[1]曹晓娟,李 强,范奎奎,等.小鼠脑干3β羟基类固醇脱氢酶1免疫阳性神经元信号检测方法的优化[J].中国预防兽医学报,2019,(12):1280-1284.[doi:0.3969/j.issn.1008-0589.201905034]
 CAO Xiao-juan,LI Qiang,FAN Kui-kui,et al.dehydrogenase 1 immunoreactive neurons in mouse brain stem[J].Chinese journal of preventive veterinary medicine,2019,(12):1280-1284.[doi:0.3969/j.issn.1008-0589.201905034]
点击复制

小鼠脑干3β羟基类固醇脱氢酶1免疫阳性神经元信号检测方法的优化()
分享到:

《中国预防兽医学报》[ISSN:1008-0589/CN:23-1417/S]

卷:
期数:
2019年12
页码:
1280-1284
栏目:
研究简报
出版日期:
2020-01-25

文章信息/Info

Title:
dehydrogenase 1 immunoreactive neurons in mouse brain stem
文章编号:
1008-0589(2019)12-1280-05
作者:
 

曹晓娟1李 强2范奎奎3潘 登3刘昊东3王 昆2海日汗1杜晨光13*

 (1. 内蒙古农业大学 职业技术学院,内蒙古 包头 014109;2. 内蒙古农业大学 兽医学院,内蒙古 呼和浩特 010018;
3. 河北省农林科学院粮油作物研究所,河北 石家庄 050000)
Author(s):
 

CAO Xiao-juan1 LI Qiang2 FAN Kui-kui3 PAN Deng3 LIU Hao-dong3 WANG Kun2 HAI Ri-han1 DU Chen-guang13*

 (1. Vocational and Technical College, Inner Mongolia Agricultural University, Baotou 014109, China;
2. College of veterinary medicine, Inner Mongolia Agricultural University, Hohhot 010018, China;
3. Institute of Grain and Oil Crops, Hebei Academy of Agricultural and Forestry Sciences,Shijiazhuang 050000, China )
Keywords:
 immunofluorescence  3β hydroxyl steroid dehydrogenase 1  mouse brainstem
分类号:
S852.4
DOI:
0.3969/j.issn.1008-0589.201905034
文献标志码:
A
摘要:
 
Abstract:
3β-hydroxysteroid dehydrogenase-1 (3β-HSD1), the key regulator of steroid synthesis, is the key regulator of steroid hormone synthesis pathway, which can ultimately affect the secretion level of androgen (testosterone). However, the distribution of 3β-HSD1 in the brain has not been reported, which limits the understanding of energy metabolism, reproduction, endocrine and other functions in the central nervous system. In order to optimize the method of detecting the distribution of brain stem 3β-HSD1 in mice brain neurons, different fixation methods  such as 4% paraformaldehyde, 20% methanol+0.2% acetone, or 4% paraformaldehyde+10 mmol/L sodium citrate, were used to  preperfuse the mice,  and frozen sections were then prepared, optimized and pretreated before sealing [1% TritonX-100+0.4% SDS perforation, 10 mmol/L citric acid]. The distribution of 3β-HSD1 in the brain tissue of mice and the influence of different fixations on the neuron positive signal were detected by immunohistochemical method. Meanwhile, western blot was used to verify the results of the immunohistochemical test. The results showed that 3β-HSD1 was mainly expressed in the major nuclei of steroid hormones in the posterior brain stem of mice, such as RPA, GRN and PB. No 3β-HSD1 positive fluorescence signal was observed in 4% paraformaldehyde fixed mice; 3β-HSD1 positive signal distribution was observed in 20% methanol+0.2% acetone fixed mice, but the fluorescence signal was vague; The mice in 4% paraformaldehyde+10 mmol/L sodium citrate were able to obtain 3β-HSD1 positive fluorescent signals with clear staining results and full neuron shape in the above nuclei in the brain stem. Western blot also confirmed the specific expression of 3β-HSD1 protein in the mouse brain stem. In conclusion, when using immunohistochemistry to detect the localization of 3β- HSD1 in the mouse brain, a full 3β-HSD1 positive fluorescence signal can be obtained in the RPA, GRN and PB nuclei of the brain stem of mice by the 4% paraformaldehyde+10 mmol/L sodium citrate fixation. The optimization of this method will provide a technical means for further study of the distribution of 3β-HSD1 in mice brain and the effect of 3β-HSD1 on the synthesis of steroids.

