骨关节炎(osteoarthritis,OA)是最常见的慢性致残性关节疾患,目前尚无针对病因的有效治疗手段。程序性坏死在多种疾病中扮演关键角色,受体相互作用蛋白质激酶3(receptor-interacting protein kinase 3,RIP3)是程序性坏死进程的关键调控...骨关节炎(osteoarthritis,OA)是最常见的慢性致残性关节疾患,目前尚无针对病因的有效治疗手段。程序性坏死在多种疾病中扮演关键角色,受体相互作用蛋白质激酶3(receptor-interacting protein kinase 3,RIP3)是程序性坏死进程的关键调控因子。有研究显示,RIP3在人与鼠骨关节炎退变软骨组织中表达水平显著上调,提示程序性坏死的发生,但RIP3在软骨中的具体病生理角色仍不明确。本研究拟对过表达RIP3前后的软骨细胞转录物组进行测序分析,探索RIP3在骨关节炎进程中发挥作用的具体机制。RNA测序结果显示,RIP3的过表达诱发软骨细胞中244个基因表达上调,277个表达下调。通过进一步构建基因间共表达作用网络,筛选出16个候选靶基因在mRNA水平进行验证,证实RIP3对磷脂酰肌醇3激酶调节亚单位5(phosphoinositide-3-kinase,regulatory subunit 5,Pik3r5)、整合素β3(integrin subunit beta 3,Itgb3)及成髓细胞瘤转录因子第2亚型(MYB proto-oncogene like 2,Mybl2)的表达上调作用最为显著。CCK-8以及乳酸脱氢酶活性检测结果表明,利用siRNA沉默Itgb3的表达可显著抑制RIP3诱发的软骨细胞活力下降及程序性坏死,同时也抑制了RIP3对软骨细胞中分解代谢相关基因Mmp1、Mmp13与Il6的表达上调作用,以及其对合成代谢相关基因Acan、Col2a1与Sox9的下调作用。本研究证实,RIP3通过上调软骨细胞中Itgb3的表达诱发软骨细胞坏死与软骨基质代谢紊乱,并最终导致软骨退变,为骨关节炎的临床治疗提供了新靶点,同时进一步明确了程序性坏死的病理生理学意义。展开更多
By analyzing recent schemes for preparing the chain cluster state,we find that the existence of long-range couplings between qubits will induce a quick decay of the fidelity of the cluster state.We propose some scheme...By analyzing recent schemes for preparing the chain cluster state,we find that the existence of long-range couplings between qubits will induce a quick decay of the fidelity of the cluster state.We propose some schemes for eliminating the effect of the next-nearest-neighbor interactions in the XY and Ising models by dynamical decoupling method.In our approach,the undesired next-nearest-neighbor interactions are effectively suppressed and the fidelity of the generated cluster state is greatly improved by the pulse operations.展开更多
Background Remodeling of the anterior cruciate ligament （ACL） graft usually takes longer than expected. Gene therapy offers a radical different approach to remodeling of the graft. In this study, the internal riboso...Background Remodeling of the anterior cruciate ligament （ACL） graft usually takes longer than expected. Gene therapy offers a radical different approach to remodeling of the graft. In this study, the internal ribosome entry site （IRES） sequence was used to construct a new recombinant adenovirus which permits co-expression of transforming growth factor-β1 （TGFβ1） and vascular endothelial growth factor 165 （VEGF165） genes （named Ad-VEGF165-IRES-TGFβ1）. We investigated the effects of the new adenovirus on the migration of and matrix synthesis by ACL fibroblasts. Methods Adenoviral vector containing TGFβ1 and VEGF165 genes was constructed. ACL fibroblasts were obtained from New Zealand white rabbits. After ACL fibroblasts were exposed to Ad-VEGF165-IRES-TGFβ1, the expression of VEGF165 and TGFβ1 proteins were assessed by enzyme-linked immunosorbent assay （ELISA） and Western blotting analysis. Bioassay of VEGF165 and TGFβ1 proteins were assessed by Western blotting analysis. Proliferation and migration of ACL fibroblasts were assessed by in vitro wound closure assay. Gene expression of collagen type Ⅰ, collagen type Ⅲ, and fibronectin mRNA among matrix markers were assessed by real-time PCR. Results The results showed the successful construction of a recombinant co-expression adenovirus vector containing TGFβ1 and VEGF165 genes. Co-expression of TGFβ1 and VEGF165 can induce relatively rapid and continuous proliferation of ACL fibroblasts and high gene expression of collagen type Ⅰ, collagen type Ⅲ, and fibronectin mRNA among matrix markers. Conclusion Co-expression of TGFβ1 and VEGF165 genes has more powerful and efficient effects on the migration of and matrix synthesis by ACL fibroblasts.展开更多
Background In vitro chondrocyte expansion is a major challenge in cell-based therapy for human articular cartilage repair. Classical culture conditions usually use animal serum as a medium supplement, which raises a n...Background In vitro chondrocyte expansion is a major challenge in cell-based therapy for human articular cartilage repair. Classical culture conditions usually use animal serum as a medium supplement, which raises a number of undesirable questions. In the present study, two kinds of defined, serum-free media were developed to expand chondrocytes in monolayer culture for the purpose of cartilage tissue engineering. Methods Bovine chondrocytes were expanded in serum-free media supplemented with fibroblast growth factor-2 and platelet-derived growth factor or fibroblast growth factor-2 and insulin-like growth factor. Expansion culture in a conventional 10% fetal bovine serum （FBS） medium served as control. Fibronectin coating was used to help cell adhesion in serum-free medium. Next, in vitro three-dimensional pellet culture was used to evaluate the chondrocyte capacity. Cell pellets were expanded in different media to re-express the differentiated phenotype （re-differentiation） and to form cartilaginous tissue. The pellets were assessed by glycosaminoglycans contents, collagen II, collagen I and collagen X immunohistological staining. Results Chondrocytes cultured in serum-free media showed no proliferation difference than cells grown with 10% FBS medium. In addition, chondrocytes expanded in both serum-free media expressed more differentiated phenotypes at the end of monolayer culture, as indicated by higher gene expression ratios of collagen type II to collagen type I. Pellets derived from chondrocytes cultured in both serum-free media displayed comparable chondrogenic capacities to pellets from cells expanded in 10% FBS medium. Conclusion These findings provide alternative culture approaches for chondrocytes in vitro expansion, which may benefit the clinical use of autologous chondrocytes implantation.展开更多
基金Supported by the National Natural Science Foundation of China under Grant Nos 60921091 and 10874170the K.C.Wong Education Foundation,Hong Kong.
