Activation of the innate immune system has been reported in HD patients, yet it is unclear whether this is due to the effect of mutant htt or represents a secondary event in response to the degeneration of neurons. As evidenced by isolated primary HD microglia culture assays, microglial cells lines with overexpression of mutant htt, isolated human HD myeloid cells, and in vivo thioglycollate assays in HD mice, our results demonstrate that myeloid cells expressing mutant htt exhibit deficits in migration to chemotactic stimuli. Our results also suggest that these deficits are dependent on mutant htt expression. Migration deficits were observed in primary microglia when isolated from early
postnatal HD mice, prior to any significant cell loss, consistent with apparent microglial dysfunction observed in pre-manifest HD patients (19
). Importantly, migration deficits were observed in monocytes and macrophages isolated from HD patients before the onset of neurological and motor symptoms, and these deficits appeared to be independent of disease stage. Similar results were observed in vivo in mouse models of HD.
The detailed molecular mechanisms responsible for migration defects in immune cells that express mutant htt are likely to be complex and remain to be fully elucidated. However, our data suggest that mutant htt perturbs the proper regulation of actin remodeling due to decreased levels of cofilin and p-cofilin. Consistent with our findings in microglia, mutant htt colocalizes with nuclear actin-cofilin rods in neurons under conditions of stress (49
). This study also detected cross-linked complexes of actin and cofilin in HD patient lymphoblasts that correlated with disease progression. Although mutant htt may directly interact with cofilin and actin in microglia and peripheral myeloid cells, cofilin was reported to physically interact only with full-length mutant htt and not mutant htt fragments (49
). On the other hand, we showed reduced cofilin levels in both mutant full-length htt– and fragment htt–expressing microglia. Because we found that the phosphatase activity also appears to be altered upon ATP stimulation, it is conceivable that mutant htt may also affect the kinase that phosphorylates cofilin and/or other components of the actin remodeling machinery. As reported, in non-stimulated conditions, membrane ruffling and actin polymerization are reduced in the dendritic spines and synapses of striatal neurons from HD mouse models (51
). Since membrane ruffling is not altered in HD microglia in the absence of stimulation, the functional effect of mutant htt on actin remodeling may be different from that in neurons. Cytoskeletal organization is needed for normal neuronal cell development and functionality, and perhaps neurons are more sensitive to mutant htt effects under basal conditions. Moreover, the effects of mutant htt and the normal functions of htt on actin remodeling may vary between neurons and immune cells. Interestingly, null mutants of htt in Dictyostelium discoideum
are also impaired in polarization and chemotaxis, suggesting that migration impairment in mutant htt–expressing immune cells could be due at least in part to the loss of WT htt function (53
Our observations of migration deficits in microglia and peripheral myeloid cells that express mutant htt may have important implications for understanding HD pathogenesis. Under basal conditions in the intact healthy brain, microglia represent “sentinels” whose highly motile processes constantly survey the brain parenchyma and make transient activity-dependent contacts with synapses (31
). One recent study suggests that microglial activation with ATP triggers astrocyte-mediated modulation of excitatory neurotransmission (54
). Microglia have dysfunctional morphological phenotypes and are aberrantly activated early in HD patients and mice. Interestingly, microglia from mice that lack both copies of the chemokine receptor Cx3cr1
), which functions in leukocyte trafficking, also have activated morphological phenotypes (55
). We show that microglia harvested from early postnatal
HD mice already have defects in process extension and migration, suggesting this impairment could contribute to the early activation phenotypes reported in the HD brain. In addition, under basal, unperturbed conditions, microglia from adult BACHD mice exhibited significant deficits in process extension and exaggerated process retraction. Since synapse levels are decreased in BACHD mice (16
) and other HD mouse models (56
), these results suggest that microglia expressing mutant htt may not be efficient in synapse surveillance, which could conceivably contribute to synaptic and network dysfunction. We also found that microglia from aged BACHD mice showed a delayed response to laser-induced injury. As microglia form the first line of defense in the CNS by orchestrating rapid responses aimed to contain pathological insults within the normal brain (30
), these results suggest that mutant htt–expressing microglia might be limited in their ability to respond to traumatic or pathological insults, such as traumatic brain injury or brain infections. Taken together, our results indicate that migration impairment in microglia may be one of the first abnormalities in the HD brain, thereby compromising the functional integrity of the HD brain in early and late stages.
Mutant htt expression in peripheral myeloid cells might also contribute to some aspects of central and peripheral pathogenesis in HD. Proinflammatory cytokines and chemokines are elevated in pre-manifest HD patients and remain elevated throughout the course of disease (11
). We hypothesize that migration deficits in immune cells may lead to inefficient transduction of cytokine and chemokine signaling mechanisms, which may explain chronic increases in proinflammatory cytokines and chemokines and microglial activation in HD patients. Indeed, IL-6, IL-8, IL-1β, TNF-α, and MCP-1 levels are elevated in HD and are predicted to augment inflammatory signals in the brain. These factors could also contribute directly to neuronal dysfunction, as receptors for many of these molecules are expressed on neurons, some of which transduce proapoptotic signaling pathways (57
). Consistent with this hypothesis, myeloid cells in mice mutant for the chemokine receptor Cx3cr1
are impaired in migration, leading to increased IL-1β levels and decreased synaptic plasticity (55
). In mice with deletion of Ccr2, whose monocytes and macrophages also do not migrate properly, serum levels of IL-6 and IL-1β are elevated (33
). Mice mutant for these chemokine receptors are also unable to clear infections, have impaired wound healing, and are unable to mount effective adaptive immune responses (33
). Unfortunately, no studies to date have systematically examined whether the incidence of infections or defects in wound healing are increased in HD patients, although chronic skin ulcers have been reported (64
). Ongoing studies in our laboratory are examining whether HD mice can efficiently clear bacterial and viral infections, and if infections influence degenerative processes in the CNS and periphery.
Several recent studies in mouse models provide additional support for the hypothesis that dysfunction of the peripheral immune system might play an important disease-modifying role in HD. Proinflammatory cytokines did not increase in YAC128 mice that received transplants of WT bone marrow, and this conferred modest behavioral and neuropathological benefits to YAC128 and BACHD mice (16
). Genetic deletion of cannabinoid receptor 2 (CB2) — a protein expressed predominantly in peripheral immune cells that regulates production of proinflammatory cytokines, likely via NF-κB (65
) — exacerbates pathogenesis in a mouse model of HD (68
). Consistent with these findings, we recently found that CB2 signaling in immune cells mediates the onset and severity of symptoms in BACHD mice and that CB2 agonists are neuroprotective in R6/2 mice, even when given at late disease stages (69
). Finally, inhibition of kynurenine 3-monooxygenase (KMO) in blood cells prevented synaptic loss and brain inflammation and increased lifespan in R6/2 mice (56
). Together these findings support a critical link between blood cells and neurodegeneration in HD.
In summary, our study provides in vitro and in vivo evidence for motility and migration deficits in immune cells from HD patients and mouse models. These deficits may partially explain the early immune dysfunction and chronic elevation of proinflammatory cytokines and chemokines in this disease. Immune cell migration impairment is known to cause recurrent infections and immune dysregulation (34
). Further studies to monitor the innate and adaptive immune responses in HD mouse models and patients are warranted. Therapeutic approaches currently used to lower proinflammatory cytokines, including antibodies to TNF-α and IL-6, might provide some benefits to HD patients and should be investigated in mouse models of HD. More broadly, disease-causing proteins in many neurodegenerative diseases caused by polyQ expansions are expressed ubiquitously, and the functional consequences of this expression in non-neuronal cells, including immune cells, merits further investigation.