The effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / Sarmila Hanim Mustafa

The effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / Sarmila Hanim Mustafa

Osteosarcoma (OS) is a rare malignant bone cancer affecting children and young adults. To date, OS still has a poor prognosis despite the use of advanced multimodal therapy in its treatment. Recurrence cases has also shown to be increased in metastatic patients especially to the lung. A possible...

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Main Author: Mustafa, Sarmila Hanim
Format: Thesis
Language:English
Published: 2020
Subjects:
Research
Experimentation
Online Access:https://ir.uitm.edu.my/id/eprint/60051/1/60051.pdf
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spelling my-uitm-ir.600512022-05-21T01:22:09Z The effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / Sarmila Hanim Mustafa 2020-08 Mustafa, Sarmila Hanim Research. Experimentation Osteosarcoma (OS) is a rare malignant bone cancer affecting children and young adults. To date, OS still has a poor prognosis despite the use of advanced multimodal therapy in its treatment. Recurrence cases has also shown to be increased in metastatic patients especially to the lung. A possible cause for this poor prognosis could be because of lack of understanding in the underlying mechanism of OS progression and metastasis. Previously, loss of Wnt/β-catenin pathway activity during osteoblast differentiation was suggested to contribute to the OS development. In addition, a metastatic condition of OS was shown to be established through the adhesion of OS cells to blood vessel mediated by α4β1 integrin. It has been known that aberrant glycosylation is associated with metastasis in various cancer by modulating the cells invasiveness and their dissemination through the extracellular matrix (ECM) layer. However, the effect of aberrant glycosylation, mainly during N-glycan processing in OS has yet to be elucidated. Therefore, the objective of this study is to investigate the effects of aberrant glycosylation towards OS invasiveness via the cell-ECM interaction. In this study, the inhibition of glycosylation was carried out by the treatment of OS cell line (MG-63) with 1-deoxynojirimycin (1-DNJ); an α- glucosidase-I/II inhibitor, and 1-deoxymannojirimycin (1-DMJ); an α-mannosidase I inhibitor. Then, its effects on glycosylation pattern, invasion ability, protein expression of α4β1 integrin and the expression level of ECM degradative enzymes genes; MMP-2, MMP-9, TIMP-1, TIMP-2 and β-catenin genes were determined. In this study, the normal osteoblast cell (hFOB1.19) was also treated with 1-DNJ and 1- DMJ and both samples were subjected to the same method as MG-63 cells except for cell invasion assay. The results showed that the inhibition of N-glycosylation by 1- DMJ enhanced the invasion of MG-63 cells through the ECM layer significantly by 3.87 fold (p<0.05). It was also found that 1-DMJ significantly (p<0.05) regulates the expression of α4β1 integrin protein in hFOB1.19 cells. qRT-PCR analysis showed that 1-DMJ downregulate the expression MMP-9 gene (2.42 fold ± 0.284; p<0.05) in MG- 63 cells. On the other hand, TIMP-1 (1.54 fold ± 0.035), TIMP-2 (1.22 fold ± 0.041) and β-catenin (1.24 fold ± 0.07) genes were significantly (p<0.05) upregulated in MG- 63 cells when compared to control. However, in hFOB1.19 cells treated with 1-DMJ, MMP-2 (48.32 fold ± 0.16), MMP-9 (4.38 fold ± 0.389), and TIMP-1 (1.45 fold ± 0.064) genes were significantly (p<0.05) downregulated when compared to control. Whereas, the expression of TIMP-2 (1.2 fold ± 0.03), and β-catenin gene (2.22 fold ± 0.232) in hFOB1.19 cells were significantly (p<0.05) upregulated. 1-DNJ was shown to downregulate the expression of MMP-2 in both cells; MG-63 (1.29 fold ± 0.067) and hFOB1.19 (55.25 fold ± 0.097) significantly (p<0.05) when compared to control. However, the expression of β-catenin gene in 1-DNJ treated hFOB1.19 cells was significantly (p<0.05) increased by 2.4 fold ± 0.103 when compared to control. This study suggested that 1-DMJ may promote the invasion of MG-63 cells through the ECM layer via the regulation of MMP-9, TIMP-1, and TIMP-2 genes. Whereas the overexpression of β-catenin genes in 1-DMJ and 1-DNJ treated hFOB1.19 cells might give an insight on the epithelial-to-mesenchymal transition (EMT) of normal osteoblast cells. This study might provide more information on the underlying mechanism of OS therefore contributes to OS prognosis improvement. 2020-08 Thesis https://ir.uitm.edu.my/id/eprint/60051/ https://ir.uitm.edu.my/id/eprint/60051/1/60051.pdf text en public masters Universiti Teknologi MARA Faculty of Medicine Ab. Rahim, Sharaniza (Assoc. Prof. Dr.)
institution Universiti Teknologi MARA
collection UiTM Institutional Repository
