Elsevier

Neurobiology of Disease

Volume 45, Issue 1, January 2012, Pages 438-449
Neurobiology of Disease

Age-dependent neurovascular abnormalities and altered microglial morphology in the YAC128 mouse model of Huntington disease

https://doi.org/10.1016/j.nbd.2011.09.003Get rights and content

Abstract

Central nervous system (CNS) inflammatory processes including microglial activation have been implicated in the pathogenesis of neurodegenerative diseases such as Huntington Disease (HD). We report age-dependent changes in striatal microglial morphology and vasculature in the YAC128 mouse model of HD. Decreases in microglial ramification along with a decrease in vessel diameter and increased vessel density and length suggest the presence of microgliosis and proangiogenic activity in YAC128 mice. Our hypothesis for this study was that the changes in microglial morphology and perturbations in vasculature may be involved in the pathogenesis of HD and that peripheral challenge with the bacterial endotoxin, lipopolysaccharide (LPS), will exacerbate these microglial and vascular changes as well as the HD phenotype in YAC128 mice at 12 months. Chronic peripheral LPS (1 mg/kg) potentiated microglial activation indicated by an increase in microglial cell body size and retraction of processes. This potentiation in microglial activation with chronic peripheral LPS challenge was paralleled with vascular remodeling including dilatation, increased vessel wall thickness, increased BBB permeability and fibrinogen deposition in YAC128 striatum. Although peripheral LPS caused an increase in microglial activation and degenerative changes in cerebrovasculature, the phenotypic hallmarks of HD in YAC128 mice such as motor coordination deficits and decreased striatal volume were not exacerbated by chronic peripheral LPS exposure. This study identifies age-dependent increases in microglial activation and angiogenesis in YAC128 at 12 months. Peripheral inflammation induced by chronic LPS causes similar changes but does not influence the HD phenotype in YAC128 mice.

Highlights

► Microglia morphology and cerebrovasculature are altered in YAC128 mouse striatum. ► Peripheral LPS stimulation potentiated microglial activation and vessel remodeling. ► The YAC128 mouse HD phenotype was unaltered by chronic peripheral LPS stimulation.

Introduction

Huntington Disease (HD) is an autosomal dominant inherited neurodegenerative disease caused by an expanded CAG repeat in the HTT gene which encodes the huntingtin protein (Ross and Tabrizi, 2011). Symptoms of HD include motor abnormalities, psychiatric disturbances and cognitive decline. Neuronal death of primarily medium spiny neurons in the caudate/putamen collectively called the striatum is a pathological hallmark of early HD. In addition, inflammation in the striatum including the presence of microglia, the immune cells of the central nervous system (CNS), with altered morphology has also been reported in areas of neuronal loss (Sapp et al., 2001) and their presence is thought to precede disease onset (Bjorkqvist et al., 2008, Tai et al., 2007).

Microglial responses differ depending on their extracellular milieu and microglial activation has been implicated in HD pathogenesis (Moller, 2010). Under normal resting conditions, microglia take on a ramified morphology characterized by a small cell body with several protruding processes and secrete factors which maintain CNS homeostasis. However, when stimulated as a result of traumatic injury or insult, microglia become rounded, retract their processes and secrete a plethora of factors which potentiate an inflammatory response to aid in tissue repair and remodeling (Hanisch and Kettenmann, 2007, Moller, 2010). If persistently activated, as occurs in neurodegeneration, microglia can become reactive, secreting neurotoxic substances leading to neuronal loss (Moller, 2010). Indeed evidence for a distinct inflammatory process including increases in inflammatory cytokines IL-1β and TNF-α in the striatum of HD patients has been reported (Silvestroni et al., 2009). Similarly, a correlation between increased microglial activation with HD progression has also been documented (Pavese et al., 2006). Furthermore, peripheral monocytes isolated from premanifest HD patients showed a strong secretory response to the inflammatory stimulus, lipopolysaccharide (LPS), a bacterial endotoxin, as compared to control subjects. This response was paralleled by a potentiated secretory response in macrophages from YAC128 and from microglia in R6/2 mouse models of HD (Bjorkqvist et al., 2008). These reports suggest that microglial activation may be involved in the pathogenesis of HD.

Systemic infections have profound effects on the diseased brain, either acutely exacerbating symptoms, or having long-term consequences particularly in elderly and demented patients (Perry, 2010). Cytokines such as TNF-α, IL-1β and IL-6 can be produced peripherally and enter the CNS resulting in microglial stimulation. These cytokines have been reported to induce a CNS inflammatory response which alters the blood brain barrier and affects neuronal function (van Gool et al., 2010). Systemic inflammation has also been associated with increased cognitive decline in Alzheimer's disease (AD) patients indicating that systemic inflammation or infection does play a role in neurodegenerative disease (Holmes et al., 2009). Peripheral immune activation occurs prior to disease onset in HD and is correlated with an increase in pro-inflammatory cytokines such as IL-6 and IL-8 in cerebrospinal fluid with increasing stage of HD (Bjorkqvist et al., 2008).

