Turnover Of Microglia Research Articles

Chronic Cortical Inflammation, Cognitive Impairment, and Immune Reactivity Associated with Diffuse Brain Injury Are Ameliorated by Forced Turnover of Microglia.

Traumatic brain injury (TBI) is associated with an increased risk of cognitive, psychiatric, and neurodegenerative complications that may develop after injury. Increased microglial reactivity following TBI may underlie chronic neuroinflammation, neuropathology, and exaggerated responses to immune challenges. Therefore, the goal of this study was to force turnover of trauma-associated microglia that develop after diffuse TBI and determine whether this alleviated chronic inflammation, improved functional recovery and attenuated reduced immune reactivity to lipopolysaccharide (LPS) challenge. Male mice received a midline fluid percussion injury (mFPI) and 7 d later were subjected to a forced microglia turnover paradigm using CSF1R antagonism (PLX5622). At 30 d postinjury (dpi), cortical gene expression, dendritic complexity, myelin content, neuronal connectivity, cognition, and immune reactivity were assessed. Myriad neuropathology-related genes were increased 30 dpi in the cortex, and 90% of these gene changes were reversed by microglial turnover. Reduced neuronal connectivity was evident 30 dpi and these deficits were attenuated by microglial turnover. TBI-associated dendritic remodeling and myelin alterations, however, remained 30 dpi independent of microglial turnover. In assessments of functional recovery, increased depressive-like behavior, and cognitive impairment 30 dpi were ameliorated by microglia turnover. To investigate microglial priming and reactivity 30 dpi, mice were injected intraperitoneally with LPS. This immune challenge caused prolonged lethargy, sickness behavior, and microglial reactivity in the TBI mice. These extended complications with LPS in TBI mice were prevented by microglia turnover. Collectively, microglial turnover 7 dpi alleviated behavioral and cognitive impairments associated with microglial priming and immune reactivity 30 dpi.SIGNIFICANCE STATEMENT A striking feature of traumatic brain injury (TBI), even mild injuries, is that over 70% of individuals have long-term neuropsychiatric complications. Chronic inflammatory processes are implicated in the pathology of these complications and these issues can be exaggerated by immune challenge. Therefore, our goal was to force the turnover of microglia 7 d after TBI. This subacute 7 d postinjury (dpi) time point is a critical transitional period in the shift toward chronic inflammatory processes and microglia priming. This forced microglia turnover intervention in mice attenuated the deficits in behavior and cognition 30 dpi. Moreover, microglia priming and immune reactivity after TBI were also reduced with microglia turnover. Therefore, microglia represent therapeutic targets after TBI to reduce persistent neuroinflammation and improve recovery.

Chronic cortical inflammation and cognitive impairment associated with diffuse brain injury is ameliorated by forced turnover of microglia

Chronic cortical inflammation and cognitive impairment associated with diffuse brain injury is ameliorated by forced turnover of microglia

An overview of microglia ontogeny and maturation in the homeostatic and pathological brain.

Microglia are the resident immune cells of the central nervous system (CNS) and are increasingly recognized as critical players in development, brain homeostasis, and disease pathogenesis. The lifespan, maintenance, proliferation, and turnover of microglia are important factors that regulate microglial behavior and affect their roles in the CNS. However, emerging evidence suggests that microglia are morphologically and phenotypically distinct in different brain areas, at different ages, and during disease. Ongoing research focuses on understanding how microglia acquire specific phenotypes in response to extrinsic cues in the environment and how phenotypes are specified by intrinsic properties of different populations of microglia. With the development of pharmacological and genetic tools that allow the investigation of microglia in vivo, there have been considerable advances in understanding molecular signatures of both homeostatic microglia and those reacting to injury and disease. Here, we review the master gene regulators that define microglia as well as discuss the evidence that microglia are heterogeneous and fall into distinct clusters that display specific intrinsic properties and perform unique tasks in different settings. Taken together, the information presented supports the idea that microglia morphology and transcriptional heterogeneity should be considered when studying the complex nature of microglia and their roles in brain health and disease.

Microglia turnover with aging and in an Alzheimer's model via long-term in vivo single-cell imaging.

To clarify the role of microglia in brain homeostasis and disease, an understanding of their maintenance, proliferation and turnover is essential. The lifespan of brain microglia, however, remains uncertain, and reflects confounding factors in earlier assessments that were largely indirect. We genetically labeled single resident microglia in living mice and then used multiphoton microscopy to monitor these cells over time. Under homeostatic conditions, we found that neocortical resident microglia were long-lived, with a median lifetime of well over 15 months; thus, approximately half of these cells survive the entire mouse lifespan. While proliferation of resident neocortical microglia under homeostatic conditions was low, microglial proliferation in a mouse model of Alzheimer's β-amyloidosis was increased threefold. The persistence of individual microglia throughout the mouse lifespan provides an explanation for how microglial priming early in life can induce lasting functional changes and how microglial senescence may contribute to age-related neurodegenerative diseases.

