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Background: Cardiac iron overload and ferroptosis greatly contribute to the poor prognosis of myocardial infarction (MI). Iron chelator is one of the most promising strategies for scavenging excessive iron and alleviating cardiac dysfunction post MI. However, various side effects of existing chemical iron chelators restrict their clinical application, which calls for a more viable and safer approach to protect against iron injury in ischemic hearts., Results: In this study, we isolated macrophage-derived extracellular vesicles (EVs) and identified macrophage-derived EVs as a novel endogenous biological chelator for iron. The administration of macrophage-derived EVs effectively reduced iron overload in hypoxia-treated cardiomyocytes and hearts post MI. Moreover, the oxidative stress and ferroptosis induced by excessive iron were considerably suppressed by application of macrophage-derived EVs. Mechanistically, transferrin receptor (TfR), which was inherited from macrophage to the surface of EVs, endowed EVs with the ability to bind to transferrin and remove excess protein-bound iron. EVs with TfR deficiency exhibited a loss of function in preventing MI-induced iron overload and protecting the heart from MI injury. Furthermore, the iron-chelating EVs were ultimately captured and processed by macrophages in the liver., Conclusions: These results highlight the potential of macrophage-derived EVs as a powerful endogenous candidate for iron chelation therapy, offering a novel and promising therapeutic approach to protect against iron overload-induced injury in MI and other cardiovascular diseases., (© 2024. The Author(s).)

Introduction: Atherosclerosis is one of the major causes of cardiovascular diseases. Lipid uptake and accumulation in macrophages play a major role in atherosclerotic plaque formation from its initiation to advanced atheroma formation. The dipeptidyl peptidase-4 (DPP-4) inhibitor Linagliptin is commonly used to lower blood glucose in type 2 diabetes patients. Recent studies report that Linagliptin has cardiovascular protective and anti-inflammatory effects., Methods: THP-1 macrophage cells were treated with 100 nM PMA for 72 hour to induce foam cell formation. The differentiated cells were exposed to 100 μg/mL ox-LDL in the presence or absence of the DPP-4 inhibitor Linagliptin. The expression levels of DPP-4 and inflammatory cytokines were detected by RT-PCR, ELISA, and Western blot experiments. The cellular ROS level was measured by staining the cells with the fluorescent probe DCFH-DA. The separation of lipoprotein fractions was achieved by high-performance liquid chromatography (HPLC). The cells were labeled with fluorescent-labeled cholesterol to measure cholesterol efflux, and lipid droplets were revealed by Nile red staining., Results: The presence of Linagliptin significantly reduced ox-LDL-induced cytokine production (IL-1β and IL-6) and ROS production. Linagliptin ameliorated ox-LDL-induced lipid accumulation and impaired cholesterol efflux in macrophages. Mechanistically, this study showed that Linagliptin mitigated ox-LDL-induced expression of the scavenger receptors CD36 and LOX-1, but not SRA. Furthermore, Linagliptin increased the expression of the cholesterol transporter ABCG1, but not ABCA1., Conclusion: Linagliptin possesses a potent inhibitory effect on THP-1 macrophage-derived foam cell formation in response to ox-LDL. This effect could be mediated through a decrease in the expression of CD36 and LOX-1 on macrophages and an increase in the expression of the cholesterol transporter ABCG1. This study indicates that the DPP-4 inhibitor Linagliptin plays a critical role in preventing foam cell formation in vitro. However, future research using an atherosclerotic animal model is necessary to determine its effectiveness and to prove its potential implication in the prevention and treatment of atherosclerosis., Competing Interests: The authors report no conflicts of interest for this work., (© 2020 Wang et al.)

