Polymorphonuclear neutrophils (PMNs) play a key role in host defense. However, their massive accumulation at the site of inflammation can delay regenerative healing processes and can initiate pathological inflammatory processes. Thus, the efficient clearance of PMNs mediated by the induction of regulated cell death is a key process preventing the development of these pathological conditions. Myeloperoxidase (MPO), a highly abundant enzyme in PMN granules, primarily connected with PMN defense machinery, is suggested to play a role in PMN-regulated cell death. However, the contribution of MPO to the mechanisms of PMN cell death remains incompletely characterized. Herein, the process of the cell death of mouse PMNs induced by three different stimuli - phorbol 12-myristate 13-acetate (PMA), opsonized streptococcus (OST), and N-formyl-met-leu-phe (fMLP) - was investigated. MPO-deficient PMNs revealed a significantly decreased rate of cell death characterized by phosphatidylserine surface exposure and cell membrane permeabilization. An inhibitor of MPO activity, 4-aminobenzoic acid hydrazide, did not exhibit a significant effect on PMA-induced cell death compared to MPO deficiency. Interestingly, only the limited activation of markers related to apoptotic cell death was observed (e.g. caspase 8 activation, Bax expression) and they mostly did not correspond to phosphatidylserine surface exposure. Furthermore, a marker characterizing autophagy, cleavage of LC3 protein, as well as histone H3 citrullination and its surface expression was observed. Collectively, the data show the ability of MPO to modulate the life span of PMNs primarily through the potentiation of cell membrane permeabilization and phosphatidylserine surface exposure.

• Klíčová slova
• annexin V, apoptosis, cell death, inflammation, myeloperoxidase, neutrophils,
• MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• neutrofily metabolismus   patologie   MeSH
• peroxidasa nedostatek   metabolismus   MeSH
• regulovaná buněčná smrt MeSH
• zánět metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• peroxidasa MeSH

Programmed cell death (PCD) pathways play a crucial role in the response of cancer cells to treatment. Their dysregulation is one of the cancer hallmarks and one of the reasons of drug resistance. Here, we studied the significance of the individual members of PCD signaling pathways in response to treatment with common anti-cancer drugs using the T-cell leukemia Jurkat cells with single or double knockouts of necroptosis and/or apoptosis genes. We identified apoptosis as the primary cell death pathway upon anti-cancer drugs treatment. The cells with knocked out either Fas-associated protein with death domain (FADD) or all executioner caspases were resistant. This resistance could be partially overcome by induction of RIP1-dependent necroptosis through TNFR1 activation using combined treatment with TNF-α and smac mimetic (LCL161). RIP1 was essential for cellular response to TNF-α and smac mimetic, but dispensable for the response to anti-cancer drugs. Here, we demonstrated the significance of FADD and executioner caspases in carrying out programmed cell death upon anti-cancer drug treatments and the ability of combined treatment with TNF-α and smac mimetic to partially overcome drug resistance of FADD and/or CASP3/7/6-deficient cells via RIP1-dependent necroptosis. Thus, a combination of TNF-α and smac mimetic could be a suitable strategy for overcoming resistance to therapy in cells unable to trigger apoptosis.

• Klíčová slova
• FADD, RIP1, RIP3, apoptosis, cancer, caspase, cell death, leukemia, necroptosis, ripoptosome,
• MeSH
• apoptóza účinky léků   genetika   MeSH
• buněčná smrt účinky léků   MeSH
• chemorezistence účinky léků   fyziologie   MeSH
• Jurkat buňky MeSH
• kaspasy genetika   MeSH
• komplex proteinů jaderného póru metabolismus   MeSH
• lidé MeSH
• nekroptóza účinky léků   genetika   MeSH
• protein FADD asociující s Fas genetika   MeSH
• proteiny vázající RNA metabolismus   MeSH
• protinádorové látky farmakologie   MeSH
• TNF-alfa farmakologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• AGFG1 protein, human MeSH Prohlížeč
• FADD protein, human MeSH Prohlížeč
• kaspasy MeSH
• komplex proteinů jaderného póru MeSH
• protein FADD asociující s Fas MeSH
• proteiny vázající RNA MeSH
• protinádorové látky MeSH
• TNF-alfa MeSH

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.

