Aldose reductase deficiency inhibits LPS-induced M1 response in macrophages by activating autophagy.
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ABSTRACT:
SUBMITTER: Cheng P
PROVIDER: S-EPMC8004403 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Pathological cardiac hypertrophy is associated with the accumulation of lipid peroxidation-derived aldehydes such as 4-hydroxy-trans-2-nonenal (HNE) and acrolein in the heart. These aldehydes are metabolized via several pathways, of which aldose reductase (AR) represents a broad-specificity route for their elimination. We tested the hypothesis that by preventing aldehyde removal, AR deficiency accentuates the pathological effects of transverse aortic constriction (TAC). We found that the levels of AR in the heart were increased in mice subjected to TAC for 2?weeks. In comparison with wild-type (WT), AR-null mice showed lower ejection fraction, which was exacerbated 2?weeks after TAC. Levels of atrial natriuretic peptide and myosin heavy chain were higher in AR-null than in WT TAC hearts. Deficiency of AR decreased urinary levels of the acrolein metabolite, 3-hydroxypropylmercapturic acid. Deletion of AR did not affect the levels of the other aldehyde-metabolizing enzyme - aldehyde dehydrogenase 2 in the heart, or its urinary product - (N-Acetyl-S-(2-carboxyethyl)-l-cystiene). AR-null hearts subjected to TAC showed increased accumulation of HNE- and acrolein-modified proteins, as well as increased AMPK phosphorylation and autophagy. Superfusion with HNE led to a greater increase in p62, LC3II formation, and GFP-LC3-II punctae formation in AR-null than WT cardiac myocytes. Pharmacological inactivation of JNK decreased HNE-induced autophagy in AR-null cardiac myocytes. Collectively, these results suggest that during hypertrophy the accumulation of lipid peroxidation derived aldehydes promotes pathological remodeling via excessive autophagy, and that metabolic detoxification of these aldehydes by AR may be essential for maintaining cardiac function during early stages of pressure overload.
| S-EPMC6205513 | biostudies-literature
Project description:Activated macrophages serve a key role in various inflammatory diseases, such as atherosclerosis and septic shock. Tripartite motif-containing protein 65 (TRIM65) has been previously reported to participate in tumor progression and lung inflammation. However, the molecular mechanisms that controls its expression under inflammatory conditions and its consequences in activated macrophages remain poorly understood. The present study first collected the tissues of C57BL/6J mice, smooth muscle cells, macrophages and endothelial cells to determine the expression and distribution of TRIM65 by reverse transcription-quantitative (RT-q) PCR and western blotting. Mouse and human macrophages were treated with LPS and C57BL/6J mice were intraperitoneally injected with LPS followed by isolation of spleen, lung, aorta and bone marrow. Following treatment, TRIM65 mRNA and protein level was examined by RT-qPCR and western blotting. The results showed that TRIM65 was highly expressed in organs of the immune system, such as the spleen, lymph node and thymus, but lowly expressed in heart, liver, brain and kidneys. TRIM65 was also highly expressed in macrophages and endothelial cells. TRIM65 mRNA and protein expression levels were found to be decreased in LPS-treated macrophages in vitro and in tissues isolated from C57BL/6J mice intraperitoneally injected with LPS in vivo. In addition, to identify the signaling pathways by which LPS regulates TRIM65 expression, inhibitors of MAPK and Akt signaling pathways were used to treat macrophages followed by examination the expression of TRIM65 by western blotting. The results demonstrated that LPS-inhibited TRIM65 expression was blocked by treatment with the ERK1/2 inhibitor U0126. Moreover, the RT-qPCR results showed that TRIM65 knockout potentiated LPS-induced expression of inflammatory cytokines in macrophages. Taken together, data from the present study suggest that LPS decreased TRIM65 expression in macrophages and C57BL/6J mouse by activating the ERK1/2 signaling pathway, whilst TRIM65 knockout promoted macrophage activation. This information may facilitate the development of potential therapeutic strategies for the prevention and treatment of inflammatory diseases, such as atherosclerosis.
