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This mainly qualitative study compared 40 families' experience of hospital and home care. This is one aspect of a randomised, controlled trial, which aimed to evaluate the clinical and cost effectiveness of a paediatric hospital at home service (HAH) for acute illness in children. This paper builds upon previous work that has aimed to examine parents' and children's views as service users. Forty families from a larger sample population took part in structured interviews. Eleven children aged 5 to 12 years took part in semistructured interviews. A drawing technique was the chosen method of augmentation in the children's interviews. Research findings showed that HAH is an acceptable alternative to hospital care where there are essentially nursing needs. Thirty-six (90%) parents and seven children stated a clear preference for HAH. The parents' preference was based on a perception that their child's illness wasn't serious or life threatening and therefore could be managed at home with appropriate support from health professionals. The social and financial costs of hospital care compared with HAH were the other main drivers, rather than a comparison of the quality of nursing care of their child. [ABSTRACT FROM AUTHOR]

Colchicine overdose is uncommon but potentially life threatening. It is a safe drug when used according to established therapeutic guidelines but causes serious systemic effects if ingested in doses that exceed the recommendations. Overdose must therefore be recognised early and treated appropriately to optimise the outcome. A fatal case of colchicine overdose caused by inappropriate self medication is reported and to the best of the authors' knowledge, there has been no report of fatal accidental overdose in the United Kingdom. The pharmacology of colchicine, the clinical features associated with overdose, and the options for treatment are discussed. [ABSTRACT FROM AUTHOR]

Background: Advancements in surgical planning, technique, and prosthesis design have improved adaptation to patient anatomy in reverse total shoulder arthroplasty (rTSA). Postoperative changes in deltoid and rotator cuff muscle length are important and may vary based on preoperative indications and prosthesis selection. The purpose of this study is to demonstrate the changes in deltoid and rotator cuff muscle length for planned rTSA using the spectrum of prosthesis configurations in both GHOA and RCA., Methods: Ten shoulder arthroplasty surgeons used preoperative planning software to plan rTSA cases for 20 subjects (10 GHOA, 10 RCA) following surgical guidelines. Each surgeon planned each case using three prosthesis configurations: (1) 8-mm lateralized glenosphere and 135° neck-shaft angle (135+8), (2) 4-mm lateralized glenosphere and 145° neck-shaft angle (145+4), and (3) 0-mm lateralized glenosphere and 155° neck-shaft angle (155+0). Pre- and postoperative deltoid and rotator cuff muscle lengths and percentage-change were calculated and compared between prosthesis configurations within each indication. Different muscle lines of action were included representing the deltoid, subscapularis, infraspinatus, and teres minor., Results: Preoperatively, the RCA cohort had significantly shorter muscle lines of action in the posterior, lateral, and anterior deltoid (P<0.001), a longer inferior subscapularis (P=0.022), and a longer teres minor (P=0.001) than the GHOA cohort. ANOVA and post-hoc analysis showed that post-planning lengths of each deltoid action line were greater in the 155+0 configuration compared to the 135+8 configuration in the RCA cohort (P<0.001, P=0.003, P=0.032, respectively), and post-planning lengths of the anterior and middle deltoid action lines were also greater for the same comparison in the GHOA cohort (P=0.004 and P=0.017, respectively). There were no significant differences in post-planning deltoid lengths between the 135+8 and 145+4 configurations in either diagnosis cohort (P>0.05). All post-planning rotator cuff muscle lengths (subscapularis, infraspinatus, and teres minor) differed significantly (P<0.001) between all prosthesis configurations in both diagnosis cohorts, with the 135+8 configuration resulting in the longest lengths and the 155+0 configuration resulting in the shortest lengths., Conclusion: Automated preoperative planning software calculates the lengths of muscle action lines, which vary between GHOA and RCA diagnoses. Varying rTSA implant geometries result in predictable differences in deltoid lengthening and rotator cuff shortening. Shoulder prostheses with a more lateralized center of rotation show greater rotator cuff muscle length and similar deltoid muscle length when compared to medialized designs with similar deltoid lengthening. Surgeons can use this software to understand the impact of implant geometry on muscle length., (Copyright © 2024. Published by Elsevier Inc.)

Toll-like receptor (TLR) innate immunity signalling protects against pathogens, but excessive or prolonged signalling contributes to a range of inflammatory conditions. Structural information on the TLR cytoplasmic TIR (Toll/interleukin-1 receptor) domains and the downstream adaptor proteins can help us develop inhibitors targeting this pathway. The small molecule o-vanillin has previously been reported as an inhibitor of TLR2 signalling. To study its mechanism of action, we tested its binding to the TIR domain of the TLR adaptor MAL/TIRAP (MAL TIR ). We show that o-vanillin binds to MAL TIR and inhibits its higher-order assembly in vitro . Using NMR approaches, we show that o-vanillin forms a covalent bond with lysine 210 of MAL. We confirm in mouse and human cells that o-vanillin inhibits TLR2 but not TLR4 signalling, independently of MAL, suggesting it may covalently modify TLR2 signalling complexes directly. Reactive aldehyde-containing small molecules such as o-vanillin may target multiple proteins in the cell.

Background: Particle-induced osteolysis resulting from polyethylene wear remains a source of implant failure in anatomic total shoulder designs. Modern polyethylene components are irradiated in an oxygen-free environment to induce cross-linking, but reducing the resulting free radicals with melting or heat annealing can compromise the component's mechanical properties. Vitamin E has been introduced as an adjuvant to thermal treatments. Anatomic shoulder arthroplasty models with a ceramic head component have demonstrated that vitamin E-enhanced polyethylene show improved wear compared with highly cross-linked polyethylene (HXLPE). This study aimed to assess the biomechanical wear properties and particle size characteristics of a novel vitamin E-enhanced highly cross-linked polyethylene (VEXPE) glenoid compared to a conventional ultrahigh-molecular-weight polyethylene (UHMWPE) glenoid against a cobalt chromium molybdenum (CoCrMo) head component., Methods: Biomechanical wear testing was performed to compare the VEXPE glenoid to UHMWPE glenoid with regard to pristine polyethylene wear and abrasive endurance against a polished CoCrMo alloy humeral head in an anatomic shoulder wear-simulation model. Cumulative mass loss (milligrams) was recorded, and wear rate calculated (milligrams per megacycle [Mc]). Under pristine wear conditions, particle analysis was performed, and functional biologic activity (FBA) was calculated to estimate particle debris osteolytic potential. In addition, 95% confidence intervals for all testing conditions were calculated., Results: The average pristine wear rate was statistically significantly lower for the VEXPE glenoid compared with the HXLPE glenoid (0.81 ± 0.64 mg/Mc vs. 7.00 ± 0.45 mg/Mc) (P < .05). Under abrasive wear conditions, the VEXPE glenoid had a statistically significant lower average wear rate compared with the UHMWPE glenoid comparator device (18.93 ± 5.80 mg/Mc vs. 40.47 ± 2.63 mg/Mc) (P < .05). The VEXPE glenoid demonstrated a statistically significant improvement in FBA compared with the HXLPE glenoid (0.21 ± 0.21 vs. 1.54 ± 0.49 (P < .05)., Conclusions: A new anatomic glenoid component with VEXPE demonstrated significantly improved pristine and abrasive wear properties with lower osteolytic particle debris potential compared with a conventional UHMWPE glenoid component. Vitamin E-enhanced polyethylene shows early promise in shoulder arthroplasty components. Long-term clinical and radiographic investigation needs to be performed to verify if these biomechanical wear properties translate to diminished long-term wear, osteolysis, and loosening., (Copyright © 2024 Journal of Shoulder and Elbow Surgery Board of Trustees. Published by Elsevier Inc. All rights reserved.)

