Gwenn A. Garden

Pathways to neuronal injury and apoptosis in HIV-associated dementia

Human immunodeficiency virus-1 (HIV-1) can induce dementia with alarming occurrence worldwide. The mechanism remains poorly understood, but discovery in brain of HIV-1-binding sites (chemokine receptors) provides new insights. HIV-1 infects macrophages and microglia, but not neurons, although neurons are injured and die by apoptosis. The predominant pathway to neuronal injury is indirect through release of macrophage, microglial and astrocyte toxins, although direct injury by viral proteins might also contribute. These toxins overstimulate neurons, resulting in the formation of free radicals and excitotoxicity, similar to other neurodegenerative diseases. Recent advances in understanding the signalling pathways mediating these events offer hope for therapeutic intervention.

Association Between Carotid Plaque Characteristics and Subsequent Ischemic Cerebrovascular Events A Prospective Assessment With MRI—Initial Results

Background and Purpose— MRI is able to quantify carotid plaque size and composition with good accuracy and reproducibility and provides an opportunity to prospectively examine the relationship between plaque features and subsequent cerebrovascular events. We tested the hypothesis that the characteristics of carotid plaque, as assessed by MRI, are possible predictors of future ipsilateral cerebrovascular events. Methods— A total of 154 consecutive subjects who initially had an asymptomatic 50% to 79% carotid stenosis by ultrasound with ≥12 months of follow-up were included in this study. Multicontrast-weighted carotid MRIs were performed at baseline, and participants were followed clinically every 3 months to identify symptoms of cerebrovascular events. Results— Over a mean follow-up period of 38.2 months, 12 carotid cerebrovascular events occurred ipsilateral to the index carotid artery. Cox regression analysis demonstrated a significant association between baseline MRI identification of the following plaque characteristics and subsequent symptoms during follow-up: presence of a thin or ruptured fibrous cap (hazard ratio, 17.0; P≤0.001), intraplaque hemorrhage (hazard ratio, 5.2; P=0.005), larger mean intraplaque hemorrhage area (hazard ratio for 10 mm2 increase, 2.6; P=0.006), larger maximum %lipid-rich/necrotic core (hazard ratio for 10% increase, 1.6; P=0.004), and larger maximum wall thickness (hazard ratio for a 1-mm increase, 1.6; P=0.008). Conclusions— Among patients who initially had an asymptomatic 50% to 79% carotid stenosis, arteries with thinned or ruptured fibrous caps, intraplaque hemorrhage, larger maximum %lipid-rich/necrotic cores, and larger maximum wall thickness by MRI were associated with the occurrence of subsequent cerebrovascular events. Findings from this prospective study provide a basis for larger multicenter studies to assess the risk of plaque features for subsequent ischemic events.

Microglia Biology in Health and Disease

Microglia cells are resident central nervous system (CNS) leukocytes that regulate innate immunity and participate in adaptive immune responses in CNS tissue. However, microglia cells also appear to play an important role during normal function of the mature nervous system. In response to injury, ischemia, and inflammatory stimuli, microglia cells assume an activated phenotype associated with proliferation, migration to the site of injury, phagocytosis of cellular debris, and elaboration (Power and Proudfoot 2001) of both neurotoxic and neurotrophic factors. Recent reports strongly suggest that regulating microglia function may be a fruitful future therapeutic target for the prevention of neurological dysfunction in a variety of CNS injuries and chronic diseases. Thus, developing a thorough understanding of extracellular signals that activate microglia as well as a complete catalogue of microglia responses to activating stimuli in both the healthy and diseased state are crucial scientific endeavors. This review presents the current understanding of the biology of microglia during normal CNS function as well as in response to CNS injury or neurodegenerative disease. In addition, microglia modulate both the activation and down-regulation of the adaptive immune response in the CNS. Evidence that microglia cells play a primary role in regulating CNS immune responses will also be discussed.

