Chondrogenesis,joint formation,and articular cartilage regeneration

The repair of joint surface defects remains a clinical challenge, as articular cartilage has a limited healing response. Despite this, articular cartilage does have the capacity to grow and remodel extensively during pre‐ and post‐natal development. As such, the elucidation of developmental mechanisms, particularly those in post‐natal animals, may shed valuable light on processes that could be harnessed to develop novel approaches for articular cartilage tissue engineering and/or regeneration to treat injuries or degeneration in adult joints. Much has been learned through mouse genetics regarding the embryonic development of joints. This knowledge, as well as the less extensive available information regarding post‐natal joint development is reviewed here and discussed in relation to their possible relevance to future directions in cartilage tissue repair and regeneration. J. Cell. Biochem. 107: 383–392, 2009. © 2009 Wiley‐Liss, Inc.     

Perlecan immunolocalizes to perichondrial vessels and canals in human fetal cartilaginous primordia in early vascular and matrix remodeling events associated with diarthrodial joint development.

The aim of this study was to ascertain how perlecan was localized in human fetal cartilaginous joint rudiment tissues. Perlecan was immunolocalized in human fetal (12-14-week-old) toe, finger, knee, elbow, shoulder, and hip joint rudiments using a monoclonal antibody to domain-1 of perlecan (MAb A76). Perlecan had a widespread distribution in the cartilaginous joint rudiments and growth plates and was also prominent in a network of convoluted hairpin loop-type vessels at the presumptive articulating surfaces of joints. Perlecan was also present in small perichondrial venules and arterioles along the shaft of the developing long bones, small blood vessels in the synovial lining and joint capsules, and in distinctive arrangements of cartilage canals in the knee, elbow, shoulder, and hip joint rudiments. Perlecan was notably absent from CD-31-positive metaphyseal vessels in the hip, knee, shoulder, and fingers. These vessels may have a role in the nutrition of the expanding cell populations in these developing joint tissues and in the establishment of the secondary centers of ossification in the long bones, which is essential for endochondral ossification.     

Toward regeneration of articular cartilage

Articular cartilage is classified as permanent hyaline cartilage and has significant differences in structure, extracelluar matrix components, gene expression profile, and mechanical property from transient hyaline cartilage found in the epiphyseal growth plate. In the process of synovial joint development, articular cartilage originates from the interzone, developing at the edge of the cartilaginous anlagen, and establishes zonal structure over time and supports smooth movement of the synovial joint through life. The cascade actions of key regulators, such as Wnts, GDF5, Erg, and PTHLH, coordinate sequential steps of articular cartilage formation. Articular chondrocytes are restrictedly controlled not to differentiate into a hypertrophic stage by autocrine and paracrine factors and extracellular matrix microenvironment, but retain potential to undergo hypertrophy. The basal calcified zone of articular cartilage is connected with subchondral bone, but not invaded by blood vessels nor replaced by bone, which is highly contrasted with the growth plate. Articular cartilage has limited regenerative capacity, but likely possesses and potentially uses intrinsic stem cell source in the superficial layer, Ranvier's groove, the intra‐articular tissues such as synovium and fat pad, and marrow below the subchondral bone. Considering the biological views on articular cartilage, several important points are raised for regeneration of articular cartilage. We should evaluate the nature of regenerated cartilage as permanent hyaline cartilage and not just hyaline cartilage. We should study how a hypertrophic phenotype of transplanted cells can be lastingly suppressed in regenerating tissue. Furthermore, we should develop the methods and reagents to activate recruitment of intrinsic stem/progenitor cells into the damaged site. Birth Defects Research (Part C) 99:192–202, 2013 . © 2013 Wiley Periodicals, Inc .     