Improving the prognosis for patients with metastatic HR+/HER2- breast cancer remains an unmet need. Patients with tumors that have progressed on endocrine therapy and/or are not eligible for endocrine therapy had limited treatment options beyond chemotherapy. Antibody-drug conjugates are a novel and promising treatment class in this setting. Datopotamab deruxtecan (Dato-DXd) consists of a TROP2-directed humanized IgG1 monoclonal antibody attached via a serum-stable cleavable linker to a topoisomerase I inhibitor payload. TROPION-Breast01 is an ongoing phase III study that is evaluating the efficacy and safety of Dato-DXd compared with investigator's choice of standard-of-care chemotherapy in patients with inoperable or metastatic HR+/HER2- breast cancer who have received one or two prior lines of systemic chemotherapy in the inoperable or metastatic setting. Clinical Trial Registration: NCT05104866 (ClinicalTrials.gov).

Bivalency is a prevalent natural mechanism to enhance receptor avidity. Various two-domain disulfide-rich peptides exhibiting bivalent action have been identified from animal venoms. A unique characteristic of these peptides is that they induce a pharmacological response different from that provoked by any of the constituent domains. The enhanced potency and avidity of such peptides is therefore a consequence of their domain fusion by a peptide linker. The role of the linker itself, beyond conjugation, remains unclear. Here, we investigate how the linker affects the bivalency of the capsaicin receptor (TRPV1) agonist DkTx. We recombinantly produced isotope labelled DkTx using a protein splicing approach, to solve the high-resolution solution structure of DkTx, revealing residual linker order stabilised by linker-domain interactions leading to biased domain orientations. The significance of this was studied using a combination of mutagenesis, spin relaxation studies and electrophysiology measurements. Our results reveal that disrupting the pre-organisation of the domains of DkTx is accompanied by reductions in potency and onset of avidity. Our findings support a model of pre-configured two-domain binding, in favour of the previously suggested sequential binding model. This highlights the significance of ordered elements in linker design and the natural evolution of these in bivalent toxins., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Despite recent treatment advances, the prognosis for patients with locally recurrent inoperable or metastatic triple-negative breast cancer (TNBC) remains poor. The antibody-drug conjugate datopotamab deruxtecan (Dato-DXd) is composed of a humanized anti-TROP2 IgG1 monoclonal antibody linked to a topoisomerase I inhibitor payload via a stable, cleavable linker. The phase III TROPION-Breast02 trial in patients previously untreated for locally recurrent inoperable or metastatic TNBC, who are not candidates for PD-1/PD-L1 inhibitors is evaluating efficacy and safety of Dato-DXd versus investigator's choice of chemotherapy (ICC). Approximately 600 patients will be randomized 1:1 to Dato-DXd 6 mg/kg iv. every 3 weeks or ICC (paclitaxel, nab-paclitaxel, carboplatin, capecitabine or eribulin mesylate). Dual primary end points are progression-free survival by blinded independent central review and overall survival.

Group heteroscedasticity is commonly observed in pseudo-bulk single-cell RNA-seq datasets and its presence can hamper the detection of differentially expressed genes. Since most bulk RNA-seq methods assume equal group variances, we introduce two new approaches that account for heteroscedastic groups, namely voomByGroup and voomWithQualityWeights using a blocked design (voomQWB). Compared to current gold-standard methods that do not account for group heteroscedasticity, we show results from simulations and various experiments that demonstrate the superior performance of voomByGroup and voomQWB in terms of error control and power when group variances in pseudo-bulk single-cell RNA-seq data are unequal., (© 2023. The Author(s).)

Multivalent ligands of ion channels have proven to be both very rare and highly valuable in yielding unique insights into channel structure and pharmacology. Here, we describe a bivalent peptide from the venom of Xibalbanus tulumensis, a troglobitic arthropod from the enigmatic class Remipedia, that causes persistent calcium release by activation of ion channels involved in muscle contraction. The high-resolution solution structure of φ-Xibalbin3-Xt3a reveals a tandem repeat arrangement of inhibitor-cysteine knot (ICK) domains previously only found in spider venoms. The individual repeats of Xt3a share sequence similarity with a family of scorpion toxins that target ryanodine receptors (RyR). Single-channel electrophysiology and quantification of released Ca 2+ stores within skinned muscle fibers confirm Xt3a as a bivalent RyR modulator. Our results reveal convergent evolution of RyR targeting toxins in remipede and scorpion venoms, while the tandem-ICK repeat architecture is an evolutionary innovation that is convergent with toxins from spider venoms., (© 2023. The Author(s).)

The paramagnetism of a lanthanoid tag site-specifically installed on a protein provides a rich source of structural information accessible by nuclear magnetic resonance (NMR) and electron paramagnetic resonance (EPR) spectroscopy. Here we report a lanthanoid tag for selective reaction with cysteine or selenocysteine with formation of a (seleno)thioether bond and a short tether between the lanthanoid ion and the protein backbone. The tag is assembled on the protein in three steps, comprising (i) reaction with 4-fluoro-2,6-dicyanopyridine (FDCP); (ii) reaction of the cyano groups with α -cysteine, penicillamine or β -cysteine to complete the lanthanoid chelating moiety; and (iii) titration with a lanthanoid ion. FDCP reacts much faster with selenocysteine than cysteine, opening a route for selective tagging in the presence of solvent-exposed cysteine residues. Loaded with Tb 3 + and Tm 3 + ions, pseudocontact shifts were observed in protein NMR spectra, confirming that the tag delivers good immobilisation of the lanthanoid ion relative to the protein, which was also manifested in residual dipolar couplings. Completion of the tag with different 1,2-aminothiol compounds resulted in different magnetic susceptibility tensors. In addition, the tag proved suitable for measuring distance distributions in double electron-electron resonance experiments after titration with Gd 3 +  ions., Competing Interests: At least one of the (co-)authors is a member of the editorial board of Magnetic Resonance. The peer-review process was guided by an independent editor, and the authors also have no other competing interests to declare., (Copyright: © 2022 Sreelakshmi Mekkattu Tharayil et al.)

Atypical teratoid/rhabdoid tumors (AT/RT) are the most common malignant brain tumors of infancy and have a dismal 4-year event-free survival (EFS) of 37%. We have previously shown that mTOR activation contributes to AT/RT's aggressive growth and poor survival. Targeting the mTOR pathway with the dual mTORC1/2 inhibitor TAK-228 slows tumor growth and extends survival in mice bearing orthotopic xenografts. However, responses are primarily cytostatic with limited durability. The aim of this study is to understand the impact of mTOR inhibitors on AT/RT signaling pathways and design a rational combination therapy to drive a more durable response to this promising therapy. We performed RNASeq, gene expression studies, and protein analyses to identify pathways disrupted by TAK-228. We find that TAK-228 decreases the expression of the transcription factor NRF2 and compromises AT/RT cellular defenses against oxidative stress and apoptosis. The BH3 mimetic, Obatoclax, is a potent inducer of oxidative stress and apoptosis in AT/RT. These complementary mechanisms of action drive extensive synergies between TAK-228 and Obatoclax slowing AT/RT cell growth and inducing apoptosis and cell death. Combination therapy activates the integrative stress response as determined by increased expression of phosphorylated EIF2α, ATF4, and CHOP, and disrupts the protective NOXA.MCL-1.BIM axis, forcing stressed cells to undergo apoptosis. Combination therapy is well tolerated in mice bearing orthotopic xenografts of AT/RT, slows tumor growth, and extends median overall survival. This novel combination therapy could be added to standard upfront therapies or used as a salvage therapy for relapsed disease to improve outcomes in AT/RT., (© 2022. The Author(s).)

