Surface membrane glycoproteins of wild-type and differentiation-inducer- resistant HL-60 cells

Surface membrane glycoproteins (SMGs) of cells from the parental wild- type HL-60 cell line and from three sublines variably cross-resistant to the granulocyte differentiation-inducing effects of retinoic acid (RA), dimethylsulfoxide (DMSO), and certain purine bases (6-thioguanine [6TG] or hypoxanthine) were studied by one-dimensional and two- dimensional gel electrophoresis. After both oligosaccharide (periodate/borotritide) and peptide (1,3,4,6-tetrachloro-3 alpha, 6 alpha-diphenylglycouril) ectolabeling procedures, striking common changes were noted in the gel electrophoretic patterns of the SMGs from the RA- and 6TG-resistant sublines compared to those from the wild-type HL-60 line or the DMSO-resistant subline. Most prominently, this included the presence in the RA- and 6TG-resistant cells of an apparent high molecular weight acidic glycoprotein(s) (mol wt, 200 to 285 kilodaltons [kD]; isoelectric point range [pl], 4.5 to 6.0) not observed in the wild-type or DMSO-resistant cells and, conversely, the presence of a lower molecular weight glycoprotein(s) (mol wt, 120 to 165 kD; pl, 4.2 to 5.9) in the wild-type and DMSO-resistant cells, which was absent or much reduced in the RA- and 6TG-resistant cells. These acidic SMGs did not change as a function of the induction of granulocyte differentiation. However, some other more basic SMGs varied as a function of granulocyte differentiation in both the wild-type and differentiation inducer-resistant sublines, including the loss of the transferrin receptor and the gain of a mol wt 55- to 60-kD neutrophil- associated protein. In the context of previously reported information, these results indicate (1) that the overall pattern of SMG changes in the RA- and 6TG-resistant cells closely resembles that associated with multidrug (pleiotropic) resistance to cytotoxic agents in a variety of mammalian cells; (2) that the RA/6TG resistance-associated SMG changes are not granulocyte differentiation stage-specific; and (3) that either the RA/6TG resistance-associated SMG changes are not related to the resistance mechanism or they are involved in the resistance/cross- resistance mechanism(s) for RA/purine bases but not for DMSO.     

Impact of ladder‐related falls on the emergency department and recommendations for ladder safety

Objectives

To describe the characteristics of patients who presented to the ED from a ladder‐related fall and their injuries, highlight the impact of ladder‐related falls on the ED, identify contributing factors of ladder falls and draw recommendations to improve ladder safety.

Methods

A prospective observational study was conducted in two EDs. Patients’ demographics and ED services used were obtained from medical records. A 53‐item questionnaire was used to gather information about the type of ladder used, ladder activity, circumstances of the fall, contributing factors and future recommendations.

Results

A total of 177 patients were recruited for this study. The typical patient was male, over the age of 50 and using a domestic ladder. The ED length of stay was between 30 min and 16 h, and was longer if patients were transferred to the short stay unit. Services most utilised in the ED included diagnostic tests, procedures and referrals to other healthcare teams. Most falls occurred because of ladder movement and slips or misstep. The major contributing factors identified were a combination of user features and flaws in ladder setup.

Conclusions

Ladder‐related falls carry a considerable burden to the ED. Recommendations include ladder safety interventions that target ladder users most at risk of falls: men, ≥50 years old and performing domestic tasks. Safety interventions should emphasise task avoidance, education and training, utilisation of safety equipment and appropriate ladder setup.     

Simulated effect of pneumococcal vaccination in the Netherlands on existing rules constructed in a non-vaccinated cohort predicting sequelae after bacterial meningitis

Background  

Previously two prediction rules identifying children at risk of hearing loss and academic or behavioral limitations after bacterial meningitis were developed. Streptococcus pneumoniae as causative pathogen was an important risk factor in both. Since 2006 Dutch children receive seven-valent conjugate vaccination against S. pneumoniae. The presumed effect of vaccination was simulated by excluding all children infected by S. pneumoniae with the serotypes included in the vaccine, from both previous collected cohorts (between 1990-1995).     

