scholarly journals Receptor-Interacting Protein Kinase 3 Inhibition Prevents Cadmium-Mediated Macrophage Polarization and Subsequent Atherosclerosis via Maintaining Mitochondrial Homeostasis

2021 ◽  
Vol 8 ◽  
Author(s):  
Jiexin Zhang ◽  
Weijing Feng ◽  
Minghui Li ◽  
Peier Chen ◽  
Xiaodong Ning ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Macrophage Polarization ◽  
Mitochondrial Fission ◽  
Underlying Mechanism ◽  
Interacting Protein ◽  
Mitochondrial Homeostasis ◽  
Cd Exposure ◽  
M1 Type

Chronic cadmium (Cd) exposure contributes to the progression of cardiovascular disease (CVD), especially atherosclerosis (AS), but the underlying mechanism is unclear. Since mitochondrial homeostasis is emerging as a core player in the development of CVD, it might serve as a potential mechanism linking Cd exposure and AS. In this study, we aimed to investigate Cd-mediated AS through macrophage polarization and know the mechanisms of Cd-caused mitochondrial homeostasis imbalance. In vitro, flow cytometry shows that Cd exposure promotes M1-type polarization of macrophages, manifested as the increasing expressions of nuclear Factor kappa-light-chain-enhancer of activated B (NF-kB) and NLR family pyrin domain containing 3 (NLRP3). Mitochondrial homeostasis tests revealed that decreasing mitochondrial membrane potential and mitophage, increasing the mitochondrial superoxide (mROS), and mitochondrial fission are involved in the Cd-induced macrophage polarization. The upregulated expressions of receptor-interacting protein kinase 3 (RIPK3) and pseudokinase-mixed lineage kinase domain-like protein (p-MLKL) were observed. Knocking out RIPK3, followed by decreasing the expression of p-MLKL, improves the mitochondrial homeostasis imbalance which effectively reverses macrophage polarization. In vivo, the oil red O staining showed that Cd with higher blood significantly aggravates AS. Besides, M1-type polarization of macrophages and mitochondrial homeostasis imbalance were observed in the aortic roots of the mice through immunofluorescence and western blot. Knocking out RIPK3 restored the changes above. Finally, the administered N-acetyl cysteine (NAC) or mitochondrial division inhibitor-1 (Mdivi-1), which decreased the mROS or mitochondrial fission, inhibited the expressions of RIPK3 and p-MLKL, attenuating AS and macrophage M1-type polarization in the Cd-treated group. Consequently, the Cd exposure activated the RIPK3 pathway and impaired the mitochondrial homeostasis, resulting in pro-inflammatory macrophage polarization and subsequent AS. Knocking out RIPK3 provided a potential therapeutic target for Cd-caused macrophage polarization and subsequent AS.

scholarly journals Chronic Cadmium Exposure Impairs Macrophage Mitochondrial Homeostasis and Promotes Macrophage Polarization Contributing to Atherosclerosis via Regulating RIPK3 Signaling

2021 ◽  
Author(s):  
Jiexin Zhang ◽  
Weijing Feng ◽  
Peier Chen ◽  
Xiaodong Ning ◽  
Caiwen Ou ◽  
...  
Keyword(s):  
Mitochondrial Dysfunction ◽  
Mitochondrial Function ◽  
Macrophage Polarization ◽  
Mitochondrial Fission ◽  
Inflammatory Factors ◽  
Mitochondrial Homeostasis ◽  
Cd Exposure ◽  
M1 Type

