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Evaluation of Caspase Activation to Assess Innate Immune Cell Death
Published on: January 20, 2023
Streptococcus pneumoniae R6x induced p38 MAPK and JNK-mediated caspase-dependent apoptosis in human endothelial cells
Philippe Dje N'Guessan1, Bernd Schmeck, Abena Ayim
1Department of Internal Medicine/Infectious Diseases, Charité - University Medicine Berlin, Germany.
Abstract:
Streptococcus pneumoniae is the major pathogen of community-acquired pneumonia and a common cause of otitis, meningitis and sepsis. During pneumococci infection accompanied with bacterial invasion and hematogenous spreading, the endothelium is directly targeted by pneumococci and their virulence factors. Therefore, we tested the hypothesis that pneumococci induced endothelial apoptosis. Unencapsulated R6x pneumococci strongly induced apoptosis of human endothelial cells both from lung microvasculature and umbilical vein, whereas an encapsulated strain D39 mainly led to necrotic cell death. Deletion of the gene coding for pneumolysin reduced pneumococci-induced apoptosis in HUVEC. Furthermore, N-acetyl-L-cysteine, an antioxidant thiol, significantly reduced apoptosis caused by R6x, and LDH release induced by D39, pointing to a role for reactive oxygen species in the pathogenesis. Apoptotic cells showed increased cleavage and activity of caspases 6 and 9 but only late activation of caspase 3. Programmed cell death could be strongly reduced by pan-caspase inhibitor zVAD. Reduced levels of Bcl2 and cytosolic increase of apoptosis-inducing factor in pneumococci-infected cells implicated involvement of mitochondrial death pathways. Caspase activation and apoptosis were abolished by cAMP elevation. Moreover, p38 mitogen-activated protein kinase and c-Jun NH(2)-terminal kinase were activated in pneumococci-infected cells and inhibitors of both kinases strongly reduced pneumococci-induced caspase activation and apoptosis. Hence, kinase- and caspase-dependence of pneumococci-induced endothelial apoptosis may bear relevance to novel therapeutic approaches to pneumococci-related disease.
Insights
Streptococcus pneumoniae infection triggers endothelial apoptosis, particularly by unencapsulated strains. This programmed cell death involves caspases and kinases, suggesting new therapeutic targets for pneumococcal diseases.
Area of Science:
- Cell Biology
- Microbiology
- Pathogenesis
Background:
- Streptococcus pneumoniae causes pneumonia, meningitis, and sepsis.
- Endothelial cells are targeted during pneumococcal infections.
- The role of pneumococci in inducing endothelial cell death requires clarification.
Purpose of the Study:
- To investigate whether Streptococcus pneumoniae induces endothelial apoptosis.
- To identify the mechanisms and pathways involved in pneumococci-induced endothelial cell death.
- To explore potential therapeutic targets for pneumococcal diseases.
Main Methods:
- Culturing human endothelial cells (lung microvasculature and umbilical vein).
- Comparing apoptosis induced by encapsulated (D39) and unencapsulated (R6x) S. pneumoniae strains.
- Assessing the role of pneumolysin, reactive oxygen species (ROS), caspases, Bcl2, apoptosis-inducing factor, cAMP, and MAP kinases (p38, JNK).
- Utilizing inhibitors (N-acetyl-L-cysteine, zVAD, kinase inhibitors) and cAMP elevation.
Main Results:
- Unencapsulated R6x induced significant endothelial apoptosis, while encapsulated D39 caused necrosis.
- Pneumolysin contributed to apoptosis; N-acetyl-L-cysteine reduced apoptosis and LDH release, indicating ROS involvement.
- Apoptosis involved caspase-6, -9, and late caspase-3 activation, mitochondrial pathways, and activation of p38 and JNK kinases.
- Caspase activation and apoptosis were inhibited by cAMP elevation and specific kinase inhibitors.
Conclusions:
- Streptococcus pneumoniae induces endothelial apoptosis through caspase- and kinase-dependent pathways.
- Reactive oxygen species and mitochondrial pathways are implicated in pneumococci-induced endothelial cell death.
- Targeting these pathways, particularly kinase and caspase activation, may offer novel therapeutic strategies for pneumococcal infections.
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