Biology subjects

Laggner, M.

Publications and source records attributed to Laggner, M..

5 recordsLinked to original sources

bioRxiv2022

Antithymocyte globulin inhibits CD8+ T cell effector functions via the paracrine induction of PDL-1 on monocytes

Antithymocyte globulins (ATG) are T cell depleting antibodies used in solid organ transplantation for induction therapy in sensitized patients with high risk of graft rejection. Previously described effects besides depletion of T cells suggest additional modes of action and identified further cellular targets. Here, we examined the transcriptional changes arising in immune cells from human blood after ex vivo stimulation with ATG on a single cell level to uncover additional mechanisms by which ATG regulates T cell activity and effector functions. Analysis of the paracrine factors present in plasma of ATG-treated whole blood revealed high levels of chemokines and cytokines including Interferon-{gamma} (IFN-{gamma}). Furthermore, we identify an increase of surface expression of programmed cell death 1 ligand 1 (PDL-1) on monocytes mediated by the released paracrine factors. In addition, we show that this induction is dependent on activation of JAK/STAT signaling via binding of IFN-{gamma} to Interferon-{gamma} receptor 1 (IFN-{gamma}R1). Lastly, we demonstrate that the modulation of the immune-regulatory axis of Programmed cell death protein 1 (PD1) on activated CD8+ T cells with PDL-1 found on monocytes mediated by ATG potently inhibits effector functions including proliferation and granzyme B release of activated T cells. Together our findings represent a novel mode of action by which ATG exerts its immunosuppressive effects. One Sentence SummaryATG increases PDL-1 on CD14+-monocytes and inhibits T cell effector functions.

immunology↗

bioRxiv2022

The effect of paracrine factors released by irradiated peripheral blood mononuclear cells on neutrophil extracellular trap formation

Neutrophil extracellular trap (NET)-formation represents an important defence mechanism for rapid clearance of infections. However, exaggerated NET formation has been shown to negatively affect tissue-regeneration after injury. As our previous studies revealed strong tissue-protective and regenerative properties of the secretome of stressed peripheral blood mononuclear cells (PBMCsec), we here investigated the influence of PBMCsec on the formation of NETs. The effect of PBMCsec on NET formation was assessed ex vivo in ionomycin stimulated neutrophils derived from healthy donors using flow cytometry, image stream analysis and quantification of released extracellular DNA. Molecular mechanisms involved in NET formation that were potentially impaired by PBMCsec treatment, including protein kinase C activity, reactive oxygen species production and peptidyl arginine deiminase 4 activity were analysed. Our results showed that PBMCsec significantly inhibited NET formation. Investigation of the different biological substance classes found in PBMCsec revealed only partial reduction of NET formation, suggesting a synergistic effect. Mechanistically, PBMCsec treatment did not interfere with calcium signalling and PKC-activation, but exerted anti-oxidant activity, as evidenced by reduced levels of reactive oxygen species and upregulation of heme oxygenase 1, hypoxia inducible-factor 1 as well as heat shock protein 27 in PBMCsec-treated neutrophils. In addition, PBMCsec strongly inhibited the activation of peptidyl arginine deiminase 4 (PAD4), ultimately leading to the inhibition of NET formation. As therapeutics antagonizing excessive NET formation are currently not available, our study provides a promising novel treatment option for a variety of conditions resulting from exaggerated NET formation.

immunology↗

bioRxiv2022

Paracrine factors of stressed peripheral blood mononuclear cells activate pro-angiogenic and anti- proteolytic processes in whole blood cells and protect the endothelial barrier