The Lifespan and Turnover of Microglia in the Human Brain

SummaryThe hematopoietic system seeds the CNS with microglial progenitor cells during the fetal period, but the subsequent cell generation dynamics and maintenance of this population have been poorly understood. We report that microglia, unlike most other hematopoietic lineages, renew slowly at a median rate of 28% per year, and some microglia last for more than two decades. Furthermore, we find no evidence for the existence of a substantial population of quiescent long-lived cells, meaning that the microglia population in the human brain is sustained by continuous slow turnover throughout adult life.

Coupled Proliferation and Apoptosis Maintain the Rapid Turnover of Microglia in the Adult Brain

SummaryMicroglia play key roles in brain development, homeostasis, and function, and it is widely assumed that the adult population is long lived and maintained by self-renewal. However, the precise temporal and spatial dynamics of the microglial population are unknown. We show in mice and humans that the turnover of microglia is remarkably fast, allowing the whole population to be renewed several times during a lifetime. The number of microglial cells remains steady from late postnatal stages until aging and is maintained by the spatial and temporal coupling of proliferation and apoptosis, as shown by pulse-chase studies, chronic in vivo imaging of microglia, and the use of mouse models of dysregulated apoptosis. Our results reveal that the microglial population is constantly and rapidly remodeled, expanding our understanding of its role in the maintenance of brain homeostasis.

In vivo imaging of microglia turnover in the mouse retina after ionizing radiation and dexamethasone treatment.

Gamma irradiation and bone marrow transplantation (BMT) are established clinical procedures for the treatment of hematologic malignancies. The radiation targets cells in the bone marrow, but injury to other tissues, including the central nervous system (CNS), have been reported. Here, we examine if anti-inflammatory treatment can mitigate the radiation-induced turnover of retinal microglia and the replacement by bone marrow-derived cells (BMDCs). Two-color chimeric mice were generated by lethal irradiation of heterozygous CX3CR1-GFP mice that express GFP in microglial cells and bone marrow transplantation from universal DsRed donor mice. Mice were treated with the corticosteroid dexamethasone; a control group received no dexamethasone treatment. The populations of resident microglia (GFP+) and BMDCs (DsRed+) were quantified by serial in vivo imaging for 10 weeks after irradiation with a confocal scanning laser ophthalmoscope that we custom-built specifically for multicolor imaging of the murine retina. Ionizing radiation resulted in loss of 75% of the resident retinal microglia population after 70 days. Recruitment of BMDCs was delayed with respect to the microglia loss, resulting in a transient depletion of the total immune cell number in the retina. With dexamethasone treatment, both the loss of the resident microglia and the infiltration of BMDCs were suppressed by at least 50%. Anti-inflammatory treatment with the corticosteroidal agent dexamethasone preserves resident microglia and minimizes recruitment of BMDCs after ionizing radiation exposure and BMT.

Brain conditioning is instrumental for successful microglia reconstitution following hematopoietic stem cell transplantation

The recent hypothesis that postnatal microglia are maintained independently of circulating monocytes by local precursors that colonize the brain before birth has relevant implications for the treatment of various neurological diseases, including lysosomal storage disorders (LSDs), for which hematopoietic cell transplantation (HCT) is applied to repopulate the recipient myeloid compartment, including microglia, with cells expressing the defective functional hydrolase. By studying wild-type and LSD mice at diverse time-points after HCT, we showed the occurrence of a short-term wave of brain infiltration by a fraction of the transplanted hematopoietic progenitors, independently from the administration of a preparatory regimen and from the presence of a disease state in the brain. However, only the use of a conditioning regimen capable of ablating functionally defined brain-resident myeloid precursors allowed turnover of microglia with the donor, mediated by local proliferation of early immigrants rather than entrance of mature cells from the circulation.

426. Cure of Established Disease and Direct Evidence of Cellular Cross-Correction in the Nervous System of Metachromatic Leukodystrophy Mice after HSC-Based Gene Therapy

