scholarly journals Stem Cell-Derived Exosomes Ameliorate Doxorubicin-Induced Muscle Toxicity through Counteracting Pyroptosis

2020 ◽  
Vol 13 (12) ◽  
pp. 450
Author(s):  
Fatima Bianca A. Dessouki ◽  
Rakesh C. Kukreja ◽  
Dinender K. Singla
Keyword(s):  
Muscle Function ◽  
Muscular Atrophy ◽  
Embryonic Stem ◽  
Mouse Embryonic Fibroblast ◽  
Negative Control ◽  
M1 Macrophages ◽  
Tnf Α ◽  
Embryonic Fibroblast ◽  
Muscle Toxicity

Doxorubicin (Dox)-induced muscle toxicity (DIMT) is a common occurrence in cancer patients; however, the cause of its development and progression is not established. We tested whether inflammation-triggered cell death, “pyroptosis” plays a role in DIMT. We also examined the potential role of exosomes derived from embryonic stem cells (ES-Exos) in attenuating DIMT. C57BL/6J mice (10 ± 2 wks age) underwent the following treatments: Control (saline), Dox, Dox+ES-Exos, and Dox+MEF-Exos (mouse-embryonic fibroblast-derived exosomes, negative control). Our results demonstrated that Dox significantly reduced muscle function in mice, which was associated with a significant increase in NLRP3 inflammasome and initiation marker TLR4 as compared with controls. Pyroptosis activator, ASC, was significantly increased compared to controls with an upregulation of specific markers (caspase-1, IL-1β, and IL-18). Treatment with ES-Exos but not MEF-Exos showed a significant reduction in inflammasome and pyroptosis along with improved muscle function. Additionally, we detected a significant increase in pro-inflammatory cytokines (TNF-α and IL-6) and inflammatory M1 macrophages in Dox-treated animals. Treatment with ES-Exos decreased M1 macrophages and upregulated anti-inflammatory M2 macrophages. Furthermore, ES-Exos showed a significant reduction in muscular atrophy and fibrosis. In conclusion, these results suggest that DIMT is mediated through inflammation and pyroptosis, which is attenuated following treatment with ES-Exos.

scholarly journals Exosomes derived from embryonic stem cells inhibit doxorubicin and inflammation-induced pyroptosis in muscle cells

2018 ◽  
Vol 96 (3) ◽  
pp. 304-307 ◽  
Author(s):  
Zahra Tavakoli Dargani ◽  
Reetu Singla ◽  
Taylor Johnson ◽  
Rakesh Kukreja ◽  
Dinender K. Singla
Keyword(s):  
Inflammatory Cytokine ◽  
In Vitro Model ◽  
Embryonic Stem ◽  
Mouse Embryonic Fibroblast ◽  
Caspase 1 ◽  
Key Players ◽  
Vitro Model ◽  
Embryonic Fibroblast ◽  
Muscle Toxicity

Doxorubicin (Dox) is an effective anticancer drug. Unfortunately, it causes cardiac and muscle toxicity due to increased oxidative stress and inflammation; however, it remains unknown whether Dox induces “pyroptosis” — an inflammation-mediated cell death. We investigated whether Dox induces pyroptosis in mouse soleus muscle (Sol 8) cells in vitro and to show the protective effect of embryonic stem cell exosomes (ES-exos) on pyroptosis. Dox and inflammation-induced in vitro model was generated. Pyroptosis was confirmed using immunohistochemistry (with putative markers caspase-1, IL-1β, and pro-inflammatory cytokine IL-18) and Western blotting of caspase-1 and IL-1β. The results show significant increase in the expression of caspase-1, IL-1β, and IL-18 following treatment with Dox, which was inhibited by ES-exos but not mouse embryonic fibroblast exosomes. Moreover, GW4869 compound inhibited functional activity of ES-exos, suggesting these vesicles are key players in the inhibition of pyroptosis. These results suggest that Dox induces inflammatory pyroptosis in Sol 8 cells, which is attenuated by ES-exos in vitro.

scholarly journals The Role of Plasmin/Plasminogen in Experimental Polymicrobial Sepsis

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 16-17
Author(s):  
Juliana Vago ◽  
Larissa Froede Brito ◽  
Lívia C.R. Teixeira ◽  
Izabela Z. Moreira ◽  
Thaís C. Miranda ◽  
...  
Keyword(s):  
Survival Rate ◽  
Vehicle Group ◽  
Experimental Sepsis ◽  
Inflammatory Infiltration ◽  
M1 Macrophages ◽  
Bacterial Counts ◽  
Inflammatory Parameters ◽  
Tnf Α ◽  
Lethal Sepsis

