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The role of neutrophils in solid tumor metastasis remains largely controversial. In preclinical models of solid tumors, both pro-metastatic and anti-metastatic effects of neutrophils have been reported. In this study, using mouse models of breast cancer, we demonstrate that the metastasis-modulating effects of neutrophils are dictated by the status of host natural killer (NK) cells. In NK cell-deficient mice, granulocyte colony-stimulating factor-expanded neutrophils show an inhibitory effect on the metastatic colonization of breast tumor cells in the lung. In contrast, in NK cell-competent mice, neutrophils facilitate metastatic colonization in the same tumor models. In an ex vivo neutrophil-NK cell-tumor cell tri-cell co-culture system, neutrophils are shown to potentially suppress the tumoricidal activity of NK cells, while neutrophils themselves are tumoricidal. Intriguingly, these two modulatory effects by neutrophils are both mediated by reactive oxygen species. Collectively, the absence or presence of NK cells, governs the net tumor-modulatory effects of neutrophils. The role of neutrophils in the regulation of tumour growth and metastasis remains controversial. Here, the authors demonstrate that neutrophils, by exerting inhibitory effects on cytotoxic NK cells, show a net pro-metastatic effect in immune-competent mice, while they are tumoricidal and anti-metastatic in NK cell-deficient hosts.

Key Points There is a growing need for animal models to carry out in vivo studies of human biological systems without putting individuals at risk. Severely immunodeficient mice engrafted with human cells and tissues, known as 'humanized' mice, facilitate progress in studies of human haematopoiesis, immunity, gene therapy, infectious diseases, cancer and regenerative medicine. Advances in the generation of humanized mice have depended on a systematic progression of genetic modifications in immunodeficient mouse hosts and on improvements in engraftment techniques. Mice homozygous for the severe combined immunodeficiency ( scid ) gene mutation or for targeted mutations at the recombination-activating gene 1 ( Rag1 ) or Rag2 loci, accompanied by a targeted mutation at the interleukin-2 receptor γ-chain ( Il2rg ) locus, support greatly increased engraftment and function of human haematopoietic stem cells (HSCs) and peripheral-blood mononuclear cells (PBMCs) compared with previous immunodeficient mouse models. The development of immunodeficient mice that are humanized by engraftment of human lymphoid tissues, HSCs and PBMCs provides an opportunity to carry out translational research on human immunity and autoimmune diseases. These models are also being used to study the biology of human pathogens responsible for AIDS and several other infectious diseases. Humanized mice are being increasingly used as hosts for human malignant cells in studies of carcinogenesis, tumour metastasis and cancer therapy. The phenotypic and functional characterization of human tumour stem cells is also being advanced through the study of humanized mice. The potential for new advances in our understanding of human immunology and other areas of human biology that is supported by studies in humanized mice remains promising. Additional genetic and technological modifications will accelerate progress towards the development of a functional human immune system in mice. The development of humanized mice over the past few decades has enabled the examination of human haematopoiesis, immunity to infectious diseases, cancer and autoantibodies in mice. But are these mice the key to translational research or is more work required? The culmination of decades of research on humanized mice is leading to advances in our understanding of human haematopoiesis, innate and adaptive immunity, autoimmunity, infectious diseases, cancer biology and regenerative medicine. In this Review, we discuss the development of these new generations of humanized mice, how they will facilitate translational research in several biomedical disciplines and approaches to overcome the remaining limitations of these models.

Immunodeficient mice engrafted with human cells and tissues have provided an exciting alternative to in vitro studies with human tissues and nonhuman primates for the study of human immunobiology. A major breakthrough in the early 2000s was the introduction of a targeted mutation in the interleukin 2 (IL-2) receptor common gamma chain (IL2rgn null ) into mice that were already deficient in T and B cells. Among other immune defects, natural killer (NK) cells are disrupted in these mice, permitting efficient engraftment with human hematopoietic cells that generate a functional human immune system. These humanized mouse models are becoming increasingly important for preclinical studies of human immunity, hematopoiesis, tissue regeneration, cancer, and infectious diseases. In particular, humanized mice have enabled studies of the pathogenesis of human-specific pathogens, including human immunodeficiency virus type 1, Epstein Barr virus, and Salmonella typhi. However, there are a number of limitations in the currently available humanized mouse models. Investigators are continuing to identify molecular mechanisms underlying the remaining defects in the engrafted human immune system and are generating \"next generation\" models to overcome these final deficiencies. This article provides an overview of some of the emerging models of humanized mice, their use in the study of infectious diseases, and some of the remaining limitations that are currently being addressed.

