Deletion of <i>Prmt5</i> in Cerebral Endothelial Cells Leads to Cerebrovascular Disease and Astrocyte Activation

doi:10.13523/j.cb.2112026

China Biotechnology

2022, Vol. 42

Issue (4): 1-8 DOI: 10.13523/j.cb.2112026

Deletion of Prmt5 in Cerebral Endothelial Cells Leads to Cerebrovascular Disease and Astrocyte Activation

NING Hui-min^1,²,ZHANG Yi-zhe²,HAN Yu-ying²,ZHANG Chong²,SONG Xiao-peng²,LIANG Shuang²,YANG Xiao^1,^2,^**(

),WANG Jun^2,^**(

)

1 School of Basic Medicine, Qingdao University, Qingdao 266071, China
2 State Key Laboratory of Proteomics, Beijing National Center for Protein Sciences (Beijing), Beijing Institute of Lifeomics, Beijing 102206, China

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Abstract

Objective: To study the role of protein arginine methyltransferase 5 (Prmt5) in cerebral vascular development and homeostasis maintenance in mice, and to investigate the effect of Prmt5 specific knockout on the central nervous system. Methods: We crossed Prmt5^fl/fl mice with SP-A Cre transgenic mice that express Cre recombinase in cerebrovascular endothelial to generate cerebrovascular endothelial cell-specific Prmt5 knockout mice. H-E staining and immunostaining were performed to observe the vascular structures of control and Prmt5^fl/fl mutant mice. Laser speckle contrast imaging was used to detect cerebral blood flow in control and mutant mice. Sulfo-NHS-Biotin was intraperitoneally injected into control and mutant mice to examine the blood brain barrier(BBB) integrity. The expression levels of astrocyte glial fibrillary acidic protein (GFAP), S100 calcium-binding protein β (S100β), complement C3 (C3), C1q, tumor necrosis factor alpha(TNF-α) and Interleukin-1 beta(IL-1β) were detected by immunofluorescence and Western blot to evaluate the activation level of astrocytes in cortex, thalamus and cerebellum of knockout mice and control mice. Furthermore, activators of astrocytes, such as C1q, TNF-α, IL-1β and other cytokines, were also detected by real-time PCR. Results: We found that cerebrovascular endothelial cell-specific Prmt5 knockout mice exhibited aberrant cerebrovascular structure, and increased the number of reactive astrocytes. The expression levels of TNF-α and IL-1β in the whole brain, as well as the C1q, TNF-α and IL-1β, were all increased in Prmt5^fl/fl mutant mice. Conclusion: Prmt5 plays an essential role in the maintenance of cerebrovascular homeostasis, suggesting that it might act as a potential therapeutic target for cerebrovascular diseases.

Key words： Cerebrovascular endothelial cell Prmt5 Astrocyte

Received: 08 December 2021 Published: 05 May 2022

ZTFLH:

Q819

Corresponding Authors: Xiao YANG,Jun WANG E-mail: yangx@bmi.ac.cn;wangjun1@bmi.ac.cn

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	Articles by authors
	NING Hui-min
	ZHANGYi-zhe
	HAN Yu-ying
	ZHANG Chong
	SONG Xiao-peng
	LIANG Shuang
	YANG Xiao
	WANG Jun

Cite this article:

NING Hui-min, ZHANGYi-zhe, HAN Yu-ying, ZHANG Chong, SONG Xiao-peng, LIANG Shuang, YANG Xiao, WANG Jun . Deletion of Prmt5 in Cerebral Endothelial Cells Leads to Cerebrovascular Disease and Astrocyte Activation. China Biotechnology, 2022, 42(4): 1-8.

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https://manu60.magtech.com.cn/biotech/10.13523/j.cb.2112026 OR https://manu60.magtech.com.cn/biotech/Y2022/V42/I4/1

Table 1 Realtime-PCR primers

Fig.1 Cerebrovascular-specific deletion of Prmt5 induces vascular lesions (a), (b) Real-time PCR (a) and Western blot (b) analysis of PRMT5 expression in primary isolated brain ECs derived from Prmt^5fl/⁺ and Prmt5^fl/fl mice. n=3, *** P<0.001 (c) H-E staining of brain tissue of Prmt5^fl/⁺ and Prmt5^fl/fl mice. Scale bar, 50 μm (d) Immunofluorescence staining of Endomucin, a marker of cerebrovascular, in brains of Prmt5^fl/⁺ and Prmt5^fl/fl mice. Scale bar, 50 μm (e) Immunostaining analyses of brain sections after Sulfo-NHS-Biotin injections. Scale bar, 500 μm (f) Dynamic monitoring of the changes of cerebral blood flow in Prmt5^fl/⁺ and Prmt5^fl/fl mice. The corresponding quantitative data is shown in (g). n = 3, * P< 0.05

Fig.2 Brain endothelial cell-specific deletion of Prmt5 leads to increased release of inflammatory cytokines (a) Real-time PCR was used to detect the changes of inflammatory factors secreted by endothelium. n=3, * P<0.05 or ** P<0.01 (b) Real-time PCR was used to detect the changes of inflammatory factors in the whole brain. n=3(4,5), ** P< 0.01 or *** P< 0.001

Fig.3 Reactive astrocyte activation in brains of Prmt5 knockout mice (a) Immunostaining analyses of brain sections for GFAP. Scale bar, 50 μm (b) Quantitative analysis of area occupied by GFAP⁺ cells. n=3, ** P< 0.01 (c) Western blot analyses of levels of C3, GFAP and GAPDH in brains of Prmt5^fl/⁺ and Prmt5^fl/fl mice (d),(e) Quantitative results of Western blot. n=3 or n=4, * P< 0.05 (f) Immunostaining analyses of brain sections for GFAP (green), S100β (syan), and C3 (red). Scale bar, 50 μm