Improved cartilage integration and interfacial strength after enzymatic treatment in a cartilage transplantation model

The objective of the present study was to investigate whether treatment of articular cartilage with hyaluronidase and collagenase enhances histological and mechanical integration of a cartilage graft into a defect. Discs of 3 mm diameter were taken from 8-mm diameter bovine cartilage explants. Both discs and annulus were either treated for 24 hours with 0.1% hyaluronidase followed by 24 hours with 10 U/ml collagenase or left untreated (controls). Discs and annulus were reassembled and implanted subcutaneously in nude mice for 5 weeks. Integration of disc with surrounding cartilage was assessed histologically and tested biomechanically by performing a push-out test. After 5 weeks a significant increase in viable cell counts was seen in wound edges of the enzyme-treated group as compared with controls. Furthermore, matrix integration (expressed as a percentage of the total interface length that was connected; mean ± standard error) was 83 ± 15% in the treated samples versus 44 ± 40% in the untreated controls. In the enzyme-treated group only, picro-Sirius Red staining revealed collagen crossing the interface perpendicular to the wound surface. Immunohistochemical analyses demonstrated that the interface tissue contained cartilage-specific collagen type II. Collagen type I was found only in a small region of fibrous tissue at the level of the superficial layer, and collagen type III was completely absent in both groups. A significant difference in interfacial strength was found using the push-out test: 1.32 ± 0.15 MPa in the enzyme-treated group versus 0.84 ± 0.14 MPa in the untreated controls. The study shows that enzyme treatment of cartilage wounds increases histological integration and improves biomechanical bonding strength. Enzymatic treatment may represent a promising addition to current techniques for articular cartilage repair.     

Generating,growing and transforming skeletal shape: insights from amphibian pharyngeal arch cartilages

Amphibians that undergo a metamorphosis provide an unparalleled opportunity to investigate how skeletal shape is generated, preserved, and transformed in development. Their pharyngeal arch (PA) cartilages, which support breathing and feeding behaviors, form embryonically from cranial neural crest cells, grow isometrically at larval stages, and abruptly change shape during metamorphosis. Further, the shape changes occur in three different ways: some adult cartilages form de novo, others emerge from within resorbing larval cartilages and some larval cartilages reshape themselves at the cellular level. Isometric growth followed by abrupt shape change is unique to amphibian PA cartilages, which suggests that the origin and evolution of amphibian metamorphosis has been influenced by the tissue properties of cartilage. This essay reviews the functional role of the PA skeleton in frogs and salamanders and presents a mechanistic framework for understanding how its shape is generated, preserved, and transformed at the levels of cell behavior and specification mechanisms.     

84 例基于三脚架结构改建的鼻尖综合整形术的临床总结

目的:总结基于鼻翼软骨三脚架结构的改建技术在鼻尖综合整形术中的应用经验。方法:从2012年09月到2015年02月间,共84例求美者在我院进行初次鼻尖综合整形术。3例为男性,81例为女性。年龄20-45岁,平均年龄31.7岁。其中鼻头肥大伴鼻背低平65例,行鼻翼软骨缝合+鼻翼软骨切除+鼻假体+自体软骨帽状移植术;鼻头肥大、鼻背低平伴鼻小柱短小19例,行自体软骨鼻小柱支撑+鼻翼软骨切除+鼻翼软骨缝合+鼻假体植入+自体软骨帽状移植术。结果:84例求美者术后随访1个月-2年,除1例病例鼻头过于肥大,鼻尖形态改善不明显以外,其余求美者鼻额角及鼻尖角度及均较术前有明显改善,鼻小柱短小组的鼻小柱长度也较术前有明显改善。所有病例切口瘢痕均不明显,无明显并发症出现。结论:针对不同鼻翼软骨发育条件下的病人,个性化的应用鼻翼软骨三脚架结构改建的鼻尖综合整形术具有较好的临床效果,须根据不同病人特点选用。     

Perlecan displays variable spatial and temporal immunolocalisation patterns in the articular and growth plate cartilages of the ovine stifle joint

Perlecan is a modular heparan sulphate and/or chondroitin sulphate substituted proteoglycan of basement membrane, vascular tissues and cartilage. Perlecan acts as a low affinity co-receptor for fibroblast growth factors 1, 2, 7, 9, binds connective tissue growth factor and co-ordinates chondrogenesis, endochondral ossification and vascular remodelling during skeletal development; however, relatively little is known of its distribution in these tissues during ageing and development. The aim of the present study was to immunolocalise perlecan in the articular and epiphyseal growth plate cartilages of stifle joints in 2-day to 8-year-old pedigree merino sheep. Perlecan was prominent pericellularly in the stifle joint cartilages at all age points and also present in the inter-territorial matrix of the newborn to 19-month-old cartilage specimens. Aggrecan was part pericellular, but predominantly an extracellular proteoglycan. Perlecan was a prominent component of the long bone growth plates and displayed a pericellular as well as a strong ECM distribution pattern; this may indicate a so far unrecognised role for perlecan in the mineralisation of hypertrophic cartilage. A significant age dependant decline in cell number and perlecan levels was evident in the hyaline and growth plate cartilages. The prominent pericellular distribution of perlecan observed indicates potential roles in cell-matrix communication in cartilage, consistent with growth factor signalling, cellular proliferation and tissue development.     

