Vhl deletion in Dmp1 -expressing cells alters MEP metabolism and promotes stress erythropoiesis.

In recent years, general hypoxia-inducible factor (HIF)-prolyl hydroxylase (PHD) enzyme inhibitors have been developed for the treatment of anemia due to renal disease and osteoporosis. However, it remains a challenge to target the HIF signaling pathway without dysregulating the skeletal and hematopoietic system. Here, we examined the effects of Vhl deletion in bone by performing longitudinal analyses of Vhl cKO mice at 3, 6, 10, and 24 weeks of age, where at 10 and 24 weeks of age, high bone mass and splenomegaly are present. Using flow cytometry, we observed increased frequency (%) of CD71 lo TER119 hi FSC lo orthochromatophilic erythroblasts and reticulocytes in 10- and 24-week-old Vhl cKO bone marrow (BM), which correlated with elevated erythropoietin levels in the BM and increased number of red blood cells in circulation. The absolute numbers of myeloerythroid progenitors (MEPs) in the BM were significantly reduced at 24 weeks. Bulk RNA-Seq of the MEPs showed upregulation of Epas1 ( Hif1a) and Efnb2 ( Hif2a) in Vhl cKO MEPs, consistent with a response to hypoxia, and genes involved in erythrocyte development, actin filament organization, and response to glucose. Additionally, histological analysis of Vhl cKO spleens revealed red pulp hyperplasia and the presence of megakaryocytes, both of which are features of extramedullary hematopoiesis (EMH). EMH in the spleen was correlated with the presence of mature stress erythroid progenitors, suggesting that stress erythropoiesis is occurring to compensate for the BM microenvironmental irregularities. Our studies implicate that HIF-driven alterations in skeletal homeostasis can accelerate erythropoiesis. Key Points: • Dysregulation of HIF signaling in Dmp1+ bone cells induces stress erythropoiesis.• Skeletal homeostasis modulates erythropoiesis.

A novel bioabsorbable drug-eluting tracheal stent.

OBJECTIVES/HYPOTHESIS: Currently available silicone and metallic stents for tracheal stenosis are associated with problems of granulations, mucus trapping, and difficult removals. Our aim was to develop a novel bioabsorbable tracheal stent with mitomycin C (MMC) drug elution to circumvent such problems. STUDY DESIGN: A randomized animal study. METHODS: Twenty-five rabbits were randomly assigned into five test groups: 1) controls (without stent), 2) silicone tubular stents (commercially available currently); 3) bioabsorbable helical stents; 4) bioabsorbable tubular stents; and 5) bioabsorbable tubular stents with MMC. Weekly tracheal endoscopy to document granulation, mucus plugging, and extent of tracheal stenosis was performed for 12 weeks. One rabbit was euthanized every 3 weeks for histological analysis of the trachea. In vitro MMC-release profiles in conditions mimicking tracheal conditions were studied. RESULTS: The bioabsorbable tubular stents with 0.1 mg MMC drug elution performed the best, with the least mucus trapping and airway obstruction due to tracheal stenosis. Tracheal stenosis was most significant for the bioabsorbable helical stents, followed by the control group without stent, the group of bioabsorbable tubular stents, and then the silicone stents. After 12 weeks, tracheal stenosis for the bioabsorbable tubular stents with MMC was only half that of the silicone stents. CONCLUSIONS: This study reports on the development of a novel bioabsorbable tracheal stent with sustained MMC drug elution for preventing tracheal stenosis. Further studies are warranted to optimize stent design and drug dosage.

Sustained high levels of interleukin-6 contribute to the pathogenesis of enterovirus 71 in a neonate mouse model.