Abstract Background: Chronic cadmium (Cd) exposure can contribute to the progression of cardiovascular disease (CVD), especially atherosclerosis (AS), but the underlying mechanism is unclear. Since mitochondrial homeostasis is emerging as a core player in the development of CVD, it might serve as a potential mechanism linking Cd exposure and AS. Here, we aimed to investigate the Cd-induced AS through macrophage polarization and tried to find out the mechanism of mitochondrial dysfunction caused by Cd exposure. Methods and results: In vitro, flow cytometry showed that Cd exposure markedly promoted M1-type polarization of macrophages, manifesting as the increasing expression of NF-kB, NLRP3 and their downstream inflammatory factors, IL-1β and IL-6. Mitochondrial function test revealed that the decreasing mitochondrial membrane potential and increasing superoxide (mROS) and mitochondrial fission were involved in Cd-induced macrophage polarization. Transmission electron microscope observation and immunofluorescence both identified the decrease of mitophage after Cd exposure. And improving mitochondrial function above significantly restored the balance of macrophage polarization. In vivo, Cd exposure was positively correlated with blood Cd concentration, and oil red O staining showed higher blood Cd significantly increased the area of AS plaques. Besides, M1-type polarization of macrophages and mitochondrial dysfunction were observed in mouse aortic roots through immunofluorescence and western blot as the dosage of Cd increasing. And the administered NAC or Mdivi-1, which decreased mROS or mitochondrial fission, markedly attenuated AS plaques and macrophage M1-type polarization in Cd-treated group. Finally, the up-regulated expressions of RIPK3 and p-MLKL were observed both in vitro and in vivo. And knocking out RIPK3 with decreasing expression of p-MLKL followed did improve mitochondrial dysfunction caused by Cd which effectively reversed macrophage polarization. Conclusion: Cd exposure activated RIPK3 pathway and impaired mitochondrial homeostasis, resulting in macrophage polarization to a pro-inflammatory phenotype and subsequent AS. These findings suggest that improving mitochondrial homeostasis may provide a potential therapeutic target for AS induced by chronic Cd exposure.

scholarly journals Drosophila Homeodomain-Interacting Protein Kinase (Hipk) Phosphorylates the Homeodomain Proteins Homeobrain, Empty Spiracles, and Muscle Segment Homeobox

2019 ◽  
Vol 20 (8) ◽  
pp. 1931
Author(s):  
Eva Louise Steinmetz ◽  
Denise Nicole Dewald ◽  
Nadine Luxem ◽  
Uwe Walldorf
Keyword(s):  
Nervous System ◽  
Protein Kinase ◽  
Mutational Analysis ◽  
Bimolecular Fluorescence Complementation ◽  
Physical Interaction ◽  
Homeodomain Proteins ◽  
Interacting Protein ◽  
Enzyme Substrate

The Drosophila homeodomain-interacting protein kinase (Hipk) is the fly representative of the well-conserved group of HIPKs in vertebrates. It was initially found through its characteristic interactions with homeodomain proteins. Hipk is involved in a variety of important developmental processes, such as the development of the eye or the nervous system. In the present study, we set Hipk and the Drosophila homeodomain proteins Homeobrain (Hbn), Empty spiracles (Ems), and Muscle segment homeobox (Msh) in an enzyme-substrate relationship. These homeoproteins are transcription factors that function during Drosophila neurogenesis and are, at least in part, conserved in vertebrates. We reveal a physical interaction between Hipk and the three homeodomain proteins in vivo using bimolecular fluorescence complementation (BiFC). In the course of in vitro phosphorylation analysis and subsequent mutational analysis we mapped several Hipk phosphorylation sites of Hbn, Ems, and Msh. The phosphorylation of Hbn, Ems, and Msh may provide further insight into the function of Hipk during development of the Drosophila nervous system.

scholarly journals The phosphorylation of CapZ-interacting protein (CapZIP) by stress-activated protein kinases triggers its dissociation from CapZ

2005 ◽  
Vol 389 (1) ◽  
pp. 127-135 ◽  
Author(s):  
Claire E. EYERS ◽  
Helen McNEILL ◽  
Axel KNEBEL ◽  
Nick MORRICE ◽  
Simon J. C. ARTHUR ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Protein Kinases ◽  
Osmotic Shock ◽  
Jurkat Cells ◽  
Mitogen Activated Protein Kinase ◽  
Interacting Protein ◽  
Capping Protein ◽  
Sb 203580

