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Neuzil, Jiri
Publications (10 of 20) Show all publications
Terman, A., Dalen, H., Eaton, J. W., Neuzil, J. & Brunka, U. (2004). Aging of cardiac myocytes in culture - Oxidative stress, lipofuscin accumulation, and mitochondrial turnover. Annals of the New York Academy of Sciences, 1019, 70-77
Open this publication in new window or tab >>Aging of cardiac myocytes in culture - Oxidative stress, lipofuscin accumulation, and mitochondrial turnover
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2004 (English)In: Annals of the New York Academy of Sciences, ISSN 0077-8923, E-ISSN 1749-6632, Vol. 1019, p. 70-77Article in journal (Refereed) Published
Abstract [en]

Oxidative stress is believed to be an important contributor to aging, mainly affecting long-lived postmitotic cells such as cardiac myocytes and neurons. Aging cells accumulate functionally effete, often mutant and enlarged mitochondria, as well as an intralysosomal undegradable pigment, lipofuscin. To provide better insight into the role of oxidative stress, mitochondrial damage, and lipofuscinogenesis in postmitotic aging, we studied the relationship between these parameters in cultured neonatal rat cardiac myocytes. It was found that the content of lipofuscin, which varied drastically between cells, positively correlated with mitochondrial damage (evaluated by decreased innermembrane potential), as well as with the production of reactive oxygen species. These results suggest that both lipofuscin accumulation and mitochondrial damage have common underlying mechanisms, likely including imperfect autophagy and ensuing lysosomal degradation of oxidatively damaged mitochondria and other organelles. Increased size of mitochondria (possibly resulting from impaired fission due to oxidative damage to mitochondrial DNA, membranes, and proteins) also may interfere with mitochondrial turnover, leading to the appearance of so-called "giant" mitochondria. This assumption is based on our observation that pharmacological inhibition of autophagy with 3-methyladenine induced only moderate accumulation of large (senescent-like) mitochondria but drastically increased numbers of small, apparently normal mitochondria, reflecting their rapid turnover and suggesting that enlarged mitochondria are poorly autophagocytosed. Overall, our findings emphasize the importance of mitochondrial turnover in postmitotic aging and provide further support for the mitochondrial-lysosomal axis theory of aging.

Keywords
aging, heart, lipofuscin, lysosomes, mitochondria, oxidative stress
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-46195 (URN)10.1196/annals.1297.015 (DOI)
Available from: 2009-10-11 Created: 2009-10-11 Last updated: 2017-12-13
Jostarndt, K., Rubic, T., Kuhn, H., Anthosen, M. W., Andera, L., Gellert, N., . . . Neuzil, J. (2004). Enzymatically modified low-density lipoprotein upregulates CD36 in low-differentiated monocytic cells in a peroxisome proliferator-activated receptor-γ-dependent way. Biochemical Pharmacology, 67(5), 841-854
Open this publication in new window or tab >>Enzymatically modified low-density lipoprotein upregulates CD36 in low-differentiated monocytic cells in a peroxisome proliferator-activated receptor-γ-dependent way
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2004 (English)In: Biochemical Pharmacology, ISSN 0006-2952, E-ISSN 1356-1839, Vol. 67, no 5, p. 841-854Article in journal (Refereed) Published
Abstract [en]

Peroxisome proliferator-activated receptor-γ (PPARγ) has been suggested to upregulate CD36. Since free oxidized polyunsaturated fatty acids are PPARγ ligands, we studied the effects of LDL modified by the simultaneous action of sPLA2 and 15-lipoxygenase (15LO) on CD36 expression and PPARγ activation in monocytic cells. Exposure of MM6 cells, which do not express CD36 or other scavenger receptors, to such enzymatically modified LDL (enzLDL) resulted in upregulation of CD36 surface protein and mRNA expression. Similar effects were observed with free 13-hydroperoxyoctadecadienoic acid but not its esterified counterpart. Less pronounced effects were observed with LDL modified by 15LO alone. Upregulation of CD36 was inversely correlated to the state of cell differentiation, as showed by lower response to enzLDL of the scavenger receptor-expressing MM6-sr and THP1 cells. Importantly, LDL modified by sPLA2 and 15LO did not efficiently induce upregulation CD36 in PPARγ-deficient macrophage-differentiated embryonic stem cells confirming a role of PPARγ in CD36 expression in cells stimulated with enzLDL. Our data show that LDL modified with physiologically relevant enzymes stimulates CD36 expression in non-differentiated monocytes and that this process involves PPARγ activation. These effects of enzLDL can be considered pro-atherogenic in the context of early atherosclerosis.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-24665 (URN)10.1016/j.bcp.2003.09.041 (DOI)6896 (Local ID)6896 (Archive number)6896 (OAI)
Available from: 2009-10-07 Created: 2009-10-07 Last updated: 2017-12-13
Buss, J. L., Neuzil, J. & Ponka, P. (2004). Oxidative stress mediates toxicity of pyridoxal isonicotinoyl hydrazone analogs. Archives of Biochemistry and Biophysics, 421(1)
Open this publication in new window or tab >>Oxidative stress mediates toxicity of pyridoxal isonicotinoyl hydrazone analogs
2004 (English)In: Archives of Biochemistry and Biophysics, ISSN 0003-9861, E-ISSN 1096-0384, Vol. 421, no 1Article in journal (Refereed) Published
Abstract [en]

