Influence of autophagy, apoptosis and their interplay in filaricidal activity of C-cinnamoyl glycosides

Parasitology ◽  
2019 ◽  
Vol 146 (11) ◽  
pp. 1451-1461 ◽  
Author(s):  
Priya Roy ◽  
Anirban Sengupta ◽  
Nikhilesh Joardar ◽  
Arindam Bhattacharyya ◽  
Nimai Chandra Saha ◽  
...  
Keyword(s):  
Cell Death ◽  
Decisive Role ◽  
Autophagic Flux ◽  
Apoptotic Pathway ◽  
Setaria Cervi ◽  
Cell Death Pathways ◽  
Marker Proteins ◽  
Apoptotic Protein ◽  
Causative Factors

AbstractThe present work aims to explore the mechanism of action of C-cinnamoyl glycoside as an antifilarial agent against the bovine filarial nematode Setaria cervi. Both apoptosis and autophagy programmed cell death pathways play a significant role in parasitic death. The generation of reactive oxygen species, alteration of the level of antioxidant components and disruption of mitochondrial membrane potential may be the causative factors that drive the parasitic death. Monitoring of autophagic flux via the formation of autophagosome and autophagolysosome was detected via CYTO ID dye. The expression profiling of both apoptotic and autophagic marker proteins strongly support the initial findings of these two cell death processes. The increased interaction of pro-autophagic protein Beclin1 with BCL-2 may promote apoptotic pathway by suppressing anti-apoptotic protein BCL-2 from its function. This in turn partially restrains the autophagic pathway by engaging Beclin1 in the complex. But overall positive increment in autophagic flux was observed. Dynamic interaction and regulative balance of these two critical cellular pathways play a decisive role in controlling disease pathogenesis. Therefore, the present experimental work may prosper the chance for C-cinnamoyl glycosides to become a potential antifilarial therapeutic in the upcoming day after detail in vivo study and proper clinical trial.

scholarly journals Autophagy Is Involved in Mesenchymal Stem Cell Death in Coculture with Chondrocytes

Cartilage ◽  
2020 ◽  
pp. 194760352094122
Author(s):  
Carlo Alberto Paggi ◽  
Amel Dudakovic ◽  
Yao Fu ◽  
Catalina Galeano Garces ◽  
Mario Hevesi ◽  
...  
Keyword(s):  
Cell Death ◽  
Rna Sequencing ◽  
Human Mscs ◽  
Chondrocyte Proliferation ◽  
Apoptotic Pathway ◽  
Primary Chondrocytes ◽  
Cartilage Formation ◽  
Cell Death Pathways

Objective Cartilage formation is stimulated in mixtures of chondrocytes and human adipose–derived mesenchymal stromal cells (MSCs) both in vitro and in vivo. During coculture, human MSCs perish. The goal of this study is to elucidate the mechanism by which adipose tissue–derived MSC cell death occurs in the presence of chondrocytes. Methods Human primary chondrocytes were cocultured with human MSCs derived from 3 donors. The cells were cultured in monoculture or coculture (20% chondrocytes and 80% MSCs) in pellets (200,000 cells/pellet) for 7 days in chondrocyte proliferation media in hypoxia (2% O2). RNA sequencing was performed to assess for differences in gene expression between monocultures or coculture. Immune fluorescence assays were performed to determine the presence of caspase-3, LC3B, and P62. Results RNA sequencing revealed significant upregulation of >90 genes in the 3 cocultures when compared with monocultures. STRING analysis showed interconnections between >50 of these genes. Remarkably, 75% of these genes play a role in cell death pathways such as apoptosis and autophagy. Immunofluorescence shows a clear upregulation of the autophagic machinery with no substantial activation of the apoptotic pathway. Conclusion In cocultures of human MSCs with primary chondrocytes, autophagy is involved in the disappearance of MSCs. We propose that this sacrificial cell death may contribute to the trophic effects of MSCs on cartilage formation.

scholarly journals The Novel ALG-2 Target Protein CDIP1 Promotes Cell Death by Interacting with ESCRT-I and VAPA/B

2021 ◽  
Vol 22 (3) ◽  
pp. 1175
Author(s):  
Ryuta Inukai ◽  
Kanako Mori ◽  
Keiko Kuwata ◽  
Chihiro Suzuki ◽  
Masatoshi Maki ◽  
...  
Keyword(s):  
Cell Death ◽  
Essential Gene ◽  
Susceptibility Gene ◽  
Target Protein ◽  
Hek293 Cells ◽  
Dependent Manner ◽  
Gene Encoding ◽  
Endosomal Sorting ◽  
Cell Death Pathways ◽  
Apoptotic Protein

