Publication: Mezenkimal kök hücre farklılaşmasında GAS6 (growth arrest specıfıc gene 6) nın rolü
Abstract
1.ÖZET Amaç: GAS6 (Growth Arrest-Specific 6) proteini, K vitaminine bağımlı protein ailesinin yeni üyelerinden biridir. GAS6, reseptör tirozin kinaz ailesinden Tyro3 (Sky), Axl ve Mer (TAM) reseptörlerinin ligandıdır. Bu tez, insan adipoz doku kaynaklı mezenkimal kök hücrelerin osteojenik, kondrojenik ve adipojenik farklılaşması sırasında ortama eklenen GAS6 proteinin etkisi inceleyen ilk çalışmadır. Gereç ve Yöntem: Estetik amaçla yapılan “liposuction” işlemi sırasında karından alınan yağın bir bölümü bu çalışmada başlangıç materyali olarak kullanıldı. Daha sonra hücreler, uygun farklılaştırma medyumları kullanılarak adipojenik, kondrojenik ve osteojenik farklılaşmaya bırakıldı. Paralel olarak yürütülen deneylerde, ortama eksojen olarak 100 ve 500 ng/ mL GAS6 proteini eklendi. Farklılaşma işlemleri sonunda spesifik hücre belirteçleri (FABP4, Kollajen Tip 1 ve Tip2) ve TAM reseptörlerinin mRNA ekspresyon düzeyleri Gerçek Zamanlı- Polimeraz Zincir Reaksiyon (GZ-PZR) ile belirlendi. Hücreler uygun boyalarla boyanarak morfolojik yönden incelendi. Bulgular: 100 ng/ mL GAS6’nın adipojenik ve kondrojenik farklılaşmayı arttırdığı; 500 ng/ mL’nin ise inhibe ettiği belirlendi. Osteojenik farklılaşma sırasında 100 ng/ mL GAS6 inhibisyona neden olurken, 500 ng/ mL’nin farklılaşmayı uyardığı saptandı. Sonuç: Bu çalışmada, mezenkimal kök hücrelerin adipojenik, kondrojenik ve osteojenik farklılaşması sırasında ortama GAS6 eklenmesinin olumlu etkileri olduğu belirlendi. Bu mekanizmaların aydınlatılması için daha kapsamlı araştırmalara ihtiyaç vardır. Growth Arrest-Specific protein 6”, mezenkimal kök hücreler, AXL, TYRO3, MER. 1.
Purpose: GAS6 (Growth Arrest-Specific 6) protein is a new member of vitamin K dependent proteins. It is a ligand of Tyro3 (Sky), Axl and Mer (TAM) receptors from the tyrosine kinase family. This thesis is the first study on the role of exogenous GAS6 added to the medium during the differantiation of mesenchymal stem cells into osteogenic, chondrogenic and adipogenic cells. Material and Method: A part of fat collected by liposuction was used for isolation of mesenchymal stem cells, which were subjected to osteogenic, adipogenic and condrogenic differentiation. 100 or 500 ng/ mL GAS6 protein was added into the differantiation medium in parallel tests. At the end of differentiation, Real Time-PCR analyses were performed to determine mRNA expression of FABP4, Collagen Type 1 and 2, and TAM receptors. The cells were stained with morphologically suitable dyes. Results: GAS6 (100 ng/ mL) had a positive effect on adipogenic and chondrogenic differentiation although 500 ng/ mL GAS6 inhibited those differentiations. 500ng/ mL GAS6 had a positive effect on osteogenic differentiation although 100 ng/ mL GAS6 dose had a negative effect. Conclusion: It was concluded that exogenous GAS6 addition to the medium has a positive effect on the differantiation of mesenchymal stem cells into osteogenic, chondrogenic and adipogenic cells. Further studies are needed to clarify the role of GAS6/ TAM system on mesenchymal stem cell differentiation. Key words: Growth Arrest-Specific protein 6, mesenchymal stem cells, AXL, TYRO3, MER.
Purpose: GAS6 (Growth Arrest-Specific 6) protein is a new member of vitamin K dependent proteins. It is a ligand of Tyro3 (Sky), Axl and Mer (TAM) receptors from the tyrosine kinase family. This thesis is the first study on the role of exogenous GAS6 added to the medium during the differantiation of mesenchymal stem cells into osteogenic, chondrogenic and adipogenic cells. Material and Method: A part of fat collected by liposuction was used for isolation of mesenchymal stem cells, which were subjected to osteogenic, adipogenic and condrogenic differentiation. 100 or 500 ng/ mL GAS6 protein was added into the differantiation medium in parallel tests. At the end of differentiation, Real Time-PCR analyses were performed to determine mRNA expression of FABP4, Collagen Type 1 and 2, and TAM receptors. The cells were stained with morphologically suitable dyes. Results: GAS6 (100 ng/ mL) had a positive effect on adipogenic and chondrogenic differentiation although 500 ng/ mL GAS6 inhibited those differentiations. 500ng/ mL GAS6 had a positive effect on osteogenic differentiation although 100 ng/ mL GAS6 dose had a negative effect. Conclusion: It was concluded that exogenous GAS6 addition to the medium has a positive effect on the differantiation of mesenchymal stem cells into osteogenic, chondrogenic and adipogenic cells. Further studies are needed to clarify the role of GAS6/ TAM system on mesenchymal stem cell differentiation. Key words: Growth Arrest-Specific protein 6, mesenchymal stem cells, AXL, TYRO3, MER.