Introduction: Sepsis is characterized by systemic inflammation and increased coagulation. Although plasmin activity is increased at the onset of sepsis, increased levels of plasminogen activator inhibitor-1 can counter-regulate it triggering an imbalance of the procoagulant and fibrinolytic systems, which results in disseminated intravascular coagulation. Objective: To evaluate the role of the plasminogen/plasmin (Plg/Pla) system in experimental sepsis. Material and Methods: Plg-deficient (Plg-/-) mice, Plg+/+ littermate controls and wild-type (WT) C57Bl/6 mice were subjected to Cecal Ligation and Puncture (CLP) to induce sepsis. In this model, a double puncture is made through the cecum using either a 30 or 18-gauge needle to induce sub-lethal or lethal sepsis, respectively. Survival rates were assessed. Inflammatory parameters and bacterial counts were evaluated in peritoneal lavage fluid and blood 12h after CLP. In other experiments WT mice were treated with Pla or Plg, employing two different therapeutic protocols. Results: In WT mice, the survival rate in sub-lethal sepsis was more than 80% at 6 days after CLP, while lethal sepsis all mice died by 2 days. Increased inflammatory infiltrates were found in peritoneal cavities of mice subjected to lethal sepsis compared to sub-lethal sepsis. The plasma levels of Plg were reduced in lethal sepsis when compared to sub-lethal sepsis, while the levels of a marker for sepsis severity, IL-6, increased. In the sub-lethal model of sepsis, the survival rate of Plg-/- mice was 40% while all Plg+/+ mice survived. In this model, increased inflammatory infiltration, predominantly neutrophils and M1 macrophages, was present in peritoneal cavities of Plg-/- mice 12h after CLP, accompanied by increased IL-6 and ALT levels, compared with Plg+/+ littermates. No genotype-dependent difference in the levels of TNF-α and IL-10 were observed. The bacterial clearance in Plg-/- mice and Plg+/+ littermates were similar. In WT mice, exogenous Pla (10 µg/mice, i.p.) administered 3h after lethal sepsis did not modify the lethality rate, but decreased inflammatory infiltration (neutrophils and M1 macrophages) at the infectious site with reduced levels of CXCL1 and ALT. No differences in the levels of TNF-α, IL-1-6 and IL-10 were observed after Pla treatment. Pla administration reduced bacterial counts in blood with no differences in peritoneal fluid. In a separate protocol, administration of Pla or Plg (10 µg/mouse, i.p.) in two bolus injections, at both 6 and 12h after lethal CLP, resulted in a significant reduction in lethality rate (~30% survival) when compared to the vehicle group (no survival). Interestingly, co-injection of Plg in combination with a broad-spectrum antibiotic (imipenem 30mg/kg, i.p.) protected approximately 80% of mice, compared with 60% protection with antibiotic alone. Conclusion: Plg/Pla exhibit a protective effect in sepsis by reducing inflammatory parameters, neutrophil recruitment and tissue damage. Keywords: sepsis, plasmin, plasminogen system, CLP model. Disclosures No relevant conflicts of interest to declare.

scholarly journals Endo-β-N-acetylglucosaminidase forms N-GlcNAc protein aggregates during ER-associated degradation in Ngly1-defective cells

2015 ◽  
Vol 112 (5) ◽  
pp. 1398-1403 ◽  
Author(s):  
Chengcheng Huang ◽  
Yoichiro Harada ◽  
Akira Hosomi ◽  
Yuki Masahara-Negishi ◽  
Junichi Seino ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Genetic Disorder ◽  
Protein Aggregates ◽  
Mouse Embryonic Fibroblast ◽  
Potential Mechanism ◽  
Functional Importance ◽  
Embryonic Fibroblast ◽  
Precise Role ◽  
In Cells

