Apoptosis and inflammation

During the last few decades it has been recognized that cell death is not the consequence of accidental injury, but is the expression of a cell suicide programme. Kerr et al. (1972) introduced the term apoptosis. This form of cell death is under the influence of hormones, growth factors and cytokines, which depending upon the receptors present on the target cells, may activate a genetically controlled cell elimination process. During apoptosis the cell membrane remains intact and the cell breaks into apoptotic bodies, which are phagocytosed. Apoptosis, in contrast to necrosis, is not harmful to the host and does not induce any inflammatory reaction. The principal event that leads to inflammatory disease is cell damage, induced by chemical/physical injury, anoxia or starvation. Cell damage means leakage of cell contents into the adjacent tissues, resulting in the capillary transmigration of granulocytes to the injured tissue. The accumulation of neutrophils and release of enzymes and oxygen radicals enhances the inflammatory reaction. Until now there has been little research into the factors controlling the accumulation and the tissue load of granulocytes and their histotoxic products in inflammatory processes. Neutrophil apoptosis may represent an important event in the control of intlamtnation. It has been assumed that granulocytes disintegrate to apoptotic bodies before their fragments are removed by local macrophages. Removal of neutrophils from the inflammatory site without release of granule contents is of paramount importance for cessation of inflammation. In conclusion, apoptotic cell death plays an important role in inflammatory processes and in the resolution of inflammatory reactions. The facts known at present should stimulate further research into the role of neutrophil, eosinophil and macrophage apoptosis in inflammatory diseases

Elevated plasma levels of endothelin are associated with the severity of sepsis and presence of shock in contrast to the levels of atrial natriuretic peptide

Immunoreactive endothelin (ETi) and atrial natriuretic peptide (ANPi) blood levels were measured by radioimmunoassay in patients with clinically defined sepsis. The interaction between these two peptides and their relation to circulatory shock and mortality were studied. All septic patients (n = 16) had significantly higher ETi (22.3 ± 11.1 pg/ml) and ANPi (398.3 ± 154.3 pg/ml) plasma concentrations compared to control subjects (ETi, 4.1 ± 1.2; ANPi, 59.1 ± 14.8 pg/ml; n = 13). ETi levels followed the severity of illness according to the APACHE II scoring system and were higher in patients who did not survive. ETi levels were significantly higher in the presence of shock and bacteraemia. Furthermore, ETi correlated well with plasma lactate (r = 0.83, p < 0.05), but not with renal function. ANPi levels did not show correlation with any of these determinants. Serial blood sampling, six consecutive days after admission, showed that ETi levels gradually decreased in normotensive patients in contrast to patients with septic shock. ANPi levels did not show systematic changes in time, and no relationship was observed between ETi and ANPi levels. These results suggest that plasma ETi levels are indicative for disease severity and might have prognostic significance. The role of ANPi during sepsis remains to be eludicated

Apoptotic cell death, detected ex vivo in peripheral blood lymphocytes of HIV-1 infected persons

In HIV-1 infection the ongoing depletion of CD4+ T-lymphocytes is believed, to a large extent, to be due to apoptosis. Until now quantitative information about in vivo apoptosis of lymphocytes in HIV-patients is scarce because of the very nature of the apoptotic process. Successful detection of apoptosis ex vivo requires the recognition of the initial phase of this process, because at a later stage the cells may not remain any longer in the circulation. We measured quantitatively the amount of early apoptotic peripheral blood lymphocytes directly ex vivo in HIV-1 infected patients using a recently described flow cytometric assay. With this method we observed in an unselected heterogenous group of twelve HIV-infected individuals a median percentage of apoptotic lymphocytes to be significantly higher than in ten healthy controls. To the best of our knowledge this is the first report of ex vivo observed increased apoptosis of peripheral blood lymphocytes in HIV-infected persons

Black Sheep, Outsiders, and the Qumran Movement : Social-Psychological Perspectives on Norm-Deviant Behaviour

Peer reviewe

Six-month outcomes from a randomized trial augmenting serotonin reuptake inhibitors with exposure and response prevention or risperidone in adults with obsessive-compulsive disorder

OBJECTIVE: To compare outcomes after 6-month maintenance treatment of adults diagnosed with obsessive-compulsive disorder (OCD) based on DSM-IV criteria who responded to acute treatment with serotonin reuptake inhibitors (SRIs) augmented by exposure and response prevention (EX/RP) or risperidone. METHOD: A randomized trial was conducted at 2 academic sites from January 2007 through December 2012. In the acute phase, 100 patients on therapeutic SRI dose with at least moderate OCD severity were randomized to 8 weeks of EX/RP, risperidone, or pill placebo. Responders entered the 6-month maintenance phase, continuing the augmentation strategy they received acutely (n = 30 EX/RP, n = 8 risperidone). Independent evaluations were conducted every month. The main outcome was the Yale-Brown Obsessive Compulsive Scale (Y-BOCS). RESULTS: Intent-to-treat analyses indicated that, after 6-month maintenance treatment, EX/RP yielded OCD outcomes that were superior to risperidone (Y-BOCS = 10.95 vs 18.70; t40 = 2.76, P = .009); more patients randomized to EX/RP met response criteria (Y-BOCS decrease \u3e/= 25%: 70% vs 20%; P \u3c .001) and achieved minimal symptoms (Y-BOC

