Contents lists available at ScienceDirect Experimental Neurology journal homepage: www.elsevier.com/locate/yexnr Research paper rhIGF-1 reduces the permeability of the blood-brain barrier following intracerebral hemorrhage in mice Derek Sunil Nowrangi a , Devin McBride a , Anatol Manaenko a , Brandon Dixon a , Jiping Tang a , John H. Zhang a,b, ⁎ a Department of Physiology, Loma Linda University School of Medicine, Risley Hall, Room 223, Loma Linda, CA 92354, USA b Department of Neurosurgery, Loma Linda University School of Medicine, Loma Linda, CA, USA ARTICLE INFO Keywords: Intracerebral hemorrhage Blood-brain barrier Insulin-like growth factor 1 Insulin-like growth factor 1 receptor Collagenase Blood-injection Intranasal ABSTRACT Disruption of the blood-brain barrier results in the formation of edema and contributes to the loss of neurological function following intracerebral hemorrhage (ICH). This study examined insulin-like growth factor-1 (IGF-1) as a treatment and its mechanism of action for protecting the blood-brain barrier after ICH in mice. 171 Male CD-1 mice were subjected to ICH via collagenase or autologous blood. A dose study for recombinant human IGF-1 (rhIGF-1) was performed. Brain water content and behavioral deficits were evaluated at 24 and 72 h after the surgery, and Evans blue extravasation and hemoglobin assay were conducted at 24 h. Western blotting was performed for the mechanism study and interventions were used targeting the IGF-1R/GSK3β/MEKK1 pathway. rhIGF-1 reduced edema and blood-brain barrier permeability, and improved neurobehavior outcomes. Western blots showed that rhIGF-1 reduced p-GSK3β and MEKK1 expression, thereby increasing occludin and claudin-5 expression. Inhibition and knockdown of IGF-1R reversed the therapeutic benefits of rhIGF-1. The findings within suggest that stimulation of the IGF-1R is a therapeutic target for ICH which may lead to improved neurofunctional and blood-brain barrier protection. 1. Introduction Intracerebral hemorrhage (ICH) is one of the most dangerous forms of stroke that currently has no effective treatments. Among the several pathological sequelae of ICH, the hematoma-induced breakdown of the blood-brain barrier (BBB) is one of the major concerns which plays a critical role in the pathophysiology of the disease (Selim and Sheth, 2015; Xi et al., 2002). Loss of endothelial tight junction proteins con- tribute to increased permeability of the BBB leading to the formation of vasogenic edema and a deterioration of neurological functions (Liu et al., 2012; Gebel Jr et al., 2002). Previous studies have demonstrated that the protection and recovery of tight junction proteins decreases the severity of the ICH injury with improved neurological outcomes (Krafft et al., 2013). Insulin-like growth factor 1 (IGF-1) is highly selective for the IGF-1 receptor (IGF-1R), which is expressed on several cell types in the brain including neurons, astrocytes, and endothelial cells (Bake et al., 2016; Liu et al., 1994; Shemer et al., 1987). Canonically, stimulation of the IGF-1R causes of the phosphatidylinositol 3-kinase (PI3K)-Akt pathway thus attenuating glycogen synthase kinase-3β (GSK3β) activation (Hetman et al., 2000; Allen et al., 2005). Previously, our lab demon- strated that preventing activation of glycogen synthase kinase-3β (GSK3β) reduces edema and damage to the BBB following injury through protecting tight junction proteins (Krafft et al., 2013). GSK3β physically binds to and activates mitogen-activated protein kinase (MAPK) kinase (MEK) kinase 1 (MEKK1), an important stimulator of the c-Jun/JNK pathway, which has been shown to increase MMP9 pro- duction (Kim et al., 2003; Feiler et al., 2011). GSK3β has also been shown to reduce expression of the tight junction proteins occludin and claudin-5, indicating an increase in the permeability of the barrier (Nitta et al., 2003). IGF-1 has been previously described to have protective effects in ischemic brain injury models, however the role of this growth factor remains unexplored in ICH. In addition, the role and mechanism by which IGF-1 and IGF-1R protects the BBB is not completely understood. We hypothesize that IGF-1 will decrease the permeability of the BBB and reduce edema in an ICH mouse model via IGF-1R stimulation, re- ducing GSK3β/MEKK1 activation, to prevent the loss of tight junction protein expression. https://doi.org/10.1016/j.expneurol.2018.11.009 Received 16 July 2018; Received in revised form 26 November 2018; Accepted 29 November 2018 ⁎ Corresponding author at: Department of Physiology, Loma Linda University School of Medicine, Risley Hall, Room 223, Loma Linda, CA 92354, USA. E-mail addresses: dnowrangi@llu.edu (D.S. Nowrangi), dmcbride@llu.edu (D. McBride), bjdixon@llu.edu (B. Dixon), jtang@llu.edu (J. Tang), johnzhang3910@yahoo.com (J.H. Zhang). Experimental Neurology 312 (2019) 72–81 Available online 30 November 2018 0014-4886/ © 2018 Published by Elsevier Inc. T