Type I IFNs drive hematopoietic stem and progenitor cell collapse via impaired proliferation and increased RIPK1-dependent cell death during shock-like ehrlichial infection.

Type I interferons (IFNα/ß) regulate diverse aspects of host defense, but their impact on hematopoietic stem and progenitor cells (HSC/HSPCs) during infection remains unclear. Hematologic impairment can occur in severe infections, thus we sought to investigate the impact of type I IFNs on hematopoiesis in a tick-borne infection with a virulent ehrlichial pathogen that causes shock-like disease. During infection, IFNα/ß induced severe bone marrow (BM) loss, blunted infection-induced emergency myelopoiesis, and reduced phenotypic HSPCs and HSCs. In the absence of type I IFN signaling, BM and splenic hematopoiesis were increased, and HSCs derived from Ifnar1-deficient mice were functionally superior in competitive BM transplants. Type I IFNs impaired hematopoiesis during infection by both limiting HSC/HSPC proliferation and increasing HSPC death. Using mixed BM chimeras we determined that type I IFNs restricted proliferation indirectly, whereas HSPC death occurred via direct IFNαR -mediated signaling. IFNαR-dependent signals resulted in reduced caspase 8 expression and activity, and reduced cleavage of RIPK1 and RIPK3, relative to Ifnar1-deficient mice. RIPK1 antagonism with Necrostatin-1s rescued HSPC and HSC numbers during infection. Early antibiotic treatment is required for mouse survival, however antibiotic-treated survivors had severely reduced HSPCs and HSCs. Combination therapy with antibiotics and Necrostatin-1s improved HSPC and HSC numbers in surviving mice, compared to antibiotic treatment alone. We reveal two mechanisms whereby type I IFNs drive hematopoietic collapse during severe infection: direct sensitization of HSPCs to undergo cell death and enhanced HSC quiescence. Our studies reveal a strategy to ameliorate the type I IFN-dependent loss of HSCs and HSPCs during infection, which may be relevant to other infections wherein type I IFNs cause hematopoietic dysfunction.

An Immature Myeloid/Myeloid-Suppressor Cell Response Associated with Necrotizing Inflammation Mediates Lethal Pulmonary Tularemia.

Inhalation of Francisella tularensis (Ft) causes acute and fatal pneumonia. The lung cytokine milieu favors exponential Ft replication, but the mechanisms underlying acute pathogenesis and death remain unknown. Evaluation of the sequential and systemic host immune response in pulmonary tularemia reveals that in contrast to overwhelming bacterial burden or cytokine production, an overt innate cellular response to Ft drives tissue pathology and host mortality. Lethal infection with Ft elicits medullary and extra-medullary myelopoiesis supporting recruitment of large numbers of immature myeloid cells and MDSC to the lungs. These cells fail to mature and die, leading to subsequent necrotic lung damage, loss of pulmonary function, and host death that is partially dependent upon immature Ly6G+ cells. Acceleration of this process may account for the rapid lethality seen with Ft SchuS4. In contrast, during sub-lethal infection with Ft LVS the pulmonary cellular response is characterized by a predominance of mature neutrophils and monocytes required for protection, suggesting a required threshold for lethal bacterial infection. Further, eliciting a mature phagocyte response provides transient, but dramatic, innate protection against Ft SchuS4. This study reveals that the nature of the myeloid cell response may be the primary determinant of host mortality versus survival following Francisella infection.

Hematopoietic stem cell loss and hematopoietic failure in severe aplastic anemia is driven by macrophages and aberrant podoplanin expression.

