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Neuroglia critically shape the brain´s response to ischemic stroke. However, their phenotypic heterogeneity impedes a holistic understanding of the cellular composition of the early ischemic lesion. Here we present a single cell resolution transcriptomics dataset of the brain´s acute response to infarction. Oligodendrocyte lineage cells and astrocytes range among the most transcriptionally perturbed populations and exhibit infarction- and subtype-specific molecular signatures. Specifically, we find infarction restricted proliferating oligodendrocyte precursor cells (OPCs), mature oligodendrocytes and reactive astrocytes, exhibiting transcriptional commonalities in response to ischemic injury. OPCs and reactive astrocytes are involved in a shared immuno-glial cross talk with stroke-specific myeloid cells. Within the perilesional zone, osteopontin positive myeloid cells accumulate in close proximity to CD44 + proliferating OPCs and reactive astrocytes. In vitro, osteopontin increases the migratory capacity of OPCs. Collectively, our study highlights molecular cross talk events which might govern the cellular composition of acutely infarcted brain tissue., (© 2024. The Author(s).)

Idiopathic subglottic stenosis (ISGS) is a rare fibrotic disease of the upper trachea with an unknown pathomechanism. It typically affects adult Caucasian female patients, leading to severe airway constrictions caused by progressive scar formation and inflammation with clinical symptoms of dyspnoea, stridor and potential changes to the voice. Endoscopic treatment frequently leads to recurrence, whereas surgical resection and reconstruction provides excellent long-term functional outcome. This study aimed to identify so far unrecognized pathologic aspects of ISGS using single cell RNA sequencing. Our scRNAseq analysis uncovered the cellular composition of the subglottic scar tissue, including the presence of a pathologic, profibrotic fibroblast subtype and the presence of Schwann cells in a profibrotic state. In addition, a pathology-associated increase of plasma cells was identified. Using extended bioinformatics analyses, we decoded pathology-associated changes of factors of the extracellular matrix. Our data identified ongoing fibrotic processes in ISGS and provide novel insights on the contribution of fibroblasts, Schwann cells and plasma cells to the pathogenesis of ISGS. This knowledge could impact the development of novel approaches for diagnosis and therapy of ISGS., Competing Interests: Authors MD, ML, DC, KK, DB, and HA were employed by Aposcience AG. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Direder, Laggner, Copic, Klas, Bormann, Schweiger, Hoetzenecker, Aigner, Ankersmit and Mildner.)

Case: A 33-year-old man with previously diagnosed lupus membranous nephropathy presented with painful swelling in both legs. Laboratory tests revealed acute kidney injury, and imaging studies by duplex ultrasound and computed tomography scan showed acute thrombosis of both renal veins, the infrahepatic inferior vena cava, and both iliofemoral venous segments. Initially, pharmacomechanical thrombolysis led to an insufficient morphological result. The therapeutic breakthrough was achieved by catheter-based mechanical thrombectomy of the infrarenal vena cava and both renal veins, which successfully cleared all affected venous segments from thrombus, paralleled by improvement of the patient's condition. However, after 1 week, the patient experienced recurrent thrombosis of the right renal vein with hemorrhagic infarction of the right kidney. After further optimization of immunomodulatory and antithrombotic therapy, a repeated catheter-based mechanical thrombectomy resulted in sustained clinical improvement and preservation of renal venous drainage and kidney function., Conclusion: Extensive acute thrombosis of both renal veins, the inferior vena cava, and both iliofemoral venous segments is a rare emergency potentially threatening kidney function. Immediate effective thrombus removal is essential to preserve kidney function and can be achieved by catheter-based mechanical thrombectomy embedded in a comprehensive immunomodulatory and antithrombotic therapeutic concept., Clinical Impact: This case demonstrated the efficacy of a catheter-based therapeutic approach in patients with extensive thrombosis of the venous system. A catheter-based approach must be embedded in a comprehensive medical therapeutic concept, which is essential to achieve a sustainable result., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Objectives: This population-based cohort study investigated mid-term outcome after surgical aortic valve replacement with a bioprosthetic or mechanical valve prosthesis in patients aged <50 years in a European social welfare state., Methods: We analysed patient data from the main social insurance carriers in Austria (2010-2020). Subsequent patient-level record linkage with national health data provided patient characteristics and clinical outcome. Survival, reoperation, myocardial infarction, heart failure, embolic stroke or intracerebral haemorrhage, bleeding other than intracerebral haemorrhage and major adverse cardiac events were evaluated as outcomes., Results: A total of 991 patients were analysed. Regarding demographics, no major differences between groups were observed. Multivariable Cox regression revealed no significant difference in overall survival (P = 0.352) with a median follow-up time of 6.2 years. Reoperation-free survival was decreased (hazard ratio = 1.560 [95% CI: 1.076-2.262], P = 0.019) and the risk for reoperation was increased (hazard ratio = 2.770 [95% CI: 1.402-5.472], P = 0.003) in patients who received bioprostheses. Estimated probability of death after reoperation was 0.23 (CL: 0.08-0.35) after 2 years and 0.34 (CL: 0.06-0.53) after 10 years over both groups. Regarding further outcomes, no significant differences between the two groups were observed., Conclusions: In patients below 50 years of age receiving aortic valve replacement, implantation of bioprostheses when compared to mechanical heart valve prostheses was associated with a significantly higher rate of reoperations and reduced reoperation-free survival. Nevertheless, we could not observe a difference in overall survival. However, long-term follow-up has to evaluate that a significantly lower rate of reoperations may translate in consistently improved long-term survival., (© The Author(s) 2023. Published by Oxford University Press on behalf of the European Association for Cardio-Thoracic Surgery.)

