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Expression of human cytokines dramatically improves reconstitution of specific human-blood lineage cells in humanized mice.

ABSTRACT: Adoptive transfer of human hematopoietic stem cells (HSCs) into mice lacking T, B and natural killer (NK) cells leads to development of human-blood lineage cells in the recipient mice (humanized mice). Although human B cell reconstitution is robust and T cell reconstitution is reasonable in the recipient mice, reconstitution of NK cells and myeloid cells is generally poor or undetectable. Here, we show that the poor reconstitution is mainly the result of a deficiency of appropriate human cytokines that are necessary for the development and maintenance of these cell lineages. When plasmid DNA encoding human IL-15 and Flt-3/Flk-2 ligand were delivered into humanized mice by hydrodynamic tail-vein injection, the expression of the human cytokine lasted for 2 to 3 weeks and elevated levels of NK cells were induced for more than a month. The cytokine-induced NK cells expressed both activation and inhibitory receptors, killed target cells in vitro, and responded robustly to a virus infection in vivo. Similarly, expression of human GM-CSF and IL-4, macrophage colony stimulating factor, or erythropoietin and IL-3 resulted in significantly enhanced reconstitution of dendritic cells, monocytes/macrophages, or erythrocytes, respectively. Thus, human cytokine gene expression by hydrodynamic delivery is a simple and efficient method to improve reconstitution of specific human-blood cell lineages in humanized mice, providing an important tool for studying human immune responses and disease progression in a small animal model.

SUBMITTER: Chen Q

PROVIDER: S-EPMC2789167 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Immunodeficient mouse-human chimeras provide a powerful approach to study host specific pathogens like Plasmodium (P.) falciparum that causes human malaria. Existing mouse models of P. falciparum infection require repeated injections of human red blood cells (RBCs). In addition, clodronate lipsomes and anti-neutrophil antibodies are injected to suppress the clearance of human RBCs by the residual immune system of the immunodeficient mice. Engraftment of NOD-scid Il2rg-/- mice with human hematopoietic stem cells leads to reconstitution of human immune cells. Although human B cell reconstitution is robust and T cell reconstitution is reasonable in the recipient mice, human RBC reconstitution is generally poor or undetectable. The poor reconstitution is mainly the result of a deficiency of appropriate human cytokines that are necessary for the development and maintenance of these cell lineages. Delivery of plasmid DNA encoding human erythropoietin and interleukin-3 into humanized mice by hydrodynamic tail-vein injection resulted in significantly enhanced reconstitution of erythrocytes. With this improved humanized mouse, here we show that P. falciparum infects de novo generated human RBCs, develops into schizonts and causes successive reinvasion. We also show that different parasite strains exhibit variation in their ability to infect these humanized mice. Parasites could be detected by nested PCR in the blood samples of humanized mice infected with P. falciparum K1 and HB3 strains for 3 cycles, whereas in other strains such as 3D7, DD2, 7G8, FCR3 and W2mef parasites could only be detected for 1 cycle. In vivo adaptation of K1 strain further improves the infection efficiency and parasites can be detected by microscopy for 3 cycles. The parasitemia ranges between 0.13 and 0.25% at the first cycle of infection, falls between 0.08 and 0.15% at the second cycle, and drops to barely detectable levels at the third cycle of infection. Compared to existing mouse models, our model generates human RBCs de novo and does not require the treatment of mice with immunomodulators.

Project description:Mouse is the most widely used animal model in biomedical research, but it remains unknown what causes the large number of differentially regulated genes between human and mouse livers identified in recent years. In this report, we aim to determine whether these divergent gene regulations are primarily caused by environmental factors or some of them are the result of cell-autonomous differences in gene regulation in human and mouse liver cells. The latter scenario would suggest that many human genes are subject to human-specific regulation and can only be adequately studied in a human or humanized system. To understand the similarity and divergence of gene regulation between human and mouse livers, we performed stepwise comparative analyses in human, mouse, and humanized livers with increased stringency to gradually remove the impact of factors external to liver cells, and used bioinformatics approaches to retrieve gene networks to ascertain the regulated biological processes. We first compared liver gene regulation by fatty liver disease in human and mouse under the condition where the impact of genetic and gender biases was minimized, and identified over 50% of all commonly regulated genes, that exhibit opposite regulation by fatty liver disease in human and mouse. We subsequently performed more stringent comparisons when a single specific transcriptional or post-transcriptional event was modulated in vitro or vivo or in liver-specific humanized mice in which human and mouse hepatocytes colocalize and share a common circulation. Intriguingly and strikingly, the pattern of a high percentage of oppositely regulated genes persists under well-matched conditions, even in the liver of the humanized mouse model, which represents the most closely matched in vivo condition for human and mouse liver cells that is experimentally achievable. Gene network analyses further corroborated the results of oppositely regulated genes and revealed substantial differences in regulated biological processes in human and mouse cells. We also identified a list of regulated lncRNAs that exhibit very limited conservation and could contribute to these differential gene regulations. Our data support that cell-autonomous differences in gene regulation might contribute substantially to the divergent gene regulation between human and mouse livers and there are a significant number of biological processes that are subject to human-specific regulation and need to be carefully considered in the process of mouse to human translation.

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Expression of human cytokines dramatically improves reconstitution of specific human-blood lineage cells in humanized mice.

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