Setting up an acute infection model for CVB1 in mice
Mustonen, Iiris (2024)
2024
Bioteknologian ja biolääketieteen tekniikan maisteriohjelma - Master's Programme in Biotechnology and Biomedical Engineering
Lääketieteen ja terveysteknologian tiedekunta - Faculty of Medicine and Health Technology
This publication is copyrighted. You may download, display and print it for Your own personal use. Commercial use is prohibited.
Hyväksymispäivämäärä
2024-05-08
Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:tuni-202404244514
https://urn.fi/URN:NBN:fi:tuni-202404244514
Tiivistelmä
Background and aims: Coxsackie B viruses (CVB) under Enterovirus (EV) genus are small yet highly contagious human pathogens. CVB infection can cause severe inflammation in multiple organs, including heart and pancreas. Infection is also linked with diseases such as type 1 diabetes (T1D) and dilated cardiopathy (DCM), among others. The pathogenic mechanisms of CVBs and their role in disease aetiologies are not well known. In addition, there are no treatments or vaccines against CVBs. The aim of this thesis is to set up an animal infection laboratory for Tampere University and to set up an acute infection model for CVB1 in mice. Animal infection model would enable research on CVB infection mechanisms, disease associations and testing for preventive strategies.
Methods: CVB1 was produced in Green Monkey Kidney (GMK) cells and purified by ultracentrifugation. Purified virus was characterized with SDS-PAGE, Western blot, Dynamic Light scattering and BCA total protein quantification. Infectivity of the virus was measured with tissue culture infectious dose 50% (TCID50) method. Animal infection laboratory was established, and operating procedures were determined according to risk assessment and mock studies, among other factors. Infection model was established by intraperitoneal inoculation of CVB1 in BALB/c mice. Infection studies were conducted in four separate studies. The model was optimized ac-cording to infectious dose, study length, and sex. The infection was determined from the amount of replicative virus in the pancreas, heart, and blood by TCID50 method. During the studies, wellbeing of the mice was evaluated with body score indexing, and weights were measured daily.
Results: Induction of systemic CVB1 infection in BALB/c mice was successful. Replicative virus was most consistently found in pancreas but was also detected from blood samples until day 4. Viremia increased during the first 4 days post infection (dpi) in female mice. Viral doses 106 and 107 TCID50 per mouse were able to cause infection in the pancreas most consistently. Replicative virus seemed to be cleared from blood and pancreas around 7 dpi. Female mice had higher viral titres in pancreas 4 dpi compared to male mice. Wellbeing of male mice deteriorated during the infections as indicated by diarrhea and ruffled fur. Also, weights decreased significantly. Female mice, on the other hand, did not show external signs of infection and the weights remained similar. Replicative virus was not found in heart at any points of the studies.
Conclusions: The aims of the thesis were successfully completed. The animal infection laboratory was set up and we were able to induce systemic CVB1 infection in mice. For further optimization of the infection model, we should use the optimized viral doses of 106 and 107 TCID50. For detection of replicative virus in pancreas, the infection should be followed 4 dpi. Moreover, our results support the use of female mice to reduce animal suffering. This infection model could further enable conduction of challenge studies, where safety and efficacy of vaccines can be tested. Moreover, established facility would support infection models with different viruses serotypes or modelling CVB induced diseases.
Methods: CVB1 was produced in Green Monkey Kidney (GMK) cells and purified by ultracentrifugation. Purified virus was characterized with SDS-PAGE, Western blot, Dynamic Light scattering and BCA total protein quantification. Infectivity of the virus was measured with tissue culture infectious dose 50% (TCID50) method. Animal infection laboratory was established, and operating procedures were determined according to risk assessment and mock studies, among other factors. Infection model was established by intraperitoneal inoculation of CVB1 in BALB/c mice. Infection studies were conducted in four separate studies. The model was optimized ac-cording to infectious dose, study length, and sex. The infection was determined from the amount of replicative virus in the pancreas, heart, and blood by TCID50 method. During the studies, wellbeing of the mice was evaluated with body score indexing, and weights were measured daily.
Results: Induction of systemic CVB1 infection in BALB/c mice was successful. Replicative virus was most consistently found in pancreas but was also detected from blood samples until day 4. Viremia increased during the first 4 days post infection (dpi) in female mice. Viral doses 106 and 107 TCID50 per mouse were able to cause infection in the pancreas most consistently. Replicative virus seemed to be cleared from blood and pancreas around 7 dpi. Female mice had higher viral titres in pancreas 4 dpi compared to male mice. Wellbeing of male mice deteriorated during the infections as indicated by diarrhea and ruffled fur. Also, weights decreased significantly. Female mice, on the other hand, did not show external signs of infection and the weights remained similar. Replicative virus was not found in heart at any points of the studies.
Conclusions: The aims of the thesis were successfully completed. The animal infection laboratory was set up and we were able to induce systemic CVB1 infection in mice. For further optimization of the infection model, we should use the optimized viral doses of 106 and 107 TCID50. For detection of replicative virus in pancreas, the infection should be followed 4 dpi. Moreover, our results support the use of female mice to reduce animal suffering. This infection model could further enable conduction of challenge studies, where safety and efficacy of vaccines can be tested. Moreover, established facility would support infection models with different viruses serotypes or modelling CVB induced diseases.