Development of Immune Cell Specific Markers is Essential to Study the Role of Immune Cells Following Ischemic Insult
Bhakta Prasad Gaire, Department of Pharmacology, College of Pharmacy and Gachon Institute of Pharmaceutical Sciences, Gachon University, Incheon
24 June 2015
In an inspiring article by Ritzel and colleagues, the functional differences between microglia and monocytes following ischemic stroke have been examined in a mouse model of transient focal cerebral ischemia. The researchers claim that they have identified distinct functional roles of brain resident microglia and infiltrating monocytes in the ischemic brain. This article not only reflects the novel role of infiltrating monocyte as an important target for the future stroke therapy, but also informs the huge debate about the functional role of brain resident microglia following ischemic insults.
Although there is a high probability of circulating immune cell infiltration into the brain following ischemia induced blood brain barrier disruption, convincing evidence suggests that the brain's resident microglia play a vital role in inflammatory response after ischemic challenge. Previous studies have reported that it is the brain's resident microglia, rather than the infiltrating immune cells, that participate in the inflammatory cascades following ischemic insult. The brain's resident microglia proliferate after ischemia and their number significantly increases after 3- 5 days in the accumulation region of the ischemic brain territory. With the help of immunostaining and an in vivo two-photon imaging technique, Li et al., revealed that cells involved in the process of microgliosis were mainly derived from proliferating resident microglia (1). Using conventional iba1 immunostaining, we have also reported that the number of activated microglia was significantly increased 1 or 3 days following a 90 min middle cerebral artery occlusion (MCAo)/reperfusion (M/R) induced focal cerebral ischemia. Furthermore, 90 min of M/R induced significant microglial proliferation 3 days after ischemia induction (2). Several other studies reveal an increase in microglia activation in post ischemic rodent brain (3, 4).
In contrast to the previous findings, Ritzel and colleagues, report that there is no significant change in microglia populations 24 h following 90 min of MCAo, and that there is a significant reduction of microglia after 72 h. Moreover, infiltrating monocytes increased 72 h after MCAo challenge. There is a lack of specific markers for immuno-histological study that could support their findings. In addition, they have proven distinct signaling pathways of monocyte and microglia at different time-points following stroke by flow cytometric analysis. Although I agree that use of iba1 immunostaining alone cannot confirm the fate of microglia following ischemia induction, and that the use of specific immune cell markers is essential when addressing the particular role of specific immune cells in ischemia, a few points need to be clarified by Ritzel and colleagues about their novel findings. The authors noticed that newly recruited monocytes expressed higher levels of TNF at 24 h; however, microglia expressed significantly greater levels by 72 h. At 72 h after ischemic challenge, the authors report a significant decrease in microglia and an increase in monocyte population. Does that mean that the TNF produced by decreased levels of microglia is still higher than that produced by an increased level of monocytes at 72 h? The phagocytic activity of microglia increased after 72 h, but monocytes predominate at 24 h and 7 days following ischemic insult.
Previous reports have suggested that microglia transform their morphology from the ramified (inactivated) to amoeboid (activated) form at 72 h following ischemic challenge (2, 5). Data regarding the microglia morphology and phagocytic activity would have been of great interests. BrdU/iba1 double positive cells are reported to increase 72 h after ischemia induction, suggesting microglia proliferation; however, Ritzel et al., suggest that these cells are newly infiltrated bone marrow monocytes and not microglia. Availability of immune cell specific markers would have allowed the authors to identify the type of immune cells that were present. The authors also mention that TNF is produced by phagocytic microglia, but evidence for TNF production by phagocytic microglia is lacking.
In conclusion, the overall informative article by Ritzel et al., is somehow overcome by misleading information regarding the exact role of brain resident microglia following ischemic challenge. This study includes truly novel findings that report a distinct role for bone marrow derived monocytes in the ischemic brain; however, these findings alone cannot eliminate the concept of microgliosis following ischemic insult. Ultimately, better immunostaining techniques and specific immune cell markers should be developed to specify the exact role of immune cells in cerebral ischemia/stroke.
References:
1. Li, T., S. Pang, Y. Yu, X. Wu, J. Guo, and S. Zhang. 2013. Proliferation of parenchymal microglia is the main source of microgliosis after ischaemic stroke. Brain : a journal of neurology 136: 3578-3588.
2. Gaire, B. P., O. W. Kwon, S. H. Park, K. H. Chun, S. Y. Kim, D. Y. Shin, and J. W. Choi. 2015. Neuroprotective effect of 6-paradol in focal cerebral ischemia involves the attenuation of neuroinflammatory responses in activated microglia. PloS one 10: e0120203.
3. Boscia, F., R. Gala, A. Pannaccione, A. Secondo, A. Scorziello, G. Di Renzo, and L. Annunziato. 2009. NCX1 expression and functional activity increase in microglia invading the infarct core. Stroke; a journal of cerebral circulation 40: 3608-3617.
4. Ito, D., K. Tanaka, S. Suzuki, T. Dembo, and Y. Fukuuchi. 2001. Enhanced expression of Iba1, ionized calcium-binding adapter molecule 1, after transient focal cerebral ischemia in rat brain. Stroke; a journal of cerebral circulation 32: 1208-1215.
5. Taylor, R. A., and L. H. Sansing. 2013. Microglial responses after ischemic stroke and intracerebral hemorrhage. Clinical & developmental immunology 2013: 746068.
