Designing Electrospun Scaffolds with Architectures ... · Designing Electrospun Scaffolds with...

1
Designing Electrospun Scaffolds with Architectures Suitable for Tendon Repair + 1 Bosworth, LA; 1 Downes, S + 1 School of Materials, The University of Manchester, Manchester, UK [email protected] ABSTRACT INTRODUCTION: There is an increasing incidence of patients suffering from chronic pain and spontaneous rupture of tendons. Whilst current interventions are able to repair tendons, e.g. suturing the two tendon ends together, their functionality is often lost. We propose synthetic, degradable scaffolds with purposefully designed architectures as an alternative, ‘off- the-shelf’ solution. In order to achieve this an understanding of tendon structure is required. Tenocytes are the main cell type in tendon and are responsible for the production of collagen Type I fibres. These fibres are arranged into fascicles of increasing size to create a highly organised, hierarchical tissue structure. The objectives were to recreate this structural composition using electrospinning and then assess the fabricated fibre scaffolds both physically and for biocompatibility. METHODS: Poly(ε-caprolactone) (PCL) (Mn 80,000; Sigma) dissolved in Acetone (Fisher Scientific) (concentration 10%w/v) was electrospun using pre- determined parameters: voltage - 20kV, flow rate – 0.05ml/min, distance to collector – 15cm. Fibres were collected on a fine-edged (width 3mm) rotating mandrel, 50RPM for 2D-random fibres or 500RPM for 2D- aligned fibres. 3D-scaffolds were developed by manually twisting aligned fibres (length 30mm) at either end. The three fibre matrices were visually assessed by Scanning Electron Microscopy (SEM) and tensile properties determined by loading to failure using an Instron 2211 with 5N load cell and 5mm/min crosshead speed. Tenocytes sourced from adult equine superficial digital flexor tendons (seeding density 50,000cm -2 ) were cultured on each scaffold type up to 14 days. Cell morphology and their interaction with the scaffold was assessed by SEM. A purpose-made defect was created in the Achilles tendon of a mouse. A single 3D-scaffold (Ø~150μm; length 3mm) was implanted into the defect. The scaffold was held in place by single knot sutures at the proximal and distal ends. Assessment of the graft in situ was determined using variable pressure-SEM immediately post-implantation and after 21 days. RESULTS SECTION: Tensile properties were most significant for 3D-scaffolds, followed by 2D-aligned fibres and weakest tensile properties were obtained for 2D-random fibres (Table 1). Orientation of PCL fibres affected cell alignment (Fig.1). Random fibres demonstrated no orientation compared to aligned and 3D-scaffold, which resulted in cells lying parallel to the fibre axis. Assessment in vivo demonstrated a good interface between the tendon tissue and 3D-scaffold (Fig.2a). After 21 days, the scaffold had been fully integrated within the tendon (Fig.2b). Table 1 – Tensile properties of electrospun PCL fibre scaffolds Electrospun PCL Structure Modulus (MPa) Tensile Strength (MPa) 2D random fibre orientation 1.54 (±0.26) 0.45 (±0.09) 2D aligned fibre orientation 4.84 (±0.13) 1.30 (±0.14) 3D fibre scaffold 14.11 (±3.76) 4.74 (±1.64) Figure 1 – SEM micrographs of tenocytes cultured on PCL fibres (5 days) with orientation (a) 2D-random, (b) 2D-aligned and (c) 3D- scaffold Figure 2 – VP-SEM micrograph of 3D-scaffold in situ immediately after implantation (a) and after 21 days (b). Arrow indicates likely position of integrated 3D-scaffold. DISCUSSION: As a method, electrospinning enables the control of fibre diameter and fibre orientation, which is ideal for mimicking highly fibrous, organised tissues, such as tendons. 3D-scaffolds yielded greatest tensile properties for this polymer/solvent combination (PCL/Acetone) and individual nanofibres conferred contact guidance to the cells. The 3D-scaffolds were proven to be biocompatible in vivo and were well positioned with good interfacial response in situ. After 21 days implantation, the scaffold was located by one of the suture knots as the scaffold had been fully integrated into the tendon due to new tissue formation. SIGNIFICANCE: This research highlights the importance of scaffold design and fabrication in order to create structures that mimic the tissues they are intended to replace, such as tendons. ACKNOWLEDGEMENTS: The Authors would like to thank the EPSRC and UMIP Premier Fund. A B C 3D Scaffold Tendon Tendon 3D Scaffold A B Poster No. 0647 ORS 2012 Annual Meeting

Transcript of Designing Electrospun Scaffolds with Architectures ... · Designing Electrospun Scaffolds with...