Reprograming of cellular metabolism is a hallmark of cancer. Altering metabolism allows cancer cells to overcome unfavorable microenvironment conditions and to proliferate and invade. Medulloblastoma is the most common malignant brain tumor of children. Genomic amplification of MYC defines a subset of poor-prognosis medulloblastoma. We performed comprehensive metabolic studies of human MYC -amplified medulloblastoma by comparing the metabolic profiles of tumor cells in three different conditions-in vitro, in flank xenografts and in orthotopic xenografts in the cerebellum. Principal component analysis showed that the metabolic profiles of brain and flank high-MYC medulloblastoma tumors clustered closely together and separated away from normal brain and in vitro MYC-amplified cells. Compared to normal brain, MYC -amplified medulloblastoma orthotopic xenograft tumors showed upregulation of the TCA cycle as well as the synthesis of nucleotides, hexosamines, amino acids and glutathione. There was significantly higher glucose uptake and usage in orthotopic xenograft tumors compared to flank xenograft tumors and cells in culture. In orthotopic tumors, glucose was the main carbon source for the de novo synthesis of glutamate, glutamine and glutathione through the TCA cycle. In vivo, the glutaminase II pathway was the main pathway utilizing glutamine. Glutathione was the most abundant upregulated metabolite in orthotopic tumors compared to normal brain. Glutamine-derived glutathione was synthesized through the glutamine transaminase K (GTK) enzyme in vivo. In conclusion, high MYC medulloblastoma cells have different metabolic profiles in vitro compared to in vivo, and key vulnerabilities may be missed by not performing in vivo metabolic analyses.

Medulloblastoma is the most common malignant pediatric brain tumor. Amplification of c-MYC is a hallmark of a subset of poor-prognosis medulloblastoma. MYC upregulates glutamine metabolism across many types of cancer. We modified the naturally occurring glutamine antagonist 6-diazo-5-oxo-l-norleucine (DON) by adding 2 promoeities to increase its lipophilicity and brain penetration creating the prodrug isopropyl 6-diazo-5-oxo-2-(((phenyl (pivaloyloxy) methoxy) - carbonyl) amino) hexanoate, termed JHU395. This prodrug was shown to have a 10-fold improved CSF-to-plasma ratio and brain-to-plasma ratio relative to DON. We hypothesized that JHU395 would have superior cell penetration compared with DON and would effectively and more potently kill MYC-expressing medulloblastoma. JHU395 treatment caused decreased growth and increased apoptosis in multiple human high-MYC medulloblastoma cell lines at lower concentrations than DON. Parenteral administration of JHU395 in Nu/Nu mice led to the accumulation of micromolar concentrations of DON in brain. Treatment of mice bearing orthotopic xenografts of human MYC-amplified medulloblastoma with JHU395 increased median survival from 26 to 45 days compared with vehicle control mice (p < 0.001 by log-rank test). These data provide preclinical justification for the ongoing development and testing of brain-targeted DON prodrugs for use in medulloblastoma., (© 2021 American Association of Neuropathologists, Inc. All rights reserved.)

The mitogen-activated protein kinase pathway is one of the most frequently altered pathways in cancer. It is involved in the control of cell proliferation, invasion, and metabolism, and can cause resistance to therapy. A number of aggressive malignancies, including melanoma, colon cancer, and glioma, are driven by a constitutively activating missense mutation (V600E) in the v-Raf murine sarcoma viral oncogene homolog B (BRAF) component of the pathway. Mitogen-activated protein kinase kinase (MEK) inhibition is initially effective in targeting these cancers, but reflexive activation of mammalian target of rapamycin (mTOR) signaling contributes to frequent therapy resistance. We have previously demonstrated that combination treatment with the MEK inhibitor trametinib and the dual mammalian target of rapamycin complex 1/2 inhibitor TAK228 improves survival and decreases vascularization in a BRAF V600E mutant glioma model. To elucidate the mechanism of action of this combination therapy and understand the ensuing tumor response, we performed comprehensive unbiased proteomic and phosphoproteomic characterization of BRAF V600E mutant glioma xenografts after short-course treatment with trametinib and TAK228. We identified 13,313 proteins and 30,928 localized phosphosites, of which 12,526 proteins and 17,444 phosphosites were quantified across all samples (data available via ProteomeXchange; identifier PXD022329). We identified distinct response signatures for each monotherapy and combination therapy and validated that combination treatment inhibited activation of the mitogen-activated protein kinase and mTOR pathways. Combination therapy also increased apoptotic signaling, suppressed angiogenesis signaling, and broadly suppressed the activity of the cyclin-dependent kinases. In response to combination therapy, both epidermal growth factor receptor and class 1 histone deacetylase proteins were activated. This study reports a detailed (phospho)proteomic analysis of the response of BRAF V600E mutant glioma to combined MEK and mTOR pathway inhibition and identifies new targets for the development of rational combination therapies for BRAF-driven tumors., Competing Interests: Conflict of interest The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

The natural flight response in shrimp is powered by rapid contractions of the abdominal muscle fibres to propel themselves backwards away from perceived danger. This muscle contraction is dependent on repetitive depolarization of muscle plasma membrane, triggering tightly spaced cytoplasmic [Ca 2+ ] transients and rapidly rising tetanic force responses. To achieve such high amplitude and high frequency of Ca 2+ transients requires a high abundance of sarcoplasmic/endoplasmic reticulum Ca 2+ ATPase (SERCA) to rapidly clear cytoplasmic Ca 2+ between each transient and an efficient Ca 2+ release system consisting of the Ryanodine Receptor (RyR), and voltage gated Ca 2+ channels (Ca V s). With the aim to expand our knowledge of muscle gene function and identify orthologous genes regulating muscle excitation-contraction (EC) coupling, this study assembled nine Penaeid shrimp muscle transcriptomes. On average, the nine transcriptomes contained 27,000 contigs, with an annotation rate of 36% and a BUSCO completeness of 70%. Despite maintaining their function, the crustacean RyR and Ca V proteins showed evidence of significant diversification from mammalian orthologs, while SERCA remained more conserved. Several key components of protein interaction were conserved, while others showed distinct crustacean specific evolutionary adaptations. Lastly, this study revealed approximately 1,000 orthologous genes involved in muscle specific processes present across all nine species., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Objective: To compare the performance of polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) tests for diagnosing Echinococcus granulosus in dog feces among national reference laboratories in Argentina, Chile, Peru, and Uruguay., Methods: National laboratories affiliated with the Ministry of Health/Agriculture of each country exchanged panels of 10 positive/negative samples obtained from their regular national surveillance programs in November 2015 - November 2016. All laboratories applied PCR; two also applied ELISA techniques. Sensitivity and specificity were determined for each laboratory and concordance of results among the laboratories was evaluated by Cohen Kappa coefficient., Results: Poor concordance (3 of 10 paired comparisons had values of Kappa > 0.4), low sensitivity and specificity across all laboratories, and poor performance of both techniques in detecting E. granulosus in canine feces was demonstrated in this study. An ex-post comparison of the laboratories' test protocols showed substantial heterogeneity that could partially explain poor concordance of results., Conclusion: The results underscore the heterogeneity of canine echinococcosis diagnosis across the region and indicate possible sources of variability. Efforts to standardize canine echinococcosis testing must be included in the plan of action for the Regional Initiative for the Control of Cystic Echinococcosis. Future comparisons with fecal samples of known parasite load are needed., Competing Interests: Conflict of interests. None declared.