language English
advisor Ab. Rahim, Sharaniza (Assoc. Prof. Dr.)
topic Research
Experimentation
spellingShingle Research
Experimentation
Mustafa, Sarmila Hanim
The effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / Sarmila Hanim Mustafa
description Osteosarcoma (OS) is a rare malignant bone cancer affecting children and young adults. To date, OS still has a poor prognosis despite the use of advanced multimodal therapy in its treatment. Recurrence cases has also shown to be increased in metastatic patients especially to the lung. A possible cause for this poor prognosis could be because of lack of understanding in the underlying mechanism of OS progression and metastasis. Previously, loss of Wnt/β-catenin pathway activity during osteoblast differentiation was suggested to contribute to the OS development. In addition, a metastatic condition of OS was shown to be established through the adhesion of OS cells to blood vessel mediated by α4β1 integrin. It has been known that aberrant glycosylation is associated with metastasis in various cancer by modulating the cells invasiveness and their dissemination through the extracellular matrix (ECM) layer. However, the effect of aberrant glycosylation, mainly during N-glycan processing in OS has yet to be elucidated. Therefore, the objective of this study is to investigate the effects of aberrant glycosylation towards OS invasiveness via the cell-ECM interaction. In this study, the inhibition of glycosylation was carried out by the treatment of OS cell line (MG-63) with 1-deoxynojirimycin (1-DNJ); an α- glucosidase-I/II inhibitor, and 1-deoxymannojirimycin (1-DMJ); an α-mannosidase I inhibitor. Then, its effects on glycosylation pattern, invasion ability, protein expression of α4β1 integrin and the expression level of ECM degradative enzymes genes; MMP-2, MMP-9, TIMP-1, TIMP-2 and β-catenin genes were determined. In this study, the normal osteoblast cell (hFOB1.19) was also treated with 1-DNJ and 1- DMJ and both samples were subjected to the same method as MG-63 cells except for cell invasion assay. The results showed that the inhibition of N-glycosylation by 1- DMJ enhanced the invasion of MG-63 cells through the ECM layer significantly by 3.87 fold (p<0.05). It was also found that 1-DMJ significantly (p<0.05) regulates the expression of α4β1 integrin protein in hFOB1.19 cells. qRT-PCR analysis showed that 1-DMJ downregulate the expression MMP-9 gene (2.42 fold ± 0.284; p<0.05) in MG- 63 cells. On the other hand, TIMP-1 (1.54 fold ± 0.035), TIMP-2 (1.22 fold ± 0.041) and β-catenin (1.24 fold ± 0.07) genes were significantly (p<0.05) upregulated in MG- 63 cells when compared to control. However, in hFOB1.19 cells treated with 1-DMJ, MMP-2 (48.32 fold ± 0.16), MMP-9 (4.38 fold ± 0.389), and TIMP-1 (1.45 fold ± 0.064) genes were significantly (p<0.05) downregulated when compared to control. Whereas, the expression of TIMP-2 (1.2 fold ± 0.03), and β-catenin gene (2.22 fold ± 0.232) in hFOB1.19 cells were significantly (p<0.05) upregulated. 1-DNJ was shown to downregulate the expression of MMP-2 in both cells; MG-63 (1.29 fold ± 0.067) and hFOB1.19 (55.25 fold ± 0.097) significantly (p<0.05) when compared to control. However, the expression of β-catenin gene in 1-DNJ treated hFOB1.19 cells was significantly (p<0.05) increased by 2.4 fold ± 0.103 when compared to control. This study suggested that 1-DMJ may promote the invasion of MG-63 cells through the ECM layer via the regulation of MMP-9, TIMP-1, and TIMP-2 genes. Whereas the overexpression of β-catenin genes in 1-DMJ and 1-DNJ treated hFOB1.19 cells might give an insight on the epithelial-to-mesenchymal transition (EMT) of normal osteoblast cells. This study might provide more information on the underlying mechanism of OS therefore contributes to OS prognosis improvement.
format Thesis
qualification_level Master's degree
author Mustafa, Sarmila Hanim
author_facet Mustafa, Sarmila Hanim
author_sort Mustafa, Sarmila Hanim
title The effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / Sarmila Hanim Mustafa
title_short The effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / Sarmila Hanim Mustafa
title_full The effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / Sarmila Hanim Mustafa
title_fullStr The effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / Sarmila Hanim Mustafa
title_full_unstemmed The effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / Sarmila Hanim Mustafa
title_sort effects of extracellular matrix glycosylation inhibition on osteosarcoma invasiveness / sarmila hanim mustafa
granting_institution Universiti Teknologi MARA
granting_department Faculty of Medicine
publishDate 2020
url https://ir.uitm.edu.my/id/eprint/60051/1/60051.pdf
_version_ 1783735076350066688

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