Jeger 2013 Abstracts of ESICM LIVES 2013: Difference between revisions
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{{Publication | {{Publication | ||
|title=Jeger V, Steinmann M, Takala J, Jakob SM, Djafarzadeh S | |title=Jeger V, Steinmann M, Takala J, Jakob SM, Djafarzadeh S (2013) TLR-2-induced mitochondrial dysfunction in cultured human hepatocytes. Abstracts of ESICM LIVES 2013 | ||
|authors=Jeger V, Steinmann M, Takala J, Jakob SM, Djafarzadeh S | |info=[http://www.zb.unibe.ch/download/eldiss/13steinmann_m.pdf PDF] | ||
|authors=Jeger V, Steinmann M, Takala J, Jakob SM, Djafarzadeh S | |||
|year=2013 | |year=2013 | ||
|journal=Abstracts of ESICM LIVES 2013 | |journal=Abstracts of ESICM LIVES 2013 | ||
|abstract= | |abstract=Hepatic mitochondrial function may be impaired in sepsis, and it has | ||
been shown that Toll-like receptors TLR-3 and TLR-4 can be involved. TLR-2 is stimulated | been shown that Toll-like receptors TLR-3 and TLR-4 can be involved. TLR-2 is stimulated by Gram positive bacteria (e.g. Listeria monocytogenes). Its stimulation is involved in cytokine upregulation (via NF-jB) and mitochondrial network fragmentation, which is associated with apoptosis. | ||
by Gram positive bacteria (e.g. Listeria monocytogenes). Its stimulation is involved in | |||
cytokine upregulation (via NF-jB) and mitochondrial network fragmentation, which is | |||
associated with apoptosis. | |||
To evaluate whether TLR-2 stimulation is implicated in hepatic mitochondrial | |||
dysfunction and apoptosis. | dysfunction and apoptosis. | ||
Human hepatocellular carcinoma cell line (HepG2) was cultured and incubated | |||
with placebo or heat-killed Listeria monocytogenes (HKLM) at concentrations of 107 | with placebo or heat-killed Listeria monocytogenes (HKLM) at concentrations of 107 HKLM/ml and 108 HKLM/ml for 1 and 24 h (n C 10). Mitochondrial respiration was measured by high-resolution respirometry (Oroboros Instruments, Innsbruck, Austria). For Western blot analysis, HepG2 cells were exposed to HKLM at concentrations of 107 HKLM/ml and 108 HKLM/ml for 4, 8, 24 and 72 h. Caspase-3 and actin protein levels were determined by sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. Statistics: paired sample t-test. | ||
HKLM/ml and 108 HKLM/ml for 1 and 24 h (n C 10). Mitochondrial respiration was | |||
measured by high-resolution respirometry (Oroboros Instruments, Innsbruck, Austria). For | |||
Western blot analysis, HepG2 cells were exposed to HKLM at concentrations of 107 | |||
HKLM/ml and 108 HKLM/ml for 4, 8, 24 and 72 h. Caspase-3 and actin protein levels were | |||
determined by sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) and | |||
Western blotting. Statistics: paired sample t-test. | |||
24 h of incubation with HKLM at a concentration of 107 HKLM/ml induced a | |||
significant reduction in complex IV-dependent mitochondrial respiration (100 ± 31 in | significant reduction in complex IV-dependent mitochondrial respiration (100 ± 31 in unstimulated vs. 84 ± 20 pmol/[s million cells] in stimulated cells; p = 0.048). Complex I-dependent (67 ± 24 in unstimulated vs. 59 ± 15 pmol/[s million cells] in stimulated cells; p = 0.15) and complex II-dependent respiration (100 ± 28 in unstimulated vs. 92 ± 23 pmol/[s million cells] in stimulated cells; ''p'' = 0.18) were not significantly affected. | ||
unstimulated vs. 84 ± 20 pmol/[s million cells] in stimulated cells; p = 0.048). Complex | |||
I-dependent (67 ± 24 in unstimulated vs. 59 ± 15 pmol/[s million cells] in stimulated | |||
cells; p = 0.15) and complex II-dependent respiration (100 ± 28 in unstimulated vs. | |||
92 ± 23 pmol/[s million cells] in stimulated cells; p = 0.18) were not significantly affected. | |||
Incubation of cells with HKLM (108 HKLM/ml for 24 h) and 1 h incubations at both | Incubation of cells with HKLM (108 HKLM/ml for 24 h) and 1 h incubations at both | ||
concentrations did not affect maximal mitochondrial respiration (Fig. 1). Active caspase-3 | concentrations did not affect maximal mitochondrial respiration (Fig. 1). Active caspase-3 protein levels were not increased under any condition. | ||
protein levels were not increased under any condition. | |||
In this study, we demonstrate that in cultured human HepG2 cells, stimulation of TLR-2 with HKLM for 24 h induces a reduction in maximal mitochondrial | |||