Pyridoxal isonicotinoyl hydrazone (PIH) and many of its analogs are effective iron chelators in vivo and in vitro, and are of interest for the treatment of secondary iron overload. Because previous work has implicated the Fe3+-chelator complexes as a determinant of toxicity, the role of iron-based oxidative stress in the toxicity of PIH analogs was assessed. The Fe3+ complexes of PIH analogs were reduced by K562 cells and the physiological reductant, ascorbate. Depletion of the antioxidant, glutathione, sensitized Jurkat T lymphocytes to the toxicity of PIH analogs and their Fe 3+ complexes, and toxicity of the chelators increased with oxygen tension. Fe3+ complexes of pyridoxal benzoyl hydrazone (PBH) and salicyloyl isonicotinoyl hydrazone (SIH) caused lipid peroxidation and toxicity in K562 cells loaded with eicosapentenoic acid (EPA), a readily oxidized fatty acid, whereas Fe(PIH)2 did not. The lipophilic antioxidant, vitamin E, completely prevented both the toxicity and lipid peroxidation caused by Fe(PBH)2 in EPA-loaded cells, indicating a causal relationship between oxidative stress and toxicity. PBH also caused concomitant lipid peroxidation and toxicity in EPA-loaded cells, both of which were reversed as its concentration increased. In contrast, PIH was inactive, while SIH was equally toxic toward control and EPA-loaded cells, without causing lipid peroxidation, indicating a much smaller contribution of oxidative stress to the mechanism of toxicity of these analogs. In summary, PIH analogs and their Fe3+ complexes are redox active in the intracellular environment. The contribution of oxidative stress to the overall mechanism of toxicity varies across the series. © 2003 Elsevier Inc. All rights reserved.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-24641 (URN)10.1016/j.abb.2003.09.044 (DOI)6827 (Local ID)6827 (Archive number)6827 (OAI)
Available from: 2009-10-07 Created: 2009-10-07 Last updated: 2017-12-13
Neuzil, J., Tomasetti, M., Mellick, A. S., Alleva, R., Salvatore, B. A., Birringer, M. & Fariss, M. W. (2004). Vitamin E analogues: A new class of inducers of apoptosis with selective anti-cancer effects. Current Cancer Drug Targets, 4(4), 355-372
Open this publication in new window or tab >>Vitamin E analogues: A new class of inducers of apoptosis with selective anti-cancer effects
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2004 (English)In: Current Cancer Drug Targets, ISSN 1568-0096, E-ISSN 1873-5576, Vol. 4, no 4, p. 355-372Article in journal (Refereed) Published
Abstract [en]

In spite of unrelenting effort, the net incidence of neoplastic diseases appears not to have been curbed. While some types of cancer have been suppressed significantly, others are either stagnating or on the increase. Therefore, the need for a cure is imperative, in particularly a drug or combination of drugs that would be selective for malignant cells, i.e. with as low secondary toxicity as possible. Recent data strongly suggest that analogues of vitamin E, epitomised by the most studied α-tocopheryl succinate (α-TOS), may meet the need for the coveted drugs with a selective anti-neoplastic effect. The reasons for this optimism are reviewed in this article.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-24640 (URN)10.2174/1568009043332943 (DOI)6826 (Local ID)6826 (Archive number)6826 (OAI)
Available from: 2009-10-07 Created: 2009-10-07 Last updated: 2017-12-13
Tomasetti, M., Rippo, M., Alleva, R., Moretti, S., Andera, L., Neuzil, J. & Procopio, A. (2004). α-tocopheryl succinate and TRAIL selectively synergise in induction of apoptosis in human malignant mesothelioma cells. British Journal of Cancer, 90(8), 1644-1653
Open this publication in new window or tab >>α-tocopheryl succinate and TRAIL selectively synergise in induction of apoptosis in human malignant mesothelioma cells
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2004 (English)In: British Journal of Cancer, ISSN 0007-0920, E-ISSN 1532-1827, Vol. 90, no 8, p. 1644-1653Article in journal (Refereed) Published
Abstract [en]