Peripheral contact to pathogen-associated molecular patterns (PAMPs) evokes a systemic innate immune response which is rapidly relayed to the central nervous system (CNS). The remarkable cellular heterogeneity of the CNS poses a significant challenge to the study of cell type and stimulus dependent responses of neural cells during acute inflammation. Here we utilized single nuclei RNA sequencing (snRNAseq), serum proteome profiling and primary cell culture methods to systematically compare the acute response of the mammalian brain to the bacterial PAMP lipopolysaccharide (LPS) and the viral PAMP polyinosinic:polycytidylic acid (Poly(I:C)), at single cell resolution. Our study unveiled convergent transcriptional cytokine and cellular stress responses in brain vascular and ependymal cells and a downregulation of several key mediators of directed blood brain barrier (BBB) transport. In contrast the neuronal response to PAMPs was limited in acute neuroinflammation. Moreover, our study highlighted the dominant role of IFN signalling upon Poly(I:C) challenge, particularly in cells of the oligodendrocyte lineage. Collectively our study unveils heterogeneous, shared and distinct cell type and stimulus dependent acute responses of the CNS to bacterial and viral PAMP challenges. Our findings highlight inflammation induced dysregulations of BBB-transporter gene expression, suggesting potential translational implications on drug pharmacokinetics variability during acute neuroinflammation. The pronounced dependency of oligodendrocytes on IFN stimulation during viral PAMP challenges, emphasizes their limited molecular viral response repertoire., Competing Interests: Declaration of Competing Interest The authors declare that the research has been performed without any conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Lithium (Li) metal is a promising anode material for lithium-ion batteries (LIBs) because of its high theoretical specific capacity of 3860 mAh g -1 and the low potential of -3.04 V versus the standard hydrogen electrode (SHE). However, these anodes rely on repeated plating and stripping of Li, which leads to consumption of Li inventory and the growth of dendrites that can lead to self-discharge and safety issues. To address these issues, as well as problems related to the volume change of these anodes, a number of different porous conductive scaffolds have been reported to create high surface area electrode on which Li can be plated reliably. While impressive results have been reported in literature, current processes typically rely on either expensive or poorly scalable techniques. Herein, we report a scalable fabrication method to create robust 3D Cu anodes using a one-step electrodeposition process. The areal loading, pore structure, and electrode thickness can be tuned by changing the electrodeposition parameters, and we show how standard mechanical calendering provides a way to further optimize electrode volume, capacity, and cycling stability. Optimized electrodes achieve high Coulombic efficiencies (CEs) of 99% during 800 cycles in half cells at a current density of 0.5 mA cm -2 with a total capacity of 0.5 mAh cm -2 . To the best of our knowledge, this is the highest value ever reported for a host for Li-metal anodes using lithium bis(trifluoromethanesulfonyl)imide LITFSI based electrolyte.

Trauma represents one of the leading causes of death worldwide. Traumatic injuries elicit a dynamic inflammatory response with systemic release of inflammatory cytokines. Disbalance of this response can lead to systemic inflammatory response syndrome or compensatory anti-inflammatory response syndrome. As neutrophils play a major role in innate immune defence and are crucial in the injury-induced immunological response, we aimed to investigate systemic neutrophil-derived immunomodulators in trauma patients. Therefore, serum levels of neutrophil elastase (NE), myeloperoxidase (MPO) and citrullinated histone H3 (CitH3) were quantified in patients with injury severity scores above 15. Additionally, leukocyte, platelet, fibrinogen and CRP levels were assessed. Lastly, we analysed the association of neutrophil-derived factors with clinical severity scoring systems. Although the release of MPO, NE and CitH3 was not predictive of mortality, we found a remarkable increase in MPO and NE in trauma patients as compared with healthy controls. We also found significantly increased levels of MPO and NE on Days 1 and 5 after initial trauma in critically injured patients. Taken together, our data suggest a role for neutrophil activation in trauma. Targeting exacerbated neutrophil activation might represent a new therapeutic option for critically injured patients., (© 2023 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Hypertrophic scars can cause pain, movement restrictions, and reduction in the quality of life. Despite numerous options to treat hypertrophic scarring, efficient therapies are still scarce, and cellular mechanisms are not well understood. Factors secreted by peripheral blood mononuclear cells (PBMCsec) have been previously described for their beneficial effects on tissue regeneration. In this study, we investigated the effects of PBMCsec on skin scarring in mouse models and human scar explant cultures at single-cell resolution (scRNAseq). Mouse wounds and scars, and human mature scars were treated with PBMCsec intradermally and topically. The topical and intradermal application of PBMCsec regulated the expression of various genes involved in pro-fibrotic processes and tissue remodeling. We identified elastin as a common linchpin of anti-fibrotic action in both mouse and human scars. In vitro, we found that PBMCsec prevents TGFβ-mediated myofibroblast differentiation and attenuates abundant elastin expression with non-canonical signaling inhibition. Furthermore, the TGFβ-induced breakdown of elastic fibers was strongly inhibited by the addition of PBMCsec. In conclusion, we conducted an extensive study with multiple experimental approaches and ample scRNAseq data demonstrating the anti-fibrotic effect of PBMCsec on cutaneous scars in mouse and human experimental settings. These findings point at PBMCsec as a novel therapeutic option to treat skin scarring.

