NADPH oxidase and aging drive microglial activation,oxidative stress,and dopaminergic neurodegeneration following systemic LPS administration

Parkinson's disease is characterized by a progressive degeneration of substantia nigra (SN) dopaminergic neurons with age. We previously found that a single systemic lipopolysaccharide (LPS, 5 mg/kg, i.p.) injection caused a slow progressive loss of tyrosine hydroxylase immunoreactive (TH+IR) neurons in SN associated with increasing motor dysfunction. In this study, we investigated the role of NADPH oxidase (NOX) in inflammation‐mediated SN neurotoxicity. A comparison of control (NOX2^+/+) mice with NOX subunit gp91^phox‐deficient (NOX2^?/?) mice 10 months after LPS administration (5 mg/kg, i.p.) resulted in a 39% (P < 0.01) loss of TH+IR neurons in NOX2^+/+ mice, whereas NOX2^?/? mice did not show a significant decrease. Microglia (Iba1+IR) showed morphological activation in NOX2^+/+ mice, but not in NOX2^?/? mice at 1 hr. Treatment of NOX2^+/+ mice with LPS resulted in a 12‐fold increase in NOX2 mRNA in midbrain and 5.5–6.5‐fold increases in NOX2 protein (+IR) in SN compared with the saline controls. Brain reactive oxygen species (ROS), determined using diphenyliodonium histochemistry, was increased by LPS in SN between 1 hr and 20 months. Diphenyliodonium (DPI), an NOX inhibitor, blocked LPS‐induced activation of microglia and production of ROS, TNFα, IL‐1β, and MCP‐1. Although LPS increased microglial activation and ROS at all ages studied, saline control NOX2^+/+ mice showed age‐related increases in microglial activation, NOX, and ROS levels at 12 and 22 months of age. Together, these results suggest that NOX contributes to persistent microglial activation, ROS production, and dopaminergic neurodegeneration that persist and continue to increase with age. © 147.     

Axon responses of embryonic stem cell‐derived dopaminergic neurons to semaphorins 3A and 3C

Class 3 Semaphorins are a subfamily of chemotropic molecules implicated in the projection of dopaminergic neurons from the ventral mesencephalon and in the formation of the nigrostriatal pathway (NSP) during embryonic development. In humans, loss of mesencephalic dopaminergic neurons leads to Parkinson's disease (PD). Cell replacement therapy with dopaminergic neurons generated from embryonic stem cells (ES‐TH⁺) is being actively explored in models of PD. Among several requisites for this approach to work are adequate reconstruction of the NSP and correct innervation of normal striatal targets by dopaminergic axons. In this work, we characterized the response of ES‐TH⁺ neurons to semaphorins 3A, 3C, and 3F and compared it with that of tyrosine hidroxylase‐positive neurons (TH⁺) obtained from embryonic ventral mesencephalon (VM‐TH⁺). We observed that similar proportions of ES‐TH⁺ and VM‐TH⁺ neurons express semaphorin receptors neuropilins 1 and 2. Furthermore, the axons of both populations responded very similarly to semaphorin exposure: semaphorin 3A increased axon length, and semaphorin 3C attracted axons and increased their length. These effects were mediated by neuropilins, insofar as addition of blocking antibodies against these proteins reduced the effects on axonal growth and attraction, and only TH⁺ axons expressing neuropilins responded to the semaphorins analyzed. The observations reported here show phenotypic similarities between VM‐TH⁺ and ES‐TH⁺ neurons and suggest that semaphorins 3A and 3C could be employed to guide axons of grafted ES‐TH⁺ in therapeutic protocols for PD. © 2009 Wiley‐Liss, Inc.     