Malignant mesothelioma (MM) is a fatal type of neoplasia with poor therapeutic prognosis, largely due to resistance to apoptosis. We investigated the apoptotic effect of α-tocopheryl succinate (α-TOS), a strong proapoptotic agent, in combination with the immunological apoptogen TNF-related apoptosis-inducing ligand (TRAIL) on both MM and nonmalignant mesothelial cells, since MM cells show low susceptibility to the clinically intriguing TRAIL. All MM cell lines tested were sensitive to α-TOS-induced apoptosis, and exerted high sensitivity to TRAIL in the presence of subapoptotic doses of the vitamin E analogue. Neither TRAIL or α-TOS alone or in combination caused apoptosis in nonmalignant mesothelial cells. Isobologram analysis of the cytotoxicity assays revealed a synergistic interaction between the two agents in MM cells and their antagonistic effect in nonmalignant mesothelial cells. TRAIL-induced apoptosis and its augmentation by α-TOS were inhibited by the caspase-8 inhibitor Z-IETD-FMK and the pan-caspase inhibitor Z-VAD-FMK. Activation of caspase-8 was required to induce apoptosis, which was amplified by α-TOS via cytochrome c release following Bid cleavage, with ensuing activation of caspase-9. Enhancement of TRAIL-induced apoptosis in MM cells by α-TOS was also associated with upregulation of the TRAIL cognate death receptors DR4 and DR5. Our results show that α-TOS and TRAIL act in synergism to kill MM cells via mitochondrial pathway, and are nontoxic to nonmalignant mesothelial cells. These findings are indicative of a novel strategy for treatment of thus far fatal MM.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-24639 (URN)10.1038/sj.bjc.6601707 (DOI)6825 (Local ID)6825 (Archive number)6825 (OAI)
Available from: 2009-10-07 Created: 2009-10-07 Last updated: 2017-12-13
Dalen, H. & Neuzil, J. (2003). a-tocopheryl succinate sensitises a T lymphoma cell line to TRAIL-induced apoptosis by suppressing NF-?B activation. British Journal of Cancer, 88(1), 153-158
Open this publication in new window or tab >>a-tocopheryl succinate sensitises a T lymphoma cell line to TRAIL-induced apoptosis by suppressing NF-?B activation
2003 (English)In: British Journal of Cancer, ISSN 0007-0920, E-ISSN 1532-1827, Vol. 88, no 1, p. 153-158Article in journal (Refereed) Published
Abstract [en]

Activation of nuclear factor-?B (NF-?B) can interfere with induction of apoptosis triggered by the tumour necrosis factor-related apoptosis-inducing ligand (TRAIL, Apo2L). Therefore, agents that suppress NF-?B activation may sensitise cells to TRAIL-dependent apoptosis. Exposure of Jurkat cells to TRAIL resulted in massive and saturable apoptosis induction, following an initial lag time. This lag was abolished by pretreatment of the cells with subapoptotic doses of a-tocopheryl succinate (a-TOS) or the proteasome inhibitor MGI32. Exposure of the cells to TRAIL led to a rapid, transient activation of NF-?B, a process that was suppressed by cell pretreatment with a-TOS or MGI32. Activation of NF-?B by TNF-a prior to TRAIL exposure increased resistance of the cells to TRAIL-mediated apoptosis. We conclude that a-TOS sensitises cells to TRAIL killing, at least in some cases, through inhibition of NF-?B activation. This further supports the possibility that this semisynthetic analogue of vitamin E is a potential adjuvant in cancer treatment, such as in the case of TRAIL-mediated inhibition of cancer.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-27722 (URN)10.1038/sj.bjc.6600683 (DOI)12460 (Local ID)12460 (Archive number)12460 (OAI)
Available from: 2009-10-08 Created: 2009-10-08 Last updated: 2017-12-13
Weber, A., Dalen, H., Andera, L., Nègre-Salvayre, A., Augé, N., Sticha, M., . . . Neuzil, J. (2003). Mitochondria play a central role in apoptosis induced by a-tocopheryl succinate, an agent with antineoplastic activity: Comparison with receptor-mediated pro-apoptotic signaling. Biochemistry, 42(14), 4277-4291
Open this publication in new window or tab >>Mitochondria play a central role in apoptosis induced by a-tocopheryl succinate, an agent with antineoplastic activity: Comparison with receptor-mediated pro-apoptotic signaling
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2003 (English)In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 42, no 14, p. 4277-4291Article in journal (Refereed) Published
Abstract [en]

