Limits...
Human mesenchymal stem cells labelled with dye-loaded amorphous silica nanoparticles: long-term biosafety, stemness preservation and traceability in the beating heart.

Gallina C, Capelôa T, Saviozzi S, Accomasso L, Catalano F, Tullio F, Martra G, Penna C, Pagliaro P, Turinetto V, Giachino C - J Nanobiotechnology (2015)

Bottom Line: However, there are still controversies on their effective biosafety on cellular systems.Optimal cell labelling is obtained after 16 h exposure of hMSCs to fluorescent 50 nm SiO2-NPs (50 µg mL(-1)); interestingly, lysosomal activation consequent to NPs storage is not associated to oxidative stress.The demonstrated biosafety, combined with proper cell labelling and visualization in histological sections, make these SiO2-NPs optimal candidates for the purpose of stem cell tracking inside heart tissue.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical and Biological Sciences, University of Turin, 10, Regione Gonzole, CAP 10043, Orbassano, TO, Italy. clara.gallina@gmail.com.

ABSTRACT

Background: Treatment of myocardial infarction with mesenchymal stem cells (MSCs) has proven beneficial effects in both animal and clinical studies. Engineered silica nanoparticles (SiO2-NPs) have been extensively used as contrast agents in regenerative medicine, due to their resistance to degradation and ease of functionalization. However, there are still controversies on their effective biosafety on cellular systems. In this perspective, the aims of the present study are: 1) to deeply investigate the impact of amorphous 50 nm SiO2-NPs on viability and function of human bone marrow-derived MSCs (hMSCs); 2) to optimize a protocol of harmless hMSCs labelling and test its feasibility in a beating heart model.

Results: Optimal cell labelling is obtained after 16 h exposure of hMSCs to fluorescent 50 nm SiO2-NPs (50 µg mL(-1)); interestingly, lysosomal activation consequent to NPs storage is not associated to oxidative stress. During prolonged culture hMSCs do not undergo cyto- or genotoxicity, preserve their proliferative potential and their stemness/differentiation properties. Finally, the bright fluorescence emitted by internalized SiO2-NPs allows both clear visualization of hMSCs in normal and infarcted rat hearts and ultrastructural analysis of cell engraftment inside myocardial tissue.

Conclusions: Overall, 50 nm SiO2-NPs display elevated compatibility with hMSCs in terms of lack of cyto- and genotoxicity and maintenance of important features of these cells. The demonstrated biosafety, combined with proper cell labelling and visualization in histological sections, make these SiO2-NPs optimal candidates for the purpose of stem cell tracking inside heart tissue.

No MeSH data available.


Related in: MedlinePlus

Imaging of SiO2-NPs -labelled hMSCs inside infarcted hearts. a Reconstruction of a 10 µm transverse slice to show the distribution of labelled hMSCs in infarcted hearts. White points: clusters of hMSCs. Red line: perimeter of cell distribution inside infarcted ventricles. b Subset illustrating labelled hMSCs (red) clustered inside ventricular tissue. Magnification 5×, scale bar 200 µm. c Representative confocal reconstruction with superposition of bright field, hMSCs (red) and nuclei (blue). The white dotted line is the perimeter of the volume representation in (d) to underline the higher co-localization between labelled hMSCs (red) and nuclei (blue) due to major cell aggregation typical of the injured area. Magnification 40×, scale bar 50 µm
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4625930&req=5

Fig6: Imaging of SiO2-NPs -labelled hMSCs inside infarcted hearts. a Reconstruction of a 10 µm transverse slice to show the distribution of labelled hMSCs in infarcted hearts. White points: clusters of hMSCs. Red line: perimeter of cell distribution inside infarcted ventricles. b Subset illustrating labelled hMSCs (red) clustered inside ventricular tissue. Magnification 5×, scale bar 200 µm. c Representative confocal reconstruction with superposition of bright field, hMSCs (red) and nuclei (blue). The white dotted line is the perimeter of the volume representation in (d) to underline the higher co-localization between labelled hMSCs (red) and nuclei (blue) due to major cell aggregation typical of the injured area. Magnification 40×, scale bar 50 µm