stimulation of TLR-2 with HKLM for 24 h induces a reduction in maximal mitochondrial | oxygen consumption of complex IV in a time- and dose-dependent manner. We observed previously reductions of complex IV-dependent respiration after TLR-3 and TLR-4 stimulation [1]. These results suggest that TLR-2, -3 and -4 signaling use similar mechanisms (e.g. mitochondrial permeability transition pore opening) to induce changes in mitochondrial respiration. HKLM did not induce any signs of early apoptosis. The clinical relevance and the pathomechanism of our findings remain to be evaluated. | ||
oxygen consumption of complex IV in a time- and dose-dependent manner. We observed | |keywords=Sepsis, Mitochondria, Cytokines, Mitochondrial Respiration, Caspase- 3, Actin, Western blot, High-resolution respirometry, Toll- like receptor 2, ''Listeria monocytogenes'', HepG2 | ||
previously reductions of complex IV-dependent respiration after TLR-3 and TLR-4 stimulation | |mipnetlab=CH Bern Djafarzadeh S | ||
[1]. These results suggest that TLR-2, -3 and -4 signaling use similar mechanisms | |||
(e.g. mitochondrial permeability transition pore opening) to induce changes in mitochondrial | |||
respiration. HKLM did not induce any signs of early apoptosis. The clinical relevance | |||
and the pathomechanism of our findings remain to be evaluated. | |||
}} | }} | ||
{{Labeling | {{Labeling | ||
|area=Respiration | |||
|organism=Human | |||
|tissues=Liver | |||
|preparations=Permeabilized cells | |||
|pathways=N, S, CIV | |||
|instruments=Oxygraph-2k | |instruments=Oxygraph-2k | ||
}} | }} | ||
Latest revision as of 16:29, 27 March 2018
| Jeger V, Steinmann M, Takala J, Jakob SM, Djafarzadeh S (2013) TLR-2-induced mitochondrial dysfunction in cultured human hepatocytes. Abstracts of ESICM LIVES 2013 |
Jeger V, Steinmann M, Takala J, Jakob SM, Djafarzadeh S (2013) Abstracts of ESICM LIVES 2013
Abstract: Hepatic mitochondrial function may be impaired in sepsis, and it has been shown that Toll-like receptors TLR-3 and TLR-4 can be involved. TLR-2 is stimulated by Gram positive bacteria (e.g. Listeria monocytogenes). Its stimulation is involved in cytokine upregulation (via NF-jB) and mitochondrial network fragmentation, which is associated with apoptosis.
To evaluate whether TLR-2 stimulation is implicated in hepatic mitochondrial dysfunction and apoptosis.
Human hepatocellular carcinoma cell line (HepG2) was cultured and incubated with placebo or heat-killed Listeria monocytogenes (HKLM) at concentrations of 107 HKLM/ml and 108 HKLM/ml for 1 and 24 h (n C 10). Mitochondrial respiration was measured by high-resolution respirometry (Oroboros Instruments, Innsbruck, Austria). For Western blot analysis, HepG2 cells were exposed to HKLM at concentrations of 107 HKLM/ml and 108 HKLM/ml for 4, 8, 24 and 72 h. Caspase-3 and actin protein levels were determined by sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. Statistics: paired sample t-test.
24 h of incubation with HKLM at a concentration of 107 HKLM/ml induced a significant reduction in complex IV-dependent mitochondrial respiration (100 ± 31 in unstimulated vs. 84 ± 20 pmol/[s million cells] in stimulated cells; p = 0.048). Complex I-dependent (67 ± 24 in unstimulated vs. 59 ± 15 pmol/[s million cells] in stimulated cells; p = 0.15) and complex II-dependent respiration (100 ± 28 in unstimulated vs. 92 ± 23 pmol/[s million cells] in stimulated cells; p = 0.18) were not significantly affected. Incubation of cells with HKLM (108 HKLM/ml for 24 h) and 1 h incubations at both concentrations did not affect maximal mitochondrial respiration (Fig. 1). Active caspase-3 protein levels were not increased under any condition.
In this study, we demonstrate that in cultured human HepG2 cells, stimulation of TLR-2 with HKLM for 24 h induces a reduction in maximal mitochondrial oxygen consumption of complex IV in a time- and dose-dependent manner. We observed previously reductions of complex IV-dependent respiration after TLR-3 and TLR-4 stimulation [1]. These results suggest that TLR-2, -3 and -4 signaling use similar mechanisms (e.g. mitochondrial permeability transition pore opening) to induce changes in mitochondrial respiration. HKLM did not induce any signs of early apoptosis. The clinical relevance and the pathomechanism of our findings remain to be evaluated. • Keywords: Sepsis, Mitochondria, Cytokines, Mitochondrial Respiration, Caspase- 3, Actin, Western blot, High-resolution respirometry, Toll- like receptor 2, Listeria monocytogenes, HepG2
• O2k-Network Lab: CH Bern Djafarzadeh S
Labels: MiParea: Respiration
Organism: Human
Tissue;cell: Liver
Preparation: Permeabilized cells
Pathway: N, S, CIV HRR: Oxygraph-2k
