A high-quality severe combined immunodeficiency (SCID) rat bioresource

Abstract

Immunodeficient animals are valuable models for the engraftment of exogenous tissues; they are widely used in many fields, including the creation of humanized animal models, as well as regenerative medicine and oncology. Compared with mice, laboratory rats have a larger body size and can more easily undergo transplantation of various tissues and organs. Considering the absence of high-quality resources of immunodeficient rats, we used the CRISPR/Cas9 genome editing system to knock out the interleukin-2 receptor gamma chain gene (Il2rg) in F344/Jcl rats—alone or together with recombination activating gene 2 (Rag2)—to create a high-quality bioresource that researchers can freely use: severe combined immunodeficiency (SCID) rats. We selected one founder rat with frame-shift mutations in both Il2rg (5-bp del) and Rag2 ([1-bp del+2-bp ins]/[7-bp del+2-bp ins]), then conducted mating to establish a line of immunodeficient rats. The immunodeficiency phenotype was preliminarily confirmed by the presence of severe thymic hypoplasia in Il2rg-single knockout (sKO) and Il2rg/Rag2-double knockout (dKO) rats. Assessment of blood cell counts in peripheral blood showed that the white blood cell count was significantly decreased in sKO and dKO rats, while the red blood cell count was unaffected. The decrease in white blood cell count was mainly caused by a decrease in lymphocytes. Furthermore, analyses of lymphocyte populations via flow cytometry showed that the numbers of B cells (CD3- CD45+) and natural killer cells (CD3- CD161+) were markedly reduced in both knockout rats. In contrast, T cells were markedly reduced but showed slightly different results between sKO and dKO rats. Notably, our immunodeficient rats do not exhibit growth retardation or gametogenesis defects. This high-quality SCID rat resource is now managed by the National BioResource Project in Japan. Our SCID rat model has been used in various research fields, demonstrating its importance as a bioresource.

In vivo tracking transplanted cardiomyocytes derived from human induced pluripotent stem cells using nuclear medicine imaging

“…This may be a technical problem, but our experience with II2rg/Rag2 -knockout rats used in this study gave us the impression that they had a higher mortality rate during and after open heart surgery than the commonly used Wistar rats (data not shown). Compared to Il2rg -knockout rats, Il2rg/Rag2 knockout rats are characterized by their inability to produce natural killer cells ( 31 ). We do not know if this is related to the cause of the high intraoperative mortality rate in this study.…”

A novelkitmutant rat enables hematopoietic stem cell engraftment without irradiation

“…This provides an invaluable tool for researchers studying HSC xenotransplantation. Beyond mice, rats with severe combined immunodeficiency (SCID) with Il2rg and/or Rag2 gene knockouts 29 have also been utilized in various research fields, such as the engraftment of cancer cells derived from patients 30 , as well as for the successful transplantation of human induced pluripotent stem cell-derived airway epithelial cell sheets 31 , miniature human livers 32 , and human mesenchymal progenitor cells within humanized bone constructs 33 . By combining the Kit V834M mutation with SCID rats, we can anticipate further advancements in the generation of humanized rat models for HSC research.…”

The 36th International Mammalian Genome Conference: A scientific gathering under the cherry blossoms in Tsukuba

“…First, Dr. Mashimo (Univ. Tokyo) provided a review of the progress in genome editing technology, and presented findings related to the generation of humanized rats and new genome editing method using Cas3 originally developed by his group (Miyasaka et al, 2022; Morisaka et al, 2019; Takeishi et al, 2020). Dr. Murakawa of Kyoto University presented topics such as the identification of cis ‐elements involved in gene expression regulation and diseases, and the use of long‐read sequencing technology to study transcription units within the genome (Hirabayashi et al, 2019).…”