The cytoplasmic peptide:N-glycanase (PNGase; Ngly1 in mice) is a deglycosylating enzyme involved in the endoplasmic reticulum (ER)-associated degradation (ERAD) process. The precise role of Ngly1 in the ERAD process, however, remains unclear in mammals. The findings reported herein, using mouse embryonic fibroblast (MEF) cells, that the ablation of Ngly1 causes dysregulation of the ERAD process. Interestingly, not only delayed degradation but also the deglycosylation of a misfolded glycoprotein was observed in Ngly1−/− MEF cells. The unconventional deglycosylation reaction was found to be catalyzed by the cytosolic endo-β-N-acetylglucosaminidase (ENGase), generating aggregation-prone N-GlcNAc proteins. The ERAD dysregulation in cells lacking Ngly1 was restored by the additional knockout of ENGase gene. Thus, our study underscores the functional importance of Ngly1 in the ERAD process and provides a potential mechanism underlying the phenotypic consequences of a newly emerging genetic disorder caused by mutation of the human NGLY1 gene.

scholarly journals Targeted Disruption of Migration Inhibitory Factor Gene Reveals Its Critical Role in Sepsis

1999 ◽  
Vol 189 (2) ◽  
pp. 341-346 ◽  
Author(s):  
Marcelo Bozza ◽  
Abhay R. Satoskar ◽  
Guosheng Lin ◽  
Bao Lu ◽  
Alison A. Humbles ◽  
...  
Keyword(s):  
Migration Inhibitory Factor ◽  
Critical Role ◽  
Embryonic Stem ◽  
Interferon Γ ◽  
Inhibitory Factor ◽  
High Dose ◽  
Wild Type ◽  
Neutrophil Accumulation ◽  
Tnf Α

To study the biologic role of migration inhibitory factor (MIF), a pleiotropic cytokine, we generated a mouse strain lacking MIF by gene targeting in embryonic stem cells. Analysis of the role of MIF during sepsis showed that MIF−/− mice were resistant to the lethal effects of high dose bacterial lipopolysaccharide (LPS), or Staphylococcus aureus enterotoxin B (SEB) with d-galactosamine and had lower plasma levels of tumor necrosis factor α (TNF-α) than did wild-type mice, but normal levels of interleukin (IL)-6 and IL-10. When stimulated with LPS and interferon γ, macrophages from MIF−/− mice showed diminished production of TNF-α, normal IL-6 and IL-12, and increased production of nitric oxide. MIF−/− animals cleared gram-negative bacteria Pseudomonas aeruginosa instilled into the trachea better than did wild-type mice and had diminished neutrophil accumulation in their bronchoalveolar fluid compared to the wild-type mice. Thioglycollate elicited peritoneal exudates in uninfected MIF−/− mice, but showed normal neutrophil accumulation. Finally, the findings of enhanced resistance to P. aeruginosa and resistance to endotoxin-induced lethal shock suggest that the counteraction or neutralization of MIF may serve as an adjunct therapy in sepsis.

scholarly journals Phenotypic and Functional Characteristics of Exosomes Derived from Irradiated Mouse Organs and Their Role in the Mechanisms Driving Non-Targeted Effects

2020 ◽  
Vol 21 (21) ◽  
pp. 8389
Author(s):  
Seda Tuncay Cagatay ◽  
Ammar Mayah ◽  
Mariateresa Mancuso ◽  
Paola Giardullo ◽  
Simonetta Pazzaglia ◽  
...  
Keyword(s):  
Membrane Vesicles ◽  
Mouse Embryonic Fibroblast ◽  
Whole Body ◽  
Recipient Mouse ◽  
Oxygen Species ◽  
Cellular Viability ◽  
Radiation Induced ◽  
Partial Body ◽  
Embryonic Fibroblast

Molecular communication between irradiated and unirradiated neighbouring cells initiates radiation-induced bystander effects (RIBE) and out-of-field (abscopal) effects which are both an example of the non-targeted effects (NTE) of ionising radiation (IR). Exosomes are small membrane vesicles of endosomal origin and newly identified mediators of NTE. Although exosome-mediated changes are well documented in radiation therapy and oncology, there is a lack of knowledge regarding the role of exosomes derived from inside and outside the radiation field in the early and delayed induction of NTE following IR. Therefore, here we investigated the changes in exosome profile and the role of exosomes as possible molecular signalling mediators of radiation damage. Exosomes derived from organs of whole body irradiated (WBI) or partial body irradiated (PBI) mice after 24 h and 15 days post-irradiation were transferred to recipient mouse embryonic fibroblast (MEF) cells and changes in cellular viability, DNA damage and calcium, reactive oxygen species and nitric oxide signalling were evaluated compared to that of MEF cells treated with exosomes derived from unirradiated mice. Taken together, our results show that whole and partial-body irradiation increases the number of exosomes, instigating changes in exosome-treated MEF cells, depending on the source organ and time after exposure.