Severe aplastic anemia (SAA) results from profound hematopoietic stem cell loss. T cells and interferon gamma (IFNγ) have long been associated with SAA, yet the underlying mechanisms driving hematopoietic stem cell loss remain unknown. Using a mouse model of SAA, we demonstrate that IFNγ-dependent hematopoietic stem cell loss required macrophages. IFNγ was necessary for bone marrow macrophage persistence, despite loss of other myeloid cells and hematopoietic stem cells. Depleting macrophages or abrogating IFNγ signaling specifically in macrophages did not impair T-cell activation or IFNγ production in the bone marrow but rescued hematopoietic stem cells and reduced mortality. Thus, macrophages are not required for induction of IFNγ in SAA and rather act as sensors of IFNγ. Macrophage depletion rescued thrombocytopenia, increased bone marrow megakaryocytes, preserved platelet-primed stem cells, and increased the platelet-repopulating capacity of transplanted hematopoietic stem cells. In addition to the hematopoietic effects, SAA induced loss of non-hematopoietic stromal populations, including podoplanin-positive stromal cells. However, a subset of podoplanin-positive macrophages was increased during disease, and blockade of podoplanin in mice was sufficient to rescue disease. Our data further our understanding of disease pathogenesis, demonstrating a novel role for macrophages as sensors of IFNγ, thus illustrating an important role for the microenvironment in the pathogenesis of SAA.

Macrophage-Lineage Cells Negatively Regulate the Hematopoietic Stem Cell Pool in Response to Interferon Gamma at Steady State and During Infection.

Bone marrow (BM) resident macrophages (Mϕs) regulate hematopoietic stem cell (HSC) mobilization; however, their impact on HSC function has not been investigated. We demonstrate that depletion of BM resident Mϕs increases HSC proliferation as well as the pool of quiescent HSCs. At the same time, during bacterial infection where BM resident Mϕs are selectively increased we observe a decrease in HSC numbers. Moreover, strategies that deplete or reduce Mϕs during infection prevent HSC loss and rescue HSC function. We previously found that the transient loss of HSCs during infection is interferon-gamma (IFNγ)-dependent. We now demonstrate that IFNγ signaling specifically in Mϕs is critical for both the diminished HSC pool and maintenance of BM resident Mϕs during infection. In addition to the IFNγ-dependent loss of BM HSC and progenitor cells (HSPCs) during infection, IFNγ reduced circulating HSPC numbers. Importantly, under infection conditions AMD3100 or G-CSF-induced stem cell mobilization was impaired. Taken together, our data show that IFNγ acts on Mϕs, which are a negative regulator of the HSC pool, to drive the loss in BM and peripheral HSCs during infection. Our findings demonstrate that modulating BM resident Mϕ numbers can impact HSC function in vivo, which may be therapeutically useful for hematologic conditions and refinement of HSC transplantation protocols.

MyD88 signaling in CD4 T cells promotes IFN-γ production and hematopoietic progenitor cell expansion in response to intracellular bacterial infection.

Hematopoietic stem and progenitor cell (HSPC) phenotype and function can change in response to infectious challenge. These changes can be mediated by cytokines, IFNs, and pathogen-associated molecules, via TLR, and are thought to promote tailored immune responses for particular pathogens. In this study, we investigated the signals that activate HSPCs during ehrlichiosis, a disease characterized by profound hematopoietic dysfunction in both humans and mice. In a mouse model of ehrlichiosis, we observed that infection-induced proliferation of bone marrow HSPCs was dependent on IFN-γ signaling and was partially dependent on MyD88. However, MyD88 was not required in HSPCs for their expansion during infection, because similar frequencies of MyD88-deficient and wild-type HSPCs proliferated in mixed bone marrow chimeric mice. MyD88-deficient mice exhibited low serum and bone marrow concentration of IFN-γ compared with wild-type mice. We next identified CD4 T cells as the primary cells producing IFN-γ in the bone marrow and demonstrated a nonredundant role for CD4-derived IFN-γ in increased HSPCs. Using mixed bone marrow chimeric mice, we identified a requirement for MyD88 in CD4 T cells for increased T-bet expression, optimal IFN-γ production, and CD4 T cell proliferation. Our data demonstrate an essential role for CD4 T cells in mediating HSPC activation in response to bacterial infection and illustrate a novel role for MyD88 signaling in CD4 T cells in this process. These findings further support the idea that IFN-γ production is essential for HSPC activation and hematopoietic responses to infection.

