Native hyaluronan produces less hypersensitivity than cross-linked hyaluronan.

Hyaluronan has been used in patients with osteoarthritis to relieve the painful symptoms associated with this condition. The native form of hyaluronan and artificially cross-linked forms of hyaluronan (such as Hylan G-F 20) are widely used brands that are approved by the Food and Drug Administration for use in patients with osteoarthritis. Clinical evidence suggests that some of these hyaluronan products may induce an antigenic reaction in some patients.

Preincubation of tissue engineered constructs enhances donor cell retention.

Cartilage tissue engineering has been the focus of considerable research. However, the fate of transplanted donor cells rarely is explored directly. In the current study, the effect of preincubating perichondrial cells into a polylactic acid scaffold before implantation into an osteochondral defect was studied.

Adhesion of perichondrial cells to a polylactic acid scaffold.

The number of chondrogenic cells available locally is an important factor in the repair process for cartilage defects. Previous studies demonstrated that the number of transplanted rabbit perichondrial cells (PC) remaining in a cartilage defect in vivo, after being carried into the site in a polylactic acid (PLA) scaffold, declined markedly within two days. This study examined the ability of in vitro culture of PC/PLA constructs to enhance subsequent biomechanical stability of the cells and the matrix content in an in vitro screening assay.

Tissue-engineered repair of osteochondral defects: effects of the age of donor cells and host tissue.

Transplantation of a tissue-engineered construct containing cells of a chondrocytic phenotype into an osteochondral defect provides a biological solution to this type of injury. Among the factors that affect cell proliferation and phenotypic expression, age is one that has not been well characterized. In this study adult and aged male donor cells, derived from perichondrium, were cultured and adsorbed into a polylactic acid (PLA) scaffold and transplanted into osteochondral defects created in adult (8- to 10-month-old) and aged (4- to 5-year-old) female rabbits.

Matrices for cartilage repair.

Techniques for repairing focal articular cartilage defects are evolving from methods that induce a local stimulation of fibrocartilaginous repair to methods that will lead to a hyaline articular cartilage repair. Mosaicplasty and autologous chondrocyte implantation are examples of the latter. A tissue engineered hyaline cartilage implant that could be used off the self would minimize the morbidity of these techniques.

Chondrocyte apoptosis and regional differential expression of nitric oxide in the medial meniscus following partial meniscectomy.

Partial medial meniscectomy leads to tibial articular cartilage degeneration. Nitric oxide (NO) production increases with the development of osteoarthritis (OA) and has been shown to have a catabolic effect on chondrocytes. Since distribution of chondrocytic and fibroblastic cell types within the total cell population comprising meniscus is region-specific, we compared NO production in the peripheral and central regions of the medial meniscus 12 weeks after partial medial meniscectomy and assessed chondrocyte apoptosis and NO production in the tibial articular cartilage.

Donor cell fate in tissue engineering for articular cartilage repair.

Articular cartilage repair is a clinical challenge because of its limited intrinsic healing potential. Considerable research has focused on tissue engineering and transplantation of viable chondrogenic cells to enhance cartilage regeneration. However, the question remains: do transplanted allogenic cells survive in the repair with time? This study assessed donor cell fate after transplantation of male New Zealand White rabbit perichondrium cell and polylactic acid constructs into osteochondral defects created in the medial femoral condyles of female New Zealand White rabbits.

High-efficiency non-viral transfection of primary chondrocytes and perichondrial cells for ex-vivo gene therapy to repair articular cartilage defects.

Background: Primary perichondrial cells and chondrocytes have been used to repair articular cartilage defects in tissue engineering studies involving various animal models. Transfection of these cells with a gene that induces chondrocytic phenotype may form an ideal method to affect tissue engineering of articular cartilage.

Design: A protocol for high-efficiency transfection of primary perichondrial and cartilage cells was optimized.

The tetrabasic KKKK(147-150) motif determines intracrine regulatory effects of PthrP 1-173 on chondrocyte PPi metabolism and matrix synthesis.

Expression of PTHrP is a major regulator of growth cartilage development and also becomes robust in osteoarthritic cartilage. We further defined how PTHrP 1-173, which we observed to be the preferentially expressed PTHrP isoform in normal and osteoarthritic cartilage, functions in chondrocytes. We transfected both immortalized human juvenile costal chondrocytes (TC28 cells) and rabbit articular chondrocytes with wild-type PTHrP 1-173 and mutants of putative PTHrP 1-173 endoproteolytic processing sites.