One Health Surveillance (OHS) implements the One Health approach to improving health by collecting data and producing information to support integrated action across the animal health, human health and environment sectors. The purpose of this study was to survey the biosurveillance community to assess its OHS practices and capabilities, its attitudes towards OHS (perceived value), and the factors that motivate its members to implement OHS practices. The authors used a convenience sample of 185 professionals from multiple domains and 44 nations. They examined the extent to which these professionals implemented OHS, gathered their opinions on the value of OHS, assessed their perceptions of the capacity to perform specific OHS tasks and identified their priorities for change. Over 85% of all respondents said that they considered OHS to be beneficial, with no significant differences between work domains or country income groups; over 50% indicated that they already applied OHS. Obtaining access to data collected by other domains was both the most frequent challenge and the most difficult to improve. The highest priority for improvement was having the ability to send and receive electronic data. Respondents from low-income or middle-income countries were more motivated to make improvements than stakeholders from high-income countries. These findings provide a snapshot of current opinions and practices and, together with suggestions for improvements from professionals in the field, can help to target priority needs for OHS information, training and resources., (© World Organisation for Animal Health (OIE), 2019)

Chronic obstructive pulmonary disease (COPD) is an incurable inflammatory lung disease that afflicts millions of people worldwide, and it is the fourth leading cause of death. Systemic comorbidities affecting the heart, skeletal muscle, bone, and metabolism are major contributors to morbidity and mortality. Given the surprising finding in large prospective clinical biomarker studies that peripheral white blood cell count is more closely associated with disease than inflammatory biomarkers, we probed the role of blood growth factors. Using the SHIP-1-deficient COPD mouse model, which manifests a syndrome of destructive lung disease and a complex of comorbid pathologies, we have identified a critical and unexpected role for granulocyte-CSF (G-CSF) in linking these conditions. Deletion of G-CSF greatly reduced airway inflammation and lung tissue destruction, and attenuated systemic inflammation, right heart hypertrophy, loss of fat reserves, and bone osteoporosis. In human clinical translational studies, bronchoalveolar lavage fluid of patients with COPD demonstrated elevated G-CSF levels. These studies suggest that G-CSF may play a central and unforeseen pathogenic role in COPD and its complex comorbidities, and identify G-CSF and its regulators as potential therapeutic targets.

Neovascular retinopathies are major causes of vision loss; yet treatments to prevent the condition are inadequate. The role of regulatory T cells in neovascular retinopathy is unknown. Here we show that in retinopathy regulatory T cells are transiently increased in lymphoid organs and the retina, but decline when neovascularization is established. The decline is prevented following regulatory T cells expansion with an IL-2/anti-IL-2 mAb complex or the adoptive transfer of regulatory T cells. Further, both approaches reduce vasculopathy (vaso-obliteration, neovascularization, vascular leakage) and alter the activation of Tmem119 + retinal microglia. Our in vitro studies complement these findings, showing that retinal microglia co-cultured with regulatory T cells exhibit a reduction in co-stimulatory molecules and pro-inflammatory mediators that is attenuated by CTLA-4 blockade. Collectively, we demonstrate that regulatory T cells are recruited to the retina and, when expanded in number, repair the vasculature. Manipulation of regulatory T cell numbers is a previously unrecognized, and promising avenue for therapies to prevent blinding neovascular retinopathies.The local immune responses in the eye are attenuated to preserve sight. Surprisingly, Deliyanti et al. show that regulatory T cells (Tregs) take an active role in protecting the eye from neovascularization in oxygen-induced retinopathy, and that interventions that augment the retinal Treg numbers reduce neovascular retinopathy in mice.

Although subjects with recurrent low back pain (LBP) demonstrate altered trunk control, the kinematic and kinetic responses of the trunk have not been carefully investigated. This study was conducted to compare the standing time, spine range of motion (ROM), and dynamic postural steadiness index (DPSI) based on visual condition between subjects with and without recurrent LBP during upright one leg standing. Sixty-three individuals participated in the study, including 34 control subjects and 29 subjects with recurrent LBP. The DPSI was a composite of the medio-lateral (MLSI), anterior-posterior (APSI), and vertical steadiness indices (VSI) on a force platform. The control group demonstrated longer standing time (s) during the eyes-open condition than the LBP group (26.82±6.03 vs. 19.87±9.36; t=2.96, p=0.01). Regarding spine ROM, visual condition was significantly different between groups (F=7.09, p=0.01) and demonstrated interactions with spine region and group (F=5.53, p=0.02). For the kinetic measures, there was a significant interaction between visual conditions and indices (F=25.30, p=0.001). In the LBP group, the DPSI was significantly correlated with the MLSI (r=0.59, p=0.002), APSI (r=0.44, p=0.03), and VSI (r=0.98, p=0.01) in the eyes-closed condition. Overall, the results of this study indicated that the LBP group decreased thorax and lumbar spine rotations during the eyes-closed condition. The LBP group also demonstrated positive correlations with the kinetic indices, enhancing dynamic postural steadiness in the eyes-closed condition in order to possibly avoid pain or further injury. This dynamic postural steadiness strategy is necessary to improve kinetic and kinematic chain reactions in the LBP group. This compensatory pattern supports the development of optimal postural correction strategies to prevent LBP recurrence and might represent a chain reaction to protect trunk control without visual input., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Larvae (maggots) of Cochliomyia hominivorax, the New World Screwworm fly, are voracious consumers of living flesh that have a negative economic impact by decreasing productivity, predisposing to other pathogens, and, in severe cases, causing death of domestic livestock. Screwworm caused extensive financial losses to the livestock industry in North America prior to its eradication. Sterile insect technique (SIT) was used to eradicate screwworm throughout North and Central America and continues to be the main tool to control it in eastern Panama. The goal of this study was to evaluate the temporal and spatial trends of screwworm myiasis cases reported in the Province of Darien and Comarca Embera (border with Colombia), Panama, from 2001 to 2011. We hypothesized that screwworm cases would vary seasonally and be spatially clustered near Colombia as a result of effective eradication strategies in Panama and the presence of an autochthonous population of flies in western Colombia. Temporal and spatial data were retrieved from COPEG-USDA records (Panama) and analysed by anova, Ripley's K function, discrete Poisson spatial statistic scan and Getis-Ord Gi*. No significant temporal trend was found, but cases were spatially distributed in four clusters. One cluster of cases occurred from 2001 to 2003 and was considered a focal temporal and spatial cluster. One cluster occurred in 2001 and 2007 indicating more rare outbreaks in an area with fewer cattle. The two remaining clusters contained cases from 2004 to 2011 and 2001 to 2011 suggesting regular breaks in the control barrier due to occasional failures of the SIT programme, difficulties implementing border quarantine strategies, livestock smuggling or the movement of infested wildlife., (© 2015 Blackwell Verlag GmbH.)