Complicated vascular access port removals: incidence, antecedents and avoidance.

PURPOSE: Port removal is usually a straightforward procedure delegated to trainees. However, some port removals are complicated by central venous catheter (CVC) fragmentation, a challenge for even experienced surgeons. This study aimed to determine the incidence of, and risk factors for, complicated port removal in children. METHODS: A single-centre study assessed the outcome of removal for all paediatric ports inserted from 1996 to 2012. Data were recorded detailing patient, insertion, device and removal characteristics. Risk factors for complicated removals were scrutinised using Chi-square tests; p < 0.05 significant. RESULTS: Of 628 ports inserted from 1996 to 2012, 443 were subsequently removed at the same centre. 8/443 (1.8%) removals were complicated by CVC fragmentation, a median of 3.3 (2.4-3.9) years after insertion. Of complicated cases, 8/8 underwent formal neck dissection, 3/8 intravascular dissection, and 1/8 endovascular retrieval. 2/8 cases have retained intravascular CVC fragments. Risk factors for complication were CVC caliber <6Fr (p < 0.001) and use duration >2 years (p < 0.001). CONCLUSION: Greatest care and senior supervision should be ensured when removing ports with CVC caliber <6Fr and/or >2 years since insertion. However, complications also occur with larger CVCs or after shorter durations. Therefore, the key to avoiding complicated port removal may simply be: preparation, preparation, neck preparation.

Type I interferons promote severe disease in a mouse model of lethal ehrlichiosis.

Human monocytic ehrlichiosis (HME) is caused by a tick-borne obligate intracellular pathogen of the order Rickettsiales. HME disease can range from mild to a fatal, toxic shock-like syndrome, yet the mechanisms regulating pathogenesis are not well understood. We define a central role for type I interferons (alpha interferon [IFN-α] and IFN-ß) in severe disease in a mouse model of fatal ehrlichiosis caused by Ixodes ovatus Ehrlichia (IOE). IFN-α and IFN-ß were induced by IOE infection but not in response to a less virulent strain, Ehrlichia muris. The major sources of type I IFNs during IOE infection were plasmacytoid dendritic cells and monocytes. Mice lacking the receptor for type I IFNs (Ifnar deficient) or neutralization of IFN-α and IFN-ß resulted in a reduced bacterial burden. Ifnar-deficient mice exhibited significantly increased survival after IOE infection, relative to that of wild-type (WT) mice, that correlated with increased type II IFN (IFN-γ) production. Pathogen-specific antibody responses were also elevated in Ifnar-deficient mice, and this required IFN-γ. Remarkably, increased IFN-γ and IgM were not essential for protection in the absence of type I IFN signaling. The direct effect of type I IFNs on hematopoietic and nonhematopoietic cells was evaluated in bone marrow chimeric mice. We observed that chimeric mice containing Ifnar-deficient hematopoietic cells succumbed to infection early, whereas Ifnar-deficient mice containing WT hematopoietic cells exhibited increased survival, despite having a higher bacterial burden. These data demonstrate that IFN-α receptor signaling in nonhematopoietic cells is important for pathogenesis. Thus, type I IFNs are induced during a rickettsial infection in vivo and promote severe disease.

Genetic reversal of the globin switch concurrently modulates both fetal and sickle hemoglobin and reduces red cell sickling.

