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Characterization of Cell Death Pathways after Allergen Challenge and Viral-induced Asthma Exacerbation

Cerps, Samuel (2016) MOBT01 20152
Degree Projects in Molecular Biology
Abstract
Introduction: Asthma is a chronic inflammatory lung disease, which are associated with airflow obstructions. Asthma exacerbations are commonly caused by respiratory viral infections and lack efficient treatment options. Rhinovirus-induced exacerbations have been characterized by occurrence of necrotic cells, which may cause pathogenic effects. We hypothesized that house dust mite (HDM)-induced allergic inflammation primes the airway to aggravated responses to viral infection involving activation of cell death pathways: apoptosis, necrosis and necroptosis.

Methods: The bronchial epithelial cell line BEAS-B was used to study apoptosis in vitro. Wild type (C57BL/6), Myd88-/-, NfκB-/-, IRF3-/- and IFN-β-/- mice were administered saline or... (More)
Introduction: Asthma is a chronic inflammatory lung disease, which are associated with airflow obstructions. Asthma exacerbations are commonly caused by respiratory viral infections and lack efficient treatment options. Rhinovirus-induced exacerbations have been characterized by occurrence of necrotic cells, which may cause pathogenic effects. We hypothesized that house dust mite (HDM)-induced allergic inflammation primes the airway to aggravated responses to viral infection involving activation of cell death pathways: apoptosis, necrosis and necroptosis.

Methods: The bronchial epithelial cell line BEAS-B was used to study apoptosis in vitro. Wild type (C57BL/6), Myd88-/-, NfκB-/-, IRF3-/- and IFN-β-/- mice were administered saline or HDM intranasally for three weeks to induce experimental asthma in vivo. Double stranded (ds) RNA was given for additional 3 days to mimic viral infection and induce exacerbation. Lung tissue apoptosis was evaluated using Western blot for cleaved caspase-3 and PARP together with TUNEL-staining on lung sections. Lung expression of RIP3 was determined to explore occurrence of necroptosis (regulated necrosis).

Results: TUNEL analysis revealed that HDM mainly induced cell death in recruited inflammatory cells which were further aggravated in IFN-β-/- mice but absent in NfκB-/- and Myd88-/- mice. Exacerbation evoked by PIC aggravated the HDM-induced lung inflammation and increased the apoptotic markers cleaved caspases-3 and PARP compared to control (p<0.05). The necrosis marker RIP3 was induced by PIC alone in animals without prior HDM challenge but was not further increased at exacerbation. Furthermore RIP3 followed the same pattern as the pan necrotic marker lactate dehydrogenase levels in bronchoalveolar lavage fluid. We could confirm that PIC induced apoptosis in BEAS-2B cells in vitro.

Conclusion: A mixture of both apoptotic and necrotic pathways are activated during exacerbation. This suggests that cell death pathways are involved in asthma exacerbations and may thus be molecular targets for therapeutic intervention. (Less)
Popular Abstract
Periods of Asthma Attacks Lead to Increased Cell Death

There is a coordinated balance between new and dying cells in the body controlled by different signals. The cells in the body can either die in a silent manner or burst in an inflammatory way that can be destructive for the organism. There are many diseases that are caused by problems in the cell death machinery involving both the silent and non-silent forms of death. Asthma is a chronic disease causing the lungs to react more easily to things that are not necessary harmful. This reaction leads to obstruction of the airways, which makes it harder to breath. Sometimes asthmatics have periods called exacerbations, during which they have more frequent asthma attack. The cold virus... (More)
Periods of Asthma Attacks Lead to Increased Cell Death