Mentions: After the profound characterization of the impact of 50 nm SiO2-NPs on viability, proliferation and stemness properties of in vitro cultured hMSCs, a further important aim of the present study was to ascertain the traceability of injected SCs in the challenging condition of a beating heart, subjected or not to ischemia/reperfusion (I/R). To this aim an ex vivo model of perfused rat heart was used. Cells employed for these investigations were harvested at t0, as this time point allowed obtaining almost the totality of the cells stained with 50 nm SiO2-NPs with no interference of culture-dependent dilution of internalized NPs, seen above (Fig. 3). In non-ischemic hearts, reconstruction of a representative transverse section highlighted that labelled cells, injected into the myocardium apex, were widespread distributed inside the walls of both right and left ventricles (Fig. 5a). Focus on a specific low magnification field underlined that hMSCs were clearly labelled by internalized fluorescent SiO2-NPs (Fig. 5b, arrowheads). In addition, confocal analysis at higher magnification (Fig. 5c) and volume reconstruction of a particular area of the same field (Fig. 5d, arrowheads) confirmed that cells were roughly dispersed inside the tissue, as often co-localization of one nucleus per labelled cell was observed. Concerning infarcted hearts, cells were injected into the apex 30 min after the ligation of the left descending coronary artery (LDCA) was removed. In these conditions, the pattern of cell distribution within the left ventricle myocardium depicted a major concentration of hMSCs close to the I/R lesion (Fig. 6a, white points) and the bright red fluorescence emitted by internalized SiO2-NPs underlined larger cell clusters (Fig. 6b, arrowheads). Higher magnification analysis (Fig. 6c) and volume reconstruction of a specific field (Fig. 6d, arrowheads) then revealed a higher number of nuclei co-localizing with red fluorescent cells respect to normally perfused hearts.Fig. 5


Human mesenchymal stem cells labelled with dye-loaded amorphous silica nanoparticles: long-term biosafety, stemness preservation and traceability in the beating heart.

Gallina C, Capelôa T, Saviozzi S, Accomasso L, Catalano F, Tullio F, Martra G, Penna C, Pagliaro P, Turinetto V, Giachino C - J Nanobiotechnology (2015)

Imaging of SiO2-NPs -labelled hMSCs inside infarcted hearts. a Reconstruction of a 10 µm transverse slice to show the distribution of labelled hMSCs in infarcted hearts. White points: clusters of hMSCs. Red line: perimeter of cell distribution inside infarcted ventricles. b Subset illustrating labelled hMSCs (red) clustered inside ventricular tissue. Magnification 5×, scale bar 200 µm. c Representative confocal reconstruction with superposition of bright field, hMSCs (red) and nuclei (blue). The white dotted line is the perimeter of the volume representation in (d) to underline the higher co-localization between labelled hMSCs (red) and nuclei (blue) due to major cell aggregation typical of the injured area. Magnification 40×, scale bar 50 µm
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4625930&req=5

