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Engineered Trehalose Permeable to Mammalian Cells.

Abazari A, Meimetis LG, Budin G, Bale SS, Weissleder R, Toner M - PLoS ONE (2015)

Bottom Line: Since mammalian cells do not synthesize nor import trehalose, the development of novel methods for efficient intracellular delivery of trehalose has been an ongoing investigation.Trehalose conjugated with 6 acetyl groups (trehalose hexaacetate or 6-O-Ac-Tre) demonstrated superior permeability in rat hepatocytes compared with regular trehalose, trehalose diacetate (2-O-Ac-Tre) and trehalose tetraacetate (4-O-Ac-Tre).Engineering of trehalose chemical structure rather than manipulating the cell, is an innocuous, cell-friendly method for trehalose delivery, with demonstrated potential for trehalose loading in different types of cells and cell lines, and can facilitate the wide-spread application of trehalose as an intracellular protective agent in biopreservation studies.

View Article: PubMed Central - PubMed

Affiliation: The Center for Engineering in Medicine, Massachusetts General Hospital, Harvard Medical School, and Shriners Hospital for Children, Boston, Massachusetts, United States of America.

ABSTRACT
Trehalose is a naturally occurring disaccharide which is associated with extraordinary stress-tolerance capacity in certain species of unicellular and multicellular organisms. In mammalian cells, presence of intra- and extracellular trehalose has been shown to confer improved tolerance against freezing and desiccation. Since mammalian cells do not synthesize nor import trehalose, the development of novel methods for efficient intracellular delivery of trehalose has been an ongoing investigation. Herein, we studied the membrane permeability of engineered lipophilic derivatives of trehalose. Trehalose conjugated with 6 acetyl groups (trehalose hexaacetate or 6-O-Ac-Tre) demonstrated superior permeability in rat hepatocytes compared with regular trehalose, trehalose diacetate (2-O-Ac-Tre) and trehalose tetraacetate (4-O-Ac-Tre). Once in the cell, intracellular esterases hydrolyzed the 6-O-Ac-Tre molecules, releasing free trehalose into the cytoplasm. The total concentration of intracellular trehalose (plus acetylated variants) reached as high as 10 fold the extracellular concentration of 6-O-Ac-Tre, attaining concentrations suitable for applications in biopreservation. To describe this accumulation phenomenon, a diffusion-reaction model was proposed and the permeability and reaction kinetics of 6-O-Ac-Tre were determined by fitting to experimental data. Further studies suggested that the impact of the loading and the presence of intracellular trehalose on cellular viability and function were negligible. Engineering of trehalose chemical structure rather than manipulating the cell, is an innocuous, cell-friendly method for trehalose delivery, with demonstrated potential for trehalose loading in different types of cells and cell lines, and can facilitate the wide-spread application of trehalose as an intracellular protective agent in biopreservation studies.

No MeSH data available.


The synthesis steps for trehalose diacetate (2-O-Ac-Tre), trehalose tetraacetate (4-O-Ac-Tre) and trehalose hexaacetate (6-O-Ac-Tre).
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pone.0130323.g001: The synthesis steps for trehalose diacetate (2-O-Ac-Tre), trehalose tetraacetate (4-O-Ac-Tre) and trehalose hexaacetate (6-O-Ac-Tre).

Mentions: Three trehalose derivatives were synthesized with 2, 4 and 6 degrees of acetylation as shown in Fig 1. To a solution of Trehalose in pyridine was added trityl chloride to give intermediate 1, which was subsequently acetylated with acetic anhydride to provide compound 2. Trityl deprotection of compound 2 was accomplished with Iron (III) chloride hexahydrate in methylene chloride to yield trehalose hexaacetate (compound 8). In a similar fashion trehalose tetraacetate (compound 9) was constructed by formation of the bis acetal (compound 3) from trehalose followed by acetylation and deprotection with acetic acid to afford trehalose tetraacetate (compound 9). Common synthetic intermediate 1 was used to access trehalose diaacetate Firstly, intermediate 1 was benzyl protected with benzyl bromide and sodium hydride in dimethyl formamide to give the fully protected dissacharide intermediate 4. Compound 4 was trityl deprotected with trifluoroacetic acid and subsequently acetylated with acetic anhydride to give intermediate 6. Finally, compound 6 was benzyl deprotected with palladium hydroxide in the presence of hydrogen to provide trehalose diaacetate (compound 7).