Background: In recent years, global public health security has been threatened by zoonotic disease emergence as exemplified by outbreaks of H5N1 and H1N1 influenza, SARS, and most recently Ebola and Zika. Additionally, endemic zoonoses, such as rabies, burden countries year after year, placing demands on limited finances and personnel. To survey the baseline status of the emerging and endemic zoonoses programmes of the Latin American and the Caribbean (LAC) countries, the Pan American Health Organization (PAHO) conducted a survey of priority emerging and endemic zoonoses, countries´ prioritization criteria and methodologies, and suggestions to strengthen countries capacities and regional approaches to zoonoses control., Methods: A fillable online questionnaire was sent to the zoonoses programme managers of the Ministries of Health (MOH) and Ministries of Agriculture (MAg) of 33 LAC countries from January to April of 2015. The questionnaire comprised 36 single, multiple choice and open-ended questions to inform the objectives of the survey. A descriptive exploratory analysis was completed., Results: Fifty-four ministries (26 MOH, 25 MAg, and 3 combined responses) in 31 LAC countries responded to the survey. Within the ministries, 22 (85%) MOH, 5 (20%) MAg, and 2 (67%) combined entities indicated they had specialized zoonoses units. For endemic zoonoses, 32 of 54 ministries responded that they conduct formal prioritization exercises, most of them annually (69%). The three priority endemic zoonoses for the MOHs were leptospirosis, rabies, and brucellosis while the three priorities for the MAgs were brucellosis, rabies, and tuberculosis. Diagnosis for rabies and leptospirosis were cited as the capacities most in need of development. The most needed cross-cutting capacity was coordination between stakeholders. For emerging zoonoses, 28 ministries performed formal prioritization exercises. The top prioritization criteria were probability of introduction into the country and impact. The three priority emerging zoonoses for the MOHs were Ebola viral disease, avian influenza, and Chikungunya while for the MAgs were avian influenza, bovine spongiform encephalopathy and West Nile virus disease. Surveillance for avian influenza and Ebola, and diagnosis for BSE were quoted as the capacities most needed. For all zoonoses, the majority of respondents (69%) ranked their relationship with the other Ministry as productive or very productive, and 31% minimally productive. Many countries requested a formal regional network, better regional communication and collaboration, and integrated surveillance., Conclusions: The survey is the first comprehensive effort to date to inform the status of zoonoses programmes in LAC. The information collected here will be used to develop a regional strategy for zoonoses (both endemic and emerging), increase efforts, advocacy, and promote prompt identification and management of EIDs and improvement of endemic programmes.

We examined the role of NFκB1 in the homeostasis and function of peripheral follicular (Fo) B cells. Aging mice lacking NFκB1 (Nfκb1(-/-)) develop lymphoproliferative and multiorgan autoimmune disease attributed in large part to the deregulated activity of Nfκb1(-/-)Fo B cells that produce excessive levels of the proinflammatory cytokine interleukin 6 (IL-6). Despite enhanced germinal center (GC) B cell differentiation, the formation of GC structures was severely disrupted in the Nfκb1(-/-)mice. Bone marrow chimeric mice revealed that the Fo B cell-intrinsic loss of NFκB1 led to the spontaneous generation of GC B cells. This was primarily the result of an increase in IL-6 levels, which promotes the differentiation of Fo helper CD4(+)T cells and acts in an autocrine manner to reduce antigen receptor and toll-like receptor activation thresholds in a population of proliferating IgM(+)Nfκb1(-/-)Fo B cells. We demonstrate that p50-NFκB1 represses Il-6 transcription in Fo B cells, with the loss of NFκB1 also resulting in the uncontrolled RELA-driven transcription of Il-6.Collectively, our findings identify a previously unrecognized role for NFκB1 in preventing multiorgan autoimmunity through its negative regulation of Il-6 gene expression in Fo B cells., (© 2016 de Valle et al.)

Maintenance of an appropriate number of plasma cells, long-lived antibody-producing cells that are derived from B cells, is essential for maintaining immunological memory while limiting disease. Plasma cell survival relies on extrinsic factors, the limited availability of which determines the size of the plasma cell population. Mice deficient in the nonreceptor tyrosine kinase Lyn are prone to an autoimmune disease that is characterized by inflammation and an excess of plasma cells (plasmacytosis). We demonstrated that the plasmacytosis was intrinsic to B cells and independent of inflammation. We also showed that Lyn attenuated signaling by signal transducer and activator of transcription 3 (STAT3) and STAT5 in response to the cytokines interleukin-6 (IL-6) and IL-3, respectively, in two previously uncharacterized plasma cell signaling pathways. Thus, in the absence of Lyn, the survival of plasma cells was improved, which enabled the plasma cells to become established in excess numbers in niches in vivo. These data identify Lyn as a key regulator of survival signaling in plasma cells, limiting plasma cell accumulation and autoimmune disease susceptibility., (Copyright © 2014, American Association for the Advancement of Science.)

SHIP-1 has an important role in controlling immune cell function through its ability to downmodulate PI3K signaling pathways that regulate cell survival and responses to stimulation. Mice deficient in SHIP-1 display several chronic inflammatory phenotypes including antibody-mediated autoimmune disease, Crohn's disease-like ileitis and a lung disease reminiscent of chronic obstructive pulmonary disease. The ileum and lungs of SHIP-1-deficient mice are infiltrated at an early age with abundant myeloid cells and the mice have a limited lifespan primarily thought to be due to the consolidation of lungs with spontaneously activated macrophages. To determine whether the myeloid compartment is the key initiator of inflammatory disease in SHIP-1-deficient mice, we examined two independent strains of mice harboring myeloid-restricted deletion of SHIP-1. Contrary to expectations, conditional deletion of SHIP-1 in myeloid cells did not result in consolidating pneumonia or segmental ileitis typical of germline SHIP-1 deficiency. In addition, other myeloid cell abnormalities characteristic of germline loss of SHIP-1, including flagrant splenomegaly and enhanced myelopoiesis, were absent in mice lacking SHIP-1 in myeloid cells. This study indicates that the spontaneous inflammatory disease characteristic of germline SHIP-1 deficiency is not initiated solely by LysM-positive myeloid cells but requires the simultaneous loss of SHIP-1 in other hematolymphoid lineages.

Aims/hypothesis: Type 1 diabetes is characterised by early peri-islet insulitis and insulin autoantibodies, followed by invasive insulitis and beta cell destruction. The immunological events that precipitate invasive insulitis are not well understood. We tested the hypothesis that B cells in diabetes-prone NOD mice drive invasive insulitis through elevated expression of CD19 and consequent enhanced uptake and presentation of beta cell membrane-bound antigens to islet invasive T cells., Methods: CD19 expression and signalling pathways in B cells from NOD and control mice were compared. Expansion of CD8(+) T cells specific for insulin and islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP) were compared in CD19-deficient and wild-type NOD mice and this was correlated with insulitis severity. The therapeutic potential of anti-CD19 treatment during the period of T cell activation was assessed for its ability to block invasive insulitis., Results: CD19 expression and signalling in B cells was increased in NOD mice. CD19 deficiency significantly diminished the expansion of CD8(+) T cells with specificity for the membrane-bound beta cell antigen, IGRP. Conversely the reduction in CD8(+) T cells with specificity for the soluble beta cell antigen, insulin, was relatively small and not significant., Conclusions/interpretation: Elevated CD19 on NOD B cells promotes presentation of the membrane-bound antigen, IGRP, mediating the expansion of autoreactive T cells specific for antigens integral to beta cells, which are critical for invasive insulitis and diabetes. Downregulating the CD19 signalling pathway in insulin autoantibody-positive individuals before the development of type 1 diabetes may prevent expansion of islet-invasive T cells and preserve beta cell mass.

