Tissue Engineering Nanotechnology and applicationsSUBMITTED BY :VISHWAMITRA KUMARROLL NO : CH8276Liu Nanobionics Lab

OutlineWhat is Tissue EngineeringWhat is NanotechnologyWhy we apply Nanotechnology to Tissue EngineeringHow is Nanotechnology applied to Tissue EngineeringDifferent nanofabrication techniquesApplications

Tissue EngineeringRemove cells from the body.Expand number in cultureSeed onto an appropriate scaffold with suitable growth factors and cytokinesPlace into cultureRe-implant engineered tissue repair damaged site

Tissue EngineeringDifferentiated cellsAdult stem cellsEmbryonic stem cells Dynamic cell seedingImproved mass transferMechanical stimuliHydrogelsNanofibrous scaffoldsSelf-assembling scaffoldsSolid freeform fabricated scaffoldsSmall moleculesGrowth factors/polypeptidesNucleic acids (DNA, siRNA, and antisense oligonucleotides)Nanotechnology and Tissue Engineering: The Scaffold, CRC Press; 1 edition (June 16, 2008)

Nanotechnology OverviewNanotechnology is a branch of science and engineering which deals with structures and devices in nanometer scale.Create & ManipulateView & CharacterizeOptical MicroscopyElectron MicroscopyAtomic Force MicroscopyNanofabrication: (Top down & Bottom up)Lithography (Optical, E-beam, NIP, DPL)Etching (Wet Etching, Plasma Etching)Deposition (Evaporation, PECVD, Electrochemical Deposition, Sputtering, etc.)Epitaxial growth (MOCVD, MBE, etc.)

Why we apply Nanotech in TE?Cells on microfibrous scaffolds have a polarized relationship, with one side of the cell attached to the scaffold, the other exposed to physiological media. In comparison, it is likely that cells are more naturally constrained by nanofibrous scaffolds.

Nanofibrous ScaffoldElectrospinningSelf-Assembly

ElectrospinningThis process involves the ejection of a charged polymer fluid onto an oppositely charged surface.Multiple polymers can be combinedControl over fiber diameter and scaffold architecture

Research on Parameters of Electrospinning ProcessSolution propertiesViscosityConductivitySurface tensionPolymer molecular weightDipole momentDielectric constantControlled variablesFlow rateElectric field strengthDistance between tip and collectorNeedle tip designCollector composition and geometryAmbient parametersTemperatureHumidityAir velocity

Tissue Engineering. May 2006, 12(5): 1197-1211.

Research on MaterialsPolyglycolic acid (PGA)Highly crystalline, hydrophilic, byproduct is glycolic acid Polylactic acid (PLA)Hydrophobic, lower melting temperature, byproduct is lactic acidPolydioxanone (PDO)Highly crystallinePolycaprolactone (PCL)Semi-crystalline properties, easily co-polymerized, byproduct caproic acidBlendsPGA-PLAPGA-PCLPLA-PCLPDO-PCL

Elastin Gelatin collagenFibrillar collagenCollagen blendsFibrinogen Synthetic polymersPGA, PLA and PLGA most commonly usedPDO most similar to Elastin collagen blend (limited by shape memory)PCL most elastic and mixed frequenlty with other material sProvide nanoscale physical featuresNatural polymersCollagen Type I & III + PDO: best possible match for blood vesselsAdvanced Drug Delivery Reviews Volume 59, Issue 14, 10 December 2007, Pages 1413-1433

Self AssemblyFigure 1: Fabrication of various peptide materials.Figure 2: Self-assembling peptides form a three-dimensional scaffold woven from nanofibers ~ 10 nm in diameter.Nature Biotechnology 21, 1171 - 1178 (2003) Representation of self-assembling peptide. Electron micrograph of three-dimensional scaffold formed in vitro. Rat hippocampal neurons form active nerve connections; each green dot represents a single synapsis. Neural cells from a rat hippocampal tissue slide migrate on the three-dimensional peptide scaffold. Cells on the polymer membrane (left) and on the peptide scaffold (right) are shown. Both glial cells (green) and neural progenitors (red) migrate into the three-dimensional peptide scaffold.Brain damage repair in hamster. The peptide scaffold was injected into the optic nerve, which was first severed with a knife. The cut was sealed by the migrating cells after 2 days. A great number of neurons form synapses. Chondrocytes from young and adult bovine encapsulated in the peptide scaffold. These cells not only produce a large amount of glycosaminoglycans (purple) and type II collagen (yellow), characteristic materials found in cartilage, but also a cartilage-like tissue in vitro53.Adult rat liver progenitor cells encapsulated in the peptide scaffold. The cells on the two-dimensional dish did not produce cytochrome P450type enzymes (left). However, cells in three-dimensional scaffolds showed cytochrome P450 activity (right).