A protein expressed in immune cells and muscle was detected in muscle extracts as a substrate for several SAPKs (stress-activated protein kinases). It interacted specifically with the F-actin capping protein CapZ in splenocytes, and was therefore termed ‘CapZIP’ (CapZ-interacting protein). Human CapZIP was phosphorylated at Ser-179 and Ser-244 by MAPKAP-K2 (mitogen-activated protein kinase-activated protein kinase 2) or MAPKAP-K3 in vitro. Anisomycin induced the phosphorylation of CapZIP at Ser-179 in Jurkat cells, which was prevented by SB 203580, consistent with phosphorylation by MAPKAP-K2 and/or MAPKAP-K3. However, osmotic shock-induced phosphorylation of Ser-179 was unaffected by SB 203580. These and other results suggest that CapZIP is phosphorylated at Ser-179 in cells by MAPKAP-K2/MAPKAP-K3, and at least one other protein kinase. Stress-activated MAP kinase family members phosphorylated human CapZIP at many sites, including Ser-68, Ser-83, Ser-108 and Ser-216. Ser-108 became phosphorylated when Jurkat cells were exposed to osmotic shock, which was unaffected by SB 203580 and/or PD 184352, or in splenocytes from mice that do not express either SAPK3/p38γ or SAPK4/p38δ. Our results suggest that CapZIP may be phosphorylated by JNK (c-Jun N-terminal kinase), which phosphorylates CapZIP to >5 mol/mol within minutes in vitro. Osmotic shock or anisomycin triggered the dissociation of CapZIP from CapZ in Jurkat cells, suggesting that phosphorylation of CapZIP may regulate the ability of CapZ to remodel actin filament assembly in vivo.

scholarly journals Vitamin D Attenuates Ischemia/Reperfusion-Induced Cardiac Injury by Reducing Mitochondrial Fission and Mitophagy

2020 ◽  
Vol 11 ◽  
Author(s):  
Tzu-Lin Lee ◽  
Ming-Hsueh Lee ◽  
Yu-Chen Chen ◽  
Yi-Chieh Lee ◽  
Tsai-Chun Lai ◽  
...  
Keyword(s):  
Vitamin D ◽  
Cell Viability ◽  
Ischemia Reperfusion ◽  
Mitochondrial Fission ◽  
Cardiac Injury ◽  
Physiological Role ◽  
H9c2 Cell ◽  
Interacting Protein ◽  
Mitochondrial Homeostasis

Myocardial infarction is the leading cause of morbidity and mortality worldwide. Although myocardial reperfusion after ischemia (I/R) is an effective method to save ischemic myocardium, it can cause adverse reactions, including increased oxidative stress and cardiomyocyte apoptosis. Mitochondrial fission and mitophagy are essential factors for mitochondrial quality control, but whether they play key roles in cardiac I/R injury remains unknown. New pharmacological or molecular interventions to alleviate reperfusion injury are currently considered desirable therapies. Vitamin D3 (Vit D3) regulates cardiovascular function, but its physiological role in I/R-exposed hearts, especially its effects on mitochondrial homeostasis, remains unclear. An in vitro hypoxia/reoxygenation (H/R) model was established in H9c2 cells to simulate myocardial I/R injury. H/R treatment significantly reduced H9c2 cell viability, increased apoptosis, and activated caspase 3. In addition, H/R treatment increased mitochondrial fission, as manifested by increased expression of phosphorylated dynein-related protein 1 (p-Drp1) and mitochondrial fission factor (Mff) as well as increased mitochondrial translocation of Drp1. Treatment with the mitochondrial reactive oxygen species scavenger MitoTEMPO increased cell viability and decreased mitochondrial fission. H/R conditions elicited excessive mitophagy, as indicated by increased expression of BCL2-interacting protein 3 (BNIP3) and light chain (LC3BII/I) and increased formation of autolysosomes. In contrast, Vit D3 reversed these effects. In a mouse model of I/R, apoptosis, mitochondrial fission, and mitophagy were induced. Vit D3 treatment mitigated apoptosis, mitochondrial fission, mitophagy, and myocardial ultrastructural abnormalities. The results indicate that Vit D3 exerts cardioprotective effects against I/R cardiac injury by protecting mitochondrial structural and functional integrity and reducing mitophagy.

