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Novel (89)Zr cell labeling approach for PET-based cell trafficking studies.

Bansal A, Pandey MK, Demirhan YE, Nesbitt JJ, Crespo-Diaz RJ, Terzic A, Behfar A, DeGrado TR - EJNMMI Res (2015)

Bottom Line: The effect of labeling on cellular viability was tested by proliferation, trypan blue, and cytotoxicity/apoptosis assays.Radioactivity concentrations of labeled cells of up to 0.5 MBq/10(6) cells were achieved without a negative effect on cellular viability.We have developed a robust, general, and biostable (89)Zr-DBN-based cell labeling strategy with promise for wide applications of PET-based non-invasive in vivo cell trafficking.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiology, Mayo Clinic, Rochester, 55905 MN USA.

ABSTRACT

Background: With the recent growth of interest in cell-based therapies and radiolabeled cell products, there is a need to develop more robust cell labeling and imaging methods for in vivo tracking of living cells. This study describes evaluation of a novel cell labeling approach with the positron emission tomography (PET) isotope (89)Zr (T 1/2 = 78.4 h). (89)Zr may allow PET imaging measurements for several weeks and take advantage of the high sensitivity of PET imaging.

Methods: A novel cell labeling agent, (89)Zr-desferrioxamine-NCS ((89)Zr-DBN), was synthesized. Mouse-derived melanoma cells (mMCs), dendritic cells (mDCs), and human mesenchymal stem cells (hMSCs) were covalently labeled with (89)Zr-DBN via the reaction between the NCS group on (89)Zr-DBN and primary amine groups present on cell surface membrane protein. The stability of the label on the cell was tested by cell efflux studies for 7 days. The effect of labeling on cellular viability was tested by proliferation, trypan blue, and cytotoxicity/apoptosis assays. The stability of label was also studied in in vivo mouse models by serial PET scans and ex vivo biodistribution following intravenous and intramyocardial injection of (89)Zr-labeled hMSCs. For comparison, imaging experiments were performed after intravenous injections of (89)Zr hydrogen phosphate ((89)Zr(HPO4)2).

Results: The labeling agent, (89)Zr-DBN, was prepared in 55% ± 5% decay-corrected radiochemical yield measured by silica gel iTLC. The cell labeling efficiency was 30% to 50% after 30 min labeling depending on cell type. Radioactivity concentrations of labeled cells of up to 0.5 MBq/10(6) cells were achieved without a negative effect on cellular viability. Cell efflux studies showed high stability of the radiolabel out to 7 days. Myocardially delivered (89)Zr-labeled hMSCs showed retention in the myocardium, as well as redistribution to the lung, liver, and bone. Intravenously administered (89)Zr-labeled hMSCs also distributed primarily to the lung, liver, and bone, whereas intravenous (89)Zr(HPO4)2 distributed to the liver and bone with no activity in the lung. Thus, the in vivo stability of the radiolabel on the hMSCs was evidenced.

Conclusions: We have developed a robust, general, and biostable (89)Zr-DBN-based cell labeling strategy with promise for wide applications of PET-based non-invasive in vivo cell trafficking.

No MeSH data available.


Related in: MedlinePlus

Assessment of (A) viability, (B) cytotoxicity, and (C) apoptosis in89Zr-labeled and unlabeled cells. No statistically significant differences were observed between 89Zr-labeled and unlabeled cells after 7 days of culture with regard to viability, cytotoxicity, or apoptosis. As positive controls, 30 μg/mL digitonin was used for assays (A) and (B), and 2 μΜ staurosporine for (C). *p < 0.05 versus assessments in 89Zr-labeled and unlabeled cells using unpaired t-test. Values are shown as mean ± standard deviation, n = 3.
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Fig3: Assessment of (A) viability, (B) cytotoxicity, and (C) apoptosis in89Zr-labeled and unlabeled cells. No statistically significant differences were observed between 89Zr-labeled and unlabeled cells after 7 days of culture with regard to viability, cytotoxicity, or apoptosis. As positive controls, 30 μg/mL digitonin was used for assays (A) and (B), and 2 μΜ staurosporine for (C). *p < 0.05 versus assessments in 89Zr-labeled and unlabeled cells using unpaired t-test. Values are shown as mean ± standard deviation, n = 3.

Mentions: The ApoTox-Glo assay showed no loss in cellular viability and no increase in cytotoxicity or apoptosis in radiolabeled cells as illustrated in Figure 3. Viability was lost, and cytotoxicity enhanced when 30 μg/mL digitonin was added to cells (positive control), and apoptosis was increased with the addition of 2 μM staurosporine (positive control).Figure 3


Novel (89)Zr cell labeling approach for PET-based cell trafficking studies.