Induction of GAP‐43 modulates neuroplasticity in PBSC (CD34+) implanted‐Parkinson's model

As a result of the progressive decrease in efficacy of drugs used to treat Parkinson's disease (PD) and the rapid development of motor complications, effective alternative treatments for PD are required. In a 6‐hydroxydopamine (6‐OHDA)‐induced Parkinson's rat model, intracerebral peripheral blood stem cell (CD34⁺) (PBSC) transplantation significantly protected dopaminergic neurons from 6‐OHDA‐induced neurotoxicity, enhanced neural repair of tyrosine hydroxylase neurons through up‐regulation of Bcl‐2, facilitated stem cell plasticity, and attenuated activation of microglia, in comparison with vehicle‐control rats. The 6‐OHDA‐lesioned hemi‐Parkinsonian rats receiving intrastriatal transplantation of PBSCs also showed: 1) enhanced glucose metabolism in the lesioned striatum and thalamus, demonstrated by [¹⁸F]fluoro‐2‐deoxyglucose positron emission tomography (FDG‐PET), 2) improved neurochemical activity as shown by proton magnetic resonance spectroscopy (¹H‐MRS), and 3) significantly reduced rotational behavior in comparison with control lesioned rats. These observations might be explained by an up‐regulation of growth‐associated protein 43 (GAP‐43) expression because improvements in neurological dysfunction were blocked by injection of MK‐801 in the PBSC‐treated group. In addition, a significant increase in neurotrophic factor expression was found in the ipsilateral hemisphere of the PBSC‐treated group. In summary, this protocol may be a useful strategy for the treatment of clinical PD. © 2009 Wiley‐Liss, Inc.     

The 6‐hydroxydopamine‐induced nigrostriatal neurodegeneration produces microglia‐like NG2 glial cells in the rat substantia nigra

Neuron/glial 2 (NG2)‐expressing cells are often referred to as oligodendrocyte precursor cells. NG2‐expressing cells have also been identified as multipotent progenitor cells. However, microglia‐like NG2 glial cells have not been fully examined in neurodegenerative disorders such as Parkinson's disease (PD). In the present study, we chose two rat models of PD, i.e., intranigral or intrastriatal injection of 6‐hydroxydopamine (6‐OHDA), since the cell bodies of dopamine (DA) neurons, which form a nigrostriatal pathway, are in the substantia nigra pars compacta (SNpc) while their nerve terminals are in the striatum. In the nigral 6‐OHDA‐injected model, activated NG2‐positive cells were detected in the SNpc but not in the striatum. In contrast, in the striatal 6‐OHDA‐injected model, these cells were detected in both the SNpc and the striatum. In both models, activated NG2‐positive cells were located close to surviving tyrosine hydroxylase (TH)‐positive neurons in the SNpc. In addition, activated NG2‐positive cells in the SNpc coexpressed ionized calcium‐binding adaptor molecule 1 (Iba1), a microglia/macrophage marker. Interestingly, these double‐positive glial cells coexpressed glial cell line‐derived neurotrophic factor (GDNF). These results suggest that microglia‐like NG2 glial cells may help protect DA neurons and may lead to new therapeutic targets in PD. © 2010 Wiley‐Liss, Inc.     

Glial cells suppress postencephalitic CD8+ T lymphocytes through PD‐L1

Engagement of the programmed death (PD)?1 receptor on activated cells by its ligand (PD‐L1) is a mechanism for suppression of activated T‐lymphocytes. Microglia, the resident inflammatory cells of the brain, are important for pathogen detection and initiation of innate immunity, however, a novel role for these cells as immune regulators has also emerged. PD‐L1 on microglia has been shown to negatively regulate T‐cell activation in models of multiple sclerosis and acute viral encephalitis. In this study, we investigated the role of glial cell PD‐L1 in controlling encephalitogenic CD8⁺ T‐lymphocytes, which infiltrate the brain to manage viral infection, but remain to produce chronic neuroinflammation. Using a model of chronic neuroinflammation following murine cytomegalovirus (MCMV)‐induced encephalitis, we found that CD8⁺ T‐cells persisting within the brain expressed PD‐1. Conversely, activated microglia expressed PD‐L1. In vitro, primary murine microglia, which express low basal levels of PD‐L1, upregulated the co‐inhibitory ligand on IFN‐γ‐treatment. Blockade of the PD‐1: PD‐L1 pathway in microglial: CD8⁺ T‐cell co‐cultures increased T‐cell IFN‐γ and interleukin (IL)?2 production. We observed a similar phenomenon following blockade of this co‐inhibitory pathway in astrocyte: CD8⁺ T‐cell co‐cultures. Using ex vivo cultures of brain leukocytes, including microglia and CD8⁺ T‐cells, obtained from mice with MCMV‐induced chronic neuroinflammation, we found that neutralization of either PD‐1 or PD‐L1 increased IFN‐γ production from virus‐specific CD8⁺ T‐cells stimulated with MCMV IE1_168–176 peptide. These data demonstrate that microglia and astrocytes control antiviral T‐cell responses and suggest a therapeutic potential of PD1: PD‐L1 modulation to manage the deleterious consequences of uncontrolled neuroinflammation. GLIA 2014;62:1582–1594     