Apoptosis-linked gene 2 (ALG-2, also known as PDCD6) is a member of the penta-EF-hand (PEF) family of Ca2+-binding proteins. The murine gene encoding ALG-2 was originally reported to be an essential gene for apoptosis. However, the role of ALG-2 in cell death pathways has remained elusive. In the present study, we found that cell death-inducing p53 target protein 1 (CDIP1), a pro-apoptotic protein, interacts with ALG-2 in a Ca2+-dependent manner. Co-immunoprecipitation analysis of GFP-fused CDIP1 (GFP-CDIP1) revealed that GFP-CDIP1 associates with tumor susceptibility gene 101 (TSG101), a known target of ALG-2 and a subunit of endosomal sorting complex required for transport-I (ESCRT-I). ESCRT-I is a heterotetrameric complex composed of TSG101, VPS28, VPS37 and MVB12/UBAP1. Of diverse ESCRT-I species originating from four VPS37 isoforms (A, B, C, and D), CDIP1 preferentially associates with ESCRT-I containing VPS37B or VPS37C in part through the adaptor function of ALG-2. Overexpression of GFP-CDIP1 in HEK293 cells caused caspase-3/7-mediated cell death. In addition, the cell death was enhanced by co-expression of ALG-2 and ESCRT-I, indicating that ALG-2 likely promotes CDIP1-induced cell death by promoting the association between CDIP1 and ESCRT-I. We also found that CDIP1 binds to vesicle-associated membrane protein-associated protein (VAP)A and VAPB through the two phenylalanines in an acidic tract (FFAT)-like motif in the C-terminal region of CDIP1, mutations of which resulted in reduction of CDIP1-induced cell death. Therefore, our findings suggest that different expression levels of ALG-2, ESCRT-I subunits, VAPA and VAPB may have an impact on sensitivity of anticancer drugs associated with CDIP1 expression.

scholarly journals Inhibiting autophagy targets human leukemic stem cells and hypoxic AML blasts by disrupting mitochondrial homeostasis

2021 ◽  
Vol 5 (8) ◽  
pp. 2087-2100
Author(s):  
Kaitlyn M. Dykstra ◽  
Hannah R. S. Fay ◽  
Ashish C. Massey ◽  
Neng Yang ◽  
Matthew Johnson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Death ◽  
Mitochondrial Respiration ◽  
Prolonged Exposure ◽  
Tumor Burden ◽  
Therapeutic Interventions ◽  
Autophagic Flux ◽  
Mitochondrial Homeostasis ◽  
Nsg Mice

Abstract Leukemia stem cells (LSCs) and therapy-resistant acute myeloid leukemia (AML) blasts contribute to the reinitiation of leukemia after remission, necessitating therapeutic interventions that target these populations. Autophagy is a prosurvival process that allows for cells to adapt to a variety of stressors. Blocking autophagy pharmacologically by using mechanistically distinct inhibitors induced apoptosis and prevented colony formation in primary human AML cells. The most effective inhibitor, bafilomycin A1 (Baf A1), also prevented the in vivo maintenance of AML LSCs in NSG mice. To understand why Baf A1 exerted the most dramatic effects on LSC survival, we evaluated mitochondrial function. Baf A1 reduced mitochondrial respiration and stabilized PTEN-induced kinase-1 (PINK-1), which initiates autophagy of mitochondria (mitophagy). Interestingly, with the autophagy inhibitor chloroquine, levels of enhanced cell death and reduced mitochondrial respiration phenocopied the effects of Baf A1 only when cultured in hypoxic conditions that mimic the marrow microenvironment (1% O2). This indicates that increased efficacy of autophagy inhibitors in inducing AML cell death can be achieved by concurrently inducing mitochondrial damage and mitophagy (pharmacologically or by hypoxic induction) and blocking mitochondrial degradation. In addition, prolonged exposure of AML cells to hypoxia induced autophagic flux and reduced chemosensitivity to cytarabine (Ara-C), which was reversed by autophagy inhibition. The combination of Ara-C with Baf A1 also decreased tumor burden in vivo. These findings demonstrate that autophagy is critical for mitochondrial homeostasis and survival of AML cells in hypoxia and support the development of autophagy inhibitors as novel therapeutic agents for AML.