The nature and significance of invertebrate cartilages revisited: distribution and histology of cartilage and cartilage-like tissues within the Metazoa

Tissues similar to vertebrate cartilage have been described throughout the Metazoa. Often the designation of tissues as cartilage within non-vertebrate lineages is based upon sparse supporting data. To be considered cartilage, a tissue should meet a number of histological criteria that include composition and organization of the extracellular matrix. To re-evaluate the distribution and structural properties of these tissues, we have re-investigated the histological properties of many of these tissues from fresh material, and review the existing literature on invertebrate cartilages. Chondroid connective tissue is common amongst invertebrates, and differs from invertebrate cartilage in the structure and organization of the cells that comprise it. Groups having extensive chondroid connective tissue include brachiopods, polychaetes, and urochordates. Cartilage is found within cephalopod mollusks, chelicerate arthropods and sabellid polychaetes. Skeletal tissues found within enteropneust hemichordates are unique in that the extracellular matrix shares many properties with vertebrate cartilage, yet these tissues are completely acellular. The possibility that this tissue may represent a new category of cartilage, acellular cartilage, is discussed. Immunoreactivity of some invertebrate cartilages with antibodies that recognize molecules specific to vertebrate bone suggests an intermediate phenotype between vertebrate cartilage and bone. Although cartilage is found within a number of invertebrate lineages, we find that not all tissues previously reported to be cartilage have the appropriate properties to merit their distinction as cartilage.     

The histology and affinities of sinacanthid fishes: primitive gnathostomes from the Silurian of China

On the basis of well preserved specimens from the Lower Silurian of the Tarim Basin, Xinjiang Uygur Autonomous Region and Shiqian County, Guizhou Province, People's Republic of China we describe in detail the histological structure of sinacanthid spines, the only known remains of a group of fishes common in Silurian strata from China. The sinacanthids have previously been assigned either to the acanthodians or to the chondrichthyans. The spine structure is composed of an outer layer of atubular dentine and an inner layer of globular calcified cartilage, and the nature and distribution of these tissues indicates that the spines were formed as a result of interaction between the endoskeleton and dermoskeleton. The tissue distribution and style of growth described herein places the sinacanthids crownwards of the placoderms, and possibly within the total group Chondrichthyes. However, before they can be firmly placed within a phylogenetic scheme, further evidence is required both on the general anatomy of sinacanthids and on the nature of chondrichthyan apomorphies.  © 2005 The Linnean Society of London, Zoological Journal of the Linnean Society , 2005, 144 , 379–386.     

Effect of a subset of adipose-derived stem cells isolated with liposome magnetic beads to promote cartilage repair

This study aimed to investigate the ability of CD146⁺ subset of ADSCs to repair cartilage defects. In this study, we prepared CD146⁺ liposome magnetic beads (CD146⁺LMB) to isolate CD146⁺ADSCs. The cells were induced for chondrogenic differentiation and verified by cartilage-specific mRNA and protein expression. Then a mouse model of cartilage defect was constructed and treated by filling the induced cartilage cells into the damaged joint, to evaluate the function of such cells in the cartilage microenvironment. Our results demonstrated that the CD146⁺LMBs we prepared were uniform, small and highly stable, and cell experiments showed that the CD146⁺LMB has low cytotoxicity to the ADSCs. ADSCs isolated with CD146⁺LMB were all CD146⁺, CD105⁺, CD166⁺ and CD73⁺. After chondrogenic induction, the cells showed significantly increased expression of cartilage markers Sox9, collagen Ⅱ and aggrecan at protein level and significantly increased Sox9, collagen Ⅱ and aggrecan at mRNA level, and the protein expression and mRNA expression of CD146⁺ADSCs group were higher than those of ADSCs group. The CD146⁺ADSCs group showed superior tissue repair ability than the ADSCs group and blank control group in the animal experiment, as judged by gross observation, histological observation and histological scoring. The above results proved that CD146⁺LMB can successfully isolate the CD146⁺ADSCs, and after chondrogenic induction, these cells successfully promoted repair of articular cartilage defects, which may be a new direction of tissue engineering.     

