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Noninvasive Monitoring of Placenta-Specific Transgene Expression by Bioluminescence Imaging

BACKGROUND: Placental dysfunction underlies numerous complications of pregnancy. A major obstacle to understanding the roles of potential mediators of placental pathology has been the absence of suitable methods for tissue-specific gene manipulation and sensitive assays for studying gene functions i...

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Autores principales: Fan, Xiujun, Ren, Peigen, Dhal, Sabita, Bejerano, Gill, Goodman, Stuart B., Druzin, Maurice L., Gambhir, Sanjiv S., Nayak, Nihar R.
Formato: Texto
Lenguaje:English
Publicado: Public Library of Science 2011
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3025029/
https://www.ncbi.nlm.nih.gov/pubmed/21283713
http://dx.doi.org/10.1371/journal.pone.0016348
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author Fan, Xiujun
Ren, Peigen
Dhal, Sabita
Bejerano, Gill
Goodman, Stuart B.
Druzin, Maurice L.
Gambhir, Sanjiv S.
Nayak, Nihar R.
author_facet Fan, Xiujun
Ren, Peigen
Dhal, Sabita
Bejerano, Gill
Goodman, Stuart B.
Druzin, Maurice L.
Gambhir, Sanjiv S.
Nayak, Nihar R.
author_sort Fan, Xiujun
collection PubMed
description BACKGROUND: Placental dysfunction underlies numerous complications of pregnancy. A major obstacle to understanding the roles of potential mediators of placental pathology has been the absence of suitable methods for tissue-specific gene manipulation and sensitive assays for studying gene functions in the placentas of intact animals. We describe a sensitive and noninvasive method of repetitively tracking placenta-specific gene expression throughout pregnancy using lentivirus-mediated transduction of optical reporter genes in mouse blastocysts. METHODOLOGY/PRINCIPAL FINDINGS: Zona-free blastocysts were incubated with lentivirus expressing firefly luciferase (Fluc) and Tomato fluorescent fusion protein for trophectoderm-specific infection and transplanted into day 3 pseudopregnant recipients (GD3). Animals were examined for Fluc expression by live bioluminescence imaging (BLI) at different points during pregnancy, and the placentas were examined for tomato expression in different cell types on GD18. In another set of experiments, blastocysts with maximum photon fluxes in the range of 2.0E+4 to 6.0E+4 p/s/cm(2)/sr were transferred. Fluc expression was detectable in all surrogate dams by day 5 of pregnancy by live imaging, and the signal increased dramatically thereafter each day until GD12, reaching a peak at GD16 and maintaining that level through GD18. All of the placentas, but none of the fetuses, analyzed on GD18 by BLI showed different degrees of Fluc expression. However, only placentas of dams transferred with selected blastocysts showed uniform photon distribution with no significant variability of photon intensity among placentas of the same litter. Tomato expression in the placentas was limited to only trophoblast cell lineages. CONCLUSIONS/SIGNIFICANCE: These results, for the first time, demonstrate the feasibility of selecting lentivirally-transduced blastocysts for uniform gene expression in all placentas of the same litter and early detection and quantitative analysis of gene expression throughout pregnancy by live BLI. This method may be useful for a wide range of applications involving trophoblast-specific gene manipulations in utero.
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spelling pubmed-30250292011-01-31 Noninvasive Monitoring of Placenta-Specific Transgene Expression by Bioluminescence Imaging Fan, Xiujun Ren, Peigen Dhal, Sabita Bejerano, Gill Goodman, Stuart B. Druzin, Maurice L. Gambhir, Sanjiv S. Nayak, Nihar R. PLoS One Research Article BACKGROUND: Placental dysfunction underlies numerous complications of pregnancy. A major obstacle to understanding the roles of potential mediators of placental pathology has been the absence of suitable methods for tissue-specific gene manipulation and sensitive assays for studying gene functions in the placentas of intact animals. We describe a sensitive and noninvasive method of repetitively tracking placenta-specific gene expression throughout pregnancy using lentivirus-mediated transduction of optical reporter genes in mouse blastocysts. METHODOLOGY/PRINCIPAL FINDINGS: Zona-free blastocysts were incubated with lentivirus expressing firefly luciferase (Fluc) and Tomato fluorescent fusion protein for trophectoderm-specific infection and transplanted into day 3 pseudopregnant recipients (GD3). Animals were examined for Fluc expression by live bioluminescence imaging (BLI) at different points during pregnancy, and the placentas were examined for tomato expression in different cell types on GD18. In another set of experiments, blastocysts with maximum photon fluxes in the range of 2.0E+4 to 6.0E+4 p/s/cm(2)/sr were transferred. Fluc expression was detectable in all surrogate dams by day 5 of pregnancy by live imaging, and the signal increased dramatically thereafter each day until GD12, reaching a peak at GD16 and maintaining that level through GD18. All of the placentas, but none of the fetuses, analyzed on GD18 by BLI showed different degrees of Fluc expression. However, only placentas of dams transferred with selected blastocysts showed uniform photon distribution with no significant variability of photon intensity among placentas of the same litter. Tomato expression in the placentas was limited to only trophoblast cell lineages. CONCLUSIONS/SIGNIFICANCE: These results, for the first time, demonstrate the feasibility of selecting lentivirally-transduced blastocysts for uniform gene expression in all placentas of the same litter and early detection and quantitative analysis of gene expression throughout pregnancy by live BLI. This method may be useful for a wide range of applications involving trophoblast-specific gene manipulations in utero. Public Library of Science 2011-01-21 /pmc/articles/PMC3025029/ /pubmed/21283713 http://dx.doi.org/10.1371/journal.pone.0016348 Text en Fan et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Fan, Xiujun
Ren, Peigen
Dhal, Sabita
Bejerano, Gill
Goodman, Stuart B.
Druzin, Maurice L.
Gambhir, Sanjiv S.
Nayak, Nihar R.
Noninvasive Monitoring of Placenta-Specific Transgene Expression by Bioluminescence Imaging
title Noninvasive Monitoring of Placenta-Specific Transgene Expression by Bioluminescence Imaging
title_full Noninvasive Monitoring of Placenta-Specific Transgene Expression by Bioluminescence Imaging
title_fullStr Noninvasive Monitoring of Placenta-Specific Transgene Expression by Bioluminescence Imaging
title_full_unstemmed Noninvasive Monitoring of Placenta-Specific Transgene Expression by Bioluminescence Imaging
title_short Noninvasive Monitoring of Placenta-Specific Transgene Expression by Bioluminescence Imaging
title_sort noninvasive monitoring of placenta-specific transgene expression by bioluminescence imaging
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3025029/
https://www.ncbi.nlm.nih.gov/pubmed/21283713
http://dx.doi.org/10.1371/journal.pone.0016348
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