The Effect Of Calcitonin On Inflammatory In Chondrocyte And P38MAPK Signaling

Objectives In order to investigate the effect of calcitonin(CT) on p38 kinase(p38) and its phosphorylation protein, c-myc, matrix metalloproteinase-13(MMP-13) and collagenⅡ(ColⅡ) in IL-1β-induced chondrocytes, we explored whether CT restrained IL-1β-induced chondrocyte damage through inhibiting p38 MAPK signaling pathway.Methods Sprague-Dawley rats’ chondrocytes were isolated and cultured. The chondrocytes were identified by Toluidin Blue staining. The second-generation chondrocytes were divided into seven groups, A is normal control group: DMEM/F12 complete medium; B is IL-1β-induced group: DMEM/F12 complete medium with 10ng/ml IL-1β of complete medium; C is CT control group: DMEM/F12 complete medium with 50ng/ml CT; D is CT treated group: DMEM/F12 complete medium with 10ng/ml IL-1β for 15 min, then plus 50ng/ml CT further. D is SB203580 treated group: DMEM/F12 complete medium with 10ng/ml IL-1β for 15 min, then plus 10ng/ml SB203580 further. F is CT and SB203580 control group: DMEM/F12 complete medium with 50ng/ml CT and 10ng/ml SB203580; G is CT and SB203580 treated group: DMEM/F12 complete medium with 10ng/ml IL-1β for 15 min,then plus 50 ng/ml CT ande 10ng/ml SB203580 further.All groups cultured into 24 hours. Then we use Western Blot to detect the expression of MMP-13, ColⅡ, p38, P-p38 and c-myc, immunocytochemistry was used to observe the expression of MMP-13, ColⅡ, p38 and P-p38.Results 1 The stain of toluidine blue and immunocytochemistry is confirmed the cell which we cultured are chondrocyte.It would be fine to use in the following experiment. 2 The result of MTT assay demonstrate the concentration of CT between 50ng/ml to 500 ng/ml have the same effect in proliferation of chondrocyte, so we choose 50 ng/ml CT into following experiment. 3 The result of Western Blot: 1) The expression of p38 have no significant difference in each group(P>0.05). The expression of P-p38 in B group is higher than other groups(P<0.05). The expression of P-p38 in D, E and G group is higher than A group but lower than B group(P<0.05). The D, E, G group have no significant difference(P>0.05). 2) The expression of MMP13 and c-myc in B group is higher than other groups(P<0.05). The expression of MMP13 and c-myc in D and E group is higher than A group but lower than B group(P<0.05). The expression of MMP13 and c-myc in G group ishigher than A group but lower than B, D and E group(P<0.05). The D and E group have no significant difference(P>0.05). 3) The expression of ColⅡ in B group is lower than other groups(P<0.05). The expression of ColⅡ in D, E and G group is lower than A group but higher than B group(P<0.05). The D, E and G group have no significant difference(P>0.05). 4) There is no significant difference in A, C and F group(P>0.05); 4 The stain of immunocytochemistry was used to detection the p38, P-p38, c-myc, MMP-13 and ColⅡ have the same result with Western Blot.Conclusions Calcitonin can improvement IL-1β-induced injury in rat chondrocytes, may be one of the mechanisms is to inhibition the activating of the p38 MAPK signaling pathway to reduce the expression of MMP-13, thereby reducing the degradation of ColⅡ.