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Hierarchical Approaches to Investigating Tissue Micromechanics Hazel Screen School of Engineering & Materials Science, Queen Mary, University of London.

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Presentation on theme: "Hierarchical Approaches to Investigating Tissue Micromechanics Hazel Screen School of Engineering & Materials Science, Queen Mary, University of London."— Presentation transcript:

1 Hierarchical Approaches to Investigating Tissue Micromechanics Hazel Screen School of Engineering & Materials Science, Queen Mary, University of London 6 th November 2008

2 Connective Tissue Function & Health Connective tissues = structural support cartilage once destroyed, is not repaired Hunter. W, 1743 Normal healing mechanisms are unavailable to damaged connective tissues

3 Investigating Tissue Micromechanics 1. Understanding tissue structure and how to help protect it from damage 2. Understand how to facilitate repair in damaged tissue

4 How to Facilitate Tissue Repair Chemical cues: Growth factors Nutrients Mechanical cues: Fluid flow Pressure Deformation

5 Regulates normal tissue homeostasis Implicated in pathological processes Implicated in repair processes Harness it for tissue engineering?? Mechanical Loading (in vitro) (in vivo) Altered Cell Response Proliferation Matrix synthesis Matrix degradation Cell/matrix orientation Mechanotransduction

6 The Hierarchy of Mechanobiology Body mechanics Joint mechanics Tissue mechanics Cell mechanics Protein mechanics

7 The Hierarchy of Mechanobiology Body mechanics Joint mechanics Tissue mechanics Cell mechanics Protein mechanics

8 How does the tissue hierarchy control mechanical properties? How does the material deform: How are strains transferred to the cells? Investigate the local mechanical environment as the mechanotransduction stimulus of interest Tissue Composition & Mechanics

9 Tissue Composition & Tissue Mechanics Articular cartilage Tendon / ligament Skin Aortic valve

10 In Situ Analysis Techniques Screen et al. (2003) Biorheol. 40, Stepper Motor Heater Pads Microscope Objective Lens Grips Specimen Medium Coverslip Screen et al. (2004) J. Eng. Med. 218, Custom designed rig for location on confocal microscope Enables tensile / compressive loading of viable tissue samples Use range of matrix & cell stains to visualise matrix components during loading

11 Tendon Structure Multi-level fibre composite Considered simple collagen tissue to study

12 Tendon Extension Mechanisms u Fibre Extension Fibre Sliding v L u Fibre Extension v L Fibre Sliding Screen et al. (2004) J. Strain 40:4,

13 Tendon Extension Mechanisms Collagen molecule Fibril Fibre Fascicle

14 Tendon Extension Mechanisms Shearing/ Sliding Extension rotation Collagen molecule Fibril Fibre Fascicle

15 What controls the fibre composite behaviour? Non-Collagenous Matrix Shape Molecule Scott (2003) J. Physiol. 553; Scott & Thomlinson (1998) J. Anat. 192; Decorin: Binds around collagen fibrils Screen et al (2005) Ann Biomed Eng 33;

16 Understanding Viscoelasticity Very rapid relaxation ; Total relaxation < 60 secs Highly viscous tissue Direct testsIncremental tests 8% 2% 4% 6% 8% Gross mechanical properties:

17 Confocal Images – Stress Relaxation

18

19 Fibre Relaxation Fibre Siding collagen fibre tenocyte nuclei Applied Extension = L Confocal Images – Stress Relaxation

20 Percentage fibre relaxation (%) Percentage between-fibre relaxation (%) Fibre RelaxationFibre Sliding TYPICAL DATA: 4 % Applied Strain Confocal Images – Stress Relaxation

21 Between-fibre displacement ( m) Fibre Relaxation 1% 2% 4% 6% 8% Fibre Sliding Fibre relaxation ( m) 1% 2% 4% 6% 8% Confocal Images – Stress Relaxation

22 How does this affect the cells? We now have some understanding of the mechanisms of extension & relaxation: What does this mean for the local strain environment throughout the sample and surrounding the cells?

23 Finite Element Approach Track coordinates of every cell Construct a Delaunay mesh of triangle elements Monitor deformation & strain in each element during relaxation Important coordinates into Matlab S Evans - Cardiff University

24 x y Finite Element Approach X displacementY displacement

25 Displacements Y displacementX displacement x y

26 Shear strain Relaxation Strains x y X strain Y strain Huge variability in response Strain seems random

27 Relaxation Strains x y x strains y strains shear strains Predominantly negative = compression Range positive & negative = Fibre sliding Wide range of shear strains

28 Relaxation Behaviour Relaxation strains far exceed the initial applied strain Values are both positive and negative Monitoring deformation of each triangle Significant sliding between cells on different fibres Sliding creates large shear strain in matrix (on cells) Loading Direction: Transverse Direction: More uniform response & predominantly negative strains Water movement out of inter-fibre spacing

29 Cell Perspective Cell processes link adjacent rows of cells: Large deflections (y strains) Compressive loading of cells (x strains)

30 Other Hierarchical Changes Confocal focus X-ray synchrotron scattering Himadri Gupta (Max Plank)

31 Synchrotron X-ray Scattering ESRF BL ID2 Peter Boesecke (Grenoble) CCD X – ray detector X - ray Load cell Small angle X – ray scattering (SAXS) setup 2 /D Microtensile tester Max load 250 g – 12 kg Strain measured with video extensometry (NON-contact)

32 Fibril Strain During Relaxation Two time constants +, - General Form Stress (MPa) Fibril strain (%) Time (Seconds) Fitting Data: + σ & + ε 10 s - σ & - ε 50 s Fitting ε constants to σ ? Fibril relaxation & stress relaxation governed by same relaxation constants

33 Two Component Viscoelastic Model E1E1 1 E2E2 2 Fixed strain 0 Voigt element Maxwell element

34 Transverse Fibril Mechanics? Same two-stage relaxation Fits same time constants Increase greater than volume conservation alone

35 Relaxation Mechanics? Fibrils Fibres Shorter Slide AXIAL TRANSVERSE Increases

36 Relaxation Behaviour Significant structural reordering during relaxation Significant movement of water Some water moves out of sample? Water moves into fibrils? Transfer from fibre to fibril space? Each level of fibre composite independent Fibril response very ordered Fibre response opposes this

37 Acknowledgements Shima Toorani Vinton Cheng Mike Kayser Jong Seto Steffi Krauss Prof Steve Greenwald Prof Julia Shelton Prof Dan Bader Prof David Lee EPSRC Tissue Science Laboratories Dr Sam Evans Dr Himadri Gupta


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