Objective To evaluate the feasibility and the value of the layered cylindric collagenhydroxyapatite composite as a scaffold for the cartilage tissue engineering after an observation of how it absorbs the chondrocytes and affe cts the cell behaviors. Methods The chondrocytes were isolated and multiplied in vitro, and then the chondrocytes were seeded onto the porous collagen／h ydro xyapatite composite scaffold and were cultured in a three-dimensional environme n t for 3 weeks. The effects of the composite scaffold on the cell adhesivity, proliferation, morphological changes, and synthesis of the extracellular matrix were observed by the phase-contrast microscopy, histology, scanning electron micros copy, and immunohistochemistry. Results The pore diameter of the upper layer of the collagen-hydroxyapatite composite scaffold was about 147 μm. and the porosity was 89%; the pore diameter of the bottom layer was about 85 μm and the porosity was 85%. The layered cylindric collagenhydroxyapatite composite scaffold had good hydrophilia. The chondrocytes that adhered to the surface of the scaffold, proliferated and migrated into the scaffold after 24 hours. The chondrocytesattached to the wall of the microholes of the scaffold maintained a rounded morphology and could secrete the extracellular matrix on the porous scaffold. Conclusion The layered cylindric collagenhydroxyapatite composite scaffold has a good cellular compatibility, and it is ber in the mechanical property than the pure collagen. It will be an ideal scaffold for the cartilage tissue enginee ring.
In order to investigate the effect of motion on repairing articular cartilage defect following autogenous periosteal graft, sixty adult rabbits were divided randomly into three groups: out-cage motion (OCM), in-cage motion (ICM) and immobilization (IMM). A defect of the articular cartilage, 1 cm x 0.5 cm in size, was made in the patellar-groove of femur of each hind limb. Free autogenous periosteal graft from the proximal tibia was sutured on the base of the left defect, while the right limb was served as control. The animals were sacrificed at 4, 8 and 12 weeks, respectively, after operation. The regeneration of the cartilage implanted was observed through gross, histology, histochemical assay and electronic microscope. The influence of different amount of motion on the chondrogenesis from the periosteal implant was also compared. The result showed that the hyaline cartilage produced from periosteal implant could be capable to repair full-thickness of articular cartilage. From statistical study, there was significant difference between OCM and ICM groups (P lt; 0.05), ICM and IMM (P lt; 0.05) as well as OCM and IMM (P lt; 0.01). It was suggested that the periosteal graft was effective in repair of defect of articular cartilage and the amount of motion was important for chondrogenesis.
ObjectiveTo explore the feasibility of three-dimensional (3D) bioprinted adipose-derived stem cells (ADSCs) combined with gelatin methacryloyl (GelMA) to construct tissue engineered cartilage.MethodsAdipose tissue voluntarily donated by liposuction patients was collected to isolate and culture human ADSCs (hADSCs). The third generation cells were mixed with GelMA hydrogel and photoinitiator to make biological ink. The hADSCs-GelMA composite scaffold was prepared by 3D bioprinting technology, and it was observed in general, and observed by scanning electron microscope after cultured for 1 day and chondrogenic induction culture for 14 days. After cultured for 1, 4, and 7 days, the composite scaffolds were taken for live/dead cell staining to observe cell survival rate; and cell counting kit 8 (CCK-8) method was used to detect cell proliferation. The composite scaffold samples cultured in cartilage induction for 14 days were taken as the experimental group, and the composite scaffolds cultured in complete medium for 14 days were used as the control group. Real-time fluorescent quantitative PCR (qRT-PCR) was performed to detect cartilage formation. The relative expression levels of the mRNA of cartilage matrix gene [(aggrecan, ACAN)], chondrogenic regulatory factor (SOX9), cartilage-specific gene [collagen type Ⅱ A1 (COLⅡA1)], and cartilage hypertrophy marker gene [collagen type ⅩA1 (COLⅩA1)] were detected. The 3D bioprinted hADSCs-GelMA composite scaffold (experimental group) and the blank GelMA hydrogel scaffold without cells (control group) cultured for 14 days of chondrogenesis were implanted into the subcutaneous pockets of the back of nude mice respectively, and the materials were taken after 4 weeks, and gross observation, Safranin O staining, Alcian blue staining, and collagen type Ⅱ immunohistochemical staining were performed to observe the cartilage formation in the composite scaffold.ResultsMacroscope and scanning electron microscope observations showed that the hADSCs-GelMA composite scaffolds had a stable and regular structure. The cell viability could be maintained at 80%-90% at 1, 4, and 7 days after printing, and the differences between different time points were significant (P<0.05). The results of CCK-8 experiment showed that the cells in the scaffold showed continuous proliferation after printing. After 14 days of chondrogenic induction and culture on the composite scaffold, the expressions of ACAN, SOX9, and COLⅡA1 were significantly up-regulated (P<0.05), the expression of COLⅩA1 was significantly down-regulated (P<0.05). The scaffold was taken out at 4 weeks after implantation. The structure of the scaffold was complete and clear. Histological and immunohistochemical results showed that cartilage matrix and collagen type Ⅱ were deposited, and there was cartilage lacuna formation, which confirmed the formation of cartilage tissue.ConclusionThe 3D bioprinted hADSCs-GelMA composite scaffold has a stable 3D structure and high cell viability, and can be induced differentiation into cartilage tissue, which can be used to construct tissue engineered cartilage in vivo and in vitro.