| S-EPMC10068263 | biostudies-literature
Project description:Microglia cells are the main mediators of neuroinflammation. Activation of microglia often aggravates the pathological process of various neurological diseases. Natural chemicals have unique advantages in inhibiting microglia-mediated neuroinflammation and improving neuronal function. Here, we examined the effects of asperosaponin VI (ASA VI) on LPS-activated primary microglia. Microglia were isolated from mice and pretreated with different doses of ASA VI, following lipopolysaccharide (LPS) administration. Activation and inflammatory response of microglia cells were evaluated by real-time fluorescence quantitative polymerase chain reaction (q-PCR), immunohistochemistry and enzyme-linked immunosorbent assay (ELISA). Signaling pathways were detected by western blotting. We found that the ASA VI inhibited the morphological expansion of microglia cells, decreased the expression and release of proinflammatory cytokines, and promoted the expression of antiinflammatory cytokines in a dose-dependent manner. ASA VI also activated PPAR-γ signaling pathway in LPS-treated microglia. The anti-inflammatory effects of ASA VI in microglia were blocked by treating PPAR-γ antagonist (GW9662). These results showed that ASA VI promote the transition of microglia cells from proinflammatory to anti-inflammatory by regulating PPAR-γ pathway.
| S-EPMC7569127 | biostudies-literature
Project description:Macrophages are found in tissues, body cavities, and mucosal surfaces. Most tissue macrophages are seeded in the early embryo before definitive hematopoiesis is established. Others are derived from blood monocytes. The macrophage lineage diversification and plasticity are key aspects of their functionality. Macrophages can also be generated from monocytes in vitro and undergo classical (LPS+IFN-γ) or alternative (IL-4) activation. In vivo, macrophages with different polarization and different activation markers coexist in tissues. Certain mouse strains preferentially promote T-helper-1 (Th1) responses and others Th2 responses. Their macrophages preferentially induce iNOS or arginase and have been called M1 and M2, respectively. In many publications, M1 and classically activated and M2 and alternatively activated are used interchangeably. We tested whether this is justified by comparing the gene lists positively [M1(=LPS+)] or negatively [M2(=LPS-)] correlated with the ratio of IL-12 and arginase 1 in transcriptomes of LPS-treated peritoneal macrophages with in vitro classically (LPS, IFN-γ) vs. alternatively activated (IL-4) bone marrow derived macrophages, both from published datasets. Although there is some overlap between in vivo M1(=LPS+) and in vitro classically activated (LPS+IFN-γ) and in vivo M2(=LPS-) and in vitro alternatively activated macrophages, many more genes are regulated in opposite or unrelated ways. Thus, M1(=LPS+) macrophages are not equivalent to classically activated, and M2(=LPS-) macrophages are not equivalent to alternatively activated macrophages. This fundamental discrepancy explains why most surface markers identified on in vitro generated macrophages do not translate to the in vivo situation. Valid in vivo M1/M2 surface markers remain to be discovered.
| S-EPMC6543837 | biostudies-literature
Project description:Objective: Exosomes (Exos) are membrane-encased vesicles derived by nearly all cell types for intercellular communication and regulation. They also received attention for their use as natural therapeutic platforms and drug delivery system. Classically activated M1 macrophages suppress tumor growth by releasing pro-inflammatory factors. This study investigated the suitability of M1-exosomes (M1-Exos) as drug carrier and their effect on the NF-κB signal pathway and further detected whether macrophages repolarization can potentiate the antitumor activities of chemotherapeutics. Methods: M1-Exos were isolated from M1-macrophages by ultracentrifugation and characterized by transmission electron, nanoparticle tracking analysis, dynamic light scattering and western blot. Then M1-Exos were used as Paclitaxel (PTX) carriers to prepare a nano-formulation (PTX- M1-Exos). A relatively simple slight sonication method was used to prepare the drug delivery system (PTX-M1-Exos). The cytotoxicity of PTX-M1-Exos on cancer cells was detected by MTT and flow cytometry in vitro. 4T1 tumor bearing mice were used to perform the therapeutic effect of PTX-M1-Exos in vivo. Results: The expression of caspase-3 in breast cancer cells was increased when co-incubated with macrophages in the presence of M1-Exos in vitro. The production of pro-inflammatory cytokines was increased after exposure of macrophages in M1-Exos. M1-Exos provided a pro-inflammatory environment which enhanced the anti-tumor activity via caspase-3 mediated pathway. The treatment of M1-Exos to the tumor bearing mice exhibit anti-tumor effects in vivo. Meanwhile, the treatment of PTX-M1-Exos demonstrated higher anti-tumor effects than the M1-Exos or PTX group. Conclusion: The results in our study indicate that the M1-Exos act as the carrier to deliver PTX into the tumor tissues, and also enhance the anti-tumor effects of chemotherapeutics in tumor bearing mice.
