193P - Humanized mice applying CD47; Prkcd; IL2rg triple KO mice exhibit enhanced human immune cell engraftment and reduced GvHD symptoms

Background

Humanized immune system mice provide a useful platform for various immune research fields. However, there have been limitations such as insufficient engraftment of human cells and development of graft-versus-host disease (GVHD). To overcome the limitations, we generated a novel immunodeficient mouse strain, CD47; Prkdc; IL2rg triple KO (STKO) mice based on SID mice (Prkdc; IL2rg double KO). Because CD47 plays an important role in engraftment of human tissues and cells, we expected improved human cell engraftment and GvHD development in STKO mice.

Methods

4-week female SID and STKO mice were applied to generate HSC hu-mice. 1X10⁵ hCD34+ HSCs were injected within 24 hours of myeloablation applying busulfan. For generating PBMC hu-mice 5X10⁶ PBMCs were injected. To overcome lethal GvHD, low dose PBMCs were applied to 4 weeks old mice. The proportion of human leukocyte markers was analyzed by flow cytometry.

Results

As a result of HSC hu-mice analysis, the transplantation efficacy was enhanced that hCD45 of STKO exceeded 50% at 8 weeks after HSC transplantation, which was more than 50% greater than that of SID. The engraftment of hCD45 and hCD3 in STKO was significantly improved compared to SID until the last measurement and GvHD symptoms were also reduced in STKO. Similarly, the PBMC hu-mice studies showed that human leukocyte transplantation efficacy and GvHD symptoms were improved in STKO. Especially, in the study applying low-dose PBMC to young mice, long-term studies were possible with minimal clinical signs despite the significant human leukocyte engraftment.

Conclusions

In the HSC hu-mice and the PBMC hu-mice analyses, the transplantation and the differentiation of immune cells were significant, and GvHD symptoms were alleviated in STKO mice. To relieve GvHD symptoms, we applied low-dose PBMC to 4 week old mice and observed minimal health problems in STKO with improved human immune cell reconstitution. In conclusion, STKO mice is expected to be a novel and useful platform for various immune studies, considering enhanced transplantation efficiency with the stable health of the animal.

Legal entity responsible for the study

The animal study was approved and monitored by the Institutional Animal Care and Use Committee of Asan Institute for Life Sciences (Seoul, Korea, IACUC No. 2022-12-123).

Funding

National Research Foundation of Korea grant (NRF-2020R1C1C1014653).

Disclosure

All authors have declared no conflicts of interest.