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MRI detection of transcriptional regulation of gene expression in transgenic mice

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.

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MRI detection of transcriptional regulation of gene expression in transgenic mice

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  1. 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

  2. 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

  3. 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

  4. Experimental regulation of Ferritin tTA tetracyclin Tissue specific VE cadherin promoter Tetracycline transactivator (tTA)

  5. Creation of dTG and sTG mice • dTG=double transgenic • sTG=single transgenic

  6. Experiment 1:tTA controlled by VE-cadherin • VE-cadherin is used in small percentage of cells  demonstrate sensitivity sTG: dTG:

  7. Experiment 2: expression in utero • VE-Cadherin expressed after day 13.5 • High expression in the liver, heart and posterior, low in brain sTG dTG

  8. 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 concentration sTG dTG

  9. Experiment 4: dynamic changes in ferritin sTG dTG • Induction of transgene with tetracycline withdrawal • Unexpected: R2 drops with expression of ferritin • Could be vacuoles

  10. 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

  11. Uses in BE • Tool for testing iron homeostasis • MRI can be used to detect gene expression with high spatial and anatomical resolution

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