Use of quantitative real-time PCR (qRT-PCR) to measure cytokine transcription and viral load in murine cytomegalovirus infection

Yajarayma J. Tang-Feldman, Angela Wojtowicz, G. Raymond Lochhead, Merica A. Hale, Yueju Li, Claire Pomeroy

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

A quantitative real-time PCR (qRT-PCR) assay was developed to measure cytokine transcription profiles and viral load during sub-clinical and clinical infection with murine cytomegalovirus (MCMV). Primers/fluorogenic probes specific for mouse cytokines and for the immediate early gene 1 (IE1) of MCMV were used to quantitate cytokine responses and viral load in various organs of MCMV infected mice. Increased mRNA levels of TNF-α, INF-γ and IL-10 were detected in the spleens, lungs and livers of clinically infected mice at 5 days post-infection. Transcription of these cytokines was 2-5-fold lower (p = 0.07 for each cytokine) in the spleens and 10-100-fold lower in the lungs (p = 0.03 for INFγ, not significant for IL-10 and TNFα) and livers (p < 0.05 for each cytokine) of sub-clinically infected mice. Clinical MCMV infection induced high levels of IL-6 in the lungs and spleens of infected animals, while no significant transcription of IL-6 was detected in any organ during sub-clinical infection (p < 0.05). The timing of peak amounts of INF-γ, IL-10 and IL-6 observed in the spleens of clinically infected mice correlated with high viral loads in these organs. Cytokine expression rose in the salivary glands later, at day 15, corresponding to the increase in salivary gland viral load. The qRT-PCR demonstrates that infection with MCMV induces an organ-specific cytokine response characterized by the production of TNF-α, INF-γ, IL-6 and IL-10 which correlates with severity of the disease (sub-clinical versus clinical) and with viral load. In summary, qRT-PCR is a sensitive and accurate method to study MCMV infection and host responses to the virus.

Original languageEnglish (US)
Pages (from-to)122-129
Number of pages8
JournalJournal of Virological Methods
Volume131
Issue number2
DOIs
StatePublished - Feb 2006

Fingerprint

Muromegalovirus
Cytomegalovirus Infections
Viral Load
Real-Time Polymerase Chain Reaction
Cytokines
Interleukin-10
Interleukin-6
Spleen
Infection
Salivary Glands
Lung
Immediate-Early Genes
Liver
Viruses
Messenger RNA

Keywords

  • Cytokine response
  • MCMV
  • Quantitative RT-PCR
  • Viral load

ASJC Scopus subject areas

  • Virology

Cite this

Use of quantitative real-time PCR (qRT-PCR) to measure cytokine transcription and viral load in murine cytomegalovirus infection. / Tang-Feldman, Yajarayma J.; Wojtowicz, Angela; Lochhead, G. Raymond; Hale, Merica A.; Li, Yueju; Pomeroy, Claire.

In: Journal of Virological Methods, Vol. 131, No. 2, 02.2006, p. 122-129.

Research output: Contribution to journalArticle

Tang-Feldman, Yajarayma J. ; Wojtowicz, Angela ; Lochhead, G. Raymond ; Hale, Merica A. ; Li, Yueju ; Pomeroy, Claire. / Use of quantitative real-time PCR (qRT-PCR) to measure cytokine transcription and viral load in murine cytomegalovirus infection. In: Journal of Virological Methods. 2006 ; Vol. 131, No. 2. pp. 122-129.
@article{212f8662130c4ea7a63f67dfd1278df5,
title = "Use of quantitative real-time PCR (qRT-PCR) to measure cytokine transcription and viral load in murine cytomegalovirus infection",
abstract = "A quantitative real-time PCR (qRT-PCR) assay was developed to measure cytokine transcription profiles and viral load during sub-clinical and clinical infection with murine cytomegalovirus (MCMV). Primers/fluorogenic probes specific for mouse cytokines and for the immediate early gene 1 (IE1) of MCMV were used to quantitate cytokine responses and viral load in various organs of MCMV infected mice. Increased mRNA levels of TNF-α, INF-γ and IL-10 were detected in the spleens, lungs and livers of clinically infected mice at 5 days post-infection. Transcription of these cytokines was 2-5-fold lower (p = 0.07 for each cytokine) in the spleens and 10-100-fold lower in the lungs (p = 0.03 for INFγ, not significant for IL-10 and TNFα) and livers (p < 0.05 for each cytokine) of sub-clinically infected mice. Clinical MCMV infection induced high levels of IL-6 in the lungs and spleens of infected animals, while no significant transcription of IL-6 was detected in any organ during sub-clinical infection (p < 0.05). The timing of peak amounts of INF-γ, IL-10 and IL-6 observed in the spleens of clinically infected mice correlated with high viral loads in these organs. Cytokine expression rose in the salivary glands later, at day 15, corresponding to the increase in salivary gland viral load. The qRT-PCR demonstrates that infection with MCMV induces an organ-specific cytokine response characterized by the production of TNF-α, INF-γ, IL-6 and IL-10 which correlates with severity of the disease (sub-clinical versus clinical) and with viral load. In summary, qRT-PCR is a sensitive and accurate method to study MCMV infection and host responses to the virus.",
keywords = "Cytokine response, MCMV, Quantitative RT-PCR, Viral load",
author = "Tang-Feldman, {Yajarayma J.} and Angela Wojtowicz and Lochhead, {G. Raymond} and Hale, {Merica A.} and Yueju Li and Claire Pomeroy",
year = "2006",
month = "2",
doi = "10.1016/j.jviromet.2005.07.013",
language = "English (US)",
volume = "131",
pages = "122--129",
journal = "Journal of Virological Methods",
issn = "0166-0934",
publisher = "Elsevier",
number = "2",

