IFN-gamma protects short-term ovarian carcinoma cell lines from CTL lysis via a CD94/NKG2A-dependent mechanism

IFN-gamma regulates the immunogenicity of target cells by increasing their expression of HLA class I molecules. This facilitates the T cell receptor-mediated recognition by CD8(+) T cells but decreases target cell sensitivity to lysis by NK cells due to engagement of inhibitory NK receptors. In this study, short-term tumor cell lines from patients with advanced ovarian carcinomas were established. We demonstrate the paradoxical finding that IFN-gamma treatment of these short-term ovarian carcinoma cell lines (OVACs) resulted in resistance of tumor cells to lysis by peptide- and allospecific CD8(+) T cells. Blocking experiments revealed that this phenomenon was dependent on enhanced inhibitory signalling via CD94/NKG2A receptors expressed on the effector cells. This was associated with increased expression of HLA-E mRNA and HLA-G at the protein level in IFN-gamma-treated OVACs. Furthermore, pulsing of untreated OVACs with the leader sequence peptide of HLA-G protected these cells from lysis by CTLs, thus mimicking the inhibitory effect of IFN-gamma. This study provides evidence that CD94/NKG2A receptors play an important role in regulating T cell activity against tumors and shows that IFN-gamma modulation of target cells may shift the balance of triggering and inhibitory signals to T cells, turning off their cytolytic activity.     

IFN-gamma regulates Fas ligand expression in human CD4+ T lymphocytes and controls their anti-mycobacterial cytotoxic functions

Fas and Fas Ligand (FasL) expression, activation-induced cell death (AICD) and mycobacterial antigen-specific cytotoxicity of peripheral T cells from patients with complete inherited IFN-gamma receptor 1 binding chain deficiency (IFN-gammaR1-/-) were investigated. Fas was equally expressed in both normal and deficient T lymphoblasts and they underwent apoptosis when stimulated with agonist anti-Fas mAb. By contrast, T lymphoblasts and CD4+ T cell clones (TCC) from deficient patients displayed a reduced surface FasL expression and resistance to AICD. CD8+ TCC from healthy and deficient patients displayed similar high level of FasL and susceptibility to AICD. In Jurkat CD4+ T cells competent to transduce IFN-gamma signaling, IFN-gamma induced surface FasL export and their Fas-dependent apoptosis. Effector T cells generated from a patient with a dominant negative mutation of IFN-gammaR1 (IFN-gammaR1DN) following stimulation with mycobacterial antigens were unable to kill MHC class II-matched, mycobacterial antigen-pulsed macrophages. Normal Fas expression in T cells and FasL in CD8+ cells may account for the absence of autoimmune disorders in these patients. Conversely, defective FasL expression on IFN-gammaR1DN CD4+ T cells impairs their cytotoxic functions and highlights a novel role for IFN-gamma signaling in the control of mycobacterial infection in humans.     

Component-resolved diagnosis in selecting patients for yellowjacket venom immunotherapy

Background

Venom immunotherapy is effective in preventing systemic allergic reactions (SARs), but the diagnosis of venom allergy is problematic.

Erythropoietin attenuates the tissue injury associated with hemorrhagic shock and myocardial ischemia

Here we investigate the effects of erythropoietin (EPO) on the tissue/organ injury caused by hemorrhagic shock (HS), endotoxic shock, and regional myocardial ischemia and reperfusion in anesthetized rats. Male Wistar rats were anesthetized with thiopental sodium (85 mg/kg i.p.) and subjected to hemorrhagic shock (HS; i.e., mean arterial blood pressure reduced to 45 mmHg for 90 min, followed by resuscitation with shed blood for 4 h), endotoxemia (for 6 h), or left anterior descending coronary artery occlusion (25 min) and reperfusion (2 h). HS and endotoxemia resulted in renal dysfunction and liver injury. Administration of EPO (300 IU/kg i.v., n = 10) before resuscitation abolished the renal dysfunction and liver injury in hemorrhagic, but not endotoxic, shock. HS also resulted in significant increases in the kidney of the activities of caspases 3, 8, and 9. This increase in caspase activity was not seen in HS rats treated with EPO. In cultured human proximal tubule cells, EPO concentration-dependently reduced the cell death and increase in caspase-3 activity caused by either ATP depletion (simulated ischemia) or hydrogen peroxide (oxidative stress). In the heart, administration of EPO (300 IU/kg i.v., n = 10) before reperfusion also caused a significant reduction in infarct size. In cultured rat cardiac myoblasts (H9C2 cells), EPO also reduced the increase in DNA fragmentation caused by either serum deprivation (simulated ischemia) or hydrogen peroxide (oxidative stress). We propose that the acute administration of EPO on reperfusion and/or resuscitation will reduce the tissue injury caused by ischemia-reperfusion of the heart (and other organs) and hemorrhagic shock.     

