Inhibition Of The Akt1-Mtorc1 Axis Alters Venous Remodeling To Improve Arteriovenous Fistula Patency

Yale University

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Yale Medicine Thesis Digital Library School of Medicine

January 2019

Inhibition Of The Akt1-Mtorc1 Axis Alters Venous Remodeling To

Improve Arteriovenous Fistula P enous Fistula Patency

Arash Fereydooni

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Recommended Citation

Fereydooni, Arash, "Inhibition Of The Akt1-Mtorc1 Axis Alters Venous Remodeling To Improve

Arteriovenous Fistula Patency" (2019). Yale Medicine Thesis Digital Library. 3899.

https://elischolar.library.yale.edu/ymtdl/3899

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Inhibition of the Akt1-mTORC1 Axis Alters Venous

Remodeling to Improve Arteriovenous Fistula Patency

A Thesis Submitted to the

Yale University School of Medicine

in Partial Fulfillment of the Requirements for the

Degree of Doctor of Medicine and

Master of Health Sciences

By

Arash Fereydooni

2020

Abstract

Arteriovenous fistulae (AVF) are the most common access created for

hemodialysis, but up to 60% do not sustain dialysis within a year, suggesting a need to

improve AVF maturation and patency. In a mouse AVF model, Akt1 regulates fistula wall

thickness and diameter. We hypothesized that inhibition of the Akt1-mTORC1 axis alters

venous remodeling to improve AVF patency. Daily intraperitoneal injections of

rapamycin reduced AVF wall thickness with no change in diameter. Rapamycin

decreased smooth muscle cell (SMC) and macrophage proliferation; rapamycin also

reduced both M1 and M2 type macrophages. AVF in mice treated with rapamycin had

reduced Akt1 and mTORC1 but not mTORC2 phosphorylation. Depletion of

macrophages with clodronate-containing liposomes was also associated with reduced

AVF wall thickness and both M1- and M2-type macrophages; however, AVF patency was

reduced. Rapamycin was associated with improved long-term patency, enhanced early

AVF remodeling and sustained reduction of SMC proliferation. These results suggest

that rapamycin improves AVF patency by reducing early inflammation and wall

thickening while attenuating the Akt1-mTORC1 signaling pathway in SMC and

macrophages. Macrophages are associated with AVF wall thickening and M2-type

macrophages may play a mechanistic role in AVF maturation. Rapamycin is a potential

translational strategy to improve AVF patency.

Acknowledgements

I am eternally indebted to my incredible mentor, Professor Alan Dardik, for his

constant support and insight; he has served as an inspiring role model and showed me

what it means to be a successful surgeon-scientist. He has invested in my career and

given me opportunities I did not deserve. I am grateful to my colleagues at Dardik Lab

for their help, particularly Dr. Jolanta Gorecka for her teamwork and willingness serve as

a valuable sounding board.

I would like to also thank my clinical mentors, Dr. Cassius Ochoa Chaar and Dr.

Naiem Nassiri, for showing me what it means to be excellent academic surgeons, to

deliver the best comprehensive care to our patients, and not to be afraid to push the

envelope and advance the field of vascular surgery. Drs. Julia Chen, Christine Deyholos,

Anand Brahmandam, Robert Botta, Jason Chin and Kristine Orion, I sincerely appreciate

your teaching, mentorship and friendship. Dr. Raul Guzman, thank you for your

leadership, support and encouragement.

I would like to thank the Howard Hughes Medical Institute, the Society for

Vascular Surgery and the American Heart Association for funding my research at Dardik

Lab. I would also like to thank the Office of Student Research for their support with my

research endeavors throughout medical school.

Most importantly, my journey to become a surgeon-scientist would not be

possible without the sacrifices of my parents, Alireza and Naimeh, who unrooted their

lives and immigrated to the United States ten years ago to provide my sisters and me

with better educational opportunities. This work is dedicated to them.

Table of Contents

1. Introduction……………………………………………………………………………………………………………1

1.1. Poor Clinical Outcomes in Arteriovenous Fistulae Utilization……………………….1

1.2. Mechanisms of Fistula Maturation and Failure…………………………………………….1

1.3. Akt1 signaling in AVF maturation…………………………………………………………………4

2. Statement of Purpose and Aims……………………………………………………………………………..6

2.1. Statement of Purpose

2.2. Aims

3. Methods…………………………………………………………………………………………………………………7

3.1. Study Approval…………………………………………………………………………………………….7

3.2. Infrarenal aorto-caval fistula………………………………………………………………………..7

3.3. Confirmation of fistula patency and measurement of fistula dilation…………..7

3.4. Histology.…………………………………………………………………………………………………….8

3.5. Immunohistochemistry and Immunofluorescence……………………………………….8

3.6. Western Blot.……………………………………………………………………………………….……10

3.7. Rapamycin and clodronate treatment…………………………………………………..……11

3.8. Adenovirus treatment………………………………………………………………………………..12

3.9. Statistics.……………………………….…………………………………………………………………..12

4. Results………………………………………………………………………………………………………………….13

4.1. Reduced AVF wall thickness, extracellular matrix deposition, SMC and

macrophages with rapamycin…………………………………………………..………………………13

4.2. Reduced M1- and M2-type macrophages with rapamycin…………………………15