Master Degree / Yüksek Lisans Tezleri

Permanent URI for this collectionhttps://hdl.handle.net/11147/3008

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Now showing 1 - 3 of 3
  • Master Thesis
    The Investigation of the Role of Nf-Κb in the Proliferation and Differentiation of Hacat Cells Containing Cx26 Kid Syndrome Mutations
    (01. Izmir Institute of Technology, 2024) İnal, Ece; Özçivici, Gülistan Meşe
    Keratit-iktiyozis-sağırlık (KID) sendromu, Connexin26 (Cx26) mutasyonlarından kaynaklanan, sağırlık ve görme bozukluklarının yanı sıra palmoplantar keratoderma (avuç içi ve ayak tabanlarında epidermisin kalınlaşması) ile karakterize edilen nadir bir genetik hastalıktır. Cx26 mutasyonları, hücre zarında kontrolsüz molekül geçişine neden olan sürekli açık, hiperaktif yarım kanalların oluşumuna yol açar. Bu durum keratinositlerin proliferasyonu ve farklılaşmasını etkilese de, Cx26 mutasyonlarından kaynaklanan epidermal değişikliklerin altında yatan mekanizmalar henüz bilinmemektedir. Proteomik çalışmalar, Cx26-D50Y KID sendromu mutasyonu içeren HaCaT keratinosit hücre hattında NF-κB sinyal yolağı proteinlerinde zenginleşme olduğunu göstermiştir. Bu nedenle, Cx26 mutant kanallarının NF-κB yolağı aracılığıyla keratinosit proliferasyon ve farklılaşma mekanizmalarını etkileyerek epidermal bozukluklara yol açabileceği hipotez edildi. Hastalarda farklı şiddetlere neden olan Cx26-G45E and Cx26-D50Y mutasyonlarını sürekli olarak ifade eden HaCaT hücrelerinin proliferasyon ve farklılaşma mekanizmaları üzerinde NF-κB sinyal yolağının rolünü araştırdık. G45E ve D50Y hücreleri sırasıyla çekirdekte bulunan en yüksek RelA ve c-Rel sinyallerini gösterdi. Ek olarak, NaSal muamelesi D50Y hücrelerinde erken ve geç apoptoz oranlarını farklı şekilde etkiledi. Dahası, erken ve geç apoptoz oranları karşılaştırıldığında G45E ve D50Y hücreleri ters yönde bir trend gösterdi. Ayrıca, NF-κB inhibisyonu, G45E hücrelerinin proliferasyon oranını azalttı. Son olarak, NaSal muamelesinden sonra G45E hücrelerinde cytokeratin10 protein seviyelerinde farklılıklar görüldü. G45E ve D50Y'de NF-κB, apoptozu farklı mekanizmalarla etkileyebilir ve D50Y'nin apoptoz mekanizmaları üzerinde farklı etkiler gösteriyor olabilir. Ayrıca, NF-κB, G45E'nin proliferasyon ve farklılaşma mekanizmalarını düzenliyor olabilir. KID sendromu için mevcut bir tedavi bulunmamaktadır. Bu nedenle, bu çalışma KID sendromunun moleküler ve hücresel mekanizmalarını anlamak açısından önemlidir.
  • Master Thesis
    The Effects of Calcium on Nf-Κb Pathway in Hacat Cells Containing Connexin26 Kid Syndrome Mutations
    (01. Izmir Institute of Technology, 2023) Yaşarbaş, Sümeyye Şüheda; Özçivici, Gülistan Meşe
    Keratitis-ichthyosis-deafness (KID) syndrome is a rare genetic disorder characterized by deafness, corneal defects, and thickened, scaly skin and associated with mutations in Connexin26 (Cx26), resulting in the formation of hyperactive hemichannels that disrupt calcium (Ca2+) transfer. Ca2+ is crucial for normal epidermal cell function and may contribute to characteristics of KID syndrome. While unregulated Ca2+ transfer through aberrant Cx26 hemichannels is known to impact keratinocyte proliferation and differentiation, the specific mechanisms remained unclear. An increase in the molecules associated with the nuclear factor-κB (NF-κB) signaling pathway was observed in the D50Y mutation of KID syndrome. This suggests that Cx26 mutant channels may disrupt keratinocyte physiology through NF-κB signaling. Our study hypothesizes that Ca2+ signals altered due to Cx26 mutations affect the NF-κB pathway, potentially contributing to KID syndrome by modifying keratinocyte cell physiology. Our study showed that NF-κB activation significantly increased in D50Y cells, linked to hyperproliferation and activation was dependent on intracellular Ca2+. This was associated with increased p65 activation and nuclear localization due to hyperactive Cx26 channels in D50Y cells. These findings reveal a direct link between aberrant Ca2+ transport through Cx26 channels due to the D50Y mutation and NF-κB activation, shedding light on the hyperproliferative characteristics of Cx26 D50Y KID syndrome. Our goal was to understand how Ca2+ mechanisms impact the NF-κB pathway, potentially altering the physiology of keratinocytes expressing D50Y and G45E mutations. This research offers insights into the potential targeting of the NF-κB pathway for treating KID syndrome caused by Cx26 mutations.
  • Master Thesis
    Deciphering Functions of Aberrant Hemichannels Formed by Connexin 26- I30n and D50y Mutations
    (Izmir Institute of Technology, 2015) Aypek, Hande; Meşe Özçivici, Gülistan
    Cells need to communicate with each other for maintenance cellular and tissue homeostasis. Gap junctions are channel-forming structures that are formed by docking of two hemichannels on the plasma membrane of adjacent cells. Connexins are subunits of gap junctions. Connexin 26 (Cx26) is one of the connexin isoform and mutations on the Cx26 gene (GJB2) cause non-syndromic and syndromic deafness. Keratitis-ichthyosis-deafness (KID) syndrome is one of the syndromic deafness disorders caused by Cx26 mutations. Among these mutations, Cx26-I30N and D50Y missense mutations were shown to form aberrant hemichannels but their effect on protein biosynthesis and functions have not studied. In this study, we aimed to decipher in vitro functions of aberrant hemichannels formed by Cx26-I30N and D50Y mutations. First of all, the effect of Cx26-I30N and D50Y mutations on localization, mRNA expression and protein synthesis properties were investigated in HeLa, N2A and HaCaT cells. Results suggested that Cx26-I30N and D50Y mutants were not able to form gap junction plaques on the plasma membrane and were localized in the Golgi apparatus. In addition, mutations resulted in a reduction in mRNA expression and protein synthesis. After, functional analysis was performed in Cx26-I30N and D50Y transfected N2A and HaCaT cells. Internal Ca2+ content measurement, measurement of released ATP, measurement of cell size and apoptosis assays were performed. Ca2+ measurement results showed that both Cx26-I30N and D50Y mutations deregulate Ca2+ balance in both N2A and HaCaT cells. Result of ATP release assay indicated that ATP amount in the extracellular environment decreased in N2A cells having Cx26-I30N and D50Y clones. Finally, apoptosis assay showed that number of necrotic cells increased when N2A cells were transfected with Cx26-I30N and D50Y constructs. Therefore, it was shown that aberrant hemichannels formed by Cx26-I30N and D50Y mutations may induce necrotic cell death by disrupting Ca2+ balance and ATP amount in cells.