Nitric oxide increases the amount of murine cytomegalovirus-DNA in mice latently infected with the virus

Abstract

At 5 wk after the intraperitoneal infection of murine cytomegalovirus (MCMV) in adult (C3H/He x BALB/c) F1 (CBF1) mice, MCMV-DNA was noted mainly in the salivary glands (SG). Since graft-versus-host reaction (GVHR) is well known as a major risk factor for CMV reactivation, parental BALB/c spleen cells were then intravenously transferred to these mice to induce GVHR. A high copy number of full-length MCMV-DNA was thus observed in the lungs as well as the hearts and livers at 4 to 6 wk after the cell transfer, whereas the copy numbers in SG after the cell transfer were comparable to those before the transfer. It was therefore suggested that the amount of MCMV-DNA noted in these organs had been increased by GVHR. However, no transcripts of MCMV-DNA could be detected. Since a marked expression of mRNA of inducible nitric oxide synthetase (iNOS) was noted in the lungs and the hearts during the course of GVHR, the role of NO in the increase of MCMV-DNA after the cell transfer was next examined. The administration of L-arginine, a substrate for NO, increased the amount of MCMV-DNA in the lungs and the hearts after the induction of GVHR. In contrast, the material which suppressed iNOS induction inhibited the emergence of MCMV-DNA. Therefore, the NO generated during the course of GVHR increased the amount of the viral DNA in the lungs and the hearts of the mice latently infected with MCMV. If the increase in the amount of viral genome is the initial step of viral reactivation from the latent state, then NO would thus trigger the initial step of the reactivation of the DNA virus. As the lungs are the major organ to produce NO after systemic stimulation, the observations of this study may explain why the lungs are the major site of CMV latency and why GVHR triggers CMV reactivation. These findings are thus considered to demonstrate that NO plays a novel role in viral infection.

Role of Nitric Oxide Synthase Type 2 in Acute Infection with Murine Cytomegalovirus

“…These cells after a new in vitro infection (multiplicity of infection (MOI), 0.005) were cultured in a 24-well plate at 1 ϫ 10 7 /well for 24 h at 37°C in a 5% CO 2 atmosphere, and then the culture supernatants were harvested. The measurement of NO (NO 2 Ϫ /NO 3 Ϫ ) in the supernatants was performed using the assay kit (NO 2 /NO 3 assay kit-Fluometric; Dojindo, Kumamoto, Japan) as described previously (43). In brief, 80 l of the sample were added to the well of a 96-well fluoroplate (Fluoroplate LS9502867; Iwaki Glass, Cjiba, Japan).…”

“…In short, the MCMV-DNA of the immediate early (IE) region was enhanced by PCR using two oligonucleotide primers (primer 1, 1701-1730; primer 2, 2371-2400) as reported previously (43). This primer pair amplified the 700-bp segment.…”

Therapeutic effect of a traditional Chinese medicine, ren-shen-yang-rong-tang (Japanese name: Ninjin'yoeito) on nitric oxide-mediated lung injury in a mouse infected with murine cytomegalovirus

“…The measurement of the serum level of NO 2 À /NO 3 À was done by a fluorometric analysis as previously reported [14]. In brief, the serum samples were filtered through a 3000 Mr cutoff filter (Ultrafree C3; Japan Milipore Ltd., Tokyo, Japan) to eliminate the protein component, and then was stocked at À 80 8C until assay.…”