Patrycja Kwiecińska

Multiple mechanisms contribute to skin barrier integrity, and emerging evidence suggests that acyl-CoA:diacylglycerol acyltransferase 1 (DGAT1) plays a key role in its regulation through both lipid and retinoid metabolism. Although DGAT1 deficiency leads to cutaneous abnormalities, the direct role of DGAT1 in keratinocyte biology remains unclear. Here, we show that DGAT1 deficiency drives extensive keratinocyte hyperproliferation and barrier dysfunction under homeostatic conditions. Transcriptomic profiling revealed upregulation of genes associated with proliferation, inflammation, and retinoic acid (RA) signaling, consistent with increased proliferation observed in vitro and in vivo . Unexpectedly, while DGAT1 expression was elevated in human psoriatic lesions and in a murine psoriasis model, DGAT1 loss reduced keratinocyte proliferation in this inflammatory condition. Further analyses identified dysregulated RA signaling as a central mediator of these effects. DGAT1-deficient keratinocytes exhibited elevated RA-target gene expression and increased expression of RA-binding proteins, indicating enhanced RA bioavailability. Our findings establish DGAT1 as a context-dependent regulator of keratinocyte proliferation, promoting growth in homeostasis while restraining it in psoriasis. By linking DGAT1 activity with RA signaling, this work uncovers a previously unrecognized pathway contributing to epidermal physiology and inflammatory skin disease.

Background: Recombinant granulocyte colony-stimulating factor (G-CSF) is the most commonly used agent for treating neutropenia and mobilizing hematopoietic stem cells (HSCs) for transplantation. However, some patients do not respond effectively to the currently used mobilization protocols. To address this, new therapeutic approaches are needed. A potential strategy is pharmacological induction of endogenous mobilizing factors via cobalt protoporphyrin IX (CoPP). CoPP mobilizes HSCs and granulocytes by increasing endogenous G-CSF, though optimal dosing and potential side effects remain unclear. Our study aimed to optimize CoPP dosing and timing, and assess its safety in mobilizing cells from bone marrow to blood. Methods: Mice were treated with different doses of CoPP, and blood cell counts, cytokine concentrations, and organ damage markers were evaluated at various time points after injection. Results: Our results show that CoPP exerts a dose-dependent mobilizing effect, with the highest G-CSF levels and number of mobilized leukocytes observed in mice treated with 10 mg/kg of CoPP. While there were no severe adverse effects, there were mild fluctuations in markers of organ function, including a reduction in blood urea nitrogen (BUN) and glucose levels during the five days of administration. Additionally, although most parameters normalized within 30 days, the decrease in BUN persisted. Mice experienced short-term weight loss following CoPP administration, but they regained their initial weight within two weeks. Conclusions: This study demonstrates that CoPP mobilizes cells from the bone marrow to the blood in a dose-dependent manner, with mild side effects, including temporary changes in biochemical markers and a sustained reduction in BUN levels.

Skin infiltration by neutrophils is a hallmark of the chronic inflammatory skin disease psoriasis, yet the mechanisms underlying neutrophil recruitment and positioning in chronically inflamed skin remain poorly understood. In this study, we demonstrate the significant impact of a total genetic deficiency of secretory leukocyte protease inhibitor (SLPI) on neutrophil migration in mouse skin. Without SLPI, neutrophils displayed an unconventional migratory pattern, characterized by altered interactions with vessel walls and reduced efficiency in extravasating from blood vessels into skin tissue during the early stages of experimental psoriasis. This was associated with changes in tissue motility, positioning neutrophils farther from the skin entry vessels and closer to the skin surface. Neutrophil diapedesis was partially dependent on SLPI within the neutrophils themselves. The impact of SLPI on neutrophil movement was further supported by the increased migration of human neutrophils in the presence of neutrophil-penetrant recombinant SLPI. Additionally, our data suggest that neutrophils with varying capacities for vessel wall interaction are released from the bone marrow into circulation in an SLPI-dependent manner. These findings establish a role for SLPI in regulating the spatiotemporal infiltration of neutrophils into the skin in psoriasis, highlighting its relevance to psoriasis pathophysiology.

Hematopoietic stem cells (HSCs) produce all blood cells throughout the lifespan of the organism. HSC requires a specialized bone marrow microenvironment, known as the niche, for proper differentiation and self-renewal. While several molecular and cellular elements of the niche are described, the precise understanding of the mechanobiology underlying HSC adhesion to the niche remains poorly understood. Here, we aim to characterize physical interactions and adhesion within the hematopoietic niche by combining cell sorting, surface functionalization, and atomic force microscopy. Using this approach, we quantified and compared the adhesion of bone marrow (BM) mesenchymal stromal cells (MSCs) to different extracellular proteins, as well as the adhesion of HSCs to MSCs. We observed that MSCs adhere with the greatest force to fibronectin in an Arg-Gly-As (RGD) motif mediated and actin cytoskeleton-dependent manner. Additionally, we showed that HSCs strongly adhere to MSC within 30 seconds and that the binding is RGD-independent. In conclusion, we demonstrated how to implement atomic force microscopy to measure the physiological interactions within the HSCs niche in a direct, specific, and quantitative way. This approach provides a more comprehensive and precise characterization of the biology of the HSC niche compared to previously used methods. Furthermore, it highlights the complexity of studying extremely small structures and rare cell populations in biophysical experiments.