There is a coordinated balance between new and dying cells in the body controlled by different signals. The cells in the body can either die in a silent manner or burst in an inflammatory way that can be destructive for the organism. There are many diseases that are caused by problems in the cell death machinery involving both the silent and non-silent forms of death. Asthma is a chronic disease causing the lungs to react more easily to things that are not necessary harmful. This reaction leads to obstruction of the airways, which makes it harder to breath. Sometimes asthmatics have periods called exacerbations, during which they have more frequent asthma attack. The cold virus commonly causes these periods because asthmatics have deficient signals involved in fighting the virus. It is not known if the signals present in an asthmatic lung contribute to cell death or if cell death is actually one of the mechanisms that cause the disease, especially exacerbations

A mouse model helps us understand the impact of cell death in the context of asthma
In an experiment, both the silent and the non-silent form of cell death were studied by looking at specific markers of cell death in an asthma and exacerbation model. A viral mimic that was used instead of a real virus, induced both the silent and the non-silent form of cell death. During exacerbation there was an increase in the expression of the markers of the silent form of cell death compared to non-asthmatic mice that received the viral mimic. It was found that an important signal involved in fighting viral infections was also needed for survival of immune cells in asthmatic mice.

These findings suggest that an asthma exacerbation caused by viral infection leads to an increased cell death compared to what we see in a healthy person just having a cold. If this increased death is manifested as death of the protective outer cell layer in the lungs together with impaired survival of cells fighting the virus, a consequence would be that viruses could reach further down in the lungs thereby causing an asthma attack. Therefore the cell death machinery could be an important drug target for treating asthma exacerbations.

Advisors: Lena Uller and Hamid Akbarshahi
Master´s Degree Project 60 credits in Molecular Biology, 2016
Department of Biology, Lund University. (Less)
Please use this url to cite or link to this publication:
author
Cerps, Samuel
supervisor
organization
course
MOBT01 20152
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
8891276
date added to LUP
2016-09-09 11:37:07
date last changed
2016-09-09 11:37:07
@misc{8891276,
  abstract     = {Introduction: Asthma is a chronic inflammatory lung disease, which are associated with airflow obstructions. Asthma exacerbations are commonly caused by respiratory viral infections and lack efficient treatment options. Rhinovirus-induced exacerbations have been characterized by occurrence of necrotic cells, which may cause pathogenic effects. We hypothesized that house dust mite (HDM)-induced allergic inflammation primes the airway to aggravated responses to viral infection involving activation of cell death pathways: apoptosis, necrosis and necroptosis.

Methods: The bronchial epithelial cell line BEAS-B was used to study apoptosis in vitro. Wild type (C57BL/6), Myd88-/-, NfκB-/-, IRF3-/- and IFN-β-/- mice were administered saline or HDM intranasally for three weeks to induce experimental asthma in vivo. Double stranded (ds) RNA was given for additional 3 days to mimic viral infection and induce exacerbation. Lung tissue apoptosis was evaluated using Western blot for cleaved caspase-3 and PARP together with TUNEL-staining on lung sections. Lung expression of RIP3 was determined to explore occurrence of necroptosis (regulated necrosis).

Results: TUNEL analysis revealed that HDM mainly induced cell death in recruited inflammatory cells which were further aggravated in IFN-β-/- mice but absent in NfκB-/- and Myd88-/- mice. Exacerbation evoked by PIC aggravated the HDM-induced lung inflammation and increased the apoptotic markers cleaved caspases-3 and PARP compared to control (p<0.05). The necrosis marker RIP3 was induced by PIC alone in animals without prior HDM challenge but was not further increased at exacerbation. Furthermore RIP3 followed the same pattern as the pan necrotic marker lactate dehydrogenase levels in bronchoalveolar lavage fluid. We could confirm that PIC induced apoptosis in BEAS-2B cells in vitro.

Conclusion: A mixture of both apoptotic and necrotic pathways are activated during exacerbation. This suggests that cell death pathways are involved in asthma exacerbations and may thus be molecular targets for therapeutic intervention.},
  author       = {Cerps, Samuel},
  language     = {eng},
  note         = {Student Paper},
  title        = {Characterization of Cell Death Pathways after Allergen Challenge and Viral-induced Asthma Exacerbation},
  year         = {2016},
}