  • Designing Electrospun Scaffolds with Architectures Suitable for Tendon Repair +1Bosworth, LA; 1Downes, S

    +1School of Materials, The University of Manchester, Manchester, UK [email protected]

    ABSTRACT INTRODUCTION: There is an increasing incidence of patients suffering from chronic pain and spontaneous rupture of tendons. Whilst current interventions are able to repair tendons, e.g. suturing the two tendon ends together, their functionality is often lost. We propose synthetic, degradable scaffolds with purposefully designed architectures as an alternative, ‘off-the-shelf’ solution. In order to achieve this an understanding of tendon structure is required. Tenocytes are the main cell type in tendon and are responsible for the production of collagen Type I fibres. These fibres are arranged into fascicles of increasing size to create a highly organised, hierarchical tissue structure. The objectives were to recreate this structural composition using electrospinning and then assess the fabricated fibre scaffolds both physically and for biocompatibility. METHODS: Poly(ε-caprolactone) (PCL) (Mn 80,000; Sigma) dissolved in Acetone (Fisher Scientific) (concentration 10%w/v) was electrospun using pre-determined parameters: voltage - 20kV, flow rate – 0.05ml/min, distance to collector – 15cm. Fibres were collected on a fine-edged (width 3mm) rotating mandrel, 50RPM for 2D-random fibres or 500RPM for 2D-aligned fibres. 3D-scaffolds were developed by manually twisting aligned fibres (length 30mm) at either end. The three fibre matrices were visually assessed by Scanning Electron Microscopy (SEM) and tensile properties determined by loading to failure using an Instron 2211 with 5N load cell and 5mm/min crosshead speed. Tenocytes sourced from adult equine superficial digital flexor tendons (seeding density 50,000cm-2) were cultured on each scaffold type up to 14 days. Cell morphology and their interaction with the scaffold was assessed by SEM. A purpose-made defect was created in the Achilles tendon of a mouse. A single 3D-scaffold (Ø~150µm; length 3mm) was implanted into the defect. The scaffold was held in place by single knot sutures at the proximal and distal ends. Assessment of the graft in situ was determined using variable pressure-SEM immediately post-implantation and after 21 days. RESULTS SECTION: Tensile properties were most significant for 3D-scaffolds, followed by 2D-aligned fibres and weakest tensile properties were obtained for 2D-random fibres (Table 1). Orientation of PCL fibres affected cell alignment (Fig.1). Random fibres demonstrated no orientation compared to aligned and 3D-scaffold, which resulted in cells lying parallel to the fibre axis. Assessment in vivo demonstrated a good interface between the tendon tissue and 3D-scaffold (Fig.2a). After 21 days, the scaffold had been fully integrated within the tendon (Fig.2b). Table 1 – Tensile properties of electrospun PCL fibre scaffolds

    Electrospun PCL Structure

    Modulus (MPa) Tensile Strength (MPa)

    2D random fibre orientation

    1.54 (±0.26)

    0.45 (±0.09)

    2D aligned fibre orientation

    4.84 (±0.13)

    1.30 (±0.14)

    3D fibre scaffold 14.11 (±3.76)

    4.74 (±1.64)

    Figure 1 – SEM micrographs of tenocytes cultured on PCL fibres (5 days) with orientation (a) 2D-random, (b) 2D-aligned and (c) 3D-scaffold

