Establishment of rat brain endothelial cells susceptible to rat cytomegalovirus ALL-03 infection
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tablishment of rat brain endothelial cells susceptible to rat cytomegalovirus ALL-03 infection Siti-Nazrina Camalxaman & Nazariah Allaudin Zeenathul & Yi-Wan Quah & Hwei-San Loh & Hassan Zuridah & Homayoun Hani & Abdul Rahman Sheikh-Omar & Mohd Lila Mohd-Azmi
Received: 6 June 2012 / Accepted: 7 September 2012 / Published online: 22 February 2013 / Editor: T. Okamoto # The Society for In Vitro Biology 2013
Abstract Endothelial cells have been implicated as key cells in promoting the pathogenesis and spread of cytomegalovirus (CMV) infection. This study describes the isolation and culture of rat brain endothelial cells (RBEC) and further evaluates the infectious potential of a Malaysian rat CMV (RCMV ALL-03) in these cultured cells. Brain tissues were mechanically fragmented, exposed to enzymatic digestion, purified by gradient density centrifugation, and cultured in vitro. Morphological characteristics and expression of von Willebrand factor (factor VIII-related antigen) verified the cells were of endothelial origin. RBEC were found to be permissive to the virus by cytopathic effects with detectable plaques formed within 7 d of infection. This was confirmed by electron microscopy examination which proved the existence of the viral particles in the infected cells. The susceptibility of the virus to these target cells under the experimental conditions described in this report provides a platform for developing a cell-culture-based experimental S.-N. Camalxaman : N. A. Zeenathul (*) : Y.-W. Quah : H. Hani : A. R. Sheikh-Omar : M. L. Mohd-Azmi Department of Veterinary Pathology and Microbiology, Faculty of Veterinary Medicine, Universiti Putra Malaysia (UPM), 43400, UPM, Serdang, Selangor, Malaysia e-mail: [email protected] H.-S. Loh School of Biosciences, The University of Nottingham Malaysia Campus, Jalan Broga, 43500 Semenyih, Selangor, Malaysia H. Zuridah Department of Medical Laboratory Technology, Faculty of Health Sciences, Universiti Teknologi MARA (UiTM), Kampus Puncak Alam, 42300 Bandar Puncak Alam, Selangor, Malaysia
model for studies of RCMV pathogenesis and allows stimulation of further studies on host cell responses imposed by congenital viral infections. Keywords Cytopathology . Isolation . Endothelial cells . Rat cytomegalovirus
Introduction Human cytomegalovirus (CMV) is a clinically significant herpes virus (Bissinger et al. 2002) that establishes a lifelong latent infection. Although relatively harmless in the healthy population, the virus is a public health nuisance and a major cause of distress in the immunoimmature or immunosuppressed individuals (Emery 2001; Varani and Landini 2011). As a consequence of maternofetal primary infection, human CMV infection may lead to severe neurological crisis and long-term disabilities which are often irreversible (Boppana et al. 1997; Ornoy and Diav-Citrin 2006; Cheeran et al. 2009). The virus is classically involved in a multifaceted interplay with the host and unanimously behaves in a parasitical manner. During early stages of infection, CMV dynamica
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