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Simple SPION incubation as an efficient intracellular labeling method for tracking neural progenitor cells using MRI.

Chen CC, Ku MC, D M J, Lai JS, Hueng DY, Chang C - PLoS ONE (2013)

Bottom Line: However, compelling evidence also shows that simple SPION incubation is not invariably ineffective.The labeling efficiency can be improved by prolonged incubation and elevated iron doses.The results showed that, following 48 hours of incubation at 75 µg/ml, nearly all NPCs exhibited visible SPION intake.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biomedical Sciences, Academic Sinica, Taipei, Taiwan.

ABSTRACT
Cellular magnetic resonance imaging (MRI) has been well-established for tracking neural progenitor cells (NPC). Superparamagnetic iron oxide nanoparticles (SPIONs) approved for clinical application are the most common agents used for labeling. Conventionally, transfection agents (TAs) were added with SPIONs to facilitate cell labeling because SPIONs in the native unmodified form were deemed inefficient for intracellular labeling. However, compelling evidence also shows that simple SPION incubation is not invariably ineffective. The labeling efficiency can be improved by prolonged incubation and elevated iron doses. The goal of the present study was to establish simple SPION incubation as an efficient intracellular labeling method. To this end, NPCs derived from the neonatal subventricular zone were incubated with SPIONs (Feridex®) and then evaluated in vitro with regard to the labeling efficiency and biological functions. The results showed that, following 48 hours of incubation at 75 µg/ml, nearly all NPCs exhibited visible SPION intake. Evidence from light microscopy, electron microscopy, chemical analysis, and magnetic resonance imaging confirmed the effectiveness of the labeling. Additionally, biological assays showed that the labeled NPCs exhibited unaffected viability, oxidative stress, apoptosis and differentiation. In the demonstrated in vivo cellular MRI experiment, the hypointensities representing the SPION labeled NPCs remained observable throughout the entire tracking period. The findings indicate that simple SPION incubation without the addition of TAs is an efficient intracellular magnetic labeling method. This simple approach may be considered as an alternative approach to the mainstream labeling method that involves the use of TAs.

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Examination of the iron content after simple SPION incubation with light microscopy, chemical analysis, and in vitro MRI.(A) No PB staining found in unlabeled cells while all the labeled ones were PB positive. (B) Little iron was detected in unlabeled cells while the labeled cells showed an averaged iron content of 5.3±1.1 pg/cell. (C) The R2* value was minimal in unlabeled cells while the labeled cells showed an averaged R2* value of 68.7±4.9 ms−1. (D) The R2 value was 6.1±0.4 ms−1 in unlabeled cells while the labeled cells showed an averaged R2 value of 12.8±0.8 ms−1. (E) The T2* map. (F) The T2 map.
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pone-0056125-g004: Examination of the iron content after simple SPION incubation with light microscopy, chemical analysis, and in vitro MRI.(A) No PB staining found in unlabeled cells while all the labeled ones were PB positive. (B) Little iron was detected in unlabeled cells while the labeled cells showed an averaged iron content of 5.3±1.1 pg/cell. (C) The R2* value was minimal in unlabeled cells while the labeled cells showed an averaged R2* value of 68.7±4.9 ms−1. (D) The R2 value was 6.1±0.4 ms−1 in unlabeled cells while the labeled cells showed an averaged R2 value of 12.8±0.8 ms−1. (E) The T2* map. (F) The T2 map.

Mentions: The simple SPION incubation method resulted in 100% PB positive NPCs while no PB positive cells were seen in unlabeled NPCs (Fig. 4A). Chemical analysis of the iron content by ICP-AES indicated the averaged iron content was 5.3±1.1 pg/cell in the labeled NPCs and 0.1±0.06 pg/cell in the unlabeled ones (Fig. 4B). In vitro MRI relaxometry indicated that the R2* values were 6.2±0.2 ms−1 and 68.7±4.9 ms−1 for unlabeled and labeled NPCs, respectively (Fig. 4C). The R2 values were 6.1±0.4 ms−1 and 12.8±0.8 ms−1 for unlabeled and labeled NPCs, respectively (Fig. 4D). The T2* and T2 maps of the NPCs solidified in agarose are shown in Fig. 4E, and Fig. 4F. The labeled NPCs appeared as hypointensities on both of the images.


