Differences in the effect of two injectable Pluronic F-127 hydrogel composites in repairing rabbit cartilage defects / 中国组织工程研究

BACKGROUND: Preliminary experiments show that bone marrow mesenchymal stem cells transfected with SOX9 gene can grow and proliferate in Pluronic F-127 hydrogel, promote the secretion of extracellular matrix, and increase the expression of cartilage matrix. OBJECTIVE: The SOX9 gene was transduced into bone marrow mesenchymal stem cells by lentivirus gene induction, and then combined with injectable Pluronic F-127 hydrogel to observe the effect of Pluronic F-127 hydrogel on repairing cartilage defects. METHODS: SOX9 gene was transfected into bone marrow mesenchymal stem cells by lentivirus gene induction. After 48 hours of transfectlon, SOX9 gene was combined with Pluronic F-127 hydrogel. Sixty New Zealand white rabbits provided by the Experimental Animal Center of Wuhan University of Science and Technology were selected to establish the models of femoral condylar cartilage defect of the right knee joint. The rabbits were randomly divided into three groups: model group without implantation of any material at the defect site, control group with implantation of non-transfected bone marrow mesenchymal stem cells and Pluronic F-127 hydrogel complex at the defect site, and experimental group with implantation of SOX9 gene-transfected bone marrow mesenchymal stem cells and Pluronic F-127 hydrogel complex at the defect site. Four and twelve weeks after operation, the defect tissues were taken for three-dimensional reconstruction of micro-CT, hematoxylin-eosin staining, Safranine O staining, type II collagen immunohistochemical staining and Wakitani soft tissue repair histological score. This study was approved by the Ethics Committee of Wuhan University of Science and Technology. RESULTS AND CONCLUSION: (1) At 12 weeks after operation, three-dimensional reconstruction of Micro-CT showed that there was no obvious repair In the defect area of the model group, and there was still a large depression In the center. In the control group, the central depression area was significantly reduced and more trabecular structures of regenerated bone were observed. In the experimental group, the defect area was basically repaired. (2) At 12 weeks after operation, hematoxylin-eosin staining showed that there was no trabecular bone structure, disordered cell distribution and no cartilage lacunae at the defect area of the model group. In the control group, more bone tissue was reconstructed, and the defect area was mainly filled with cartilage-like tissue and fibrous tissue. In the experimental group, bone tissue was reconstructed adequately, and the defect area was mainly filled with chondroid cells and chondroid extracellular matrix. Cells arranged columnariy, similar to the surrounding cartilage. (3) At 12 weeks after surgery, Safranine O staining and collagen II immunohistochemical staining results showed that a small amount of glycosamlnoglycan was observed, but no type II collagen was found in the model group. The expression of glycosaminoglycan and type II collagen was more in the control group. The expression of glycosaminoglycan and type II collagen was highest In the experimental group compared with the other two groups. (4) The histological score of Wakitani soft tissue repair in the experimental group was higher than that in the control group and model group (P < 0.05). (5) The results suggested that Pluronic F-127 hydrogel complex loaded with SOX9 gene transfected bone marrow mesenchymal stem cells can promote the repair of cartilage defects.

Effects of the Chinese herbal compound, Tengmei decoction, on inflammatory injury in joints of rat models of type Ⅱcollagen-induced arthritis / 中国比较医学杂志

Objective To explore the effects of Chinese herbal compound, Tengmei decoction, on type II collagen-induced arthritis ( CIA) in rats, and to examine the changes of arthritis index ( AI) , limb swelling, joint tissue inflammatory infiltration, and the effects on immune-inflammatory factors.Methods Sprague-Dawley rat models of arthritis were successfully established by intradermal injection of type II collagen and Freund’ s complete adjuvant.The model rats were randomly divided into model group, positive drug group, and high-and low-dose Chinese medicine groups, 6 rats in each group.The intervention and treatment period was 12 weeks.To measure weekly the anteroposterior and transverse diameters of the rear ankles and wrists, the transverse diameter of the claw foot palm pad, the thickness and the highest point width of hind limb plantar joint swelling, and to evaluate the integrated scores of joints and limbs swelling using a vernier caliper.Results ①Compared with the normal group, the total arthritis scores and hind limbs AI scores of the model group were significantly increased ( P <0.05 or P <0.01 ) .The left forelimb AI scores were significantly increased during 10 -12 weeks ( P <0.05 ) .The anteroposterior and transverse diameters of the left hind limb, the thickness of the highest point measurement of the left hind foot pad metatarsal were significantly increased ( P<0.05 or P<0.01) in different time periods between 1-12 weeks.Compared with the model group, the total scores and the left hind limb joints AI scores of the high-and low-dose drug groups were decreased after 6 weeks (P<0.05).②Compared with the normal control group, levels of mRNA transcription and protein expression of IL-6 and TNF-αwere significantly up-regulated ( P<0.01 ) in the model group.Compared with the model group, the levels of mRNA transcription and the expression of IL-6 and TNF-αproteins were significantly down-regulated in the positive group and Chinese medicine groups ( P <0.01 ) .③ Histological examination showed that the low-dose TCM significantly improved the CIA synovial hyperplasia and inflammatory cell infiltration.Conclusions The molecular mechanism of Chinese herbal compound Tengmei decotion in improving joint pathological injury of CIA rat models may be related to its inhibitory effect on the high expression of immune-inflammatory factors in the synovial tissue of CIA rats.