We previously reported initial clinical results of post-transcriptional gene silencing of BCL11A expression (NCT03282656) reversing the fetal to adult hemoglobin switch. A goal of this approach is to increase fetal hemoglobin (HbF) expression while coordinately reducing sickle hemoglobin (HbS) expression. The resulting combinatorial effect should prove effective in inhibiting HbS polymerization at lower physiologic oxygen values thereby mitigating disease complications. Here we report results of exploratory single-cell analysis of patients in which BCL11A is targeted molecularly and compare results with cells of patients treated with hydroxyurea (HU), the current standard of care. We use single-cell assays to assess HbF, HbS, oxygen saturation, and hemoglobin polymer content in RBCs for nine gene therapy trial subjects (BCLshmiR, median HbF% = 27.9) and compare them to 10 HU-treated subjects demonstrating high and comparable levels of HbF (HU High Responders, median HbF% = 27.0). All BCL11A patients achieved the primary endpoint for NCT03282656, which was defined by an absolute neutrophil count greater than or equal to 0.5 × 109 cells/L for three consecutive days, achieved within 7 weeks following infusion. Flow cytometric assessment of single-RBC HbF and HbS shows fewer RBCs with high HbS% that would be most susceptible to sickling in BCLshmiR vs. HU High Responders: median 42% of RBCs with HbS%>70% in BCLshmiR vs. 61% in HU High Responders (p = 0.004). BCLshmiR subjects also demonstrate more RBCs resistant to HbS polymerization at lower physiologic oxygen tension: median 32% vs. 25% in HU High Responders (p = 0.006). Gene therapy-induced BCL11A down-regulation reverses the fetal-to-adult hemoglobin switch and induces RBCs with higher HbF%, lower HbS%, and greater resistance to deoxygenation-induced polymerization in clinical trial subjects compared with a cohort of highly responsive hydroxyurea-treated subjects.

Systemic macrophage depletion inhibits Helicobacter bilis-induced proinflammatory cytokine-mediated typhlocolitis and impairs bacterial colonization dynamics in a BALB/c Rag2-/- mouse model of inflammatory bowel disease.

Helicobacter bilis, an enterohepatic helicobacter, is associated with chronic hepatitis in aged immunocompetent inbred mice and inflammatory bowel disease (IBD) in immunodeficient mice. To evaluate the role of macrophages in H. bilis-induced IBD, Rag2(-/-) BALB/c or wild-type (WT) BALB/c mice were either sham dosed or infected with H. bilis Missouri strain under specific-pathogen-free conditions, followed by an intravenous injection of a 0.2-ml suspension of liposomes coated with either phosphate-buffered saline (control) or clodronate (a macrophage depleting drug) at 15 weeks postinfection (wpi). At 16 wpi, the ceca of H. bilis-infected Rag2(-/-) mice treated with control liposomes had significantly higher histopathological lesional scores (for cumulative typhlitis index, inflammation, edema, epithelial defects, and hyperplasia) and higher counts of F4/80(+) macrophages and MPO(+) neutrophils compared to H. bilis-infected Rag2(-/-) mice treated with clodronate liposomes. In addition, cecal quantitative PCR analyses revealed a significant suppression in the expression of macrophage-related cytokine genes, namely, Tnfa, Il-1ß, Il-10, Cxcl1, and iNos, in the clodronate-treated H. bilis-infected Rag2(-/-) mice compared to the H. bilis-infected Rag2(-/-) control mice. Finally, cecal quantitative PCR analyses also revealed a significant reduction in bacterial colonization in the clodronate-treated Rag2(-/-) mice. Taken together, our results suggest that macrophages are critical inflammatory cellular mediators for promoting H. bilis-induced typhlocolitis in mice.

CCR5 maintains macrophages in the bone marrow and drives hematopoietic failure in a mouse model of severe aplastic anemia.

