Osteojenik Ortam ile Demineralize Dentinde Kültür Edilen DPKH'lerden Büyüme Faktörlerinin Uzun Süreli Salım Profili ve Odontojenik Farklılaşma Üzerine Etkisi

Abstract

Kemik morfogenetik proteini (BMP-2) ve vasküler endotel büyüme faktörünün (VEGF) dentin ve kök hücrelerden salınarak, diş pulpa kök hücrelerinin (DPKH) odontogenik faklılaşmasına katıldıkları bilinmektedir. Bununla birlikte, demine- ralize dentin üzerindeki DPKH’lerden VEGF ve BMP-2’nin uzun süreli salım profilleri henüz osteojenik farklılaşma ortamında araştırılmamıştır. Bu çalışmada, insan diş dentini % 15’lik etilendiamintetraasetik asit (EDTA) ile 15 dakika demineralize edildikten sonra insan diş pulpasın- dan izole edilen DPKH’ler demineralize dentin üzerinde 28 gün kültür edilmiş ve VEGF ve BMP-2’nin salım miktarları ve bunların odontojenik farklılaşması üzerindeki etkileri araştırılmıştır. Sonuçlar, hücrelerin demineralize dentin yüzeyine olduk- ça iyi tutunduklarını ve çoğaldıklarını göstermiştir. 28 gün sonunda DPKH’lerin varlığında, demineralize dentinden 0.45 ng/ mL BMP-2 salımı gözlenirken, hücre varlığında 18.5 ng/mL VEGF salımı olmuştur. Büyüme faktörlerinin bu miktarları Dentin Matriks Protein-1 (DMP-1) ifadesinde 10.7 katlık artış sağlarken, Kemik Sialoprotein (BSP) ifadesinde bir artış olmamıştır. Hücre farklılaşmasını artırmak için osteojenik ortama rekombinat büyüme faktörlerinden (rhBMP-2 ve rhVEGF) 10 ng/mL eklenmiş ve DMP-1 ve BSP ifadeleri değerlendirilmiştir. Kontrol grubu ile kıyaslandığında, rhBMP-2 ve rhVEGF eklenmesi, DMP-1 ifadesinde sırasıyla 32.9 ve 26.4 kat artışa neden olmuştur (p<0.05). Diğer taraftan, rhBMP-2 ve rhVEGF, BSP ifa- desinde sırasıyla 4.7 ve 3.6 katlık artış sağlamıştır (p<0.05). DPKH'lerin odontojenik farklılaşmasının artırılmasında ortama eksojen VEGF ve BMP-2’nin eklenmesinin yararlı olacağı sonucuna varılmıştır. 

Keywords

• pulpa rejenerasyonu,dental pulpa kök hücresi,vasküler endotel büyüme faktörü,kemik morfogenetik proteini,odontogenik farklılaşma

Project Number

THD-2015-7664

Long-Term Release Profiles of Growth Factors From DPSCs on Demineralized Dentin Under Osteogenic Environment and Their Effects on Odontogenic Differentation

Abstract

It is known that bone morphogenetic protein (BMP-2) and vascular endothelial growth factor (VEGF) are released from dentin and stem cells, concomitantly participate in odontogenic differentiation of dental pulp stem cells (DPSCs). However, long-term release profiles of VEGF and BMP-2 from DPSCs on demineralized dentin have not yet been investigated under osteogenic differentiation conditions. In this study, after demineralization of human dentin by 15% ethylenediaminetetraacetic acid (EDTA) for 15 min, isolated human DPSCs were cultured on demineralized dentin for 28 days and the release amount of VEGF and BMP-2 and their effects on odontogenic differentiation of cells was investigated. Results showed that cells well attached to demineralized dentin surfaces and proliferated very well. At the end of day 28, BMP-2 and VEGF release from demineralized dentin was determined as 0.45 and 18.5 ng/mL respectively in the presence of cells. The amount of released growth factors caused 10.7-fold increase in Dentin Matrice Protein 1 (DMP-1) expression whereas Bone Sialoprotein (BSP) expression was not upregulated. In an attempt to increase the odontogenic related gene expressions, 10 ng/mL of each recombinant growth factors (rhBMP-2 and rhVEGF) were added into osteogenic medium and DMP-1 and BSP expression were evaluated. As compared to the control group, in terms of DMP-1, rhBMP-2 and rhVEGF resulted in 32.9-fold and 26.4-fold increase, respectively (p<0.05). On the other hand, rhBMP-2 and rhVEGF caused a 4.7-fold and 3.6-fold increase of BSP expression, respectively (p<0.05). As a result, it has been concluded that to increase the odontogenic differentiation of DPSCs, exogenous growth factors should be added to the osteogenic medium.

Keywords

• pulp regeneration,stem cells,vascular endothelial growth factor,bone morphogenetic protein,odontogenic differentiation

Supporting Institution

Hacettepe University Scientific Research Projects Coordination Unit

Project Number

THD-2015-7664

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