Caspase Cascades in Human Immunodeficiency Virus-Associated Neurodegeneration

Many patients infected with human immunodeficiency virus-1 (HIV-1) develop a syndrome of neurologic deterioration known as HIV-associated dementia (HAD). Neurons are not productively infected by HIV-1; thus, the mechanism of HIV-induced neuronal injury remains incompletely understood. Several investigators have observed evidence of neuronal injury, including dendritic degeneration, and apoptosis in CNS tissue from patients with HAD. Caspase enzymes, proteases associated with the process of apoptosis, are synthesized as inactive proenzymes and are activated in a proteolytic cascade after exposure to apoptotic signals. Here we demonstrate that HAD is associated with active caspase-3-like immunoreactivity that is localized to the soma and dendrites of neurons in affected regions of the human brain. Additionally, the cascade of caspase activation was studied using anin vitro model of HIV-induced neuronal apoptosis. Increased caspase-3 proteolytic activity and mitochondrial release of cytochrome c were observed in cerebrocortical cultures exposed to the HIV coat protein gp120. Specific inhibitors of both the Fas/tumor necrosis factor-α/death receptor pathway and the mitochondrial caspase pathway prevented gp120-induced neuronal apoptosis. Caspase inhibition also prevented the dendrite degeneration observed in vivo in transgenic mice with CNS expression of HIV/gp120. These findings suggest that pharmacologic interventions aimed at the caspase enzyme pathways may be beneficial for the prevention or treatment of HAD.

Bergmann glia expression of polyglutamine-expanded ataxin-7 produces neurodegeneration by impairing glutamate transport

Non-neuronal cells may be pivotal in neurodegenerative disease, but the mechanistic basis of this effect remains ill-defined. In the polyglutamine disease spinocerebellar ataxia type 7 (SCA7), Purkinje cells undergo non-cell-autonomous degeneration in transgenic mice. We considered the possibility that glial dysfunction leads to Purkinje cell degeneration, and generated mice that express ataxin-7 in Bergmann glia of the cerebellum with the Gfa2 promoter. Bergmann glia–specific expression of mutant ataxin-7 was sufficient to produce ataxia and neurodegeneration. Expression of the Bergmann glia–specific glutamate transporter GLAST was reduced in Gfa2-SCA7 mice and was associated with impaired glutamate transport in cultured Bergmann glia, cerebellar slices and cerebellar synaptosomes. Ultrastructural analysis of Purkinje cells revealed findings of dark cell degeneration consistent with excitotoxic injury. Our studies indicate that impairment of glutamate transport secondary to glial dysfunction contributes to SCA7 neurodegeneration, and suggest a similar role for glial dysfunction in other polyglutamine diseases and SCAs.

p53-Dependent Cell Death Signaling in Neurons

The p53 tumor suppressor gene is a sequence-specific transcription factor that activates the expression of genes engaged in promoting growth arrest or cell death in response to multiple forms of cellular stress. p53 expression is elevated in damaged neurons in acute models of injury such as ischemia and epilepsy and in brain tissue samples derived from animal models and patients with chronic neurodegenerative diseases. p53 deficiency or p53 inhibition protects neurons from a wide variety of acute toxic insults. Signal transduction pathways associated with p53-induced neuronal cell death are being characterized, suggesting that intervention may prove effective in maintaining neuronal viability and restoring function following neural injury and disease.

Microglia in human immunodeficiency virus-associated neurodegeneration

Infection with the human immunodeficiency virus (HIV) is associated with a syndrome of cognitive and motor abnormalities that may develop in the absence of opportunistic infections. Neurons are not productively infected by HIV. Thus, one hypothesis to explain the pathophysiology of HIV‐associated dementia (HAD) suggests that signals released from other infected cell types in the CNS secondarily lead to neuronal injury. Microglia are the predominant resident CNS cell type productively infected by HIV‐1. Neurologic dysfunction in HAD appears to be a consequence of microglial infection and activation. Several neurotoxic immunomodulatory factors are released from infected and activated microglia, leading to altered neuronal function, synaptic and dendritic degeneration, and eventual neuronal apoptosis. This review summarizes findings from clinical/pathological studies, animal models, and in vitro models of HAD. Most of these studies support the hypothesis that altered microglial physiology is the nidus for a cascade of events leading to neuronal dysfunction and death. Several molecular mediators of neuronal injury in HAD that emanate from microglia have been identified, and strategies for altering the impact of these neurotoxins are discussed. GLIA 40:240–251, 2002.

