Dietary polyphenols, which are present in Aronia melanocarpa, have been associated with various beneficial effects on human health including antioxidant, antiviral, and anti-inflammatory activities. We aimed to investigate the immunomodulatory effects of aronia juice polyphenols in a randomized placebo-controlled human intervention study and cell culture experiments. A total of 40 females were asked to consume either 200 mL of aronia juice or a placebo drink for six weeks and were investigated again after a washout period of another six weeks. We observed that only half of the participants tolerated the aronia juice well (Vt) and the other half reported complaints (Vc). The placebo (P) was generally tolerated with one exception (p = 0.003). Plasma polyphenol levels increased significantly in Vt after the intervention (p = 0.024) but did neither in P nor in Vc. Regulatory T cell (Treg) frequencies remained constant in Vt and P during the intervention, whereas Tregs decreased in Vc (p = 0.018). In cell culture, inhibiting effects of ferulic acid (p = 0.0005) and catechin (p = 0.0393) on the differentiation of Tregs were observed as well as reduced activation of CD4-T cells in ferulic acid (p = 0.0072) and aronia juice (p = 0.0163) treated cells. Interestingly, a CD4⁺CD25⁻FoxP3⁺ cell population emerged in vitro in response to aronia juice, but not when testing individual polyphenols. In conclusion, our data strengthen possible individual hormetic effects, the importance of the food matrix for bioactivity, and the need for further investigations on possible impacts of specific physiological features such as the gut microbiota in the context of personalized nutrition.

Endoplasmic reticulum (ER) stress is a key feature of lipid-laden macrophages and contributes to the development of atherosclerotic plaques. Blood platelets are known to interact with macrophages and fine-tune effector functions such as inflammasome activation and phagocytosis. However, the effect of platelets on ER stress induction is unknown. The objective of this study is to elucidate the potential of platelets in regulating ER stress in macrophages in vitro. Bone marrow-derived macrophages and RAW 264.7 cells were incubated with isolated murine platelets, and ER stress and inflammation markers were determined by reverse transcription-quantitative polymerase chain reaction, Western blotting, and enzyme-linked immunosorbent assay. ER morphology was investigated by electron microscopy. Cell viability, lipid accumulation, and activation were measured by flow cytometry. To gain mechanistic insights, coincubation experiments were performed with platelet decoys/releasates as well as lipopolysaccharide, blocking antibodies, and TLR4 inhibitors. Coincubation of platelets and macrophages led to elevated levels of ER stress markers (BIP, IRE1α, CHOP, and XBP1 splicing) in murine and human macrophages, which led to a pronounced enlargement of the ER. Macrophage ER stress was accompanied by increased release of proinflammatory cytokines and intracellular lipid accumulation, but not cell death. Platelet decoys, but not platelet releasates or lysate from other cells, phenocopied the effect of platelets. Blocking TLR4 inhibited inflammatory activation of macrophages but did not affect ER stress induction by platelet coincubation. To our knowledge, this study is the first to demonstrate that platelets induce ER stress and unfolded protein response in macrophages by heat-sensitive membrane proteins, independent of inflammatory activation of macrophages.

BACKGROUND: Perivascular adipose tissue (PVAT) fine-tunes blood vessel contractility and vascular homeostasis. During obesity and atherosclerosis, PVAT becomes dysfunctional and loses its anticontractile potential. Previously, we reported that global knockout of adipose triglyceride lipase (ATGL), the major enzyme responsible for the breakdown of triglycerides, has the potential to modify PVAT functions. To address the causal relationship between PVAT lipolysis and blood vessel contractility, we analyzed ex vivo vasomotor function of mice with tissue-specific rescue/overexpression or knockout of ATGL in adipose tissue. METHODS: To generate mice lacking ATGL in all tissues except for adipose tissue (ATGL knockout with adipocyte-specific expression of ATGL [A+/AKO]), we crossed adipocyte ATGL-rescued (A+) mice with ATGL-deficient (ATGL knockout [AKO]) mice. Body weight, plasma levels of fatty acids, and blood glucose were compared between A+/AKO and AKO mice. Ex vivo vasoreactivity studies were performed in the absence and presence of PVAT to test for acute and chronic effects of PVAT on vascular function. RESULTS: Adipocyte-rescued AKO mice (A+/AKO) had significantly less amounts of PVAT than AKO controls while displaying moderate ATGL expression. A+/AKO aortas exhibited decreased anticontractile effects of PVAT compared with AKO aortas. This effect on contractile function was observed in an agonist-specific manner without affecting smooth muscle cell function or endothelium-dependent relaxation. Assessment of cardiac function using the Langendorff setup revealed that adipocyte ATGL selectively modulated vascular contractility without affecting systolic or diastolic performance. Studies using mice that express ATGL solely in cardiac muscle and adipocyte-specific ATGL knockout mice verified our findings in A+/AKO mice, revealing acute and chronic effects of adipocyte lipolysis on vasoreactivity. CONCLUSIONS: We provide the first evidence that changes in adipocyte lipolysis have the potential to regulate blood vessel contractility. Ablation of ATGL in adipocytes decreases vascular contractility and, thus, has the potential to prevent PVAT dysfunction in obesity and atherosclerosis.