The epidermal barrier of mammals is initially formed during embryonic development and continuously regenerated by the differentiation and cornification of keratinocytes in postnatal life. Cornification is associated with the breakdown of organelles and other cell components by mechanisms which are only incompletely understood. Here, we investigated whether heme oxygenase 1 (HO-1), which converts heme into biliverdin, ferrous iron and carbon monoxide, is required for normal cornification of epidermal keratinocytes. We show that HO-1 is transcriptionally upregulated during the terminal differentiation of human keratinocytes in vitro and in vivo. Immunohistochemistry demonstrated expression of HO-1 in the granular layer of the epidermis where keratinocytes undergo cornification. Next, we deleted the Hmox1 gene, which encodes HO-1, by crossing Hmox1 -floxed and K14-Cre mice. The epidermis and isolated keratinocytes of the resulting Hmox1 f/f K14-Cre mice lacked HO-1 expression. The genetic inactivation of HO-1 did not impair the expression of keratinocyte differentiation markers, loricrin and filaggrin. Likewise, the transglutaminase activity and formation of the stratum corneum were not altered in Hmox1 f/f K14-Cre mice, suggesting that HO-1 is dispensable for epidermal cornification. The genetically modified mice generated in this study may be useful for future investigations of the potential roles of epidermal HO-1 in iron metabolism and responses to oxidative stress.

Background: Antithymocyte globulins (ATG) are T cell-depleting antibodies used in solid organ transplantation for induction therapy in sensitized patients with a high risk of graft rejection. Previously described effects besides the depletion of T cells have suggested additional modes of action and identified further cellular targets., Methods: We examined the transcriptional changes arising in immune cells from human blood after ex vivo stimulation with ATG at the single-cell level to uncover additional mechanisms by which ATG regulates T cell activity and effector functions., Findings: Analysis of the paracrine factors present in the plasma of ATG-treated whole blood revealed high levels of chemokines and cytokines, including interferon-γ (IFN-γ). Furthermore, we identified an increase in the surface expression of the programmed death ligand 1 (PDL-1) on monocytes mediated by the released paracrine factors. In addition, we showed that this induction is dependent on the activation of JAK/STAT signaling via the binding of IFN-γ to interferon-γ receptor 1 (IFN-γR1). Lastly, we demonstrated 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 the proliferation and granzyme B release of activated T cells., Interpretation: Together, our findings represent a novel mode of action by which ATG exerts its immunosuppressive effects.

Recently, a specific Schwann cell type with profibrotic and tissue regenerative properties that contributes to keloid formation has been identified. In the present study, we reanalyzed published single-cell RNA sequencing (scRNA-seq) studies of keloids, healthy skin, and normal scars to reliably determine the specific gene expression profile of keloid-specific Schwann cell types in more detail. We were able to confirm the presence of the repair-like, profibrotic Schwann cell type in the datasets of all three studies and identified a specific gene-set for these Schwann cells. In contrast to keloids, in normal scars, the number of Schwann cells was not increased, nor was their gene expression profile distinctly different from that of Schwann cells of normal skin. In addition, our bioinformatics analysis provided evidence for a role of transcription factors of the AP1, STAT, and KLF families, and members of the IER genes in the dedifferentiation process of keloidal Schwann cells. Together, our analysis strengthens the role of the profibrotic Schwann cell type in the formation of keloids. Knowledge of the exact gene expression profile of these Schwann cells will facilitate their identification in other organs and diseases., (© 2022. The Author(s).)

Neutrophil extracellular trap (NET)-formation represents an important defence mechanism for the rapid clearance of infections. However, exaggerated NET formation has been shown to negatively affect tissue-regeneration after injury. As our previous studies revealed the 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. The effect of PBMCsec on molecular mechanisms involved in NET formation, including Ca-flux, protein kinase C activity, reactive oxygen species production, and protein 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 a partial reduction in 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 and hypoxia inducible-factor 1 in PBMCsec-treated neutrophils. In addition, PBMCsec strongly inhibited the activation of protein arginine deiminase 4 (PAD4), ultimately leading to the inhibition of NET formation. As therapeutics antagonizing excessive NET formation are not currently available, our study provides a promising novel treatment option for a variety of conditions resulting from exaggerated NET formation.