Figure 3: Lipid, peptide and protein scaffold nanowires.Figure 4: Microlenses and fiber-optics fabricated from protein scaffolds.Self AssemblyNature Biotechnology 21, 1171 - 1178 (2003)

Self-Assembling Peptide Scaffolds for Regenerative MedicineSAPNS heals the brain in young animals.SAPNS allows axons to regenerate through the lesion site in brain.PNAS March 28, 2006 vol. 103 no. 13 5054-5059

Phase separationThis process involves dissolving of a polymer in a solvent at a high temperature followed by a liquidliquid or solidliquid phase separation induced by lowering the solution temperatureCapable of wide range of geometry and dimensions include pits, islands, fibers, and irregular pore structuresSimpler than self-assemblya) powder, b) scaffolds with continuous network, c) foam with closed poresSEM of nanofibrous scaffold with interconnected spherical macroporesAdvanced Drug Delivery Reviews Volume 59, Issue 14, 10 December 2007, Pages 1413-1433

Carbon NanotubeCell tracking and labelingSensing cellular behaviorAugmenting cellular behaviorAugmenting cellular behaviorCytotoxicityMurine myoblast stem cells incubated with DNA-encapsulated nanotubesneuron bridging an array of carbon nanotubes thereby creating neural networks.Biomaterials Volume 28, Issue 2, January 2007, Pages 344-353

Block CoploymerSynthetic scheme of block copolymers.Science 30 May 1997: Vol. 276. no. 5317, pp. 1401 - 1404Nature 388, 860-862 (28 August 1997) Gelsol transition curves.In vitro release profile of FITC-labelled dextran (Mr 20,000) from PEOPLLAPEO (Mr 5,0002,0405,000) triblock copolymer. Injectable drug-delivery system

Printing TechnologyNanoimprinting LithographyOrgan PrintingContact Printing

Nanoimprinting LithographyProf. Stephen Y. ChouThermal-sensitive PolymerOptical-sensitive Polymer

Nanopattern-induced changes in morphology and motility of smooth muscle cellsSMC morphologyAlignment and elongation characterizationWound healing assay for cell motilityBrdU cell proliferation assayBiomaterials Volume 26, Issue 26, September 2005, Pages 5405-5413

Organ printing: computer-aided jet-based 3D tissue engineeringFig. 1. Fusion of embryonic myocardial ring. Myocardium rings were cut fromStage 1516 HH chick ventricle, containing only myocardium, endocardium andsome intervening matrix. Isolated rings beat steadily for several days; (a) adjacentapposed rings fused overnight and (b) beat as one. (c). Schematic representationof principle of organ printing technology: placing of cell aggregates layer by layerin solidifying thermo-reversible gel with sequential cell aggregate fusion andmorphing into 3D tube.Fig. 2. Cell printer and images of printed cells and tissue constructs.Fig. 3. (a) Printed bagel-like ring that consists of several layers of sequentially(layer-by-layer) deposited collagen type 1 gel. (b) Manually printed living tube withradial branches from the chick 27stage HH embryonic heart cushion tissue placedin 3D collagen type 1 gel.Trends Biotechnol. 2003 Apr;21(4):157-61.

Contact PrintingJ. Am. Chem. Soc., 2005, 127 (48), pp 1677416775Advanced Materials Volume 19 Issue 24, Pages 4338 - 4342

SummaryNanofibrous ScaffoldElectrospinningSelf-AssemblyNanoporous ScaffoldPhase SeparationCarbon NanotubeBlock CopolymerPrintingNanoimprinting Lithography Organ Printing Contact Printing

Lab Chip, 2004, 4, 98 - 103

APPLICATIONS1)Coronary Heart Disease Myocardial InfarctionCongestive Heart FailureDysfunctional Heart ValvesPeripheral Vascular DisordersAbdominal Aortic Aneurysms2)Neurological StrokeParkinsons DiseaseAlzheimers DiseaseEpilepsyTraumatic Brain and Spinal Cord InjuryMultiple Sclerosis3)OrthopedicNon-union FracturesCartilage Damage and RepairLigament DamageVertebral Disc DamageBone Graft Materials4)UrologicalIncontinenceKidney DiseaseBladder5)Skin/IntegumentaryBurnsDiabetic UlcersVenous Ulcerslastic Surgery