HIV associated neurodegeneration requires p53 in neurons and microglia

HIV infection of the central nervous system leads to HIV‐associated dementia (HAD) in a substantial subset of infected individuals. The pathogenesis of neuronal dysfunction in HAD is not well understood, but previous studies have demonstrated evidence for activation of apoptotic pathways. The tumor suppressor transcription factor p53 is an apical mediator of neuronal apoptosis following a variety of injurious stimuli. To determine whether p53 participates in HAD, we exposed cerebrocortical cultures from wild‐type and p53 deficient mice to the neurotoxic HIV envelope protein gp120. Using neuron/microglia co‐culture of mixed p53 genotype, we observed that both neurons and microglia require p53 for gp120 induced neuronal apoptosis. Additionally, accumulation of p53 protein in neurons was recently reported in post‐ mortem cortical tissue from a small group of HAD patients. Using a much larger cohort of HAD cases, we extend this finding and report that p53 protein also increases in non‐neuronal cells, including microglia. Taken together these findings demonstrate a novel role for p53 in the microglial response to gp120. Additionally, these findings, in conjunction with a recent report that monocytes expressing HIV‐Tat also secrete neurotoxins that promote p53 activation, suggest that distinct HIV proteins may converge on the p53 pathway to promote neurotoxicity.

A Simple Composite Phenotype Scoring System for Evaluating Mouse Models of Cerebellar Ataxia

We describe a protocol for the rapid and sensitive quantification of disease severity in mouse models of cerebella ataxia. It is derived from previously published phenotype assessments in several disease models, including spinocerebellar ataxias, Huntington s disease and spinobulbar muscular atrophy. Measures include hind limb clasping, ledge test, gait and kyphosis. Each measure is recorded on a scale of 0-3, with a combined total of 0-12 for all four measures. The results effectively discriminate between affected and non-affected individuals, while also quantifying the temporal progression of neurodegenerative disease phenotypes. Measures may be analyzed individually or combined into a composite phenotype score for greater statistical power. The ideal combination of the four described measures will depend upon the disorder in question. We present an example of the protocol used to assess disease severity in a transgenic mouse model of spinocerebellar ataxia type 7 (SCA7). Albert R. La Spada and Gwenn A. Garden contributed to this manuscript equally.

Autophagy activation and enhanced mitophagy characterize the Purkinje cells of pcd mice prior to neuronal death

Purkinje cells are a class of specialized neurons in the cerebellum, and are among the most metabolically active of all neurons, as they receive immense synaptic stimulation, and provide the only efferent output from the cerebellum. Degeneration of Purkinje cells is a common feature of inherited ataxias in humans and mice. To understand Purkinje neuron degeneration, investigators have turned to naturally occurring Purkinje cell degeneration phenotypes in mice to identify key regulatory proteins and cellular pathways. The Purkinje cell degeneration (pcd) mouse is a recessive mutant characterized by complete and dramatic post-natal, cell autonomous Purkinje neuron degeneration and death. As the basis of Purkinje cell death in pcd is unresolved, and contradictory data has emerged for the role of autophagy in Purkinje cell degeneration, we studied the mechanism of Purkinje cell death in pcd mice. BAX null status did not suppress Purkinje neuron death in pcd mice, indicating that classic apoptosis is not responsible for Purkinje cell loss. Interestingly, LC3 Western blot analysis and GFP-LC3 immunostaining of degenerating pcd cerebellum revealed activation of the autophagy pathway. Ultrastructural studies confirmed increased autophagy pathway activity in Purkinje cells, and yielded evidence for mitophagy, in agreement with LC3 immunoblotting of cerebellar fractions. As p62 levels were decreased in pcd cerebellum, our findings suggest that pcd Purkinje cell neurons can execute effective autophagy. However, our results support a role for dysregulated autophagy activation in pcd, and suggest that increased or aberrant mitophagy contributes to the Purkinje cell degeneration in pcd mice.