Integrin expression is upregulated during early healing in a canine intrasynovial flexor tendon repair and controlled passive motion model.

To explore crucial early molecular events involved in contact healing of the intrasynovial flexor tendon, integrin expression was evaluated at the transcriptional and post-transcriptional levels during the first two weeks following injury, repair and controlled passive motion in a canine model. Specifically, immunohistochemical and reverse transcription polymerase chain reaction (RT-PCR) techniques were employed to evaluate expression of the fibronectin, vitronectin and endothelial cell binding integrin receptor subunits alpha5, alphav and alpha6, along with the common beta1 subunit. The two techniques revealed increasing expression of the four subunits over the two week post-repair period.

Nonviral in vivo gene therapy for tissue engineering of articular cartilage and tendon repair.

Heretofore, nonviral methods have been used primarily for in vitro transfection of cultured cell lines. These methods were substantially less efficient when compared with the use of viruses, particularly when used in vivo. Herein a three-step, highly efficient method of nonviral gene delivery is presented.

Novel method for the quantitative assessment of cell migration: a study on the motility of rabbit anterior cruciate (ACL) and medial collateral ligament (MCL) cells.

A novel method of quantitating cell migration has been proposed for the potential utilization of tissue engineered scaffolds. Applying Alt's conservation law to describe the motion of first passage ACL and MCL cells, we have developed a quantitative method to assess innate differences in the motility of cells from these two ligamentous tissues. In this study, first passage ACL and MCL cells were cultured from four mature New Zealand white rabbits.

Transforming growth factor beta one (TGF-beta 1) enhancement of the chondrocytic phenotype in aged perichondrial cells: an in vitro study.

Background: Perichondrium is recognized as a tissue with chondrogenic potential yielding cells which can be used for osteochondral repair. Factors which influence the proliferative ability and chondrocytic phenotype of such cells include age and presence of specific growth factors, i.e.

PT-12, a putative ras-activated proliferation-dependent gene, is expressed in patellar tendon and not in anterior cruciate ligament.

We describe a gene (PT-12) that is expressed in the patellar tendon and not in the anterior cruciate ligament. We used a recently developed polymerase chain reaction-based subtractive cDNA analysis to discover genes that are overexpressed in the patellar tendon but not expressed in the anterior cruciate ligament; the long-term objective was to find genes that are central to the self-repair of the patellar tendon, in contrast with the inability of the anterior cruciate ligament to launch a repair response following injury. PT-12 is a homologue of human S2 or mouse LLRep3 ribosomal genes, which are known to be overexpressed in highly proliferating cells.

Regulation of alpha(v)beta3 and alpha5beta1 integrin receptors by basic fibroblast growth factor and platelet-derived growth factor-BB in intrasynovial flexor tendon cells.

Integrins are important players in soft tissue healing as molecules that mediate communication between cells and extracellular matrix. Thus, the regulation of the expression of these molecules would be important during wound repair. To explore the regulatory roles of specific growth factors on integrin expression by intrasynovial flexor tendon cells, the present study assessed the in vitro effects of basic fibroblast growth factor and platelet derived growth factor-BB on expression of the alpha5beta1 and alpha(v)beta3 integrins in these cells.

The effects of hyaluronan on matrix metalloproteinase-3 (MMP-3), interleukin-1beta(IL-1beta), and tissue inhibitor of metalloproteinase-1 (TIMP-1) gene expression during the development of osteoarthritis.

Objective: To assess the influence of intra-articular injection of hyaluronan (HA) on expression of matrix metalloproteinase-3 (MMP-3), interleukin-1beta(IL-1beta), and tissue inhibitor of metalloproteinase-1 (TIMP-1) in cartilage and synovium during the process of osteoarthritis (OA).

Design: Eighteen mature New Zealand white rabbits underwent unilateral anterior cruciate ligament transection (ACLT) and were divided into two groups. The first group (HA injection group) received 0.

Age-related changes in the expression of cadherin-11, the mesenchyme specific calcium-dependent cell adhesion molecule.

