Involvement of the notch pathway in the regulation of matrix metalloproteinase 13 and the dedifferentiation of articular chondrocytes in murine cartilage

Abstract

Objective. To demonstrate the activation of the Notch signaling pathway during changes in the phenotype of chondrocytes in vitro, and to assess the influence of Notch on the production of chondrocyte markers.Methods. Serial monolayer primary cultures of murine articular chondrocytes (MACs), as a model of chondrocyte dedifferentiation, were prepared. MACs were cultured with or without a Notch inhibitor and transfected with different Notch-expressing vectors. The Notch pathway and chondrocyte marker profiles were assessed by quantitative reverse transcriptionpolymerase chain reaction, immunoblotting, and immunocytochemistry.Results. Successive passages of MACs resulted in a loss of type II collagen and aggrecan (chondrocyte differentiation markers), an increase in type I collagen (dedifferentiation marker), an increase in Notch ligands, and augmented target gene activity. The Notch inhibitor decreased the type II collagen protein content but had no effect on Col2a1 messenger RNA, while transfection with the constitutive active forms of the Notch1 receptor led to a decrease in type II collagen in transfected cells. In assays to investigate the mechanism of type II collagen breakdown, matrix metalloproteinase 13 (MMP-13) synthesis was regulated in a Notchdependent manner, whereas MMP-2 synthesis was unchanged.Conclusion. The Notch signaling pathway is associated with decreased type II collagen production during the dedifferentiation of MACs in vitro. This may be correlated with the increase in MMP-13 production linked to activation of Notch.

Human Schlafen 5 (SLFN5) Is a Regulator of Motility and Invasiveness of Renal Cell Carcinoma Cells

“…Notably, one of the studies showed a cooperation of Notch and transforming growth factor ␤ (TGF-␤) signaling pathways in the process (39). As both MMP-1 and MMP-13 expression levels are known to be regulated by TGF-␤-induced signals (40) and Notch signaling has been shown to affect MMP-13 expression (41,42), it is possible that SLFN5 controls MMP-1 and MMP-13 expression, as well as cell motility and invasion through its effects on NOTCH/TGF-␤ signaling. Expression of MMPs can also be modulated by additional pathways (43).…”

Inflammatory Cytokines Induce NOTCH Signaling in Nucleus Pulposus Cells

“…These results are consistent with previous observations that showed increased NOTCH signaling in stem cells and other cell types in response to cytokines (24 -27). In osteoarthritic cartilage, a condition linked to production of proinflammatory cytokines, the role of NOTCH1, JAGGED1, and HES5 has been associated to the expression of a reparative phenotype (28), although increase in NOTCH signaling promotes dedifferentiated phenotype of articular chondrocytes (29). Likewise, in the NP, the activity of this signaling system, in the presence of TNF-␣ and IL-1␤ possibly indicates an attempt by the cells to promote tissue repair.…”

Proinflammatory Actions of Visfatin/Nicotinamide Phosphoribosyltransferase (Nampt) Involve Regulation of Insulin Signaling Pathway and Nampt Enzymatic Activity

“…Reverse transcription was performed on 1 g of total RNA, using the Omniscript RT kit (Qiagen). Messenger RNA (mRNA) for IR was quantified using the LightCycler LC480 (Roche Diagnostics) as described (25). Specific primers for complementary DNA (cDNA) were chosen with the LightCycler Probe Design 2 program.…”