Imaging the spatial distribution of transgene expression in the lungs with positron emission tomography

Richard, Jean-Christophe; Lc, Welch; Dp, Schuster

doi:10.1038/sj.gt.3302117

Cited by 24 publications

(15 citation statements)

References 32 publications

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“…Figure 2 demonstrates this to be the case by showing a strong correlation between radioactivity data obtained from the final image at the end of the imaging session with lung tissue radioactivity measured ex vivo in lungs placed in a gamma well counter (Fig. 2). [Note that the slope of 0.35 is consistent with previous results (38) and with the known density of lung tissue (ϳ0.3 g/ml) in vivo (29). The PET data are expressed in units per Images at right are merged microCT and microPET images after coregistration, using fiducial markers on both the CT and PET images.…”

Section: F]fdg By Marrow Inflammatory Cells)supporting

confidence: 72%

Molecular imaging of lung glucose uptake after endotoxin in mice

Zhou

Kozlowski

Goodrich

et al. 2005

American Journal of Physiology-Lung Cellular and Molecular Phys

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Positron emission tomographic imaging after administration of the glucose analog fluorine-18 fluorodeoxyglucose ([18F]FDG) may be useful to study neutrophilic inflammation of the lungs. In this study, we sought to determine the specificity of the increase in lung [18F]FDG uptake after intraperitoneal endotoxin (Etx) for neutrophil influx into mouse lungs and to determine the regulation of glucose uptake after Etx by Toll-like receptors (TLRs) and TNF-α. Lung tissue radioactivity measurements by imaging were validated against counts in a gamma well counter. Glucose uptake was quantified as the [18F]FDG tissue-to-blood radioactivity ratio (TBR) after validating this measure against the “gold standard” measure of glucose uptake, the “net influx rate constant.” TBR measurements were made in a control group (no intervention), a group administered Etx, and a group administered Etx plus an additional agent (e.g., vinblastine) or Etx administered to a mutant mouse strain. The glucose uptake measurements were compared with measurements of myeloperoxidase. Increases in TBR after Etx were significantly but not completely eliminated by neutrophil depletion with vinblastine. Increases in TBR after Etx were consistent with signaling via either TLR-4 or TLR-2 (the latter probably secondary to peptidoglycan contaminants in Etx preparation) and were decreased by drug inhibition of TLR-4 but not by inhibition of TNF-α. Thus molecular imaging can be used to noninvasively monitor biological effects of Etx on lungs in mice, and changes in lung glucose uptake can be used to monitor effects of anti-inflammatory agents. Such imaging capacity provides a powerful new paradigm for translational “mouse-to-human” pulmonary research.

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Section: F]fdg By Marrow Inflammatory Cells)supporting

confidence: 72%

Molecular imaging of lung glucose uptake after endotoxin in mice

Zhou

Kozlowski

Goodrich

et al. 2005

American Journal of Physiology-Lung Cellular and Molecular Phys

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“…Our results indicate the HSV1-TK-targeted inactivating mutations in the NLS suppress nuclear import of its protein and further increase the total cellular enzymatic activity. These results also correspond to the published results of other research groups (25,26). In mammalian cells, most intracellular proteins are degraded in the cytoplasm (27,28).…”

Section: Discussionsupporting

confidence: 81%

Generation of Destabilized Herpes Simplex Virus Type 1 Thymidine Kinase as Transcription Reporter for PET Reporter Systems in Molecular–Genetic Imaging