Dendritic cells can capture and transfer retroviruses in vitro across synaptic cell-cell contacts to uninfected cells, a process called trans-infection. Whether trans-infection contributes to retroviral spread in vivo remains unknown. Here, we visualize how retroviruses disseminate in secondary lymphoid tissues of living mice. We demonstrate that murine leukemia virus (MLV) and human immunodeficiency virus (HIV) are first captured by sinus-lining macrophages. CD169/Siglec-1, an I-type lectin that recognizes gangliosides, captures the virus. MLV-laden macrophages then form long-lived synaptic contacts to trans-infect B-1 cells. Infected B-1 cells subsequently migrate into the lymph node to spread the infection through virological synapses. Robust infection in lymph nodes and spleen requires CD169, suggesting that a combination of fluid-based movement followed by CD169-dependent trans-infection can contribute to viral spread.

In acute myeloid leukemia, a sub-population of quiescent cancer cells, called leukemia stem cells, is thought to be responsible for chemotherapy resistance and eventual recurrence of the disease. Saito et al . show that treatment with granulocyte colony-stimulating factor can overcome resistance to standard therapy by inducing cell cycle entry of the leukemia stem cells. Cancer stem cells have been proposed to be important for initiation, maintenance and recurrence of various malignancies, including acute myeloid leukemia (AML) 1 , 2 , 3 . We have previously reported 4 that CD34 + CD38 − human primary AML stem cells residing in the endosteal region of the bone marrow are relatively chemotherapy resistant. Using a NOD/SCID/IL2rγ null mouse model of human AML, we now show that the AML stem cells in the endosteal region are cell cycle quiescent and that these stem cells can be induced to enter the cell cycle by treatment with granulocyte colony-stimulating factor (G-CSF). In combination with cell cycle-dependent chemotherapy, G-CSF treatment significantly enhances induction of apoptosis and elimination of human primary AML stem cells in vivo . The combination therapy leads to significantly increased survival of secondary recipients after transplantation of leukemia cells compared with chemotherapy alone.

Sphingolipids, lipids with a common sphingoid base (also termed long chain base) backbone, play essential cellular structural and signaling functions. Alterations of sphingolipid levels have been implicated in many diseases, including neurodegenerative disorders. However, it remains largely unclear whether sphingolipid changes in these diseases are pathological events or homeostatic responses. Furthermore, how changes in sphingolipid homeostasis shape the progression of aging and neurodegeneration remains to be clarified. We identified two mouse strains, flincher (fln) and toppler (to), with spontaneous recessive mutations that cause cerebellar ataxia and Purkinje cell degeneration. Positional cloning demonstrated that these mutations reside in the Lass1 gene. Lass1 encodes (dihydro)ceramide synthase 1 (CerS1), which is highly expressed in neurons. Both fln and to mutations caused complete loss of CerS1 catalytic activity, which resulted in a reduction in sphingolipid biosynthesis in the brain and dramatic changes in steady-state levels of sphingolipids and sphingoid bases. In addition to Purkinje cell death, deficiency of CerS1 function also induced accumulation of lipofuscin with ubiquitylated proteins in many brain regions. Our results demonstrate clearly that ceramide biosynthesis deficiency can cause neurodegeneration and suggest a novel mechanism of lipofuscin formation, a common phenomenon that occurs during normal aging and in some neurodegenerative diseases.