Diagnostic evaluation of planar and tomographic ventilation/perfusion lung images in patients with suspected pulmonary emboli

Planar lung ventilation/perfusion scintigraphy (V/P(PLANAR)) is a standard method for diagnosis of pulmonary embolism (PE). The goals of this study were to test whether the diagnostic information of ventilation/perfusion tomography (V/P(SPET)) applied in clinical routine might enhance information compared with V/P(PLANAR) and to streamline data processing for the demands of clinical routine. This prospective study includes 53 patients suspected for PE referred for lung scintigraphy. After inhalation of (99m)Tc-DTPA planar ventilation imaging was followed by tomography, using a dual-head gamma camera. (99m)Tc-MAA was injected i.v. for perfusion tomography followed by planar imaging. Patients were examined in supine position, unchanged during V/P tomography. Two reviewers evaluated V/P(PLANAR) and V/P(SPET) images separately and randomly. Mismatch points were calculated on the basis of extension of perfusion defects with preserved ventilation. Patients were followed up clinically for at least 6 months. With V/P(SPET) the number of patients with PE was higher and 53% more mismatch points were found. In V/P(SPET) interobserver variation was less compared with V/P(PLANAR). Ancillary findings were observed by both techniques in half of the patients but more precisely interpreted with V/P(SPET). V/P(SPET) shows more and better delineated mismatch defects, improved quantification and less interobserver variation compared with V/P(PLANAR). V/P(SPET) is amenable to implementation for clinical routine and suitable even when there is demand for a high patient throughput.     

Extracellular and intracellular barriers in non-viral gene delivery.

Complexes of DNA with cationic lipids and cationic polymers are frequently used for gene transfer. Extracellular interactions of the complexes with anionic glycosaminoglycans (GAGs) may interfere with gene transfer. Interactions of GAGs with carrier DNA complexes have been studied using tests for DNA relaxation (ethidium bromide intercalation), DNA release (electrophoresis), and transfection (pCMVbGal transfer into RAA smooth muscle cells). Several cationic lipid formulations (DOTAP, DOTAP/Chol, DOTAP/DOPE, DOTMA/DOPE, DOGS) and cationic polymers (fractured dendrimer, polyethylene imines 25 and 800 kDa, polylysines 20 and 200 kDa) were tested. Polycations condensed DNA more effectively than monovalent lipids. Hyaluronic acid did not release or relax DNA in any complex, but it inhibited transfection by some polyvalent systems (PEI, dendrimers, DOGS). Gene transfer by other carriers was not affected by hyaluronic acid. Sulfated GAGs (heparan sulfate, chondroitin sulfates B and C) completely blocked transfection, except in the case of liposomes with DOPE. Sulfated GAGs relaxed and released DNA from some complexes, but these events were not prerequisites for the inhibition of transfection. Furthermore, preliminary results suggest that cell surface GAGs, particularly heparan sulfate, inhibit gene transfer by cationic lipids and polymers.     

A safe place with space for learning: Experiences from an interprofessional training ward

Interprofessional learning in a real ward context effectively increases collaborative and professional competence among students. However, less is known on the processes behind this. The aim of this study was to explore medical, nurse, physiotherapy, and occupational therapy students’ perspectives on the process of their own learning at an interprofessional training ward (IPTW). We performed a qualitative content analysis on free-text answers of 333 student questionnaires from the years 2004 to 2011. Two main themes emerged: first, students found that the IPTW provided an enriching learning environment—a safe place with space. It included authentic and relevant patients, well-composed and functioning student teams, competent and supportive supervisors, and adjusted ward structures to support learning. Second, they developed an awareness of their own development with faith in the future—from chaos to clarity. It included personal, professional, and interprofessional development towards a comprehensive view of practice and a faith in their ability to work as professionals in the future. Our findings are discussed with a social constructivist perspective. This study suggests that when an IPTW provides a supportive and permissive learning environment with possibilities to interact with one another—a safe place with space—it enables students to move from insecurity to faith in their abilities—from chaos to clarity. However, if the learning environment is impaired, the students’ development could be halted.     

Paramedic field instructors: an approach to training the newest paramedics while maintaining the interest of the most successful senior paramedics

A unique approach to providing orientation and supervised field experience for newly graduated paramedics and for the continuing education and recognition of experienced, skilled, operational paramedics is described. A group of 30 paramedic field instructors (PFI) was selected following application, development of criteria for selection, and interviews. This program had a positive effect in both realms during its first year of operation in the emergency medical services system in which it was implemented.