鼻中隔软骨联合耳软骨雕塑鼻尖在假体隆鼻中应用效果

目的:探讨假体隆鼻时应用鼻中隔软骨联合耳软骨雕塑鼻尖的临床效果。方法:选取2013年1月至2014年3月在我院进行隆鼻术的患者98例,按照随机数表法将其分成对照组和实验组,每组49人。对照组患者采用单纯的假体隆鼻,实验组患者采用假体隆鼻并使用鼻中隔软骨联合耳软骨雕塑鼻尖。评价手术的满意程度及手术前后患者的鼻长、鼻尖高度、鼻尖角的变化情况。结果:实验组患者的治疗满意度为91.84%,明显高于对照组的40.82%,差异具有统计学意义(P0.05)。实验组患者术后鼻长、鼻尖高度、鼻尖角均优于手术前(P0.05),且显著优于对照组,差异均有统计学意义(P0.05)。结论:应用鼻中隔软骨联合耳软骨雕塑鼻尖在假体隆鼻中具有良好的应用效果,满意度较高,值得在临床上推广使用。     

In vitro extracellular matrix accumulation of nasal and articular chondrocytes for intervertebral disc repair

Chondrocyte based regenerative therapies for intervertebral disc repair such as Autologous Disc Cell Transplantation (ADCT, CODON) and allogeneic juvenile chondrocyte implantation (NuQu^®, ISTO Technologies) have demonstrated good outcomes in clinical trials. However concerns remain with the supply demand reconciliation and issues surrounding immunoreactivity which exist for allogeneic-type technologies. The use of stem cells is challenging due to high growth factor requirements, regulatory barriers and differentiation towards a stable phenotype. Therefore, there is a need to identify alternative non-disc cell sources for the development and clinical translation of next generation therapies for IVD regeneration. In this study, we compared Nasal Chondrocytes (NC) as a non-disc alternative chondrocyte source with Articular Chondrocytes (AC) in terms of cell yield, morphology, proliferation kinetics and ability to produce key extracellular matrix components under 5% and 20% oxygen conditions, with and without exogenous TGF-β supplementation.Results indicated that NC maintained proliferative capacity with high amounts of sGAG and lower collagen accumulation in the absence of TGF-β supplementation under 5% oxygen conditions. Importantly, osteogenesis and calcification was inhibited for NC when cultured in IVD-like microenvironmental conditions. The present study provides a rationale for the exploration of nasal chondrocytes as a promising, potent and clinically feasible autologous cell source for putative IVD repair strategies.     

Primary cilia: Versatile regulator in cartilage development

Cartilage is a connective tissue in the skeletal system and has limited regeneration ability and unique biomechanical reactivity. The growth and development of cartilage can be affected by different physical, chemical and biological factors, such as mechanical stress, inflammation, osmotic pressure, hypoxia and signalling transduction. Primary cilia are multifunctional sensory organelles that regulate diverse signalling transduction and cell activities. They are crucial for the regulation of cartilage development and act in a variety of ways, such as react to mechanical stress, mediate signalling transduction, regulate cartilage‐related diseases progression and affect cartilage tumorigenesis. Therefore, research on primary cilia‐mediated cartilage growth and development is currently extremely popular. This review outlines the role of primary cilia in cartilage development in recent years and elaborates on the potential regulatory mechanisms from different aspects.     

