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MRI detection of transcriptional regulation of gene expression in transgenic mice. Batya Cohen, Keren Ziv, Vicki Plaks, Tomer Israely, Vyacheslav Kalchenko, Alon Harmelin, Laura Benjamin, and Michal Neeman. Derek Ju and John Kucharczyk 20.309 December 3, 2010. - PowerPoint PPT Presentation
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MRI detection of transcriptional regulation of gene expression in
transgenic mice
Batya Cohen, Keren Ziv, Vicki Plaks, Tomer Israely, Vyacheslav Kalchenko, Alon Harmelin,
Laura Benjamin, and Michal Neeman
Derek Ju and John Kucharczyk
20.309
December 3, 2010
Goal: noninvasive detection of reporter gene expression in live
animals•Current: Fluorescent, bioluminescent, and
enzyme reporters for optical imaging and PET▫High sensitivity▫Low spatial and anatomical resolution
•Future: MRI detection of reporter genes▫High 3D spatial and anatomical resolution▫Multiple contrast mechanisms
Ferritin: the ideal reporter gene?
•Iron storage protein that is essential to life
•Ferritin substantially elevates R2 in tissue▫R2 = transverse relaxation
rate of water, 1/T2
•Linear dependence of R2 on the magnetic field
•High R2 at low iron loading
Experimental regulation of Ferritin
Tissue specific VE cadherin promoter
Tetracycline transactivator (tTA)
tTAtetracyclin
Experiment 1:tTA controlled by VE-cadherin
•VE-cadherin is used in small percentage of cells demonstrate sensitivity
sTG:
dTG:
Experiment 2: expression in utero•VE-Cadherin expressed after day 13.5•High expression in the liver, heart and
posterior, low in brain
dTG
sTG
Experiment 3:over expression in liver•No difference between dTG and sTG•EGFP demonstrated over expression of
ferritin in dTG•Liver maintains homeostasis of iron
concentrationsTG dTG
Experiment 4: dynamic changes in ferritin
•Induction of transgene with tetracycline withdrawal
•Unexpected: R2 drops with expression of ferritin
•Could be vacuoles
sTG
dTG
Conclusions
•Could detect expression in VE cells•Could detect organ specific expression in
embryos•Ferritin regulation prevents detection in
liver•Dynamic changes in liver ferritin results
in decreased R2