Abstract
Background. In our model of dilated cardiomyopathy (DCM), cardiac dilatation and hypertrophy developed after inoculation of encephalomyocarditis virus (EMCV), but the infectious virus was isolated only early after infection. In this study, we investigated whether viral RNA could be detected at later times using the polymerase chain reaction (PCR). Methods and Results. In the in vitro study, FL (human amnion) cells infected with EMCV were harvested for RNA extraction, and viral cDNA was synthesized by reverse transcription with random hexamers. Using oligonucleotide primers with homology to the 5′ noncoding region of EMCV, we enzymatically amplified a 121-base pair band, which was homologous to a probe specific for EMCV as demonstrated by Southern blot hybridization. The sensitivity of this PCR technique was at the level of about 102-103 copies of viral RNA genome. In the in vivo study, four-week-old DBA/2 mice were inoculated with EMCV intraperitoneally (10 pfu/mouse) and killed on days 1, 2, 3, 5, 7, 10, 14, 18, 28, 60, and 90. The hearts were divided into three parts for purification of total RNA, histopathological examination, and to culture for infectious virus. The infectious virus was isolated from the heart after the second day but never after the 14th day. The viral genome was detectable by PCR on the second day, when very little mononuclear cell infiltration around the blood vessels was histologically visible. Positive PCR signals were observed in all hearts through day 14. Viral RNA was also detected in four of six 28-day samples, four of six 60-day samples, and two of seven 90-day samples when diffuse myocardial fibrosis was prominent, but myocardial necrosis or cellular infiltration had disappeared. Conclusions. The persistence of EMCV RNA was shown by PCR in the chronic stage of EMCV-induced myocarditis, a time when the inflammatory reaction had largely subsided. The PCR is a potentially useful method to test possible viral etiologies in idiopathic heart muscle disease or DCM.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 522-530 |
| Number of pages | 9 |
| Journal | Circulation |
| Volume | 86 |
| Issue number | 2 |
| DOIs | |
| State | Published - Aug 1992 |
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Keywords
- Cardiomyopathy, dilated
- Myocarditis, viral
- Polymerase chain reaction
- Virus, encephalomyocarditis
ASJC Scopus subject areas
- Cardiology and Cardiovascular Medicine
- Physiology (medical)
Cite this
Cardiac persistence of cardioviral RNA detected by polymerase chain reaction in a murine model of dilated cardiomyopathy. / Kyu, Bun Sho; Matsumori, Akira; Sato, Yukihito; Okada, Ikutaro; Chapman, Nora M.; Tracy, Steven.
In: Circulation, Vol. 86, No. 2, 08.1992, p. 522-530.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Cardiac persistence of cardioviral RNA detected by polymerase chain reaction in a murine model of dilated cardiomyopathy
AU - Kyu, Bun Sho
AU - Matsumori, Akira
AU - Sato, Yukihito
AU - Okada, Ikutaro
AU - Chapman, Nora M.
AU - Tracy, Steven
PY - 1992/8
Y1 - 1992/8
N2 - Background. In our model of dilated cardiomyopathy (DCM), cardiac dilatation and hypertrophy developed after inoculation of encephalomyocarditis virus (EMCV), but the infectious virus was isolated only early after infection. In this study, we investigated whether viral RNA could be detected at later times using the polymerase chain reaction (PCR). Methods and Results. In the in vitro study, FL (human amnion) cells infected with EMCV were harvested for RNA extraction, and viral cDNA was synthesized by reverse transcription with random hexamers. Using oligonucleotide primers with homology to the 5′ noncoding region of EMCV, we enzymatically amplified a 121-base pair band, which was homologous to a probe specific for EMCV as demonstrated by Southern blot hybridization. The sensitivity of this PCR technique was at the level of about 102-103 copies of viral RNA genome. In the in vivo study, four-week-old DBA/2 mice were inoculated with EMCV intraperitoneally (10 pfu/mouse) and killed on days 1, 2, 3, 5, 7, 10, 14, 18, 28, 60, and 90. The hearts were divided into three parts for purification of total RNA, histopathological examination, and to culture for infectious virus. The infectious virus was isolated from the heart after the second day but never after the 14th day. The viral genome was detectable by PCR on the second day, when very little mononuclear cell infiltration around the blood vessels was histologically visible. Positive PCR signals were observed in all hearts through day 14. Viral RNA was also detected in four of six 28-day samples, four of six 60-day samples, and two of seven 90-day samples when diffuse myocardial fibrosis was prominent, but myocardial necrosis or cellular infiltration had disappeared. Conclusions. The persistence of EMCV RNA was shown by PCR in the chronic stage of EMCV-induced myocarditis, a time when the inflammatory reaction had largely subsided. The PCR is a potentially useful method to test possible viral etiologies in idiopathic heart muscle disease or DCM.
AB - Background. In our model of dilated cardiomyopathy (DCM), cardiac dilatation and hypertrophy developed after inoculation of encephalomyocarditis virus (EMCV), but the infectious virus was isolated only early after infection. In this study, we investigated whether viral RNA could be detected at later times using the polymerase chain reaction (PCR). Methods and Results. In the in vitro study, FL (human amnion) cells infected with EMCV were harvested for RNA extraction, and viral cDNA was synthesized by reverse transcription with random hexamers. Using oligonucleotide primers with homology to the 5′ noncoding region of EMCV, we enzymatically amplified a 121-base pair band, which was homologous to a probe specific for EMCV as demonstrated by Southern blot hybridization. The sensitivity of this PCR technique was at the level of about 102-103 copies of viral RNA genome. In the in vivo study, four-week-old DBA/2 mice were inoculated with EMCV intraperitoneally (10 pfu/mouse) and killed on days 1, 2, 3, 5, 7, 10, 14, 18, 28, 60, and 90. The hearts were divided into three parts for purification of total RNA, histopathological examination, and to culture for infectious virus. The infectious virus was isolated from the heart after the second day but never after the 14th day. The viral genome was detectable by PCR on the second day, when very little mononuclear cell infiltration around the blood vessels was histologically visible. Positive PCR signals were observed in all hearts through day 14. Viral RNA was also detected in four of six 28-day samples, four of six 60-day samples, and two of seven 90-day samples when diffuse myocardial fibrosis was prominent, but myocardial necrosis or cellular infiltration had disappeared. Conclusions. The persistence of EMCV RNA was shown by PCR in the chronic stage of EMCV-induced myocarditis, a time when the inflammatory reaction had largely subsided. The PCR is a potentially useful method to test possible viral etiologies in idiopathic heart muscle disease or DCM.
KW - Cardiomyopathy, dilated
KW - Myocarditis, viral
KW - Polymerase chain reaction
KW - Virus, encephalomyocarditis
UR - http://www.scopus.com/inward/record.url?scp=0026667050&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0026667050&partnerID=8YFLogxK
U2 - 10.1161/01.CIR.86.2.522
DO - 10.1161/01.CIR.86.2.522
M3 - Article
C2 - 1322254
AN - SCOPUS:0026667050
VL - 86
SP - 522
EP - 530
JO - Circulation
JF - Circulation
SN - 0009-7322
IS - 2
ER -