Tissue-regenerative properties have been attributed to secreted paracrine factors derived from stem cells and other cell types. In particular, the secretome of γ-irradiated peripheral blood mononuclear cells (PBMCsec) has been shown to possess high tissue-regenerative and proangiogenic capacities in a variety of preclinical studies. In light of future therapeutic intravenous applications of PBMCsec, we investigated the possible effects of PBMCsec on white blood cells and endothelial cells lining the vasculature. To identify changes in the transcriptional profile, whole blood was drawn from healthy individuals and stimulated with PBMCsec for 8 h ex vivo before further processing for single-cell RNA sequencing. PBMCsec 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 proangiogenic 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. Together, 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.

Background: IgE-mediated hypersensitivity is becoming increasingly prevalent and activation of mast cells and basophils represent key events in the pathophysiology of allergy. We have previously reported that the secretome of γ-irradiated peripheral blood mononuclear cells (PBMCsec) exerts beneficial anti-inflammatory effects. Yet, its ability to alleviate allergic symptoms has not been investigated so far., Methods: Several experimental in vitro and in vivo models have been used in this basic research study. A murine ear swelling model was used to study the effects of PBMCsec on 48/80-induced mast cell degranulation in vivo. The transcriptional profile of murine mast cells was analysed by single cell RNA sequencing (scRNAseq). Mast cell activation was studied in vitro using primary skin mast cells. Basophils from individuals allergic to birch pollens were used to investigate basophile activation by allergens. Transcriptomic and lipidomic analyses were used to identify mRNA expression and lipid species present in PBMCsec, respectively., Findings: Topical application of PBMCsec on mouse ears (C57BL/6) significantly reduced tissue swelling following intradermal injection of compound 48/80, an inducer of mast cell degranulation. Single cell RNA sequencing of PBMCsec-treated murine dermal mast cells (Balb/c) revealed a downregulation of genes involved in immune cell degranulation and Fc-receptor signalling. In addition, treatment of primary human dermal mast cells with PBMCsec strongly inhibited compound 48/80- and α-IgE-induced mediator release in vitro. Furthermore, PBMCsec remarkably attenuated allergen driven activation of basophils from allergic individuals. Transcriptomic analysis of these basophils showed that PBMCsec downregulated a distinct gene battery involved in immune cell degranulation and Fc-receptor signalling, corroborating results obtained from dermal mast cells. Finally, we identified the lipid fraction of PBMCsec as the major active ingredient involved in effector cell inhibition., Interpretation: Collectively, our data demonstrate that PBMCsec is able to reduce activation of mast cells and basophils, encouraging further studies on the potential use of PBMCsec for treating allergy., Funding: Austrian Research Promotion Agency (852748 and 862068, 2015-2019), Vienna Business Agency (2343727, 2018-2020), Aposcience AG, Austrian Federal Ministry of Education, Science and Research (SPA06/055), Danube Allergy Research Cluster, Austrian Science Fund (I4437 and P32953)., Competing Interests: Declaration of interests The Medical University of Vienna has claimed financial interest. HJA holds patents related to this work (WO2010079086A1; WO2010070105A1; EP3502692A1; WO2021130305A1). MM hold a patent related to this work (WO2021130305A1). ML, DC, VV, AG, MD, KK, DB, AP, and HJA are affiliated with the company Aposcience AG, a manufacturer of PBMCsec. All other authors declare no potential conflicts of interest., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Abbreviations: ATG7: autophagy related 7; BODIPY: boron dipyrromethene; DAG: diacyl glycerides; DBI: diazepam binding inhibitor; GFP: green fluorescent protein; KRT14: keratin 14; HPLC-MS: high performance liquid chromatography-mass spectrometry; LD: lipid droplet; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MSI: mass spectrometric imaging; ORO: Oil Red O; PC: phosphatidylcholine; PE: phosphatidylethanolamine; PG: preputial gland; PLIN2: perilipin 2; PtdIns: phosphatidylinositol; PL: phospholipids; POPC: 1-palmitoyl-2-oleoyl-PC; PS: phosphatidylserine; qRT-PCR: quantitative reverse transcribed PCR; SG: sebaceous gland; scRNAseq: single-cell RNA sequencing; TAG: triacylglycerides; TLC: thin layer chromatography.

Keloids are disfiguring, hypertrophic scars with yet poorly understood pathomechanisms, which could lead to severe functional impairments. Here we analyzed the characteristics of keloidal cells by single cell sequencing and discovered the presence of an abundant population of Schwann cells that persisted in the hypertrophic scar tissue after wound healing. In contrast to normal skin, keloidal Schwann cells show a unique, pro-fibrotic phenotype. Our data support the hypothesis that keloidal Schwann cells contribute to the formation of the extracellular matrix and are able to affect M2 polarization of macrophages. Indeed, we show that macrophages in keloids predominantly display a M2 polarization and produce factors that inhibit Schwann cell differentiation. This study suggests the contribution of a Schwann cell - macrophage cross-talk to the continuous expansion of keloids, and that targeting Schwann cells might represent an interesting novel treatment option for keloids., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Burn injuries elicit a unique and dynamic stress response which can lead to burn injury progression. Though neutrophils represent crucial players in the burn-induced immunological events, the dynamic secretion pattern and systemic levels of neutrophil-derived factors have not been investigated in detail so far. Serum levels of neutrophil elastase (NE), myeloperoxidase (MPO), citrullinated histone H3 (CitH3), and complement factor C3a were quantified in burn victims over 4 weeks post injury. Furthermore, the potential association with mortality, degree of burn injury, and inhalation trauma was evaluated. In addition, leukocyte, platelet, neutrophil, and lymphocyte counts were assessed. Lastly, we analyzed the association of neutrophil-derived factors with clinical severity scoring systems. Serum levels of NE, MPO, CitH3, and C3a were remarkably elevated in burn victims compared to healthy controls. Leukocyte and neutrophil counts were significantly increased on admission day and day 1, while relative lymphocytes were decreased in the first 7 days post burn trauma. Though neutrophil-derived factors did not predict mortality, patients suffering from 3rd degree burn injuries displayed increased CitH3 and NE levels. Accordingly, CitH3 and NE were elevated in cases with higher abbreviated burn severity indices (ABSI). Taken together, our data suggest a role for neutrophil activation and NETosis in burn injuries and burn injury progression. Targeting exacerbated neutrophil activation might represent a new therapeutic option for severe cases of burn injury., (© 2022. The Author(s).)

