Real-time polymerase chain reaction and laser capture microdissection

An efficient combination tool for Chlamydophila pneumoniae DNA quantification and localization of infection in atherosclerotic lesions

Alessandra Ciervo, F. Mancini, P. Sale, A. Russo, A. Cassone

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Chlamydophila pneumoniae has been implicated in atherosclerosis, but the role of this obligate intracellular pathogen in the development of the above pathology is still unclear. In particular, its presence and quantitative distribution within lesional areas has not yet been defined. We studied 18 carotid biopsies obtained from patients undergoing endoartherectomy. By laser microdissection (LCM), two different sites (intra-plaque and plaque-adjacent areas) were taken from each lesion, and the presence and quantity of the pathogen DNA were determined by real-time polymerase chain reaction (Real-time PCR). A total of 8 plaques, exclusively, from patients with unstable angina, were positive in real-time PCR. The bacterial DNA was detected in both lesional areas of 3 plaques which contained the highest number of DNA copies (1,900 to 2,200 copy numbers), while C. pneumoniae DNA was detected only in the intra-plaque area of the other 5 positive (500 to 1,600 copy numbers). No C. pneumoniae DNA was found in the other 10 plaques of which 6 were from patients with unstable angina and 4 from stable angina patients. No DNA from Helicobacter pylori or Cytomegalovirus was found in any plaque. This is the first report where both the target lesion and an adjacent reference site were evaluated for the presence of C. pneumoniae DNA by the combination of LCM and Real-time PCR assays. The integration of these two methodologies offer an excellent tool for in situ studies and may help to elucidate the putative role of C. pneumoniae in atherosclerosis.

Original languageEnglish
Pages (from-to)421-428
Number of pages8
JournalInternational Journal of Immunopathology and Pharmacology
Volume21
Issue number2
Publication statusPublished - Apr 2008

Fingerprint

Laser Capture Microdissection
Chlamydophila pneumoniae
Real-Time Polymerase Chain Reaction
DNA
Infection
Unstable Angina
Atherosclerosis
Bacterial DNA
Microdissection
Stable Angina
Cytomegalovirus
Helicobacter pylori
Lasers
Pathology
Biopsy

Keywords

  • Atheromatous plaques
  • Chlamydophila pneumoniae
  • Endoartherectomy
  • Laser capture microdissection
  • Real-time PCR

ASJC Scopus subject areas

  • Pharmacology

Cite this

@article{e91af4c0b9b0466aa099697b79f5cb93,
title = "Real-time polymerase chain reaction and laser capture microdissection: An efficient combination tool for Chlamydophila pneumoniae DNA quantification and localization of infection in atherosclerotic lesions",
abstract = "Chlamydophila pneumoniae has been implicated in atherosclerosis, but the role of this obligate intracellular pathogen in the development of the above pathology is still unclear. In particular, its presence and quantitative distribution within lesional areas has not yet been defined. We studied 18 carotid biopsies obtained from patients undergoing endoartherectomy. By laser microdissection (LCM), two different sites (intra-plaque and plaque-adjacent areas) were taken from each lesion, and the presence and quantity of the pathogen DNA were determined by real-time polymerase chain reaction (Real-time PCR). A total of 8 plaques, exclusively, from patients with unstable angina, were positive in real-time PCR. The bacterial DNA was detected in both lesional areas of 3 plaques which contained the highest number of DNA copies (1,900 to 2,200 copy numbers), while C. pneumoniae DNA was detected only in the intra-plaque area of the other 5 positive (500 to 1,600 copy numbers). No C. pneumoniae DNA was found in the other 10 plaques of which 6 were from patients with unstable angina and 4 from stable angina patients. No DNA from Helicobacter pylori or Cytomegalovirus was found in any plaque. This is the first report where both the target lesion and an adjacent reference site were evaluated for the presence of C. pneumoniae DNA by the combination of LCM and Real-time PCR assays. The integration of these two methodologies offer an excellent tool for in situ studies and may help to elucidate the putative role of C. pneumoniae in atherosclerosis.",
keywords = "Atheromatous plaques, Chlamydophila pneumoniae, Endoartherectomy, Laser capture microdissection, Real-time PCR",
author = "Alessandra Ciervo and F. Mancini and P. Sale and A. Russo and A. Cassone",
year = "2008",
month = "4",
language = "English",
volume = "21",
pages = "421--428",
journal = "International Journal of Immunopathology and Pharmacology",
issn = "0394-6320",
publisher = "Biomedical Research Press s.a.s.",
number = "2",