Objective To construct recombinant lentiviral expression vectors of porcine transforming growth factor β1 (TGF-β1) gene and transfect bone marrow mesenchymal stem cells (BMSCs) so as to provide TGF-β1 gene-modified BMSCs for bone and cartilage tissue engineering. Methods The TGF-β1 cDNA was extracted and packed into lentiviral vector, and positive clones were identified by PCR and gene sequencing, then the virus titer was determined. BMSCs were isolated frombone marrow of the 2-month-old Bama miniature pigs (weighing 15 kg), and the 2nd and 3rd generations of BMSCs wereharvested for experiments. BMSCs were then transfected by TGF-β1 recombinant lentiviral vectors (TGF-β1 vector group)respectively at multi pl icity of infection (MOI) of 10, 50, 70, 100, and 150; then the effects of transfection were detected bylaser confocal microscope and Western blot was used to determine the optimal value of MOI. BMSCs transfected by empty vector (empty vector group) and non-transfected BMSCs (non-transfection group) were used as control group. RT-PCR, immunocytochemistry, and ELISA were performed to detect the expressions of TGF-β1 mRNA, TGF-β1 protein, and collagen type II. Results Successful construction of recombinant lentiviral vectors of porcine TGF-β1 gene was identified by PCR and gene sequencing, and BMSCs were successfully transfected by TGF-β1 recombinant lentiviral vectors. Green fluorescence was observed by laser confocal microscope. Western blot showed the optimal value of MOI was 70. The expression of TGF-β1 mRNA was significantly higher in TGF-β1 vector group than in empty vector group and non-transfection group (P lt; 0.05). Immunocytochemistry results revealed positive expression of TGF-β1 protein and collagen type II in BMSCs of TGF-β1 vector group, but negative expression in empty vector group and non-transfection group. At 21 days after transfection, high expression of TGF-β1 protein still could be detected by ELISA in TGF-β1 vector group. Conclusion TGF-β1 gene can be successfully transfected into BMSCs via lentiviral vectors, and long-term stable expression of TGF-β1 protein can be observed, prompting BMSCs differentiation into chondrocytes.
Objective To observe the long-term clinical results of repairing large articular cartilage defects of the hip and the knee with free autogeneous periosteum. Methods Based on the results of experimental studies, the authors used free autogeneous periosteum transplantation and postoperative continuous passive motion (CPM) to repair large articular cartilaginous defects in 52 patientsfrom February 1987 to August 1995. Of 37 patients with complete follow-up data, 16 had congenital dislocation of the hip, 6traumatic arthritis of hip, 1 femoral head destruction following mild infection, 2 ankylosing spondylitis, 6 intra-articular fracture of the knee, 4 arthritisof the knee and 2 stiff knee following joint infection. The patients with dislocation of hip were given relieving traction before operation. The cartilages of pathological changes were excised to bleeding bone. The defects were repairedwith periosteum removing from tibia. CPM were immediately applied for 4-6 weeksand no bearing was allowed 6 months after discharge. The silicon membrane was taken out in the 6th month. Results Thirty-seven patients (17 males, 20 females) were followed up 7-15 years with an average of 10.5 years. The functional evaluation referred to joint pain degree，joint mobile range，daily activity and X-ray findings. The results were excellence in 11 patients , good in 18 patients , poor in 8 patients. Conclusion The method to repair articular cartilage defect with free autogeneous -periosteum is effective and may be applied clinically.
Cartilage surface fibrosis is an early sign of osteoarthritis and cartilage surface damage is closely related to load. The purpose of this study was to study the relationship between cartilage surface roughness and load. By applying impact, compression and fatigue loads on fresh porcine articular cartilage, the rough value of cartilage surface was measured at an interval of 10 min each time and the change rule of roughness before and after loading was obtained. It was found that the load increased the roughness of cartilage surface and the increased value was related to the load size. The time of roughness returning to the initial condition was related to the load type and the load size. The impact load had the greatest influence on the roughness of cartilage surface, followed by the severe fatigue load, compression load and mild fatigue load. This article provides reference data for revealing the pathogenesis of early osteoarthritis and preventing and treating articular cartilage diseases.
