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Host-pathogen interactions in chronic infections and the development of novel therapies

Betina, Nawel LU (2017) KEMT30 20171
Department of Chemistry
Abstract
Wound healing is a process that ensures regeneration and repair of damaged tissue. This process can be interrupted, leading to chronic wounds, and one of the factors that can interrupt it is infection. The bacterium Pseudomonas aeruginosa is prevalent in chronic wounds, and as it is antibiotic resistant it makes chronic wounds infected with this pathogen a challenging clinical problem.
For a better understanding of causes behind wound healing failure when a wound is infected by P. aeruginosa we studied elastase, which is one of several virulence factors of P. aeruginosa. This protein is secreted by most strains of this species and is characterized as zinc metalloprotease. The present study relies on the isolation and purification of... (More)
Wound healing is a process that ensures regeneration and repair of damaged tissue. This process can be interrupted, leading to chronic wounds, and one of the factors that can interrupt it is infection. The bacterium Pseudomonas aeruginosa is prevalent in chronic wounds, and as it is antibiotic resistant it makes chronic wounds infected with this pathogen a challenging clinical problem.
For a better understanding of causes behind wound healing failure when a wound is infected by P. aeruginosa we studied elastase, which is one of several virulence factors of P. aeruginosa. This protein is secreted by most strains of this species and is characterized as zinc metalloprotease. The present study relies on the isolation and purification of elastase for investigations of a potential difference in the enzymatic activity of elastase between four different strains of P. aeruginosa, the PAO1 (wild type strain), PAOA1 (elastase A mutant), PAOB1 (elastase B mutant), and PAOA1B1 (elastase A and B double mutant). Also, to understand the interplay between elastase and the host by clarifying the interaction between elastase and both the immune and the coagulation systems.
Results of the purity analysis, using tricine gels together with results of gelatin zymography show that the purification of active elastase B using fast protein liquid chromatography was successful. Our results from an azocasein assay together with the gelatin zymography of both P. aeruginosa conditioned medium and purified elastase show that elastase B dominates the elastolytic activity of P. aeruginosa. Interestingly, results of this study disagree with the hypothesis that elastase A can enhance the virulence of elastase B. Contrary, the present data suggest that elastase A represses virulence of both elastase B and the toxin pyocyanin. (Less)
Popular Abstract
Skin is an organ, which acts as a physical barrier between the body and the environment. It provides protection against pathogen invasion by regulating skin immune system. A skin wound can be defined as any damage or disruption of the normal anatomical structure and function of the skin, and it can be classified into acute wounds that heal in an ordered pathway within a time frame of 30 days, and chronic wounds which take longer time to heal or even fail the healing process.
The combination between the high prevalence of chronic wounds, their treatment costs, frequency of the bacterium Pseudomonas aeruginosa in chronic wounds, and its resistance to antibiotics, makes wounds infected with this bacterium a real issue for both patients and... (More)
Skin is an organ, which acts as a physical barrier between the body and the environment. It provides protection against pathogen invasion by regulating skin immune system. A skin wound can be defined as any damage or disruption of the normal anatomical structure and function of the skin, and it can be classified into acute wounds that heal in an ordered pathway within a time frame of 30 days, and chronic wounds which take longer time to heal or even fail the healing process.
The combination between the high prevalence of chronic wounds, their treatment costs, frequency of the bacterium Pseudomonas aeruginosa in chronic wounds, and its resistance to antibiotics, makes wounds infected with this bacterium a real issue for both patients and society. For this reason elastase, which is one of the most prominent protein degrading enzyme in disease produced by P. aeruginosa, is studied in this project. A better understanding of its degrading activity might help in finding new treatments of non-healing wounds.
This study aims at isolating and purifying elastase, and then comparing the protein degrading activity of four different strains of P. aeruginosa: normal strain, an elastase A deficient mutant, an elastase B deficient mutant, and a mutant lacking both elastase A and B. A further objective is to elucidate and understand the role of elastase in interactions between the bacterium and the host.
The results of purity assays combined with enzymatic activity assays confirm the successful isolation of purified active elastase B. In addition, our results show that elastase B dominates the elastase protein degrading activity of P. aeruginosa. Contradictory to previous studies, results reveal that, elastase A represses the activity of elastase B and the production of pyocyanin, which is a toxin excreted by the bacterium. (Less)
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author
Betina, Nawel LU
supervisor
organization
course
KEMT30 20171
year
type
H2 - Master's Degree (Two Years)
subject
keywords
Pseudomonas aeruginosa, chronic wounds, elastase, enzymatic activity, cytokines, protein science, proteinvetenskap
language
English
id
8927277
date added to LUP
2017-10-31 09:34:42
date last changed
2017-10-31 09:34:42
@misc{8927277,
  abstract     = {Wound healing is a process that ensures regeneration and repair of damaged tissue. This process can be interrupted, leading to chronic wounds, and one of the factors that can interrupt it is infection. The bacterium Pseudomonas aeruginosa is prevalent in chronic wounds, and as it is antibiotic resistant it makes chronic wounds infected with this pathogen a challenging clinical problem.
For a better understanding of causes behind wound healing failure when a wound is infected by P. aeruginosa we studied elastase, which is one of several virulence factors of P. aeruginosa. This protein is secreted by most strains of this species and is characterized as zinc metalloprotease. The present study relies on the isolation and purification of elastase for investigations of a potential difference in the enzymatic activity of elastase between four different strains of P. aeruginosa, the PAO1 (wild type strain), PAOA1 (elastase A mutant), PAOB1 (elastase B mutant), and PAOA1B1 (elastase A and B double mutant). Also, to understand the interplay between elastase and the host by clarifying the interaction between elastase and both the immune and the coagulation systems.
Results of the purity analysis, using tricine gels together with results of gelatin zymography show that the purification of active elastase B using fast protein liquid chromatography was successful. Our results from an azocasein assay together with the gelatin zymography of both P. aeruginosa conditioned medium and purified elastase show that elastase B dominates the elastolytic activity of P. aeruginosa. Interestingly, results of this study disagree with the hypothesis that elastase A can enhance the virulence of elastase B. Contrary, the present data suggest that elastase A represses virulence of both elastase B and the toxin pyocyanin.},
  author       = {Betina, Nawel},
  keyword      = {Pseudomonas aeruginosa,chronic wounds,elastase,enzymatic activity,cytokines,protein science,proteinvetenskap},
  language     = {eng},
  note         = {Student Paper},
  title        = {Host-pathogen interactions in chronic infections and the development of novel therapies},
  year         = {2017},
}