Generation and Characterization of Induced Pluripotent Stem Cell Lines From Multiple Sclerosis Patients and Healthy Individuals

Begentaş, Onur Can
Multiple Sclerosis (MS) is an autoimmune disease characterized by inflammation, demyelination, and axonal damage resulting in neurodegeneration in the central nervous system (CNS). Relapsing-remitting MS (RRMS) is the most common form of the disease known to affect more than 2 million individuals globally, and the prevalence of the disease is increasing worldwide. As a chronic condition without a cure, RRMS manifests in a relapsing-remitting form with sporadic attacks suddenly appearing, causing neurological dysfunction to proceed by a recovery phase where all symptoms suddenly disappear. Although the factors causing or contributing to MS initiation/progression are not well understood, previous literature has implicated many potential risk factors for the etiology of the disease, including Epstein-Barr virus, smoking, and Vitamin D deficiency. Another potential risk factor implicated in RRMS is sex. Epidemiological studies strongly suggest that MS is a sexually biased disease that is more common in females. Moreover, the disease progression and severity appear to be sexually dimorphic since the progression and the symptoms are worse in males; however, more inflammatory in females. Importantly, research suggests that biological sex differences beyond gender-related factors may be crucial in RRMS; however, how biological sex differences affect the disease initiation or progression remain unknown. The hallmark of MS is demyelination, which is the loss of the myelin sheath around axons in the CNS. Oligodendrocytes are the glial cells generating myelin sheath to axons in CNS. It is well established that autoimmune response in MS leads to damage and potentially loss of oligodendrocytes, eventually leading to neurodegeneration. Current therapeutic approaches for MS are only limited to eliminating the symptoms caused by the autoimmune attacks. While such disease-modifying therapies help suppress autoimmune attacks and eventually lower the frequency of attacks, there are no available therapeutics to induce remyelination. Clearly, there is a need for more advanced approaches in MS that would induce remyelination besides immunosuppression. To develop effective therapeutic options that can induce remyelination to complement existing immunosuppressive drugs, a better understanding of cellular processes in remyelination is needed. To facilitate such studies, reliable model systems are necessary for mechanism and drug discovery studies. Human induced pluripotent stem cells (iPSCs) based approaches can provide such systems. Due to their nature and ability to form every possible cell type from the three embryonic germ layers, they can now be easily generated from patients’ somatic cells to generate patient-specific cell types. iPSCs can be utilized to obtain cell types that are normally very hard to obtain on a large scale, especially for CNS disease modeling. Indeed, numerous studies have demonstrated that iPSCs can be successfully differentiated to neuronal and glial cells, and importantly such cells exhibit disease-specific phenotypes in culture. MS iPSCs have also been generated by various groups, and cells differentiated from the iPSCs exhibit disease characteristics, suggesting that these cells can be utilized for MS studies. The main goal in this thesis was to generate and fully characterize iPSC lines from male and female patients and matched controls to facilitate future studies. Towards this goal, blood samples were obtained from age, and sex-matched RRMS patients (3 females and 3 males) and 6 healthy individuals (3 females and 3 males) and Peripheral Blood Mononuclear Cells (PBMCs) were isolated. Following, the PBMCs were reprogrammed using Sendai virus-based reprogramming method, and a total of 12 iPSC lines were established, and their pluripotent characteristics were experimentally characterized. The established iPSC lines will serve as excellent tools for modeling RRMS, enabling us to test multiple approaches to understand the mechanisms governing sex-specific differences and remyelination.


Generation and characterization of human induced pluripotent stem cell line METUi001-A from a 25-year-old male patient with relapsing-remitting multiple sclerosis
Koc, Dilara; Begentaş, Onur Can; Yurtogullari, Sukran; Temel, Musa; Akcali, Kamil Can; Demirkaya, Seref; Kiriş, Erkan (2021-05-01)
Multiple sclerosis is a chronic disease characterized by inflammation, demyelination, and axonal damage in the central nervous system. Here, we established an induced pluripotent stem cell (iPSC) line METUi001-A from the peripheral blood mononuclear cells of a 25-year-old male individual with clinically diagnosed Relapsing-Remitting Multiple Sclerosis (RRMS) using the integration-free Sendai reprogramming method. We demonstrated that the iPSCs are free of exogenous Sendai reprogramming vectors, have a norma...
Investigation of Glutathione S-Transferase (GST) expression and activity in mouse with Multiple Sclerosis (MS)
Arçak, Deniz; Adalı, Orhan; Evin, Emre; Department of Biology (2022-5-11)
Multiple Sclerosis (MS) of unknown etiopathogenesis is a chronic demyelinating disease of the central nervous system. It mainly destroys myelin in the brain and spinal cord. Non-traumatic injuries have been observed in this disease for young adults. Various factors affect MS, but oxidative stress is one of the most important causes of demyelination. Glutathione S- Transferases (GSTs) can be described as a versatile enzyme family of eukaryotic and prokaryotic phase II metabolic isoenzymes. They have enzymati...
Evin, Emre; Adalı, Orhan; Department of Biology (2021-8)
Multiple sclerosis (MS) is a complicated, recurrent, and often progressive inflammatory demyelinating autoimmune disease of the central nervous system, yet etiopathogenesis remains unsolved. MS frequency increases with increasing latitude, leading to a hypothesis that MS is inversely correlated with the duration and intensity of sunlight and vitamin D concentrations. In this study, the relationships between vitamin D supplementation, MS, VDR, and vitamin D metabolizing CYP enzymes, including CYP2R1, CYP27A1...
Relapsing-Remitting Multiple Sclerosis diagnosis from cerebrospinal fluids via Fourier transform infrared spectroscopy coupled with multivariate analysis
Yonar, Dilek; Ocek, Levent; Tiftikcioglu, Bedile Irem; Zorlu, Yasar; Severcan, Feride (2018-01-18)
Multiple sclerosis (MS) is a chronic, progressive, inflammatory and degenerative disease of central nervous system. Here, we aimed to develop a method for differential diagnosis of Relapsing-Remitting MS (RRMS) and clinically isolated syndrome (CIS) patients, as well as to identify CIS patients who will progress to RRMS, from cerebrospinal fluid (CSF) by infrared (IR) spectroscopy and multivariate analysis. Spectral analyses demonstrated significant differences in the molecular contents, especially in the l...
Association analysis of cholesterol 7-alpha hydroxylase (CYP7A1)and cholesterol 24-hydroxlase (CYP46A1) genetic polymorphisms and multiple sclerosis risk in Turkish population
Sezer, Eda; Kaya, Zeki; Department of Molecular Biology and Genetics (2019)
Multiple Sclerosis (MS) is the most common demyelinating disorder of the central nervous system. Under the effects of certain environmental factors, MS develops in genetically susceptible individuals. People with MS have significantly lower vitamin D levels. UV-B radiation catalyzes the photo-conversion of 7-dehydrocholesterol, produced in cholesterol production pathway, to vitamin D in the skin. Cholesterol 7α-hydroxylase (CYP7A1) in the liver and Cholesterol 24S-hydroxylase (CYP46A1) in the brain are resp...
Citation Formats
O. C. Begentaş, “Generation and Characterization of Induced Pluripotent Stem Cell Lines From Multiple Sclerosis Patients and Healthy Individuals,” M.S. - Master of Science, Middle East Technical University, 2021.