Acute myocardial infarction (AMI) is a result of cardiac non-perfusion and leads to cardiomyocyte necrosis, inflammation, and compromised cardiac performance. Here, we showed that the secretome of γ-irradiated peripheral blood mononuclear cells (PBMCsec) improved heart function in a porcine AMI model and displayed beneficial long- and short-term effects. As an AMI is known to strongly affect gene regulation of the ischemia non-affected heart muscle and distal organs, we employed a transcriptomics approach to further study the immediate molecular events orchestrated using the PBMCsec in myocardium, liver, and spleen 24 h post ischemia. In the infarcted area, the PBMCsec mainly induced genes that were essential for cardiomyocyte function and simultaneously downregulated pro-inflammatory genes. Interestingly, genes associated with pro-inflammatory processes were activated in the transition zone, while being downregulated in the remote zone. In the liver, we observed a pronounced inhibition of immune responses using the PBMCsec, while genes involved in urea and tricarboxylic cycles were induced. The spleen displayed elevated lipid metabolism and reduced immunological processes. Together, our study suggested several types of pharmacodynamics by which the PBMCsec conferred immediate cardioprotection. Furthermore, our data supported the assumption that an AMI significantly affects distal organs, suggesting that a holistic treatment of an AMI, as achieved by PBMCsec, might be highly beneficial.

Despite recent advances in understanding skin scarring, mechanisms triggering hypertrophic scar formation are still poorly understood. In the present study, we investigate mature human hypertrophic scars and developing scars in mice at single cell resolution. Compared to normal skin, we find significant differences in gene expression in most cell types present in scar tissue. Fibroblasts show the most prominent alterations in gene expression, displaying a distinct fibrotic signature. By comparing genes upregulated in murine fibroblasts during scar development with genes highly expressed in mature human hypertrophic scars, we identify a group of serine proteases, tentatively involved in scar formation. Two of them, dipeptidyl-peptidase 4 (DPP4) and urokinase (PLAU), are further analyzed in functional assays, revealing a role in TGFβ1-mediated myofibroblast differentiation and over-production of components of the extracellular matrix in vitro. Topical treatment with inhibitors of DPP4 and PLAU during scar formation in vivo shows anti-fibrotic activity and improvement of scar quality, most prominently after application of the PLAU inhibitor BC-11. In this study, we delineate the genetic landscape of hypertrophic scars and present insights into mechanisms involved in hypertrophic scar formation. Our data suggest the use of serine protease inhibitors for the treatment of skin fibrosis., (© 2021. The Author(s).)

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 the 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 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 a 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.

Technological advances in membrane technology, catalysis, and electrochemical energy storage require the fabrication of controlled pore structures at ever smaller length scales. It is therefore important to develop processes allowing for the fabrication of materials with controlled submicron porous structures. We propose a combination of colloidal lithography and chemical vapor deposition of carbon nanotubes to create continuous straight pores with diameters down to 100 nm in structures with thicknesses of more than 300 μm. These structures offer unique features, including continuous and parallel pores with aspect ratios in excess of 3000, a low pore tortuosity, good electrical conductivity, and electrochemical stability. We demonstrate that these structures can be used in Li-ion batteries by coating the carbon nanotubes with Si as an active anode material., Competing Interests: The authors declare no competing financial interest., (© 2021 The Authors. Published by American Chemical Society.)

Ichthyoses comprise a broad spectrum of keratinization disorders due to hereditary defects of cornification. Until now, mutations in more than 50 genes, mostly coding for structural proteins involved in epidermal barrier formation, have been identified as causes for different types of these keratinization disorders. However, due to the high heterogeneity and difficulties in the establishment of valid experimental models, research in this field remains challenging and translation of novel findings to clinical practice is difficult. In this review, we provide an overview of existing models to study hereditary cornification defects with focus on ichthyoses and palmoplantar keratodermas.