Top of pageAbstract Metachromatic Leukodystrophy (MLD) is an inborn lipidosis caused by deficiency of the lysosomal enzyme arylsulfatase A (ARSA). It is characterized by storage of undegraded sulfatides in the CNS and PNS, leading to severe demyelination. We showed that transplantation of LV-transduced Hematopoietic Stem Cells (HSC) allowed: i) targeting therapeutic genes to CNS and PNS by turnover of microglia and endoneurial macrophages by HSC-derived cells; ii) preventing functional and neuropathological manifestations of MLD in the mouse model, when applied at pre-symptomatic stage (A. Biffi et al., J. Clin. Invest 2004, 113(8): 1118|[ndash]|29). Here we assessed whether this approach can be effective not only in preventing, but also in correcting already established MLD manifestations. By 5|[ndash]|6 months of age, MLD mice display motor conduction abnormalities in CNS and PNS, impaired motor coordination and learning, and sulfatide storage in the NS. We compared the outcome of transplant performed before disease onset with transplantation of ARSA-transduced HSC in 6|[ndash]|7 months old, already symptomatic, MLD recipients. We reached ARSA over-expression in the hematopoietic system of transplanted mice. Neurophysiological evaluations performed on 12 months old mice demonstrated normalization of motor stimuli conduction along CNS and PNS. Rotarod test showed, by comparison with age matched sham-treated MLD mice, protection from further worsening of motor coordination and learning abilities. Remarkably, 12 months old treated mice showed improved performance respect to 6 months old MLD controls and a similar performance to that of age-matched wild type mice. These data indicated that ex vivo gene therapy not only prevented disease progression, but also corrected already established functional deficits. In order to assess the mechanism responsible for the widespread correction of CNS pathology, and define the contribution of ARSA over-expression, we performed immunohistochemical studies of the brain of MLD mice transplanted with HSC expressing a HA-tagged ARSA. Confocal analysis showed the widespread occurrence of tagged enzyme within HSC-derived microglia and resident CNS neurons, oligodendrocytes and astrocytes. Importantly, ARSA-HA was correctly sorted to the lysosomal compartment in all cell types, proving the occurrence of in vivo enzyme transfer. Because these findings were associated to complete reversal of the neuronal degeneration which affects the hippocampus and cerebellum of aging MLD mice, in vivo enzyme transfer achieved robust metabolic cross-correction of the crucial CNS cell targets of the disease. Overall, these results demonstrate that transplantation of LV-transduced autologous HSC represents a promising therapeutic strategy for correcting MLD and possibly other neurodegenerative disorders in humans.

41. Ex Vivo Gene Therapy Provides a Unique Therapeutic Benefit and Corrects Metachromatic (MLD) and Globoid (GLD) Leukodystrophy in the Mouse Models

MLD and GLD are inborn lipidosis caused by the deficiency of the lysosomal enzymes arylsulfatase A (ARSA) and galactocerebrosidase (GALC). They are characterized by storage of undegraded sulfatides in the CNS and PNS, leading to progressive demyelination and early death. Using Lentiviral Vectors (LV), we efficiently transduced Hematopoietic Stem Cells (HSC) ex vivo and evaluated the potential of their progeny to target therapeutic genes to the CNS and PNS of transplanted mice, and correct MLD and GLD phenotypes in the mouse models. We proved extensive transgene expression in the CNS and PNS of transplanted mice due to progressive turnover of microglia and endoneurial macrophages by HSC-derived cells. Recruitment of these cells was consistently observed independently from the conditioning protocol, and it was faster and more robust in MLD and GLD mice. By transplanting HSC transduced with the ARSA gene, we reconstituted enzyme activity in the hematopoietic system of MLD mice at supranormal levels and prevented all mayor disease manifestations. Transplanted mice were completely protected from the development of motor conduction impairment (normalization of motor conduction velocity and F wave latency, and of central conduction time), of learning and coordination deficits (normalization of the performance at rotarod test), and of neuropathological abnormalities (prevention of sulfatide accumulation and demyelination in CNS and PNS) typical of the disease. Remarkably, ex vivo gene therapy had a significantly better therapeutic impact than wild-type HSC transplantation on all examined parameters, highlighting the crucial role of enzyme over-expression and indicating the likely occurrence of in vivo cross-correction. Confocal analysis of the CNS of treated mice showed the presence of tagged ARSA enzyme within the lysosomes both of the transduced HSC-derived microglia cells and of resident neurons, demonstrating in vivo enzyme transfer and, most likely, cross-correction. To further evaluate the therapeutic impact of this approach we challenged it with the acute and devastating evolution of GLD in the mouse model. Neonate GLD mice were transplanted with HSC transduced with the GALC gene, supplemented with accessory Sca1- cells to reduce short-term toxicity of conditioning. We observed a dramatic improvement in survival of transplanted mice, which reached an average of 135 days, as compared to 40 days of untreated GLD mice and 80 days of mice transplanted with wild type bone marrow, providing striking evidence, in another disease setting, of the therapeutic advantage of ex vivo gene therapy versus allogeneic BMT. We observed a significant improvement in the GLD phenotype (absence of leg palsy and delayed, minor twitching) and prevention of mayor histopathological signs of the disease (demyelination, axonal loss). Overall, these results demonstrate that transplantation of LV-transduced autologous HSC represents a new, potentially efficacious therapeutic strategy for MLD, GLD and possibly other neurodegenerative disorders.