Signal transduction and metabolism in chondrocytes is modulated by lactoferrin.

OBJECTIVE: Activation of granulocytes causes a considerable rise in the concentration of lactoferrin (Lf) in synovial fluid (SF). We here investigate consequences thereof on signal transduction and the balance between catabolic and anabolic metabolism in chondrocytes. METHODS: Signal transduction was analysed in cultured chondrocytes by immunodetection of mitogen activated protein kinases (MAPK) and analysis of Smad2 translocation to the nucleus. Expression levels of matrix metalloproteinases (MMPs) and of aggrecan were measured by reverse-transcription-PCR. The proteolytic activity of MMPs was ascertained by zymography. Expression of the low-density-lipoprotein-receptor-related-protein-1 (LRP-1), a Lf receptor for signalling, was assayed by immunohistochemistry in cartilage and in cultured chondrocytes by immunoblotting. RESULTS: We found LRP-1 expressed in dedifferentiated chondrocytes in culture and in cartilage tissue preferentially on the articular surface where it can encounter Lf within SF. Lf stimulated proliferation of chondrocytes, comparable to transforming growth factor-beta1 (TGFbeta1) and activated p38 and the extracellular-signal regulated-kinases 1/2 (ERK1/2) within minutes. Surprisingly, Lf induced nuclear Smad2 translocation, a signal pathway ascribed to TGFbeta receptor activation. Lf significantly increased the levels of catabolic indicators such as MMP1, MMP2, MMP3 and MMP13 and inhibited aggrecan synthesis. CONCLUSION: Lf is a robust regulator of chondrocyte metabolism, comparable to TGFbeta1. The catabolic influence together with the proliferative stimulus indicates a function as an early phase cytokine, enhancing MMPs, necessary for degradation of damaged tissue and stimulating proliferation of chondrocytes, necessary for reconstruction.

Assorted effects of TGFbeta and chondroitinsulfate on p38 and ERK1/2 activation levels in human articular chondrocytes stimulated with LPS.

OBJECTIVES: Inadequate cellular response of chondrocytes to stress frequently terminates in osteoarthritis (OA). Adequate response is fundamentally modulated by concerted cytokine signaling events, directing degradation and synthesis of cartilage on articular surfaces where and whenever necessary. Transforming growth factor (TGF)beta is a prominent mediator in cartilage anabolism, although particular catabolic activities are occasionally reported. Clearly, before the TGFbeta signal gets through to the gene regulatory machinery, cross talk with modulators occurs. METHOD: We tested the hypothesis whether chondroitinsulfate (CS) modulates cell signaling. TGFbeta and/or soluble CS was added to human articular chondrocytes (HACs) and activation of p38 and extracellular signal related kinase (ERK)1/2 was determined by immunoblot analysis. Expression levels of mRNA of matrix metalloproteinase (MMP)-2, -3 and -13 were determined by real-time polymerase chain reaction (PCR). RESULTS: No significant effects were observed unless cells were stimulated with lipopolysaccharide (LPS), invigorating catabolic metabolism in chondrocytes. LPS effects, however, were profoundly modulated by TGFbeta, CS and both applied in combination. Most prominent, the silencing of p38 stress signal by CS was superimposable to that of TGFbeta. Phospho-ERK1/2 levels were raised by TGFbeta three-fold over LPS induced levels. In contrast, CS treatment, alone or combined with TGFbeta, reduced phosphorylation significantly below LPS induced levels. Finally, suppression of LPS induced MMP-13 mRNA levels resulted with CS. CONCLUSION: Soluble CS modulates signaling events in chondrocytes concurrent with MMP-13 down regulation. The effects observed suggest a feedback signaling mechanism cross talking with TGFbeta-signal pathways and may serve an explanation, on the cellular level, for the beneficial effects found in clinical studies with pharmacologic application of CS.