    Figure 2 – VP-SEM micrograph of 3D-scaffold in situ immediately after implantation (a) and after 21 days (b). Arrow indicates likely position of integrated 3D-scaffold. DISCUSSION: As a method, electrospinning enables the control of fibre diameter and fibre orientation, which is ideal for mimicking highly fibrous, organised tissues, such as tendons. 3D-scaffolds yielded greatest tensile properties for this polymer/solvent combination (PCL/Acetone) and individual nanofibres conferred contact guidance to the cells. The 3D-scaffolds were proven to be biocompatible in vivo and were well positioned with good interfacial response in situ. After 21 days implantation, the scaffold was located by one of the suture knots as the scaffold had been fully integrated into the tendon due to new tissue formation. SIGNIFICANCE: This research highlights the importance of scaffold design and fabrication in order to create structures that mimic the tissues they are intended to replace, such as tendons. ACKNOWLEDGEMENTS: The Authors would like to thank the EPSRC and UMIP Premier Fund.

    A B

    C

    3D Scaffold

    Tendon

    Tendon

    3D Scaffold

    A

    B

    Poster No. 0647 • ORS 2012 Annual Meeting

    Main MenuSearchProgramAuthor IndexKeyword IndexCopyrightSession Number 001: Stem Cells and ProgenitorsSession Number 002: Tendon and Ligament: Biology & DevelopmentSession Number 003: Molecular Influences on Cartilage BiomechanicsSession Number 004: Inflammation in Osteoarthritis and Immune-Medicated ArthritisSession Number 005: Cell and Molecular Biomechanics: OsteocytesSession Number 006: Intervertebral Disc: Stem Cells & Cell LineSession Number 007: Tendon and Ligament: RegenerationSession Number 008: Cell and Molecular Biomechanics: Physical EffectsSession Number 009: Experimental Osteoarthritis Models: Lubrication and PainSession Number 010: Fracture Healing - QualitySpotlight Session 011: SpineSpotlight Session 012: ACL ReconstructionSpotlight Session 013: Non-Coding RNA and Posttranscriptional Regulation in OASpotlight Session 014: Tissue Reactions to WearSpotlight Session 015: OsteoporosisSession Number 016: Intervertebral DiscSession Number 017: ACL Injury MechanicsSession Number 018: Cartilage and Meniscus RepairSession Number 019: Cell SignalingSession Number 020: Bone - AgingSession Number 021: Bone and Spine BiomechanicsSession Number 022: Tendon BiomechanicsSession Number 023: Bone MechanicsSession Number 024: Cartilage Degradation Biomarkers and ImagingSession Number 025: Fracture Healing ModulationSpotlight Session 026: Current Trends in Joint ReplacementSpotlight Session 027: Tendon and Ligament: Translational ResearchSpotlight Session 028: microRNASpotlight Session 029: Regenerative Approaches for OsteoarthritisSpotlight Session 030: Muscle/BoneSession Number 031: Hip and Femoroacetabular ImpingementSession Number 032: Knee and Hip Finite Element ModalitySession Number 033: The Post-Traumatic Joint Experimental ModelsSession Number 034: Muscle and NerveSession Number 035: Tissue Engineering I: Cartilage and BoneSession Number 036: Biomaterials Stem Cells and Growth FactorsSession Number 037: MeniscusSession Number 038: Osteolysis and Implant FixationSession Number 039: Tissue Engineering II: TendonSession Number 040: Polymeric and Nano-BiomaterialsSession Number 041: Foot and AnkleSession Number 042: Growth Factors and Bone FixationSession Number 043: Knee, ACL, Patello-Femoral JointSession Number 044: Tumors and DiseasesSession Number 045: Orthopaedic InfectionSession Number 046: Cartilage MechanicsSession Number 047: ShoulderSession Number 048: SpineSession Number 049: New Polyethylene Implant WearSession Number 050: Knee ArthroplastyNIRA 1: Cartilage and SpineNIRA 2: Soft TissueNIRA 3: Cartilage Biology and OsteoarthritisNIRA 4: Fracture RepairNIRA 5: BonePS1 Bone MechanicsPS1 Skeletal Growth and Development - Developmenal BiologyPS1 Cell