文摘By analyzing recent schemes for preparing the chain cluster state,we find that the existence of long-range couplings between qubits will induce a quick decay of the fidelity of the cluster state.We propose some schemes for eliminating the effect of the next-nearest-neighbor interactions in the XY and Ising models by dynamical decoupling method.In our approach,the undesired next-nearest-neighbor interactions are effectively suppressed and the fidelity of the generated cluster state is greatly improved by the pulse operations.
文摘Background Remodeling of the anterior cruciate ligament （ACL） graft usually takes longer than expected. Gene therapy offers a radical different approach to remodeling of the graft. In this study, the internal ribosome entry site （IRES） sequence was used to construct a new recombinant adenovirus which permits co-expression of transforming growth factor-β1 （TGFβ1） and vascular endothelial growth factor 165 （VEGF165） genes （named Ad-VEGF165-IRES-TGFβ1）. We investigated the effects of the new adenovirus on the migration of and matrix synthesis by ACL fibroblasts. Methods Adenoviral vector containing TGFβ1 and VEGF165 genes was constructed. ACL fibroblasts were obtained from New Zealand white rabbits. After ACL fibroblasts were exposed to Ad-VEGF165-IRES-TGFβ1, the expression of VEGF165 and TGFβ1 proteins were assessed by enzyme-linked immunosorbent assay （ELISA） and Western blotting analysis. Bioassay of VEGF165 and TGFβ1 proteins were assessed by Western blotting analysis. Proliferation and migration of ACL fibroblasts were assessed by in vitro wound closure assay. Gene expression of collagen type Ⅰ, collagen type Ⅲ, and fibronectin mRNA among matrix markers were assessed by real-time PCR. Results The results showed the successful construction of a recombinant co-expression adenovirus vector containing TGFβ1 and VEGF165 genes. Co-expression of TGFβ1 and VEGF165 can induce relatively rapid and continuous proliferation of ACL fibroblasts and high gene expression of collagen type Ⅰ, collagen type Ⅲ, and fibronectin mRNA among matrix markers. Conclusion Co-expression of TGFβ1 and VEGF165 genes has more powerful and efficient effects on the migration of and matrix synthesis by ACL fibroblasts.
文摘Background In vitro chondrocyte expansion is a major challenge in cell-based therapy for human articular cartilage repair. Classical culture conditions usually use animal serum as a medium supplement, which raises a number of undesirable questions. In the present study, two kinds of defined, serum-free media were developed to expand chondrocytes in monolayer culture for the purpose of cartilage tissue engineering. Methods Bovine chondrocytes were expanded in serum-free media supplemented with fibroblast growth factor-2 and platelet-derived growth factor or fibroblast growth factor-2 and insulin-like growth factor. Expansion culture in a conventional 10% fetal bovine serum （FBS） medium served as control. Fibronectin coating was used to help cell adhesion in serum-free medium. Next, in vitro three-dimensional pellet culture was used to evaluate the chondrocyte capacity. Cell pellets were expanded in different media to re-express the differentiated phenotype （re-differentiation） and to form cartilaginous tissue. The pellets were assessed by glycosaminoglycans contents, collagen II, collagen I and collagen X immunohistological staining. Results Chondrocytes cultured in serum-free media showed no proliferation difference than cells grown with 10% FBS medium. In addition, chondrocytes expanded in both serum-free media expressed more differentiated phenotypes at the end of monolayer culture, as indicated by higher gene expression ratios of collagen type II to collagen type I. Pellets derived from chondrocytes cultured in both serum-free media displayed comparable chondrogenic capacities to pellets from cells expanded in 10% FBS medium. Conclusion These findings provide alternative culture approaches for chondrocytes in vitro expansion, which may benefit the clinical use of autologous chondrocytes implantation.