a-Tocopheryl succinate (a-TOS) is a semisynthetic vitamin E analogue with high pro-apoptotic and anti-neoplastic activity [Weber, T et al. (2002) Clin. Cancer Res. 8, 863-869]. Previous studies suggested that it acts through destabilization of subcellular organelles, including mitochondria, but compelling evidence is missing. Cells treated with a-TOS showed altered mitochondrial structure, generation of free radicals, activation of the sphingomyelin cycle, relocalization of cytochrome c and Smac/Diablo, and activation of multiple caspases. A pan-caspase inhibitor suppressed caspase-3 and -6 activation and phosphatidyl serine externalization, but not decrease of mitochondrial membrane potential or generation of radicals. For a-TOS, but not Fas or TRAIL, apoptosis was suppressed by caspase-9 inhibition, while TRAIL- and Fas-resistant cells overexpressing cFLIP or CrmA were susceptible to a-TOS. The central role of mitochondria was confirmed by resistance of mtDNA-deficient cells to a-TOS, by regulation of a-TOS apoptosis by Bcl-2 family members, and by anti-apoptotic activity of mitochondrially targeted radical scavengers. Co-treatment with a-TOS and anti-Fas IgM showed their cooperative effect, probably by signaling via different, convergent pathways. These data provide an insight into the molecular mechanism, by which a-TOS kills malignant cells, and advocate its testing as a potential anticancer agent or adjuvant.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-27723 (URN)10.1021/bi020527j (DOI)12461 (Local ID)12461 (Archive number)12461 (OAI)
Available from: 2009-10-08 Created: 2009-10-08 Last updated: 2017-12-13
Terman, A., Dalen, H., Eaton, J. W., Neuzil, J. & Brunk, U. (2003). Mitochondrial recycling and aging of cardiac myocytes: The role of autophagocytosis. Experimental Gerontology, 38(8), 863-876
Open this publication in new window or tab >>Mitochondrial recycling and aging of cardiac myocytes: The role of autophagocytosis
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2003 (English)In: Experimental Gerontology, ISSN 0531-5565, E-ISSN 1873-6815, Vol. 38, no 8, p. 863-876Article in journal (Refereed) Published
Abstract [en]

The mechanisms of mitochondrial alterations in aged post-mitotic cells, including formation of so-called 'giant' mitochondria, are poorly understood. To test whether these large mitochondria might appear due to imperfect autophagic mitochondrial turnover, we inhibited autophagocytosis in cultured neonatal rat cardiac myocytes with 3-methyladenine. This resulted in abnormal accumulation of mitochondria within myocytes, loss of contractility, and reduced survival time in culture. Unlike normal aging, which is associated with slow accumulation of predominantly large defective mitochondria, pharmacological inhibition of autophagy caused only moderate accumulation of large (senescent-like) mitochondria but dramatically enhanced the numbers of small mitochondria, probably reflecting their normally more rapid turnover. Furthermore, the 3-methyladenine-induced accumulation of large mitochondria was irreversible, while small mitochondria gradually decreased in number after withdrawal of the drug. We, therefore, tentatively conclude that large mitochondria selectively accumulate in aging post-mitotic cells because they are poorly autophagocytosed. Mitochondrial enlargement may result from impaired fission, a possibility supported by depressed DNA synthesis in large mitochondria. Nevertheless, enlarged mitochondria retained immunoreactivity for cytochrome c oxidase subunit 1, implying that mitochondrial genes remain active in defective mitochondria. Our findings suggest that imperfect autophagic recycling of these critical organelles may underlie the progressive mitochondrial damage, which characterizes aging post-mitotic cells.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-27710 (URN)10.1016/S0531-5565(03)00114-1 (DOI)12448 (Local ID)12448 (Archive number)12448 (OAI)
Available from: 2009-10-08 Created: 2009-10-08 Last updated: 2017-12-13
Buss, J. L., Neuzil, J., Gellert, N., Weber, C. & Ponka, P. (2003). Pyridoxal isonicotinoyl hydrazone analogs induce apoptosis in hematopoietic cells due to their iron-chelating properties. Biochemical Pharmacology, 65(2), 161-172
Open this publication in new window or tab >>Pyridoxal isonicotinoyl hydrazone analogs induce apoptosis in hematopoietic cells due to their iron-chelating properties
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2003 (English)In: Biochemical Pharmacology, ISSN 0006-2952, E-ISSN 1356-1839, Vol. 65, no 2, p. 161-172Article in journal (Refereed) Published
Abstract [en]

