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Reduction of human chorionic gonadotropin beta subunit expression by modified U1 snRNA caused apoptosis in cervical cancer cells.

Jankowska A, Gunderson SI, Andrusiewicz M, Burczynska B, Szczerba A, Jarmolowski A, Nowak-Markwitz E, Warchol JB - Mol. Cancer (2008)

Bottom Line: This study documents the molecular presence of human chorionic gonadotropin beta subunit in uterine cervix cancer tissues and investigates a novel technique to reduce hCGbeta levels based on expression of a modified U1 snRNA as a method to study the hormone's role in biology of human cervical cancer cells cultured in vitro.Reduction of hCGbeta levels resulted in a significantly increased apoptosis rate with almost 90% of cells transfected with modified anti-hCGbeta U1 snRNAs showing morphological changes characteristic of the apoptotic process.These data suggest that human chorionic gonadotropin beta subunit may act as a tumor growth-stimulating factor.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell Biology, University of Medical Sciences, Rokietnicka 5D, Poznan, Poland. ajanko@amp.edu.pl

ABSTRACT

Background: Secretion of human chorionic gonadotropin, especially its beta subunit by malignant trophoblastic tumors and varieties of tumors of different origin is now well documented; however the role of hCG in tumorogenesis is still unknown.

Results: This study documents the molecular presence of human chorionic gonadotropin beta subunit in uterine cervix cancer tissues and investigates a novel technique to reduce hCGbeta levels based on expression of a modified U1 snRNA as a method to study the hormone's role in biology of human cervical cancer cells cultured in vitro. The property of U1 snRNA to block the accumulation of specific RNA transcript when it binds to its donor sequence within the 3' terminal exon was used. The first 10 nucleotides of the human U1 snRNA gene, which normally binds to the 5'ss in pre-mRNA were replaced by a sequence complementary to a 10-nt segment in the terminal exon of the hCGbeta mRNA. Three different 5' end-mutated U1 snRNA expression plasmids were tested, each targeting a different sequence in the hCGbeta mRNA, and we found each one blocked the expression of hCGbeta in HeLa cells, a cervix carcinoma cell line, as shown by immunohistochemistry and qRT-PCR. Reduction of hCGbeta levels resulted in a significantly increased apoptosis rate with almost 90% of cells transfected with modified anti-hCGbeta U1 snRNAs showing morphological changes characteristic of the apoptotic process.

Conclusion: These data suggest that human chorionic gonadotropin beta subunit may act as a tumor growth-stimulating factor.

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Effect of transfection with anti-hCGβ U1 snRNA on apoptotic rate in HeLa cells. FACS analyses using propidium iodide staining was performed for DNA content measurement. Apoptotic population was measured as the percentage of total cell population with sub-G1 DNA content. Results shown represents experiments performed for control cells and cells transfected with plasmid 702P and 702P/767A, respectively.
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Figure 6: Effect of transfection with anti-hCGβ U1 snRNA on apoptotic rate in HeLa cells. FACS analyses using propidium iodide staining was performed for DNA content measurement. Apoptotic population was measured as the percentage of total cell population with sub-G1 DNA content. Results shown represents experiments performed for control cells and cells transfected with plasmid 702P and 702P/767A, respectively.

Mentions: Changes in cell cycles distribution were monitored by flow cytometry. The inhibition of hCGβ expression by anti-hCGβ constructs result in increase in distribution of cells at sub-G1 peak – apoptosis peak (Figure 6). The percentage of apoptotic cells was increased to 26.65%, 23.59% and 22.48% for 702P/767A, 702P/767B and 702P/767C plasmids respectively (Table 1).


Reduction of human chorionic gonadotropin beta subunit expression by modified U1 snRNA caused apoptosis in cervical cancer cells.

Jankowska A, Gunderson SI, Andrusiewicz M, Burczynska B, Szczerba A, Jarmolowski A, Nowak-Markwitz E, Warchol JB - Mol. Cancer (2008)

Effect of transfection with anti-hCGβ U1 snRNA on apoptotic rate in HeLa cells. FACS analyses using propidium iodide staining was performed for DNA content measurement. Apoptotic population was measured as the percentage of total cell population with sub-G1 DNA content. Results shown represents experiments performed for control cells and cells transfected with plasmid 702P and 702P/767A, respectively.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2335103&req=5

Figure 6: Effect of transfection with anti-hCGβ U1 snRNA on apoptotic rate in HeLa cells. FACS analyses using propidium iodide staining was performed for DNA content measurement. Apoptotic population was measured as the percentage of total cell population with sub-G1 DNA content. Results shown represents experiments performed for control cells and cells transfected with plasmid 702P and 702P/767A, respectively.
Mentions: Changes in cell cycles distribution were monitored by flow cytometry. The inhibition of hCGβ expression by anti-hCGβ constructs result in increase in distribution of cells at sub-G1 peak – apoptosis peak (Figure 6). The percentage of apoptotic cells was increased to 26.65%, 23.59% and 22.48% for 702P/767A, 702P/767B and 702P/767C plasmids respectively (Table 1).

Bottom Line: This study documents the molecular presence of human chorionic gonadotropin beta subunit in uterine cervix cancer tissues and investigates a novel technique to reduce hCGbeta levels based on expression of a modified U1 snRNA as a method to study the hormone's role in biology of human cervical cancer cells cultured in vitro.Reduction of hCGbeta levels resulted in a significantly increased apoptosis rate with almost 90% of cells transfected with modified anti-hCGbeta U1 snRNAs showing morphological changes characteristic of the apoptotic process.These data suggest that human chorionic gonadotropin beta subunit may act as a tumor growth-stimulating factor.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell Biology, University of Medical Sciences, Rokietnicka 5D, Poznan, Poland. ajanko@amp.edu.pl

ABSTRACT

Background: Secretion of human chorionic gonadotropin, especially its beta subunit by malignant trophoblastic tumors and varieties of tumors of different origin is now well documented; however the role of hCG in tumorogenesis is still unknown.

Results: This study documents the molecular presence of human chorionic gonadotropin beta subunit in uterine cervix cancer tissues and investigates a novel technique to reduce hCGbeta levels based on expression of a modified U1 snRNA as a method to study the hormone's role in biology of human cervical cancer cells cultured in vitro. The property of U1 snRNA to block the accumulation of specific RNA transcript when it binds to its donor sequence within the 3' terminal exon was used. The first 10 nucleotides of the human U1 snRNA gene, which normally binds to the 5'ss in pre-mRNA were replaced by a sequence complementary to a 10-nt segment in the terminal exon of the hCGbeta mRNA. Three different 5' end-mutated U1 snRNA expression plasmids were tested, each targeting a different sequence in the hCGbeta mRNA, and we found each one blocked the expression of hCGbeta in HeLa cells, a cervix carcinoma cell line, as shown by immunohistochemistry and qRT-PCR. Reduction of hCGbeta levels resulted in a significantly increased apoptosis rate with almost 90% of cells transfected with modified anti-hCGbeta U1 snRNAs showing morphological changes characteristic of the apoptotic process.

Conclusion: These data suggest that human chorionic gonadotropin beta subunit may act as a tumor growth-stimulating factor.

Show MeSH
Related in: MedlinePlus