] Song JM,Hahn J,Kim SH,et al.Efficacy of treatments for infantile spasms:a systematic review[J].Clin Neuropharmacol,2017,40(2):63-84.
[9] Auvin S,Cilio MR,Vezzani A.Current understanding and neurobiology of epileptic encephalopathies[J].Neurobiol Dis,2016,92(Pt A):72-89.
[10] Kurata H,Terashima H,Nakashima M,et al.Characterization of SPATA5-related encephalopathy in early childhood[J].Clin Genet,2016,90(5):437-444.
[11] Sahu JK,Vaddi VK,Negi S.Review of west syndrome:concerns on optimum dose of adrenocorticotrophic hormone[J].Clin Drug Investig,2018,38(2):1-2.
[12] Mcdonald WC,Banerji N,Mcdonald KN,et al.Steroidogenic factor 1,pit-1,and adrenocorticotropic hormone:a rational starting place for the immuno-histochemical characterization of pituitary adenoma[J].Arch Pathol Lab Med,2017,141(1):104-112.
[13] Shi C,Wang F,Tong A,et al.NFKB2mutation in common variable immunodeficiency and isolated adrenocorticotropic hormone deficiency:a case report and review of literature[J].Medicine,2016,95(40):e5081.
[14] Nhr AC,Shehata MA,Hauser AS,et al.The orphan G protein-coupled receptor GPR139is activated by the peptides:adrenocorticotropic hormone(ACTH),α-,andβ-melanocyte stimulating hormone(α-MSH,andβ-MSH),and the conserved core motif HFRW[J].Neurochem Int,2016,102(1):105-113.
[15] Slaats J,Ten Oever J,van de Veerdonk FL,et al.IL-1β/IL-6/CRP and IL-18/ferritin:distinct inflammatory programs in Infections[J]. PLoS Pathog,2016,12(12):e1005973.
[16] Oztas B,Sahin D,Kir H,et al.The effect of leptin,ghrelin,and neuropeptide-Y on serum Tnf-Α,IL-1β,IL-6,Fgf-2,galanin levels and oxidative stress in an experimental generalized convulsive seizure model[J].Neuropeptides,2017,61(2):31-37.
[17] Garcia I,Bhullar PK,Tepe B,et al.Local corticotropin releasing hormone(CRH)signals to its receptor CRHR1during postnatal development of the mouse olfactory bulb[J].Brain Struct Funct,2016,221(1):1-20.
[18] Donner NC,Siebler PH,Johnson DT,et al.Serotonergic systems in the balance:CRHR1and CRHR2differentially control stress-induced serotonin synthesis[J].Psychoneuroendocrinology,2016,63:178-190.
[19] Jing Z,Chen Z,Tian J,et al.miR-34battenuates traumainduced anxiety-like behavior by targeting CRHR1[J].Int J Mol Med,2017,40(1):90-100.
[20] Inda C,Pa DSC,Bonfiglio JJ,et al.Different cAMP sources are critically involved in G protein-coupled receptor CRHR1signaling[J].J Cell Biol,2016,214(2):181-195.
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