Tissue regenerative properties have been attributed to secreted paracrine factors derived from stem cells and other cell types. Especially, the secretome of {gamma}-irradiated peripheral blood mononuclear cells (PBMCsec) has been shown to possess high tissue-regenerative and pro-angiogenic capacities in a variety of preclinical studies. In the light of future therapeutic intravenous applications of PBMCsec, we investigated possible effects of PBMCsec on circulating white blood cells and endothelial cells lining the vasculature. MethodsTo identify changes in the transcriptional profile of white blood cells treated with PBMCSec, whole blood was drawn from healthy individuals and stimulated with PBMCsec for 8 hours ex vivo before further processing for single cell RNA sequencing (scRNAseq). In addition, we performed in vitro assay to confirm findings arising from the transcriptional profiling. ResultsAddition of PBMCsec to whole blood significantly altered the gene signature of granulocytes (17 genes), T-cells (45 genes), B-cells (72 genes) and most prominently monocytes (322 genes). We detected a strong upregulation of several tissue-regenerative and pro-angiogenic cyto- and chemokines in monocytes, including VEGFA, CXCL1 and CXCL5. Intriguingly, inhibitors of endopeptidase activity, such as SERPINB2, were also strongly induced. Measurement of the trans-endothelial electrical resistance of primary human microvascular endothelial cells revealed a strong barrier-protective effect of PBMCsec after barrier disruption. ConclusionTogether, we show that PBMCsec induces angiogenic and proteolytic processes in the blood and is able to attenuate endothelial barrier damage. These regenerative properties suggest that systemic application of PBMCsec might be a promising novel strategy to restore damaged organs.

cell biology↗

bioRxiv2021

Transcriptional differences of lipid-metabolizing enzymes in sebocytes derived from sebaceous glands of the skin and pre-putial glands

Sebaceous glands are adnexal structures, which critically contribute to skin homeostasis and the establishment of a functional epidermal barrier. Sebocytes, the main cell population found within the sebaceous glands, are highly specialized lipid-producing cells. Sebaceous gland-resembling tissue structures are also found in male rodents in form of preputial glands. Similar to sebaceous glands, they are composed of lipid-specialized sebocytes. Due to a lack of adequate organ culture models for skin sebaceous glands and the fact that preputial glands are much larger and easier to handle, previous studies have used preputial glands as a model for skin sebaceous glands. Here, we compared both types of sebocytes, using a single cell RNA sequencing approach, to unravel potential similarities and differences between the two sebocyte populations. In spite of common gene expression patterns due to general lipid-producing properties, we found significant differences in the expression levels of genes encoding enzymes involved in the biogenesis of specialized lipid classes. Specifically, genes critically involved in the mevalonate pathway, including squalene synthase, as well as the sphingolipid salvage pathway, such as ceramide synthase, (acid) sphingomyelinase or acid and alkaline ceramidases, were significantly less expressed by preputial gland sebocytes. Together, our data revealed tissue-specific sebocyte populations, indicating major developmental, functional as well as biosynthetic differences between both glands. The use of preputial glands as surrogate model to study skin sebaceous glands is therefore limited, and major differences between both glands need to be carefully considered before planning an experiment.

molecular biology↗

bioRxiv2020

Single cell landscape of hypertrophic scars identifies serine proteases as key regulators of myofibroblast differentiation

Despite recent advances in understanding skin scarring, mechanisms triggering hypertrophic scar formation are still poorly understood. In the present study we performed single-cell sequencing of mature human hypertrophic scars and developing scars in mice. Compared to normal skin, we found significant differences in gene expression in most cell types present in scar tissue. Fibroblasts (FBs) showed the most prominent alterations in gene expression, displaying a distinct fibrotic signature. By comparing genes upregulated in murine FBs during scar development with genes highly expressed in mature human hypertrophic scars, we identified a group of serine proteases, tentatively involved in scar formation. Two of them, dipeptidyl-peptidase 4 (DPP4) and urokinase (PLAU), were further analyzed in functional assays, revealing a role in TGF{beta}1-mediated myofibroblast differentiation and over-production of components of the extracellular matrix (ECM) without interfering with the canonical TGF{beta}1-signaling pathway. In this study, we delineate the genetic landscape of hypertrophic scars and present new insights into mechanisms involved in hypertrophic scar formation. Our data suggest the use of serine protease inhibitors for the treatment of skin fibrosis.

cell biology↗