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Lysosomal targeting with stable and sensitive fluorescent probes (Superior LysoProbes): applications for lysosome labeling and tracking during apoptosis.

Chen X, Bi Y, Wang T, Li P, Yan X, Hou S, Bammert CE, Ju J, Gibson KM, Pavan WJ, Bi L - Sci Rep (2015)

Bottom Line: The use of Superior LysoProbes facilitates the direct visualization of the lysosomal response to lobaplatin elicited in human chloangiocarcinoma (CCA) RBE cells, using confocal laser scanning microscopy.Additionally, we have characterized the role of lysosomes in autophagy, the correlation between lysosome function and microtubule strength, and the alteration of lysosomal morphology during apoptosis.Our findings indicate that Superior LysoProbes offer numerous advantages over previous reagents to examine the intracellular activities of lysosomes.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Michigan Technological University, Houghton, MI 49931.

ABSTRACT
Intracellular pH plays an important role in the response to cancer invasion. We have designed and synthesized a series of new fluorescent probes (Superior LysoProbes) with the capacity to label acidic organelles and monitor lysosomal pH. Unlike commercially available fluorescent dyes, Superior LysoProbes are lysosome-specific and are highly stable. The use of Superior LysoProbes facilitates the direct visualization of the lysosomal response to lobaplatin elicited in human chloangiocarcinoma (CCA) RBE cells, using confocal laser scanning microscopy. Additionally, we have characterized the role of lysosomes in autophagy, the correlation between lysosome function and microtubule strength, and the alteration of lysosomal morphology during apoptosis. Our findings indicate that Superior LysoProbes offer numerous advantages over previous reagents to examine the intracellular activities of lysosomes.

No MeSH data available.


Related in: MedlinePlus

Intracellular distribution of a control compound, rhodamine-lactose (1 μM) compared to MitoTracker (80 nM) and LysoTracker Green (2 μM).HeLa cells were imaged on an inverted laser scanning fluorescent microscope (Olympus) using a 60 × oil immersion objective lens.
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f5: Intracellular distribution of a control compound, rhodamine-lactose (1 μM) compared to MitoTracker (80 nM) and LysoTracker Green (2 μM).HeLa cells were imaged on an inverted laser scanning fluorescent microscope (Olympus) using a 60 × oil immersion objective lens.

Mentions: We hypothesized that the N-linked glycan moiety of Superior LysoProbes would enhance lysosomal targeting. The fluorescence of Superior LysoProbes I–IV was distributed in a punctate pattern only in lysosomes rather than diffusely throughout the cell. Conversely, the fluorescent images of the precursor of Superior LysoProbes (Scheme S1) was non-punctate and diffuse after several hours, suggestive of rapid diffusion leading to reduced probe sensitivity. Thus, the N-linked glycan moieties appear to enhance selective lysosomal aggregation. To further confirm our hypothesis, we synthesized an additional control compound, rhodamine-lactose (lacking N-linked glycan; chemical structure shown in Fig. 4). This derivative did not target to lysosomes. As shown in Fig. 5, rhodamine-lactose and LysoTracker did not co-localize. However, rhodamine-lactose did co-localize with MitoTracker. These results confirmed the requirement for an N-linked glycan moiety for efficient lysosomal-targeting and intracellular retention.


Lysosomal targeting with stable and sensitive fluorescent probes (Superior LysoProbes): applications for lysosome labeling and tracking during apoptosis.

Chen X, Bi Y, Wang T, Li P, Yan X, Hou S, Bammert CE, Ju J, Gibson KM, Pavan WJ, Bi L - Sci Rep (2015)

Intracellular distribution of a control compound, rhodamine-lactose (1 μM) compared to MitoTracker (80 nM) and LysoTracker Green (2 μM).HeLa cells were imaged on an inverted laser scanning fluorescent microscope (Olympus) using a 60 × oil immersion objective lens.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4355733&req=5

f5: Intracellular distribution of a control compound, rhodamine-lactose (1 μM) compared to MitoTracker (80 nM) and LysoTracker Green (2 μM).HeLa cells were imaged on an inverted laser scanning fluorescent microscope (Olympus) using a 60 × oil immersion objective lens.
Mentions: We hypothesized that the N-linked glycan moiety of Superior LysoProbes would enhance lysosomal targeting. The fluorescence of Superior LysoProbes I–IV was distributed in a punctate pattern only in lysosomes rather than diffusely throughout the cell. Conversely, the fluorescent images of the precursor of Superior LysoProbes (Scheme S1) was non-punctate and diffuse after several hours, suggestive of rapid diffusion leading to reduced probe sensitivity. Thus, the N-linked glycan moieties appear to enhance selective lysosomal aggregation. To further confirm our hypothesis, we synthesized an additional control compound, rhodamine-lactose (lacking N-linked glycan; chemical structure shown in Fig. 4). This derivative did not target to lysosomes. As shown in Fig. 5, rhodamine-lactose and LysoTracker did not co-localize. However, rhodamine-lactose did co-localize with MitoTracker. These results confirmed the requirement for an N-linked glycan moiety for efficient lysosomal-targeting and intracellular retention.

Bottom Line: The use of Superior LysoProbes facilitates the direct visualization of the lysosomal response to lobaplatin elicited in human chloangiocarcinoma (CCA) RBE cells, using confocal laser scanning microscopy.Additionally, we have characterized the role of lysosomes in autophagy, the correlation between lysosome function and microtubule strength, and the alteration of lysosomal morphology during apoptosis.Our findings indicate that Superior LysoProbes offer numerous advantages over previous reagents to examine the intracellular activities of lysosomes.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Michigan Technological University, Houghton, MI 49931.

ABSTRACT
Intracellular pH plays an important role in the response to cancer invasion. We have designed and synthesized a series of new fluorescent probes (Superior LysoProbes) with the capacity to label acidic organelles and monitor lysosomal pH. Unlike commercially available fluorescent dyes, Superior LysoProbes are lysosome-specific and are highly stable. The use of Superior LysoProbes facilitates the direct visualization of the lysosomal response to lobaplatin elicited in human chloangiocarcinoma (CCA) RBE cells, using confocal laser scanning microscopy. Additionally, we have characterized the role of lysosomes in autophagy, the correlation between lysosome function and microtubule strength, and the alteration of lysosomal morphology during apoptosis. Our findings indicate that Superior LysoProbes offer numerous advantages over previous reagents to examine the intracellular activities of lysosomes.

No MeSH data available.


Related in: MedlinePlus