Comparison of the detrimental features of microglia and infiltrated macrophages in traumatic brain injury: A study using a hypnotic bromovalerylurea

Abstract

Microglia and blood-borne macrophages in injured or diseased brains are difficult to distinguish because they share many common characteristics. However, the identification of microglia-specific markers and the use of flow cytometry have recently made it easy to discriminate these types of cells. In this study, we analyzed the features of blood-borne macrophages, and activated and resting microglia in a rat traumatic brain injury (TBI) model. Oxidative injury was indicated in macrophages and neurons in TBI lesions by the presence of 8-hydroxy-2'-deoxyguanosine (8-OHdG). Generation of mitochondrial reactive oxygen species (ROS) was markedly observed in granulocytes and macrophages, but not in activated or resting microglia. Dihydroethidium staining supported microglia not being the major source of ROS in TBI lesions. Furthermore, macrophages expressed NADPH oxidase 2, interleukin-1β (IL-1β), and CD68 at higher levels than microglia. In contrast, microglia expressed transforming growth factor β1 (TGFβ1), interleukin-6 (IL-6), and tumor necrosis factor α at higher levels than macrophages. A hypnotic, bromovalerylurea (BU), which has anti-inflammatory effects, reduced both glycolysis and mitochondrial oxygen consumption. BU administration inhibited chemokine CCL2 expression, accumulation of monocytes/macrophages, 8-OHdG generation, mitochondrial ROS generation, and proinflammatory cytokine expression, and markedly ameliorated the outcome of the TBI model. Yet, BU did not inhibit microglial activation or expression of TGFβ1 and insulin-like growth factor 1 (IGF-1). These results indicate that macrophages are the major aggravating cell type in TBI lesions, in particular during the acute phase. Activated microglia may even play favorable roles. Reduction of cellular energy metabolism in macrophages and suppression of CCL2 expression in injured tissue may lead to amelioration of TBI.

Favorable and unfavorable roles of microglia and macrophages in the pathologic central nervous system

“…Moreover, TMEM119-expressing microglia in AD brains either do not or only weakly express the polarized markers CD80, CD163, or CD206. In a TBI model, ramified or homeostatic microglia express TMEM119 at higher levels than do activated microglia [10] . Therefore, TMEM119 is particularly suitable for identifying homeostatic microglia with a ramified shape in both the normal and the injured mature brain.…”

“…Because of faint expression of CD45 by homeostatic microglia, immunohistochemical detection of this expression tends to be difficult. Conversely, macrophages and neutrophils express CD45 rather strongly [10] . However, weak CD45 expression can be detected easily by flow cytometry and is an advantage in flow cytometry analysis; microglia can be defined as CD11b + / CD45 lo and macrophages as CD11b + /CD45 hi .…”

“…Conversely, blood-borne macrophages and neutrophils, the latter of which infiltrate in abundance, do not have processes even though they may have a polygonal shape and short spikes [27] . Thus, microglia can be distinguished from blood-borne cells via morphological observation [10] . This may be the simplest method for specific identification of microglia.…”

“…Therefore, microglia express a myeloid cell marker CD11b and a panleukocyte marker CD45. Based on this finding, microglia have been analyzed by flow cytometry using antibodies to CD11b and CD45 [10,39] . Because of faint expression of CD45 by homeostatic microglia, immunohistochemical detection of this expression tends to be difficult.…”

“…This assumption originated from the historical observation by Rio-Hortega [8] , who reported that ramified microglia in the normal mature brain can, in severely damaged brains, turn into phagocytes exhibiting spherical shapes of the same morphology as that of blood-borne macrophages [5] . Therefore, numerous studies have described CD11b-expresssing (in the case of the rat brain, a monoclonal antibody OX-42-immunoreactive) spherical cells in severely injured brains in the acute phase as activated microglia, although most of them should be recognized as invading neutrophils [9,10] .…”