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Freeze-thaw treatment effects on the dynamic mechanical properties of articular cartilage.

Szarko M, Muldrew K, Bertram JE - BMC Musculoskelet Disord (2010)

Bottom Line: Both subzero storage temperature as well as freezing rate were compared using control samples (4°C) and samples stored at either -20°C or -80°C as well as samples first snap frozen in liquid nitrogen (-196°C) prior to storage at -80°C.Mechanical changes shown are likely due to ice lens creation, where frost heave effects may have caused collagen damage.That storage to -20°C and -80°C did not alter the mechanical properties of articular cartilage shows that when combined with a rapid thawing protocol to 37.5°C, the tissue may successfully be stored at subzero temperatures.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Biomedical Sciences, St. George's, University of London, London, UK.

ABSTRACT

Background: As a relatively non-regenerative tissue, articular cartilage has been targeted for cryopreservation as a method of mitigating a lack of donor tissue availability for transplant surgeries. In addition, subzero storage of articular cartilage has long been used in biomedical studies using various storage temperatures. The current investigation studies the potential for freeze-thaw to affect the mechanical properties of articular cartilage through direct comparison of various subzero storage temperatures.

Methods: Both subzero storage temperature as well as freezing rate were compared using control samples (4°C) and samples stored at either -20°C or -80°C as well as samples first snap frozen in liquid nitrogen (-196°C) prior to storage at -80°C. All samples were thawed at 37.5°C to testing temperature (22°C). Complex stiffness and hysteresis characterized load resistance and damping properties using a non-destructive, low force magnitude, dynamic indentation protocol spanning a broad loading rate range to identify the dynamic viscoelastic properties of cartilage.

Results: Stiffness levels remained unchanged with exposure to the various subzero temperatures. Hysteresis increased in samples snap frozen at -196°C and stored at -80°C, though remained unchanged with exposure to the other storage temperatures.

Conclusions: Mechanical changes shown are likely due to ice lens creation, where frost heave effects may have caused collagen damage. That storage to -20°C and -80°C did not alter the mechanical properties of articular cartilage shows that when combined with a rapid thawing protocol to 37.5°C, the tissue may successfully be stored at subzero temperatures.

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Related in: MedlinePlus

Diagram illustrating the experimental equipment. The electrodynamic vibrator (Ling Dynamics, GB) was used to drive the indenter rod. The displacement transducer mounted between the vibrator and rigid indenting rod measured actual displacements of the indenter. The piezoelectric force transducer measured force on the non-articulating side of the sample. The sample was tested while submerged in isotonic PBS. The Stanford SR780 Signal analyzer was used to generate sinusoidal waveforms and analyze voltage signals from the displacement transducer and force beam. The signal amplifier ensured that an adequate signal was produced even though low magnitude forces were used. The oscilloscope was used for identifying pre-load levels and to confirm contact between the indenter and sample surface.
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Figure 1: Diagram illustrating the experimental equipment. The electrodynamic vibrator (Ling Dynamics, GB) was used to drive the indenter rod. The displacement transducer mounted between the vibrator and rigid indenting rod measured actual displacements of the indenter. The piezoelectric force transducer measured force on the non-articulating side of the sample. The sample was tested while submerged in isotonic PBS. The Stanford SR780 Signal analyzer was used to generate sinusoidal waveforms and analyze voltage signals from the displacement transducer and force beam. The signal amplifier ensured that an adequate signal was produced even though low magnitude forces were used. The oscilloscope was used for identifying pre-load levels and to confirm contact between the indenter and sample surface.

Mentions: Mechanical properties were assessed by applying a dynamically varying, cyclic compression to the articular surface and comparing this with the output of a load cell supporting the sample. The differences between the phase of the applied waveform and that reaching the support was attributed to the intervening specimen. Movement of the indenter was produced by an electrodynamic vibrator (Ling Dynamics, UK). A displacement transducer mounted between the vibrator and the rigid indenter beam monitored displacement of the indenter and indicated the excitation signal (equal amplitude sinusoids). A custom built piezoelectric force beam, with a resolution of 1 × 10-4 N and a frequency response of 2500 Hz, measured force at the base of the specimen, providing the analysis signal. The electrodynamic vibrator was controlled by the function generator component of a spectrum analyzer (Stanford model SR780). The excitation and analysis signals were then compared using Fast Fourier Transform (FFT) analysis within the spectrum analyzer to determine specimen properties. This arrangement is shown in Figure 1 and has been previously used as reliable, sensitive way to measure dynamic mechanical properties of viscoelastic materials [18]. Maximum indenter displacements reached 0.01 mm creating strains less than 5% and producing loading forces of 28.45 N. Output signals from each transducer were amplified prior to input to the spectrum analyzer to increase resolution.


