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The ultrastructure of fibronectin fibers pulled from a protein monolayer at the air-liquid interface and the mechanism of the sheet-to-fiber transition.

Mitsi M, Handschin S, Gerber I, Schwartländer R, Klotzsch E, Wepf R, Vogel V - Biomaterials (2014)

Bottom Line: The ultrastructural characterization is then contrasted with previous FRET studies that characterized the molecular strain within these manually pulled fibers.Particularly relevant for stretch-dependent binding studies is the finding that the interior fiber surfaces are accessible to nanoparticles smaller than 10 nm.In summary, our study discovers the underpinning mechanism by which highly hierarchically structured fibers can be generated with unique mechanical and mechano-chemical properties, a concept that might be extended to other bio- or biomimetic polymers.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Applied Mechanobiology, Vladimir-Prelog-Weg 4, ETH Zurich, Switzerland.

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The swelling effect induced by ionic strength on the fiber structure is reversible. Fibers were pulled out of a 0.4 mg/ml fibronectin solution in PBS and remained in PBS (A) or incubated with H2O (C) prior to fixation. Alternatively, fibers were pulled out of the H2O solution and remained in H2O (B) or incubated with PBS (D) prior to fixation. All samples were chemically fixed with glutaraldehyde, dehydrated in ethanol and embedded in Epon. 50 nm sections were observed by TEM. (E) Fibers were pulled out of a 0.4 mg/ml fibronectin solution in PBS onto microfabricated PDMS trenches. Using a MEMS sensor, force–extension curves were determined for the segments of the fibers freely suspended over the wells. Measurements were performed in PBS (black curves), after treating the fibers with H2O (red curves) and after rehydrating them again with PBS (blue curves). Curves for an average of 12 fibers are shown. Scale bar: 200 nm.
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fig5: The swelling effect induced by ionic strength on the fiber structure is reversible. Fibers were pulled out of a 0.4 mg/ml fibronectin solution in PBS and remained in PBS (A) or incubated with H2O (C) prior to fixation. Alternatively, fibers were pulled out of the H2O solution and remained in H2O (B) or incubated with PBS (D) prior to fixation. All samples were chemically fixed with glutaraldehyde, dehydrated in ethanol and embedded in Epon. 50 nm sections were observed by TEM. (E) Fibers were pulled out of a 0.4 mg/ml fibronectin solution in PBS onto microfabricated PDMS trenches. Using a MEMS sensor, force–extension curves were determined for the segments of the fibers freely suspended over the wells. Measurements were performed in PBS (black curves), after treating the fibers with H2O (red curves) and after rehydrating them again with PBS (blue curves). Curves for an average of 12 fibers are shown. Scale bar: 200 nm.

Mentions: The distinct features of the fibers pulled out of fibronectin solutions in H2O, as compared to solutions in PBS, suggest that electrostatic interactions during fiber assembly regulate primarily the interlamellar spacing. But once the structure has been formed, can it still respond to alterations of the ionic strength of the environment? Indeed, fibers that were pulled out of solutions of fibronectin in H2O had a very similar organization to fibers initially pulled out of PBS solutions and then incubated with H2O for as short times as 1 min (Fig. 5A, B). Swelled structures, with few spiral segments that show large free spaces, abundant ring-like structures and discontinuities in the protein lamellae were observed in both types of fibers. Similarly, fibers pulled out of H2O solutions and subsequently incubated with PBS were indistinguishable from fibers pulled directly out of PBS (Fig. 5C, D). Force-extension curves (Fig. 5E) were measured for fibers pulled out of PBS (black lines) and deposited onto microfabricated trenches, by using a MEMS device as described previously [16]. The measurements were repeated on the same set of fibers after they were incubated with H2O (red lines) and once again after they were re-incubated with PBS (blue lines). All three sets of force–extension curves, as presented for an average of 12 fibers, were identical (Fig. 5E), suggesting that the tunable morphological differences induced by hydrating the fibers in buffers of different ionic strength are not associated with any material loss.