Tissue reaction to sealing materials: different view at biocompatibility

The biodegradability of root canal sealers in areas other than the root canal system is crucial to the overall success rate of endodontic treatment. The aim of the present study was to investigate, the cell and tissue reaction to GuttaFlow and AHPlus, both in vitro and in vivo. For the in vitro experiments the materials were incubated with Human Periodontal Ligament Fibroblasts and cell proliferation and cytotoxicity analyses were performed. Additional fluorescence-microscope stainings were carried out in order to visualize cell growth and morphology. For assessment of the tissue reaction to the materials a subcutaneous implantation model in Wistar rats was employed and the inflammatory response to the materials was visualized by means of general and specific histology after 6 weeks. Human gingival fibroblasts proliferation seemed to be dependent upon dental material and cultivation time. After an incubation period of 96 hrs AHPlus proved to be significantly (p<0.002) more cytotoxic than GuttaFlow, as only a small number of fibroblasts survived on AHPlus. In vivo, GuttaFlow was surrounded by a fibrous capsule and no degradation took place, while AHPlus induced a well-vascularized granulation tissue in which the material was phagocyted by macrophages. The results of this study demonstrate that a potential cytotoxic effect of a sealing material may beneficial in order to have antibacterial properties and induce self degradation when accidentally extruded over the apical foramen.     

CXCR6 and CXCL16 in liver disease

BackgroundChemokines are small molecules that act through G-protein coupled receptors to mediate primarily lymphocyte migration. CXCL16, which interacts with only one receptor (CXCR6), can mediate lymphocyte recruitment and has been implicated in various disease conditions. Steatohepatitis, caused by metabolic syndrome or alcohol misuse, is the commonest cause of liver disease in the UK. We investigated the role of CXCL16 and CXCR6 in the development of steatohepatitis.MethodsExpression of CXCL16 in whole liver and isolated cells was investigated with real-time PCR and immunohistochemistry. Serum and supernatant concentrations of soluble CXCL16 were measured with ELISA. Expression of CXCR6 on lymphocytes was investigated with flow cytometry. Lymphocyte adhesion was assessed with freshly isolated lymphocytes from liver or peripheral blood flowed over confluent layer of isolated human hepatic sinusoidal endothelium (HSEC).FindingsWhole liver expression of CXCL16 was increased relative to normal liver in fatty liver disease with increasing expression seen with increasing steatohepatitis and fibrosis. Immunohistochemistry showed CXCL16 expressed throughout regenerative nodules in both alcoholic and non-alcoholic liver disease. Isolated HSEC, biliary epithelial cells, and hepatoma cell lines increased expression of CXCL16 and released soluble CXCR6 in response to pro-inflammatory cytokines, particularly the combination of tumour necrosis factor α and interferon γ. Peripheral blood lymphocyte CXCR6 expression was confined to CD4 cells; however in the liver CD8+ cells and CD56+ cells more commonly expressed CXCR6. Inhibition of CXCR6 or CXCL16 inhibited transmigration of lymphocytes across HSEC.InterpretationCXCL16 is expressed in diseased liver where it has a role in the transmigration of lymphocytes across endothelium. This may represent a new therapeutic target in liver disease.FundingUK Medical Research Council.     

Involvement of PKCα and G-protein-coupled receptor kinase 2 in agonist-selective desensitization of μ-opioid receptors in mature brain neurons

Background and purpose:

The ability of an agonist to induce desensitization of the µ-opioid receptor (MOR) depends upon the agonist used. Furthermore, previous data suggest that the intracellular mechanisms underlying desensitization may be agonist-specific. We investigated the mechanisms underlying MOR desensitization, in adult mammalian neurons, caused by morphine (a partial agonist in this system) and DAMGO (a high-efficacy agonist).

Experimental approach:

MOR function was measured in locus coeruleus neurons, by using whole-cell patch-clamp electrophysiology, in rat and mouse brain slices (both wild-type and protein kinase C (PKC)α knockout mice). Specific isoforms of PKC were inhibited by using inhibitors of the receptors for activated C-kinase (RACK), and in vivo viral-mediated gene-transfer was used to transfect neurons with dominant negative mutants (DNMs) of specific G-protein-coupled receptor kinases (GRKs).

Key results:

Morphine-induced desensitization was attenuated by using RACK inhibitors that inhibit PKCα, but not by other isoform-specific inhibitors. Further, the PKC component of morphine-induced desensitization was absent in locus coeruleus neurons from PKCα knockout mice. The PKC-enhanced morphine-induced desensitization was not affected by over-expression of a GRK2 dominant negative mutant (GRK2 DNM). In contrast, DAMGO-induced MOR desensitization was independent of PKC activity but was reduced by over-expression of the GRK2 DNM but not by that of a GRK6 DNM.

Conclusions and implications:

In mature mammalian neurons, different MOR agonists can induce MOR desensitization by different mechanisms, morphine by a PKCα-mediated, heterologous mechanism and DAMGO by a GRK-mediated, homologous mechanism. These data represent functional selectivity at the level of receptor desensitization.