Salmonella enterica serotype Typhimurium is a food-borne pathogen that also selectively grows in tumours and functionally decreases P-glycoprotein (P-gp), a multidrug resistance transporter. Here we report that the Salmonella type III secretion effector, SipA, is responsible for P-gp modulation through a pathway involving caspase-3. Mimicking the ability of Salmonella to reverse multidrug resistance, we constructed a gold nanoparticle system packaged with a SipA corona, and found this bacterial mimic not only accumulates in tumours but also reduces P-gp at a SipA dose significantly lower than free SipA. Moreover, the Salmonella nanoparticle mimic suppresses tumour growth with a concomitant reduction in P-gp when used with an existing chemotherapeutic drug (that is, doxorubicin). On the basis of our finding that the SipA Salmonella effector is fundamental for functionally decreasing P-gp, we engineered a nanoparticle mimic that both overcomes multidrug resistance in cancer cells and increases tumour sensitivity to conventional chemotherapeutics. S. Typhimurium can grow selectively on tumours and decreases the cellular levels of the multidrug resistance transporter Pgp. Here, the authors reveal the SipA-dependent mechanism of Pgp down-regulation and produce a SipA-based gold nanoparticle that increases sensitivity to the anticancer drug doxorubicin.

Background Gene disruption in mouse embryonic stem cells or zygotes is a conventional genetics approach to identify gene function in vivo. However, because different gene disruption strategies use different mechanisms to disrupt genes, the strategies can result in diverse phenotypes in the resulting mouse model. To determine whether different gene disruption strategies affect the phenotype of resulting mutant mice, we characterized Rhbdf1 mouse mutant strains generated by three commonly used strategies—definitive-null, targeted knockout (KO)-first, and CRISPR/Cas9. Results We find that Rhbdf1 responds differently to distinct KO strategies, for example, by skipping exons and reinitiating translation to potentially yield gain-of-function alleles rather than the expected null or severe hypomorphic alleles. Our analysis also revealed that at least 4% of mice generated using the KO-first strategy show conflicting phenotypes. Conclusions Exon skipping is a widespread phenomenon occurring across the genome. These findings have significant implications for the application of genome editing in both basic research and clinical practice.

Type 2 diabetes is characterized by insulin resistance, hyperglycemia, and progressive β cell dysfunction. Excess glucose and lipid impair β cell function in islet cell lines, cultured rodent and human islets, and in vivo rodent models. Here, we examined the mechanistic consequences of glucotoxic and lipotoxic conditions on human islets in vivo and developed and/or used 3 complementary models that allowed comparison of the effects of hyperglycemic and/or insulin-resistant metabolic stress conditions on human and mouse islets, which responded quite differently to these challenges. Hyperglycemia and/or insulin resistance impaired insulin secretion only from human islets in vivo. In human grafts, chronic insulin resistance decreased antioxidant enzyme expression and increased superoxide and amyloid formation. In human islet grafts, expression of transcription factors NKX6.1 and MAFB was decreased by chronic insulin resistance, but only MAFB decreased under chronic hyperglycemia. Knockdown of NKX6.1 or MAFB expression in a human β cell line recapitulated the insulin secretion defect seen in vivo. Contrary to rodent islet studies, neither insulin resistance nor hyperglycemia led to human β cell proliferation or apoptosis. These results demonstrate profound differences in how excess glucose or lipid influence mouse and human insulin secretion and β cell activity and show that reduced expression of key islet-enriched transcription factors is an important mediator of glucotoxicity and lipotoxicity.

Acute myelogenous leukemia (AML) is the most common adult leukemia, characterized by the clonal expansion of immature myeloblasts initiating from rare leukemic stem (LS) cells 1 , 2 , 3 . To understand the functional properties of human LS cells, we developed a primary human AML xenotransplantation model using newborn nonobese diabetic/severe combined immunodeficient/interleukin (NOD/SCID/IL)2rγ null mice carrying a complete null mutation of the cytokine γc upon the SCID background 4 . Using this model, we demonstrated that LS cells exclusively recapitulate AML and retain self-renewal capacity in vivo . They home to and engraft within the osteoblast-rich area of the bone marrow, where AML cells are protected from chemotherapy-induced apoptosis. Quiescence of human LS cells may be a mechanism underlying resistance to cell cycle–dependent cytotoxic therapy. Global transcriptional profiling identified LS cell–specific transcripts that are stable through serial transplantation. These results indicate the potential utility of this AML xenograft model in the development of novel therapeutic strategies targeted at LS cells.