Expression of Cx43 in the ligament fibroblasts derived from patients presenting ossification of posterior longitudinal ligament and its effect during ossification

English Keywords:Ossification of the posterior longitudinal ligament  Cx43  Osteocalcin  Alkaline phosphatase  Collagen Ⅰ

English Abstract:

  【Abstract】 Objectives: To investigate the expression of connexin43(Cx43) in the ligament fibroblasts derived from patients presenting ossification of posterior longitudinal ligament(OPLL) and its effect during ossification. Methods: Fifteen patients presenting with OPLL and 15 non-OPLL patients underwent anterior decompression between January 2013 and December 2013. Specimens of the posterior longitudinal ligament were collected intraoperatively. Tissue fragment cell culture was performed. The mouse anti-vimentin antibody was used to identify the cultured cells via immunocytochemistry and immunofluorescence(ICC/IF). The mRNA expressions of osteoblast-specific genes of osteocalcin(OCN), alkaline phosphatase(ALP) and type Ⅰ collagen(COL Ⅰ) were detected in OPLL and non-OPLL cells by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). The protein expression of Cx43 was detected via western blotting. And after 72h, when RNA interference against Cx43 was performed in OPLL cells, the expressions of indexes mentioned above were compared again between transfection and non-transfection group. Results: The mRNA expression of OCN, ALP and COL Ⅰ was 1.36±0.21, 0.53±0.18 and 1.64±0.37. It was significantly higher compared with non-OPLL group, with expression as 0.78±0.21, 0.29±0.13 and 1.01±0.26. The protein expression of Cx43(1.00±0.30) in OPLL group was also obvious higher than the non-OPLL group(0.48±0.18). Furthermore, after 72h of transfection targeting Cx43, result of western blot showed a down-regulation of Cx43 protein expression by almost 69%(P<0.01). The mRNA expression of OCN, ALP and COL Ⅰ was 1.54±0.33, 1.32±0.36, 1.86±0.44 in non-transfection group and 0.91±0.32, 0.47±0.21, 0.88±0.29 in non-transfection group. The difference was significant. Conclusions: There is a higher expression of Cx43 in ligament fibroblasts derived from OPLL, which indicates that Cx43 plays an important role during the ossification of the ligament fibroblasts.