scholarly journals Human Monocyte Recognition of Adenosine-Based Cyclic Dinucleotides Unveils the A2a GαsProtein-Coupled Receptor Tonic Inhibition of Mitochondrially Induced Cell Death

2014 ◽  
Vol 35 (2) ◽  
pp. 479-495 ◽  
Author(s):  
Marie Tosolini ◽  
Frédéric Pont ◽  
Delphine Bétous ◽  
Emmanuel Ravet ◽  
Laetitia Ligat ◽  
...  
Keyword(s):  
Cell Death ◽  
Mammalian Cells ◽  
Mitochondrial Permeability Transition ◽  
Human Monocyte ◽  
Inverse Agonist ◽  
Apoptotic Pathway ◽  
Cyclic Dinucleotides ◽  
Agonist Ligands

Cyclic dinucleotides are important messengers for bacteria and protozoa and are well-characterized immunity alarmins for infected mammalian cells through intracellular binding to STING receptors. We sought to investigate their unknown extracellular effects by adding cyclic dinucleotides to the culture medium of freshly isolated human blood cellsin vitro. Here we report that adenosine-containing cyclic dinucleotides induce the selective apoptosis of monocytes through a novel apoptotic pathway. We demonstrate that these compounds are inverse agonist ligands of A2a, a Gαs-coupled adenosine receptor selectively expressed by monocytes. Inhibition of monocyte A2a by these ligands induces apoptosis through a mechanism independent of that of the STING receptors. The blockade of basal (adenosine-free) signaling from A2a inhibits protein kinase A (PKA) activity, thereby recruiting cytosolic p53, which opens the mitochondrial permeability transition pore and impairs mitochondrial respiration, resulting in apoptosis. A2a antagonists and inverse agonist ligands induce apoptosis of human monocytes, while A2a agonists are antiapoptotic.In vivo, we used a mock developing human hematopoietic system through NSG mice transplanted with human CD34+cells. Treatment with cyclic di-AMP selectively depleted A2a-expressing monocytes and their precursors via apoptosis. Thus, monocyte recognition of cyclic dinucleotides unravels a novel proapoptotic pathway: the A2a Gαsprotein-coupled receptor (GPCR)-driven tonic inhibitory signaling of mitochondrion-induced cell death.

scholarly journals Loss of Heterochromatin Protein 1 (HP1) chromodomain function in mammalian cells by intracellular antibodies causes cell death

2002 ◽  
Vol 115 (9) ◽  
pp. 1803-1813 ◽  
Author(s):  
Ilaria Filesi ◽  
Alessio Cardinale ◽  
Sjaak van der Sar ◽  
Ian G. Cowell ◽  
Prim B. Singh ◽  
...  
Keyword(s):  
Cell Death ◽  
Mammalian Cells ◽  
Wide Spectrum ◽  
Human Fibroblasts ◽  
Protein S ◽  
Single Chain ◽  
Heterochromatin Protein 1 ◽  
Apoptotic Pathway

The chromodomain (CD) is a highly conserved motif present in a variety of animal and plant proteins, and its probable role is to assemble a variety of macromolecular complexes in chromatin. The importance of the CD to the survival of mammalian cells has been tested. Accordingly, we have ablated CD function using two single-chain intracellular Fv (scFv) fragments directed against non-overlapping epitopes within the HP1 CD motif. The scFv fragments can recognize both CD motifs of HP1 and Polycomb (Pc) in vitro and, when expressed intracellularly, interact with and dislodge the HP1 protein(s) from their heterochromatin localization in vivo. Mouse and human fibroblasts expressing anti-chromodomain scFv fragments show a cell-lethal phenotype and an apoptotic morphology becomes apparent soon after transfection. The mechanism of cell death appears to be p53 independent, and the cells are only partly rescued by incubation with the wide spectrum caspase inhibitor Z-VAD fmk. We conclude that expression of anti-chromodomain intracellular antibodies is sufficient to trigger a p53-independent apoptotic pathway that is only partly dependent on the known Z-VAD-inhibitable caspases, suggesting that CD function is essential for cell survival.

scholarly journals Oncogenic signaling inhibits c-FLIPL expression and its non-apoptotic function during ECM-detachment