Simple SPION incubation as an efficient intracellular labeling method for tracking neural progenitor cells using MRI.

Chen CC, Ku MC, D M J, Lai JS, Hueng DY, Chang C - PLoS ONE (2013)

Examination of the iron content after simple SPION incubation with light microscopy, chemical analysis, and in vitro MRI.(A) No PB staining found in unlabeled cells while all the labeled ones were PB positive. (B) Little iron was detected in unlabeled cells while the labeled cells showed an averaged iron content of 5.3±1.1 pg/cell. (C) The R2* value was minimal in unlabeled cells while the labeled cells showed an averaged R2* value of 68.7±4.9 ms−1. (D) The R2 value was 6.1±0.4 ms−1 in unlabeled cells while the labeled cells showed an averaged R2 value of 12.8±0.8 ms−1. (E) The T2* map. (F) The T2 map.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3585319&req=5

pone-0056125-g004: Examination of the iron content after simple SPION incubation with light microscopy, chemical analysis, and in vitro MRI.(A) No PB staining found in unlabeled cells while all the labeled ones were PB positive. (B) Little iron was detected in unlabeled cells while the labeled cells showed an averaged iron content of 5.3±1.1 pg/cell. (C) The R2* value was minimal in unlabeled cells while the labeled cells showed an averaged R2* value of 68.7±4.9 ms−1. (D) The R2 value was 6.1±0.4 ms−1 in unlabeled cells while the labeled cells showed an averaged R2 value of 12.8±0.8 ms−1. (E) The T2* map. (F) The T2 map.
Mentions: The simple SPION incubation method resulted in 100% PB positive NPCs while no PB positive cells were seen in unlabeled NPCs (Fig. 4A). Chemical analysis of the iron content by ICP-AES indicated the averaged iron content was 5.3±1.1 pg/cell in the labeled NPCs and 0.1±0.06 pg/cell in the unlabeled ones (Fig. 4B). In vitro MRI relaxometry indicated that the R2* values were 6.2±0.2 ms−1 and 68.7±4.9 ms−1 for unlabeled and labeled NPCs, respectively (Fig. 4C). The R2 values were 6.1±0.4 ms−1 and 12.8±0.8 ms−1 for unlabeled and labeled NPCs, respectively (Fig. 4D). The T2* and T2 maps of the NPCs solidified in agarose are shown in Fig. 4E, and Fig. 4F. The labeled NPCs appeared as hypointensities on both of the images.

Bottom Line: However, compelling evidence also shows that simple SPION incubation is not invariably ineffective.The labeling efficiency can be improved by prolonged incubation and elevated iron doses.The results showed that, following 48 hours of incubation at 75 µg/ml, nearly all NPCs exhibited visible SPION intake.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biomedical Sciences, Academic Sinica, Taipei, Taiwan.

ABSTRACT
Cellular magnetic resonance imaging (MRI) has been well-established for tracking neural progenitor cells (NPC). Superparamagnetic iron oxide nanoparticles (SPIONs) approved for clinical application are the most common agents used for labeling. Conventionally, transfection agents (TAs) were added with SPIONs to facilitate cell labeling because SPIONs in the native unmodified form were deemed inefficient for intracellular labeling. However, compelling evidence also shows that simple SPION incubation is not invariably ineffective. The labeling efficiency can be improved by prolonged incubation and elevated iron doses. The goal of the present study was to establish simple SPION incubation as an efficient intracellular labeling method. To this end, NPCs derived from the neonatal subventricular zone were incubated with SPIONs (Feridex®) and then evaluated in vitro with regard to the labeling efficiency and biological functions. The results showed that, following 48 hours of incubation at 75 µg/ml, nearly all NPCs exhibited visible SPION intake. Evidence from light microscopy, electron microscopy, chemical analysis, and magnetic resonance imaging confirmed the effectiveness of the labeling. Additionally, biological assays showed that the labeled NPCs exhibited unaffected viability, oxidative stress, apoptosis and differentiation. In the demonstrated in vivo cellular MRI experiment, the hypointensities representing the SPION labeled NPCs remained observable throughout the entire tracking period. The findings indicate that simple SPION incubation without the addition of TAs is an efficient intracellular magnetic labeling method. This simple approach may be considered as an alternative approach to the mainstream labeling method that involves the use of TAs.

Show MeSH