The restricted potential for recovery of growth plate chondrogenesis and longitudinal bone growth following exposure to pro-inflammatory cytokines

Abstract

Childhood chronic inflammatory disease can be associated with transient and permanent growth retardation. This study examined the potential for spontaneous growth recovery following pro-inflammatory cytokine exposure. Murine ATDC5 chondrogenic cells and postnatal metatarsals were exposed to interleukin (IL)-1 , IL-6 and tumour necrosis factor-(TNF ), and their growth and proliferative capacity were determined following recovery. TNF and IL-1 reduced chondrocyte proliferation and aggrecan and collagen types II and X expression at minimum concentrations of 10 ng/ml and 0·1 ng/ml respectively. TNF but not IL-1 exposure led to increased caspase-3 activity and altered cellular morphology, consistent with reduced viability. Cytokine exposure particularly inhibited proteoglycan synthesis. This effect was dose and duration dependent. Compared with the control, IL-1 and TNF led to a 71% and 45% reduction in metatarsal growth after 8 days of exposure respectively (P<0·05). An additive effect of IL-1 combined with TNF was observed (110% decrease; P<0·05). Metatarsals exposed to IL-1 or TNF individually for a 2-day period, and allowed to recover spontaneously in the absence of cytokines for a further 6 days, showed normal growth trajectories. In combination, growth was 59% lower (P<0·01) compared with control metatarsals at the end of the recovery period. Exposure to the combination for 4 days followed by a 4-day recovery period resulted in 87% decrement compared with controls (P<0·05). IL-6 did not alter any parameter studied. IL-1 and TNF exert diverse inhibitory effects on ATDC5 chondrocyte dynamics and metatarsal growth. The extent of recovery following cytokine exposure depends on the duration of exposure, and may be incomplete following longer periods of exposure.

Ceramide inhibition of chondrocyte proliferation and bone growth is IGF-I independent

“…Standardisation of [ 3 H]thymidine uptake to protein content (Phornphutkul et al 2006) confirms that an actual reduction in proliferation was observed following ceramide exposure, rather than solely a reduced uptake due to fewer cells through increased apoptosis. Both TNF-a and IL-1b exposure have also been reported to inhibit proliferation and induce apoptosis in ATDC5 cells (MacRae et al 2006b), and the results of this present study now suggest that ceramide may be acting as a second messenger in both processes. Both IL-1b and TNFa have also been shown to markedly reduce the gene expression of collagen II, collagen X and aggrecan (MacRae et al 2006b), but in our present study ceramide did not alter the gene expression of markers of chondrogenesis and differentiation (sox 9, collagen II and collagen X).…”

“…Both TNF-a and IL-1b exposure have also been reported to inhibit proliferation and induce apoptosis in ATDC5 cells (MacRae et al 2006b), and the results of this present study now suggest that ceramide may be acting as a second messenger in both processes. Both IL-1b and TNFa have also been shown to markedly reduce the gene expression of collagen II, collagen X and aggrecan (MacRae et al 2006b), but in our present study ceramide did not alter the gene expression of markers of chondrogenesis and differentiation (sox 9, collagen II and collagen X). This suggests that ceramide generation does not affect differentiation in growth plate chondrocytes.…”

“…Whilst previous studies have shown that IL-1b and TNFa inhibit growth plate chondrocyte differentiation and induce cell death (Martensson et al 2004, MacRae et al 2006b), the direct effects of ceramide on growth plate chondrocytes have yet to be reported. In this study, the ATDC5 chondrogenic cell line was used to characterise and compare the effects of ceramide on cell proliferation, differentiation and apoptosis.…”

“…This effect has been reported to be independent of the concurrent reduction in growth associated with glucocorticoid therapy (Tynjala et al 2006). TNF-a and IL-1b may directly inhibit growth plate chondrocyte dynamics as well as longitudinal growth in vitro (Martensson et al 2004, MacRae et al 2006b), however, the underlying mechanisms that lead to these effects are unclear.…”

IKKβ in postnatal perichondrium remotely controls endochondral ossification of the growth plate through downregulation of MCP-5

“…Then, we examined the expression of genes encoding cytokines that were reported to affect endochondral ossification. 17,[29][30][31] As shown in Figure 6c, among the cytokines examined, Il-1β and Mcp-5 displayed big differences between Ikkβ Prx1KO and control mice. Although IL-1β is reported to inhibit the differentiation of chondrocytes, 29,32 it was downregulated in the perichondrium of Ikkβ Prx1KO mice and thus fails to explain the phenotype.…”