Connexin43, the major gap junction protein of astrocytes, is down-regulated in inflamed white matter in an animal model of multiple sclerosis

Abstract

Both multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE), its animal model, involve inflammatory attack on central nervous system (CNS) white matter, leading to demyelination and axonal damage. Changes in astrocytic morphology and function are also prominent features of MS and EAE. Resting astrocytes form a network that is interconnected through gap junctions, composed mainly of connexin43 (Cx43) protein. Although astrocytic gap junctional connectivity is known to be altered in many CNS pathologies, little is known about Cx43 expression in inflammatory demyelinating disease. Therefore, we evaluated the expression of Cx43 in spinal cords of EAE mice compared with healthy controls. Lumbar ventral white matter areas were heavily infiltrated with CD11β-immunoreactive monocytes, and within these infiltrated regions loss of Cx43 immunoreactivity was evident. These regions also showed axonal dystrophy, demonstrated by the abnormally dephosphorylated heavy-chain neurofilament proteins. Astrocytes in these Cx43-depleted lesions were strongly glial fibrillary acidic protein reactive. Significant loss (38%) of Cx43 protein in EAE mouse at the lumbar portion of spinal cords was confirmed by Western blot analysis. Decreased Cx43 transcript level was also observed on cDNA microarray analysis. In addition to changes in Cx43 expression, numerous other genes were altered, including those encoding adhesion and extracellular matrix proteins. Our data support the notion that, in addition to damage of myelinating glia, altered astrocyte connectivity is a prominent feature of inflammatory demyelination.Keywords white matter; inflammation; immunohistochemistry; microarray; experimental autoimmune encephalomyelitis; Cx43Multiple sclerosis (MS) is an immune-cell-mediated inflammatory demyelinating disease of the central nervous system (CNS;Miller and Karpus, 1994;Ewing and Bernard, 1998;Hohlfeld and Wekerle, 2001). MS and experimental autoimmune encephalomyelitis (EAE), an animal model of MS (Honegger and Langemann, 1989;Glabinski et al., 1997;Deloire-Grassin et al., 2000), are characterized by white matter (WM) inflammation accompanied by disturbed central nerve conductance and motor and sensory impairments (Juhler, 1988;Whitaker and Mitchell, 1997 Pathological findings in MS and EAE include infiltration of activated peripheral inflammatory cells into the CNS (Bauer et al., 1995;Owens et al., 1998;Imrich and Harzer, 2001), loss of myelin (Adams and Kubik, 1952;Raine et al., 1984;Whitaker and Mitchell, 1997), axonal damage (Ferguson et al., 1997;Trapp et al., 1998;Kornek et al., 2000;Bitsch et al., 2000;Deloire-Grassin et al., 2000), axonal loss (D'amelio et al., 1990;Landete et al., 2000;Bjartmar et al., 2001), edema (Claudio et al., 1990Levine et al., 1966), and astrocytic activation (Smith et al., 1983;Eng et al., 1992;Rafalowska et al., 1992). Notably, swelling of astrocytes is an early key event that precedes macrophage infiltration to the CNS in EAE (Smith et al., 1984;Eng et al., 1989Eng et al., , ...

Similar Transcriptomic Alterations in Cx43 Knockdown and Knockout Astrocytes

“…As an initial approach to explore this question in the Cx43 null setting, we have compared transcriptomic alterations with respect to wild-type cortical astrocytes (hereafter denoted by WT) in Cx43 null astrocytes (KO) and WT astrocytes acutely treated (48 h) with Cx43 siRNA (SI). Our findings indicate significantly similar transcriptional changes in Cx43 KO astrocytes and those treated acutely with siRNA, from which we infer that such changes may play important roles both in disorders related to hereditary connexin deficiency (for reviews see Gerido and White 2004;Harris 2007;Willecke et al 2002) and in pathological settings in which connexin expression is acutely, chronically, or intermittently altered, such as myocardial infarction (for reviews see Fishman 2005;Severs et al 2006) and multiple sclerosis (Brand-Schieber et al 2005;Roscoe et al 2007). …”

Integrated transcriptomic response to cardiac chronic hypoxia: translation regulators and response to stress in cell survival

“…An indirect confirmation of the "see-saw" model of the transcriptomic recovery came recently when two promyelinating treatments were reported to increase the level of the gap junction protein Cx43 (Roscoe et al 2007), whereas we have shown previously a significant downregulation of Cx43 in AT-EAE mouse spinal cord, an animal model of the human multiple sclerosis (Brand-Schieber et al 2005;Iacobas et al 2008b). In principle, the "seesaw" model can be considered as confirmed by the rescue obtained through overexpressing a positive partner or underexpressing a negative partner of the gene whose deletion caused the disease.…”

Astrocytes in the tempest of multiple sclerosis

“…Once monocytes invade CNS parenchyma, the ongoing inflammation affects astrocytes. In heavily infiltrated areas of CNS massive loss of connexin43 (Cx43) expression is evident in EAE [24]. The loss of Cx43 affects astrocyte connectivity, as networking of astrocytes through gap junctions is dependent on this molecule.…”

“…The loss of Cx43 affects astrocyte connectivity, as networking of astrocytes through gap junctions is dependent on this molecule. Importantly, regions of interrupted astrocytic communication also show axonal dystrophy, demonstrated by the abnormally dephosphorylated heavy-chain neurofilament proteins [24]. Astrocytes in these Cx43-depleted lesions are strongly GFAP-positive, which is a prominent characteristic of reactive astrogliosis [93].…”