Role of Cx43-Mediated NFкB Signaling Pathway in Ossification of Posterior Longitudinal Ligament: An In Vivo and In Vitro Study.
METHODS
In vivo and in vitro experiments.


OBJECTIVE
To illustrate the further molecular mechanism of Cx43-mediated osteoblastic differentiation of ligament cells.


BACKGROUND
Ossification of the posterior longitudinal ligament (OPLL) is one of the main causes of myelopathy in Asians, but its etiology has not been clarified. We have previously found the mechanical stress can upregulate Cx43 expression in ligament cells, which transduces mechanical signal to promote osteoblastic differentiation.


METHODS
The posterior longitudinal ligaments were collected intraoperatively. Ligament fibroblasts were isolated and cultured, and an in vitro mechanical loading model was established. In vivo and in vitro expression levels of Cx43 protein were compared between OPLL and non-OPLL patients. The activation of nuclear factor (NF)-κB (p65) signal and related inflammatory responses were detected in ligament cells under mechanical loading. The mechanical stress-induced inflammatory response and osteoblastic differentiation of OPLL cells were investigated after the treatment with Cx43 siRNA or NFкB (p65) inhibitor.


RESULTS
We first confirmed higher Cx43 levels in both in vivo ligament tissue from OPLL patients and in vitro cultured OPLL cells. We also found NFκB (p65) signal and related inflammatory response were activated by mechanical stimulation. The activation of NFκB (p65) signal was dependent upon Cx43, as its knockdown reduced signal. Moreover, treatment with Cx43 siRNA or NFкB (p65) inhibitor significantly decreased the mechanical-induced inflammation response, but partly attenuated mechanical-stimulated osteoblastic differentiation of OPLL cells.


CONCLUSIONS
Cx43-mediated NFкB (p65) signal played an important role in mechanical stress-induced OPLL by transduction of mechanical signal, while giving rise to the activation of inflammatory response in ligament fibroblastsLevel of Evidence: N/A.
Department of Spine Surgery, Changzheng Hospital, Second Military Medical University, Shanghai, China.
Dechun
D
Chen
Yu
Y
Chen
Tiefeng
T
Li
Lei
L
Shi
Mengxiao
M
Pan
Deyu
D
Chen
D000328:Adult; D000469:Alkaline Phosphatase / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D018031:Connexin 43 / Q000235:genetics / Q000378:metabolism*; D005260:Female; D005347:Fibroblasts / Q000502:physiology*; D055785:Gene Knockdown Techniques; D006801:Humans; D007249:Inflammation / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017887:Ossification of Posterior Longitudinal Ligament / Q000209:etiology*; D034741:RNA, Small Interfering / Q000494:pharmacology; D015398:Signal Transduction; D013314:Stress, Mechanical; D013312:Stress, Physiological; D015854:Up-Regulation
D018031:Connexin 43; C500216:GJA1 protein, human; D016328:NF-kappa B; D034741:RNA, Small Interfering; D000469:Alkaline Phosphatase