Severe aplastic anemia (SAA) is an acquired, T cell-driven bone marrow (BM) failure disease characterized by elevated interferon gamma (IFNγ), loss of hematopoietic stem cells (HSCs), and altered BM microenvironment, including dysfunctional macrophages (MΦs). T lymphocytes are therapeutic targets for treating SAA, however, the underlying mechanisms driving SAA development and how innate immune cells contribute to disease remain poorly understood. In a murine model of SAA, increased beta-chemokines correlated with disease and were partially dependent on IFNγ. IFNγ was required for increased expression of the chemokine receptor CCR5 on MΦs. CCR5 antagonism in murine SAA improved survival, correlating with increased platelets and significantly increased platelet-biased CD41hi HSCs. T cells are key drivers of disease, however, T cell-specific CCR5 expression and T cell-derived CCL5 were not necessary for disease. CCR5 antagonism reduced BM MΦs and diminished their expression of Tnf and Ccl5, correlating with reduced frequencies of IFNγ-secreting BM T cells. Mechanistically, CCR5 was intrinsically required for maintaining BM MΦs during SAA. Ccr5 expression was significantly increased in MΦs from aged mice and humans, relative to young counterparts. Our data identify CCR5 signaling as a key axis promoting the development of IFNγ-dependent BM failure, particularly relevant in aging where Ccr5 expression is elevated.

Restored Macrophage Function Ameliorates Disease Pathophysiology in a Mouse Model for IL10 Receptor-deficient Very Early Onset Inflammatory Bowel Disease.

BACKGROUND AND AIMS: Mutations in IL10 or the IL10 receptor lead to very early onset [VEO] inflammatory bowel disease [IBD], a life-threatening disease which is often unresponsive to conventional medication. Recent studies have demonstrated that defective IL-10 receptor signalling in innate immune cells is a key driver of severe intestinal inflammation in VEO-IBD. Specifically, IL10 unresponsiveness of macrophages, which govern the tight balance between pro- and anti-inflammatory responses in the intestinal system, plays a central role in the events leading to excessive inflammatory responses and the development of IBD. METHODS AND RESULTS: We here evaluated haematopoietic stem cell gene therapy in a VEO-IBD mouse model and demonstrated that the therapeutic response closely correlates with gene correction of the IL10 signalling pathway in intestinal macrophages. This finding prompted us to evaluate the therapeutic efficacy of macrophage transplantation in the Il10rb-/- VEO-IBD mouse model. A 6-week regimen employing a combination of depletion of endogenous hyperinflammatory macrophages followed by intraperitoneal administration of wild-type [WT] macrophages significantly reduced colitis symptoms. CONCLUSIONS: In summary, we show that the correction of the IL10 receptor defect in macrophages, either by genetic therapy or transfer of WT macrophages to the peritoneum, can ameliorate disease-related symptoms and potentially represent novel treatment approaches for VEO-IBD patients.

Major complications of jejunal feeding in children.

AIM OF THE STUDY: The aim of the study was to identify major gastrointestinal complications associated with direct jejunal feeding. We hypothesized that jejunal feeding may cause life-threatening surgical complications in a minority of patients. METHODS: All patients undergoing jejunal feeding between 1/2008 and 1/2018 at a pediatric surgical unit were identified retrospectively. Data sought from records included demographics, comorbidities, indications, feeding strategies, adverse events, and follow-up. Major surgical complications were defined by Clavien-Dindo grade ≥ IIIb and involving the GI tract (excluding changes of jejunal tube). MAIN RESULTS: 197 patients were identified (110 female). Median age (IQR) at initiation of jejunal feeding months was 5.6 (6-164) months. 122 were neurologically impaired. The most frequent indications were: GERD/gastroparesis (n = 114), prophylaxis/treatment of Superior Mesenteric Artery (SMA) syndrome (N.B. our center is a national spinal deformity unit) (n = 47), congenital anomalies of aerodigestive anatomy (n = 17), and malignancy (n = 7). 125 patients were managed with nasojejunal feeding alone: gastrojejunal tube (n = 51) and via Roux-en-Y jejunostomy (n = 21). There were 14 significant gastrointestinal complications (n = 11 grade > IIIb) identified among 12 patients, of whom 8 required bowel resections, and 2 died as a result: nonmechanical bowel ischemia (n = 7), intussusception (n = 4), and volvulus (n = 3). CONCLUSION: This series highlights the major complications of jejunal feeding, including a significant yet underreported risk of gut compromise. Patients undergoing jejunal feeding had a 6.1% risk of developing major surgical complications (of note, 3.6% developed bowel ischemia of unknown etiology). Susceptible children were comorbid, fragile, and neurologically impaired. These findings should influence parental discussions and informed consent before embarking upon jejunal feeding. LEVEL OF EVIDENCE: Level IV prognosis study.