Objective To construct a new type of self-assembling peptide nanofiber scaffolds—RGDmx, and to study the cell compatibility of the new scaffolds and the proliferation and chondrogenic differentiation of precartilaginous stem cells(PSCs) in scaffolds. Methods PSCs were separated and purified from newborn Sprague Dawley rats by magnetic activated cell sorting and indentified by immunohistochemistry and immunofluorescent staining. The RGDmx were constructed by mixing KLD-12 and KLD-12-PRG at volume ratio of 1 ∶ 1. PSCs at passage 3 were seeded into the KLD-12 scaffold (control group) and RGDmx scaffold (experimental group). The proliferation of PSCs in 2 groups were observed with the method of cell counting kit (CCK) -8 after 1, 3, 7, and 14 days after culture. The RGDmx were constructed by mixing KLD-12-PRG and KLD-12 at different volume ratios of 0, 20%, 40%, 60%, 80%, and 100% and the prol iferation of PSCs was also observed. The complete chondrogenic medium (CCM) was used to induce chondrogenic differentiation of PSCs in different scaffolds. The differentiation of PSCs was observed by toluidine blue staining and RT-PCR assay. Results PSCs were separated and purified successfully, which were identified by immunohistochemistry and immunofluorescent staining methods. The results of CCK-8 showed that the absorbance (A) value in the experimental group increased gradually and reached the highest at 7 days; the A value in the experimental group was significantly higher than that in the control group at 7 days and 14 days (P lt; 0.05). Meanwhile, the A value in the RGDmx scaffold with a volume ratio of 40% was significantly higher than those in others (P lt; 0.05). After 14 days of induction culture with CCM, the toluidine blue staining results were positive in 2 groups; the results of RT-PCR showedthat the expression levels of collagen type II and the aggrecan in the experimental group were significantly higher than those in the control group (P lt; 0.05). Conclusion The self-assembling peptide nanofiber scaffold—RGDmx is an ideal scaffold for tissue engineer because it has good cell compatibility and more effective properties of promoting the differentiation of PSCs to chondrocytes.
ObjectiveTo investigate the clinical efficacy of glucosamine hydrochloride tablets in treating knee cartilage injury resulting from rheumatoid arthritis. MethodsWe selected 200 knee cartilage injury patients with rheumatoid arthritis treated in our hospital from January 2011 to January 2015 as the research subjects. They were divided into control group (n=98) and observation group (n=102) according to the time of admission. The control group was treated with conventional disease modifying anti-rheumatic drugs (DMARDs), while the observation group was treated with glucosamine hydrochloride tablets on the basis of DMARDs. The treatment effect was evaluated and compared between the two groups of patients 18, 36 and 54 weeks after the treatment. ResultsFifty-four weeks later, knee pain score of the observation group was better than that of the control group, and the difference was statistically significant (P ＜ 0.05) . The observation group had a lower Noyes evaluation level than the control group, and the difference was statistically significant (P ＜ 0.05) . Adverse reaction in the observation group was 3.92% and it was 3.06% in the control group, and the difference between the two groups was not statistically significant (P ＞ 0.05) . ConclusionGlucosamine hydrochloride tablets combined with conventional anti-rheumatic treatment is effective for knee cartilage injury caused by rheumatoid arthritis, which can promote cartilage repair, and it is worthy of clinical application.
ObjectiveTo review the imaging evaluation, treatment progress, and controversy related to developmental dysplasia of the hip (DDH) in adolescents and adults. Methods The domestic and abroad hot issues related to adolescents and adults with DDH in recent years, including new imaging techniques for assessing cartilage, controversies over the diagnosis and treatment of borderline DDH (BDDH), and the improvement and prospect of peracetabular osteotomy (PAO) were summarized and analyzed. ResultsDDH is one of the main factors leading to hip osteoarthritis. As the understanding of the pathological changes of DDH continues to deepen, the use of delayed gadolinium-enhanced MRI of cartilage can further evaluate the progress of osteoarthritis and predict the prognosis after hip preservation. There are still controversies about the diagnosis and treatment of BDDH. At the same time, PAO technology and concepts are still being improved. ConclusionCartilage injury and bony structure determine the choice of surgical methods and postoperative prognosis of hip preservation surgery. The hip preservation of adolescent and adult DDH patients will move towards the goal of individualization and accuracy.
ObjectiveTo review clinical application and research progress of different types of intelligent responsive hydrogels in repairing articular cartilage injury. MethodsThe animal experiments and clinical studies of different types of intelligent responsive hydrogels for repairing articular cartilage injury were summarized by reviewing relevant literature at home and abroad. ResultsThe intrinsic regenerative capacity of articular cartilage following injury is limited. Intelligent responsive hydrogels, including those that are temperature-sensitive, light-sensitive, enzyme-responsive, pH-sensitive, and other stimuli-responsive hydrogels, can undergo phase transitions in response to specific stimuli, thereby achieving optimal functionality. These hydrogels can fill the injured cartilage area, promote the proliferation and differentiation of chondrocytes, and expedite the repair of the damaged site. With advancements in cartilage tissue engineering materials research, intelligent responsive hydrogels offer a novel approach and promising potential for the treatment of cartilage injuries. ConclusionIntelligent responsive hydrogel is a kind of flexible, controllable, efficient, and stable polymer, which has similar structure and functional properties to articular cartilage, and has become one of the important biomaterials for cartilage repair. However, there is still a lack of unified treatment standards and simple and efficient preparation technology.