• Klíčová slova
• Autophagosome, LC3, cancer, flux, lysosome, macroautophagy, neurodegeneration, phagophore, stress, vacuole,
• MeSH
• autofagie * fyziologie   MeSH
• autofagozomy MeSH
• biologické markery MeSH
• biotest normy   MeSH
• lidé MeSH
• lyzozomy MeSH
• proteiny spojené s autofagií metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• směrnice MeSH
• Názvy látek
• biologické markery MeSH
• proteiny spojené s autofagií MeSH

Cell viability and cytotoxicity assays are highly important for drug screening and cytotoxicity tests of antineoplastic or other therapeutic drugs. Even though biochemical-based tests are very helpful to obtain preliminary preview, their results should be confirmed by methods based on direct cell death assessment. In this study, time-dependent changes in quantitative phase-based parameters during cell death were determined and methodology useable for rapid and label-free assessment of direct cell death was introduced. The goal of our study was distinction between apoptosis and primary lytic cell death based on morphologic features. We have distinguished the lytic and non-lytic type of cell death according to their end-point features (Dance of Death typical for apoptosis versus swelling and membrane rupture typical for all kinds of necrosis common for necroptosis, pyroptosis, ferroptosis and accidental cell death). Our method utilizes Quantitative Phase Imaging (QPI) which enables the time-lapse observation of subtle changes in cell mass distribution. According to our results, morphological and dynamical features extracted from QPI micrographs are suitable for cell death detection (76% accuracy in comparison with manual annotation). Furthermore, based on QPI data alone and machine learning, we were able to classify typical dynamical changes of cell morphology during both caspase 3,7-dependent and -independent cell death subroutines. The main parameters used for label-free detection of these cell death modalities were cell density (pg/pixel) and average intensity change of cell pixels further designated as Cell Dynamic Score (CDS). To the best of our knowledge, this is the first study introducing CDS and cell density as a parameter typical for individual cell death subroutines with prediction accuracy 75.4% for caspase 3,7-dependent and -independent cell death.

• MeSH
• algoritmy MeSH
• apoptóza * účinky léků   MeSH
• buněčná smrt * účinky léků   MeSH
• buňky ultrastruktura   MeSH
• časosběrné zobrazování metody   MeSH
• časové faktory MeSH
• kultivované buňky MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• optické zobrazování metody   MeSH
• počet buněk MeSH
• statistické modely MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Lysosome-activated apoptosis represents an alternative method of overcoming tumor resistance compared to traditional forms of treatment. Pulsed magnetic fields open a new avenue for controlled and targeted initiation of lysosomal permeabilization in cancer cells via mechanical actuation of magnetic nanomaterials. In this study we used a noninvasive tool; namely, a benchtop pulsed magnetic system, which enabled remote activation of apoptosis in liver cancer cells. The magnetic system we designed represents a platform that can be used in a wide range of biomedical applications. We show that liver cancer cells can be loaded with superparamagnetic iron oxide nanoparticles (SPIONs). SPIONs retained in lysosomal compartments can be effectively actuated with a high intensity (up to 8 T), short pulse width (~15 µs), pulsed magnetic field (PMF), resulting in lysosomal membrane permeabilization (LMP) in cancer cells. We revealed that SPION-loaded lysosomes undergo LMP by assessing an increase in the cytosolic activity of the lysosomal cathepsin B. The extent of cell death induced by LMP correlated with the accumulation of reactive oxygen species in cells. LMP was achieved for estimated forces of 700 pN and higher. Furthermore, we validated our approach on a three-dimensional cellular culture model to be able to mimic in vivo conditions. Overall, our results show that PMF treatment of SPION-loaded lysosomes can be utilized as a noninvasive tool to remotely induce apoptosis.

• Klíčová slova
• apoptosis, lysosomal death pathways, lysosomal membrane permeabilization, magnetic nanoparticles, pulsed magnetic field,
• Publikační typ
• časopisecké články MeSH

Oxadiazines are heterocyclic compounds containing N-N-O or N-N-C-O system within a six membered ring. These structures have been up to now exclusively prepared via organic synthesis. Here, we report the discovery of a natural oxadiazine nocuolin A (NoA) that has a unique structure based on 1,2,3-oxadiazine. We have identified this compound in three independent cyanobacterial strains of genera Nostoc, Nodularia, and Anabaena and recognized the putative gene clusters for NoA biosynthesis in their genomes. Its structure was characterized using a combination of NMR, HRMS and FTIR methods. The compound was first isolated as a positive hit during screening for apoptotic inducers in crude cyanobacterial extracts. We demonstrated that NoA-induced cell death has attributes of caspase-dependent apoptosis. Moreover, NoA exhibits a potent anti-proliferative activity (0.7-4.5 μM) against several human cancer lines, with p53-mutated cell lines being even more sensitive. Since cancers bearing p53 mutations are resistant to several conventional anti-cancer drugs, NoA may offer a new scaffold for the development of drugs that have the potential to target tumor cells independent of their p53 status. As no analogous type of compound was previously described in the nature, NoA establishes a novel class of bioactive secondary metabolites.