Aged marrow macrophages expand platelet-biased hematopoietic stem cells via Interleukin1B.

The bone marrow microenvironment (BMME) contributes to the regulation of hematopoietic stem cell (HSC) function, though its role in age-associated lineage skewing is poorly understood. Here we show that dysfunction of aged marrow macrophages (Mφs) directs HSC platelet-bias. Mφs from the marrow of aged mice and humans exhibited an activated phenotype, with increased expression of inflammatory signals. Aged marrow Mφs also displayed decreased phagocytic function. Senescent neutrophils, typically cleared by marrow Mφs, were markedly increased in aged mice, consistent with functional defects in Mφ phagocytosis and efferocytosis. In aged mice, Interleukin 1B (IL1B) was elevated in the bone marrow and caspase 1 activity, which can process pro-IL1B, was increased in marrow Mφs and neutrophils. Mechanistically, IL1B signaling was necessary and sufficient to induce a platelet bias in HSCs. In young mice, depletion of phagocytic cell populations or loss of the efferocytic receptor Axl expanded platelet-biased HSCs. Our data support a model wherein increased inflammatory signals and decreased phagocytic function of aged marrow Mφs induce the acquisition of platelet bias in aged HSCs. This work highlights the instructive role of Mφs and IL1B in the age-associated lineage-skewing of HSCs, and reveals the therapeutic potential of their manipulation as antigeronic targets.

Macrophages: Key regulators of steady-state and demand-adapted hematopoiesis.

Hematopoietic stem cell (HSC) function is required for balanced blood production throughout life; it is thus essential to understand the mechanisms regulating this highly dynamic process. Bone marrow-resident macrophages (Mϕs) have recently emerged as an important component of the HSC niche, where they contribute to regulating HSC and progenitor cell (HSPC) mobilization and function. Here we review the role of macrophages (Mϕs) on immune cell production, HSPC pool size, and mobilization at steady state and under inflammatory conditions. Inflammation induces marked changes in hematopoiesis to restrict or promote generation of specific cell lineages, and this often has a negative impact on HSC function. Cytokines and growth factors induced during inflammation influence hematopoiesis by acting directly on HSPCs and/or by modulating niche cell function. We focus particular attention on the opposing effects of two key inflammatory proteins, interferon-γ and granulocyte-colony stimulating factor, in regulating bone marrow-resident macrophages (Mϕs) and HSPCs. Macrophages (Mϕs) are essential for tissue homeostasis, and here we highlight their emerging role as a central regulator of both steady-state and demand-adapted hematopoiesis.

Swallowed magnets and batteries: a dangerous but not unexpected attraction.

An 18-month-old boy was witnessed swallowing a cluster of five magnetic toy balls. He was coincidentally noted on plain x-rays to have also recently swallowed a watch battery and a small screw. Initial outpatient management with serial review and x-rays was unsuccessful, and delayed inpatient surgical care by 9 days. Although the child never manifested features of systemic or gastrointestinal upset, emergency laparotomy confirmed a resultant jejunocolic fistula. This case demonstrates how clinical assessment of children who have swallowed magnets separately from each other can be falsely reassuring, and highlights the potential dangers of outpatient management. We recommend children who have swallowed separately >1 magnetic objects (or >1 objects capable of magnetic attraction) be managed as inpatients with active observation and timely foreign body removal.