Doxycycline restrains glia and confers neuroprotection in a 6‐OHDA Parkinson model

Neuron–glia interactions play a key role in maintaining and regulating the central nervous system. Glial cells are implicated in the function of dopamine neurons and regulate their survival and resistance to injury. Parkinson's disease is characterized by the loss of dopamine neurons in the substantia nigra pars compacta, decreased striatal dopamine levels and consequent onset of extrapyramidal motor dysfunction. Parkinson's disease is a common chronic, neurodegenerative disorder with no effective protective treatment. In the 6‐OHDA mouse model of Parkinson's disease, doxycycline administered at a dose that both induces/represses conditional transgene expression in the tetracycline system, mitigates the loss of dopaminergic neurons in the substantia nigra compacta and nerve terminals in the striatum. This protective effect was associated with: (1) a reduction of microglia in normal mice as a result of doxycycline administration per se; (2) a decrease in the astrocyte and microglia response to the neurotoxin 6‐OHDA in the globus pallidus and substantia nigra compacta, and (3) the astrocyte reaction in the striatum. Our results suggest that doxycycline blocks 6‐OHDA neurotoxicity in vivo by inhibiting microglial and astrocyte expression. This action of doxycycline in nigrostriatal dopaminergic neuron protection is consistent with a role of glial cells in Parkinson's disease neurodegeneration. The neuroprotective effect of doxycycline may be useful in preventing or slowing the progression of Parkinson's disease and other neurodegenerative diseases linked to glia function.     

Neuromelanin Activates Microglia and Induces Degeneration of Dopaminergic Neurons: Implications for Progression of Parkinson’s Disease

In Parkinson’s disease (PD), there is a progressive loss of neuromelanin (NM)-containing dopamine neurons in substantia nigra (SN) which is associated with microgliosis and presence of extracellular NM. Herein, we have investigated the interplay between microglia and human NM on the degeneration of SN dopaminergic neurons. Although NM particles are phagocytized and degraded by microglia within minutes in vitro, extracellular NM particles induce microglial activation and ensuing production of superoxide, nitric oxide, hydrogen peroxide (H₂O₂), and pro-inflammatory factors. Furthermore, NM produces, in a microglia-depended manner, neurodegeneration in primary ventral midbrain cultures. Neurodegeneration was effectively attenuated with microglia derived from mice deficient in macrophage antigen complex-1, a microglial integrin receptor involved in the initiation of phagocytosis. Neuronal loss was also attenuated with microglia derived from mice deficient in phagocytic oxidase, a subunit of NADPH oxidase, that is responsible for superoxide and H₂O₂ production, or apocynin, an NADPH oxidase inhibitor. In vivo, NM injected into rat SN produces microgliosis and a loss of tyrosine hydroxylase neurons. Thus, these results show that extracellular NM can activate microglia, which in turn may induce dopaminergic neurodegeneration in PD. Our study may have far-reaching implications, both pathogenic and therapeutic.     