Recombinant granulocyte colony-stimulating factor (G-CSF) is the most commonly used agent for treating neutropenia and mobilizing hematopoietic stem cells (HSCs) for transplantation. However, some patients do not respond effectively to existing mobilization protocols. To address this, the development of new therapeutic approaches is necessary. One potential strategy is the pharmacological induction of endogenous mobilizing factors, which can be achieved through the administration of cobalt protoporphyrin IX (CoPP). CoPP induces mobilization of HSCs and granulocytes by increasing endogenous G-CSF production, though the optimal dosing and potential side effects remain unclear. The aim of our study was to optimize the dose and timing of CoPP administration and evaluate its safety in mobilizing cells from the bone marrow to the blood. Our results show that CoPP exerts a dose-dependent mobilizing effect, with the highest G-CSF levels and number of mobilized leukocytes observed in mice treated with 10 mg/kg of CoPP. While there were no severe adverse effects, there were mild fluctuations in markers of liver and kidney function, including a slight reduction in urea nitrogen (BUN) and glucose levels during the five days of administration. Additionally, although most parameters normalized within 30 days after treatment, the decrease in BUN persisted. Mice experienced short-term weight loss following CoPP administration, but they regained their initial weight within two weeks. By day 30, leukocyte counts, hematopoietic stem and progenitor cells (HSPCs) in bone marrow, and G-CSF concentration in the blood had returned to baseline. This study demonstrates that CoPP mobilizes cells from the bone marrow to the blood in a dose-dependent manner, with mild side effects, including temporary changes in biochemical markers and a sustained reduction in BUN levels.

Background: Neutrophils are key players in the innate immune system, actively migrating to sites of inflammation in the highly energetic process of chemotaxis. In this study, we focus on the role of acyl-CoA: diacylglycerol acyltransferase 1 (DGAT1), an enzyme that catalyzes the synthesis of triglycerides, the major form of stored energy, in neutrophil chemotaxis. Methods and results: Using a mouse model of psoriasis, we show that DGAT1-deficiency reduces energy-demanding neutrophil infiltration to the site of inflammation, but this inhibition is not caused by decreased glycolysis and reduced ATP production by neutrophils lacking DGAT1. Flow cytometry and immunohistochemistry analysis demonstrate that DGAT1 also does not influence lipid accumulation in lipid droplets during inflammation. Interestingly, as has been shown previously, a lack of DGAT1 leads to an increase in the concentration of retinoic acid, and here, using real-time PCR and publicly-available next-generation RNA sequencing datasets, we show the upregulation of retinoic acid-responsive genes in Dgat1KO neutrophils. Furthermore, supplementation of WT neutrophils with exogenous retinoic acid mimics DGAT1-deficiency in the inhibition of neutrophil chemotaxis in in vitro transwell assay. Conclusions: These results suggest that impaired skin infiltration by neutrophils in Dgat1KO mice is a result of the inhibitory action of an increased concentration of retinoic acid, rather than impaired lipid metabolism in DGAT1-deficient mice.

The members of the ZC3H12/MCPIP/Regnase family of RNases have emerged as important regulators of inflammation. In contrast to Regnase-1, -2 and -4, a thorough characterization of Regnase-3 (Reg-3) has not yet been explored. Here we demonstrate that Reg-3 differs from other family members in terms of NYN/PIN domain features, cellular localization pattern and substrate specificity. Together with Reg-1, the most comprehensively characterized family member, Reg-3 shared IL-6, IER-3 and Reg-1 mRNAs, but not IL-1β mRNA, as substrates. In addition, Reg-3 was found to be the only family member which regulates transcript levels of TNF, a cytokine implicated in chronic inflammatory diseases including psoriasis. Previous meta-analysis of genome-wide association studies revealed Reg-3 to be among new psoriasis susceptibility loci. Here we demonstrate that Reg-3 transcript levels are increased in psoriasis patient skin tissue and in an experimental model of psoriasis, supporting the immunomodulatory role of Reg-3 in psoriasis, possibly through degradation of mRNA for TNF and other factors such as Reg-1. On the other hand, Reg-1 was found to destabilize Reg-3 transcripts, suggesting reciprocal regulation between Reg-3 and Reg-1 in the skin. We found that either Reg-1 or Reg-3 were expressed in human keratinocytes in vitro. However, in contrast to robustly upregulated Reg-1 mRNA levels, Reg-3 expression was not affected in the epidermis of psoriasis patients. Taken together, these data suggest that epidermal levels of Reg-3 are negatively regulated by Reg-1 in psoriasis, and that Reg-1 and Reg-3 are both involved in psoriasis pathophysiology through controlling, at least in part different transcripts.

Epithelia in the skin, gut and other environmentally exposed organs display a variety of mechanisms to control microbial communities and limit potential pathogenic microbial invasion. Naturally occurring antimicrobial proteins/peptides and their synthetic derivatives (here collectively referred to as AMPs) reinforce the antimicrobial barrier function of epithelial cells. Understanding how these AMPs are functionally regulated may be important for new therapeutic approaches to combat microbial infections. Some AMPs are subject to redox-dependent regulation. This review aims to: (i) explore cysteine-based redox active AMPs in skin and intestine; (ii) discuss casual links between various redox environments of these barrier tissues and the ability of AMPs to control cutaneous and intestinal microbes; (iii) highlight how bacteria, through intrinsic mechanisms, can influence the bactericidal potential of redox-sensitive AMPs.