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Increased Expression of Slit2 and its Robo Receptors During Astroglial Scar Formation After Transient Focal Cerebral Ischemia in Rats

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Abstract

Slit2, a secreted glycoprotein, has recently been implicated in the post-ischemic astroglial reaction. The objective of this study was to investigate the temporal changes and cellular localization of Slit2 and its receptors, Robo1, Robo2, and Robo4, in a rat transient focal ischemia model induced by middle cerebral artery occlusion. We used double- and triple-immunolabeling to determine the cell-specific changes in Slit2 and its receptors during a 10-week post-ischemia period. The expression profiles of Slit2 and the Robo receptors shared overlapping expression patterns in sham-operated and ischemic striatum. Constitutive expression of Slit2 and Robo receptors was observed in striatal neurons with weak intensity, whereas in rats reperfused after ischemic insults, these immunoreactivities were increased in reactive astrocytes. Astroglial induction of Slit2 and Robo in the peri-infarct region was distinct on days 7–14 after reperfusion and thereafter increased progressively throughout the 10-week experimental period. Slit2 and Robo were prominently expressed in the perinuclear cytoplasm and main processes of reactive astrocytes forming the astroglial scar. This observation was confirmed by quantification of the mean fluorescence intensity of Slit2 and Robo receptors over reactive astrocytes localized at the edge of the infarct area. However, activated microglia/macrophages in the peri-infarct area were devoid of any specific labeling for Slit2 and Robo. Thus, our data revealed a selective and sustained induction of Slit2 and Robo in astrocytes localized throughout the astroglial scar after ischemic stroke, suggesting that Slit2/Robo signaling participates in glial scar formation and brain remodeling following ischemic injury.

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Abbreviations

Robo:

Roundabout

MCAO:

Middle cerebral artery occlusion

GFAP:

Glial fibrillary acidic protein

Iba1:

Ionized calcium-binding adaptor molecule 1

NeuN:

Neuronal nuclear antigen

CNS:

Central nervous system

PB:

Phosphate buffer

FJB:

Fluoro-Jade B

DARPP-32:

Dopamine- and cyclic AMP-regulated phosphoprotein, apparent molecular weight 32 kDa

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Acknowledgments

This research was supported by the Basic Science Research Program through the National Research Foundation of Korea, funded by the Ministry of Science, ICT, and future Planning (NRF-2014R1A2A1A11050246).

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Authors and Affiliations

  1. Department of Anatomy, Catholic Neuroscience Institute, College of Medicine, The Catholic University of Korea, 505 Banpo-dong, Socho-gu, Seoul, 137-701, South Korea

    Xuyan Jin, Yoo-Jin Shin, Tae-Ryong Riew, Jeong-Heon Choi & Mun-Yong Lee

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  1. Xuyan Jin
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Correspondence to Mun-Yong Lee.

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All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All experimental procedures were conducted in accordance with the Laboratory Animal Welfare Act, the Guide for the Care and Use of Laboratory Animals, and the Guidelines and Policies for Rodent Survival Surgery, and were approved by the IACUC (Institutional Animal Care and Use Committee) at the College of Medicine, The Catholic University of Korea (Approval Number: CUMC-2015-0213-01). All efforts were made to minimize animal suffering and to reduce the number of animals used.

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Jin, X., Shin, YJ., Riew, TR. et al. Increased Expression of Slit2 and its Robo Receptors During Astroglial Scar Formation After Transient Focal Cerebral Ischemia in Rats. Neurochem Res 41, 3373–3385 (2016). https://doi.org/10.1007/s11064-016-2072-0

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