Objective: Macrophage infiltration in kidney is a major pathological feature of diabetic nephropathy (DN), which has been demonstrated associate with macrophages autophagy homeostasis. However, the relationships between autophagy and the infiltration response related of macrophages adhesion and migration are unknown. This study aims to investigate the impact of macrophages adhesion and migration by modulating autophagy. Methods: In vivo, rats were randomly distributed into control (NC) and DN groups. The pathological changes in renal tissue were assessed, and expression of CD68, LC3, P62 were analyzed. In vitro, RAW264.7 cells were divided into NC and high glucose (HG) groups. The capacity of macrophages adhesion migration and the expression of autophagy markers were observed with and without autophagy modulators (rapamycin, 3-methyladenine, chloroquine, and bafilomycin A1 for RAPA, 3-MA, CQ, BAFA). The macrophages autophagosome and the process of degradation and fusion of autophagosome-lysosome were observed by electron microscopy. Results: In vivo, renal injury is aggravated in diabetic rat compared with NC group. The autophagy level is inhibited in renal tissues of DN group with the increasing expression of CD68 and P62, while expression level of LC3 decreased ( p  < .05). In vitro, HG and 3-MA reduce the numbers of autophagosome of macrophages to inhibit autophagy level with decrease expression of LC3 and Beclin-1, but increase expression of P62, which promote the adhesion and migration capacity of macrophages ( p  < .05). Moreover, CQ and BAFA suppress autophagy level by inhibiting the process of autophagosome-lysosome degradation and fusion of macrophages, as well as the expression of LC3 and Beclin-1. We notice an increase expression of P62 by CQ and BAFA stimulation ( p  < .05). CQ and BAFA further facilitate the adhesion and migration capacity of macrophages. However, RAPA increases the numbers of macrophages autophagosome that inhibited by HG, resulting in a recovery of autophagy level with increase expression of LC3 and Beclin-1, whereas a reduction expression of P62, which lead to inhibition of adhesion and migration of macrophages induced by HG ( p  < .05) Conclusions: High glucose efficiently reduced the level of macrophage autophagy, following macrophages adhesion and migration enhanced when autophagy is suppressed. Activation of autophagosome improve the level of autophagy, but leading to a reduction of the macrophages adhesion and migration. While, inhibiting the process of degradation and fusion of autophagosome-lysosome suppress the level of autophagy and promote the macrophages adhesion and migration. These results indicate that high glucose may play an important role in macrophages adhesion and migration through modulating autophagy activities in diabetic nephropathy.

This study aimed to examine the macrophage phenotype and its relationship to renal function and histological changes in human DN and the effect of TREM-1 on high-glucose-induced macrophage activation. We observed that in renal tissue biopsies, the expression of CD68 and M1 was apparent in the glomeruli and interstitium, while accumulation of M2 and TREM-1 was primarily observed in the interstitium. The numbers of CD68, M1, and M2 macrophages infiltrating in the DN group were increased in a process-dependent manner compared with the control group, and the intensities of the infiltrates were proportional to the rate of subsequent decline in renal function. M1 macrophages were recruited into the kidney at an early stage (I+IIa) of DN. The M1-to-M2 macrophage ratio peaked at this time, whereas M2 macrophages predominated at later time points (III) when the percentage of M1/M2 macrophages was at its lowest level. In an in vitro study, we showed that under high glucose conditions, macrophages began to up-regulate their expression of TREM-1, M1, and marker iNOS and decreased the M2 marker MR. However, the above effects of high-glucose were abolished when TREM-1 expression was inhibited by TREM-1 siRNA. In conclusion, our study demonstrated that there was a positive correlation between the M1/M2 activation state and the progress of DN, and TREM-1 played an important role in high-glucose-induced macrophage phenotype transformation., Competing Interests: The authors have declared that no competing interests exist.

Aim: Imbalance of M1/M2 macrophages phenotype activation is a key point in diabetic nephropathy (DN). This study aimed to investigate whether active vitamin D (VD) suppresses macrophage transition to the M1 phenotype via inhibiting the high glucose-induced STAT-1 phosphorylation to reduce TREM-1 expression., Methods: In vivo, pathological changes in kidney tissue were detected and the expression of CD68 TREM-1, STAT-1, M1 makers, and M2 makers were acquired in renal tissue of patients with DN and 18w DN rats. In vitro, RAW 264.7 cells were incubated in the presence of high glucose with or without VD . Silencing and overexpression of TREM-1 and silencing and activate of STAT-1 were explored to elucidate the underlying mechanism. The expression of TREM-1 and STAT-1 and the changes of macrophage phenotype were examined separately by western blot and immunofluorescence staining., Results: (a) Expression of TREM-1, p-STAT-1, and M1 markers (iNOS and TNF-α) were increased and positively correlated in kidneys from patients with DN. (b) In DN rats, the enlargement of glomerular surface area, expansion of glomerular mesangial matrix, the expression of CD68, TREM-1, p-STAT-1, and M1 marker (iNOS) were significantly increased in comparison with the normal control group, whereas above changes were markedly decreased in the diabetic group treated with the VD group. (c) In vitro, VD significantly decreased high glucose-induced CD68, TREM-1, p-STAT-1, and M1 marker (iNOS) expression. However, above-mentioned effects of VD are abolished when TREM-1 is overexpressed or STAT-1 is activated. Reductions in STAT-1 expression decreased the TREM-1 expression., Conclusion: VD can inhibit macrophage transition to the M1 phenotype through the STAT-1/TREM-1 pathway., (© 2018 Wiley Periodicals, Inc.)