Cell-free secretomes represent a promising new therapeutic avenue in regenerative medicine, and γ-irradiation of human peripheral blood mononuclear cells (PBMCs) has been shown to promote the release of paracrine factors with high regenerative potential. Recently, the use of alternative irradiation sources, such as artificially generated β- or electron-irradiation, is encouraged by authorities. Since the effect of the less hazardous electron-radiation on the production and functions of paracrine factors has not been tested so far, we compared the effects of γ- and electron-irradiation on PBMCs and determined the efficacy of both radiation sources for producing regenerative secretomes. Exposure to 60 Gy γ-rays from a radioactive nuclide and 60 Gy electron-irradiation provided by a linear accelerator comparably induced cell death and DNA damage. The transcriptional landscapes of PBMCs exposed to either radiation source shared a high degree of similarity. Secretion patterns of proteins, lipids, and extracellular vesicles displayed similar profiles after γ- and electron-irradiation. Lastly, we detected comparable biological activities in functional assays reflecting the regenerative potential of the secretomes. Taken together, we were able to demonstrate that electron-irradiation is an effective, alternative radiation source for producing therapeutic, cell-free secretomes. Our study paves the way for future clinical trials employing secretomes generated with electron-irradiation in tissue-regenerative medicine., Competing Interests: The Medical University of Vienna has claimed financial interest. H.J.A. holds patents related to this work (WO 2010/079086 A1, WO 2010/070105 A1, EP 3502692, European Patent Office application #19165340.1). All other authors declare no competing interests., (© 2021 The Author(s).)

Background: Chronic obstructive pulmonary disease (COPD) is a major cause of morbidity and mortality and is characterized by episodes of acute exacerbations. Finding a systemic biomarker that reliably predicts outcome after an acute exacerbation remains a major challenge. Heat shock protein 27 (HSP27) has been previously studied in COPD, however, urine excretion trajectory and prognostic value after an exacerbation is unknown., Methods: In this retrospective post hoc analysis of a prospective study that included 253 COPD patients who were hospitalized for acute exacerbation, 207 patients were analyzed. Urine and serum were sampled at admission, discharge, and 180 days after discharge; urine excretion trajectory was analyzed and correlated with clinicopathological and survival data., Results: HSP27 urine excretion increased after an exacerbation episode [1.8% admission, 1.8% discharge, 2.3% 180 days after discharge (P=0.091)]. In severely ill patients (GOLD IV) this course was even more distinct [1.6% admission, 2.1% discharge, 2.8% 180 days after discharge (P=0.007)]. Furthermore, fractional HSP27 urine excretion at discharge was increased in GOLD IV patients (P=0.031). In Kaplan-Meier and univariable Cox proportional hazard models patients with HSP27 urine excretion below 0.845% showed significantly worse survival at 30, 90 and 180 days after discharge. In a multivariable Cox proportional hazard model including established COPD outcome parameters fractional HSP27 urine excretion remained a significant predictor of survival at 30 and 90 days after discharge. Comparing this model to our already published model that includes HSP27 serum concentration we could show that fractional HSP27 urine excretion performs better in short-term survival., Conclusions: Our findings provide novel information about fractional HSP27 urine excretion trajectory in acute exacerbation of COPD. Fractional HSP27 urine excretion may be significantly reduced during an episode of acute exacerbation in COPD patients and may be used as a predictor of short-term all-cause mortality., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/atm-20-3683). MF reports personal fees from Astra Zeneca, personal fees from Boehringer Ingelheim, outside the submitted work. The other authors have no conflicts of interest to declare., (2021 Annals of Translational Medicine. All rights reserved.)

The role of microRNAs (miRNAs) during keratinocyte (KC) differentiation and in skin diseases with epidermal phenotypes has attracted strong interest over the past few years. However, combined mRNA and miRNA expression analyses to elucidate the intricate mRNA-miRNA networks of KCs at different stages of differentiation have not been performed yet. In the present study, we investigated the dynamics of miRNA and mRNA expression during KC differentiation in vitro and in normal and psoriatic epidermis. While we identified comparable numbers of up- and downregulated mRNAs (49% and 51%, respectively), miRNAs were predominantly upregulated (76% vs 24%) during KC differentiation. Further bioinformatics analyses suggested an important inhibitory role for miR-155 in KC differentiation, as it was repressed during KC differentiation in normal skin but strongly upregulated in the epidermis of psoriatic skin lesions. Mimicking the inflammatory milieu of psoriatic skin in vitro, we could show that the pro-inflammatory cytokines IL17, IL1β and INFγ synergistically upregulated miR-155 expression in KCs. Forced over-expression of miR-155 in human in vitro skin models specifically reduced the expression of loricrin (LOR) in KCs, indicating that miR-155 interferes with the establishment of a normal epidermal barrier. Together, our data indicate that downregulation of miR-155 during KC differentiation is a crucial step for epidermal barrier formation. Furthermore, its strong upregulation in psoriatic lesions suggests a contributing role of miR-155 in the altered keratinocyte differentiation observed in psoriasis. Therefore, miR-155 represents as a potential target for treating psoriatic skin lesions.