Single low doses of MPTP decrease tyrosine hydroxylase expression in the absence of overt neuron loss

Parkinson's disease (PD) is the second most common age-related neurodegenerative disease. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a prototypical neurotoxicant used in mice to mimic primary features of PD pathology including striatal dopamine depletion and dopamine neuron loss in the substantia nigra pars compacta (SNc). In the literature, there are several experimental paradigms involving multiple doses of MPTP that are used to elicit dopamine neuron loss. However, a recent study reported that a single low dose caused significant loss of dopamine neurons. Here, we determined the effect of a single intraperitoneal injection of one of three doses of MPTP (0.1, 2 and 20 mg/kg) on dopamine neurons, labeled by tyrosine hydroxylase (TH⁺), and total neuron number (Nissl⁺) in the SNc using unbiased stereological counting. Data reveal a significant loss of neurons in the SNc (TH⁺ and Nissl⁺) only in the group treated with 20 mg/kg MPTP. Groups treated with lower dose of MPTP (0.1 and 2 mg/kg) only showed significant loss of TH⁺ neurons rather than TH⁺ and Nissl⁺ neurons. Striatal dopamine levels were decreased in the groups treated with 2 and 20 mg/kg MPTP and striatal terminal markers including, TH and the dopamine transporter (DAT), were only decreased in the groups treated with 20 mg/kg MPTP. These data demonstrate that lower doses of MPTP likely result in loss of TH expression rather than actual dopamine neuron loss in the SN. This finding reinforces the need to measure both total neuron number along with TH⁺ cells in determining dopamine neuron loss.     

Down‐regulation of microglia and NG2‐positive cells reaction in trimethyltin‐injured hippocampus of rats treated with myelin basic protein‐reactive T cells: Possible contribution to the neuroprotective effect of T cells

In our previous investigations, we demonstrated that CD4⁺ antimyelin basic protein (MBP) T cells protect hippocampal neurons against trimethyltin‐induced damage. We hypothesized involvement of T cells, interacting with the various glial populations activated during the neurodegeneration process. In this study, we employ immunocytochemical methods to investigate the influence of administration of T cells on the response of microglia and of NG2⁺ cells to trimethyltin (TMT)‐induced damage. Female Lewis rats were treated with anti‐MBP CD4⁺ T cells (4 million per animal, i.v) 24 hr after TMT (8 mg/kg, i.p) intoxication. TMT caused degeneration of CA4 hipppocampal neurons and evoked an abundant reaction of microglial and NG2⁺ cells in the injured region. The cells changed morphology into the activated state, and the number of OX42⁺ and NG2⁺ cells increased about 4.5‐fold and 3‐fold, respectively, relative to controls as assessed on day 21 after TMT treatment. Additionally, the cells of ameboid morphology, which expressed NG2 or microglial antigens, appeared in the zone of neurodegeneration. Furthermore, certain cells of ameboid phenotype shared both antigens. In rats treated with T cells, down‐regulation of the activation of both glial classes and reduction of formation of their ameboid forms was observed. The number of the total OX42⁺ and NG2⁺ cells decreased by 21% and 54%, respectively, and the number of their ameboid forms decreased by 46% and 73%, respectively. Our data suggest that the diminished activation of microglia and NG2⁺ cells, particularly the reduced number of their ameboid forms, may contribute to the neuroprotective effect of T cells. © 2009 Wiley‐Liss, Inc.     

Activated microglia in a rat stroke model express NG2 proteoglycan in peri‐infarct tissue through the involvement of TGF‐β1

We investigated activated microglia in ischemic brain lesions from rats that had been subjected to transient middle cerebral artery occlusion. Activated microglia expressing NG2 chondroitin sulfate proteoglycan (NG2) were found only in the narrow zone (demarcation zone) that demarcated the peri‐infarct tissue and ischemic core. NG2^? activated microglia were abundantly distributed in the peri‐infarct tissue outside the demarcation zone. NG2⁺ microglia but not NG2^? microglia expressed both CD68 and a triggering receptor expressed on myeloid cells 2 (TREM‐2), suggesting that NG2⁺ microglia eliminated apoptotic neurons. In fact, NG2⁺ microglia often attached to degenerating neurons and sometimes internalized NeuN⁺ or neurofilament protein⁺ material. Kinetic studies using quantitative real‐time RT‐PCR revealed that expression of transforming growth factor‐β1 (TGF‐β1) was most evident in the ischemic core; with this marker produced mainly by macrophages located in this region. TGF‐β receptor mRNA expression peaked at 3 days post reperfusion (dpr) in the peri‐infarct tissue, including the demarcation zone. Primary cultured rat microglia also expressed the receptor mRNA. In response to TGF‐β1, primary microglia enhanced the expression of NG2 protein and TREM‐2 mRNA as well as migratory activity. A TGF‐β1 inhibitor, SB525334, abolished these effects. The present results suggest that TGF‐β1 produced in the ischemic core diffused toward the peri‐infarct tissue, driving activated microglial cells to eliminate degenerating neurons. Appropriate control of NG2⁺ microglia in the demarcation zone might be a novel target for the suppression of secondary neurodegeneration in the peri‐infarct tissue. GLIA 2014;62:185–198     

