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

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

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

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

A ritka kromoszóma-rendellenességek és a fetoplacentaris mozaikosság jelentősége a praenatalis diagnosztikában a nem invazív szűrővizsgálatok tükrében

Introduction and objective: To determine the prevalence of microscopically visible de novo atypical chromosomal aberrations and fetoplacental mosaicism in a prenatal tertial referral center, and to investigate the maternal and fetal characteristics in connection with genomewide non-invasive prenatal screening. Method: Retrospective cohort study from 2014 to 2019 of pregnancies with invasive genetic analysis. Results: In the cohort of 2504 cases, the proportion of CVS was 53.3%. We diagnosed 200 chromosomal aberrations (8%), including 13.5% of de novo rare chromosomal aberrations (14 rare autosomal trisomies, 6 polyploidies, 5 structural aberrations and 2 small supernumerary marker chromosomes). The rate of fetoplacental mosaicism was 12.4%/77.8% in common/atypical chromosomal aberrations (p = 0.001) and confined to placenta in 17/40 cases. Associated ultrasound abnormalities were abnormal nuchal translucency and major malformations in 58% and 35% with common trisomies and 11% (p = 0.006) and 67% (p = 0.047) with true mosaic cases of rare abnormalities, respectively. Major ultrasound malformation was facial dysmorphism with rare aberrations. Potential application of genomewide non-invasive prenatal test in atypical chromosomal aberrations presumably would have been false-positive in 12 cases (44%), false-negative in 1 case (3.7%), and would have early detected 2 cases of rare autosomal trisomies (7.4%) without ultrasound anomalies. Conclusion: Structural ultrasound malformations with normal nuchal translucency thickness may be indicative of rare chromosomal aberrations. Application of genomewide non-invasive prenatal test is an alternative of early diagnostic methods with a potential of discordant results due to mosaicism. Knowledge about the presence of fetoplacental mosaicism influences risk estimation and genetic counseling, detailed cytogenetic characterization is of utmost importance. © Szerző(k)

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

Coronary MR angiography at 3T: fat suppression versus water-fat separation

Objectives: To compare Dixon water-fat suppression with spectral pre-saturation with inversion recovery (SPIR) at 3T for coronary magnetic resonance angiography (MRA) and to demonstrate the feasibility of fat suppressed coronary MRA at 3T without administration of a contrast agent. Materials and methods: Coronary MRA with Dixon water-fat separation or with SPIR fat suppression was compared on a 3T scanner equipped with a 32-channel cardiac receiver coil. Eight healthy volunteers were examined. Contrast-to-noise ratio (CNR), signal-to-noise ratio (SNR), right coronary artery (RCA), and left anterior descending (LAD) coronary artery sharpness and length were measured and statistically compared. Two experienced cardiologists graded the visual image quality of reformatted Dixon and SPIR images (1: poor quality to 5: excellent quality). Results: Coronary MRA images in healthy volunteers showed improved contrast with the Dixon technique compared to SPIR (CNR blood-fat: Dixon = 14.9 ± 2.9 and SPIR = 13.9 ± 2.1; p = 0.08, CNR blood-myocardium: Dixon = 10.2 ± 2.7 and SPIR = 9.11 ± 2.6; p = 0.1). The Dixon method led to similar fat suppression (fat SNR with Dixon: 2.1 ± 0.5 vs. SPIR: 2.4 ± 1.2, p = 0.3), but resulted in significantly increased SNR of blood (blood SNR with Dixon: 19.9 ± 4.5 vs. SPIR: 15.5 ± 3.1, p < 0.05). This means the residual fat signal is slightly lower with the Dixon compared to the SIPR technique (although not significant), while the SNR of blood is significantly higher with the Dixon technique. Vessel sharpness of the RCA was similar for Dixon and SPIR (57 ± 7 % vs. 56 ± 9 %, p = 0.2), while the RCA visualized vessel length was increased compared to SPIR fat suppression (107 ± 21 vs. 101 ± 21 mm, p < 0.001). For the LAD, vessel sharpness (50 ± 13 % vs. 50 ± 7 %, p = 0.4) and vessel length (92 ± 46 vs. 90 ± 47 mm, p = 0.4) were similar with both techniques. Consequently, the Dixon technique resulted in an improved visual score of the coronary arteries in the water fat separated images of healthy subjects (RCA: 4.6 ± 0.5 vs. 4.1 ± 0.7, p = 0.01, LAD: 4.1 ± 0.7 vs. 3.5 ± 0.8, p = 0.007). Conclusions: Dixon water-fat separation can significantly improve coronary artery image quality without the use of a contrast agent at 3T

Endogenous Galectin-9 Suppresses Apoptosis in Human Rheumatoid Arthritis Synovial Fibroblasts

Galectin-9 (Gal9) has been postulated to have anti-infammatory properties based on the ability of exogenous Gal9 to induce apoptosis in synovial fbroblasts in animal models of rheumatoid arthritis (RA). Here we aimed to assess the potential role of endogenous Galectins, including Gal9, in the infammatory pathology of the RA synovium in humans. Firstly expression of Galectins 1–9 was determined in synovial fbroblasts (RASF) and dermal fbroblasts (DF) isolated from RA patients, the latter representing a non-infamed site. We then further challenged the cells with pro-infammatory TLR agonists and cytokines and assessed Galectin expression. Gal9 was found to be diferentially and abundantly expressed in RASF compared to DF. Agonists of TLR3 and TLR4, along with IFNgamma were also found to induce Gal9 expression in RASF. siRNA was then used to knock-down Gal9 expression in RASF and the efects of this on apoptosis and cell viability were assessed. Increased apoptosis was observed in RASF following Gal9 knock-down. We conclude that, unlike exogenous Gal9, endogenous Gal9 is protective against apoptosis and enhances synovial fbroblast viability suggesting that its role in RA is both pathogenic and pro-infammatory