Vitalant Research Institute San Francisco, California, United States
Background/Case Studies: Humanized mouse models are vital for preclinical research, but poor survival of human blood cells limits their effectiveness. Human AB serum (HABS) has been shown to reduce clearance of human cells in immunodeficient mice. To explore this further, we assessed the in vivo and in vitro effects of HABS on human red blood cells (hRBCs) survival. Using transfused NSG mice and macrophage phagocytosis assays, we found that HABS significantly prolonged hRBC circulation and reduced phagocytic uptake, highlighting its potential to improve human cell modeling in preclinical studies.
Study
Design/Methods: Two cohorts of 8–10-week-old NSG mice were used to evaluate the survival of hRBCs). hRBCs from healthy donors were transfused into mice, and blood samples were analyzed for hematocrit and hRBCs counts. In the first cohort, mice received intraperitoneal (IP) injections of human, mouse, or fetal bovine serum (FBS). The second cohort assessed the effects of HABS titration and route of administration (intravenous [IV] vs. IP) on hRBCs survival. Blood was collected at multiple time points post-transfusion and analyzed by flow cytometry. To investigate potential mechanisms, in vitro phagocytosis assays were conducted using bone marrow-derived macrophages from BALB/c and NSG mice. CFSE-labeled, opsonized hRBCs were co-cultured with BMDMs in the presence or absence of HABS. Phagocytosis was quantified at 10, 20, and 30 minutes, with 20 minutes providing optimal resolution. Statistical analyses were performed in Prism, with significance defined as P ≤ 0.05.
Results/Findings: Transfusion of hRBCs in HABS-treated NSG mice significantly improved cell survival over 3 hours compared to controls. The area under the curve (AUC) for hRBCs with PBS was 8,169.0 ± 416.5, while HABS-treated mice reached 14,192.0 ± 1,675. Among tested sera, only HABS improved survival (AUC: 13,293.0 ± 1,777); NSG and BALB/c serum, as well as fetal bovine serum, had no effect. A single HABS dose enabled detection of viable hRBCs up to 12 hours, with no added benefit from repeated doses. IV transfusion of hRBCs suspended in 300–400 μL of HABS yielded the best survival, with IV or combined IV plus IP administration outperforming IP injection alone. In vitro phagocytosis assays demonstrated that HABS significantly reduced uptake of hRBCs by both NSG and BALB/c macrophages, suggesting a mechanism by which HABS inhibits murine innate immune clearance of human cells.
Conclusions: These findings demonstrate that HABS significantly improves the in vivo survival of transfused hRBCs in NSG mice. A single HABS dose enables detection of circulating hRBCs for up to 12 hours post-transfusion. In vitro data support the hypothesis that HABS inhibits macrophage-mediated clearance of hRBCs, contributing to enhanced survival. Future studies are needed to identify the active HABS components responsible for this protective effect.