Effects of 3,3',5‐triiodothyronine on microglial functions

l ‐tri‐iodothyronine (3, 3', 5–triiodothyronine; T3) is an active form of the thyroid hormone (TH) essential for the development and function of the CNS. Though nongenomic effect of TH, its plasma membrane–bound receptor, and its signaling has been identified, precise function in each cell type of the CNS remained to be investigated. Clearance of cell debris and apoptotic cells by microglia phagocytosis is a critical step for the restoration of damaged neuron‐glia networks. Here we report nongenomic effects of T3 on microglial functions. Exposure to T3 increased migration, membrane ruffling and phagocytosis of primary cultured mouse microglia. Injection of T3 together with stab wound attracted more microglia to the lesion site in vivo. Blocking TH transporters and receptors (TRs) or TRα‐knock‐out (KO) suppressed T3‐induced microglial migration and morphological change. The T3‐induced microglial migration or membrane ruffling was attenuated by inhibiting G_i/o‐protein as well as NO synthase, and subsequent signaling such as phosphoinositide 3‐kinase (PI3K), mitogen‐activated protein kinase (MAPK)/extracellular signal‐regulated kinase (ERK). Inhibitors for Na⁺/K⁺‐ATPase, reverse mode of Na⁺/Ca²⁺ exchanger (NCX), and small‐conductance Ca²⁺‐dependent K⁺ (SK) channel also attenuated microglial migration or phagocytosis. Interestingly, T3‐induced microglial migration, but not phagocytosis, was dependent on GABA_A and GABA_B receptors, though GABA itself did not affect migratory aptitude. Our results demonstrate that T3 modulates multiple functional responses of microglia via multiple complex mechanisms, which may contribute to physiological and/or pathophysiological functions of the CNS. GLIA 2015:63:906–920     

Extensive exploration of a novel rat model of Parkinson's disease using partial 6‐hydroxydopamine lesion of dopaminergic neurons suggests new therapeutic approaches

Parkinson's disease (PD) is characterized by the degeneration of dopaminergic (DA) neurons constituting the nigrostriatal pathway. Neuroinflammation, related to microglial activation, plays an important role in this process. Exploration of animal models of PD using neuroimaging modalities allows to better understand the pathophysiology of the disease. Here, we fully explored a moderate lesion model in the rat in which 6‐hydroxydopamine was unilaterally delivered in three sites along the striatum. The degenerative process was assessed through in vivo Positron Emission Tomography (PET) imaging and in vitro autoradiographic quantitation of the striatal dopamine transporter (DAT) and immunostaining of tyrosine hydroxylase (TH). The microglial activation was studied through in vitro autoradiographic quantitation of the 18 kDa translocator protein (TSPO) in the striatum and CD11b staining in the SN. In addition, a targeted metabolomics exploration was performed in both these structures using mass spectrometry coupled to HPLC. Our results showed a reproducible decrease in the striatal DAT density associated with a reduction in the number of TH‐positive cells in the SN and striatum, reflecting a robust moderate degeneration of nigrostriatal DA neurons. In addition, we observed strong microglia activation in both the striatum and SN ipsilateral to the lesion, highlighting that this moderate degeneration of DA neurons was associated with a marked neuroinflammation. Our metabolomics studies revealed alterations of specific metabolites and metabolic pathways such as carnitine, arginine/proline, and histidine metabolisms. These results bring new insights in the PD mechanism knowledge and new potential targets for future therapeutic strategies.     