and Molecular Biomechanics - Physical Effects on CellsPS1 Bone FracturePS1 Fracture FixationPS1 ImagingPS1 Cancer/TumorsPS1 Bone BiologyPS1 Bone - Growth FactorsPS1 Bone - Osteoporosis & Metabolic Bone DiseasePS1 Progenitors & Stem CellsPS1 Biomaterials - BioactivePS1 Tissue Engineering - Soft TissuePS1 Cartilage/Meniscus/Synovium - Genetics/GenomicsPS1 Cartilage/Meniscus/Synovium - Cartilage & Matrix ProteinsPS1 Cartilage/Meniscus/Synovium - Cartilage Matrix DegradationPS1 Cartilage/Meniscus/Synovium - CytokinesPS1 Cartilage/Meniscus/Synovium - Growth Factors & AgingPS1 Cartilage/Meniscus/Synovium - Cartilage RepairPS1 Cartilage/Meniscus/Synovium - Cartilage Repair - Cell Based ApproachesPS1 Cartilage/Meniscus/Synovium - MeniscusPS1 Cell & Molecular Imaging of CartilagePS1 Cartilage/Meniscus/Synovium - OsteoarthritisPS1 Cartilage/Meniscus/Synovium - Cartilage MechanicsPS1 Cartilage/Meniscus/Synovium - MechanobiologyPS1 Disease Process Knee - Joint MechanicsPS1 Disease Process HipPS1 Gait & KinematicsPS1 Foot & AnklePS1 Infection & InflammationPS1 TraumaPS1 Total Hip Replacement - GeneralPS1 Total Knee Replacement - General/WearPS1 Arthroplasty - Osteolysis Wear DebrisPS1 Arthroplasty - Implant FixationPS1 Total Knee Replacement - KinematicsPS1 Arthroplasty - Finite Element Analysis/HipPS1 Polyethylene Wear - UHMWPEPS1 Arthroplasty - Finite Element Analysis - KneePS1 Total Hip Replacement - ImplantPS1 Implant - Uni-Knee ReplacementPS1 Hip/Knee Arthroplasty - Surgical Navigation & RoboticsPS1 Implant Wear - Vitamin E PolyethylenePS1 Implant Wear - Metal on Metal & Ceramic BearingsPS1 SpinePS1 Spine BiomechanicsPS1 Spine - Intervertebral Disc BiomechanicsPS1 Spine - Intervertebral Disc BiologyPS1 Spinal TherapeuticsPS1 Upper Extremity - Shoulder & ElbowPS1 Upper Extremity - Hand & WristPS1 Muscle/NervePS1 Ligament & Tendon MechanicsPS1 Ligament & Tendon BiologyPS2 Bone MechanicsPS2 PublicationsPS2 Cell & Molecular Biomechanics - Physical Effects on CellsPS2 Bone FracturePS2 Fracture FixationPS2 ImagingPS2 Cancer/TumorsPS2 Bone BiologyPS2 Bone - Growth FactorsPS2 Bone - OsteoporosisPS2 Progenitors and Stem CellsPS2 Biomaterials - BioactivePS2 Tissue Engineering - Soft TissuePS2 Cartilage/Meniscus/Synovium - Gene TherapyPS2 Cartilage/Meniscus/Synovium - Cartilage & Matrix ProteinsPS2 Cartilage/Meniscus/Synovium - Immune-Mediated ArthritisPS2 Cartilage/Meniscus/Synovium - Cartilage Matrix DegradationPS2 Cartilage/Meniscus/Synovium - CytokinesPS2 Cartilage/Meniscus/Synovium - Growth Factors & AgingPS2 Cartilage/Meniscus/Synovium - Cartilage RepairPS2 Cartilage/Meniscus/Synovium - Cartilage Repair - Cell-Based ApproachesPS2 Cartilage/Meniscus/Synovium - MeniscusPS2 Cartilage/Meniscus/Synovium - Cell and Molecular Imaging of CartilagePS2 Cartilage/Meniscus/Synovium - OsteoarthritisPS2 Cartilage/Meniscus/Synovium - Cartilage MechanicsPS2 Cartilage/Meniscus/Synovium - MechanobiologyPS2 Disease Process Knee - Joint MechanicsPS2 Disease Process HipPS2 Gait and Kinematics, KinesiologyPS2 Foot and AnklePS2 Infection and InflammationPS2 Total Hip Replacement - Resurfacing and Metal on MetalPS2 Total Knee Reconstruction - General/WearPS2 Arthroplasty - Osteolysis Wear DebrisPS2 Arthroplasty - Implant FixationPS2 Total Knee Replacement - KinematicsPS2 Arthroplasty - Finite Element Analysis - HipPS2 Polyethylene Wear - UHMWPEPS2 Arthroplasty - Finite Element Anlysis KneePS2 Total Hip Replacement ImplantPS2 Implant - Uni-Knee ReplacementPS2 Arthroplasty Hip & Knee - Surgical Navigation and RoboticsPS2 Implant Wear - Vitamin E PolyethylenePS2 Implant Wear - Metal on Metal and Ceramic BearingsPS2 SpinePS2 Spine BiomechanicsPS2 Spine - Intervertebral Disc BiomechanicsPS2 Spine - Intervertebral Disc BiologyPS2 Spine - Spinal TherapeuticsPS2 Upper Extremity - Shoulder and ElbowPS2 Upper Extremity - Hand and WristPS2 MusclePS2 Ligament and Tendon MechanicsPS2 Ligament and Tendon Biology