• MeSH
• apoptóza účinky léků   MeSH
• chromatografie kapalinová MeSH
• HeLa buňky MeSH
• hmotnostní spektrometrie MeSH
• lidé MeSH
• magnetická rezonanční spektroskopie MeSH
• molekulární struktura MeSH
• multigenová rodina MeSH
• oxaziny chemie   farmakologie   MeSH
• sekvence aminokyselin MeSH
• sekvenční homologie aminokyselin MeSH
• sinice chemie   MeSH
• spektroskopie infračervená s Fourierovou transformací MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• nocuolin A MeSH Prohlížeč
• oxaziny MeSH

Progressing malignancies establish robust immunosuppressive networks that operate both systemically and locally. In particular, as tumors escape immunosurveillance, they recruit increasing amounts of myeloid and lymphoid cells that exert pronounced immunosuppressive effects. These cells not only prevent the natural recognition of growing neoplasms by the immune system, but also inhibit anticancer immune responses elicited by chemo-, radio- and immuno therapeutic interventions. Throughout the past decade, multiple strategies have been devised to counteract the accumulation or activation of tumor-infiltrating immunosuppressive cells for therapeutic purposes. Here, we review recent preclinical and clinical advances on the use of small molecules that target the immunological tumor microenvironment for cancer therapy. These agents include inhibitors of indoleamine 2,3-dioxigenase 1 (IDO1), prostaglandin E2, and specific cytokine receptors, as well as modulators of intratumoral purinergic signaling and arginine metabolism.

• Klíčová slova
• Adenosine, IDO1, PGE2, Tregs, myeloid-derived suppressor cells, tumor-associated macrophages,
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

• Klíčová slova
• LC3, autolysosome, autophagosome, chaperone-mediated autophagy, flux, lysosome, macroautophagy, phagophore, stress, vacuole,
• MeSH
• autofagie * fyziologie   MeSH
• biotest metody   normy   MeSH
• lidé MeSH
• počítačová simulace MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Research Support, N.I.H., Extramural MeSH
• směrnice MeSH

The term "immunogenic cell death" (ICD) is commonly employed to indicate a peculiar instance of regulated cell death (RCD) that engages the adaptive arm of the immune system. The inoculation of cancer cells undergoing ICD into immunocompetent animals elicits a specific immune response associated with the establishment of immunological memory. Only a few agents are intrinsically endowed with the ability to trigger ICD. These include a few chemotherapeutics that are routinely employed in the clinic, like doxorubicin, mitoxantrone, oxaliplatin, and cyclophosphamide, as well as some agents that have not yet been approved for use in humans. Accumulating clinical data indicate that the activation of adaptive immune responses against dying cancer cells is associated with improved disease outcome in patients affected by various neoplasms. Thus, novel therapeutic regimens that trigger ICD are urgently awaited. Here, we discuss current combinatorial approaches to convert otherwise non-immunogenic instances of RCD into bona fide ICD.

• Klíčová slova
• ATP, HMGB1, autophagy, calreticulin, endoplasmic reticulum stress, type I interferon,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

The use of patient-derived dendritic cells (DCs) as a means to elicit therapeutically relevant immune responses in cancer patients has been extensively investigated throughout the past decade. In this context, DCs are generally expanded, exposed to autologous tumor cell lysates or loaded with specific tumor-associated antigens (TAAs), and then reintroduced into patients, often in combination with one or more immunostimulatory agents. As an alternative, TAAs are targeted to DCs in vivo by means of monoclonal antibodies, carbohydrate moieties or viral vectors specific for DC receptors. All these approaches have been shown to (re)activate tumor-specific immune responses in mice, often mediating robust therapeutic effects. In 2010, the first DC-based preparation (sipuleucel-T, also known as Provenge®) has been approved by the US Food and Drug Administration (FDA) for use in humans. Reflecting the central position occupied by DCs in the regulation of immunological tolerance and adaptive immunity, the interest in harnessing them for the development of novel immunotherapeutic anticancer regimens remains high. Here, we summarize recent advances in the preclinical and clinical development of DC-based anticancer therapeutics.

• Klíčová slova
• DC, dendritic cell, DC-based vaccination, FDA, Food and Drug Administration, IFN, interferon, MRC1, mannose receptor, C type 1, MUC1, mucin 1, TAA, tumor-associated antigen, TLR, Toll-like receptor, Toll-like receptor agonists, Treg, regulatory T cell, WT1, Wilms tumor 1, antigen cross-presentation, autophagy, iDC, immature DC, immunogenic cell death, mDC, mature DC, pDC, plasmacytoid DC, regulatory T cells,
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH