Chandipura virus infection in mice: the role of toll like receptor 4 in pathogenesis.
BACKGROUND
The susceptibility of mice and humans to Chandipura virus infection is age-dependent. Upon experimental infection, mice secrete significant amounts of proinflammatory cytokines. Similarly, children who recover from natural infection with the virus show significant amounts of TNF-α production, suggesting that innate immunity plays a major role in the response to Chandipura virus. Toll-like receptors (TLR) are key host molecules involved in innate immune responses in infections. Therefore, the aim of this study was to examine the role of TLR in the response to Chandipura virus infection.


METHODS
The mouse monocyte-macrophage cell line, RAW 264.7, and C3H/HeJ mice were used as models. Micro array techniques were used to identify the type of TLR involved in the response to infection. The results were validated by examining TLR expression using flow cytometry and by measuring the levels of proinflammatory cytokines and nitric oxide (NO) in the culture supernatants using bead assays and the Griess method, respectively. The pathogenic role of Toll-like receptor 4 (TLR4) was studied in a TLR4 mutant strain of mice -C3H/HeJ and the results compared with those from wild-type mice- C3H/CaJ. The pathogenic effects of NO were studied by treating experimentally infected mice with the NO inhibitor, aminoguanidine (AG).


RESULTS
The micro array results showed that TLR4 was regulated after Chandipura virus infection. At high multiplicities of infection (10 MOI), RAW cells up- regulated cell surface expression of TLR4 and secreted significant amounts of TNF-α, MCP-1, IL-10 and IL-12 and NO. The survival rate of C3H/HeJ mice was higher than those of wild-type C3H/CaJ mice. The survived C3H/HeJ mice secreted significant quantity of MCP-1 and IFN-γ cytokines and cleared virus from brain. Similarly, the survival rate of AG-treated mice was higher than those of the untreated controls.


CONCLUSIONS
Chandipura virus regulates TLR4, which leads to the secretion of proinflammatory cytokines and NO by infected RAW cells. Difference in survival rate in TLR4 mutant mice and nitric oxide inhibitor treated mice, confirmed the role of these molecules in disease pathogenesis. The result is significant in clinical management and designing antiviral intervention for Chandipura virus infection.
National Institute of Virology Kerala Unit, Medical College Hospital Complex, Vandanam, Alappuzha, Kerala 688005, India. anukumar@icmr.org.in
Balakrishnan
B
Anukumar
Prajakta
P
Shahir
D000818:Animals; D002460:Cell Line; D016207:Cytokines / Q000378:metabolism; D004195:Disease Models, Animal; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D008264:Macrophages / Q000276:immunology / Q000821:virology; D051379:Mice; D008809:Mice, Inbred C3H; D046228:Microarray Analysis; D009569:Nitric Oxide / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000172:deficiency / Q000276:immunology*; D018116:Vesiculovirus / Q000472:pathogenicity*
D016207:Cytokines; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; D009569:Nitric Oxide