Freeze-thaw treatment effects on the dynamic mechanical properties of articular cartilage.

Szarko M, Muldrew K, Bertram JE - BMC Musculoskelet Disord (2010)

Diagram illustrating the experimental equipment. The electrodynamic vibrator (Ling Dynamics, GB) was used to drive the indenter rod. The displacement transducer mounted between the vibrator and rigid indenting rod measured actual displacements of the indenter. The piezoelectric force transducer measured force on the non-articulating side of the sample. The sample was tested while submerged in isotonic PBS. The Stanford SR780 Signal analyzer was used to generate sinusoidal waveforms and analyze voltage signals from the displacement transducer and force beam. The signal amplifier ensured that an adequate signal was produced even though low magnitude forces were used. The oscilloscope was used for identifying pre-load levels and to confirm contact between the indenter and sample surface.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2958988&req=5

Figure 1: Diagram illustrating the experimental equipment. The electrodynamic vibrator (Ling Dynamics, GB) was used to drive the indenter rod. The displacement transducer mounted between the vibrator and rigid indenting rod measured actual displacements of the indenter. The piezoelectric force transducer measured force on the non-articulating side of the sample. The sample was tested while submerged in isotonic PBS. The Stanford SR780 Signal analyzer was used to generate sinusoidal waveforms and analyze voltage signals from the displacement transducer and force beam. The signal amplifier ensured that an adequate signal was produced even though low magnitude forces were used. The oscilloscope was used for identifying pre-load levels and to confirm contact between the indenter and sample surface.
Mentions: Mechanical properties were assessed by applying a dynamically varying, cyclic compression to the articular surface and comparing this with the output of a load cell supporting the sample. The differences between the phase of the applied waveform and that reaching the support was attributed to the intervening specimen. Movement of the indenter was produced by an electrodynamic vibrator (Ling Dynamics, UK). A displacement transducer mounted between the vibrator and the rigid indenter beam monitored displacement of the indenter and indicated the excitation signal (equal amplitude sinusoids). A custom built piezoelectric force beam, with a resolution of 1 × 10-4 N and a frequency response of 2500 Hz, measured force at the base of the specimen, providing the analysis signal. The electrodynamic vibrator was controlled by the function generator component of a spectrum analyzer (Stanford model SR780). The excitation and analysis signals were then compared using Fast Fourier Transform (FFT) analysis within the spectrum analyzer to determine specimen properties. This arrangement is shown in Figure 1 and has been previously used as reliable, sensitive way to measure dynamic mechanical properties of viscoelastic materials [18]. Maximum indenter displacements reached 0.01 mm creating strains less than 5% and producing loading forces of 28.45 N. Output signals from each transducer were amplified prior to input to the spectrum analyzer to increase resolution.

Bottom Line: Both subzero storage temperature as well as freezing rate were compared using control samples (4°C) and samples stored at either -20°C or -80°C as well as samples first snap frozen in liquid nitrogen (-196°C) prior to storage at -80°C.Mechanical changes shown are likely due to ice lens creation, where frost heave effects may have caused collagen damage.That storage to -20°C and -80°C did not alter the mechanical properties of articular cartilage shows that when combined with a rapid thawing protocol to 37.5°C, the tissue may successfully be stored at subzero temperatures.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Biomedical Sciences, St. George's, University of London, London, UK.

ABSTRACT

Background: As a relatively non-regenerative tissue, articular cartilage has been targeted for cryopreservation as a method of mitigating a lack of donor tissue availability for transplant surgeries. In addition, subzero storage of articular cartilage has long been used in biomedical studies using various storage temperatures. The current investigation studies the potential for freeze-thaw to affect the mechanical properties of articular cartilage through direct comparison of various subzero storage temperatures.

Methods: Both subzero storage temperature as well as freezing rate were compared using control samples (4°C) and samples stored at either -20°C or -80°C as well as samples first snap frozen in liquid nitrogen (-196°C) prior to storage at -80°C. All samples were thawed at 37.5°C to testing temperature (22°C). Complex stiffness and hysteresis characterized load resistance and damping properties using a non-destructive, low force magnitude, dynamic indentation protocol spanning a broad loading rate range to identify the dynamic viscoelastic properties of cartilage.

Results: Stiffness levels remained unchanged with exposure to the various subzero temperatures. Hysteresis increased in samples snap frozen at -196°C and stored at -80°C, though remained unchanged with exposure to the other storage temperatures.

Conclusions: Mechanical changes shown are likely due to ice lens creation, where frost heave effects may have caused collagen damage. That storage to -20°C and -80°C did not alter the mechanical properties of articular cartilage shows that when combined with a rapid thawing protocol to 37.5°C, the tissue may successfully be stored at subzero temperatures.

Show MeSH
Related in: MedlinePlus