Background: Since numerous pathological conditions are evoked by unwanted dendritic cell (DC) activity, therapeutic agents modulating DC functions are of great medical interest. In regenerative medicine, cellular secretomes have gained increasing attention and valuable immunomodulatory properties have been attributed to the secretome of γ-irradiated peripheral blood mononuclear cells (PBMCs). Potential effects of the PBMC secretome (PBMCsec) on key DC functions have not been elucidated so far., Methods: We used a hapten-mediated murine model of contact hypersensitivity (CH) to study the effects of PBMCsec on DCs in vivo. Effects of PBMCsec on human DCs were investigated in monocyte-derived DCs (MoDC) and ex vivo skin cultures. DCs were phenotypically characterised by transcriptomics analyses and flow cytometry. DC function was evaluated by cytokine secretion, antigen uptake, PBMC proliferation and T-cell priming., Findings: PBMCsec significantly alleviated tissue inflammation and cellular infiltration in hapten-sensitized mice. We found that PBMCsec abrogated differentiation of MoDCs, indicated by lower expression of classical DC markers CD1a, CD11c and MHC class II molecules. Furthermore, PBMCsec reduced DC maturation, antigen uptake, lipopolysaccharides-induced cytokine secretion, and DC-mediated immune cell proliferation. Moreover, MoDCs differentiated with PBMCsec displayed diminished ability to prime naïve CD4 + T-cells into T H 1 and T H 2 cells. Furthermore, PBMCsec modulated the phenotype of DCs present in the skin in situ. Mechanistically, we identified lipids as the main biomolecule accountable for the observed immunomodulatory effects., Interpretation: Together, our data describe DC-modulatory actions of lipids secreted by stressed PBMCs and suggest PBMCsec as a therapeutic option for treatment of DC-mediated inflammatory skin conditions., Funding: This research project was supported by the Austrian Research Promotion Agency (Vienna, Austria; grant "APOSEC" 862068; 2015-2019) and the Vienna Business Agency (Vienna, Austria; grant "APOSEC to clinic" 2343727)., Competing Interests: Declaration of Competing Interest The Medical University of Vienna has claimed financial interest and HJA holds patents related to this work (WO 2010/079,086 A1, WO 2010/070,105, PCT/EP2018/085,955, EPO 19,165,340.1, EPO application 19,219,342.3). All other authors declare no potential conflicts of interest., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Advances in the synthesis and assembly of nanomaterials offer a unique opportunity to purposefully design structures according to the requirements of the targeted applications. This paper shows a process to create robust 3D carbon nanotube (CNT) structures, which provide an electrically conductive support for nanoparticle coating. We describe a process to reliably fabricate robust honeycomb structures with walls made out of aligned CNTs. We present a design of experimental analysis of this fabrication process and discuss methods to coat these honeycombs with BiVO 4 for solar fuel applications. The proposed honeycomb structure allows for an efficient transport of electrons through the electrode, as well as an enhanced light-electrode interaction. Finally, we demonstrate that the developed CNT electrodes can survive harsh BiVO 4 synthesis conditions and can subsequently be used as photoelectrodes for solar water splitting.

Controlling the arrangement and interface of nanoparticles is essential to achieve good transfer of charge, heat, or mechanical load. This is particularly challenging in systems requiring hybrid nanoparticle mixtures such as combinations of organic and inorganic materials. This work presents a process to coat vertically aligned carbon nanotube (CNT) forests with metal oxide nanoparticles using microwave-assisted hydrothermal synthesis. Hydrothermal processes normally damage delicate CNT forests, which is addressed here by a combination of lithographic patterning, transfer printing, and reduction of the synthesis time. This process is applied for the fabrication of structured Li-ion battery (LIB) electrodes where the aligned CNTs provide a straight electron transport path through the electrode and the hydrothermal coating process is used to coat the CNTs with conversion anode materials for LIBs. These nanoparticles are anchored on the surface of the CNTs and batteries fabricated following this process show a fourfold longer cyclability. Finally, this process is used to create thick electrodes (350 µm) with a gravimetric capacity of over 900 mAh g -1 ., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Peripheral blood mononuclear cells (PBMCs) have been shown to produce and release a plethora of pro-angiogenetic factors in response to γ-irradiation, partially accounting for their tissue-regenerative capacity. Here, we investigated whether a certain cell subtype of PBMCs is responsible for this effect, and whether the type of cell death affects the pro-angiogenic potential of bioactive molecules released by γ-irradiated PBMCs. PBMCs and PBMC subpopulations, including CD4 + and CD8 + T cells, B cells, monocytes, and natural killer cells, were isolated and subjected to high-dose γ-irradiation. Transcriptome analysis revealed subpopulation-specific responses to γ-irradiation with distinct activation of pro-angiogenic pathways, cytokine production, and death receptor signalling. Analysis of the proteins released showed that interactions of the subsets are important for the generation of a pro-angiogenic secretome. This result was confirmed at the functional level by the finding that the secretome of γ-irradiated PBMCs displayed higher pro-angiogenic activity in an aortic ring assay. Scanning electron microscopy and image stream analysis of γ-irradiated PBMCs revealed distinct morphological changes, indicative for apoptotic and necroptotic cell death. While inhibition of apoptosis had no effect on the pro-angiogenic activity of the secretome, inhibiting necroptosis in stressed PBMCs abolished blood vessel sprouting. Mechanistically, we identified tumor necrosis factor (TNF) receptor superfamily member 1B as the main driver of necroptosis in response to γ-irradiation in PBMCs, which was most likely mediated via membrane-bound TNF-α. In conclusion, our study demonstrates that the pro-angiogenic activity of the secretome of γ-irradiated PBMCs requires interplay of different PBMC subpopulations. Furthermore, we show that TNF-dependent necroptosis is an indispensable molecular process for conferring tissue-regenerative activity and for the pro-angiogenic potential of the PBMC secretome. These findings contribute to a better understanding of secretome-based therapies in regenerative medicine.

