A novel cell line from spontaneously immortalized murine microglia

Abstract

Background Purified microglia cultures are useful tools to study microglial behavior in vitro. Microglial cell lines serve as an attractive alternative to primary microglia culture, circumventing the costly and lengthy preparation of the latter. However, immortalization by genetic or pharmacologic manipulations may show altered physiology from primary microglia. New Method A novel microglial cell line was isolated from a primary glial culture of postnatal murine cerebral cortices. The culture contained a population of spontaneously transformed microglia that continued to divide without genetic or pharmacological manipulations. After several clones were isolated, one particular clone, SIM-A9, was analyzed for its microglial characteristics. Results SIM-A9 cells expressed macrophage/microglia-specific proteins, CD68 and Iba1. SIM-A9 cells were responsive to exogenous inflammatory stimulation with lipopolysaccharide and β-amyloid, triggering tyrosine kinase-based and NFκB signaling cascades as well as TNFα secretion. SIM-A9 cells also exhibited phagocytic uptake of fluorescent labeled β-amyloid and bacterial bioparticles. Furthermore, lipopolysaccharide increased the levels of inducible nitric oxide synthase and cyclooxygenase-2, whereas IL-4 stimulation increased arginase-1 levels demonstrating that SIM-A9 cells are capable of switching their profiles to pro- or anti-inflammatory phenotypes, respectively. Comparison with Existing Methods The use of SIM-A9 cells avoids expensive and lengthy procedures required for the preparation of primary microglia. Spontaneously immortalized SIM-A9 cells are expected to behave more comparably to primary microglia than virally transformed or pharmacologically induced microglial cell lines. Conclusions SIM-A9 cells exhibit key characteristics of cultured primary microglia and may serve as a valuable model system for the investigation of microglial behavior in vitro.

Mesenchymal Stromal Cell-derived Extracellular Vesicles Promote Myeloid-biased Multipotent Hematopoietic Progenitor Expansion via Toll-Like Receptor Engagement

“…Vector supernatant was harvested 36, 48, and 72 h later, filtered through a 0.45-m filter, and pooled. For transduction, SIM-A9 microglia cells (57) were washed and resuspended in medium with 8 g/ml protamine sulfate (MP Biomedicals). Transductions took place overnight at 37°C followed by wash and maintenance in culture.…”

Fluorescent flavonoids for endoplasmic reticulum cell imaging

“…The ER selectivity was also examined on SIM-A9 cells (semi adherent microglial cells). 29 As a type of glial cell located throughout the brain, microglial cells account for 10–15% of all cells found within the brain, 30 and ER stress plays a significant role in the microglial cell death. 31 Although the ER distribution in GL261 cells is quite different from that in SIM-A9 cells (Fig.…”

An immortalized microglial cell line (Mocha) derived from rat cochlea

“…Mocha cells are different from existing microglial cell lines, such as the mouse brain-derived SIM-A9 (Nagamoto-Combs, et al, 2014), N9 (Righi, et al, 1989), and BV2 cell lines (Blasi, et al, 1990) as well as the human HMO6 cell line (Nagai, et al, 2001). Mocha cells are derived from the rat cochlea and are immortalized with the 12S E1A gene using a replication-incompetent retroviral vector, which allows for in vivo studies without the confounding issue of host retroviral infection.…”