| S-EPMC6485189 | biostudies-literature
Project description:Diabetic nephropathy (DN) is a chronic diabetic microvascular complication. Hyperactivity of the polyol pathway is involved in the pathogenesis of DN. Aldose reductase (AR), the rate-limiting enzyme of the polyol pathway, is expected to be an effective target in the treatment of DN. WJ-39 is a novel inhibitor of AR. The present study aimed at exploring the effects of WJ-39 in DN. DN was induced in rats by injecting 30?mg/kg streptozotocin (STZ). After 14 weeks, WJ-39 (10, 20, and 40?mg/kg) was intragastrically administered to the rats for 12 weeks. Treatment with WJ-39 significantly inhibited AR activation and ameliorated renal dysfunction and fibrosis in DN rats. WJ-39 reduced oxidative stress in the kidneys of DN rats by activating the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. WJ-39 suppressed the activation of the nuclear factor-kappa B (NF-?B) pathway and the nucleotide-binding and oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome to reduce the secretion of inflammatory factors. Rat mesangial cells (RMCs) were cultured under hyperglycemic conditions. WJ-39 abrogated the high glucose- (HG-) induced, excessive production of reactive oxygen species (ROS) and inflammatory factors. However, transfection with Nrf2 small interfering RNA abolished the effects of WJ-39. WJ-39 also blocked the transforming growth factor-?1/Smad pathway to reduce the production of glomerular extracellular matrix proteins, ultimately reducing fibrogenesis in DN. Our results show that WJ-39 ameliorated renal injury in DN rats, and its effects on oxidative stress and inflammation were associated with the activation of Nrf2 signaling. Thus, WJ-39 and its mechanism of amelioration of renal lesions in DN rats by reducing renal inflammation, oxidative stress, and fibrosis injury could be an effective strategy for the treatment of DN.
| S-EPMC7277034 | biostudies-literature
Project description:Aims/hypothesisThe aim of the study was to investigate the effects of genetic deficiency of aldose reductase in mice on the development of key endpoints of diabetic nephropathy.MethodsA line of Ar (also known as Akr1b3)-knockout (KO) mice, a line of Ar-bitransgenic mice and control C57BL/6 mice were used in the study. The KO and bitransgenic mice were deficient for Ar in the renal glomeruli and all other tissues, with the exception of, in the bitransgenic mice, a human AR cDNA knockin-transgene that directed collecting-tubule epithelial-cell-specific AR expression. Diabetes was induced in 8-week-old male mice with streptozotocin. Mice were further maintained for 17 weeks then killed. A number of serum and urinary variables were determined for these 25-week-old mice. Periodic acid-Schiff staining, western blots, immunohistochemistry and protein kinase C (PKC) activity assays were performed for histological analyses, and to determine the levels of collagen IV and TGF-β1 and PKC activities in renal cortical tissues.ResultsDiabetes-induced extracellular matrix accumulation and collagen IV overproduction were completely prevented in diabetic Ar-KO and bitransgenic mice. Ar deficiency also completely or partially prevented diabetes-induced activation of renal cortical PKC, TGF-β1 and glomerular hypertrophy. Loss of Ar results in a 43% reduction in urine albumin excretion in the diabetic Ar-KO mice and a 48% reduction in the diabetic bitransgenic mice (p < 0.01).Conclusions/interpretationGenetic deficiency of Ar significantly ameliorated development of key endpoints linked with early diabetic nephropathy in vivo. Robust and specific inhibition of aldose reductase might be an effective strategy for the prevention and treatment of diabetic nephropathy.
