ABSTRACT OF THE THESIS

Myt1l Contributes to Reprogramming of Mature Hepatocytes to a Beta-cell Like State

by

Melissa Tran

Master of Science in Biology

University of California San Diego, 2020

Professor Marc Montminy, Chair

Professor Gen-Sheng Feng, Co-Chair

The goal of regenerative medicine is to replenish damaged and diseased cells in the body. In order to do so, mature cells are encouraged to convert towards another cell type. One way of doing this is by overexpressing specific genes using viral vector delivery. Previous research has shown that the beta cell specific transcription factors Pdx-1, NeuroD1, MafA, Insm1, Isl1, and Pax6 are able to shift liver cells and exocrine pancreas cells towards a beta cell-like state. In this experiment, we analyze different combinations of these lineage determining transcription factors and their reprogramming effects on mature hepatocytes. We were also prompted to investigate Myt1l, a gene that has previously only been found in neuronal tissue. Previous work done in the lab, however, has shown it to be highly enriched in islets, which may suggest that the gene plays a critical role in beta cell function. Here we show that combining the transcription factors Pdx-1, NeuroD1, MafA, Insm1, Isl1, Pax6 and Myt1l leads to a substantial shift towards a beta cell-like state based on the increased expression of IAPP, a beta cell marker that is co-expressed with insulin to regulate blood glucose levels with satiety signals. Furthermore, our results suggest that Myt1l is an important factor converting mature hepatocytes towards a more beta cell-like state. Based on transcriptomic characterization, experimental conditions in the presence of Myt11 when compared to the experimental conditions without Myt1l display an upregulation of beta cell markers that are closely related to insulin secretion. These observations provide evidence that Myt1l is a key beta cell reprogramming factor.

Introduction: Over the past decade, medical students have witnessed a decline in the opportunities to perform technical skills during their clinical years. Ultrasound-guided central venous access (USG-CVA) is a critical procedure commonly performed by emergency medicine, anesthesia, and general surgery residents, often during their first month of residency. However, the acquisition of skills required to safely perform this procedure is often deficient upon graduation from medical school. To ameliorate this lack of technical proficiency, ultrasound simulation models have been introduced into undergraduate medical education to train venous access skills. Criticisms of simulation models are the innate lack of realistic tactile qualities, as well as the lack of anatomical variances when compared to living patients. The purpose of our investigation was to design and evaluate a life-like and reproducible training model for USG-CVA using a fresh cadaver.

Methods: This was a cross-sectional study at an urban academic medical center. An 18-point procedural knowledge tool and an 18-point procedural skill evaluation tool were administered during a cadaver lab at the beginning and end of the surgical clerkship. During the fresh cadaver lab, procedure naïve third-year medical students were trained on how to perform ultrasound-guided central venous access of the femoral and internal jugular vessels. Preparation of the fresh cadaver model involved placement of a thin-walled latex tubing in the anatomic location of the femoral and internal jugular vein respectively.

Results: Fifty-six third-year medical students participated in this study during their surgical clerkship. The fresh cadaver model provided high quality and lifelike ultrasound images despite numerous cannulation attempts. Technical skill scores improved from an average score of 3 to 12 (p<0.001) and procedural knowledge scores improved from an average score of 4 to 8 (p<0.001).

Conclusion: The use of this novel cadaver model prevented extravasation of fluid, maintained ultrasound-imaging quality, and proved to be an effective educational model allowing third-year medical students to improve and maintain their technical skills.