Farnesoid X Receptor Protects Murine Lung against IL-6-promoted Ferroptosis Induced by Polyriboinosinic-Polyribocytidylic Acid.

Various infections trigger a storm of proinflammatory cytokines in which IL-6 acts as a major contributor and leads to diffuse alveolar damage in patients. However, the metabolic regulatory mechanisms of IL-6 in lung injury remain unclear. Polyriboinosinic-polyribocytidylic acid [poly(I:C)] activates pattern recognition receptors involved in viral sensing and is widely used in alternative animal models of RNA virus-infected lung injury. In this study, intratracheal instillation of poly(I:C) with or without an IL-6-neutralizing antibody model was combined with metabonomics, transcriptomics, and so forth to explore the underlying molecular mechanisms of IL-6-exacerbated lung injury. We found that poly(I:C) increased the IL-6 concentration, and the upregulated IL-6 further induced lung ferroptosis, especially in alveolar epithelial type II cells. Meanwhile, lung regeneration was impaired. Mechanistically, metabolomic analysis showed that poly(I:C) significantly decreased glycolytic metabolites and increased bile acid intermediate metabolites that inhibited the bile acid nuclear receptor farnesoid X receptor (FXR), which could be reversed by IL-6-neutralizing antibody. In the ferroptosis microenvironment, IL-6 receptor monoclonal antibody tocilizumab increased FXR expression and subsequently increased the Yes-associated protein (YAP) concentration by enhancing PKM2 in A549 cells. FXR agonist GW4064 and liquiritin, a potential natural herbal ingredient as an FXR regulator, significantly attenuated lung tissue inflammation and ferroptosis while promoting pulmonary regeneration. Together, the findings of the present study provide the evidence that IL-6 promotes ferroptosis and impairs regeneration of alveolar epithelial type II cells during poly(I:C)-induced murine lung injury by regulating the FXR-PKM2-YAP axis. Targeting FXR represents a promising therapeutic strategy for IL-6-associated inflammatory lung injury.

CD34+ cell-derived fibroblast-macrophage cross-talk drives limb ischemia recovery through the OSM-ANGPTL signaling axis.

CD34+ cells improve the perfusion and function of ischemic limbs in humans and mice. However, there is no direct evidence of the differentiation potential and functional role of these cells in the ischemic muscle microenvironment. Here, we combined the single-cell RNA sequencing and genetic lineage tracing technology, then provided exact single-cell atlases of normal and ischemic limb tissues in human and mouse, and consequently found that bone marrow (BM)-derived macrophages with antigen-presenting function migrated to the ischemic site, while resident macrophages underwent apoptosis. The macrophage oncostatin M (OSM) regulatory pathway was specifically turned on by ischemia. Simultaneously, BM CD34+-derived proregenerative fibroblasts were recruited to the ischemia niche, where they received macrophage-released OSM and promoted angiopoietin-like protein-associated angiogenesis. These findings provided mechanisms on the cellular events and cell-cell communications during tissue ischemia and regeneration and provided evidence that CD34+ cells serve as fibroblast progenitors promoting tissue regeneration.

B cell-derived anti-beta 2 glycoprotein I antibody mediates hyperhomocysteinemia-aggravated hypertensive glomerular lesions by triggering ferroptosis.