Helicobacter pylori infection does not promote hepatocellular cancer in a transgenic mouse model of hepatitis C virus pathogenesis.

Helicobacter pylori (H. pylori) and hepatitis C virus (HCV) infect millions of people and can induce cancer. We investigated if H. pylori infection promoted HCV-associated liver cancer. Helicobacter-free C3B6F1 wild-type (WT) and C3B6F1-Tg(Alb1-HCVN)35Sml (HT) male and female mice were orally inoculated with H. pylori SS1 or sterile media. Mice were euthanized at ~12 mo postinoculation and samples were collected for analyses. There were no significant differences in hepatocellular tumor promotion between WT and HT mice; however, HT female mice developed significantly larger livers with more hepatic steatosis than WT female mice. H. pylori did not colonize the liver nor promote hepatocellular tumors in WT or HT mice. In the stomach, H. pylori induced more corpus lesions in WT and HT female mice than in WT and HT male mice, respectively. The increased corpus pathology in WT and HT female mice was associated with decreased gastric H. pylori colonization, increased gastric and hepatic interferon gamma expression, and increased serum Th1 immune responses against H. pylori. HT male mice appeared to be protected from H. pylori-induced corpus lesions. Furthermore, during gastric H. pylori infection, HT male mice were protected from gastric antral lesions and hepatic steatosis relative to WT male mice and these effects were associated with increased serum TNF-α. Our findings indicate that H. pylori is a gastric pathogen that does not promote hepatocellular cancer and suggest that the HCV transgene is associated with amelioration of specific liver and gastric lesions observed during concurrent H. pylori infection in mice.

Colitis and colon cancer in WASP-deficient mice require helicobacter species.

BACKGROUND: Wiskott-Aldrich syndrome protein-deficient patients and mice are immunodeficient and can develop inflammatory bowel disease. The intestinal microbiome is critical to the development of colitis in most animal models, in which Helicobacter spp. have been implicated in disease pathogenesis. We sought to determine the role of Helicobacter spp. in colitis development in Wiskott-Aldrich syndrome protein-deficient (WKO) mice. METHODS: Feces from WKO mice raised under specific pathogen-free conditions were evaluated for the presence of Helicobacter spp., after which a subset of mice were rederived in Helicobacter spp.-free conditions. Helicobacter spp.-free WKO animals were subsequently infected with Helicobacter bilis. RESULTS: Helicobacter spp. were detected in feces from WKO mice. After rederivation in Helicobacter spp.-free conditions, WKO mice did not develop spontaneous colitis but were susceptible to radiation-induced colitis. Moreover, a T-cell transfer model of colitis dependent on Wiskott-Aldrich syndrome protein-deficient innate immune cells also required Helicobacter spp. colonization. Helicobacter bilis infection of rederived WKO mice led to typhlitis and colitis. Most notably, several H. bilis-infected animals developed dysplasia with 10% demonstrating colon carcinoma, which was not observed in uninfected controls. CONCLUSIONS: Spontaneous and T-cell transfer, but not radiation-induced, colitis in WKO mice is dependent on the presence of Helicobacter spp. Furthermore, H. bilis infection is sufficient to induce typhlocolitis and colon cancer in Helicobacter spp.-free WKO mice. This animal model of a human immunodeficiency with chronic colitis and increased risk of colon cancer parallels what is seen in human colitis and implicates specific microbial constituents in promoting immune dysregulation in the intestinal mucosa.

Prevalence of murine Helicobacter spp. Infection is reduced by restocking research colonies with Helicobacter-free mice.