The immunological barrier is among the most significant barriers in vivo. Macrophages and dendritic cells play a crucial role in immune responses, involving phagocytosis, antigen presentation, and triggering adaptive responses. Nanoscale drug-delivery vehicles, such as polymer-encapsulated lipid-bilayer nanodiscs, are of particular interest in the development of new therapeutic approaches, but require well-characterized human in vitro cell models. To this end, the present study differentiated human monocytes into two distinct states, resting macrophages and immature dendritic-like cells (iDCs). These cells served as model systems to assess the efficacy of lipid-bilayer nanodiscs encapsulated by anionic glyco-DIBMA (diisobutylene–maleic acid) or electroneutral sulfo-DIBMA polymers. Nanodisc–cell interaction studies—including cell viability, reactive oxygen species production, cytokine release, particle uptake, and activation marker expression—demonstrated that immune responses depend sensitively on the cell type and polymer and thus on the surface charge of the nanodiscs. Sulfo-DIBMA nanodiscs induced minimal immune cell activation, accompanied by cytokine release and reduced uptake of the nanodiscs by immune cells. In contrast, glyco-DIBMA nanodiscs exhibited increased interactions with cells, elicited pro-inflammatory immune responses, and promoted iDC maturation. This involved co-stimulatory and antigen-presenting molecules, potentially leading to T-cell activation. These findings underscore the potential of glyco-DIBMA nanodiscs to modulate immune responses through receptor-specific interactions, paving the way for immunotherapeutic strategies.

Atherosclerosis is a chronic, inflammatory disease of the vessel wall where triggered immune cells bind to inflamed endothelium, extravasate and sustain local inflammation. Leukocyte adhesion and extravasation are mediated by adhesion molecules expressed by activated endothelial cells, like intercellular adhesion molecule 1 (ICAM-1). Extracellular adherence protein (Eap) from Staphylococcus aureus binds to a plethora of extracellular matrix proteins, including ICAM-1 and its ligands macrophage-1 antigen (Mac-1, α_Mβ₂) and lymphocyte function-associated antigen 1 (LFA-1, α_Lβ₂), thereby disrupting the interaction between leukocytes and endothelial cells. We aimed to use Eap to inhibit the interaction of leukocytes with activated endothelial cells in settings of developing and established atherosclerosis in apolipoprotein E (ApoE) deficient mice on high-fat diet. In developing atherosclerosis, Eap treatment reduced circulating platelet-neutrophil aggregates as well as infiltration of T cells and neutrophils into the growing plaque, accompanied by reduced formation of neutrophil extracellular traps (NETs). However, plaque size did not change. Intervention treatment with Eap of already established plaques did not result in cellular or morphological plaque changes, whereas T cell infiltration was increased and thereby again modulated by Eap. We conclude that although Eap leads to cellular changes in developing plaques, clinical implications might be limited as patients are usually treated at a more advanced stage of disease progression. Hence, usage of Eap might be an interesting mechanistic tool for cellular infiltration during plaque development in basic research but not a clinical target.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is the most common chronic liver disease, characterized by endoplasmic reticulum (ER) stress, which impairs hepatic lipoprotein metabolism. Blood platelets can modulate cellular responses to ER stress and accumulate in the liver during MASLD, contributing to disease development through interaction with liver-resident (immune) cells. Given the role of platelets in modulating hepatic ER stress in vitro, this study aims to investigate their effects on hepatic ER stress and steatosis. C57BL/6 mice (n = 6-12) were depleted for platelets and injected with tunicamycin (1 μg/g) to induce ER stress-induced hepatic steatosis. Hepatic platelet infiltration and activation was investigated by flow cytometry, ELISA, and electron microscopy. Livers were analyzed for ER stress markers by quantitative PCR, Western blotting, proteomics, and flow cytometry. Thereafter, platelet-hepatocyte co-cultures (n=4) and in vivo blocking of GPIbα (n = 5-10) were used for further mechanistic investigations. Our study demonstrates that hepatic ER stress leads to 1.8-fold accumulation (p = 0.007) of platelets. Depletion of platelets decreased ER stress-induced liver steatosis by 35% (p <0.0001). Mechanistically, we found that platelet depletion ameliorated the initial ER stress response and increased the influx of neutrophils into the liver. Inhibiting the platelet-specific receptor glycoprotein 1bα (GPIbα) partly mimicked the effects of platelet depletion, leading to impaired hepatic platelet recruitment and reduced triglyceride accumulation. This study reveals a new role of platelets in hepatic ER stress and the concomitant effect on hepatic steatosis. This paper shows that blood platelets are capable of regulating specific metabolic functions in liver cells (so-called ER stress) and thereby contribute to the development of hepatic steatosis.