Advanced

Immunohistochemical characterization of the innate and adaptive immune response during influenza a infections in human infants and mice

Lidén, Evelina (2017) MOBT01 20162
Degree Projects in Molecular Biology
Abstract
Background. Lower tract respiratory infections are one of the most prevalent health problems worldwide. Influenza A is a main contributor to these infections, and infants are one of the risk groups for infection and developing serious lower respiratory tract complications. Better treatment for infants is needed, but little is known about the immune response that occurs in the lungs during infection. The aim of this study was to establish immunohistochemistry protocols and use them to characterize immune cell profiles in influenza A infected human infant and mouse lungs.

Methods. Twenty-two antibodies were tested on influenza A infected human infant lungs, and fourteen antibodies were tested on influenza A infected mice lungs. Optimal... (More)
Background. Lower tract respiratory infections are one of the most prevalent health problems worldwide. Influenza A is a main contributor to these infections, and infants are one of the risk groups for infection and developing serious lower respiratory tract complications. Better treatment for infants is needed, but little is known about the immune response that occurs in the lungs during infection. The aim of this study was to establish immunohistochemistry protocols and use them to characterize immune cell profiles in influenza A infected human infant and mouse lungs.

Methods. Twenty-two antibodies were tested on influenza A infected human infant lungs, and fourteen antibodies were tested on influenza A infected mice lungs. Optimal staining was achieved by optimizing primary antibody concentration, chromogen detection systems, and pretreatment methods. Antibody staining quality was evaluated and the antibody protocols that gave adequate staining results were used for subsequent immunohistochemistry.

Results. Optimal immunohistochemistry protocols were achieved for seventeen of the twenty-two antibodies tested on infant lungs, and seven of the fourteen antibodies tested on mice lungs. In influenza A infected mice lungs an increase was seen in B cells and CD3+ T cells, while macrophages and eosinophils were unchanged. There were several distributional differences from healthy mice lungs such as the formation of numerous B cell aggregates, an increase in CD3+ T cells surrounding infected airways, and macrophage redistribution from the alveolar parenchyma to infected airways. The influenza A infected infant lung showed a predominance of CD8+ T cells over CD4+ T cells, extensive distribution of macrophages in the alveolar parenchyma, very few dendritic cells, and a very high density of neutrophils. In addition, more T cells than B cells were present, natural killer cells were found throughout the lung, and a high amount of mast cells and eosinophils was observed.

Conclusion. Protocols were established for all main immune cells and lung structures in influenza A infected human infant lungs, however further protocol optimization is needed to get a full picture of the immune reaction in mice. The immune cell protocols optimized for human infants can be applied to a larger study, and include infant lungs with other viral infections such as respiratory syncytial virus and adenovirus.
Keywords. Immunohistochemistry, protocol, paraffin, immunology, innate immune system, adaptive immune system, Influenza A, human, infant, mouse. (Less)
Popular Abstract
Microscopic Wars - Influenza A in Infants-

Influenza A, also known as the flu, is a virus that most of us have had a close encounter with. For most people, falling ill with the flu just adds up to a few inconvenient days stuck in bed, but for some the flu can lead to serious lower respiratory tract infection (LRTI) complications – and nobody really knows how to treat it. LRTIs are among the most prevalent public health problems worldwide, causing more disease and death that any other infectious disease, yet there are currently few effective treatments for LRTIs caused by viruses such as influenza A.

Influenza A infections in infants
Infants are especially susceptible to influenza infections, averaging 5-6 infections per year, and... (More)
Microscopic Wars - Influenza A in Infants-

Influenza A, also known as the flu, is a virus that most of us have had a close encounter with. For most people, falling ill with the flu just adds up to a few inconvenient days stuck in bed, but for some the flu can lead to serious lower respiratory tract infection (LRTI) complications – and nobody really knows how to treat it. LRTIs are among the most prevalent public health problems worldwide, causing more disease and death that any other infectious disease, yet there are currently few effective treatments for LRTIs caused by viruses such as influenza A.

