Centrifugal Force Induces Human Ligamentum Flavum Fibroblasts Inflammation Through Activation of JNK and p38 Pathways

Abstract

Inflammation has been proposed to be an important causative factor in ligamentum flavum hypertrophy. However, the mechanisms of mechanical load on inflammation of ligamentum flavum remain unclear. In this study, we used an in vitro model of human ligamentum flavum fibroblasts subjected to centrifugal force to elucidate the effects of mechanical load on cultured human ligamentum flavum fibroblasts; we further studied its molecular and biochemical mechanisms. Human ligamentum flavum fibroblasts were obtained from six patients undergoing lumbar spine surgery. Monolayer cultures of human ligamentum flavum fibroblasts were subjected to different magnitudes of centrifugal forces. Cell viability, cell death, biochemical response, and molecular response to centrifugal forces were analyzed. It was found that centrifugal stress significantly suppressed cell viability without inducing cell death. Centrifugal force at 67.1 g/cm(2) for 60 min significantly increases the production of prostaglandin E2 and nitric oxide as well as gene expression of proinflammatory cytokines, including interleukin (IL)-1α, IL-1β and IL-6, showed that centrifugal force-dependent induction of cyclooxygense-2 and inducible NO synthase required JNK and p38 mitogen-activated protein kinase, but not ERK 1/2 activities. This study suggested that centrifugal force does induce inflammatory responses in human ligamentum flavum fibroblasts. The activation of both JNK and p38 mitogen-activated protein kinase mechanotransduction cascades is a crucial intracellular mechanism that mediates cyclooxygense-2/prostaglandin E2 and inducible NO synthase/nitric oxide production.

From sentinel cells to inflammatory culprits: cancer‐associated fibroblasts in tumour‐related inflammation

“…While there are few cancer-related studies that directly connect mechano-sensing by fibroblasts with pro-inflammatory signalling, several studies performed in other fields have reported that activation of fibroblasts by biomechanical forces induces pro-inflammatory signalling: For example, periodontal fibroblasts respond to compression forces by increased TNF-α production at the compression site, resulting in the activation of CD4 + T cells and facilitating bone resorption [47]. Furthermore, application of mechanical forces induces fibroblasts to express several proinflammatory cytokines including IL-1α, IL-1β, and IL-6 [48]. Kook et al reported that application of tensile forces on fibroblasts stimulated mRNA expression of collagen I and MMP-1, typical of activated fibroblasts, in an ERK-NF-κB signalling-dependent manner [49], thus linking mechanical forces with inflammatory signalling.…”

Tissue transglutaminase is involved in mechanical load–induced osteogenic differentiation of human ligamentum flavum cells

“…Mechanical stress has also been evidenced to play important role in the progression of ligamentum flavum degeneration (11,12). In addition to that, our previous study demonstrated that mechanical load increased the production of PGE2 and NO as well as pro-inflammatory cytokines, IL-1α, IL-1β, and IL-6, gene expression in human ligamentum flavum cells (4). Taken together, it prompted us to further explore the critical mechanism underlying mechanical loading and the calcification of ligamentum flavum.…”

“…In this study, ligamentum flavum cells were exposed to mechanical load by applying a well-established in vitro compression model of centrifugation of cultured cells. Since there is still no documented data to show the magnitude of loading on ligamentum flavum that mediates the degenerative process, the magnitude of loading applied in this study is experimentally based on our previous work (4). Our previous study demonstrated a distinguished cascade of inflammatory response when ligamentum flavum cells were subjected to compressive stress (4).…”

“…Since there is still no documented data to show the magnitude of loading on ligamentum flavum that mediates the degenerative process, the magnitude of loading applied in this study is experimentally based on our previous work (4). Our previous study demonstrated a distinguished cascade of inflammatory response when ligamentum flavum cells were subjected to compressive stress (4). In this study, mechanical load only, BMP-2 only, and mechanical load with concurrent BMP-2 were all shown to significantly increase ALP activity on day 14 followed by an enhanced mineralization of ligamentum flavum cells.…”

“…As centrifugal force represents a constant hydrostatic pressure that mimics compression force, while static loading in other tissues has been shown to be damaging and may lead to ultimate tendon failure (3); we employed it to simulate a constant biomechanical force faced by ligamentum flavum within this experimental context. In our previous work, we demonstrated that the ligamentum flavum responds to such kind of mechanical load through activation of JNK and p38 pathways (4). In this study, the parameters chosen (applied) were based on prior experience with some modulation for better enhancement of the result.…”