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Vet Res Commun (2012) 36:195199 DOI 10.1007/s11259-012-9525-y
SHORT COMMUNICATION
The role of cytoskeleton, components of inositol phospholipid signaling pathway and iron in Ehrlichia canis in vitro proliferation
Marcelo Arantes Levenhagen &
Rosiane Nascimento Alves & Susana Elisa Rieck &
Marcelo Bahia Labruna & Marcelo Emlio Beletti
Accepted: 2 March 2012 /Published online: 14 March 2012 # Springer Science+Business Media B.V. 2012
Abstract Ehrlichia canis, etiologic agent of Canine Monocytic Ehrlichiosis, is an obligatory intracellular bacterium that parasitizes monocytes and macrophages. In this study we analyzed the role of the cytoskeleton specifically actin microfilaments and microtubules, components of inositol phospholipid signaling pathway such as phospholipase C (PLC), protein kinase (PTK) and calcium channels as well as the role of iron in the E. canis proliferation in DH82 cells. Different inhibitory compounds were used for each component: Cytochalasin D (inhibits actin polymerization), Nocodazole (inhibits microtubule polymerization), Neomycin (PLC inhibitor), Genistein (PTK inhibitor), Verapamil (calcium channel blocker) and Deferoxamine (iron chelator). We observed a significant decrease in the total number of bacteria in infected cells treated suggesting that these cellular components analized are essentials to E. canis proliferation.
Keywords Cell signaling . Ehrlichia canis . DH82 cells
Introduction
Ehrlichia canis, etiologic agent of Canine Monocytic Ehrlichiosis (CME), is a pleomorphic Gram-negative, obliga-tory intracellular bacterium found isolated (elementary bodies) or in compact inclusions within the cytoplasmic vacuoles (morulae) of monocytes and macrophages (Dumler et al. 2001).
The cellular internalization, proliferation and propagation processes of Ehrlichia sp. involves the activation of different cell signaling pathways specific to each bacterium, culminating in nutrient acquisition and lysosomal evasion (Rikihisa 2006). Inclusions containing Ehrlichia chaffeensis exhibit characteristics of early endosomes, including transferrin receptor (TfR). Importantly, TfR is required for lysosomal avoidance and iron acquisition, and both are important for bacterial multiplication as well as spread to other cells (Rikihisa 2010).
Other cell signaling mechanisms in E. chaffensis include phospholipase C (PLC)-2 activation, tyrosine phosphorylation, inositol 1,4,5-triphosphate (IP3) production and a moderate increase in intracellular calcium (Rikihisa 2006). The cytoskeleton is also involved in internalization and proliferation, as this process is inhibited by taxol and colchicines (microtubules inhibitors), and cytochalasin D (an actin filament inhibitor) (Lin et al. 2002). Rikihisa et al. (1994), observed that cytoskeleton envelopment also occurs during the cellular trafficking of...