Hsieh¹,

Chen²,

Wang³

et al. 2007

J Nucl Med

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Herpes simplex virus type 1 thymidine kinase (HSV1-TK) is a widely used reporter for in vivo noninvasive monitoring of therapeutic gene expression, immune cell trafficking, and proteinprotein interactions in various animal systems. However, the stability of HSV1-TK limits its application in studies that require rapid turnover of the reporter. The purpose of this study was to create a destabilized HSV1-TK as a transcription reporter that allows for dynamic studies of short-time-scale gene expression events. Methods: A destabilized HSV1-TK was created by targeting inactivating mutations in the nuclear localization signal of HSV1-TK and fusing the degradation domain of mouse ornithine decarboxylase to the C-terminal end. The protein or enzyme stability was determined by Western blot analysis and HSV1-TK enzyme activity assay, respectively. The proteasome inhibition assay was used to test whether the rapid turnover of the destabilized HSV1-TK was processed in a 26S proteasomedependent manner. The suitability of destabilized HSV1-TK as a transcription reporter was tested by linking it to a tetracyclineturnoff-expressing system. The dynamic transcriptional events mediating a series of doxycycline inductions were monitored by destabilized HSV1-TK or by native HSV1-TK and were determined by an in vitro HSV1-TK enzyme activity assay and in vivo small-animal PET imaging. Results: The destabilized HSV1-TK, unlike wild-type HSV1-TK, was unstable in the presence of cycloheximide and had a short half-life of protein and enzyme activity. The rapid turnover of the destabilized HSV1-TK was processed in a 26S proteasome-dependent manner. Furthermore, the destabilized HSV1-TK had low cytotoxicity when it was highly expressed in living cells. The results of dynamic gene expression studies in vitro and in vivo showed that the destabilized HSV1-TK is an optimal reporter for monitoring short-time-scale dynamic transcriptional events mediating a series of doxycycline inductions, whereas the wild-type HSV1-TK is not optimal to achieve this purpose. Conclusion: The use of destabilized HSV1-TK as a transcription reporter together with a molecular probe, which has a short physical and biologic half-life, allows more direct monitoring of transcription induction and easier monitoring of its coincidence with other biochemical changes.

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“…Furthermore, this molecular imaging strategy has been used effectively to study the intrapulmonary distribution of vector delivery, and to compare different vector vehicles (19). The current combinations of reporter imaging genes and probes should be considered to be 'first generation' systems; improvements can be anticipated as the relative value and limitations of each system are defined and newer combinations are developed.…”

Section: Discussionmentioning

confidence: 99%

“…Using viral delivery of a mutant Herpes simplex virus type-1 thymidine kinase gene (mHSV1-tk 1 ) to subsequently trap and accumulate a radioactively labeled PET reporter probe, our group has recently used PET imaging to characterize the onset and duration of transgene expression, the intrapulmonary spatial distribution of a transgene and the relative efficacy of different methods of vector delivery in vivo in normal rodent lungs (18)(19)(20)(21). To date, however, no study has shown that PET imaging can be used to visualize transgene expression during disease, and more specifically, during acute lung injury or solid organ transplantation.…”

Section: Introductionmentioning

confidence: 99%

In Vivo Molecular Imaging Characterizes Pulmonary Gene Expression During Experimental Lung Transplantation

Dharmarajan¹,

Hayama²,

Kozlowski

et al. 2005

American Journal of Transplantation

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Experimental gene therapy is a promising strategy to prevent ischemia-reperfusion (I/R) injury and allograft rejection after lung transplantation, and methods will eventually be needed to characterize pulmonary transgene expression in vivo in humans. Therefore, we studied positron emission tomography (PET) as a means of performing in vivo molecular imaging in rodent models of lung transplantation. Rats were transfected endotracheally with adenovirus encoding a fusion gene of a mutant Herpes simplex virus-1 thymidine kinase and the green fluorescent protein gene (the former serving as an imaging reporter gene). Twenty-four hours after transfection, lungs were transplanted in groups representing normal transplantation, I/R injury and acute allograft rejection. Imaging was obtained either 24 h after transplantation to study reperfusion injury or 4 days after transplantation to study graft rejection. After imaging, lungs were excised and analyzed for thymidine kinase activity. Imaging detected transgene expression in transplanted lungs even in the presence of acute rejection or I/R injury. The PET imaging signal correlated with in vitro lung tissue assays of thymidine kinase activity (r 2 = 0.534). Thus, noninvasive molecular imaging with PET is a feasible, sensitive and quantitative method for characterizing pulmonary transgene expression in experimental lung transplantation.

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Imaging the spatial distribution of transgene expression in the lungs with positron emission tomography

Cited by 24 publications

References 32 publications

Molecular imaging of lung glucose uptake after endotoxin in mice

Molecular imaging of lung glucose uptake after endotoxin in mice

Generation of Destabilized Herpes Simplex Virus Type 1 Thymidine Kinase as Transcription Reporter for PET Reporter Systems in Molecular–Genetic Imaging

In Vivo Molecular Imaging Characterizes Pulmonary Gene Expression During Experimental Lung Transplantation

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