Biology subjects

Behrends, J.

Publications and source records attributed to Behrends, J..

4 recordsLinked to original sources

bioRxiv2025

High-Purity Enrichment of Extracellular Vesicles from Diverse Sources by Conventional and Image-Based Fluorescence Activated Cell Sorters for Robust Downstream Applications

Selective enrichment of extracellular vesicle (EV) subpopulations from the heterogeneous EV pool is essential for understanding their characteristic biological functions and exploiting their potential as diagnostic and prognostic biomarkers. However, isolation of specific EV-subsets remains challenging. Fluorescence-Activated Cell Sorting (FACS) has emerged as promising technique for EV subpopulations enrichment, despite limitations associated to their small size. Although FACS-based EV sorting has been reported, a broadly applicable and systematically validated workflow is still lacking. Here, we describe and validate an optimized workflow for the sorting and analysis of EVs derived from diverse species, tissues, blood and cell culture systems. Using two advanced flow cytometric cell sorters, the BD FACSAria Fusion, and the BD FACSDiscover S8, we systematically evaluated key technical parameters, including nozzle size, sample dilutions, and sorting mode. The optimized workflow enabled efficient enrichment of differently labelled EV populations of interest, achieving near-100% purity, including rare subsets representing less than 10% of the total EV pool, while maintaining compatibility with downstream analyses. Sorted EV populations were characterized by high-sensitivity imaging flow cytometry, transmission electron microscopy, and liquid chromatography-tandem mass spectrometry. This workflow provides a robust framework for EV subset isolation and characterization, supporting both fundamental EV research and translational biomarker applications.

cell biology↗

bioRxiv2024

Ablation of B cell-derived IL-10 increases tuberculosis resistance

Due to the historical dogma, that host defense against intracellular pathogens is mediated by cell-mediated immunity, B cells have been considered unimportant in providing protection against Mycobacterium tuberculosis (Mtb) and remained understudied for decades. However, emerging evidence suggests a more complex and multifaceted role for B cells in tuberculosis (TB) immunity. They accumulate at the side of infection in both animal models and human TB patients, suggesting a potential link to protective immunity. Still, the diverse roles of B cells in TB immunity continue to be unraveled. Apart from antibodies, B cells produce a wide range of cytokines, which can influence the local immune response. Here we addressed the relevance of interleukin 10 (IL-10) secreting B cells in long-term control of the Mtb Beijing strain HN878. Our research highlights the previously unknown role of B cell-derived IL-10 as a negative regulator of protective immunity in TB. For the first time, we demonstrate that mice lacking B cell-derived IL-10 show increased resistance to aerosol Mtb infection, as evidenced by a delayed onset of clinical symptoms and prolonged survival. Notably, this effect was significantly more pronounced in males compared to females, and was accompanied by male-specific immune alterations, indicating a previously unknown sex-specific regulatory role of B cell-derived IL-10 during Mtb infection.

immunology↗

bioRxiv2024

Sex differences in vaccine induced immunity and protection against Mycobacterium tuberculosis

Tuberculosis (TB) is a disease that has evolved with humankind for millennia, causing approximately 1.3 million deaths worldwide per annum. Although increased male affliction for TB and other infections were long known from an epidemiological perspective, our mechanistic understanding of the underlying immunological divergences is relatively recent. As such, there is insufficient knowledge regarding the sexually dimorphic immune response to TB vaccines, where no accepted correlates of protection are yet available. In this context, our goal was to explore how individual sex influences the protective effects of TB vaccines. For this purpose, we vaccinated female and male C57BL/6 mice with Bacille Calmette-Guerin (BCG) and two recombinant derivatives, VPM1002 and BCG{Delta}BCG1419c, to analyse their protective efficacy against challenge with Mycobacterium tuberculosis HN878. We found poor efficacy of BCG in males and the ability of next generation vaccine candidates to improve protection specifically in males. To determine the underlying mechanisms for the differences in survival upon vaccination between females and males, as well as, among different vaccine candidates, we analysed the distribution and persistence of the vaccine strains, in addition to vaccine-induced immune responses at various time points in draining lymph nodes and spleen. We identified sex specific differences in CD8 T cell proliferation in response to mycobacterial antigens ex vivo, 90 days post-vaccination, that associates with vaccine mediated protection against HN878. By integrating our multi-parametric datasets into principal component analysis, followed by extraction of high-variance features, we have uncovered an additional significant association of early CD4 T cell responses with late CD8 T cell responses as well as with survival post HN878 infection. In addition, we have also identified specific clusters of responding CD8 T cells in spleen post-vaccination, that are globally deficient in males as compared to females, irrespective of the BCG strain administered.

immunology↗

bioRxiv2020

WNT6-ACC2-induced accumulation of triacylglycerol rich lipid droplets is exploited by Mycobacterium tuberculosis

In view of emerging drug-resistant tuberculosis, host directed therapies are urgently needed to improve treatment outcomes with currently available anti-tuberculosis therapies. One option is to interfere with the formation of lipid-laden “foamy” macrophages in the infected host. Here, we provide evidence that WNT6, a member of the evolutionary conserved WNT signaling pathway, promotes foam cell formation by regulating key lipid metabolic genes including acetyl-CoA carboxylase-2 (ACC2) during pulmonary TB. In addition, we demonstrate that Mycobacterium tuberculosis (Mtb) facilitates its intracellular growth and dissemination in the host by exploiting the WNT6-ACC2 pathway. Using genetic and pharmacological approaches, we show that lack of functional WNT6 or ACC2 significantly reduces intracellular TAG levels, Mtb growth and necrotic cell death of macrophages. In combination with the anti-TB drug isoniazid, pharmacological inhibition of ACC2 improved anti-mycobacterial treatment in vitro and in vivo. Therefore, we propose the WNT6-ACC2 signaling pathway as a promising target for a host-directed therapy to reduce intracellular replication of Mtb by modulating neutral lipid metabolism.Competing Interest StatementDrs. N. Reiling and J. Brandenburg (Research Center Borstel, Leibniz Lung Center, 23845 Borstel, Germany) have filed a patent application entitled ACC inhibitors as means and methods for treating mycobacterial diseases(WO2018007430A1, patent pending).View Full Text

immunology↗