    /ColorImageDict > /JPEG2000ColorACSImageDict > /JPEG2000ColorImageDict > /AntiAliasGrayImages false /CropGrayImages true /GrayImageMinResolution 300 /GrayImageMinResolutionPolicy /OK /DownsampleGrayImages true /GrayImageDownsampleType /Bicubic /GrayImageResolution 300 /GrayImageDepth -1 /GrayImageMinDownsampleDepth 2 /GrayImageDownsampleThreshold 1.50000 /EncodeGrayImages true /GrayImageFilter /DCTEncode /AutoFilterGrayImages true /GrayImageAutoFilterStrategy /JPEG /GrayACSImageDict > /GrayImageDict > /JPEG2000GrayACSImageDict > /JPEG2000GrayImageDict > /AntiAliasMonoImages false /CropMonoImages true /MonoImageMinResolution 1200 /MonoImageMinResolutionPolicy /OK /DownsampleMonoImages true /MonoImageDownsampleType /Bicubic /MonoImageResolution 1200 /MonoImageDepth -1 /MonoImageDownsampleThreshold 1.50000 /EncodeMonoImages true /MonoImageFilter /CCITTFaxEncode /MonoImageDict > /AllowPSXObjects false /CheckCompliance [ /None ] /PDFX1aCheck false /PDFX3Check false /PDFXCompliantPDFOnly false /PDFXNoTrimBoxError true /PDFXTrimBoxToMediaBoxOffset [ 0.00000 0.00000 0.00000 0.00000 ] /PDFXSetBleedBoxToMediaBox true /PDFXBleedBoxToTrimBoxOffset [ 0.00000 0.00000 0.00000 0.00000 ] /PDFXOutputIntentProfile () /PDFXOutputConditionIdentifier () /PDFXOutputCondition () /PDFXRegistryName () /PDFXTrapped /False

    /CreateJDFFile false /Description > /Namespace [ (Adobe) (Common) (1.0) ] /OtherNamespaces [ > /FormElements false /GenerateStructure false /IncludeBookmarks false /IncludeHyperlinks false /IncludeInteractive false /IncludeLayers false /IncludeProfiles false /MultimediaHandling /UseObjectSettings /Namespace [ (Adobe) (CreativeSuite) (2.0) ] /PDFXOutputIntentProfileSelector /DocumentCMYK /PreserveEditing true /UntaggedCMYKHandling /LeaveUntagged /UntaggedRGBHandling /UseDocumentProfile /UseDocumentBleed false >> ]>> setdistillerparams> setpagedevice