Hyperhomocysteinemia (HHcy) is a risk factor for chronic kidney diseases (CKDs) that affects about 85% CKD patients. HHcy stimulates B cells to secrete pathological antibodies, although it is unknown whether this pathway mediates kidney injury. In HHcy-treated 2-kidney, 1-clip (2K1C) hypertensive murine model, HHcy-activated B cells secreted anti-beta 2 glycoprotein I (ß2GPI) antibodies that deposited in glomerular endothelial cells (GECs), exacerbating glomerulosclerosis and reducing renal function. Mechanistically, HHcy 2K1C mice increased phosphatidylethanolamine (PE) (18:0/20:4, 18:0/22:6, 16:0/20:4) in kidney tissue, as determined by lipidomics. GECs oxidative lipidomics validated the increase of oxidized phospholipids upon Hcy-activated B cells culture medium (Hcy-B CM) treatment, including PE (18:0/20:4 + 3[O], PE (18:0a/22:4 + 1[O], PE (18:0/22:4 + 2[O] and PE (18:0/22:4 + 3[O]). PE synthases ethanolamine kinase 2 (etnk2) and ethanolamine-phosphate cytidylyltransferase 2 (pcyt2) were increased in the kidney GECs of HHcy 2K1C mice and facilitated polyunsaturated PE synthesis to act as lipid peroxidation substrates. In HHcy 2K1C mice and Hcy-B CM-treated GECs, the oxidative environment induced by iron accumulation and the insufficient clearance of lipid peroxides caused by transferrin receptor (TFR) elevation and down-regulation of SLC7A11/glutathione peroxidase 4 (GPX4) contributed to GECs ferroptosis of the kidneys. In vivo, pharmacological depletion of B cells or inhibition of ferroptosis mitigated the HHcy-aggravated hypertensive renal injury. Consequently, our findings uncovered a novel mechanism by which B cell-derived pathogenic anti-ß2GPI IgG generated by HHcy exacerbated hypertensive kidney damage by inducing GECs ferroptosis. Targeting B cells or ferroptosis may be viable therapeutic strategies for ameliorating lipid peroxidative renal injury in HHcy patients with hypertensive nephropathy.

T lymphocyte-derived extracellular vesicles aggravate abdominal aortic aneurysm by promoting macrophage lipid peroxidation and migration via pyruvate kinase muscle isozyme 2.

T lymphocyte and macrophage infiltration in the aortic wall is critical for abdominal aortic aneurysm (AAA). However, how T lymphocytes interact with macrophages in the pathogenesis of AAA remains largely uncharacterized. In an elastase-induced murine AAA model, we first found that the expression of pyruvate kinase muscle isozyme 2 (PKM2), the last rate-limiting enzyme in glycolysis, was increased in infiltrated T lymphocytes of vascular lesions. T lymphocyte-specific PKM2 deficiency in mice (LckCrePKM2fl/fl) or intraperitoneal administration of the sphingomyelinase inhibitor GW4869 caused a significant attenuation of the elastase-increased aortic diameter, AAA incidence, elastic fiber disruption, matrix metalloproteinases (MMPs) expression, and macrophage infiltration in the vascular adventitia compared with those in PKM2fl/fl mice. Mechanistically, extracellular vesicles (EVs) derived from PKM2-activated T lymphocytes elevated macrophage iron accumulation, lipid peroxidation, and migration in vitro, while macrophages treated with EVs from PKM2-null T lymphocytes or pretreated with the lipid peroxidation inhibitors ferrostatin-1 (Fer-1), liproxstatin-1 (Lip-1), or the iron chelating agent deferoxamine mesylate (DFOM) reversed these effects. In vascular lesions of elastase-induced LckCrePKM2fl/fl mice with AAA, the oxidant system weakened, with downregulated 4-hydroxynonenal (4-HNE) levels and strengthened antioxidant defense systems with upregulated glutathione peroxidase 4 (GPX4) and cystine/glutamate antiporter solute carrier family 7 member 11 (Slc7a11) expressions in macrophages. High-throughput metabolomics showed that EVs derived from PKM2-activated T lymphocytes contained increased levels of polyunsaturated fatty acid (PUFA)-containing phospholipids, which may provide abundant substrates for lipid peroxidation in target macrophages. More importantly, upregulated T lymphocyte PKM2 expression was also found in clinical AAA subjects, and EVs isolated from AAA patient plasma enhanced macrophage iron accumulation, lipid peroxidation, and migration ex vivo. Therefore, from cell-cell crosstalk and metabolic perspectives, the present study shows that PKM2-activated T lymphocyte-derived EVs may drive AAA progression by promoting macrophage redox imbalance and migration, and targeting the T lymphocyte-EV-macrophage axis may be a potential strategy for early warning and treating AAA.

Exosomes and Atherogenesis.

Myocardial infarction and ischemic stroke are the leading causes of mortality worldwide. Atherosclerosis is their common pathological foundation. It is known that atherosclerosis is characterized by endothelial activation/injury, accumulation of inflammatory immune cells and lipid-rich foam cells, followed by the development of atherosclerotic plaque. Either from arterial vessel wall or blood circulation, endothelial cells, smooth muscle cells, macrophages, T-lymphocytes, B-lymphocytes, foam cells, and platelets have been considered to contribute to the pathogenesis of atherosclerosis. Exosomes, as natural nano-carriers and intercellular messengers, play a significant role in modulation of cell-to-cell communication. Under physiological or pathological conditions, exosomes can deliver their cargos including donor cell-specific proteins, lipids, and nucleic acids to target cells, which in turn affect the function of the target cells. In this review, we will describe the pathophysiological significance of various exosomes derived from different cell types associated with atherosclerosis, and the potential applications of exosome in clinical diagnosis and treatment.