Objective: Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease that is characterized by the production of antinuclear antibodies (ANAs) and leads to immune complex deposition in the kidneys and nephritis. Lyn tyrosine kinase is a regulator of antibody-mediated autoimmune disease, as evidenced by studies in gene-targeted mice and as suggested in genome-wide association studies in SLE. Like SLE patients, Lyn-deficient mice have increased levels of interleukin-6 (IL-6). Deletion of IL-6 from Lyn-deficient mice abrogates levels of inflammation, pathogenic autoantibodies, and nephritis. The purpose of this study was to assess the role of IL-6 trans-signaling in autoimmune disease by overexpressing soluble gp130Fc (sgp130Fc) in a mouse model., Methods: The effect of overexpression of sgp130Fc on immune cell phenotypes was determined by flow cytometry in young and aged mice with lupus, and ANAs were measured by enzyme-linked immunosorbent assay. Glomerulonephritis was assessed by histopathologic analysis, by measuring the glomerular area and the blood urea nitrogen concentration, and by immunohistochemistry. Immunofluorescence defined renal immune complex and complement deposition. The acute-phase response was determined by quantitative real-time polymerase chain reaction., Results: In contrast to removing IL-6, impaired IL-6 trans-signaling had little effect on many immune cell abnormalities in Lyn-/- mice. Pathogenic ANAs and kidney deposition of immune complexes were also unaltered by sgp130Fc. However, sgp130Fc overexpression led to diminished macrophage expansion, reduced glomerular leukocyte infiltration, reduced complement fixation, significantly attenuated glomerulonephritis, and improved renal function in Lyn-deficient mice., Conclusion: Our results reveal key roles of leukocytes, complement, and the innate immune system in mediating glomerulonephritis, and they implicate IL-6 trans-signaling in this process. We suggest that targeting this pathway may be an effective adjunct to B cell depletion in SLE treatment., (Copyright © 2013 by the American College of Rheumatology.)

Lyn is involved in erythropoietin (Epo)-receptor signaling and erythroid homeostasis. Downstream pathways influenced following Lyn activation and their significance to erythropoiesis remain unclear. To address this, we assessed a gain-of-function Lyn mutation (Lyn(up/up)) on erythropoiesis and Epo receptor signaling. Adult Lyn(up/up) mice were anemic, with dysmorphic red cells (spherocyte-like, acanthocytes) in their circulation, indicative of hemolytic anemia and resembling the human disorder chorea acanthocytosis. Heterozygous Lyn(+/up) mice became increasingly anemic with age, indicating that the mutation was dominant. In an attempt to overcome this anemia, extramedullary erythropoiesis was activated. As the mice aged, the levels of different immature erythroid populations changed, indicating compensatory mechanisms to produce more erythrocytes were dynamic. Changes in Epo signaling were observed in Lyn(+/up) erythroid cell lines and primary CD71(+) Lyn(up/up) erythroblasts, including significant alterations to the phosphorylation of Lyn, the Epo receptor, Janus kinase 2, Signal Transducer and Action of Transcription-5, GRB2-associated-binding protein-2, Akt, and Forkhead box O3. As a consequence of altered Lyn signaling, Lyn(+/up) cells remained viable in the absence of Epo but displayed delayed Epo-induced differentiation. These data demonstrate that Lyn gene dosage and activity are critical for normal erythropoiesis; constitutively active Lyn alters Epo signaling, which in turn produces erythroid defects.

Atopy and autoimmunity are usually considered opposed immunological manifestations. Lyn(-/-) mice develop lupus-like autoimmune disease yet have coexistent intrinsic allergic traits and are prone to severe, persistent asthma induced exogenously. Recently it has been proposed that the Th2 environment and IgE auto-Abs promotes autoimmune disease in Lyn(-/-) mice. To examine this apparent contradiction, we derived Lyn(-/-) mice with a null mutation in STAT6, a regulator of Th2 immunity that integrates signaling from the IL-4/IL-13 receptor complex. Atopy and spontaneous peritoneal eosinophilia, characteristic of Lyn(-/-) mice, were lost in young Lyn(-/-)STAT6(-/-) mice; however, autoimmune disease was markedly exacerbated. At a time-point where Lyn(-/-) mice showed only mild autoimmune disease, Lyn(-/-)STAT6(-/-) mice had maximal titres of IgG and IgA auto-Abs, impaired renal function, myeloid expansion and a highly activated T cell compartment. Remarkably, low level IgE auto-Abs but not IgG1 auto-Abs were a feature of some aged Lyn(-/-)STAT6(-/-) mice. Furthermore, aged Lyn(-/-)STAT6(-/-) mice showed dramatically increased levels of serum IgE but minimal IgG1, suggesting that class-switching to IgE can occur in the absence of an IgG1 intermediate. The results show that Lyn-deficient mice can overcome the effects of disabling Th2 immunity, highlighting the importance of Lyn in controlling Th2 responses. Our data also indicates that, under certain conditions, STAT6-independent factors can promote IgE class-switching. This work has important clinical implications as many experimental therapies designed for the treatment of asthma or atopy are based on targeting the STAT6 axis, which could potentially reveal life endangering autoimmunity or promote atopy in susceptible individuals., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Ab-mediated autoimmune disease is multifaceted and may involve many susceptibility loci. The majority of autoimmune patients are thought to have polymorphisms in a number of genes that interact in different combinations to contribute to disease pathogenesis. Studies in mice and humans have implicated the Lyn protein tyrosine kinase as a regulator of Ab-mediated autoimmune disease. To examine whether haploinsufficiency of Lyn gives rise to cellular and clinical manifestations of autoimmune disease, we evaluated the phenotype of Lyn(+/-) mice. We find that their B cell compartment is significantly perturbed, with reduced numbers of marginal zone and transitional stage 2 B cells, expansion of plasma cells, downregulation of surface IgM, and upregulation of costimulatory molecules. Biochemical studies show that Lyn(+/-) B cells have defects in negative regulation of signaling, whereas Lyn(+/-) mice develop IgG autoantibodies and glomerulonephritis with age. Because Lyn has a pivotal role in the activation of inhibitory phosphatases, we generated mice harboring double heterozygous loss-of-function mutations in Lyn and SHP-1 or Lyn and SHIP-1. Partial inactivation of SHP-1 or SHIP-1 amplifies the consequence of Lyn haploinsufficiency, leading to an accelerated development of autoantibodies and disease. Our data also reveal that the BALB/c background is protective against autoimmune-mediated glomerulonephritis, even in the face of high titer autoantibodies, whereas the C57BL/6 background is susceptible. This study demonstrates that Lyn is a haploinsufficient gene in autoimmune disease and importantly shows that quantitative genetic variation in Lyn-regulated pathways can mirror the complete loss of a single critical inhibitory molecule.

