The Effect Of Cyclic Pressure At Different Amplitudes On Tissue Engineering Cartilage

ObjectiveTo explore the effect of cyclic pressure at different amplitudes on theconstruction of tissue-engineering cartilage in bioreactor.MethodsKill 4 3-to 5-day-old New Zealand rabbits. Under sterile conditions,articular cartilage was taken from the rabbits .And shear it to the size ofabout 1mm×1mm×1mm. Then chondrocytes were isolated from thearticular cartilage by digesting with 0.25% trypsin and 0.2% typeⅡcollagenase in sequence. After expansion in vitro, the 2nd passagechondrocytes were seeded into PLGA scaffolds with the size of5mm×5mm and cover slips. After adherence, the scaffolds and cover slipswere both divided into 4 groups,each group had 10 scaffolds.All sampleswere cultured in bioreactor(Celdyne HFB-40).Four different cultureconditions were examined: a control group(no pressure),a 200Kpa group(0～200kPa, 0.1Hz),a 100Kpa group(0～100Kpa, 0.1Hz),a 50Kpagroup(0～50Kpa, 0.1Hz).Each group was cultured for 2 weeks. After twoweeks, the level of GAG in each sample was examined. Macroscopicalanalysis, histological analysis of HE stain and TB stain,andimmunohistochemical stain were performed.ResultThere were significant differences among the 4 groups. The 200kPagroup was morphologically the thickest and has smooth surface. Thehistology observation of HE stain shows: the cartilage celles weresurrounded by amount of extracellular matrix, multiplicated in mostprosperity, arranged close and were well-distributed. TB stain shows theEMC was colored with deep blue. Immunohistochemical stain showspositive dyeing with buffy. Also the typeⅡcollagen dyeing area waslageest(82.44±1.87%)and produced the most GAG(476.33±25.66μg) in 4groups, the 100Kpa group following closely, the control group is the worst.The samples of group was morphologically the thinnest and the surfacewas no smooth. The histology observation of HE stain shows: the cartilagecelles were surrounded by little of extracellular matrix, multiplicatedslowly, arranged disperse and were aniso-distributed. TB stain shows theEMC was colored with blue. Immunohistochemical stain shows positivedyeing with buffy. But the typeⅡcollagen dyeing area was smallest(41.05±6.28%) and produced the least GAG(220.00±12.14μg) in 4 groups. ConclusionThe results indicate that the cyclic pressure(0～200kPa, 0.1Hz) bestpromotes chondrocytes to synthesis and secrete extracellular matrix just asGAG and typeⅡcollagen in 4 groups.This suggests that cyclic pressure atdifferent amplitudes have important effects on metabolism of tissue--engineering cartilage.