Hyaline articular cartilage, the load-bearing tissue of the joint, has very limited repair and regeneration capacities. The lack of efficient treatment modalities for large chondral defects has motivated attempts to engineer cartilage constructs in vitro by combining cells, scaffold materials and environmental factors, including growth factors, signaling molecules, and physical influences. Despite promising experimental approaches, however, none of the current cartilage repair strategies has generated long lasting hyaline cartilage replacement tissue that meets the functional demands placed upon this tissue in vivo. The reasons for this are diverse and can ultimately result in matrix degradation, differentiation or integration insufficiencies, or loss of the transplanted cells and tissues. This article aims to systematically review the different causes that lead to these impairments, including the lack of appropriate differentiation factors, hypertrophy, senescence, apoptosis, necrosis, inflammation, and mechanical stress. The current conceptual basis of the major biological obstacles for persistent cell-based regeneration of articular cartilage is discussed, as well as future trends to overcome these limitations. IntroductionStructure and function of articular cartilage Articular cartilage is a highly specialized tissue that protects the bones of diarthrodial joints from forces associated with load bearing and impact, and allows nearly frictionless motion between the articulating surfaces [1,2]. The extracellular matrix (ECM) of articular cartilage is distinct from that of other connective tissues, consisting of an intricate network containing predominantly fibrillar collagens and proteoglycans. The collagens, types II, IX and XI, form a fibrous framework that gives the tissue its shape, strength and tensile stiffness [3]. Collagen type VI is found pericellularly around chondrocytes [4], and collagen type X is found in calcifying cartilage [5]. Although collagen type I is the most prevalent collagen throughout the body, the primary constituent of the articular cartilage matrix is type II, comprising 80% to 90% of the collagen content [3]. The proteoglycans in articular cartilage in their most abundant form exist as large hydrophilic aggregates, which contain the fluid component and control its movement. The level of compaction of the proteoglycans within the collagen lattice will determine their level of hydration and, in turn, the stiffness of the articular cartilage. The synthesis, incorporation and degradation of ECM proteins are orchestrated by chondrocytes that populate the matrix at low density [3]. Because articular cartilage is avascular, nutrients for the chondrocytes are supplied from the capillaries of the synovium and must diffuse into the synovial fluid and then into the cartilage matrix. Coordinated synthesis and proteolytic breakdown of certain ECM components by chondrocytes enables certain components of the cartilage matrix to undergo turnover and maintenance [3]. Factors that impair cho...

Signal peptides (SP) are key determinants for targeting glycoproteins to the secretory pathway. Here we describe the involvement in particle maturation as an additional function of a viral glycoprotein SP. The SP of foamy virus (FV) envelope glycoprotein is predicted to be unusually long. Using an SP-specific antiserum, we demonstrate that its proteolytic removal occurs posttranslationally by a cellular protease and that the major N-terminal cleavage product, gp18, is found in purified viral particles. Analysis of mutants in proposed signal peptidase cleavage positions and N-glycosylation sites revealed an SP about 148 amino acids (aa) in length. FV particle release from infected cells requires the presence of cognate envelope protein and cleavage of its SP sequence. An N-terminal 15-aa SP domain with two conserved tryptophan residues was found to be essential for the egress of FV particles. While the SP N terminus was found to mediate the specificity of FV Env to interact with FV capsids, it was dispensable for Env targeting to the secretory pathway and FV envelopemediated infectivity of murine leukemia virus pseudotypes.Signal peptides (SP) are key determinants for targeting and membrane insertion of secretory and membrane proteins (reviewed in reference 25). They can be removed co-or posttranslationally by the cellular membrane-bound signal peptidase or may, if not cleaved, serve as membrane anchors for proteins with distinct membrane orientations. In general, SP are composed of three domains, of which a central 6-to 15-amino-acid (aa)-long hydrophobic domain (h-domain) is the most essential. An N-terminal polar domain (n-domain) usually of net positive charge shows high variability in overall length, ranging from 15 to more than 50 aa. The composition and structure of the n-domain influences protein orientation in the membrane. The polar C-terminal domain (c-domain) often contains helixbreaking as well as small uncharged residues in positions -3 and -1 which determine the site of SP cleavage. In most cases, SP cleavage is thought to occur cotranslationally; however, for some proteins, e.g., the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein gp160, SP cleavage occurs inefficiently and very late after translocation (21). A basic amino acid stretch in the n-domain of gp160 is responsible for this phenomenon and believed to influence folding and exit of HIV-1 Env from the endoplasmic reticulum (ER) (21). Recent studies revealed that SP bear specific information accounting for distinct functions in targeting and membrane insertion or even for defined metabolic pathways after their cleavage from the parent protein (reviewed in reference 25). The HIV-1 SP Env , for example, is further processed by the signal peptidase, leading to the release of an SP fragment into the cytosol, where it binds to calmodulin (26). The function of this process in viral replication is not known.Foamy viruses (FV), as studied with the prototype member human foamy virus (HFV), follow a replication cycle which is charact...

Foamy virus (FV) vectors that have minimal cis-acting sequences and are devoid of residual viral gene expression were constructed and analyzed by using a packaging system based on transient cotransfection of vector and different packaging plasmids. Previous studies indicated (i) that FV gag gene expression requires the presence of the R region of the long terminal repeat and (ii) that RNA from packaging constructs is efficiently incorporated into vector particles. Mutants with changes in major 5 splice donor (SD) site located in the R region identified this sequence element as responsible for regulating gag gene expression by an unidentified mechanism. Replacement of the FV 5 SD with heterologous splice sites enabled expression of the gag and pol genes. The incorporation of nonvector RNA into vector particles could be reduced to barely detectable levels with constructs in which the human immunodeficiency virus 5 SD or an unrelated intron sequence was substituted for the FV 5 untranslated region and in which gag expression and pol expression were separated on two different plasmids. By this strategy, efficient vector transfer was achieved with constructs that have minimal genetic overlap.