The ultrastructure of fibronectin fibers pulled from a protein monolayer at the air-liquid interface and the mechanism of the sheet-to-fiber transition.

Mitsi M, Handschin S, Gerber I, Schwartländer R, Klotzsch E, Wepf R, Vogel V - Biomaterials (2014)

The swelling effect induced by ionic strength on the fiber structure is reversible. Fibers were pulled out of a 0.4 mg/ml fibronectin solution in PBS and remained in PBS (A) or incubated with H2O (C) prior to fixation. Alternatively, fibers were pulled out of the H2O solution and remained in H2O (B) or incubated with PBS (D) prior to fixation. All samples were chemically fixed with glutaraldehyde, dehydrated in ethanol and embedded in Epon. 50 nm sections were observed by TEM. (E) Fibers were pulled out of a 0.4 mg/ml fibronectin solution in PBS onto microfabricated PDMS trenches. Using a MEMS sensor, force–extension curves were determined for the segments of the fibers freely suspended over the wells. Measurements were performed in PBS (black curves), after treating the fibers with H2O (red curves) and after rehydrating them again with PBS (blue curves). Curves for an average of 12 fibers are shown. Scale bar: 200 nm.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

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fig5: The swelling effect induced by ionic strength on the fiber structure is reversible. Fibers were pulled out of a 0.4 mg/ml fibronectin solution in PBS and remained in PBS (A) or incubated with H2O (C) prior to fixation. Alternatively, fibers were pulled out of the H2O solution and remained in H2O (B) or incubated with PBS (D) prior to fixation. All samples were chemically fixed with glutaraldehyde, dehydrated in ethanol and embedded in Epon. 50 nm sections were observed by TEM. (E) Fibers were pulled out of a 0.4 mg/ml fibronectin solution in PBS onto microfabricated PDMS trenches. Using a MEMS sensor, force–extension curves were determined for the segments of the fibers freely suspended over the wells. Measurements were performed in PBS (black curves), after treating the fibers with H2O (red curves) and after rehydrating them again with PBS (blue curves). Curves for an average of 12 fibers are shown. Scale bar: 200 nm.
Mentions: The distinct features of the fibers pulled out of fibronectin solutions in H2O, as compared to solutions in PBS, suggest that electrostatic interactions during fiber assembly regulate primarily the interlamellar spacing. But once the structure has been formed, can it still respond to alterations of the ionic strength of the environment? Indeed, fibers that were pulled out of solutions of fibronectin in H2O had a very similar organization to fibers initially pulled out of PBS solutions and then incubated with H2O for as short times as 1 min (Fig. 5A, B). Swelled structures, with few spiral segments that show large free spaces, abundant ring-like structures and discontinuities in the protein lamellae were observed in both types of fibers. Similarly, fibers pulled out of H2O solutions and subsequently incubated with PBS were indistinguishable from fibers pulled directly out of PBS (Fig. 5C, D). Force-extension curves (Fig. 5E) were measured for fibers pulled out of PBS (black lines) and deposited onto microfabricated trenches, by using a MEMS device as described previously [16]. The measurements were repeated on the same set of fibers after they were incubated with H2O (red lines) and once again after they were re-incubated with PBS (blue lines). All three sets of force–extension curves, as presented for an average of 12 fibers, were identical (Fig. 5E), suggesting that the tunable morphological differences induced by hydrating the fibers in buffers of different ionic strength are not associated with any material loss.

Bottom Line: The ultrastructural characterization is then contrasted with previous FRET studies that characterized the molecular strain within these manually pulled fibers.Particularly relevant for stretch-dependent binding studies is the finding that the interior fiber surfaces are accessible to nanoparticles smaller than 10 nm.In summary, our study discovers the underpinning mechanism by which highly hierarchically structured fibers can be generated with unique mechanical and mechano-chemical properties, a concept that might be extended to other bio- or biomimetic polymers.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Applied Mechanobiology, Vladimir-Prelog-Weg 4, ETH Zurich, Switzerland.

Show MeSH
Related in: MedlinePlus