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Matyas Abel Tsegaye ◽  
Jianping He ◽  
Kyle McGeehan ◽  
Ireland M. Murphy ◽  
Mati Nemera ◽  
...  
Keyword(s):  
Cell Death ◽  
Cancer Cells ◽  
Protein C ◽  
Tumor Tissue ◽  
Oncogenic Signaling ◽  
Inhibitory Protein ◽  
Cell Death Pathways ◽  
Apoptotic Protein ◽  
Cellular Context ◽  
Context Dependent

AbstractInhibition of programmed cell death pathways is frequently observed in cancer cells where it functions to facilitate tumor progression. However, some proteins involved in the regulation of cell death function dichotomously to both promote and inhibit cell death depending on the cellular context. As such, understanding how cell death proteins are regulated in a context-dependent fashion in cancer cells is of utmost importance. We have uncovered evidence that cellular FLICE-like Inhibitory Protein (c-FLIP), a well-known anti-apoptotic protein, is often downregulated in tumor tissue when compared to adjacent normal tissue. These data argue that c-FLIP may have activity distinct from its canonical role in antagonizing cell death. Interestingly, we have discovered that detachment from extracellular matrix (ECM) serves as a signal to elevate c-FLIP transcription and that oncogenic signaling blocks ECM-detachment-induced c-FLIP elevation. In addition, our data reveal that downregulation of c-FLIP promotes luminal filling in mammary acini and that c-FLIP overexpression in cancer cells inhibits colony formation in cells exposed to ECM-detachment. Taken together, our study reveals an unexpected, non-apoptotic role for c-FLIP during ECM-detachment and raises the possibility that c-FLIP may have context-dependent roles during tumorigenesis.

scholarly journals DOP86 An IFN-STAT1-MLKL axis drives programmed necrosis of Paneth cells in Crohn’s ileitis

2020 ◽  
Vol 14 (Supplement_1) ◽  
pp. S125-S126
Author(s):  
L HARTMANN ◽  
B Siegmund ◽  
C Weidinger ◽  
C Becker ◽  
M F Neurath ◽  
...  
Keyword(s):  
Cell Death ◽  
Intestinal Inflammation ◽  
Paneth Cell ◽  
Signalling Pathways ◽  
Caspase 8 ◽  
Programmed Necrosis ◽  
Cell Death Pathways ◽  
Small Intestinal

Abstract Background Interferons (IFNs) are immune-modulatory cytokines expressed by epithelial and mucosal cells in response to viral and bacterial infection. Just recently, we discovered a correlation between IFN-λ expression and disease activity, including small intestinal inflammation and Paneth cell dysfunction, in human Crohn’s disease patients. On a molecular level, we uncovered that IFN-λ mediates epithelial cell death, in particular, Paneth cell death by a programmed necrosis, dependent on STAT1 activation and controlled by caspase-8. These results suggested that IFN-λ can be considered as a pathogenic cytokine in Crohn′s ileitis and should be considered as a new and promising target for future therapeutic intervention for this particular subtype of IBD. Our central question is now by which pathways interferon-regulated programmed necrosis of epithelial cells contributes to intestinal inflammation and how these mechanisms could be targeted for future therapeutic intervention. Methods We use a mouse model for Crohn’s Disease like inflammation and Paneth cell death that has a specific deletion of Caspase-8 in intestinal epithelial cells (Casp8∆IEC). We stimulate small intestinal organoids derived from Casp8∆IEC mice with IFNs in vitro and we overexpress IFN-λ in these mice in vivo by hydrodynamic tail vein injection of an IFN-λ expression vector. Furthermore, we use JAK-inhibitors to impede pharmacologically cell death pathways in the pathogenesis of intestinal inflammation in vitro and in vivo. Results We uncovered that gene expression of the cell death mediators Mlkl and Caspase-8 is dependent on IFN-λ-mediated JAK-STAT1 signalling. The non-specific pan JAK-inhibitor Tofacitinib is able to attenuate gene expression of Mlkl and Caspase-8 in vitro as well as in vivo. It prevents non-apoptotic as well as apoptotic cell death of small intestinal organoids stimulated with IFN-λ and is sufficient to prevent small intestinal tissue destruction in Casp8∆IEC mice challenged with IFN-λ. Additionally, we use the selective JAK1-inhibitor Filgotinib to limit the targeted JAK-STAT signalling pathways to only JAK1-STAT1 signalling and thus reduce side effects of the inhibitor on other signalling pathways. This had a similar effect as Tofacitinib suggesting that IFN controls MLKL-mediated cell death via JAK1. Conclusion In summary, our results indicate that targeting IFN-λ-mediated JAK-STAT1 signalling by the small-molecules Tofacitinib and Filgotinib impedes induction of Mlkl and Caspase-8-mediated cell death pathways. Therefore, JAK1 inhibitors such as Filgotinib might represent a promising novel therapy that may be sufficient to achieve efficacy particularly in Crohn′s ileitis patients who display elevated IFN-l serum levels.