}

TY - JOUR

T1 - Real-time polymerase chain reaction and laser capture microdissection

T2 - An efficient combination tool for Chlamydophila pneumoniae DNA quantification and localization of infection in atherosclerotic lesions

AU - Ciervo, Alessandra

AU - Mancini, F.

AU - Sale, P.

AU - Russo, A.

AU - Cassone, A.

PY - 2008/4

Y1 - 2008/4

N2 - Chlamydophila pneumoniae has been implicated in atherosclerosis, but the role of this obligate intracellular pathogen in the development of the above pathology is still unclear. In particular, its presence and quantitative distribution within lesional areas has not yet been defined. We studied 18 carotid biopsies obtained from patients undergoing endoartherectomy. By laser microdissection (LCM), two different sites (intra-plaque and plaque-adjacent areas) were taken from each lesion, and the presence and quantity of the pathogen DNA were determined by real-time polymerase chain reaction (Real-time PCR). A total of 8 plaques, exclusively, from patients with unstable angina, were positive in real-time PCR. The bacterial DNA was detected in both lesional areas of 3 plaques which contained the highest number of DNA copies (1,900 to 2,200 copy numbers), while C. pneumoniae DNA was detected only in the intra-plaque area of the other 5 positive (500 to 1,600 copy numbers). No C. pneumoniae DNA was found in the other 10 plaques of which 6 were from patients with unstable angina and 4 from stable angina patients. No DNA from Helicobacter pylori or Cytomegalovirus was found in any plaque. This is the first report where both the target lesion and an adjacent reference site were evaluated for the presence of C. pneumoniae DNA by the combination of LCM and Real-time PCR assays. The integration of these two methodologies offer an excellent tool for in situ studies and may help to elucidate the putative role of C. pneumoniae in atherosclerosis.

AB - Chlamydophila pneumoniae has been implicated in atherosclerosis, but the role of this obligate intracellular pathogen in the development of the above pathology is still unclear. In particular, its presence and quantitative distribution within lesional areas has not yet been defined. We studied 18 carotid biopsies obtained from patients undergoing endoartherectomy. By laser microdissection (LCM), two different sites (intra-plaque and plaque-adjacent areas) were taken from each lesion, and the presence and quantity of the pathogen DNA were determined by real-time polymerase chain reaction (Real-time PCR). A total of 8 plaques, exclusively, from patients with unstable angina, were positive in real-time PCR. The bacterial DNA was detected in both lesional areas of 3 plaques which contained the highest number of DNA copies (1,900 to 2,200 copy numbers), while C. pneumoniae DNA was detected only in the intra-plaque area of the other 5 positive (500 to 1,600 copy numbers). No C. pneumoniae DNA was found in the other 10 plaques of which 6 were from patients with unstable angina and 4 from stable angina patients. No DNA from Helicobacter pylori or Cytomegalovirus was found in any plaque. This is the first report where both the target lesion and an adjacent reference site were evaluated for the presence of C. pneumoniae DNA by the combination of LCM and Real-time PCR assays. The integration of these two methodologies offer an excellent tool for in situ studies and may help to elucidate the putative role of C. pneumoniae in atherosclerosis.

KW - Atheromatous plaques

KW - Chlamydophila pneumoniae

KW - Endoartherectomy

KW - Laser capture microdissection

KW - Real-time PCR

UR - http://www.scopus.com/inward/record.url?scp=47249102044&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=47249102044&partnerID=8YFLogxK

M3 - Article

VL - 21

SP - 421

EP - 428

JO - International Journal of Immunopathology and Pharmacology

JF - International Journal of Immunopathology and Pharmacology

SN - 0394-6320

IS - 2

ER -