J Bacteriol Virol.  2016 Dec;46(4):221-230. 10.4167/jbv.2016.46.4.221.

Role of Rotavirus Enterotoxin NSP4 in the Inflammatory Response in Murine Macrophage RAW 264.7 Cells

• Kim SA
• Than VT
• Kim W

• KMID: 2366867
• DOI: http://doi.org/10.4167/jbv.2016.46.4.221

Abstract

The rotavirus nonstructural glycoprotein, NSP4, has been identified as the first viral enterotoxin capable of inducing diarrhea. To investigate the biological function of NSP4 in the inflammatory process, a cDNA from human rotavirus (Wa strain) RNA segment 10 was amplified by RT-PCR, cloned into TA vector, and subsequently subcloned into pET23b expression plasmid. The expression of NSP4 protein was determined by SDS-PAGE and Western blotting, then, the protein was purified by affinity chromatography on Ni-NTA-agarose column. The inflammatory effects of NSP4, namely, production of nitric oxide (NO), pro-inflammatory cytokines (IL-1β, IL-6, IL-10, and TNF-α), and prostaglandin E2 (PGE₂), was evaluated using NSP4-stimulated RAW 264.7 murine macrophages and compared with those observed after stimulation with lipopolysaccharide (LPS). The levels of IL-1β, IL-6, and TNF-α were significantly increased, and those of NO and PGE₂ also increased in NSP4-stimulated RAW 264.7 cells. These findings indicate that NSP4 plays an important role in the inflammatory response observed during rotavirus infection.

Keyword

Rotavirus; NSP4; RAW 264.7 murine macrophage; Inflammation

MeSH Terms

Blotting, Western
Chromatography, Affinity
Clone Cells
Cytokines
Diarrhea
Dinoprostone
DNA, Complementary
Electrophoresis, Polyacrylamide Gel
Enterotoxins*
Glycoproteins
Humans
Inflammation
Interleukin-10
Interleukin-6
Macrophages*
Nitric Oxide
Plasmids
RAW 264.7 Cells*
RNA
Rotavirus Infections
Rotavirus*
Cytokines
DNA, Complementary
Dinoprostone
Enterotoxins
Glycoproteins
Interleukin-10
Interleukin-6
Nitric Oxide
RNA