Ectopic overexpression of Nanog induces tumorigenesis in non-tumorous fibroblasts

2016 ◽  
Vol 397 (3) ◽  
pp. 249-255 ◽  
Author(s):  
Yo Seph Park ◽  
Judee Grace E. Nemeño ◽  
Na Young Choi ◽  
Jeong Ik Lee ◽  
Kisung Ko ◽  
...  
Keyword(s):  
Stem Cells ◽  
Morphological Changes ◽  
Embryonic Stem ◽  
Mouse Embryonic Fibroblast ◽  
Tumor Formation ◽  
Proliferation Rate ◽  
Fibroblast Cells ◽  
Reprogramming Factors ◽  
Embryonic Fibroblast

Abstract Key regulatory genes in pluripotent stem cells are of interest not only as reprogramming factors but also as regulators driving tumorigenesis. Nanog is a transcription factor involved in the maintenance of embryonic stem cells and is one of the reprogramming factors along with Oct4, Sox2, and Lin28. Nanog expression has been detected in different types of tumors, and its expression is a poor prognosis for cancer patients. However, there is no clear evidence that Nanog is functionally involved in tumorigenesis. In this study, we induced overexpression of Nanog in mouse embryonic fibroblast cells and subsequently assessed their morphological changes, proliferation rate, and tumor formation ability. We found that Nanog overexpression induced immortalization of mouse embryonic fibroblast cells (MEFs) and increased their proliferation rate in vitro. We also found that formation of tumors after subcutaneous injection of retroviral-Nanog infected MEFs (N-MEFs) into athymic mouse. Cancer-related genes such as Bmi1 were expressed at high levels in N-MEFs. Hence, our results demonstrate that Nanog is able to transform normal somatic cells into tumor cells.

scholarly journals A methanolic extract of Trigonella foenum-graecum (fenugreek) seeds regulates markers of macrophage polarization.

2015 ◽  
Vol 5 (12) ◽  
pp. 417 ◽  
Author(s):  
Nurudeen Hassan ◽  
Cathryn Withycombe ◽  
Maninder Ahluwalia ◽  
Andrew Thomas ◽  
Keith Morris
Keyword(s):  
Methanolic Extract ◽  
Macrophage Polarization ◽  
Diabetic Patients ◽  
M1 Macrophages ◽  
Fenugreek Seeds ◽  
Tissue Microenvironment ◽  
Trigonella Foenum Graecum ◽  
Tnf Α ◽  
Cellular Mediators

Background: Macrophages are key cellular mediators in diabetes-related inflammation. Molecular cues such as cytokines found in the tissue microenvironment regulates the polarization of macrophages into an M1 (pro-inflammatory) or M2 (immunoregulatory) phenotype. Recent evidence suggests that M1 macrophages in diabetic patients may contribute to the complications associated with the disease such as atherosclerosis. Trigonella foenum- graecum (Tfg: fenugreek) seeds have been used in traditional medicine in Asia, Africa and the Middle-East for their alleged anti-diabetic properties.Objective: To identify the molecular mechanism(s) through which Tfg seeds exert their effects, we investigated the role of a crude methanolic extract of Tfg (FME) seeds on macrophage polarization in vitro.Method: THP-1 macrophages (Mϕ) were treated with gBSA in the presence/absence of FME and the release and expression of M1 and M2 markers/cytokines were analysed. The role of FME on NF-κB activity was also explored using transfected HEK-293T cells.Results: This study found that the FME significantly (P<0.05) decreased gBSA-induced secretion of M1 cytokines (TNF-α, IL-1β, IL-6 and IL-8) in THP-1 Mϕ cells. In the presence of gBSA, FME also significantly increased the gene expression of the M2 marker Dectin-1, but had no effect on IL-10, IL-1Ra.  FME also significantly decreased TNF-α induced NF-kB reporter activity.Conclusion: These results suggest that FME can regulate the expression of M1 and M2 markers in THP-1 Mϕ cells. This may be potentially through the modulation of NF-kB activity. Further work should be carried out to identify precise mechanism(s) involved in the effects of FME and Tfg seeds.Keywords: chronic inflammation, macrophage polarization, diabetes, glycated BSA, THP-1 cells, Trigonella foenum graecum, fenugreek seeds, NF-κB, 

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