scholarly journals The Adapter Protein ZIP Binds Grb14 and Regulates Its Inhibitory Action on Insulin Signaling by Recruiting Protein Kinase Cζ

2002 ◽  
Vol 22 (20) ◽  
pp. 6959-6970 ◽  
Author(s):  
Bertrand Cariou ◽  
Dominique Perdereau ◽  
Katia Cailliau ◽  
Edith Browaeys-Poly ◽  
Véronique Béréziat ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Insulin Receptor ◽  
Insulin Signaling ◽  
Inhibitory Action ◽  
Catalytic Domain ◽  
Inhibitory Effect ◽  
Negative Regulator ◽  
Interacting Protein

ABSTRACT Grb14 is a member of the Grb7 family of adapters and acts as a negative regulator of insulin-mediated signaling. Here we found that the protein kinase Cζ (PKCζ) interacting protein, ZIP, interacted with Grb14. Coimmunoprecipitation experiments demonstrated that ZIP bound to both Grb14 and PKCζ, thereby acting as a link in the assembly of a PKCζ-ZIP-Grb14 heterotrimeric complex. Mapping studies indicated that ZIP interacted through its ZZ zinc finger domain with the phosphorylated insulin receptor interacting region (PIR) of Grb14. PKCζ phosphorylated Grb14 under in vitro conditions and in CHO-IR cells as demonstrated by in vivo labeling experiments. Furthermore, Grb14 phosphorylation was increased under insulin stimulation, suggesting that the PKCζ-ZIP-Grb14 complex is involved in insulin signaling. The PIR of Grb14, which also interacts with the catalytic domain of the insulin receptor (IR) and inhibits its activity, was preferentially phosphorylated by PKCζ. Interestingly, the phosphorylation of Grb14 by PKCζ increased its inhibitory effect on IR tyrosine kinase activity in vitro. The role of ZIP and Grb14 in insulin signaling was further investigated in vivo in Xenopus laevis oocytes. In this model, ZIP potentiated the inhibitory action of Grb14 on insulin-induced oocyte maturation. Importantly, this effect required the recruitment of PKCζ and the phosphorylation of Grb14, providing in vivo evidences for a regulation of Grb14-inhibitory action by ZIP and PKCζ. Together, these results suggest that Grb14, ZIP, and PKCζ participate in a new feedback pathway of insulin signaling.

Targeted Disruption of the Murine Homeodomain-Interacting Protein Kinase-2 Causes Growth Deficiency In Vivo and Cell Cycle Arrest In Vitro

2009 ◽  
Vol 28 (4) ◽  
pp. 161-167 ◽  
Author(s):  
Francesco Trapasso ◽  
Rami I. Aqeilan ◽  
Rodolfo Iuliano ◽  
Rosa Visone ◽  
Eugenio Gaudio ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Kinase ◽  
Cell Cycle Arrest ◽  
Targeted Disruption ◽  
Interacting Protein ◽  
Growth Deficiency ◽  
Cycle Arrest

scholarly journals Yi-Qi-Jian-Pi Formula Suppresses RIPK1/RIPK3-Complex-Dependent Necroptosis of Hepatocytes Through ROS Signaling and Attenuates Liver Injury in Vivo and in Vitro