6)DentalMissing teethPeriodontal disease7)Organ TransplantationLiverHeartKidneyPancreas8)OphthalmologyCorneaRetina9)GastrointestinalEsophagusStomachSmall IntestineColon10)Ear, Nose and Throat/Respiratory/CardiopulmonaryTracheaRespiratory Epithelial Cells (Nasal Turbinates)

11)CancerUrology (Bladder, kidney/Renal cell, prostate)Neurology (Brain/Glioblastoma/Glioma, CNS, head/Neck)Obstetrics/Gynecology (Breast, pelvic, ovarian, endometrial)Orthopedic (Chordoma/Bone)Gastrointestinal/Gastroenterology (Colorectal, gastric, pancreas)ENT (Esophageal, oral, pharynx, olfactory)Hematopoietic (Leukemia, lymphoma)Respiratory (Lung/Mesothelioma)Dermatology (Melanoma/Skin)

Carbon nanotubes are among the numerous candidates for tissue engineering scaffolds since they are biocompatible, resistant tobiodegradation and can be functionalized withbiomolecules. However, the possibility of toxicity with non-biodegradable nano-materials is not fully understood.

ReferencesNanotechnology and Tissue Engineering: The Scaffold, CRC Press; 1 edition (June 16, 2008)Quynh P. Pham, Upma Sharma, Ph.D., Dr. Antonios G. Mikos, Electrospinning of Polymeric Nanofibers for Tissue Engineering Applications: A Review, Tissue Engineering. May 2006, 12(5): 1197-1211.Catherine P. Barnes, Scott A. Sell, Eugene D. Boland, David G. Simpson, Gary L. Bowlin, Nanofiber technology: Designing the next generation of tissue engineering scaffolds, Advanced Drug Delivery Reviews, Volume 59, Issue 14, Intersection of Nanoscience and Modern Surface Analytical Methodology, 10 December 2007, Pages 1413-1433, ISSN 0169-409X, DOI: 10.1016/j.addr.2007.04.022. Shuguang Zhang, Fabrication of novel biomaterials through molecular self-assembly, Nature Biotechnology 21, 1171 - 1178 (2003) Rutledge G. Ellis-Behnke, Yu-Xiang Liang, Si-Wei You, David K. C. Tay, Shuguang Zhang, Kwok-Fai So, and Gerald E. Schneider, Nano neuro knitting: Peptide nanofiber scaffold for brain repair and axon regeneration with functional return of vision PNAS 2006 103 (13) 5054-5059Benjamin S. Harrison, Anthony Atala, Carbon nanotube applications for tissue engineering, Biomaterials, Volume 28, Issue 2, Cellular and Molecular Biology Techniques for Biomaterials Evaluation, January 2007, Pages 344-353, ISSN 0142-9612, DOI: 10.1016/j.biomaterials.2006.07.044.Miri Park, Christopher Harrison, Paul M. Chaikin, Richard A. Register, Douglas H. Adamson, Block Copolymer Lithography: Periodic Arrays of ~1011 Holes in 1Square Centimeter, Science 30 May 1997: Vol. 276. no. 5317, pp. 1401 - 1404Byeongmoon Jeong, You Han Bae, Doo Sung Lee and Sung Wan Kim, Biodegradable block copolymers as injectable drug-delivery systems, Nature 388, 860-862 (28 August 1997)Evelyn K.F. Yim, Ron M. Reano, Stella W. Pang, Albert F. Yee, Christopher S. Chen, Kam W. Leong, Nanopattern-induced changes in morphology and motility of smooth muscle cells, Biomaterials, Volume 26, Issue 26, September 2005, Pages 5405-5413, ISSN 0142-9612, DOI: 10.1016/j.biomaterials.2005.01.058.Mironov V, Boland T, Trusk T, Forgacs G, Markwald RR.Organ printing: computer-aided jet-based 3D tissue engineering. Trends Biotechnol. 2003 Apr;21(4):157-61. Yu, A. A.; Stellacci, F., Contact Printing beyond Surface Roughness: Liquid Supramolecular Nano-Stamping, Advanced Materials, 19, 4338-4342, 2007Yu A.A., Savas T., Cabrini S., diFabrizio E., Smith H.I., Stellacci F., High resolution printing of DNA features on poly(methyl methacrylate) substrates using supramolecular nano-stamping, J. Am. Chem. Soc., 127, 16774-16775, 2005

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