Analogs of pyridoxal isonicotinoyl hydrazone (PIH) are of interest as iron chelators for the treatment of secondary iron overload and cancer. PIH, salicylaldehyde isonicotinoyl hydrazone (SIH), and 2-hydroxy-1-naphthylaldehyde isonicotinoyl hydrazone (NIH), the toxicity of which vary over two orders of magnitude, were selected for a study of their mechanisms of toxicity. PIH analogs and their iron complexes caused concentration- and time-dependent apoptosis in Jurkat T lymphocytes and K562 cells. Bcl-2 overexpression was partially anti-apoptotic, suggesting mitochondrial mediation of apoptosis. Since the pan-caspase inhibitor zVAD-fmk did not reduce lysosomal and mitochondrial destabilization, these events occur upstream of caspase activation. In contrast, phosphatidylserine externalization and the development of apoptotic morphology were inhibited significantly, indicating the role of caspases in mediating these later events. Since overexpression of CrmA had no effect on apoptosis, caspase-8 is not likely involved. Fe3+ complexes of SIH and NIH, which accumulated in 59Fe-labeled mouse reticulocytes during incubation with the chelators, also caused apoptosis. BSA, which promotes release of the complexes from cells, reduced the toxicity of SIH and NIH, suggesting that the induction of apoptosis by PIH analogs involves toxic effects mediated by their Fe3+ complexes. Moreover, analogs of these agents lacking the iron-chelating moiety were non-toxic. ⌐ 2002 Published by Elsevier Science Inc.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-27895 (URN)10.1016/S0006-2952(02)01512-5 (DOI)12655 (Local ID)12655 (Archive number)12655 (OAI)
Available from: 2009-10-08 Created: 2009-10-08 Last updated: 2017-12-13
Birringer, M., EyTina, J. H., Salvatore, B. A. & Neuzil, J. (2003). Vitamin E analogues as inducers of apoptosis: Structure-function relation. British Journal of Cancer, 88(12), 1948-1955
Open this publication in new window or tab >>Vitamin E analogues as inducers of apoptosis: Structure-function relation
2003 (English)In: British Journal of Cancer, ISSN 0007-0920, E-ISSN 1532-1827, Vol. 88, no 12, p. 1948-1955Article in journal (Refereed) Published
Abstract [en]

Recent results show that a-tocopheryl succinate (a-TOS) is a proapoptotic agent with antineoplastic activity. As modifications of the vitamin E (VE) molecule may affect its apoptogenic activity, we tested a number of newly synthesised VE analogues using malignant cell lines. Analogues of a-TOS with lower number of methyl substitutions on the aromatic ring were less active than a-TOS. Replacement of the succinyl group with a maleyl group greatly enhanced the activity, while it was lower for the glutaryl esters. Methylation of the free succinyl carboxyl group on a-TOS and d-TOS completely prevented the apoptogenic activity of the parent compounds. Both Trolox and its succinylated derivative were inactive. a-tocotrienol (a-T3 H) failed to induce apoptosis, while ?-T3 H was apoptogenic, and more so when succinylated. Shortening the aliphatic side chain of ?-T3 by one isoprenyl unit increased its activity. Neither phytyl nor oleyl succinate caused apoptosis. These findings show that modifications of different functional moieties of the VE molecule can enhance apoptogenic activity. It is hoped that these observations will lead to the synthesis of analogues with even higher apoptogenic and, consequently, antineoplastic efficacy.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-27896 (URN)10.1038/sj.bjc.6600981 (DOI)12656 (Local ID)12656 (Archive number)12656 (OAI)
Available from: 2009-10-08 Created: 2009-10-08 Last updated: 2017-12-13
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