2021 ◽  
Vol 12 ◽  
Author(s):  
Feixia Wang ◽  
Li Tang ◽  
Baoyu Liang ◽  
Chun Jin ◽  
Liyuan Gao ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Liver Injury ◽  
Protective Effect ◽  
Molecular Mechanisms ◽  
Positive Role ◽  
Interacting Protein ◽  
Ros Signaling ◽  
Tnf Α

Acute-on-chronic liver failure (ACLF) is described as a characteristic of acute jaundice and coagulation dysfunction. Effective treatments for ACLF are unavailable and hence are urgently required. We aimed to define the effect of Yi-Qi-Jian-Pi Formula (YQJPF) on liver injury and further examine the molecular mechanisms. In this study, we established CCl4-, LPS-, and d-galactosamine (D-Gal)-induced ACLF rat models in vivo and LPS- and D-Gal-induced hepatocyte injury models in vitro. We found that YQJPF significantly ameliorates liver injury in vivo and in vitro that is associated with the regulation of hepatocyte necroptosis. Specifically, YQJPF decreased expression of receptor-interacting protein kinase 1 (RIPK1), receptor-interacting protein kinase 3 (RIPK3) and pseudokinase mixed lineage kinase domain-like (MLKL) to inhibit the migration of RIPK1 and RIPK3 into necrosome. YQJPF also reduces the expression of inflammatory cytokines IL-6, IL-8, IL-1β, and TNF-α, which were regulated by RIPK3 mediates cell death. RIPK1 depletion was found to enhance the protective effect of YQJPF. Furthermore, we showed that YQJPF significantly downregulates the mitochondrial reactive oxygen species (ROS) production and mitochondrial depolarization, with ROS scavenger, 4-hydroxy-TEMPO treatment recovering impaired RIPK1-mediated necroptosis and reducing the expression of IL-6, IL-8, IL-1β, and TNF-α. In summary, our study revealed the molecular mechanism of protective effect of YQJPF on hepatocyte necroptosis, targeting RIPK1/RIPK3-complex-dependent necroptosis via ROS signaling. Overall, our results provided a novel perspective to indicate the positive role of YQJPF in ACLF.

scholarly journals MSC-derived miR-181b Overexpressing Exosomes Enhanced Osteointegration by Enhancing M2 Polarization of Macrophages via Targeting the PRKCD/AKT Pathway

2020 ◽  
Author(s):  
Wei Liu ◽  
Muyu Yu ◽  
Feng Chen ◽  
Dong Xie ◽  
Longqing Wang ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Macrophage Polarization ◽  
Underlying Mechanism ◽  
Fluorescent Labeling ◽  
M2 Macrophage ◽  
Implant Loosening ◽  
M2 Polarization ◽  
Tnf Α

Abstract Background: Many patients suffer from implant loosening after the implantation of titanium alloy caused by immune response to the foreign bodies and this could inhibit osteogenesis, which could possibly give rise to poor osteointegration and there is currently no appropriate solution in clinical practice. Exosomes overexpressing miRNA has been proven to be a suitable candidate for solving this problem. In this study, we explored whether miR-181b could exert beneficial effect on promoting M2 macrophage polarization, thus inhibiting inflammation as well as promoting osteogenesis and elaborated the underlying mechanism in vitro. Furthermore, we aimed to find whether exosomes overexpressing miR-181b (Exo-181b) could enhance osteointegration in vivo.Methods: In vitro and in vivo studies were carried out for assessing the anti-inflammatory and pro-osteogenesis effect of miR-181b. In vitro, ELISA was applied for the detection of the inflammation factors levels including IL-6, TNF-α, as well as IL-10 and the percentage of M1 or M2 polarization was determined by flow cytometry. Also, qRT-PCR was used for the detection of the relative gene expression of the CCR7, CD206, Arg-1, iNOS, VEGF and BMP-2 genes. Western blotting was applied for detecting the protein expression of PRKCD, AKT and p-AKT. In vivo, we established air pouch model for evaluating the effect of Exo-181b on macrophage polarization and distal femoral bone defect model was established for determining the osteointegration effect of Exo-181b by MicroCT, sequential fluorescent labeling and histological analysis. Results: In vitro, we firstly verified that miR-181b significantly enhanced M2 polarization and inhibited inflammation by suppressing PRKCD and activating p-AKT. Then, in vivo, we verified that Exo-181b enhanced M2 polarization, reduced the inflammatory response and enhanced osteointegration. Conclusions: MiR-181b could suppress inflammatory response by regulating the PRKCD/AKT signaling pathway and promoting M2 polarization, which further promoting osteogenesis of hBMSC in vitro and Exo-181b could promote osteointegration in vivo.