Delayed Long Bone Formation in Hyperthermia-exposed Mouse Embryos / 체질인류학회지

Maternal hyperthermia, which is currently confirmed as one of major causative factors inducing growth retardation, congenital anomalies and abortion, is known to influence normal development of CNS and various organ system. In addition, maternal hyperthermia could induce severe developmental defects including development of the limb. However, it is not clearly identified how maternal hyperthermia affects the expression of chondrogenesis-related proteins in developing limb of mouse. Thus, this study is aimed to investigate the effects of the maternal hyperthermia on the expression of a various proteins in developing upper limb. To elucidate it, ICR mice were used in this study, and the animals were divided into control and heat shock groups. The heat shock treatment was given to embryonic day (ED) 8. The animals were sacrificed on ED 11, 13, 15 and 17, and the humerus were removed. Chondrogenesis-related factors such as FGF8, SOX9 and collagen II were detected on ED 11, 13 and 15 using western blot and immunohistochemistry. Developing humerus on ED 17 was stained with alizarin red S and alcian blue. The expression of FGF8 of heat shock groups was continued even though the development was succeeded. SOX9 expression in heat shock groups was significantly elevated on ED 13 compared to the control embryos. In addition, collagen II expression of heat groups was significantly higher than that of the control group on ED 13 and 15. The results of this study suggest that hyperthermia causes delayed endochondral ossification in long bone through continuous expression of FGF8, SOX9 and collagen II proteins even though the endochondral ossification is succeeded.

Delayed Long Bone Formation in Hyperthermia-exposed Mouse Embryos / 체질인류학회지

Maternal hyperthermia, which is currently confirmed as one of major causative factors inducing growth retardation, congenital anomalies and abortion, is known to influence normal development of CNS and various organ system. In addition, maternal hyperthermia could induce severe developmental defects including development of the limb. However, it is not clearly identified how maternal hyperthermia affects the expression of chondrogenesis-related proteins in developing limb of mouse. Thus, this study is aimed to investigate the effects of the maternal hyperthermia on the expression of a various proteins in developing upper limb. To elucidate it, ICR mice were used in this study, and the animals were divided into control and heat shock groups. The heat shock treatment was given to embryonic day (ED) 8. The animals were sacrificed on ED 11, 13, 15 and 17, and the humerus were removed. Chondrogenesis-related factors such as FGF8, SOX9 and collagen II were detected on ED 11, 13 and 15 using western blot and immunohistochemistry. Developing humerus on ED 17 was stained with alizarin red S and alcian blue. The expression of FGF8 of heat shock groups was continued even though the development was succeeded. SOX9 expression in heat shock groups was significantly elevated on ED 13 compared to the control embryos. In addition, collagen II expression of heat groups was significantly higher than that of the control group on ED 13 and 15. The results of this study suggest that hyperthermia causes delayed endochondral ossification in long bone through continuous expression of FGF8, SOX9 and collagen II proteins even though the endochondral ossification is succeeded.

Transforming Growth Factor-beta1 and Bone Morphogenetic Protein-2 Upregulates Matrix Synthesis and Chondrogenic Phenotype in Intervertebral Disc Cells / 대한척추외과학회지

OBJECTIVES: To determine effect of transforming growth factor-beta1 and bone morphogenetic protein-2 in matrix synthesis and expression of chondrogenic phenotype in human intervertebral disc cells. MATERIALS AND METHODS: The intervertebral disc cells were harvested and cultured from the surgical patients for the degenerative disc disease. TGF-beta1 was purchased from R&D and BMP-2 was produced by transfection of pcDNA3.1/Hygro/BMP-2 to CHO cell using Lipofectamine 2000. rhBMP-2 was separated by Heparin-Sepharose A chromatography. TGF-bata1 and BMP-2 were administered to culture. Proteoglycan synthesis was assessed by 35S incorporation and expression of matrix mRNA was analyzed by RT-PCR for collagen I, collagen II, aggrecan, and osteocalcin. RESULTS: TGF-bata1 and BMP-2 showed increased proteoglycan synthesis and expression of collagen I, collagen II and aggrecan mRNA in dose dependent manner respectively. There was no recognizable synergistic effect in matrix synthesis and matrix mRNA expression. Throughout dosage, expression of osteogenic phenotype (osteocalcin mRNA) was not noted. CONCLUSION: TGF-beta1 and BMP-2 proved to be effective anabolic agent for maximizing matrix synthesis without evidence of osteogenesis.