Engineered Trehalose Permeable to Mammalian Cells.

Abazari A, Meimetis LG, Budin G, Bale SS, Weissleder R, Toner M - PLoS ONE (2015)

The synthesis steps for trehalose diacetate (2-O-Ac-Tre), trehalose tetraacetate (4-O-Ac-Tre) and trehalose hexaacetate (6-O-Ac-Tre).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4482662&req=5

pone.0130323.g001: The synthesis steps for trehalose diacetate (2-O-Ac-Tre), trehalose tetraacetate (4-O-Ac-Tre) and trehalose hexaacetate (6-O-Ac-Tre).
Mentions: Three trehalose derivatives were synthesized with 2, 4 and 6 degrees of acetylation as shown in Fig 1. To a solution of Trehalose in pyridine was added trityl chloride to give intermediate 1, which was subsequently acetylated with acetic anhydride to provide compound 2. Trityl deprotection of compound 2 was accomplished with Iron (III) chloride hexahydrate in methylene chloride to yield trehalose hexaacetate (compound 8). In a similar fashion trehalose tetraacetate (compound 9) was constructed by formation of the bis acetal (compound 3) from trehalose followed by acetylation and deprotection with acetic acid to afford trehalose tetraacetate (compound 9). Common synthetic intermediate 1 was used to access trehalose diaacetate Firstly, intermediate 1 was benzyl protected with benzyl bromide and sodium hydride in dimethyl formamide to give the fully protected dissacharide intermediate 4. Compound 4 was trityl deprotected with trifluoroacetic acid and subsequently acetylated with acetic anhydride to give intermediate 6. Finally, compound 6 was benzyl deprotected with palladium hydroxide in the presence of hydrogen to provide trehalose diaacetate (compound 7).

Bottom Line: Since mammalian cells do not synthesize nor import trehalose, the development of novel methods for efficient intracellular delivery of trehalose has been an ongoing investigation.Trehalose conjugated with 6 acetyl groups (trehalose hexaacetate or 6-O-Ac-Tre) demonstrated superior permeability in rat hepatocytes compared with regular trehalose, trehalose diacetate (2-O-Ac-Tre) and trehalose tetraacetate (4-O-Ac-Tre).Engineering of trehalose chemical structure rather than manipulating the cell, is an innocuous, cell-friendly method for trehalose delivery, with demonstrated potential for trehalose loading in different types of cells and cell lines, and can facilitate the wide-spread application of trehalose as an intracellular protective agent in biopreservation studies.

View Article: PubMed Central - PubMed

Affiliation: The Center for Engineering in Medicine, Massachusetts General Hospital, Harvard Medical School, and Shriners Hospital for Children, Boston, Massachusetts, United States of America.

ABSTRACT
Trehalose is a naturally occurring disaccharide which is associated with extraordinary stress-tolerance capacity in certain species of unicellular and multicellular organisms. In mammalian cells, presence of intra- and extracellular trehalose has been shown to confer improved tolerance against freezing and desiccation. Since mammalian cells do not synthesize nor import trehalose, the development of novel methods for efficient intracellular delivery of trehalose has been an ongoing investigation. Herein, we studied the membrane permeability of engineered lipophilic derivatives of trehalose. Trehalose conjugated with 6 acetyl groups (trehalose hexaacetate or 6-O-Ac-Tre) demonstrated superior permeability in rat hepatocytes compared with regular trehalose, trehalose diacetate (2-O-Ac-Tre) and trehalose tetraacetate (4-O-Ac-Tre). Once in the cell, intracellular esterases hydrolyzed the 6-O-Ac-Tre molecules, releasing free trehalose into the cytoplasm. The total concentration of intracellular trehalose (plus acetylated variants) reached as high as 10 fold the extracellular concentration of 6-O-Ac-Tre, attaining concentrations suitable for applications in biopreservation. To describe this accumulation phenomenon, a diffusion-reaction model was proposed and the permeability and reaction kinetics of 6-O-Ac-Tre were determined by fitting to experimental data. Further studies suggested that the impact of the loading and the presence of intracellular trehalose on cellular viability and function were negligible. Engineering of trehalose chemical structure rather than manipulating the cell, is an innocuous, cell-friendly method for trehalose delivery, with demonstrated potential for trehalose loading in different types of cells and cell lines, and can facilitate the wide-spread application of trehalose as an intracellular protective agent in biopreservation studies.

No MeSH data available.