Circular dichroism spectroscopy has become a powerful tool to characterise proteins and other biomolecules. For heterogeneous samples such as those present for interacting proteins, typically only average spectroscopic features can be resolved. Here we overcome this limitation by using free-flow microfluidic size separation in-line with synchrotron radiation circular dichroism to resolve the secondary structure of each component of a model protein mixture containing monomers and fibrils. To enable this objective, we have integrated far-UV compatible measurement chambers into PDMS-based microfluidic devices. Two architectures are proposed so as to accommodate for a wide range of concentrations. The approach, which can be used in combination with other bulk measurement techniques, paves the way to the study of complex mixtures such as the ones associated with protein misfolding and aggregation diseases including Alzheimer's and Parkinson's diseases.

Carbon nanotube (CNT)-based filters have the potential to revolutionize water treatment because of their high capacity and fast kinetics in sorption of organic, inorganic, and biological pollutants. To date, CNT filters either rely on CNTs dispersed in liquids, which are difficult to recover and cause safety concerns, or on CNT buckypaper, which offers high efficiency, but suffers from an intrinsic trade-off between filter permeability and capacity. Here, a new approach is presented that bypasses this trade-off and achieves buckypaper-like efficiency combined with filter-column-like permeability and capacity. For this, CNTs are first assembled into porous microspheres and then are packed into microfluidic column filters. These microcolumns exhibit large flow-through filtration efficiencies, while maintaining membrane permeabilities an order of magnitude larger then CNT buckypaper and specific permeabilities double that of activated carbon for similar flowrates (232 000 L m -2 h -1 bar -1 , 1.23 × 10 -12 m 2 ). Moreover, in a test to remove sodium dodecyl sulfate (SDS) from water, these microstructured CNT columns outperform activated carbon columns. This improved filtration efficiency and permeability is an important step toward a broader implementation of CNT-based filtration devices., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The flexible batteries that are needed to power flexible circuits and displays remain challenging, despite considerable progress in the fabrication of such devices. Here, it is shown that flexible batteries can be fabricated using arrays of carbon nanotube microstructures, which decouple stress from the energy-storage material. It is found that this battery architecture imparts exceptional flexibility (radius ≈ 300 μm), high rate (20 A g(-1) ), and excellent cycling stability., (© 2016 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

An advanced battery architecture composed of 3D carbon nanotube (CNT) current collectors is used to mitigate stresses in flexible batteries. On Page 6705, C. George, M. De Volder, and co-workers describe the fabrication process and characteristics of this new generation of ultraflexible batteries, which show high rate and cyclablility. These batteries may find applications in the powering of flexible displays and logics., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Liquid crystalline polymers have recently been engineered to exhibit complex macroscopic shape adaptivity, including optically- and thermally driven bending, self-sustaining oscillation, torsional motion, and three-dimensional folding. Miniaturization of these novel materials is of great interest for both fundamental study of processing conditions and for the development of shape-changing microdevices. Here, we present a scalable method for high-fidelity replica molding of glassy liquid crystalline polymer networks (LCNs), by vacuum-assisted replica molding, along with magnetic field-induced control of the molecular alignment. We find that an oxygen-free environment is essential to establish high-fidelity molding with low surface roughness. Identical arrays of homeotropic and polydomain LCN microstructures are fabricated to assess the influence of molecular alignment on the elastic modulus (E = 1.48 GPa compared to E = 0.54 GPa), and side-view imaging is used to quantify the reversible thermal actuation of individual LCN micropillars by high-resolution tracking of edge motion. The methods and results from this study will be synergistic with future advances in liquid crystalline polymer chemistry, and could enable the scalable manufacturing of stimuli-responsive surfaces for applications including microfluidics, tunable optics, and surfaces with switchable wetting and adhesion.

High performance active materials are of rapidly growing interest for applications including soft robotics, microfluidic systems, and morphing composites. In particular, paraffin wax has been used to actuate miniature pumps, solenoid valves, and composite fibers, yet its deployment is typically limited by the need for external volume constraint. We demonstrate that compact, high-performance paraffin actuators can be made by confining paraffin within vertically aligned carbon nanotube (CNT) films. This large-stroke vertical actuation is enabled by strong capillary interaction between paraffin and CNTs and by engineering the CNT morphology by mechanical compression before capillary-driven infiltration of the molten paraffin. The maximum actuation strain of the corrugated CNT-paraffin films (∼0.02-0.2) is comparable to natural muscle, yet the maximum stress is limited to ∼10 kPa by collapse of the CNT network. We also show how a CNT-paraffin film can serve as a self-activating thermal interface that closes a gap when it is heated. These new CNT-paraffin film actuators could be produced by large-area CNT growth, infiltration, and lamination methods, and are attractive for use in miniature systems due to their self-contained design.