This study intends to investigate the effect of active vitamin D (VD) on the expression of triggering receptor expressed on myeloid cells-1 (TREM-1) in the renal tissues of diabetic nephropathy (DN) rats and to explore the impact of TREM-1 on macrophage adhesion and migration. We find that the expressions of TREM-1 and CD68 protein are higher in DN rats compared with rats in the normal control group and that these changes are decreased in the DN + VD group. In vitro, the capacity for macrophage adhesion and migration and the expression of TREM-1 are increased under high-glucose conditions, but VD inhibits this progress. TREM-1 siRNA decreases high-glucose-induced macrophage adhesion and migration, whereas over-expression of TREM-1 inhibits its action. However, VD cannot suppress high glucose-induced TREM-1 expression and macrophage adhesion and migration when TREM-1 is over-expressed. These results demonstrate that VD can suppress macrophage adhesion and migration by reducing the expression of TREM-1., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Imbalance of M1/M2 macrophages phenotype activation is a key point in diabetic nephropathy (DN). Macrophages mainly exhibit M1 phenotype, which contributes to the inflammation and fibrosis in DN. Studies indicate that autophage plays an important role in M1/M2 activation. However, the effect of mitophage on M1/M2 macrophage phenotype transformation in DN is unknown. This study investigates the role of mitophage on macrophage polarization in DN. In vivo experiments show that macrophages are exhibited to M1 phenotype and display a lower level of mitophagy in the kidney of streptozocin (STZ)-induced diabetic rats. Additionally, inducible nitric oxide synthase (iNOS) expression is positive correlated with the P62 expression, while negative correlated with LC3. Electronic microscope analysis shows mitochondria swelling, crista decrease and lysosome reduction in DN rats compared with NC rats. In vitro, RAW264.7 macrophages switch to M1 phenotype under high glucose conditions. Mitophagy is downregulated in such high glucose induced M1 macrophages. Furthermore, macrophages tend to switch to the M1 phenotype, expressing higher iNOS and TNF-α when impair mitophagy by 3-MA. Rapamycin, an activator of mitophagy, signifcantly blocks high-glucose induced M1 makers (iNOS and TNF-α) expression, meanwhile enhances M2 makers (MR and Arg-1) expression. These results demonstrate that mitophage participates in the regulation of M1/M2 macrophage phenotype in diabetic nephropathy., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Objective To investigate the effect of microRNA-155 on inflammatory response and lipid uptake of macrophages after the cells are stimulated by ox-LDL and its potential mechanism. Methods Macrophage RAW264.7 cells were treated with 0, 25, 50 and 100 μg/mL ox-LDL for 24 hours or with 50 μg/mL ox-LDL for 0, 6, 12, 24 hours. The level of miR-155 was evaluated in all above samples through real-time quantitative PCR. In our research, RAW264.7 cells were divided into six groups: control group, ox-LDL group, ox-LDL/negative control group, ox-LDL/anti-miR-155 group, ox-LDL/shRNA negative control group and ox-LDL/PPARγ-shRNA group. Oil red O staining was used to observe lipid uptake in the cells. Filipin staining was used to evaluate the cellular uptake of ox-LDL. Cholesterol testing was performed to examine the levels of total cholesterol (TC) and free cholesterol (FC). Real-time quantitative PCR was done to detect the expressions of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and IL-6 mRNAs. According to study purpose, we explored the potential mechanisms of miR-155 inhibitor (including control group, ox-LDL group, ox-LDL/negative control group and ox-LDL/miR-155 inhibitor group), miR-155 mimic (including negative control group and miR-155 mimic group), and PPARγ shRNA (including control group, ox-LDL group, ox-LDL/shRNA negative control group and ox-LDL/PPARγ shRNA group) in ox-LDL-treated RAW264.7 cells through evaluating the expressions of p-STAT3, PPARγ, CD36 and NF-κBp65 using Western blotting. Results Ox-LDL stimulation increased the relative expression of miR-155 in a dose- and time-dependent manner. Through oil red O staining, Filipin staining, cholesterol testing and real-time PCR experiment, we found the relative absorbance, levels of TC and FC, filipin fluorescence intensity, and levels of TNF-α, IL-1β and IL-6 mRNAs were significantly lower in ox-LDL/anti-miR-155 group than in ox-LDL and ox-LDL/negative control group. Similarly, the relative absorbance, levels of TC and FC, filipin fluorescence intensity and levels of TNF-α, IL-1β and IL-6 mRNAs were significantly lower in ox-LDL/ PPARγ shRNA group than in ox-LDL group and ox-LDL/shRNA negative control group. The expressions of p-STAT3, PPARγ, CD36 and NF-κBp65 proteins were suppressed in ox-LDL/anti-miR-155 group as compared with ox-LDL group and ox-LDL/negative control group. Similarly, p-STAT3, PPARγ, CD36 and NF-κBp65 protein levels decreased in ox-LDL/PPARγ shRNA as compared with ox-LDL/vector group. Moreover, p-STAT3, PPARγ, CD36 and NF-κBp65 protein levels were higher in miR-155 mimic group than in negative control group. Conclusion Mediated by PPARγ, miR-155 induced inflammation response and lipid uptake of macrophages via STAT3/NF-κB signal pathway and CD36.