Shikonin attenuates hyperhomocysteinemia-induced CD4+ T cell inflammatory activation and atherosclerosis in ApoE-/- mice by metabolic suppression.

T cell metabolic activation plays a crucial role in inflammation of atherosclerosis. Shikonin (SKN), a natural naphthoquinone with anti-inflammatory activity, has shown to exert cardioprotective effects, but the effect of SKN on atherosclerosis is unclear. In addition, SKN was found to inhibit glycolysis via targeting pyruvate kinase muscle isozyme 2 (PKM2). In the present study, we investigated the effects of SKN on hyperhomocysteinemia (HHcy)-accelerated atherosclerosis and T cell inflammatory activation in ApoE-/- mice and the metabolic mechanisms in this process. Drinking water supplemented with Hcy (1.8 g/L) was administered to ApoE-/- mice for 2 weeks and the mice were injected with SKN (1.2 mg/kg, i.p.) or vehicle every 3 days. We showed that SKN treatment markedly attenuated HHcy-accelerated atherosclerosis in ApoE-/- mice and significantly decreased inflammatory activated CD4+ T cells and proinflammatory macrophages in plaques. In splenic CD4+ T cells isolated from HHcy-ApoE-/- mice, SKN treatment significantly inhibited HHcy-stimulated PKM2 activity, interferon-γ secretion and the capacity of these T cells to promote macrophage proinflammatory polarization. SKN treatment significantly inhibited HHcy-stimulated CD4+ T cell glycolysis and oxidative phosphorylation. Metabolic profiling analysis of CD4+ T cells revealed that Hcy administration significantly increased various glucose metabolites as well as lipids and acetyl-CoA carboxylase 1, which were reversed by SKN treatment. In conclusion, our results suggest that SKN is effective to ameliorate atherosclerosis in HHcy-ApoE-/- mice and this is at least partly associated with the inhibition of SKN on CD4+ T cell inflammatory activation via PKM2-dependent metabolic suppression.

T-cell-derived extracellular vesicles regulate B-cell IgG production via pyruvate kinase muscle isozyme 2.

Intercellular communication between lymphocytes plays a fundamental role in numerous immune responses. Previously, we demonstrated that hyperhomocysteinemia (HHcy) induced T cell intracellular glycolytic-lipogenic reprogramming and IFN-γ secretion via pyruvate kinase muscle isozyme 2 (PKM2) to accelerate atherosclerosis. Usually, B cells partially obtain help from T cells in antibody responses. However, whether PKM2 activation in T cells regulates B cell antibody production is unknown. Extracellular vesicles (EVs) are important cellular communication vehicles. Here, we found that PKM2 activator TEPP46-stimulated T-cell-derived EVs promoted B-cell IgG secretion. Conversely, EVs secreted from PKM2-null T cells were internalized into B cells and markedly inhibited B-cell mitochondrial programming, activation, and IgG production. Mechanistically, lipidomics analyses showed that increased ceramides in PKM2-activated T-cell EVs were mainly responsible for enhanced B cell IgG secretion induced by these EVs. Finally, quantum dots (QDs) were packaged with PKM2-null T cell EVs and anti-CD19 antibody to exert B-cell targeting and inhibit IgG production, eventually ameliorating HHcy-accelerated atherosclerosis in vivo. Thus, PKM2-mediated EV ceramides in T cells may be an important cargo for T-cell-regulated B cell IgG production, and QD-CD19-PKM2-null T cell EVs hold high potential to treat B cell overactivation-related diseases.-Yang, J., Dang, G., Lü, S., Liu, H., Ma, X., Han, L., Deng, J., Miao, Y., Li, X., Shao, F., Jiang, C., Xu, Q., Wang, X., Feng, J. T-cell-derived extracellular vesicles regulate B-cell IgG production via pyruvate kinase muscle isozyme 2.