Phagocytosis and digestion of pH-sensitive fluorescent dye (Eos-FP) transfected E. coli in whole blood assays from patients with severe sepsis and septic shock

The function of phagocytic and antigen presenting cells is of crucial importance to sustain immune competence against infectious agents as well as malignancies. We here describe a reproducible procedure for the quantification of phagocytosis by leukocytes in whole blood. For this, a pH-sensitive green-fluorescent protein- (GFP) like dye (Eos-FP) is transfected into infectious microroganisms. After UV-irradiation, the transfected bacteria emit green (≈5160 nm) and red (≈581 nm) fluorescent light at 490 nm excitation. Since the red fluorescent light is sensitive to acidic pH, the phagocytosed bacteria stop emitting red fluorescent light as soon as the phagosomes fuse with lysosomes. The green fluorescence is maintained in the phagolysosome until pathogen degradation is completed. Fluorescence emission can be followed by flow cytometry with filter settings documenting fluorescence 1 (FL 1, FITC) and fluorescence 2 (FL 2, phycoerythrin, PE). Eos-FP transfected bacteria can also be traced within phagocytes using microscopical techniques. A standardized assay has been developed which is suitable for clinical studies by providing clinicians with syringes pre-filled with fixed and appropriately UV-irradiated Eos-FP E. coli (TruCulture™). After adding blood or body fluids to these containers and starting the incubation at 37°C, phagocytosis by granulocytes proceeds over time. Cultures can be terminated at a given time by lysing red blood cells followed by flow cytometry. A pilot study demonstrated that Eos-FP E. coli phagocytosis and digestion was up-regulated in the majority of patients with either severe sepsis or septic shock as compared to healthy donors (p < 0.0001 after o/n incubation). Following treatment with recombinant human granulocyte colony-stimulating factor (rhG-CSF) in selected patients with sepsis, phagolysosome fusion appeared to be accelerated

Genetic Variation Shapes Protein Networks Mainly through Non-transcriptional Mechanisms

Variation in the levels of co-regulated proteins that function within networks in an outbred yeast population is not driven by variation in the corresponding transcripts

Latent class analysis of sexual health markers among men and women participating in a British probability sample survey.

BACKGROUND: Despite known associations between different aspects of sexual health, it is not clear how patterning of adverse sexual health varies across the general population. A better understanding should contribute towards more effective problem identification, prevention and treatment. We sought to identify different clusters of sexual health markers in a general population, along with their socio-demographic, health and lifestyle correlates. METHODS: Data came from men (N = 5113) and women (N = 7019) aged 16-74 who reported partnered sexual activity in the past year in Britain's third National Survey of Sexual Attitudes and Lifestyles, undertaken in 2010-2012. Latent class analysis used 18 self-reported variables relating to adverse sexual health outcomes (STI and unplanned pregnancy, non-volitional sex, and sexual function problems). Correlates included socio-demographics, early debut, alcohol/drug use, depression, and satisfaction/distress with sex life. RESULTS: Four classes were found for men (labelled Good Sexual Health 83%, Wary Risk-takers 4%, Unwary Risk-takers 4%, Sexual Function Problems 9%); six for women (Good Sexual Health 52%, Wary Risk-takers 2%, Unwary Risk-takers 7%, Low Interest 29%, Sexual Function Problems 7%, Highly Vulnerable 2%). Regardless of gender, Unwary Risk-takers reported lower STI/HIV risk perception and more condomless sex than Wary Risk-takers, but both were more likely to report STI diagnosis than Good Sexual Health classes. Highly Vulnerable women reported abortion, STIs and functional problems, and more sexual coercion than other women. Distinct socio-demographic profiles differentiated higher-risk classes from Good Sexual Health classes, with depression, alcohol/drug use, and early sexual debut widely-shared correlates of higher-risk classes. Females in higher-risk classes, and men with functional problems, evaluated their sex lives more negatively than those with Good Sexual Health. CONCLUSIONS: A greater prevalence and diversity of poor sexual health appears to exist among women than men in Britain, with more consistent effects on women's subjective sexual well-being. Shared health and lifestyle characteristics of higher-risk groups suggest widespread benefits of upstream interventions. Several groups could benefit from tailored interventions: men and women who underestimate their STI/HIV risk exposure, women distressed by low interest in sex, and women experiencing multiple adverse outcomes. Distinctive socio-demographic profiles should assist with identification and targeting