Sağlıklı İnsan Uyarılmış Pluripotent Kök Hücrelerinden Kortikal Nöron Elde Edilmesi
Temel Özkılıç, Melis
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Induced pluripotent stem cells (iPSC) are a unique resource for personalized treatment, disease modeling and drug trials. Target tissue and organ formation can be achieved with various transcription factors or small molecules from blood and fibroblast cells taken from the patient. Tissues and organs such as kidney, heart, liver, brain, etc. can be differentiated. The most interesting of these is undoubtedly the brain. Unlike other tissues and/or organs, it is a difficult area to study due to the lack of biopsy opportunity. After pluripotency is achieved by transferring Yamanaka factors to the blood and/or fibroblast sample taken from the patient, differentiation into nerve cells or organoids can be achieved with appropriate differentiation protocols. The aim of this thesis is to obtain cortical neuron from healthy human pluripotent stem cells (iPSCs) within 12- 15 days using six small molecules. In this context, characterized healthy human uPPCs were propagated in culture. Pluipotentiation was confirmed by qPCR and immunofluorescence staining to check the preservation of markers. For neuronal differentiation experiments, the cells were treated with cocktails of six small molecules at different doses every day. The morphologic status of the cells was monitored daily by light microscopy. At the end of the protocol, RNA was collected from the cells and immunofluorescent staining was performed. As a result of qPCR experiments, a significant increase in the expression of cortical neuron precursors DCX, FOXG1 and REELIN was observed in the cells obtained compared to the positive control. MAP2 and TUJ1 positive cells were observed in immunofluorescence staining.
- Kök Hücre Bilimleri