| S-EPMC3071933 | biostudies-literature
Project description:The bacterial endotoxin lipopolysaccharide (LPS) is known to induce release of arachidonic acid (AA) and its metabolic products, which play important roles in the inflammatory process. We have shown earlier that LPS-induced signals in macrophages are mediated by aldose reductase (AR). Here we have investigated the role of AR in LPS-induced release of AA metabolites and their modulation using a potent pharmacological inhibitor, fidarestat, and AR siRNA ablation in RAW264.7 macrophages and AR-knockout mouse peritoneal macrophages and heart tissue. Inhibition or genetic ablation of AR prevented the LPS-induced synthesis and release of AA metabolites such as PGE2, TXB, PGI2, and LTBs in macrophages. LPS-induced activation of cPLA2 was also prevented by AR inhibition. Similarly, AR inhibition also prevented the calcium ionophore A23187-induced cPLA2 and LTB4 in macrophages. Further, AR inhibition by fidarestat prevented the expression of AA-metabolizing enzymes such as COX-2 and LOX-5 in RAW264.7 cells and AR-knockout mouse-derived peritoneal macrophages. LPS-induced expression of AA-metabolizing enzymes and their catalyzed metabolic products was significantly lower in peritoneal macrophages and heart tissue from AR-knockout mice. LPS-induced activation of redox-sensitive signaling intermediates such as MAPKs, transcription factor NF-κB, and EGR-1, a transcriptional regulator of mPGES-1, which in collaboration with COX-2 leads to the production of PGE2, was also significantly prevented by AR inhibition. Taken together, our results indicate that AR mediates LPS-induced inflammation by regulating the AA-metabolic pathway and thus provide a novel role for AR inhibition in preventing inflammatory complications such as sepsis.
| S-EPMC3188329 | biostudies-literature
Project description:Diverse organisms accumulate organic osmolytes to adapt to hyperosmotic stress. The molecular basis of eukaryotic gene osmoregulation remains obscure. Aldose reductase [AR; alditol:NAD(P)+ 1-oxidoreductase, EC 1.1.1.21], which catalyzes the conversion of glucose to sorbitol (an organic osmolyte), is induced in renal medullary cells under hyperosmotic conditions. Elevated extracellular NaCl increases AR mRNA transcription in PAP-HT25 cells, a cell line derived from the rabbit renal papilla. We have cloned and characterized the rabbit AR gene to determine how it is regulated by hyperosmolality. The length of the gene, not including 5' or 3' flanking regions, is approximately 14.7 kilobases (kb) organized into 10 exons and 9 introns. The transcription start site is 36 base pairs upstream of the initiator methionine codon. A 5-kb fragment containing approximately 3.5 kb of 5' flanking region was isolated. The 3.5-kb sequence was examined for basal promoter activity and hyperosmotic response in luciferase reporter gene constructs. A 235-base-pair fragment (base pairs -208 to +27) was able to drive the downstream reporter gene in transfected PAP-HT25 cells under isoosmotic conditions (300 mosmol/kg of H2O). When this fragment plus the remaining upstream sequence (from approximately base pair -3429 to base pair +27) was used, cells in hyperosmotic medium (500 mosmol/kg of H2O) showed about 40-fold induction of luciferase expression compared with cells in isoosmotic medium. The upstream fragment (from approximately base pair -3429 to base pair -192) also conferred osmotic response to a heterologous promoter (B19). This finding evidences putative osmotic response element(s) (OREs) within a specific DNA fragment in a eukaryotic genome. Identification and characterization of OREs within this fragment and their associated trans-acting factors should reveal the molecular mechanisms of gene regulation in osmotic stress.
| S-EPMC45098 | biostudies-other
Project description:Murine macrophages activated by the Toll-like receptor 4 agonist lipopolysaccharide (LPS) polarize to the M1 type by inducing proinflammatory marker proteins and changing their energy metabolism to increased aerobic glycolysis and reduced respiration. We here show that the aliphatic isothiocyanate sulforaphane (Sfn) diminishes M1 marker expression (IL-1β, IL-6, TNF-α, iNOS, NO, and ROS) and leads to highly energetic cells characterized by both high glycolytic and high respiratory activity as assessed by extracellular flux analysis. Focusing on a potential connection between high glycolytic activity and low IL-1β expression in M1 (LPS/Sfn) macrophages, we reveal that Sfn impedes the moonlighting function of pyruvate kinase M2 (PKM2) in M1 macrophages. Sfn limits mono/dimerization and nuclear residence of PKM2 accompanied by reduced HIF-1α levels, Stat3 phosphorylation at tyrosine 705, and IL-1β expression while preserving high levels of cytosolic PKM2 tetramer with high glycolytic enzyme activity. Sfn prevents glutathionylation of PKM2 in LPS-stimulated macrophages which may account for the reduced loss of PKM2 tetramer. Overall, we uncover PKM2 as a novel affected hub within the anti-inflammatory activity profile of Sfn.
| S-EPMC9380596 | biostudies-literature