Most academic research colonies of mice are endemically infected with enterohepatic Helicobacter spp. (EHS). We evaluated EHS prevalence in surveillance mice before and after a 10-y period of requiring that imported mice be free of EHS by embryo transfer rederivation or purchase from approved vendors. In 2009, composite fecal samples from CD1 surveillance mice representing colony health in 57 rooms located in 6 facilities were evaluated for EHS infection by using PCR assays. Fecal samples were screened with primers designed to detect all known EHS, and positive samples were further assayed by using primers specific for H. hepaticus, H. bilis, H. rodentium, and H. typhlonicus. Most EHS were detected in surveillance mice within the first month of dirty bedding exposure, with prevalence ranging from 0% to 64% as monoinfections or, more commonly, infections with multiple EHS. Compared with 1999 prevalence data, EHS remained endemic in colonies importing the lowest number of EHS-free mice. EHS were absent or the prevalence was greatly reduced in colonies receiving the highest percentage of EHS-free mice. This study demonstrates that the management decision to require exclusive importation of EHS-free mice reduced EHS prevalence on an institutional scale without intensive labor and expense associated with other techniques or interference with research objectives.

17ß-estradiol and tamoxifen prevent gastric cancer by modulating leukocyte recruitment and oncogenic pathways in Helicobacter pylori-infected INS-GAS male mice.

Helicobacter pylori infection promotes male predominant gastric adenocarcinoma in humans. Estrogens reduce gastric cancer risk and previous studies showed that prophylactic 17ß-estradiol (E2) in INS-GAS mice decreases H. pylori-induced carcinogenesis. We examined the effect of E2 and tamoxifen (TAM) on H. pylori-induced gastric cancer in male and female INS-GAS mice. After confirming robust gastric pathology at 16 weeks postinfection (WPI), mice were implanted with E2, TAM, both E2 and TAM, or placebo pellets for 12 weeks. At 28 WPI, gastric histopathology, gene expression, and immune cell infiltration were evaluated and serum inflammatory cytokines measured. After treatment, no gastric cancer was observed in H. pylori-infected males receiving E2 and/or TAM, whereas 40% of infected untreated males developed gastric cancer. E2, TAM, and their combination significantly reduced gastric precancerous lesions in infected males compared with infected untreated males (P < 0.001, 0.01, and 0.01, respectively). However, TAM did not alter female pathology regardless of infection status. Differentially expressed genes from males treated with E2 or TAM (n = 363 and n = 144, Q < 0.05) associated highly with cancer and cellular movement, indicating overlapping pathways in the reduction of gastric lesions. E2 or TAM deregulated genes associated with metastasis (PLAUR and MMP10) and Wnt inhibition (FZD6 and SFRP2). Compared with controls, E2 decreased gastric mRNA (Q < 0.05) and serum levels (P < 0.05) of CXCL1, a neutrophil chemokine, leading to decreased neutrophil infiltration (P < 0.01). Prevention of H. pylori-induced gastric cancer by E2 and TAM may be mediated by estrogen signaling and is associated with decreased CXCL1, decreased neutrophil counts, and downregulation of oncogenic pathways.

Right-sided diaphragmatic hernia in infants after liver transplantation.

Liver transplantation is just as successful in infants as in older children, but more challenging. This relates to the low weight of the recipients and to their rapidly deteriorating clinical condition (malnutrition and end-stage liver disease) ( J Pediatr 1990;117:205-210; BMJ 1993;307:825-828; Ann Surg 1996;223:658-664; Transplantation 1997;64:242-248; J Pediatr Surg 1998;33:20-23). In addition, higher rates of diaphragmatic complications have been shown to significantly correlate with a younger age ( Transplantation 2002;73:228-232; Transpl Int 1998;11:281-283; Pediatr Transplant 2000;4:39-44), but diaphragmatic hernia has never been reported as a complication of liver transplantation. In this report, 2 patients who developed diaphragmatic hernia after liver transplantation are presented. The possible role of several contributing factors resulting in diaphragmatic hernia is discussed. These factors include (1) diaphragm thinness related to low weight and malnutrition, (2) direct trauma at operation (dissection and diathermy), (3) increased abdominal pressure after transplantation caused by the use of a slightly oversized liver graft, and (4) the medial positioning of the partial liver graft in the abdomen.