Macrophage filopodia, which are dynamic nanotube-like protrusions, have mainly been studied in the context of pathogen clearance. The mechanisms by which they facilitate intercellular communication and mediate tissue inflammation remain poorly understood. Here, we show that macrophage filopodia produce a unique membrane structure called "filopodial tip vesicle" (FTV) that originate from the tip of macrophages filopodia. Filopodia tip-derived particles contain numerous internal-vesicles and function as cargo storage depots via nanotubular transport. Functional studies indicate that the shedding of FTV from filopodia tip allows the delivery of many molecular signalling molecules to fibroblasts. We observed that FTV derived from M1 macrophages and high glucose (HG)-stimulated macrophages (HG/M1-ftv) exhibit an enrichment of the chemokine IL11, which is critical for fibroblast transdifferentiation. HG/M1-ftv induce renal interstitial fibrosis in diabetic mice, while FTV inhibition or targeting FTV IL11- alleviates renal interstitial fibrosis, suggesting that the HG/M1-ftvIL11 pathway may be a novel mechanism underlying renal fibrosis in diabetic nephropathy. Collectively, FTV release could represent a novel function by which filopodia contribute to cell biological processes, and FTV is potentially associated with macrophage filopodia-related fibrotic diseases. 8XMabVtTRQ4kBojsCRSFmY Video Abstract [ABSTRACT FROM AUTHOR]

Macrophages, especially their activation state, are closely related to the progression of diabetic nephropathy. Classically activated macrophages (M1) are proinflammatory effectors, while alternatively activated macrophages (M2) exhibit anti-inflammatory properties. 1,25-Dihydroxyvitamin D3 has renoprotective roles that extend beyond the regulation of mineral metabolism, and PPARγ, a nuclear receptor, is essential for macrophage polarization. The present study investigates the effect of 1,25-dihydroxyvitamin D3 on macrophage activation state and its underlying mechanism in RAW264.7 cells. We find that, under high glucose conditions, RAW264.7 macrophages tend to switch to the M1 phenotype, expressing higher iNOS and proinflammatory cytokines, including TNFα and IL-12. While 1,25-dihydroxyvitamin D3 significantly inhibited M1 activation, it enhanced M2 macrophage activation; namely, it upregulated the expression of MR, Arg-1, and the anti-inflammatory cytokine IL-10 but downregulated the M1 markers. However, the above effects of 1,25-dihydroxyvitamin D3 were abolished when the expression of VDR and PPARγ was inhibited by VDR siRNA and a PPARγ antagonist. In addition, PPARγ was also decreased upon treatment with VDR siRNA. The above results demonstrate that active vitamin D promoted M1 phenotype switching to M2 via the VDR-PPARγ pathway. [ABSTRACT FROM AUTHOR]