Engineering Stromal Tissue For Use In Clinical Cornea Transplantation Kelsey Henderson, Quinn Otte, Matthew Lopez “Where there is no vision, there is no hope.” –George Washington Carver
Engineering Stromal Tissue For Use In Clinical Cornea Transplantation
Kelsey Henderson, Quinn Otte, Matthew Lopez
“Where there is no vision, there is no hope.” –George Washington Carver
The EyeSensory organPart of the
nervous system
http://www.buzzle.com/articles/causes-of-absent-red-reflex-in-newborns.html
The CorneaFunctions: • Protection• Oxygen diffusion • Refraction of light
http://www.aafp.org/afp/2013/0115/p114.html
The StromaRelatively thick- 465μmKeratocyctes
Cornea Diseases/ProblemsFuch’s dystrophy: KeratoconisMacular corneal dystrophyCorneal ulcers Injury
http://www.uhbristol.nhs.uk/patients-and-visitors/your-hospitals/bristol-eye-hospital/bristol-laser-vision/what-is-keratoconus/
Current TreatmentDonor transplant: keratoplasty
Penetrating Endothelial Deep Anterior Lamellar
Problems:Lack of
donorsLimited
lifespan of tissue
Potential for disease transfer http://www.patienteducationcenter.org/articles/corneal-transplant/
Engineering Considerations
Biocompatibility
Degradation
Cell Sources
Material Choice
Optical Clarity
Oxygen Diffusion
Major ConsiderationsOxygen Diffusion
No vascular network Oxygen is supplied to cornea by air
Optical Clarity Parallel fibrils within lamella Orthogonally aligned lamellae Constant diameter of lamellae Water content of stroma
Degradation Material should degrade at the same rate that cells are
proliferating
Current synthetic optionsBiocompatibility
Clarity Degradation
Cell adhesionandproliferation
Oxygen diffusion
Mechanical strength
ACMP ✔ ✖ ✔ -- -- ✔PVA-AM ✔ ✔ -- -- -- ✖PVA-COL ✔ ✔ -- ✔ -- ✔CV ✔ ✔ ✔ ✔ -- ✔Silk Fibroin
✔ ✔ ✔ ✔ ✔ ✔
Our proposed optionHarvest DPCs from
patientSynthesize silk fibroin
sheetsCouple silk fibroin
with RGDhttp://www.prolifebiobank.com/dental_stem_cells.php
Short time water vapor annealing followed by enzymatic pretreatment with protease XIV
Our proposed optionDifferentiate DPCs then seed cells onto silk fibroinCulture silk sheets and layer to stromal thicknessReady for implantation by clinician
http://www.techautos.com/2012/10/02/researchers-announce-artificial-cornea-clinical-trials-under-way/
Testing our strategyIn vitro:
Biocompatibility testsPresence of corneal fibroblasts and scar tissueVascularization and haze
Differentiation testsGene and protein expression
Degradation tests
Testing our strategyIn vitro:
Cell compatibility/seeding testsProduction of ECM components such as types I, V,
and VI collagen, proteoglycans, keratocan, lumican and keratan sulfate
Live/dead assayClarity tests
Uniform size/spacing of lamellaeParallel orientation of fibrilsOrthogonal orientation of lamellae
Gas diffusion tests
Testing our strategyIn vivo:
BiocompatibilityECM productionDegradation
https://www.jax.org/jax-mice-and-services/find-and-order-jax-mice
ConclusionsThe need for cornea tissue is continually increasing
Current treatment involves autologous donor sourcesStromal layer is most difficult to engineerMajor considerations include oxygen perfusion,
optical clarity and degradationA silk biomaterial is proposed along with dental
pulp stem cells differentiated into keratocytesEventual goal is to create an engineered corneal
tissue that can be used in clinical applications
Questions?Thank you for your attention