Preparation And Properties Of Novel Scaffold For Cartilage Tissue Engineering

Tissue engineering technology which made modern medicine step out of the scope o f organ transplantion and enterd into a new era of organ reconstruction emerged in the 1 980s of last century.The technology of cartilage tissue engineering which serves as a imp ortant branch of tissue engineering developed fast and made progress in recent years.How ever,many key issues still needed to be resolved and the most important question is to d evelop a new scaffold.In this dissertation,we initially used a new type of biodegradable material-P(3-hydro xybutyrate-co-3-hydroxyoctanoate)[PHBHOx] for preparation of cartilage tissue engineering scaffold,PHBHOx which served as main material was composited with acellular cartilage matrix(ACM) to fabricate cartilage tissue engineering scaffold,the properties of PHBHOx/ ACM scaffold was measured and evaluated,the target was to successfully fabricate compo site scaffold which meet the needs of cartilage tissue engineering and expand the materia1 areas of cartilage tissue engineering scaffold.Above all,fresh hog articular cartilage from knee joints was sliced in axenic conditio ns and then was treated sequentially with Triton X-100,hypotonic solution Tris-HCl,DNA enzyme,RNA enzyme for decellularization,the ACM was assessed by HE, Masson and To luidine Blue staining ultimately.After that,the ACM was shattered into powder in low tem perature and was composited with PHBHOx in different ratio[0%(control group),2%,4 %,6%,8%,10%] to fabricate scaffold with solvent-casting,particulate-leaching methods using NaCl particles(particle diameter was between100um and 300um) as pore-foaming ag ent.The aperture,porosity, hydrophilic property,degradation ratio,pH stabilization of the biod egradable fluid and biological mechanical strength of the scaffold was measured via diffe rent methods in order to screening the most excellent scaffold in properties which meet t he needs of cartilage tissue engineering. Chondrocytes isolated from hog articular cartilage was cultured and the third generation chondrocytes was used in in vitro experiment.After2 weeks'culture,we used scanning electron microscope(SEM) and MTT test to observe and analyse the growth state of chondrocytes on scaffold.The result revealed that HE staining have shown that the effect of decellularization was very good,Masson and Toluidine Blue staining have shown that the extracellular mat rix (collagen and glycosaminoglycan) was well reserved,the scaffold was three-dimension al and looked like sponge under SEM observation,image analyzer showed that the averag e aperture of scaffold was 206.91±52.33um,special gravity method showed that the porosi ty of scaffold was between (71.51±4.66)% and (83.53±3.21)%,the hydrophilic propert y of scaffold was all above 100%,the degradation property of scaffold performed excellen tly in in vitro degradation experiment,the pH value of the degradated solution was very s table,the biological mechanical strength of scaffold was between (3.20±0.41) MPa and (7. 90±0.22) MPa by using a universal testing machine,the properties mentioned above all co uld be regulated by altering the porogenic agent or composite ratio of material.Hog chon drocytes cultured with enzyme digestion method has complementary advantages in cell nu mber and livability and could be a stable method for cell culture.In in vitro experiment of 2 weeks,chondrocytes on the scaffold adhered tightly,part of chondrocytes have held o ut pseudopodium and stretched on the surface of scaffold under SEM observation,MTT te st proved that chondrocytes grew well in the scaffold.All of the experiment results proved that the novel PHBHOx/ACM scaffold had exc ellent properties and met the needs of cartilage tissue engineering and could be used as a new material for cartilage tissue engineering.