Downregulation of miR‐7116‐5p in microglia by MPP+ sensitizes TNF‐α production to induce dopaminergic neuron damage

Activation of microglia resulting in exacerbated inflammation expression plays an important role in degeneration of dopaminergic (DA) neurons in the pathogenesis of Parkinson's disease (PD). However, how this enhanced inflammation is induced in microglia remains largely unclear. Here, in the mouse PD model induced by 1‐methyl‐4‐phenyl‐1,2,3,6‐tetra hydropyridine (MPTP), we found that miR‐7116‐5p in microglia has a crucial role in this inflammation. 1‐methyl‐4‐phenylpyridinium (MPP⁺) is uptaken by microglia through organic cation transporter 3 (OCT3) to downregulate miR‐7116‐5p, an miRNA found to target tumor necrosis factor alpha (TNF‐α). Production of TNF‐α in microglia is specifically potentiated by MPP⁺ via downregulation of miR‐7116‐5p to elicit subsequent inflammatory responses. Furthermore, enhancement of miR‐7116‐5p expression in microglia in mice inhibits the production of TNF‐α and the activation of glia, and further prevents loss of DA neurons. Together, our studies suggest that MPP⁺ suppresses miR‐7116‐5p level in microglia and potentiates TNF‐α production and inflammatory responses to contribute to DA neuron damage.     

The dual dopamine‐glutamate phenotype of growing mesencephalic neurons regresses in mature rat brain

Coexpression of tyrosine hydroxylase (TH) and vesicular glutamate transporter 2 (VGLUT2) mRNAs in the ventral tegmental area (VTA) and colocalization of these proteins in axon terminals of the nucleus accumbens (nAcb) have recently been demonstrated in immature (15‐day‐old) rat. After neonatal 6‐hydroxydopamine (6‐OHDA) lesion, the proportion of VTA neurons expressing both mRNAs and of nAcb terminals displaying the two proteins was enhanced. To determine the fate of this dual phenotype in adults, double in situ hybridization and dual immunolabeling for TH and VGLUT2 were performed in 90‐day‐old rats subjected or not to the neonatal 6‐OHDA lesion. Very few neurons expressed both mRNAs in the VTA and substantia nigra (SN) of P90 rats, even after neonatal 6‐OHDA. Dually immunolabeled terminals were no longer found in the nAcb of normal P90 rats and were exceedingly rare in the nAcb of 6‐OHDA‐lesioned rats, although they had represented 28% and 37% of all TH terminals at P15. Similarly, 17% of all TH terminals in normal neostriatum and 46% in the dopamine neoinnervation of SN in 6‐OHDA‐lesioned rats were also immunoreactive for VGLUT2 at P15, but none at P90. In these three regions, all dually labeled terminals made synapse, in contradistinction to those immunolabeled for only TH or VGLUT2 at P15. These results suggest a regression of the VGLUT2 phenotype of dopamine neurons with age, following normal development, lesion, or sprouting after injury, and a role for glutamate in the establishment of synapses by these neurons. J. Comp. Neurol. 517:873–891, 2009. © 2009 Wiley‐Liss, Inc.     

Cx3cr1‐deficiency exacerbates alpha‐synuclein‐A53T induced neuroinflammation and neurodegeneration in a mouse model of Parkinson's disease

