Limits...
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.


Fitting results of the proposed diffusion-reaction model to the data.With 3 fitting parameters, the model (solid lines) closely followed the averaged uptake data for trehalose (squares) and total (n)-O-Ac-Tre (circles), which were obtained by incubation with (A) 5mM, (B) 15 mM, and (C) 30 mM 6-O-Ac-Tre. Inset Figures: Predicted increase in the concentration of 6- (black solid line), 5- (dark-gray solid line), 4- (gray solid line), 3- (gray dashed line), 2- (gray dash-dotted line) and 1-O-Ac-Tre (gray dotted line). These results were obtained by averaging the data from at least 3 animals and the error bars represent the animal-to-animal variability.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4482662&req=5

pone.0130323.g004: Fitting results of the proposed diffusion-reaction model to the data.With 3 fitting parameters, the model (solid lines) closely followed the averaged uptake data for trehalose (squares) and total (n)-O-Ac-Tre (circles), which were obtained by incubation with (A) 5mM, (B) 15 mM, and (C) 30 mM 6-O-Ac-Tre. Inset Figures: Predicted increase in the concentration of 6- (black solid line), 5- (dark-gray solid line), 4- (gray solid line), 3- (gray dashed line), 2- (gray dash-dotted line) and 1-O-Ac-Tre (gray dotted line). These results were obtained by averaging the data from at least 3 animals and the error bars represent the animal-to-animal variability.

Mentions: The data on accumulation of trehalose and acetylated trehalose in hepatocytes were obtained using cells isolated from at least 3 different animals, and demonstrated a similar trend. However, parameters such as size/age/health and variability in the isolation procedure on different days could potentially affect the function of the hepatocytes from different animals. For the purpose of calculating average values for diffusion and conversion of Act-Tre, the average values are plotted in one figure (Fig 4) where the large error bars represent the data range for visual evaluation of the goodness of the fit. The individual data sets are reported in S1 Fig.


Engineered Trehalose Permeable to Mammalian Cells.

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

Fitting results of the proposed diffusion-reaction model to the data.With 3 fitting parameters, the model (solid lines) closely followed the averaged uptake data for trehalose (squares) and total (n)-O-Ac-Tre (circles), which were obtained by incubation with (A) 5mM, (B) 15 mM, and (C) 30 mM 6-O-Ac-Tre. Inset Figures: Predicted increase in the concentration of 6- (black solid line), 5- (dark-gray solid line), 4- (gray solid line), 3- (gray dashed line), 2- (gray dash-dotted line) and 1-O-Ac-Tre (gray dotted line). These results were obtained by averaging the data from at least 3 animals and the error bars represent the animal-to-animal variability.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0130323.g004: Fitting results of the proposed diffusion-reaction model to the data.With 3 fitting parameters, the model (solid lines) closely followed the averaged uptake data for trehalose (squares) and total (n)-O-Ac-Tre (circles), which were obtained by incubation with (A) 5mM, (B) 15 mM, and (C) 30 mM 6-O-Ac-Tre. Inset Figures: Predicted increase in the concentration of 6- (black solid line), 5- (dark-gray solid line), 4- (gray solid line), 3- (gray dashed line), 2- (gray dash-dotted line) and 1-O-Ac-Tre (gray dotted line). These results were obtained by averaging the data from at least 3 animals and the error bars represent the animal-to-animal variability.
Mentions: The data on accumulation of trehalose and acetylated trehalose in hepatocytes were obtained using cells isolated from at least 3 different animals, and demonstrated a similar trend. However, parameters such as size/age/health and variability in the isolation procedure on different days could potentially affect the function of the hepatocytes from different animals. For the purpose of calculating average values for diffusion and conversion of Act-Tre, the average values are plotted in one figure (Fig 4) where the large error bars represent the data range for visual evaluation of the goodness of the fit. The individual data sets are reported in S1 Fig.

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.