参考文献/References:

[1]Rossier B C, Baker M E, Studer R A. Epithelial sodium transport and its control by aldosterone: the story of our internal environment revisited [J]. Physiol Rev, 2015, 95(1): 297-340.
[2]Moors M, Williams T A, Deinum J, et al. Steroid hormone production in patients with aldosterone producing adenomas [J]. Horm Metab Res, 2015, 47(13): 967-972.
[3]Garg R, Adler G K. Aldosterone and the mineralocorticoid receptor: Risk factors for cardiometabolic disorders [J]. Curr Hypertens Rep, 2015, 17(7): 1-8.
[4]Rosas-Arellano A, Villalobos-González J B, Palma-Tirado L, et al. A simple solution for antibody signal enhancement in immunofluorescence and triple immunogold assays [J]. Histochem Cell Biol, 2016, 146(4): 421-430.
[5]姜煜东,李文思,余翀,等. 小鼠心室肌脱细胞化细胞外基质薄片的制备和评价[J]. 生理学报,2014,66(6):709- 717.
[6]Gu Ai-hua, Ji Gui-xiang, Long Yan, et al. Assessment of an association between an aryl hydrocarbon receptor Gene (AHR) polymorphism and risk o f male infertility [J]. J Toxicol Sci, 2011, 122(2): 415-421.
[7]Targoff I N, Johnson A E, Miller F W. Antibody to signal recognition particle in polymyositis [J]. Arthritis Rheum, 1990, 33(9): 1361-1370.
[8]Büttnerennever J. The rat brain in stereotaxic coordinates, 3rd edn [J]. J Anat, 1997, 191(Pt 2): 315.
[9]范奎奎,李强,杨文雅,等. Western blot鉴定背肩胛棕色脂肪黑皮质素4型受体的研究[J]. 畜牧兽医学报,2018, 49(4):841-846.
[10]Li Xiao-jing, Fan Kui-kui, Li Qiang, et al. Melanocortin 4 receptor-mediated effects of amylin on thermogenesis and regulation of food intake [J]. Diabetes Metab Res Rev, 2019: e3149.
[11]郭玲,董兵卫,黄明. 卵巢肿瘤快速冰冻切片与石蜡切片病理诊断的对比分析[J]. 实用癌症杂志,2017,32(12):2077-2079.
[12]金云涛,郭莹莹,韩勇权,等. 固定动物组织冰冻切片技术的探讨[J]. 科学技术创新,2015,(22):114.
[13]Hofmann A, Peitzsch M, Brunssen C, et al. Elevated steroid hormone production in the db/db mouse model of obesity and type 2 Diabetes [J]. Horm Metab Res, 2016, 49(01): 43-49.
[14]孙海梅,尚宏伟,张立新,等. 不同固定液对大鼠脾石蜡切片HE染色标本的影响[J]. 中国医学装备,2011,08(10):48-50.
[15]蔡翠珠,张红,贾晓云. 胰腺冰冻切片免疫组化中组织的固定方法比较[J]. 中国现代医药杂志,2008,10(7):78-79.
[16]Hinman J D, Rasband M N, Carmichael S T. Remodeling of the axon initial segment after focal cortical and white matter stroke[J]. Stroke, 2012, 44(1): 182-189.
[17]张政,王露. 类固醇激素反应性慢性淋巴细胞性炎症伴脑桥-小脑血管周围强化征临床特点及影像学分析[J]. 海峡药学,2014,(11):206-207.
[18]顾桂宝,鞠躬. 大鼠终纹床核向结合臂旁核的投射 Ⅲ.神经降压肽、促肾上腺皮质激素释放因子肽能通路研究[J]. 神经解剖学杂志,1993,(1):118-122.
                            (本文编辑:李 娜;英文编辑:王玉娥

更新日期/Last Update: 2020-01-19