Fig6: Imaging of SiO2-NPs -labelled hMSCs inside infarcted hearts. a Reconstruction of a 10 µm transverse slice to show the distribution of labelled hMSCs in infarcted hearts. White points: clusters of hMSCs. Red line: perimeter of cell distribution inside infarcted ventricles. b Subset illustrating labelled hMSCs (red) clustered inside ventricular tissue. Magnification 5×, scale bar 200 µm. c Representative confocal reconstruction with superposition of bright field, hMSCs (red) and nuclei (blue). The white dotted line is the perimeter of the volume representation in (d) to underline the higher co-localization between labelled hMSCs (red) and nuclei (blue) due to major cell aggregation typical of the injured area. Magnification 40×, scale bar 50 µm
Mentions: After the profound characterization of the impact of 50 nm SiO2-NPs on viability, proliferation and stemness properties of in vitro cultured hMSCs, a further important aim of the present study was to ascertain the traceability of injected SCs in the challenging condition of a beating heart, subjected or not to ischemia/reperfusion (I/R). To this aim an ex vivo model of perfused rat heart was used. Cells employed for these investigations were harvested at t0, as this time point allowed obtaining almost the totality of the cells stained with 50 nm SiO2-NPs with no interference of culture-dependent dilution of internalized NPs, seen above (Fig. 3). In non-ischemic hearts, reconstruction of a representative transverse section highlighted that labelled cells, injected into the myocardium apex, were widespread distributed inside the walls of both right and left ventricles (Fig. 5a). Focus on a specific low magnification field underlined that hMSCs were clearly labelled by internalized fluorescent SiO2-NPs (Fig. 5b, arrowheads). In addition, confocal analysis at higher magnification (Fig. 5c) and volume reconstruction of a particular area of the same field (Fig. 5d, arrowheads) confirmed that cells were roughly dispersed inside the tissue, as often co-localization of one nucleus per labelled cell was observed. Concerning infarcted hearts, cells were injected into the apex 30 min after the ligation of the left descending coronary artery (LDCA) was removed. In these conditions, the pattern of cell distribution within the left ventricle myocardium depicted a major concentration of hMSCs close to the I/R lesion (Fig. 6a, white points) and the bright red fluorescence emitted by internalized SiO2-NPs underlined larger cell clusters (Fig. 6b, arrowheads). Higher magnification analysis (Fig. 6c) and volume reconstruction of a specific field (Fig. 6d, arrowheads) then revealed a higher number of nuclei co-localizing with red fluorescent cells respect to normally perfused hearts.Fig. 5

Bottom Line: However, there are still controversies on their effective biosafety on cellular systems.Optimal cell labelling is obtained after 16 h exposure of hMSCs to fluorescent 50 nm SiO2-NPs (50 µg mL(-1)); interestingly, lysosomal activation consequent to NPs storage is not associated to oxidative stress.The demonstrated biosafety, combined with proper cell labelling and visualization in histological sections, make these SiO2-NPs optimal candidates for the purpose of stem cell tracking inside heart tissue.

View Article: PubMed Central - PubMed

Affiliation: Department of Clinical and Biological Sciences, University of Turin, 10, Regione Gonzole, CAP 10043, Orbassano, TO, Italy. clara.gallina@gmail.com.

ABSTRACT

Background: Treatment of myocardial infarction with mesenchymal stem cells (MSCs) has proven beneficial effects in both animal and clinical studies. Engineered silica nanoparticles (SiO2-NPs) have been extensively used as contrast agents in regenerative medicine, due to their resistance to degradation and ease of functionalization. However, there are still controversies on their effective biosafety on cellular systems. In this perspective, the aims of the present study are: 1) to deeply investigate the impact of amorphous 50 nm SiO2-NPs on viability and function of human bone marrow-derived MSCs (hMSCs); 2) to optimize a protocol of harmless hMSCs labelling and test its feasibility in a beating heart model.

Results: Optimal cell labelling is obtained after 16 h exposure of hMSCs to fluorescent 50 nm SiO2-NPs (50 µg mL(-1)); interestingly, lysosomal activation consequent to NPs storage is not associated to oxidative stress. During prolonged culture hMSCs do not undergo cyto- or genotoxicity, preserve their proliferative potential and their stemness/differentiation properties. Finally, the bright fluorescence emitted by internalized SiO2-NPs allows both clear visualization of hMSCs in normal and infarcted rat hearts and ultrastructural analysis of cell engraftment inside myocardial tissue.

Conclusions: Overall, 50 nm SiO2-NPs display elevated compatibility with hMSCs in terms of lack of cyto- and genotoxicity and maintenance of important features of these cells. The demonstrated biosafety, combined with proper cell labelling and visualization in histological sections, make these SiO2-NPs optimal candidates for the purpose of stem cell tracking inside heart tissue.

No MeSH data available.


Related in: MedlinePlus