scholarly journals Downregulation of PUMA underlies resistance to FGFR1 inhibitors in the stem cell leukemia/lymphoma syndrome

2020 ◽  
Vol 11 (10) ◽  
Author(s):  
Yun Liu ◽  
Baohuan Cai ◽  
Yating Chong ◽  
Hualei Zhang ◽  
Chesley-Anne Kemp ◽  
...  
Keyword(s):  
Cell Death ◽  
Kinase Inhibitors ◽  
Leukemic Cells ◽  
Direct Role ◽  
Genetic Changes ◽  
Cell Death Pathways ◽  
Pten Deletion ◽  
Induced Apoptosis ◽  
Resistant Cells

Abstract Resistance to molecular therapies frequently occur due to genetic changes affecting the targeted pathway. In myeloid and lymphoid leukemias/lymphomas resulting from constitutive activation of FGFR1 kinases, resistance has been shown to be due either to mutations in FGFR1 or deletions of PTEN. RNA-Seq analysis of the resistant clones demonstrates expression changes in cell death pathways centering on the p53 upregulated modulator of apoptosis (Puma) protein. Treatment with different tyrosine kinase inhibitors (TKIs) revealed that, in both FGFR1 mutation and Pten deletion-mediated resistance, sustained Akt activation in resistant cells leads to compromised Puma activation, resulting in suppression of TKI-induced apoptosis. This suppression of Puma is achieved as a result of sequestration of inactivated p-Foxo3a in the cytoplasm. CRISPR/Cas9 mediated knockout of Puma in leukemic cells led to an increased drug resistance in the knockout cells demonstrating a direct role in TKI resistance. Since Puma promotes cell death by targeting Bcl2, TKI-resistant cells showed high Bcl2 levels and targeting Bcl2 with Venetoclax (ABT199) led to increased apoptosis in these cells. In vivo treatment of mice xenografted with resistant cells using ABT199 suppressed leukemogenesis and led to prolonged survival. This in-depth survey of the underlying genetic mechanisms of resistance has identified a potential means of treating FGFR1-driven malignancies that are resistant to FGFR1 inhibitors.

scholarly journals FTY720 induces non-canonical phosphatidylserine externalization and cell death in acute myeloid leukemia

2019 ◽  
Vol 10 (11) ◽  
Author(s):  
Megan M. Young ◽  
Van Bui ◽  
Chong Chen ◽  
Hong-Gang Wang
Keyword(s):  
Cell Death ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Membrane Integrity ◽  
Mechanistic Study ◽  
Flip Flop ◽  
Anticancer Efficacy ◽  
Cell Death Pathways ◽  
Acute Myeloid

Abstract FTY720 (fingolimod) is a FDA-approved sphingosine analog that is phosphorylated in vivo to modulate sphingosine-1-phosphate receptor (S1PR) signaling for immunosuppression in patients with refractory multiple sclerosis. FTY720 also exhibits promising anticancer efficacy in several preclinical models. While FTY720-induced cytotoxicity is not due to S1PR signaling, the mechanism remains unclear and is reported to occur through various cell death pathways. Here, we performed a systematic, mechanistic study of FTY720-induced cell death in acute myeloid leukemia (AML). We found that FTY720 induced cell death in a panel of genetically diverse AML cell lines that was accompanied by rapid phosphatidylserine (PS) externalization. Importantly, FTY720-induced PS exposure was not due to any direct effects on plasma membrane integrity and was independent of canonical signaling by regulated cell death pathways known to activate lipid flip-flop, including caspase-dependent apoptosis/pyroptosis, necroptosis, ferroptosis, and reactive oxygen species-mediated cell death. Notably, PS exposure required cellular vacuolization induced by defects in endocytic trafficking and was suppressed by the inhibition of PP2A and shedding of Annexin V-positive subcellular particles. Collectively, our studies reveal a non-canonical pathway underlying PS externalization and cell death in AML to provide mechanistic insight into the antitumor properties of FTY720.

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