Fluorescence characterization of clinically-important bacteria

Healthcare-associated infections (HCAI/HAI) represent a substantial threat to patient health during hospitalization and incur billions of dollars additional cost for subsequent treatment. One promising method for the detection of bacterial contamination in a clinical setting before an HAI outbreak occurs is to exploit native fluorescence of cellular molecules for a hand-held, rapid-sweep surveillance instrument. Previous studies have shown fluorescence-based detection to be sensitive and effective for food-borne and environmental microorganisms, and even to be able to distinguish between cell types, but this powerful technique has not yet been deployed on the macroscale for the primary surveillance of contamination in healthcare facilities to prevent HAI. Here we report experimental data for the specification and design of such a fluorescence-based detection instrument. We have characterized the complete fluorescence response of eleven clinically-relevant bacteria by generating excitation-emission matrices (EEMs) over broad wavelength ranges. Furthermore, a number of surfaces and items of equipment commonly present on a ward, and potentially responsible for pathogen transfer, have been analyzed for potential issues of background fluorescence masking the signal from contaminant bacteria. These include bedside handrails, nurse call button, blood pressure cuff and ward computer keyboard, as well as disinfectant cleaning products and microfiber cloth. All examined bacterial strains exhibited a distinctive double-peak fluorescence feature associated with tryptophan with no other cellular fluorophore detected. Thus, this fluorescence survey found that an emission peak of 340nm, from an excitation source at 280nm, was the cellular fluorescence signal to target for detection of bacterial contamination. The majority of materials analysed offer a spectral window through which bacterial contamination could indeed be detected. A few instances were found of potential problems of background fluorescence masking that of bacteria, but in the case of the microfiber cleaning cloth, imaging techniques could morphologically distinguish between stray strands and bacterial contamination

Effector cell mediated cytotoxicity measured by intracellular Granzyme B release in HIV infected subjects

CD8+ cytotoxic T lymphocyte (CTL) activity is currently believed to be one of the key immunologic mechanisms responsible for the prevention or attenuation of HIV-1 infection. The induction of CD8+ T cell activation may also result in the production of soluble or non-classical lytic factors that are associated with protection from infection or slower disease progression. Traditionally, CD8+ CTL responses have been measured by the classic chromium release assay, monitoring the ability of T cells (Effector cells) to lyse radiolabelled HLA – matched “target cells” that express the appropriate antigen-MHC complex. This method is not only labor intensive, semi quantitative assay at best, but also needs fresh, non-cryopreserved cells. Recently, cytokine specific ELISPOT assays or tetrameric MHC-I/ peptide complexes have utilized to directly quantitate circulating CD8+ effector cells, and these assays are more sensitive, quantitative and reproducible than the traditional CTL lysis assay and can also be performed on cryopreserved cells. Although these are reproducible assays for the assessment of soluble antiviral activity secreted by activated T cell populations they can be extremely expensive to perform. We have used FACS Analysis to measure Granzyme B release as a function of cell mediated cytotoxicity. This method helps quantitate the CTL activity and also identifies the phenotype of the cells elucidating this immune response. The method described not only monitors immunological response but also is also simple to perform, precise and extremely time efficient and is ideal for screening a large number of samples

Apoptosis- and necrosis-induced changes in light attenuation measured by optical coherence tomography

Optical coherence tomography (OCT) was used to determine optical properties of pelleted human fibroblasts in which necrosis or apoptosis had been induced. We analysed the OCT data, including both the scattering properties of the medium and the axial point spread function of the OCT system. The optical attenuation coefficient in necrotic cells decreased from 2.2 ± 0.3 mm−1 to 1.3 ± 0.6 mm−1, whereas, in the apoptotic cells, an increase to 6.4 ± 1.7 mm−1 was observed. The results from cultured cells, as presented in this study, indicate the ability of OCT to detect and differentiate between viable, apoptotic, and necrotic cells, based on their attenuation coefficient. This functional supplement to high-resolution OCT imaging can be of great clinical benefit, enabling on-line monitoring of tissues, e.g. for feedback in cancer treatment

Inhibition of GATA2 restrains cell proliferation and enhances apoptosis and chemotherapy mediated apoptosis in human GATA2 overexpressing AML cells

GATA2, a zinc finger transcription factor predominantly expressed in hematopoietic cells, acts as an essential regulator of hematopoietic stem cell generation, survival and functionality. Loss and gain of GATA2 expression has been implicated in myelodysplastic syndrome and acute myeloid leukemia (AML) yet the precise biological impact of GATA2 expression on human AML cell fate decisions remains ambiguous. Herein, we performed large-scale bioinformatics that demonstrated relatively frequent GATA2 overexpression in AML patients as well as select human AML (or AML-like) cell lines. By using shRNAi to target GATA2 in these AML cell lines, and an AML cell line expressing normal levels of GATA2, we found that inhibition of GATA2 caused attenuated cell proliferation and enhanced apoptosis exclusively in AML cell lines that overexpress GATA2. We proceeded to pharmacologically inhibit GATA2 in concert with AML chemotherapeutics and found this augmented cell killing in AML cell lines that overexpress GATA2, but not in an AML cell line expressing normal levels of GATA2. These data indicate that inhibition of GATA2 enhances chemotherapy-mediated apoptosis in human AML cells overexpressing GATA2. Thus, we define novel insights into the oncogenic role of GATA2 in human AML cells and suggest the potential utilization of transient GATA2 therapeutic targeting in AML