scholarly journals A novel delivery nanobiotechnology: engineered miR-181b exosomes improved osteointegration by regulating macrophage polarization

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Wei Liu ◽  
Muyu Yu ◽  
Feng Chen ◽  
Longqing Wang ◽  
Cheng Ye ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Macrophage Polarization ◽  
Underlying Mechanism ◽  
M2 Macrophage ◽  
Implant Loosening ◽  
M2 Polarization ◽  
M2 Macrophage Polarization ◽  
Osteogenesis In Vitro

Abstract Background Many patients suffer from implant loosening after the implantation of titanium alloy caused by immune response to the foreign bodies and this could inhibit the following osteogenesis, which could possibly give rise to aseptic loosening and poor osteointegration while there is currently no appropriate solution in clinical practice. Exosome (Exo) carrying miRNA has been proven to be a suitable nanocarrier for solving this problem. In this study, we explored whether exosomes overexpressing miR-181b (Exo-181b) could exert beneficial effect on promoting M2 macrophage polarization, thus inhibiting inflammation as well as promoting osteogenesis and elaborated the underlying mechanism in vitro. Furthermore, we aimed to find whether Exo-181b could enhance osteointegration. Results In vitro, we firstly verified that Exo-181b significantly enhanced M2 polarization and inhibited inflammation by suppressing PRKCD and activating p-AKT. Then, in vivo, we verified that Exo-181b enhanced M2 polarization, reduced the inflammatory response and enhanced osteointegration. Also, we verified that the enhanced M2 polarization could indirectly promote the migration and osteogenic differentiation by secreting VEGF and BMP-2 in vitro. Conclusions Exo-181b could suppress inflammatory response by promoting M2 polarization via activating PRKCD/AKT signaling pathway, which further promoting osteogenesis in vitro and promote osteointegration in vivo. Graphic abstract

Cobalt-doped Ti surface promotes immunomodulation

2021 ◽  
Author(s):  
Xiaoming Yang ◽  
Chi Zhang ◽  
Tao Zhang ◽  
Jin Xiao
Keyword(s):  
Macrophage Polarization ◽  
Cobalt Content ◽  
Inflammatory Reactions ◽  
Cobalt Ions ◽  
Novel Strategy ◽  
Plasma Electrolytic ◽  
Peo Coatings ◽  
M1 Type

Abstract Here, cobalt-doped plasma electrolytic oxidation (PEO) coatings with different cobalt contents were prepared on Ti implants. The cobalt ions in the PEO coating exhibited a slow and sustainable release and thus showed excellent biocompatibility and enhanced cell adhesion. In vitro ELISA and RT-PCR assays demonstrated that the cobalt-loaded Ti showed immunomodulatory functions to macrophages and upregulated the expression of anti-inflammatory (M1 type) genes and downregulated expression levels of pro-inflammatory (M2 type) genes compared with that of pure Ti sample. High cobalt content induced increased macrophage polarization into the M2 type. Furthermore, the findings from the in vivo air pouch model suggested that cobalt-loaded Ti could mitigate inflammatory reactions. The present work provides a novel strategy to exploit the immunomodulatory functions of implant materials.

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