Animal models are vital to the study of transfusion and development of new blood products. Post-transfusion recovery of human blood components can be studied in mice, however, there is a need to identify strains that can best tolerate xenogeneic transfusions, as well as to optimize such protocols. Specifically, the importance of using immunodeficient mice, such as NOD.Cg- Prkdc scid Il2rg tm1Wjl /SzJ (NSG) mice, to study human transfusion has been questioned. In this study, strains of wild-type and NSG mice were compared as hosts for human transfusions with outcomes quantified by flow cytometric analyses of CD235a + erythrocytes, CD45 + leukocytes, and CD41 + CD42b + platelets. Complete blood counts were evaluated as well as serum cytokines by multiplexing methods. Circulating human blood cells were maintained better in NSG than in wild-type mice. Lethargy and hemoglobinuria were observed in the first hours in wild-type mice along with increased pro-inflammatory cytokines/chemokines such as monocyte chemoattractant protein-1, tumor necrosis factor α, keratinocyte-derived chemokine (KC or CXCL1), and interleukin-6, whereas NSG mice were less severely affected. Whole blood transfusion resulted in rapid sequestration and then release of human cells back into the circulation within several hours. This rebound effect diminished when only erythrocytes were transfused. Nonetheless, human erythrocytes were found in excess of mouse erythrocytes in the liver and lungs and had a shorter half-life in circulation. Variables affecting the outcomes of transfused erythrocytes were cell dose and mouse weight; recipient sex did not affect outcomes. The sensitivity and utility of this xenogeneic model were shown by measuring the effects of erythrocyte damage due to exposure to the oxidizer diamide on post-transfusion recovery. Overall, immunodeficient mice are superior models for xenotransfusion as they maintain improved post-transfusion recovery with negligible immune-associated side effects.

The cryopreservation of hematopoietic cells using dimethyl sulfoxide (DMSO) and serum is a common procedure used in transplantation. However, DMSO has clinical and biological side effects due to its toxicity, and serum introduces variation and safety risks. Inspired by natural antifreeze proteins, a novel class of ice-interactive cryoprotectants was developed. The corresponding DMSO-, protein-, and serum-free cryopreservation media candidates were screened through a series of biological assays using human cell lines, peripheral blood cells, and bone marrow cells. XT-Thrive-A and XT-Thrive-B were identified as lead candidates to rival cryopreservation with 10% DMSO in serum based on post-thaw cell survival and short-term proliferation assays. The effectiveness of the novel cryopreservation media in freezing hematopoietic stem cells from human whole bone marrow was assessed by extreme limiting dilution analysis in immunodeficient mice. Stem cell frequencies were measured 12 weeks after transplant based on bone marrow engraftment of erythroid, myeloid, B-lymphoid, and CD34+ progenitors measured by flow cytometry. The recovered numbers of cryopreserved stem cells were similar among XT-Thrive A, XT-Thrive B, and DMSO with serum groups. These findings show that cryoprotectants developed through biomimicry of natural antifreeze proteins offers a substitute for DMSO-based media for the cryopreservation of hematopoietic stem cells.

Background aims: The cryopreservation of hematopoietic stem cells (HSCs) in dimethyl sulfoxide (DMSO) is used widely, but DMSO toxicity in transplant patients and the effects of DMSO on the normal function of cryopreserved cells are concerns. To address these issues, in vitro and clinical studies have explored using reduced concentrations of DMSO for cryopreservation. However, the effect of reducing DMSO concentration on the efficient cryopreservation of HSCs has not been directly measured. Methods: Cryopreservation of human bone marrow using 10%, 7.5% and 5% DMSO concentrations was examined. Cell counting, flow cytometry and colony assays were used to analyze different cell populations. The recovery of stem cells was enumerated using extreme limiting dilution analysis of long-term multi-lineage engraftment in immunodeficient mice. Four different methods of analyzing human engraftment were compared to ascertain stem cell engraftment: (i) engraftment of CD33 + myeloid, CD19 + B-lymphoid, CD235a + erythroid and CD34 + progenitors; (ii) engraftment of the same four populations plus CD41 + CD42b + platelets; (iii) engraftment of CD34 ++ CD133 + cells; and (iv) engraftment of CD34 ++ CD38 À cells. Results: Hematopoietic colony-forming, CD34 ++/+ , CD34 ++ CD133 + and CD34 ++ CD38 À cells were as well preserved with 5% DMSO as they were with the higher concentrations tested. The estimates of stem cell frequencies made in the xenogeneic transplant model did not show any significant detrimental effect of using lower concentrations of DMSO. Comparison of the different methods of gauging stem cell engraftment in mice led to different estimates of stem cell numbers, but overall, all measures found that reduced concentrations of DMSO supported the cryopreservation of HSCs. Conclusion: Cryopreservation of HSCs in DMSO concentrations as low as 5% is effective.

The advent of antiretroviral therapy (ART) has dramatically reduced the morbidity and mortality for human immunodeficiency virus (HIV)-infected individuals with access to healthcare in resource-rich countries. However, despite years of potent therapy, eradication of infection is not achieved due to the persistence of HIV latently infected cells during treatment. 1 Accumulating evidence suggest that "non-AIDS" cardiovascular, renal, and hepatic diseases are amplified by HIV infection, and the immune system may exhibit premature senescence even among patients with complete viral