Cadherin-11 is a calcium-dependent cell adhesion molecule that is expressed in cells of the mesenchymal lineage during embryonic development. In this study we show, for the first time, that cadherin-11 gene is expressed in the bone marrow and bone cells obtained from rabbits of various age groups. Furthermore, a quantitative measurement of gene expression revealed that cadherin-11 was expressed in young rabbits (6 week-old: open epiphysis) at a level of 6.

Bone morphogenetic proteins and bFGF exert opposing regulatory effects on PTHrP expression and inorganic pyrophosphate elaboration in immortalized murine endochondral hypertrophic chondrocytes (MCT cells).

A fundamental question in endochondral development is why the expression of parathyroid hormone-related protein (PTHrP), which inhibits chondrocyte maturation and mineralization, becomes attenuated at the stage of chondrocyte hypertrophy. To address this question, we used clonal, phenotypically stable SV40-immortalized murine endochondral chondrocytes that express a growth-arrested hypertrophic phenotype in culture (MCT cells). Addition of individual cytokines to the medium of MCT cells revealed that bone morphogenetic protein (BMP)-6, which commits chondrocytes to hypertrophy, markedly inhibited PTHrP production.

Analysis of heat shock proteins and cytokines expressed during early stages of osteoarthritis in a mouse model.

Objective: Osteoarthritis (OA) is a debilitating disease of the joints. The joints of affected individuals are characterized by a progressive degeneration of articular cartilage leading to inflammation and pain. The expression of heat shock proteins (HSPs) is a ubiquitous self-protective mechanism of all cells under stress, furthermore, the synovium of osteoarthritic individuals contains high levels of cytokines.

Chondrogenic phenotype of perichondrium-derived chondroprogenitor cells is influenced by transforming growth factor-beta 1.

Our laboratory has developed a method for the repair of osteochondral defects by implanting cultured perichondrial cells attached to a biodegradable polylactic acid scaffold. The success of this approach depends in part on the proliferative characteristics and the phenotype of the implanted cells. Transforming growth factor-beta 1 has been reported to influence these parameters in several mesenchymal-derived tissues in vitro and in vivo.

A complex that contains proteins binding to the PSE and TATA sites in a human U6 small nuclear RNA promoter.

The proximal promoter of a human U6 small nuclear RNA (snRNA)-encoding gene contains two separate elements, the proximal sequence element (PSE) and the TATA box. We investigated the interaction of the PSE- and TATA-binding proteins (PBP and TBP) with normal and mutant U6 proximal promoters using an electrophoretic mobility shift assay. We detected a complex containing both PBP and TBP bound to the wild-type U6 promoter.

Binding and activation of the promoter for the neural cell adhesion molecule by Pax-8.

The neural cell adhesion molecule (N-CAM), is expressed in definite spatiotemporal patterns during development. To identify factors that may influence place-dependent n-cam gene expression, we have studied the binding and activation of the n-cam promoter by Pax-8, a member of the Pax family of transcription factors. Pax-8 increased n-cam promoter activity 13.

Regulation in vitro of an L-CAM enhancer by homeobox genes HoxD9 and HNF-1.

Previous studies have shown that in vitro expression of the neural cell adhesion molecule (N-CAM) can be regulated by the products of homeobox genes HoxB9, -B8, and -C6. N-CAM is a Ca(2+)-independent immunoglobulin-related CAM that plays an important role in neural development. In the present study, we investigated whether the liver cell adhesion molecule (L-CAM) a member of the Ca(2+)-dependent CAM family (cadherins) is also regulated by homeobox-containing genes.

The transcriptional start site for a human U6 small nuclear RNA gene is dictated by a compound promoter element consisting of the PSE and the TATA box.

Transcription of vertebrate U6 snRNA genes by RNA polymerase III requires two sequence elements in the proximal promoter region: the PSE (proximal sequence element, found in snRNA promoters transcribed by RNA polymerase II) and the TATA element (found in many mRNA promoters). The locations of the PSE and the TATA box are important determinants for transcriptional start site selection in their respective RNA polymerase II promoters. In vertebrate U6 genes the PSE and the TATA elements are located in approximately the same positions as in the polymerase II transcribed genes, but their respective roles in initiation site selection are unknown.