Journal of Neuroinflammation
Development of Immune Cell Specific Markers is Essential to Study the Role of Immune Cells Following Ischemic Insult
24 June 2015
In an inspiring article by Ritzel and colleagues, the functional differences between microglia and monocytes following ischemic stroke have been examined in a mouse model of transient focal cerebral ischemia. The researchers claim that they have identified distinct functional roles of brain resident microglia and infiltrating monocytes in the ischemic brain. This article not only reflects the novel role of infiltrating monocyte as an important target for the future stroke therapy, but also informs the huge debate about the functional role of brain resident microglia following ischemic insults.
Although there is a high probability of circulating immune cell infiltration into the brain following ischemia induced blood brain barrier disruption, convincing evidence suggests that the brain's resident microglia play a vital role in inflammatory response after ischemic challenge. Previous studies have reported that it is the brain's resident microglia, rather than the infiltrating immune cells, that participate in the inflammatory cascades following ischemic insult. The brain's resident microglia proliferate after ischemia and their number significantly increases after 3- 5 days in the accumulation region of the ischemic brain territory. With the help of immunostaining and an in vivo two-photon imaging technique, Li et al., revealed that cells involved in the process of microgliosis were mainly derived from proliferating resident microglia (1). Using conventional iba1 immunostaining, we have also reported that the number of activated microglia was significantly increased 1 or 3 days following a 90 min middle cerebral artery occlusion (MCAo)/reperfusion (M/R) induced focal cerebral ischemia. Furthermore, 90 min of M/R induced significant microglial proliferation 3 days after ischemia induction (2). Several other studies reveal an increase in microglia activation in post ischemic rodent brain (3, 4).
In contrast to the previous findings, Ritzel and colleagues, report that there is no significant change in microglia populations 24 h following 90 min of MCAo, and that there is a significant reduction of microglia after 72 h. Moreover, infiltrating monocytes increased 72 h after MCAo challenge. There is a lack of specific markers for immuno-histological study that could support their findings. In addition, they have proven distinct signaling pathways of monocyte and microglia at different time-points following stroke by flow cytometric analysis. Although I agree that use of iba1 immunostaining alone cannot confirm the fate of microglia following ischemia induction, and that the use of specific immune cell markers is essential when addressing the particular role of specific immune cells in ischemia, a few points need to be clarified by Ritzel and colleagues about their novel findings. The authors noticed that newly recruited monocytes expressed higher levels of TNF at 24 h; however, microglia expressed significantly greater levels by 72 h. At 72 h after ischemic challenge, the authors report a significant decrease in microglia and an increase in monocyte population. Does that mean that the TNF produced by decreased levels of microglia is still higher than that produced by an increased level of monocytes at 72 h? The phagocytic activity of microglia increased after 72 h, but monocytes predominate at 24 h and 7 days following ischemic insult.
Previous reports have suggested that microglia transform their morphology from the ramified (inactivated) to amoeboid (activated) form at 72 h following ischemic challenge (2, 5). Data regarding the microglia morphology and phagocytic activity would have been of great interests. BrdU/iba1 double positive cells are reported to increase 72 h after ischemia induction, suggesting microglia proliferation; however, Ritzel et al., suggest that these cells are newly infiltrated bone marrow monocytes and not microglia. Availability of immune cell specific markers would have allowed the authors to identify the type of immune cells that were present. The authors also mention that TNF is produced by phagocytic microglia, but evidence for TNF production by phagocytic microglia is lacking.
In conclusion, the overall informative article by Ritzel et al., is somehow overcome by misleading information regarding the exact role of brain resident microglia following ischemic challenge. This study includes truly novel findings that report a distinct role for bone marrow derived monocytes in the ischemic brain; however, these findings alone cannot eliminate the concept of microgliosis following ischemic insult. Ultimately, better immunostaining techniques and specific immune cell markers should be developed to specify the exact role of immune cells in cerebral ischemia/stroke.
References:
1. Li, T., S. Pang, Y. Yu, X. Wu, J. Guo, and S. Zhang. 2013. Proliferation of parenchymal microglia is the main source of microgliosis after ischaemic stroke. Brain : a journal of neurology 136: 3578-3588.
2. Gaire, B. P., O. W. Kwon, S. H. Park, K. H. Chun, S. Y. Kim, D. Y. Shin, and J. W. Choi. 2015. Neuroprotective effect of 6-paradol in focal cerebral ischemia involves the attenuation of neuroinflammatory responses in activated microglia. PloS one 10: e0120203.
3. Boscia, F., R. Gala, A. Pannaccione, A. Secondo, A. Scorziello, G. Di Renzo, and L. Annunziato. 2009. NCX1 expression and functional activity increase in microglia invading the infarct core. Stroke; a journal of cerebral circulation 40: 3608-3617.
4. Ito, D., K. Tanaka, S. Suzuki, T. Dembo, and Y. Fukuuchi. 2001. Enhanced expression of Iba1, ionized calcium-binding adapter molecule 1, after transient focal cerebral ischemia in rat brain. Stroke; a journal of cerebral circulation 32: 1208-1215.
5. Taylor, R. A., and L. H. Sansing. 2013. Microglial responses after ischemic stroke and intracerebral hemorrhage. Clinical & developmental immunology 2013: 746068.
Competing interests
No