Systemic challenge with LPS in mice is known to activate the innate immune system and cause sickness behavior thought to be due to increased synthesis of inflammatory mediators in the CNS (Bluthe et al., 2000a, Bluthe et al., 2000b). Peripherally, injection of LPS in mice induces a persistent pro-inflammatory state in the CNS for several months after a single challenge (Qin et al., 2007). Furthermore, systemic injection of LPS has been shown to exaggerate the sickness behavior response in a mouse model of prion disease with significant neurodegeneration and causes a phenotypic switch in microglia from a “primed” state where they are activated but with minimal cytokine synthesis to a state where significant production of proinflammatory cytokines such as IL-1β occurs (Combrinck et al., 2002). Several mechanisms through which systemic LPS triggers a CNS inflammatory response have been proposed and include propagation of inflammatory cytokines, prostaglandins or of LPS itself across either an intact or altered blood brain barrier (BBB) (Banks and Robinson, 2010, Gaillard et al., 2003, Qin et al., 2007, Singh and Yiang, 2004, Xaio et al., 2001) or causing release of inflammatory substances from cells that make up the BBB such as endothelial cells (Verma et al., 2006).

Neurovascular abnormalities have been identified in a number of neurodegenerative disorders and may be secondary to defects in signaling factors between vessels and neurons, termed angioneurins (Zacchigna et al., 2008). For instance, morphological changes in brain blood vessels have been implicated in AD and normal aging (Bailey et al., 2004). To date, the role of the neurovasculature changes in HD has not been investigated. Abnormalities in cerebrovasculature such as decreased cerebral blood flow in the caudate have been reported from imaging studies in both symptomatic and presymptomatic HD patients (Harris et al., 1996, Harris et al., 1999), however, the BBB is thought to be intact in HD (Pan and Kastin, 2007). In animal models, the BBB is compromised as a result of 3-NP intrastriatal injection (Duran-Vilaregut et al., 2010). Furthermore, vessel lumen narrowing and increased angiogenesis in post-mortem HD brain tissue have been documented (Vis et al., 1998). Brain derived neurotrohpic factor (BDNF), which is a neurotrophic factor known to be decreased in serum of HD patients (Squitieri et al., 2009), has also been shown to stimulate angiogenesis. Conversely, release of vascular endothelial growth factor (VEGF), which regulates vessel growth in healthy and diseased tissue and stimulates neurogenesis, is reportedly increased from mutant huntingtin striatal cells (Niatsetskaya et al., 2010). These studies suggest that neurovascular changes and impaired crosstalk between vessels and neurons may play a role in HD pathogenesis.

Several studies indicate that chronic systemic LPS challenge exacerbates pathology and behavioral disturbances in animal models of neurodegeneration (Cunningham et al., 2005, Kitazawa et al., 2005, Nguyen et al., 2004), however, the effect of chronic peripheral LPS stimulation on HD pathogenesis is unknown. Since data indicate that peripheral immune system activation has been shown to precede HD onset (Bjorkqvist et al., 2008), we examined age-dependent changes in microglial activation and cerebrovasculature in the striatum of the YAC128 mouse, an early model of HD. We then determined the effects of chronic peripheral LPS challenge on the HD phenotype in YAC128 mouse. We hypothesized that chronic systemic LPS challenge would increase microglial activation and cerebrovasculature changes and accelerate or exacerbate the progression of the HD phenotype in YAC128 mice.

Section snippets

Animals

YAC128 mice of mixed gender and their non-transgenic littermates maintained on the FVB/N (Charles River, Wilmington, MA) strain were used for these experiments (Slow et al., 2003). Mice were group housed with a normal 12 h light–dark cycle in a clean facility and given free access to food and water. All experiments were carried out in accordance with protocols approved by UBC Animal Care and Use Committee.

Tissue processing

Mice were injected with heparin prior to terminal anesthetization by ip injection of 2.5%

Age-dependent alterations in morphology of YAC128 mice

Microglia, under normal resting and surveillance conditions, typically display a spindle shaped, ramified morphology and when activated, microglia retract their processes and transform into large ameboid shaped cells and migrate to the site of injury (Hanisch and Kettenmann, 2007, Moller, 2010). We investigated whether any morphological age-dependent changes occur in microglia in the striatum of YAC128 mice. Representative photomicrographs of Iba1 positive microglia in striatum of 3, 6 and 12 

Discussion

In the present study, we identify significant age-dependent changes in microglia and cerebral blood vessels in the striatum of the YAC128 mouse model of HD. Our hypothesis was that peripheral administration of the pro-inflammatory stimulus LPS would exacerbate these changes and modify the HD phenotype of YAC128 mice. Our main findings are that age-dependent changes including altered microglial morphology (Fig. 1, Fig. 3) and perturbations in the vasculature (Fig. 2, Fig. 4) occur in YAC128

Conclusions

Our findings indicate that the YAC128 mouse model of early HD recapitulates changes in microglial morphology observed in human HD and provide the first description of neurovascular abnormalities in a mouse model of HD. Chronic peripheral inflammation induced by LPS exacerbates the microglial morphological changes and induces neurovascular disruption but does not alter the HD phenotype of YAC128 mice. The role of chronic peripheral inflammation at later stages of the disease merits further

Acknowledgments

We would like to thank our colleagues for useful comments and discussion. We would also like to thank Danny Rogers and Ge Lu for excellent technical assistance. This study was supported by CHDI, The Canadian Institutes for Health Research, Pacific Alzheimer Research Foundation and the Huntington Society of Canada. M.R.H. is a Killam University Professor and holds a Canada Research Chair in Human Genetics.

The authors declare no relevant financial interests.

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