Enterovirus 71 (EV71) is the major causative agent of hand, foot, and mouth disease (HFMD) in young children and has been consistently associated with the most severe complications of the disease, including central nervous system inflammation and pulmonary edema. Increasing frequency and amplitude of EV71 outbreaks have raised awareness and concerns worldwide. Previous reports proposed that overwhelming virus replication combined with the induction of massive proinflammatory cytokines is responsible for the pathogenicity of EV71. Specifically, elevated interleukin-6 (IL-6) levels were observed consistently in patients and strongly correlated with disease severity. In this study, we show in the neonate mouse model that sustained high levels of IL-6 produced upon EV71 infection lead to severe tissue damage and eventually death of the animals. Administration of anti-IL-6 neutralizing antibodies after the onset of the clinical symptoms successfully improved the survival rates and clinical scores of the infected hosts. Compared to untreated infected controls, anti-IL-6-treated mice displayed reduced tissue damage, absence of splenic atrophy, and increased immune cell activation. In addition, markedly elevated systemic levels of IL-10 were measured in the protected animals. Furthermore, there was no significant difference in virus titers between anti-IL-6-treated mice and untreated mice, indicating that the anti-IL-6 antibody-mediated protection is independent of the virus load. Our findings thus demonstrate that IL-6 plays a major role in EV71-induced immunopathogenesis. As there is still neither vaccine nor treatment available against EV71, anti-IL-6 antibody treatment represents a potential therapeutic approach to providing protection from the most severe complications of the disease.

A non mouse-adapted dengue virus strain as a new model of severe dengue infection in AG129 mice.

The spread of dengue (DEN) worldwide combined with an increased severity of the DEN-associated clinical outcomes have made this mosquito-borne virus of great global public health importance. Progress in understanding DEN pathogenesis and in developing effective treatments has been hampered by the lack of a suitable small animal model. Most of the DEN clinical isolates and cell culture-passaged DEN virus strains reported so far require either host adaptation, inoculation with a high dose and/or intravenous administration to elicit a virulent phenotype in mice which results, at best, in a productive infection with no, few, or irrelevant disease manifestations, and with mice dying within few days at the peak of viremia. Here we describe a non-mouse-adapted DEN2 virus strain (D2Y98P) that is highly infectious in AG129 mice (lacking interferon-alpha/beta and -gamma receptors) upon intraperitoneal administration. Infection with a high dose of D2Y98P induced cytokine storm, massive organ damage, and severe vascular leakage, leading to haemorrhage and rapid death of the animals at the peak of viremia. In contrast, very interestingly and uniquely, infection with a low dose of D2Y98P led to asymptomatic viral dissemination and replication in relevant organs, followed by non-paralytic death of the animals few days after virus clearance, similar to the disease kinetic in humans. Spleen damage, liver dysfunction and increased vascular permeability, but no haemorrhage, were observed in moribund animals, suggesting intact vascular integrity, a cardinal feature in DEN shock syndrome. Infection with D2Y98P thus offers the opportunity to further decipher some of the aspects of dengue pathogenesis and provides a new platform for drug and vaccine testing.

Behavioural testing and histological assessments of rabbit spinal cord following intrathecal administration of ondansetron.

1. Intrathecal injection of ondansetron has the potential to reduce opioid-related side-effects. The aim of the present study was to determine whether this route of administration produces neuraxial injury. 2. Adult, non-pregnant female New Zealand white rabbits received a single bolus injection of a low (40 microg) or high (4.0 mg) dose of ondansetron into the intrathecal space between the 4th and 5th lumbar vertebrae. In some cases, ondansetron was coadministered with morphine (5 microg/kg). Control animals received a bolus injection of normal saline. Behavioural assessments were conducted at 1 and 24 h to determine overt changes in arousal and mobility, followed by histological evaluation of the excised spinal cord. 3. Of 45 animals investigated, 10 rabbits exhibited modest behavioural evidence of spinal injury, the incidence of which was equally distributed between the treatment groups. Haematoxylin and eosin, along with HAM56, staining of cross-sections of the cervical, thoracic and upper and lower lumbar areas revealed mild signs of inflammation. This, too, was equally distributed between the treatment groups, suggesting that any observed neuraxial injury was the result of needle trauma and not ondansetron neurotoxicity. 4. Collectively, these negative findings support conducting further experiments to fully assess the clinical usefulness of intrathecal ondansetron administration.