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KOÇ UNIVERSITY

GRADUATE SCHOOL OF SCIENCES & ENGINEERING

CHEMICAL AND BIOLOGICAL ENGINEERING

MS THESIS DEFENSE BY MÜKRİME BİRGÜL AKOLPOĞLU

 

Title: Development of Biomimetic Approaches for Immunoisolation of Functional Islets

 

Speaker: Mükrime Birgül Akolpoğlu

 

Time: September 10, 2018, 11.00

 

Place: ENG B11

Koç University

Rumeli Feneri Yolu

Sariyer, Istanbul

 

Thesis Committee Members:

Assoc. Prof. Seda Kızılel (Advisor, Koç University)

Asst. Prof. Cem Albayrak (Koç University)

Prof. Nihan Nugay (Boğaziçi University)

 

Abstract:

Type 1 diabetes (T1D) is a chronic autoimmune disease of pancreas, where insulin secretion function is lost due to destruction of insulin secreting β cells. People with T1D have high blood glucose levels, while their cells are deprived from glucose for their metabolic actions. Islets are “islands of β cells” and responsible for glucose metabolism in the body. Islet transplantation has transitioned from an experimental and occasionally-employed strategy to a routine clinical therapy for T1D in the past decade. Nevertheless, lifelong immunosuppression and the necessity of multiple transplantations limit wider application of this strategy. Islet immunoisolation techniques have emerged to eliminate immunosuppression permanently or to reduce immunosuppressive drug doses significantly. The concept of immunoisolation is to create a local secure microenvironment for islets to prevent graft rejection. Regulation of immune system can be done by several material-based and biologic strategies. While material-based strategies focus on preserving islets within semi-permeable membranes that allow nutrient exchange and block immune system components, biological strategies deal with regulation of immune system through chemokines and manipulation of immune cells. This thesis focuses on two of those strategies to create tolerable grafts and overcome islet graft rejection. We firstly engineered pseudoislets, or islet organoids, that are composed of insulin secreting β cells (β-TC-6) and hepatic stellate cells (HSCs). HSCs have the ability to secrete ECM proteins, angiogenesis factors and expand regulatory T cell (Treg) population in their vicinity. Tregs are crucial in modulating the immune system by suppressing and downregulating actions of effector T cells. A macrophage derived chemokine, CCL22, also has the ability to recruit Tregs and provide a local immunosuppression. We combined these two concepts and transfected Tregs with CCL22 gene. Then we prepared insulin-secreting multicellular organoids with β-TC-6 and CCL22-transfected HSCs. Implantation of these multicellular organoids to diabetic animal model resulted in more than 5-fold increase in Treg recruitment towards implantation site. This result suggested that tolerable grafts could be fabricated through modulation of the immune system not only for islet therapy but also for other cell/organ transplantation therapies. To create another type of immunoisolation for islets, we also designed biocompatible hydrogels and formed ultra-thin coatings around islet organoids. Lipid group baring microgels were synthesized by water-in-water emulsion (W/W) followed by photopolymerization steps. We optimized microgel diameter by changing emulsion and photopolymerization parameters. The smallest microgel diameter (2,13 µm) was achieved at 60% ultrasonication power for 30 minutes. In W/W emulsion and photopolymerization steps, each reaction step was carried out in aqueous solutions at physiological pH values. We prepared β-TC-6 organoids by hanging drop method and deposited lipid-microgels on organoid surface via non-covalent hydrophobic interactions. Unlike covalent bonding, hydrophobic interactions between lipid functionalities in our microgels and phospholipid bilayer on cell membranes present no harm to cells. Hydrophobic interactions do not damage membrane proteins and perturb the integrity of the membrane. We studied the coating of microgels with two different lipid concentrations, 2.5 and 5 mM. Furthermore, we coated islet organoids with two different microgel concentrations, 10 and 20 mg/ml. We observed that coating efficiency was higher when lipid concentration was increased due to more hydrophobic interactions. We obtained high and similar metabolic activity and viability for microgel coated islet organoids compared to non-coated controls. Insulin secretion functionality was also preserved, meaning that our approach of immunoisolation allowed us to engineer functional insulin secreting organoids. Our findings suggest that biological and material-based immunotherapeutic strategies hold great potential for islet transplantation in T1D treatment.

 

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