Influenza A infections in infants
Infants are especially susceptible to influenza infections, averaging 5-6 infections per year, and are at a significantly higher risk for serious complications from influenza infections compared to older children and adults. There is currently a need for better treatment of infants with LRTI complications because of influenza A, but to do so more research needs to be done on how the infant immune system reacts during the viral infection.

One of the biggest problems with developing better treatment is that much of our knowledge about influenza comes from research done on mice or assumptions based on the adult immune system, however this information is not enough as the infant immune system is vastly different from both mice and adults. This is not to say that mice are not valuable research components, in fact they are excellent tools and experiments can be manipulated to better mimic influenza infections in infants.

Characterizing the immune system response
In this study we set out to develop a way to characterize the immune system reaction to influenza A in infants and mice by optimizing immunohistochemistry protocols for antibodies that target immune cells of the innate and adaptive immune system, and using it to piece together what the immune system is doing.

To approach this, a repertoire of antibodies that covered all the key immune cells and lung structures were chosen to be tested using immunohistochemistry. The results were used to characterize the immune response and provided quantitative data for investigating the immune response in infants infected with influenza A, and a good starting point for doing the same with mice models.

For the future
The ability to characterize and analyze the complete immune response to influenza A has the potential to give insight to how the infant immune system reacts to the virus, with hopes that it can lead to a better understanding and treatment of LRTIs due to influenza A in infants.

Main supervisor: Michiko Mori
Co-supervisors: Caroline Sandén, Jonas Erjefält
Master’s Pegree Project 60 hp in Molecular Biology, 2017
Department of Biology, Lund University (Less)
Please use this url to cite or link to this publication:
author
Lidén, Evelina
supervisor
organization
course
MOBT01 20162
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
8906349
date added to LUP
2017-05-02 16:20:24
date last changed
2017-05-03 08:23:00
@misc{8906349,
  abstract     = {Background. Lower tract respiratory infections are one of the most prevalent health problems worldwide. Influenza A is a main contributor to these infections, and infants are one of the risk groups for infection and developing serious lower respiratory tract complications. Better treatment for infants is needed, but little is known about the immune response that occurs in the lungs during infection. The aim of this study was to establish immunohistochemistry protocols and use them to characterize immune cell profiles in influenza A infected human infant and mouse lungs. 

Methods. Twenty-two antibodies were tested on influenza A infected human infant lungs, and fourteen antibodies were tested on influenza A infected mice lungs. Optimal staining was achieved by optimizing primary antibody concentration, chromogen detection systems, and pretreatment methods. Antibody staining quality was evaluated and the antibody protocols that gave adequate staining results were used for subsequent immunohistochemistry. 

Results. Optimal immunohistochemistry protocols were achieved for seventeen of the twenty-two antibodies tested on infant lungs, and seven of the fourteen antibodies tested on mice lungs. In influenza A infected mice lungs an increase was seen in B cells and CD3+ T cells, while macrophages and eosinophils were unchanged. There were several distributional differences from healthy mice lungs such as the formation of numerous B cell aggregates, an increase in CD3+ T cells surrounding infected airways, and macrophage redistribution from the alveolar parenchyma to infected airways. The influenza A infected infant lung showed a predominance of CD8+ T cells over CD4+ T cells, extensive distribution of macrophages in the alveolar parenchyma, very few dendritic cells, and a very high density of neutrophils. In addition, more T cells than B cells were present, natural killer cells were found throughout the lung, and a high amount of mast cells and eosinophils was observed. 
 
Conclusion. Protocols were established for all main immune cells and lung structures in influenza A infected human infant lungs, however further protocol optimization is needed to get a full picture of the immune reaction in mice. The immune cell protocols optimized for human infants can be applied to a larger study, and include infant lungs with other viral infections such as respiratory syncytial virus and adenovirus. 
Keywords. Immunohistochemistry, protocol, paraffin, immunology, innate immune system, adaptive immune system, Influenza A, human, infant, mouse.},
  author       = {Lidén, Evelina},
  language     = {eng},
  note         = {Student Paper},
  title        = {Immunohistochemical characterization of the innate and adaptive immune response during influenza a infections in human infants and mice},
  year         = {2017},
}