Bansal A, Pandey MK, Demirhan YE, Nesbitt JJ, Crespo-Diaz RJ, Terzic A, Behfar A, DeGrado TR - EJNMMI Res (2015)

Assessment of (A) viability, (B) cytotoxicity, and (C) apoptosis in89Zr-labeled and unlabeled cells. No statistically significant differences were observed between 89Zr-labeled and unlabeled cells after 7 days of culture with regard to viability, cytotoxicity, or apoptosis. As positive controls, 30 μg/mL digitonin was used for assays (A) and (B), and 2 μΜ staurosporine for (C). *p < 0.05 versus assessments in 89Zr-labeled and unlabeled cells using unpaired t-test. Values are shown as mean ± standard deviation, n = 3.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4401478&req=5

Fig3: Assessment of (A) viability, (B) cytotoxicity, and (C) apoptosis in89Zr-labeled and unlabeled cells. No statistically significant differences were observed between 89Zr-labeled and unlabeled cells after 7 days of culture with regard to viability, cytotoxicity, or apoptosis. As positive controls, 30 μg/mL digitonin was used for assays (A) and (B), and 2 μΜ staurosporine for (C). *p < 0.05 versus assessments in 89Zr-labeled and unlabeled cells using unpaired t-test. Values are shown as mean ± standard deviation, n = 3.
Mentions: The ApoTox-Glo assay showed no loss in cellular viability and no increase in cytotoxicity or apoptosis in radiolabeled cells as illustrated in Figure 3. Viability was lost, and cytotoxicity enhanced when 30 μg/mL digitonin was added to cells (positive control), and apoptosis was increased with the addition of 2 μM staurosporine (positive control).Figure 3

Bottom Line: The effect of labeling on cellular viability was tested by proliferation, trypan blue, and cytotoxicity/apoptosis assays.Radioactivity concentrations of labeled cells of up to 0.5 MBq/10(6) cells were achieved without a negative effect on cellular viability.We have developed a robust, general, and biostable (89)Zr-DBN-based cell labeling strategy with promise for wide applications of PET-based non-invasive in vivo cell trafficking.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiology, Mayo Clinic, Rochester, 55905 MN USA.

ABSTRACT

Background: With the recent growth of interest in cell-based therapies and radiolabeled cell products, there is a need to develop more robust cell labeling and imaging methods for in vivo tracking of living cells. This study describes evaluation of a novel cell labeling approach with the positron emission tomography (PET) isotope (89)Zr (T 1/2 = 78.4 h). (89)Zr may allow PET imaging measurements for several weeks and take advantage of the high sensitivity of PET imaging.

Methods: A novel cell labeling agent, (89)Zr-desferrioxamine-NCS ((89)Zr-DBN), was synthesized. Mouse-derived melanoma cells (mMCs), dendritic cells (mDCs), and human mesenchymal stem cells (hMSCs) were covalently labeled with (89)Zr-DBN via the reaction between the NCS group on (89)Zr-DBN and primary amine groups present on cell surface membrane protein. The stability of the label on the cell was tested by cell efflux studies for 7 days. The effect of labeling on cellular viability was tested by proliferation, trypan blue, and cytotoxicity/apoptosis assays. The stability of label was also studied in in vivo mouse models by serial PET scans and ex vivo biodistribution following intravenous and intramyocardial injection of (89)Zr-labeled hMSCs. For comparison, imaging experiments were performed after intravenous injections of (89)Zr hydrogen phosphate ((89)Zr(HPO4)2).

Results: The labeling agent, (89)Zr-DBN, was prepared in 55% ± 5% decay-corrected radiochemical yield measured by silica gel iTLC. The cell labeling efficiency was 30% to 50% after 30 min labeling depending on cell type. Radioactivity concentrations of labeled cells of up to 0.5 MBq/10(6) cells were achieved without a negative effect on cellular viability. Cell efflux studies showed high stability of the radiolabel out to 7 days. Myocardially delivered (89)Zr-labeled hMSCs showed retention in the myocardium, as well as redistribution to the lung, liver, and bone. Intravenously administered (89)Zr-labeled hMSCs also distributed primarily to the lung, liver, and bone, whereas intravenous (89)Zr(HPO4)2 distributed to the liver and bone with no activity in the lung. Thus, the in vivo stability of the radiolabel on the hMSCs was evidenced.

Conclusions: We have developed a robust, general, and biostable (89)Zr-DBN-based cell labeling strategy with promise for wide applications of PET-based non-invasive in vivo cell trafficking.

No MeSH data available.


Related in: MedlinePlus