Monogenic diabetes modelling using genetically engineered human pluripotent stem cells

Tutkimustuotos: OpinnäyteVäitöskirjaArtikkelikokoelma

Abstrakti

Diabetes mellitus is a metabolic disorder characterised by inadequate insulin secretion necessary for regulating blood glucose levels. The aetiology of developing diabetes is heterogenous and diverges across its types. Nevertheless, the ultimate endpoint of most types is the deterioration of insulin-secreting pancreatic β-cells, culminating in the imperative for exogenous insulin as a replacement therapy. The emergence of human pluripotent stem cells (hPSCs) facilitated the generation of unlimited numbers of stem-cell-derived islets (SC-islets) that closely resemble adult human islets, via mimicking the signalling ques discerned from rodent and human pancreatic development. The primary purpose of SC-islets is to serve as a renewable source for cell-replacement therapy, obviating the requirement of insulin administration and improving the normal blood glucose time-in-range, thereby, mitigating diabetes complications. Not only do SC-islets permit cell therapy but also the opportunity to study β-cell development and function, elucidating diverse causative mechanisms underlying diabetes. Given that hPSCs can be derived from diabetes patients and propagated indefinitely in vitro, they resemble an attractive model, particularly when combined with genetic engineering, to study diabetes-associated genes. These genes can be thoroughly investigated by either correcting mutations in patient-derived stem cells or introducing them in healthy ones, generating isogenic cell lines. Employing isogenic cell lines circumvents the variability of different genetic backgrounds when using cell lines from different donors, highlighting the impact of the genetic mutation under study. This well-controlled model is extremely appreciated in the context of monogenic diabetes, which is a type of diabetes caused by single gene mutations. Herein, we utilised CRISPR genome-editing tools to create and study isogenic stem-cell models of three diabetes-associated genes: RFX6, YIPF5 and TYK2. Regulatory factor X 6 (RFX6) is a transcription factor that regulates pancreatic β-cell development and function. While homozygous mutations in RFX6 cause neonatal diabetes, heterozygous ones predispose to type 2 diabetes. We pinpointed how heterozygous RFX6 mutation cause RFX6 haploinsufficiency leading to reduced β-cell maturation markers and intracellular calcium levels, along with reduced insulin secretion, consistent with the increased risk of variant carriers to develop diabetes. YIP1 family member 5 (YIPF5) is a transmembrane protein that is involved in cargo trafficking between ER and Golgi. Recessive mutations in YIPF5 cause neonatal/early-onset diabetes. We demonstrated how the lack or mutation of YIPF5 resulted in proinsulin retention in the ER, marked ER stress and β-cell failure, leading to diabetes. Tyrosine kinase 2 (TYK2) is a type 1 diabetes risk gene that plays a critical role in type I interferon (IFN-I) signalling. We studied the role of TYK2 in β-cell development and response to IFNα. We showed that loss of TYK2 reduced the number of endocrine precursors by perturbing KRAS and neurogenin 3 expression. However, in mature SC-islets, loss or inhibition of TYK2 prevented IFNα-induced response and enhanced their survival against T-cell cytotoxicity, emphasising the potential of TYK2 inhibition as a preventive therapy for type 1 diabetes. In conclusion, this thesis sheds light on the potential of genetically engineered isogenic SC-islets to elucidate different mechanisms of diabetes, stemming from anomalies in β-cell development, function, or survival.
Alkuperäiskielienglanti
Valvoja/neuvonantaja
  • Otonkoski, Timo, Valvoja
JulkaisupaikkaHelsinki
Kustantaja
Painoksen ISBN978-951-51-9938-6
Sähköinen ISBN978-951-51-9937-9
TilaJulkaistu - 2024
OKM-julkaisutyyppiG5 Tohtorinväitöskirja (artikkeli)

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