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Development of microglia in mouse neopallial cell cultures
1Department of Anatomy, University of Saskatchewan, Saskatoon, Canada.
Abstract:
Microglia develop in cultures initiated from disaggregated neopallial cells of newborn C3H/HeJ mice when the cultures are subjected to nutritional deprivation for 10 or more days (Hao et al: Int J Dev Neurosci 9:1-14, 1991). In the present experiments, the cultures were pulsed with BrdU for 3 hours at different times during incubation and then the cells were immunoreacted with antibodies against BrdU, GFAP, and CR3 receptor. The dividing cells (BrdU+) were found to be either GFAP+ or GFAP-, but not Mac-1+/BrdU+. Infection of proliferating cells after 2 or more days of incubation with replication-deficient retroviral vector containing E. coli lacZ reporter gene resulted in many labeled astroglia cell clones but no labeled microglia. However, when cells were infected right after disaggregation of neopallium, labeled Mac-1+ microglia were found. When Mac-1+ cells in a suspension of disaggregated neopallial cells were killed using complement mediated lysis before setting up the cultures, Mac-1+ microglia developed, in spite of the treatment. We conclude that in cultures initiated from mouse neopallium there are MAC-1-/GFAP- microglia progenitor cells which do not divide in nutritionally deprived cultures but can transform into Mac-1+ microglia under the influence of astroglia-derived trophic factors. Microglia, which become Mac-1+ (i.e., express CR3 receptor), proliferate extensively in the presence of CSF-1 (which is produced by astroglia).
Insights
Newly identified microglia progenitor cells (Mac-1-/GFAP-) transform into microglia under astroglia influence. These cells proliferate extensively with colony-stimulating factor-1 (CSF-1) in mouse neopallial cultures.
Area of Science:
- Neuroscience
- Immunology
- Developmental Biology
Background:
- Microglia development in vitro is crucial for understanding neuroinflammation and brain development.
- Previous studies suggested microglia arise from specific progenitor cells under certain culture conditions.
Purpose of the Study:
- To identify and characterize microglia progenitor cells in mouse neopallial cultures.
- To elucidate the factors influencing microglia development and proliferation in vitro.
Main Methods:
- Primary cultures of disaggregated mouse neopallial cells were established.
- Cell proliferation was assessed using Bromodeoxyuridine (BrdU) incorporation.
- Immunoreactivity for BrdU, Glial Fibrillary Acidic Protein (GFAP), and Mac-1 (CD11b/CR3 receptor) was analyzed.
- Replication-deficient retroviral vectors carrying the E. coli lacZ reporter gene were used for cell lineage tracing.
Main Results:
- Proliferating cells (BrdU+) were GFAP+ or GFAP-, but not Mac-1+.
- Retroviral labeling of proliferating cells after 2 days yielded labeled astroglia but no microglia.
- Labeling immediately after disaggregation or pre-treatment to deplete Mac-1+ cells still resulted in microglia development.
- Astroglia-derived factors, particularly colony-stimulating factor-1 (CSF-1), promoted Mac-1+ microglia proliferation.
Conclusions:
- Mouse neopallial cultures contain Mac-1-/GFAP- microglia progenitor cells.
- These progenitors require astroglia-derived trophic factors, like CSF-1, to differentiate into Mac-1+ microglia.
- The progenitor cells do not divide under nutritional deprivation but can be activated by astroglia.