Although great progress has been made in delineating lung dendritic cell and lymphocyte subpopulations, similar advances in lung macrophages (MΦs) have been hampered by their intrinsic autofluorescence, cell plasticity, and the complexities of monocyte-MΦ compartmentalization. Using spectral scanning, we define alveolar MΦ autofluorescence characteristics, which has allowed us to develop an alternative flow cytometry method. Using this methodology, we show that mouse lung MΦs form distinct subpopulations during acute inflammation after challenge with LPS or influenza virus, and in chronic inflammatory lung disease consequent to SHIP-1 deletion. These subpopulations are distinguished by differential Mac-1 and CD11c integrin expression rather than classical M1 or M2 markers, and display differential gene signatures ex vivo. Whereas the resolution of acute inflammation is characterized by restoration to a homogenous population of CD11c(high)Mac-1(neg/low) MΦs reflective of lung homeostasis, chronic inflammatory lung disease associated with SHIP-1 deficiency is accompanied by an additional subpopulation of CD11c(high)Mac-1(pos) MΦs that tracks with lung disease in susceptible genetic background SHIP-1(-/-) animals and disease induction in chimeric mice. These findings may help better understand the roles of MΦ subpopulations in lung homeostasis and disease.

Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease characterized by the production of autoantibodies against nuclear components. Lyn-deficient mice are an excellent animal model of SLE manifesting clinical, pathological and biochemical features seen in the human disease. They develop autoreactive antibodies, glomerulonephritis and show generalized inflammation, and their B cells have a hyperactive phenotype. Since loss of Lyn confers hyper-activation of phosphoinositide 3-kinase (PI3K) signaling, we studied the effect of down-modulating PI3K in Lyn-deficient mice. We found that heterozygous inactivation of the p110δ isoform of PI3K was sufficient to restrain disease in Lyn-deficient mice, leading to significantly decreased autoantibody development and autoimmune-mediated kidney pathology, and improved survival. Intriguingly, haploinsufficiency of p110δ did not dampen signaling in Lyn-deficient B cells. However, plasma cell numbers, serum immunoglobulin titers, inflammation and T cell signaling and activation were significantly moderated in Lyn(-/-)p110δ(+/KD) mice. Importantly, we have shown that haploinsufficiency of p110δ has minor effects on the B cell compartment per se but leads to significant defects in T cell activation and B cell class-switching. These studies suggest that agents targeting p110δ PI3K need not achieve full blockade of the enzyme to be of great benefit in the treatment of SLE., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Background: Topical anesthesia of the upper airway is often recommended when difficulty in airway management is anticipated. There are published reports, however, of administration of topical anesthesia resulting in complete loss of airway control. Adverse effects are mostly attributed to interference with involuntary protective airway reflexes, while gross motor function itself generally is thought to be preserved. We hypothesized that if motor control is affected, measurable quantitative changes in vocalization should follow the use of topical anesthesia., Methods: A prospective, crossover, randomized, double-blind study was conducted, in which 24 healthy volunteers each performed 2 vocal exercises, while having their glottic appearance recorded digitally via fiberoptic nasendoscopy. Subjects gargled with 3 test solutions on separate occasions (placebo, 2% lidocaine, and 4% lidocaine) and repeated the vocal exercises and nasendoscopy. The angle between the vocal cords was measured using MB-Ruler®, and the Laryngograph Speech Studio® software was used for vocal parameter analysis., Results: The only significant changes in voice quality occurred between the control and test groups (P = 0.014). No difference could be found between the placebo and lidocaine groups., Conclusions: Although gargling with local anesthetic affected vocalization, no pharmacological effect attributable to local anesthetic was observed.

Alternatively activated M2 macrophages are implicated as both regulators and agents of lung disease, but their control is poorly understood. SHIP-1 is a 5' inositol phosphatase that negatively regulates the PI3K signaling pathway implicated in inflammation. SHIP-1-deficient mice have defects in hematopoiesis and B cell development, and die prematurely due to consolidation of lungs with M2-skewed macrophages. SHIP-1 is thought to restrain M2 macrophage polarization, with deregulated M2 skewing coinciding with severe lung disease in SHIP-1-deficient mice. To determine the influence of genetic background on the lung phenotype in SHIP-1(-/-) mice, we backcrossed the SHIP-1 null mutation onto C57BL/6 (Th2-resistant) and BALB/c (Th2-prone) backgrounds. Remarkably, we found that inflammatory lung disease was severe in C57.SHIP-1(-/-) mice, but absent in BALB.SHIP-1(-/-) mice. C57.SHIP-1(-/-), but not BALB.SHIP-1(-/-) mice had greatly increased myeloid progenitors, myeloid hyperplasia, markedly enhanced numbers of activated alveolar macrophages, and elevated amounts of Th2 and proinflammatory cytokines in bronchoalveolar lavage fluid and serum, suggesting that deregulated cytokine production induced disease. C57.SHIP-1(-/-) mice also developed severe B cell-dependent autoimmune disease, which was markedly attenuated on the BALB/c background. These data demonstrate that, contrary to current concepts, loss of SHIP-1 alone is not sufficient to cause lung inflammation, with disease only manifest on a permissive genetic background. This finding questions the nature of the lung disease in SHIP-1(-/-) mice, suggesting that its M2 classification is not strictly correct. Future identification of disease-promoting loci might reveal determinants of comorbid lung disease and autoimmunity and uncover potentially useful therapeutic targets.

In a recessive ENU mutagenesis screen for embryonic lethality, we identified a mouse pedigree with a missense mutation of SHIP1 (SHIP1(el20)) leading to an amino acid substitution I641T in the inositol-5'-phosphatase domain that represses phosphatidylinositol-3-kinase signaling. Despite detectable expression of functional SHIP1 protein, the phenotype of homozygous SHIP1(el20/el20) mice was more severe than gene-targeted SHIP1-null (SHIP1(-/-)) mice. Compared with age-matched SHIP1(-/-) mice, 5-week-old SHIP1(el20/el20) mice had increased myeloid cells, serum IL-6 levels, marked reductions in lymphoid cells, and died by 7 weeks of age with infiltration of the lungs by activated macrophages. Bone marrow transplantation demonstrated that these defects were hematopoietic-cell-autonomous. We show that the el20 mutation reduces expression in SHIP1(el20/el20) macrophages of both SHIP1 and s-SHIP, an isoform of SHIP1 generated by an internal promoter. In contrast, SHIP1(-/-) macrophages express normal levels of s-SHIP. Compound heterozygous mice (SHIP1(-/el20)) had the same phenotype as SHIP1(-/-) mice, thus providing genetic proof that the more severe phenotype of SHIP1(el20/el20) mice is probably the result of concomitant loss of SHIP1 and s-SHIP. Our results suggest that s-SHIP synergizes with SHIP1 for suppression of macrophage activation, thus providing the first evidence for a role of s-SHIP in adult hematopoiesis.

mTOR complex 1 (mTORC1; mammalian target of rapamycin [mTOR] in complex with raptor) is a key regulator of protein synthesis and cell growth in response to nutrient amino acids. Here we report that inositol polyphosphate multikinase (IPMK), which possesses both inositol phosphate kinase and lipid kinase activities, regulates amino acid signaling to mTORC1. This regulation is independent of IPMK's catalytic function, instead reflecting its binding with mTOR and raptor, which maintains the mTOR-raptor association. Thus, IPMK appears to be a physiologic mTOR cofactor, serving as a determinant of mTORC1 stability and amino acid-induced mTOR signaling. Substances that block IPMK-mTORC1 binding may afford therapeutic benefit in nutrient amino acid-regulated conditions such as obesity and diabetes., (Copyright © 2011 Elsevier Inc. All rights reserved.)

