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Although the low power of our study prevents definitive conclusions, ACI and mosaicplasty are cartilage repair techniques that are clinically equivalent and similar in performance. The high percentage of spontaneous improvement ((1/3) of the patients) observed after simple debridement calls into question the need for prompt surgical treatment of patients with lesions similar to those included in this clinical trial. Moreover, this finding warrants further investigation, ideally through randomized clinical trials in which patients subjected to debridement alone are compared with patients undergoing reconstructive surgery.

The SLNB procedure does not appear inferior to conventional ALND for the subset of patients here considered.

The SF conditions have proven valuable to prime cell proliferation and to sustain proper commitment in chondrocytes from aged patients. This culturing approach may represent a therapeutic chance extendable to a range of patients normally excluded from clinical protocols based on autologous chondrocyte implantation (ACI).

Abstract. This report deals with the quantitation of both mRNA and transcription activity of type I collagen gene and of three cartilage-specific collagens (types II, IX, and X) during in vitro differentiation of chick chondrocytes. Differentiation was obtained by transferal to suspension culture of dedifferentiated cells passaged for 3 wk as adherent cells. The type I collagen mRNA, highly represented in the dedifferentiated cells, rapidly decreased during chondrocyte differentiation. On the contrary, types II and IX collagen mRNAs sharply increased within the first week of suspension culture, peaked in the second week, and thereafter began to decrease. This decrease was particularly significant for type IX collagen mRNA. The level of type X collagen mRNA progressively increased during the course of the culture, reached its maximal value after 3-4 wk, and decreased only at a later stage of cell differentiation.As determined by in vitro run-off transcription assays, all these changes in collagen mRNA levels could be attributed to parallel modifications in the relative rate of transcription of the corresponding collagen genes.We suggest that chicken chondrocyte differentiation proceeds through at least two different steps: (a) first, transition from a stage characterized by a high level of type I collagen mRNA to a stage characterized by predominance of types II and IX collagen mRNAs; (b) later, transition to a stage characterized by the highest level of type X collagen mRNA.RGANOGENESlS of long bones involves cellular differentiation and continuous synthesis and remodeling of the extracellular matrix. During the first stages of chick development, mesenchymal cells in the limb buds differentiate, possibly through distinct regulatory steps (22), and condense to form a blastema from which cartilaginous bones develop. This process is characterized by profound changes in the cell morphology accompanied by the synthesis of novel extracellular macromolecules, among which are cartilage-specific proteoglycans and collagens (7). Changes in the levels of types I and II collagens and of their mRNAs during chondrogenesis in vivo and in vitro were detected with the use of specific antibodies and cDNA probes (27,10, 11). Cartilaginous bones grow because of cell proliferation and deposition of new extracellular matrix. Proliferating chondrocytes progressively mature into hypertrophic nondividing cells. Hypertrophy of chondrocytes, first detectable in the diaphysis and after in the epiphyseal regions, is accompanied by the onset of type X collagen synthesis (2, 20). Hypertrophic cartilage undergoes calcification, is invaded by blood vessels and osteogenic cells, and is replaced by bone tissue (18). To investigate at the molecular level the differentiation of hypertrophic cartilage chondrocytes we have used an in vitro system starting with tibial chondrocytes of early stage chick embryos (3). In this system, chondrocytes obtained by enzymatic dissociation of stage 28-30 (9) embryo tibiae, passaged on tissue culture dishes, assum...

Cytotoxic T lymphocyte antigen-4 (CTLA-4) is the major negative regulator of T-cell responses, although growing evidence supports its wider role as an immune attenuator that may also act in other cell lineages. Here, we have analyzed the expression of CTLA-4 in human monocytes and monocyte-derived dendritic cells (DCs), and the effect of its engagement on cytokine production and T-cell stimulatory activity by mature DCs. CTLA-4 was highly expressed on freshly isolated monocytes, then down-modulated upon differentiation toward immature DCs (iDCs) and it was markedly upregulated on mature DCs obtained with different stimulations (lipopolysaccharides [LPS], Poly:IC, cytokines). In line with the functional role of CTLA-4 in T cells, treatment of mDCs with an agonistic anti-CTLA-4 mAb significantly enhanced secretion of regulatory interleukin (IL)-10 but reduced secretion of IL-8/IL-12 pro-inflammatory cytokines, as well as autologous CD4+ T-cell proliferation in response to stimulation with recall antigen purified protein derivative (PPD) loaded-DCs. Neutralization of IL-10 with an anti-IL-10 antibody during the mDCs-CD4+ T-cell co-culture partially restored the ability of anti-CTLA-4-treated mDCs to stimulate T-cell proliferation in response to PPD. Taken together, our data provide the first evidence that CTLA-4 receptor is expressed by human monocyte-derived mDCs upon their full activation and that it exerts immune modulatory effects.

Abstract. Chondrocytes from chicken embryo tibia can be maintained in culture as adherent cells in Coon's modified Ham's F-12 medium supplemented with 10% FCS. In this condition, they dedifferentiate, losing type II collagen expression in favor of type I collagen synthesis. Their differentiation to hypertrophy can be obtained by transferring them to suspension culture. Differentiation is evidenced by the shift from type I to type II and type IX collagen synthesis and the following predominant expression of type X collagen, all markers of specific stages of the differentiation process. To identify the factors required for differentiation, we developed a serum-free culture system where only the addition of triiodothyronine (T3; 10 -u M), insulin (60 ng/ml), and dexamethasone (10 -9 M) to the F-12 medium was sufficient to obtain hypertrophic chondrocytes. In this hormonal context, chondrocytes display the same changes in the pattern of protein synthesis as described above. For proper and complete cell maturation, T3 and insulin concentrations cannot be modified. Insulin cannot be substituted by insulinlike growth factor-I, but dexamethasone concentration can be decreased to 10 -12 M without chondrogenesis being impaired. In the latter case, the expression of type X collagen and its mRNA are inversely proportional to dexamethasone concentration. When ascorbic acid is added to the hormone-supplemented medium, differentiating chondrocytes organize their matrix leading to a cartilage-like structure with hypertrophic chondrocytes embedded in lacunae. However, this structure does not present detectable calcification, at variance with control cultures maintained in FCS. Accordingly, in the presence of the hormone mixture, the differentiating chondrocytes have low levels of alkaline phosphatase activity. This report indicates that T3 and insulin are primary factors involved in the onset and progression of chondrogenesis, while dexamethasone supports cell viability and modulates some differentiated functions.