title: Macrophages are essential for maintaining a M2 protective response early after ischemic brain injury.
abstract: Resident microglia and recruited macrophages are major contributors to the post-ischemic inflammatory response. Initially considered functionally homogeneous populations, data now suggest distinct but still controversial roles after brain injury. Using a model of conditional monocyte/macrophage depletion we studied the contribution of these myeloid cells to brain lesion progression after ischemia, and their influence on the ischemic inflammatory environment. Male CD11b-DTR transgenic mice, expressing the human diphtheria toxin receptor under the control of the CD11b promoter, were treated with diphtheria toxin to induce monocyte/macrophage depletion. Twenty four hours later the middle cerebral artery was permanently occluded. The ischemic lesion was measured 24h after injury. At the same time microglia and macrophage activation and polarization were assessed by quantitative immunohistochemistry and confocal microscopy for CD45high, CD11b, CD68, CD16/32, iNOS, Arg1, Ym1, and CD206, and gene expression was investigated on CD11b+sorted cells. Depletion of monocytes/macrophages worsened the ischemic lesion within 24h after the ischemic insult. This effect was associated with higher M1/M2 polarization ratio in the ischemic lesion. Moreover, depletion increased the expression of M1 phenotypic markers on CD11b positive cells. Gene expression on CD11b+sorted cells indicated a selective increase of iNOS and lower Arg1 mRNA expression than in non depleted mice. Depletion of monocytes/macrophages increases the ischemic lesion, an effect accompanied by an increase in the M1/M2 polarization ratio of microglia and macrophages in the ischemic area. Thus in ischemic injury recruited monocytes/macrophages may control an excessive M1 pro-inflammatory response, suggesting their ability to drive M2 protective polarization.
authors.affiliation: IRCCS - Istituto di Ricerche Farmacologiche Mario Negri, Department of Neuroscience, via La Masa 19, 20156 Milan, Italy. Electronic address: carlo.perego@marionegri.it.
authors.forename: Carlo
authors.initials: C
authors.lastname: Perego
authors.affiliation: IRCCS - Istituto di Ricerche Farmacologiche Mario Negri, Department of Neuroscience, via La Masa 19, 20156 Milan, Italy; Neurosurgical Intensive Care Unit, Fondazione IRCCS Ca' Granda/Ospedale Maggiore Policlinico, Via Francesco Sforza 28, 20122 Milan, Italy. Electronic address: stefano.fumagalli@marionegri.it.
authors.forename: Stefano
authors.initials: S
authors.lastname: Fumagalli
authors.affiliation: IRCCS - Istituto di Ricerche Farmacologiche Mario Negri, Department of Neuroscience, via La Masa 19, 20156 Milan, Italy. Electronic address: elisa.zanier@marionegri.it.
authors.forename: Elisa R
authors.initials: ER
authors.lastname: Zanier
authors.affiliation: IRCCS - Istituto di Ricerche Farmacologiche Mario Negri, Department of Neuroscience, via La Masa 19, 20156 Milan, Italy; Neurosurgical Intensive Care Unit, Fondazione IRCCS Ca' Granda/Ospedale Maggiore Policlinico, Via Francesco Sforza 28, 20122 Milan, Italy. Electronic address: erika.carlino@marionegri.it.
authors.forename: Erika
authors.initials: E
authors.lastname: Carlino
authors.affiliation: Department of Oncology, Flow Cytometry Unit, IRCCS - Istituto di Ricerche Farmacologiche Mario Negri, via La Masa 19, 20156 Milan, Italy. Electronic address: nicolo.panini@marionegri.it.
authors.forename: Nicolò
authors.initials: N
authors.lastname: Panini
authors.affiliation: Department of Oncology, Flow Cytometry Unit, IRCCS - Istituto di Ricerche Farmacologiche Mario Negri, via La Masa 19, 20156 Milan, Italy. Electronic address: eugenio.erba@marionegri.it.
authors.forename: Eugenio
authors.initials: E
authors.lastname: Erba
authors.affiliation: IRCCS - Istituto di Ricerche Farmacologiche Mario Negri, Department of Neuroscience, via La Masa 19, 20156 Milan, Italy. Electronic address: desimoni@marionegri.it.
authors.forename: Maria-Grazia
authors.initials: MG
authors.lastname: De Simoni
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D001119:Arginase / Q000378:metabolism; D020520:Brain Infarction / Q000209:etiology; D001930:Brain Injuries / Q000209:etiology / Q000473:pathology*; D002545:Brain Ischemia / Q000150:complications*; D039481:CD11b Antigen / Q000235:genetics; D016764:Cell Polarity / Q000187:drug effects / Q000502:physiology; D004167:Diphtheria Toxin / Q000494:pharmacology; D004195:Disease Models, Animal; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D066257:Heparin-binding EGF-like Growth Factor / Q000235:genetics; D037102:Lectins / Q000378:metabolism; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000473:pathology*; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D017628:Microglia / Q000187:drug effects; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism
chemical_list: D015703:Antigens, CD; D039481:CD11b Antigen; D004167:Diphtheria Toxin; D066257:Heparin-binding EGF-like Growth Factor; D037102:Lectins; C496319:NOS2 protein, human; D052247:Nitric Oxide Synthase Type II; C429541:Chil3 protein, mouse; D001619:beta-N-Acetylhexosaminidases; C544448:Arg1 protein, mouse; D001119:Arginase
keywords: Brain ischemic injury; Inflammation; Macrophage depletion; Microglia