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Neutrophils are key players of the immune system and possess an arsenal of effector functions, including the ability to form and expel neutrophil extracellular traps (NETs) in a process termed NETosis. During NETosis, the nuclear DNA/chromatin expands until it fills the whole cell and is released into the extracellular space. NETs are composed of DNA decorated with histones, proteins, or peptides, and NETosis is implicated in many diseases. Resolving the structure of the nucleus in great detail is essential to understand the underlying processes, but so far, superresolution methods have not been applied. Here, we developed an expansion-microscopy-based method and determined the spatial distribution of chromatin/DNA, histone H1, and nucleophosmin with an over fourfold improved resolution (<40-50 nm) and increased information content. It allowed us to identify the punctate localization of nucleophosmin in the nucleus and histone-rich domains in NETotic cells with a size of 54-66 nm. The technique could also be applied to components of the nuclear envelope (lamins B1 and B2) and myeloperoxidase, providing a complete picture of nuclear composition and structure. In conclusion, expansion microscopy enables superresolved imaging of the highly dynamic structure of nuclei in immune cells.

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The measures taken to cope with the COVID-19 pandemic by governments worldwide have vast consequences on all areas of life. To assess the impact of the COVID-19 pandemic on long-term career development, we evaluated the work-family balance of medical doctors at nine German university clinics. The results indicate a severely disturbed work-family balance, which was mostly due to insufficient childcare, based on restrictions in school operations and childcare. Despite the newly created emergency childcare options, aiming to ensure the functioning of the "systematically important" professional groups, medical doctors feel that they are not sufficiently supported by the measures taken by local governments. Women, in particular, see their professional development at risk. Our results underline that proper and flexible childcare is essential for the career advancement of female medical doctors and is particularly important in times of crises such as the current COVID-19 pandemic. At university medicine clinics, increased work time flexibility and optimized schooling and childcare are needed to promote the career development of female as well as male medical doctors in the early stage of their careers.

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BACKGROUND: Homologous and heterologous SARS-CoV-2 vaccinations yield different spike protein-directed humoral and cellular immune responses. This study aimed to explore their currently unknown interdependencies. METHODS: COV-ADAPT is a prospective, observational cohort study of 417 healthcare workers who received vaccination with homologous ChAdOx1 nCoV-19, homologous BNT162b2 or with heterologous ChAdOx1 nCoV-19/BNT162b2. We assessed humoral (anti-spike-RBD-IgG, neutralizing antibodies, and avidity) and cellular (spike-induced T-cell interferon-γ release) immune responses in blood samples up to 2 weeks before (T1) and 2-12 weeks following secondary immunization (T2). RESULTS: Initial vaccination with ChAdOx1 nCoV-19 resulted in lower anti-spike-RBD-IgG compared with BNT162b2 (70 ± 114 vs. 226 ± 279 BAU/ml, p < .01) at T1. Booster vaccination with BNT162b2 proved superior to ChAdOx1 nCoV-19 at T2 (anti-spike-RBD-IgG: ChAdOx1 nCoV-19/BNT162b2 2387 ± 1627 and homologous BNT162b2 3202 ± 2184 vs. homologous ChAdOx1 nCoV-19 413 ± 461 BAU/ml, both p < .001; spike-induced T-cell interferon-γ release: ChAdOx1 nCoV-19/BNT162b2 5069 ± 6733 and homologous BNT162b2 4880 ± 7570 vs. homologous ChAdOx1 nCoV-19 1152 ± 2243 mIU/ml, both p < .001). No significant differences were detected between BNT162b2-boostered groups at T2. For ChAdOx1 nCoV-19, no booster effect on T-cell activation could be observed. We found associations between anti-spike-RBD-IgG levels (ChAdOx1 nCoV-19/BNT162b2 and homologous BNT162b2) and T-cell responses (homologous ChAdOx1 nCoV-19 and ChAdOx1 nCoV-19/BNT162b2) from T1 to T2. Additionally, anti-spike-RBD-IgG and T-cell response were linked at both time points (all groups combined). All regimes yielded neutralizing antibodies and increased antibody avidity at T2. CONCLUSIONS: Interdependencies between humoral and cellular immune responses differ between common SARS-CoV-2 vaccination regimes. T-cell activation is unlikely to compensate for poor humoral responses.

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Extracorporeal Shock Wave Therapy (ESWT) is used clinically in various disorders including chronic wounds for its pro-angiogenic, proliferative, and anti-inflammatory effects. However, the underlying cellular and molecular mechanisms driving therapeutic effects are not well characterized. Macrophages play a key role in all aspects of healing and their dysfunction results in failure to resolve chronic wounds. We investigated the role of ESWT on macrophage activity in chronic wound punch biopsies from patients with non-healing venous ulcers prior to, and two weeks post-ESWT, and in macrophage cultures treated with clinical shockwave intensities (150-500 impulses, 5 Hz, 0.1 mJ/mm2). Using wound area measurements and histological/immunohistochemical analysis of wound biopsies, we show ESWT enhanced healing of chronic ulcers associated with improved wound angiogenesis (CD31 staining), significantly decreased CD68-positive macrophages per biopsy area and generally increased macrophage activation. Shockwave treatment of macrophages in culture significantly boosted uptake of apoptotic cells, healing-associated cytokine and growth factor gene expressions and modulated macrophage morphology suggestive of macrophage activation, all of which contribute to wound resolution. Macrophage ERK activity was enhanced, suggesting one mechanotransduction pathway driving events. Collectively, these in vitro and in vivo findings reveal shockwaves as important regulators of macrophage functions linked with wound healing. This immunomodulation represents an underappreciated role of clinically applied shockwaves, which could be exploited for other macrophage-mediated disorders.

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