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  • Silencing the Cytoskeleton Protein Iba1 (Ionized Calcium Binding Adapter Protein 1) Interferes with BV2 Microglia Functioning.

Silencing the Cytoskeleton Protein Iba1 (Ionized Calcium Binding Adapter Protein 1) Interferes with BV2 Microglia Functioning.

Cellular and molecular neurobiology (2020-01-18)
Roxana-Olimpia Gheorghe, Alexandru Deftu, Alexandru Filippi, Andreea Grosu, Melania Bica-Popi, Marioara Chiritoiu, Gabriela Chiritoiu, Cristian Munteanu, Luigi Silvestro, Violeta Ristoiu
ABSTRACT

Iba1 (ionized calcium binding adapter protein 1) is a cytoskeleton protein specific only for microglia and macrophages, where it acts as an actin-cross linking protein. Although frequently regarded as a marker of activation, its involvement in cell migration, membrane ruffling, phagocytosis or in microglia remodeling during immunological surveillance of the brain suggest that Iba1 is not a simple cytoskeleton protein, but a signaling molecule involved in specific signaling pathways. In this study we investigated if Iba1 could also represent a drug target, and tested the hypothesis that its specific silencing with customized Iba1-siRNA can modulate microglia functioning. The results showed that Iba1-silenced BV2 microglia migrate less due to reduced proliferation and cell adhesion, while their phagocytic activity and P2x7 functioning was significantly increased. Our data are the proof of concept that Iba1 protein is a new microglia target, which opens a new therapeutic avenue for modulating microglia behavior.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Thiazolyl Blue Tetrazolium Bromide, 98%
Sigma-Aldrich
GenElute Mammalian Total RNA Miniprep Kit, sufficient for 70 purifications
Sigma-Aldrich
Trypsin-EDTA solution, 0.25%, sterile-filtered, BioReagent, suitable for cell culture, 2.5 g porcine trypsin and 0.2 g EDTA, 4Na per liter of Hanks′ Balanced Salt Solution with phenol red
Sigma-Aldrich
Penicillin-Streptomycin, Solution Stabilized, with 5,000 units penicillin and 5mg streptomycin/mL, 0.1 μm filtered, BioReagent, suitable for cell culture
Sigma-Aldrich
Formaldehyde solution, ACS reagent, 37 wt. % in H2O, contains 10-15% Methanol as stabilizer (to prevent polymerization)
Sigma-Aldrich
On-Column DNase I Digestion Set, sufficient for 70 preparations
Sigma-Aldrich
Pyrvinium pamoate salt hydrate, ≥98% (HPLC)