The kidneys are responsible for filtering out toxins, reabsorption of water, maintaining ion balance, as well as many other functions. Damage to the kidneys can be harmful to the body and even life threatening. Sudden damage to the kidney is categorized as acute kidney injury (AKI), while damage to the kidney over a long period of time, is known as chronic kidney disease (CKD). End-stage renal disease (ESRD) or kidney failure, is the last stage of CKD. At this stage, the kidneys are damaged to the point where they cannot function adequately for survival. Currently there is no cure for ESRD and kidneys at this stage require dialysis or a kidney transplant. Therefore, research on kidney diseases, development, and mechanisms will help reduce the number of patients with ESRD.
Kidney development has been studied for decades but there are still many unanswered questions about nephrogenesis mechanisms, genetic factors, and biochemical pathways. Kidney organoids are a versatile tool used to study the functions of human kidneys. They originate from human pluripotent stem cells (hPSCs) which are derived from skin cells. After certain growth factors are added they function as miniature kidneys in a dish which acts as a platform for research in nephrogenesis, drug therapies, fibrosis, mutations and response to injury. Since these organoids are derived from hPSC, genome editing techniques like CRISPR/Cas9 are able to introduce modifications and mutations to replicate certain diseases or specific differentiations.
The aim of this project is to develop tools to better understand kidney development and ultimately improve kidney organoid models to make advances against kidney diseases, injuries, and complications. In this project we make genetic modifications to hPSCs using the iCRISPR platform to provide new experimental tools in kidney organoids. To create the iCas9 hPSC line we utilize a knockin strategy into the AAVS1 locus. We also worked on generating a Nephron Progenitor Cell (NPC) – specific fluorescent reporter hPSC line corresponding with CITED1. Our goal is to provide tools for potential use in CRISPR screening experiments, create the ability to have live imaging of NPC behavior, and provide easy identification and isolation of NPCs for downstream analysis.
| Identifer | oai:union.ndltd.org:bu.edu/oai:open.bu.edu:2144/36601 |
| Date | 13 June 2019 |
| Creators | Nip, Allan |
| Contributors | Stearns-Kurosawa, Deborah, McCracken, Kyle |
| Source Sets | Boston University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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