Growth differentiation factor 5 in cartilage and osteoarthritis: A possible therapeutic candidate

Growth differentiation factor 5 (GDF‐5) is essential for cartilage development and homeostasis. The expression and function of GDF‐5 are highly associated with the pathogenesis of osteoarthritis (OA). OA, characterized by progressive degeneration of joint, particularly in cartilage, causes severe social burden. However, there is no effective approach to reverse the progression of this disease. Over the past decades, extensive studies have demonstrated the protective effects of GDF‐5 against cartilage degeneration and defects. Here, we summarize the current literature describing the role of GDF‐5 in development of cartilage and joints, and the association between the GDF‐5 gene polymorphisms and OA susceptibility. We also shed light on the protective effects of GDF‐5 against OA in terms of direct GDF‐5 supplementation and modulation of the GDF‐5‐related signalling. Finally, we discuss the current limitations in the application of GDF‐5 for the clinical treatment of OA. This review provides a comprehensive insight into the role of GDF‐5 in cartilage and emphasizes GDF‐5 as a potential therapeutic candidate in OA.     

Articular cartilage vesicles contain RNA

Small membrane-bound extracellular organelles known as articular cartilage matrix vesicles (ACVs) participate in pathologic mineralization in osteoarthritic articular cartilage. ACVs are also present in normal cartilage, although they have no known functions other than mineralization. Recently, RNA was identified in extracellular vesicles derived from mast cells, suggesting that such vesicles might carry coding information from cell to cell. We found that ACVs from normal porcine and human articular cartilage and primary chondrocyte conditioned media contained 1 μg RNA/80 μg ACV protein. No DNA could be detected. RT-PCR of ACV RNA demonstrated the presence of full length mRNAs for factor XIIIA, type II transglutaminase, collagen II, aggrecan, ANKH and GAPDH. RNA in intact ACVs was resistant to RNase, despite the fact that ACV preparations contained measurable levels of active RNases. Significantly, radiolabeled RNA in ACVs could be transferred to unlabeled chondrocytes by co-incubation and produced changes in levels of chondrocyte enzymes and proteins. The demonstration that ACVs contain mRNAs suggests that they may function to shuttle genetic information between articular cells and indicate novel functions for these structures in articular cartilage.     

Study on the Microstructure of Human Articular Cartilage/Bone Interface

For improving the theory of gradient microstructure of cartilage/bone interface, human distal femurs were studied. Scanning Electron Microscope (SEM), histological sections and MicroCT were used to observe, measure and model the microstructure of cartilage/bone interface. The results showed that the cartilage/bone interface is in a hierarchical structure which is composed of four different tissue layers. The interlocking of hyaline cartilage and calcified cartilage and that of calcified cartilage and subchondral bone are in the manner of “protrusion-pore” with average diameter of 17.0 μm and 34.1 μm respectively. In addition, the cancellous bone under the cartilage is also formed by four layer hierarchical structure, and the adjacent layers are connected by bone trabecula in the shape of H, I and Y, forming a complex interwoven network structure. Finally, the simplified structure model of the cartilage/bone interface was proposed according to the natural articular cartilage/bone interface. The simplified model is a 4-layer gradient biomimetic structure, which corresponds to four different tissues of natural cartilage/bone interface. The results of this work would be beneficial to the design of bionic scaffold for the tissue engineering of articular cartilage/bone.     

Sequence of chondrocranial development in the oriental fire bellied toad Bombina orientalis

The vertebrate head as a major novelty is directly linked to the evolutionary success of the vertebrates. Sequential information on the embryonic pattern of cartilaginous head development are scarce, but important for the understanding of its evolution. In this study, we use the oriental fire bellied toad, Bombina orientalis, a basal anuran to investigate the sequence and timing of larval cartilaginous development of the head skeleton from the appearance of mesenchymal Anlagen in post-neurulation stages until the premetamorphic larvae. We use different methodological approaches like classic histology, clearing and staining, and antibody staining to examine the larval skeletal morphology. Our results show that in contrast to other vertebrates, the ceratohyals are the first centers of chondrification. They are followed by the palatoquadrate and the basihyal. The latter later fuses to the ceratohyal and the branchial basket. Anterior elements like Meckel's cartilage and the rostralia are delayed in development and alter the ancestral anterior posterior pattern observed in other vertebrates. The ceratobranchials I–IV, components of the branchial basket, follow this strict anterior–posterior pattern of chondrification as reported in other amphibians. Chondrification of different skeletal elements follows a distinct pattern and the larval skeleton is nearly fully developed at Gosner Stage 28. We provide baseline data on the pattern and timing of early cartilage development in a basal anuran species, which may serve as guidance for further experimental studies in this species as well as an important basis for the understanding of the evolutionary changes in head development among amphibians and vertebrates.