RESUMO
CONTEXT: During osteoarthritis (OA), chondrocytes undergo de-differentiation, resulting in the acquisition of a fibroblast-like morphology, decreased expression of collagen type II (colII) and aggrecan, and increased expression of collagen type I (colI), metalloproteinase 13 (MMP13) and nitric oxide synthase (eNOS). Notch signaling plays a crucial role during embryogenesis. Several studies showed that Notch is expressed in adulthood. OBJECTIVE: The aim of our study was to confirm the involvement of Notch signaling in human OA at in vitro and ex vivo levels. MATERIALS AND METHODS: Normal human articular chondrocytes were cultured during four passages either treated or not with a Notch inhibitor: DAPT. Human OA cartilage was cultured with DAPT for five days. Chondrocytes secreted markers and some Notch pathway components were analyzed using Western blotting and qPCR. RESULTS: Passaging chondrocytes induced a decrease in the cartilage markers: colII and aggrecan. DAPT-treated chondrocytes and OA cartilage showed a significant increase in healthy cartilage markers. De-differentiation markers, colI, MMP13 and eNOS, were significantly reduced in DAPT-treated chondrocytes and OA cartilage. Notch1 expression was proportional to colI, MMP13 and eNOS expression and inversely proportional to colII and aggrecan expression in nontreated cultured chondrocytes. Notch ligand: Jagged1 increased in chondrocytes culture. DAPT treatment resulted in reduced Jagged1 expression. Notch target gene HES1 increased during chondrocyte culture and was reduced when treated with DAPT. CONCLUSION: Targeting Notch signaling during OA might lead to the restitution of the typical chondrocyte phenotype and even to chondrocyte redifferentiation during the pathology.
Assuntos
Desdiferenciação Celular/genética , Osteoartrite/genética , Receptores Notch/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/biossíntese , Cartilagem Articular/crescimento & desenvolvimento , Cartilagem Articular/metabolismo , Técnicas de Cultura de Células , Diferenciação Celular/genética , Condrócitos/metabolismo , Colágeno Tipo II/metabolismo , Dipeptídeos/administração & dosagem , Proteínas de Homeodomínio/biossíntese , Humanos , Osteoartrite/metabolismo , Osteoartrite/patologia , Transporte Proteico/genética , Receptores Notch/antagonistas & inibidores , Receptores Notch/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição HES-1RESUMO
To analyze the effect of intramedullary reaming on fracture healing, we investigated whether or not cortical reamings contain viable bone cells. There are several tissue components contained in medullary reamings including blood, bone marrow and cortical bone. This study is focused on the cortical reamings, which are produced during reaming of the medullary cavity. They may stimulate fracture healing but it is still unclear if they contain vital bone cells. We tested the hypothesis that these cortical reamings are a source of viable bone cells and compared cell cultures with cultivated cells from iliac crest biopsies. Responses of protein content and ALP activity to vitD stimulation in the cells were considered as properties of viability. Ten in tact living sheep femora were fully reamed and the cortical reamings were cultivated in a standard manner and compared with cultivated cells from ipsi-lateral iliac crest biopsies from the same animals. Cells started to grow from the reamings as well as the iliac crest within 2-5 days, and covered the entire culture flask within 9-13 days. Protein content and ALP activity in cells from both reamings and iliac crest were significantly responsive to vitD stimulation. Cortical reamings from intramedullary nailing have osteoblastic potential and contain living bone cells similar to bone cells from the iliac crest. These findings may further explain the superior healing of fractures, treated with reamed nailing.