The second messenger phosphatidylinositol (3,4,5)-trisphosphate (PIP(3)), formed by the p110 family of PI3-kinases, promotes cellular growth, proliferation, and survival, in large part by activating the protein kinase Akt/PKB. We show that inositol polyphosphate multikinase (IPMK) physiologically generates PIP(3) as well as water soluble inositol phosphates. IPMK deletion reduces growth factor-elicited Akt signaling and cell proliferation caused uniquely by loss of its PI3-kinase activity. Inhibition of p110 PI3-kinases by wortmannin prevents IPMK phosphorylation and activation. Thus, growth factor stimulation of Akt signaling involves PIP(3) generation through the sequential activations of the p110 PI3-kinases and IPMK. As inositol phosphates inhibit Akt signaling, IPMK appears to act as a molecular switch, inhibiting or stimulating Akt via its inositol phosphate kinase or PI3-kinase activities, respectively. Drugs regulating IPMK may have therapeutic relevance in influencing cell proliferation.

Lyn kinase, a member of the Src family of tyrosine kinases, functions as both a positive and negative regulator of B cell activation. In the absence of Lyn, BCR signaling is unregulated, leading to perturbed B cell development, hyperactive B cells, and lethal Ab-mediated autoimmune disease. We have generated a mutant mouse pedigree, termed Mld4, harboring a novel mutation in the gene encoding Lyn, which renders the protein devoid of kinase activity. Despite similarities between the phenotypes of Lyn(Mld4/Mld4) and Lyn(-/-) mice, the spectrum of defects in Lyn(Mld4/Mld4) mice is less severe. In particular, although defects in the B cell compartment are similar, splenomegaly, myeloid expansion, and autoantibody production, characteristic of Lyn(-/-) mice, are absent or mild in Lyn(Mld4/Mld4) mice. Critically, immune complex deposition and complement activation in Lyn(Mld4/Mld4) glomeruli do not result in fulminant glomerulonephritis. Our data suggest that BCR hypersensitivity is insufficient for the development of autoimmune disease in Lyn(-/-) mice and implicate other cell lineages, particularly proinflammatory cells, in autoimmune disease progression. Furthermore, our results provide evidence for an additional role for Lyn kinase, distinct from its catalytic activity, in regulating intracellular signaling pathways.

Background: In 2005, guidance on how to prevent wrong site surgery in the form of a national safety alert was issued to all NHS hospital trusts in England and Wales by the National Patient Safety Agency., Objective: To investigate the response to the alert among clinicians in England and Wales 12-15 months after it had been issued., Methods: A before-after study, using telephone/face-to-face interviews with consultant surgeons and senior nurses in ophthalmology, orthopaedics and urology in 11 NHS hospitals in England & Wales in the year prior to the alert and 12-15 months after. The interviews were coded and analysed thematically., Results: The study revealed marked heterogeneity in organisational processes in response to a national alert. There was a significant change in surgeons' self-reported practice, with only 48% of surgeons routinely marking patients prior to the alert and 85% after (p<0.001). However, inter-specialty differences remained and change in practice was not always matched by change in attitude. Compliance with the detailed recommendations about how marking should be carried out was inconsistent. There were unintended consequences in terms of greater bureaucracy and concerns about diffusion of responsibility and hastily performed marking to enable release of patients from wards., Conclusion: The alert was effective in promoting presurgical marking and encouraging awareness of safety issues in relation to correct site surgery. However, care should be taken to monitor unintended consequences and whether change is sustained. Greater flexibility for local adaptation coupled with better design and early testing of safety alerts prior to national dissemination may facilitate more sustainable changes in practice.

Background: Hip fractures are common in the elderly and have a high 30 day postoperative mortality. The ability to recognize patients at high risk of poor outcomes before operation would be an important clinical advance. This study has determined key prognostic factors predicting 30 day mortality in a hip fracture population, and incorporated them into a scoring system to be used on admission., Methods: A cohort study was conducted at the Queen's Medical Centre, Nottingham, over a period of 7 yr. Complete data were collected from 4967 patients and analysed. Forward univariate logistic regression was used to select the independent predictor variables of 30 day mortality, and then multivariate logistic regression was applied to the data to construct and validate the scoring system., Results: The variables found to be independent predictors of mortality at 30 days were: age (66-85 yr, > or =86 yr), sex (male), number of co-morbidities (> or =2), mini-mental test score (< or =6 out of 10), admission haemoglobin concentration (< or =10 g dl(-1)), living in an institution, and presence of malignant disease. These variables were subsequently incorporated into a risk score, the Nottingham Hip Fracture Score. The number of deaths observed at 30 days, and the number of deaths predicted by the scoring system, indicated good concordance (chi(2) test, P=0.79). The area (SE) under the receiver operating characteristic curve was 0.719 (0.018), which demonstrated a reasonable predictive value for the score., Conclusions: We have developed and validated a scoring system that reliably predicts the probability of mortality at 30 days for patients after hip fracture.

Phosphoinositide (PI) 3-kinases play an important role in regulating the adhesive function of a variety of cell types through affinity modulation of integrins. Two type I PI 3-kinase isoforms (p110 beta and p110 gamma) have been implicated in G(i)-dependent integrin alpha(IIb)beta(3) regulation in platelets, however, the mechanisms by which they coordinate their signaling function remains unknown. By employing isoform-selective PI 3-kinase inhibitors and knock-out mouse models we have identified a unique mechanism of PI 3-kinase signaling co-operativity in platelets. We demonstrate that p110 beta is primarily responsible for G(i)-dependent phosphatidylinositol 3,4-bisphosphate (PI(3,4)P(2)) production in ADP-stimulated platelets and is linked to the activation of Rap1b and AKT. In contrast, defective integrin alpha(IIb)beta(3) activation in p110 gamma(-/-) platelets was not associated with alterations in the levels of PI(3,4)P(2) or active Rap1b/AKT. Analysis of the effects of active site pharmacological inhibitors confirmed that p110 gamma principally regulated integrin alpha(IIb)beta(3) activation through a non-catalytic signaling mechanism. Inhibition of the kinase function of PI 3-kinases, combined with deletion of p110 gamma, led to a major reduction in integrin alpha(IIb)beta(3) activation, resulting in a profound defect in platelet aggregation, hemostatic plug formation, and arterial thrombosis. These studies demonstrate a kinase-independent signaling function for p110 gamma in platelets. Moreover, they demonstrate that the combined catalytic and non-catalytic signaling function of p110 beta and p110 gamma is critical for P2Y(12)/G(i)-dependent integrin alpha(IIb)beta(3) regulation. These findings have potentially important implications for the rationale design of novel antiplatelet therapies targeting PI 3-kinase signaling pathways.

Cervical spine injuries in paediatric patients following trauma are extremely rare. The National Emergency X-Radiography Utilization Study (NEXUS) guidelines are a set of clinical criteria used to guide physicians in identifying trauma patients requiring cervical spine imaging. It is validated for use in children. A case of a child who did not fulfil the NEXUS criteria for imaging but was found to have a cervical spine fracture is reported.