Parkinson's disease (PD) is the second most common neurodegenerative disorder characterized by the degeneration of dopaminergic neurons of the substantia nigra and the accumulation of protein aggregates, called Lewy bodies, where the most abundant is alpha‐synuclein (α‐SYN). Mutations of the gene that codes for α‐SYN (SNCA), such as the A53T mutation, and duplications of the gene generate cases of PD with autosomal dominant inheritance. As a result of the association of inflammation with the neurodegeneration of PD, we analyzed whether overexpression of wild‐type α‐SYN (α‐SYN^WT) or mutated α‐SYN (α‐SYN^A53T) are involved in the neuronal dopaminergic loss and inflammation process, along with the role of the chemokine fractalkine (CX3CL1) and its receptor (CX3CR1). We generated in vivo murine models overexpressing human α‐SYN^WT or α‐SYN^A53T in wild type (Cx3cr1^+/+) or deficient (Cx3cr1^–/–) mice for CX3CR1 using unilateral intracerebral injection of adeno‐associated viral vectors. No changes in CX3CL1 levels were observed by immunofluorescence or analysis by qRT‐PCR in this model. Interestingly, the expression α‐SYN^WT induced dopaminergic neuronal death to a similar degree in both genotypes. However, the expression of α‐SYN^A53T produced an exacerbated neurodegeneration, enhanced in the Cx3cr1^–/– mice. This neurodegeneration was accompanied by an increase in neuroinflammation and microgliosis as well as the production of pro‐inflammatory markers, which were exacerbated in Cx3cr1^–/– mice overexpressing α‐SYN^A53T. Furthermore, we observed that in primary microglia CX3CR1 was a critical factor in the modulation of microglial dynamics in response to α‐SYN^WT or α‐SYN^A53T. Altogether, our study reveals that CX3CR1 plays an essential role in neuroinflammation induced by α‐SYN^A53T.     

The majority of newly generated cells in the adult mouse substantia nigra express low levels of Doublecortin,but their proliferation is unaffected by 6‐OHDA‐induced nigral lesion or Minocycline‐mediated inhibition of neuroinflammation

Parkinson's disease is characterized by a selective loss of dopaminergic neurons in the substantia nigra (SN). However, whether regenerative endogenous neurogenesis is taking place in the mammalian SN of parkinsonian and non‐parkinsonian brains remains of debate. Here, we tested whether proliferating cells in the SN and their neurogenic potential would be affected by anti‐inflammatory treatment under physiological conditions and in the 6‐hydroxy‐dopamine (6‐OHDA) Parkinson's disease mouse model. We report that the majority of newly generated nigral cells are positive for Doublecortin (Dcx), which is an often used marker for neural progenitor cells. Yet, Dcx expression levels in these cells were much lower than in neural progenitor cells of the subventricular zone and the dentate gyrus neural progenitor cells. Furthermore, these newly generated nigral cells are negative for neuronal lineage markers such as TuJ1 and NeuN. Therefore, their neuronal commitment is questionable. Instead, we found evidence for oligodendrogenesis and astrogliosis in the SN. Finally, neither short‐term nor long‐term inhibition of neuroinflammation by Minocycline‐ or 6‐OHDA‐induced lesion affected the numbers of newly generated cells in our disease paradigm. Our findings of adult generated Dcx⁺ cells in the SN add important data for understanding the cellular composition and consequently the regenerative capacity of the SN.     

Role of Fcgamma receptors in nigral cell injury induced by Parkinson disease immunoglobulin injection into mouse substantia nigra

Immune/inflammatory factors have been implicated in the pathogenesis of Parkinson's disease (PD). Immunoglobulin G (IgG) from patients with PD can induce injury of dopaminergic neurons following stereotaxic injection into rat substantia nigra (SN). The PD IgG can be demonstrated in vitro to activate microglia via the Fcgamma receptor (Fcgamma R) and induce dopaminergic cell injury. To confirm the involvement of microglia and their Fcgamma R in IgG-induced lesions of SN in vivo we analyzed the tyrosine hydroxylase (TH)-positive cell loss in SN par compacta (SNpc) in mice lacking Fcgamma receptors (Fcgamma R(-/-)) and wild type (Fcgamma R(+/+)). At 1 day after stereotaxic injection of PD IgG into the SN of Fcgamma R(+/+) mice there was a 27% increase in the number of CD11b-positive microglial cells and no significant loss of TH-positive cells. At 14 days after the stereotaxic injection, the number of microglial cells was increased by 42%, accompanied by a 40% loss of TH-positive neurons in the SNpc. PD IgG injection in Fcgamma R(-/-) mice resulted in no significant increase of microglia and no loss of TH-positive cells in the SNpc at any time point. The injection of F(ab')(2) fragments of PD IgG was able to induce TH-positive neuronal loss in the SNpc only when the injected animals raised antibodies against the injected human IgG fragments, which confirmed the importance of the Fcgamma R in microglial activation and nigral injury.