[1]	Sweeney M D, Zhao Z, Montagne A, et al. Blood-brain barrier: from physiology to disease and back. Physiological Reviews, 2019, 99(1): 21-78. doi: 10.1152/physrev.00050.2017

[2]	Bernstein D L, Gajghate S, Reichenbach N L, et al. Let-7g counteracts endothelial dysfunction and ameliorating neurological functions in mouse ischemia/reperfusion stroke model. Brain, Behavior, and Immunity, 2020, 87: 543-555. doi: S0889-1591(19)31341-8 pmid: 32017988

[3]	Zhao Z, Nelson A R, Betsholtz C, et al. Establishment and dysfunction of the blood-brain barrier. Cell, 2015, 163(5): 1064-1078. doi: 10.1016/j.cell.2015.10.067

[4]	Blanc R S, Richard S. Arginine methylation: the coming of age. Molecular Cell, 2017, 65(1): 8-24. doi: 10.1016/j.molcel.2016.11.003

[5]	Hamard P J, Santiago G E, Liu F, et al. PRMT5 regulates DNA repair by controlling the alternative splicing of histone-modifying enzymes. Cell Reports, 2018, 24(10): 2643-2657. doi: 10.1016/j.celrep.2018.08.002

[6]	Webb L M, Sengupta S, Edell C, et al. Protein arginine methyltransferase 5 promotes cholesterol biosynthesis-mediated Th17 responses and autoimmunity. The Journal of Clinical Investigation, 2020, 130(4): 1683-1698. doi: 10.1172/JCI131254

[7]	Zhang T, Günther S, Looso M, et al. Prmt5 is a regulator of muscle stem cell expansion in adult mice. Nature Communications, 2015, 6: 7140. doi: 10.1038/ncomms8140 pmid: 26028225

[8]	Liu M, Yao B, Gui T, et al. PRMT5-dependent transcriptional repression of c-Myc target genes promotes gastric cancer progression. Theranostics, 2020, 10(10): 4437-4452. doi: 10.7150/thno.42047

[9]	Meng F W, Shi L, Cheng X, et al. Surfactant protein A promoter directs the expression of Cre recombinase in brain microvascular endothelial cells of transgenic mice. Matrix Biology, 2007, 26(1): 54-57. doi: 10.1016/j.matbio.2006.09.003

[10]	Li Z H, Xu J P, Song Y, et al. PRMT5 prevents dilated cardiomyopathy via suppression of protein O-GlcNAcylation. Circulation Research, 2021, 129(9): 857-871. doi: 10.1161/CIRCRESAHA.121.319456

[11]	Grammas P, Ovase R. Inflammatory factors are elevated in brain microvessels in Alzheimer’s disease. Neurobiology of Aging, 2001, 22(6): 837-842. pmid: 11754990

[12]	Grammas P, Samany P G, Thirumangalakudi L. Thrombin and inflammatory proteins are elevated in Alzheimer’s disease microvessels: implications for disease pathogenesis. Journal of Alzheimer’s Disease: JAD, 2006, 9(1): 51-58.

[13]	Liddelow S A, Guttenplan K A, Clarke L E, et al. Neurotoxic reactive astrocytes are induced by activated microglia. Nature, 2017, 541(7638): 481-487. doi: 10.1038/nature21029

[14]	King A, Szekely B, Calapkulu E, et al. The increased densities, but different distributions, of both C3 and S100A10 immunopositive astrocyte-like cells in Alzheimer’s disease brains suggest possible roles for both A1 and A2 astrocytes in the disease pathogenesis. Brain Sciences, 2020, 10(8): 503. doi: 10.3390/brainsci10080503

[15]	Grammas P, Martinez J, Sanchez A, et al. A new paradigm for the treatment of Alzheimer’s disease: targeting vascular activation. Journal of Alzheimer’s Disease: JAD, 2014, 40(3): 619-630.

[16]	Zamanian J L, Xu L, Foo L C, et al. Genomic analysis of reactive astrogliosis. The Journal of Neuroscience, 2012, 32(18): 6391-6410. doi: 10.1523/JNEUROSCI.6221-11.2012

[17]	Stogsdill J A, Ramirez J, Liu D, et al. Astrocytic neuroligins control astrocyte morphogenesis and synaptogenesis. Nature, 2017, 551(7679): 192-197. doi: 10.1038/nature24638

[18]	Jaudon F, Chiacchiaretta M, Albini M, et al. Kidins220/arms controls astrocyte calcium signaling and neuron-astrocyte communication. Cell Death and Differentiation, 2020, 27(5): 1505-1519. doi: 10.1038/s41418-019-0431-5 pmid: 31624352

[19]	Sofroniew M V, Vinters H V. Astrocytes: biology and pathology. Acta Neuropathologica, 2010, 119(1): 7-35. doi: 10.1007/s00401-009-0619-8 pmid: 20012068

[20]	Yang J H, Vitery M D C, Chen J N, et al. Glutamate-releasing SWELL1 channel in astrocytes modulates synaptic transmission and promotes brain damage in stroke. Neuron, 2019, 102(4): 813-827.e6. doi: 10.1016/j.neuron.2019.03.029

[21]	Díaz-García C M, Mongeon R, Lahmann C, et al. Neuronal stimulation triggers neuronal glycolysis and not lactate uptake. Cell Metabolism, 2017, 26(2): 361-374.e4. doi: S1550-4131(17)30421-7 pmid: 28768175

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