}

TY - JOUR

T1 - Use of quantitative real-time PCR (qRT-PCR) to measure cytokine transcription and viral load in murine cytomegalovirus infection

AU - Tang-Feldman, Yajarayma J.

AU - Wojtowicz, Angela

AU - Lochhead, G. Raymond

AU - Hale, Merica A.

AU - Li, Yueju

AU - Pomeroy, Claire

PY - 2006/2

Y1 - 2006/2

N2 - A quantitative real-time PCR (qRT-PCR) assay was developed to measure cytokine transcription profiles and viral load during sub-clinical and clinical infection with murine cytomegalovirus (MCMV). Primers/fluorogenic probes specific for mouse cytokines and for the immediate early gene 1 (IE1) of MCMV were used to quantitate cytokine responses and viral load in various organs of MCMV infected mice. Increased mRNA levels of TNF-α, INF-γ and IL-10 were detected in the spleens, lungs and livers of clinically infected mice at 5 days post-infection. Transcription of these cytokines was 2-5-fold lower (p = 0.07 for each cytokine) in the spleens and 10-100-fold lower in the lungs (p = 0.03 for INFγ, not significant for IL-10 and TNFα) and livers (p < 0.05 for each cytokine) of sub-clinically infected mice. Clinical MCMV infection induced high levels of IL-6 in the lungs and spleens of infected animals, while no significant transcription of IL-6 was detected in any organ during sub-clinical infection (p < 0.05). The timing of peak amounts of INF-γ, IL-10 and IL-6 observed in the spleens of clinically infected mice correlated with high viral loads in these organs. Cytokine expression rose in the salivary glands later, at day 15, corresponding to the increase in salivary gland viral load. The qRT-PCR demonstrates that infection with MCMV induces an organ-specific cytokine response characterized by the production of TNF-α, INF-γ, IL-6 and IL-10 which correlates with severity of the disease (sub-clinical versus clinical) and with viral load. In summary, qRT-PCR is a sensitive and accurate method to study MCMV infection and host responses to the virus.

AB - A quantitative real-time PCR (qRT-PCR) assay was developed to measure cytokine transcription profiles and viral load during sub-clinical and clinical infection with murine cytomegalovirus (MCMV). Primers/fluorogenic probes specific for mouse cytokines and for the immediate early gene 1 (IE1) of MCMV were used to quantitate cytokine responses and viral load in various organs of MCMV infected mice. Increased mRNA levels of TNF-α, INF-γ and IL-10 were detected in the spleens, lungs and livers of clinically infected mice at 5 days post-infection. Transcription of these cytokines was 2-5-fold lower (p = 0.07 for each cytokine) in the spleens and 10-100-fold lower in the lungs (p = 0.03 for INFγ, not significant for IL-10 and TNFα) and livers (p < 0.05 for each cytokine) of sub-clinically infected mice. Clinical MCMV infection induced high levels of IL-6 in the lungs and spleens of infected animals, while no significant transcription of IL-6 was detected in any organ during sub-clinical infection (p < 0.05). The timing of peak amounts of INF-γ, IL-10 and IL-6 observed in the spleens of clinically infected mice correlated with high viral loads in these organs. Cytokine expression rose in the salivary glands later, at day 15, corresponding to the increase in salivary gland viral load. The qRT-PCR demonstrates that infection with MCMV induces an organ-specific cytokine response characterized by the production of TNF-α, INF-γ, IL-6 and IL-10 which correlates with severity of the disease (sub-clinical versus clinical) and with viral load. In summary, qRT-PCR is a sensitive and accurate method to study MCMV infection and host responses to the virus.

KW - Cytokine response

KW - MCMV

KW - Quantitative RT-PCR

KW - Viral load

UR - http://www.scopus.com/inward/record.url?scp=30644479574&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=30644479574&partnerID=8YFLogxK

U2 - 10.1016/j.jviromet.2005.07.013

DO - 10.1016/j.jviromet.2005.07.013

M3 - Article

VL - 131

SP - 122

EP - 129

JO - Journal of Virological Methods

JF - Journal of Virological Methods

SN - 0166-0934

IS - 2

ER -