Aim To evaluate the effect of TEGDMA on human gingival fibroblasts (HGFs) in vitro co-cultured with Streptococcus mitis, focusing on the signalling pathways underlying cell tissue remodelling and inflammatory response processes. Methodology b1 integrin expression was evaluated by means of imaging flow cytometry. The Western blot technique was used to investigate the expression of protein kinase C (PKC), extracellular signal- regulated kinase (ERK), matrix metalloproteinase 9 (MMP9) and 3 (MMP3). RT-PCR was performed to quantify nuclear factor-kb subunits (Nf-kb1, ReLa), IkB kinase b (IkBkB), cyclooxygenase II (COX-2) and tumour necrosis factor-a (TNF-a) mRNA levels. Statistical analysis was performed using the analysis of variance (ANOVA). Results When HGFs are co-cultured with S. mitis, b1 integrin intensity, phosphorylated PKC (p-PKC), activated ERK (p-ERK), IkBkB mRNA level and MMP9 expression increased (for all molecules P < 0.05 HGFs versus HGFs co-cultured with S. mitis). A higher level of MMP3 in HGFs treated with TEGDMA was recorded (P < 0.05 HGFs versus HGFs exposed to TEGDMA). COX-2 inflammatory factor mRNA level appeared higher in HGFs exposed to 1 mmol L 1 TEGDMA (P < 0.01 HGFs versus HGFs exposed to TEGDMA), whereas TNF-a gene expression was higher in HGFs co-cultured with S. mitis (P < 0.05 HGFs versus HGFs co-cultured with S. mitis). Conclusions b1 integrin triggered the signalling pathway, transduced by p-PKCa and involving ERK 1 and 2 and MMPs. This pathway resulted in an unbalanced equilibrium in tissue remodelling process, along with inflammatory response when HGFs are exposed to bacteria or biomaterial alone. On the contrary, the TEGDMA/S. mitis combination restored the balance between extracellular matrix deposition and degradation and prevented an inflammatory response. Keywords: co-culture, HGFs, inflammation, matrix metalloproteinases, tissue remodelling, b1 integrin
A dual role for β1 integrin in an in vitro Streptococcus mitis/human gingival fibroblasts co-culture model in response to TEGDMA
DI NISIO, Chiara;DE COLLI, MARIANNA;DI GIACOMO, Viviana;DI VALERIO, Valentina;MARCONI, GUYA DILETTA;GALLORINI, MARIALUCIA;DI GIULIO, MARA;CATALDI, Amelia;ZARA, SUSI
2015-01-01
Abstract
Aim To evaluate the effect of TEGDMA on human gingival fibroblasts (HGFs) in vitro co-cultured with Streptococcus mitis, focusing on the signalling pathways underlying cell tissue remodelling and inflammatory response processes. Methodology b1 integrin expression was evaluated by means of imaging flow cytometry. The Western blot technique was used to investigate the expression of protein kinase C (PKC), extracellular signal- regulated kinase (ERK), matrix metalloproteinase 9 (MMP9) and 3 (MMP3). RT-PCR was performed to quantify nuclear factor-kb subunits (Nf-kb1, ReLa), IkB kinase b (IkBkB), cyclooxygenase II (COX-2) and tumour necrosis factor-a (TNF-a) mRNA levels. Statistical analysis was performed using the analysis of variance (ANOVA). Results When HGFs are co-cultured with S. mitis, b1 integrin intensity, phosphorylated PKC (p-PKC), activated ERK (p-ERK), IkBkB mRNA level and MMP9 expression increased (for all molecules P < 0.05 HGFs versus HGFs co-cultured with S. mitis). A higher level of MMP3 in HGFs treated with TEGDMA was recorded (P < 0.05 HGFs versus HGFs exposed to TEGDMA). COX-2 inflammatory factor mRNA level appeared higher in HGFs exposed to 1 mmol L 1 TEGDMA (P < 0.01 HGFs versus HGFs exposed to TEGDMA), whereas TNF-a gene expression was higher in HGFs co-cultured with S. mitis (P < 0.05 HGFs versus HGFs co-cultured with S. mitis). Conclusions b1 integrin triggered the signalling pathway, transduced by p-PKCa and involving ERK 1 and 2 and MMPs. This pathway resulted in an unbalanced equilibrium in tissue remodelling process, along with inflammatory response when HGFs are exposed to bacteria or biomaterial alone. On the contrary, the TEGDMA/S. mitis combination restored the balance between extracellular matrix deposition and degradation and prevented an inflammatory response. Keywords: co-culture, HGFs, inflammation, matrix metalloproteinases, tissue remodelling, b1 integrinI documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
