OXGR1 overexpression as putative prostate cancer promoting factor
Järvinen, Sampsa (2017)
Järvinen, Sampsa
2017
Bioteknologian tutkinto-ohjelma - Degree Programme in Biotechnology
Lääketieteen ja biotieteiden tiedekunta - Faculty of Medicine and Life Sciences
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Hyväksymispäivämäärä
2017-11-26
Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:uta-201712012848
https://urn.fi/URN:NBN:fi:uta-201712012848
Tiivistelmä
Background and aims: Prostate cancer is the most common cancer diagnosed in men. While it often has indolent course, and diagnosed men more commonly perish to comorbidities, it remains the third-leading cause of cancer death in men. Current diagnostic tools fail to reliably distinguish aggressive cancer cases from indolent ones and new tools with better prognostic value are needed to address this. Preferably, a well-informed treatment choice based on histopathological findings and molecular characteristics of cancer could be made to achieve best possible treatment outcome. To succeed in this, molecular mechanisms of prostate cancer must be better understood. Based on gene expression profiles from clinical samples, overexpression of OXGR1 could be a cancer promoting factor in certain prostate cancer subtypes. The aim of this study was to characterize effects of OXGR1 overexpression to growth rate of two prostate cancer cell lines in varying key substrate concentrations.
Methods: OXGR1 overexpressing PC-3 and LNCaP prostate cancer cells lines were created previously by stable transfections with pcDNA3.1 vectors having OXGR1 under constitutive promoter. Expression profiles of cells were characterized with RT-qPCR, western blot and immunofluorescence staining. Growth rate experiments were performed in well plates using different concentrations of glucose, glutamine and alpha-ketoglutarate, which is a ligand for OXGR1, in growth medium. Cells were imaged at intervals and growth rate was defined by measuring surface area programmatically.
Results: All transfected cell lines had high OXGR1 mRNA levels compared to controls. In LNCaP cell lines western blot results showed clear increase in protein expression of transfected lines. In transfected LNCaP cell lines an increase in growth rate was seen as the alpha-ketoglutarate concentration was increased. In PC-3 cell lines the protein levels were similarly low regardless of transfection status. OXGR1 overexpression had no effect on growth rate of PC-3 cells in any of the growth conditions. OXGR1 antibody used in western blot experiments produced a clear band over 10 kilodaltons larger than the predicted band size.
Conclusions: These results show promise that OXGR1 overexpression could have significance in prostate cancer promotion and that stable OXGR1 expressing LNCaP cell lines can be used to study this further. However, proper binding of the OXGR1 antibody must be validated and the cause of shift in band size elucidated. Studies that take more molecular interactions into account are probably needed to clarify the role of OXGR1 overexpression in prostate cancer. This could possibly be done by performing OXGR1 overexpression studies in prostate cancer mouse model.
Methods: OXGR1 overexpressing PC-3 and LNCaP prostate cancer cells lines were created previously by stable transfections with pcDNA3.1 vectors having OXGR1 under constitutive promoter. Expression profiles of cells were characterized with RT-qPCR, western blot and immunofluorescence staining. Growth rate experiments were performed in well plates using different concentrations of glucose, glutamine and alpha-ketoglutarate, which is a ligand for OXGR1, in growth medium. Cells were imaged at intervals and growth rate was defined by measuring surface area programmatically.
Results: All transfected cell lines had high OXGR1 mRNA levels compared to controls. In LNCaP cell lines western blot results showed clear increase in protein expression of transfected lines. In transfected LNCaP cell lines an increase in growth rate was seen as the alpha-ketoglutarate concentration was increased. In PC-3 cell lines the protein levels were similarly low regardless of transfection status. OXGR1 overexpression had no effect on growth rate of PC-3 cells in any of the growth conditions. OXGR1 antibody used in western blot experiments produced a clear band over 10 kilodaltons larger than the predicted band size.
Conclusions: These results show promise that OXGR1 overexpression could have significance in prostate cancer promotion and that stable OXGR1 expressing LNCaP cell lines can be used to study this further. However, proper binding of the OXGR1 antibody must be validated and the cause of shift in band size elucidated. Studies that take more molecular interactions into account are probably needed to clarify the role of OXGR1 overexpression in prostate cancer. This could possibly be done by performing OXGR1 overexpression studies in prostate cancer mouse model.