Presentation Type
Poster
Discipline Track
Biomedical Science
Abstract Type
Research/Clinical
Abstract
Background: In the Rio Grande Valley (RGV), Hispanics constitute the predominant ethnic group, accounting for over 94% of the population. Projections for 2023 indicate liver cancer to rank fifth in male and seventh in female mortality. Hepatocellular carcinoma (HCC) manifests as a significant concern within the Hispanic community, influenced by a spectrum of factors—biogenetic, social, and cultural—that intertwine with genetic predispositions and dietary practices. The incidence of HCC is intertwined with various illnesses like hepatitis, liver cirrhosis, drug-induced liver injury (DILI), adding to the burden of cancer. Over the years, HCC prevalence in the RGV has surged by over 35%. A critical factor contributing to the high mortality rates is the development of drug resistance in the primary treatments. Notably, Y-Box Binding Protein 1 (YBX1) exhibits over-expression in HCC based on TCGA data, belonging to a protein superfamily governing mRNA translation. Further exploration of this protein might unveil insights into the mechanisms underpinning treatment resistance in HCC. Understanding its interactions with other pathways and genes could offer pivotal revelations for advancing therapeutic approaches.
Methods: SKHEP-1 HCC cells were obtained and cultured following the guidelines provided by ATCC. An over-expression plasmid for YBX1 (Lentiviral), featuring a Puromycin selection marker and GFP, was custom-designed and acquired from abmgood. The control and over-expression plasmids were introduced into the cells using Lipofectamine 2000. Transfected cells expressing GFP were isolated through FACS enrichment and subsequently maintained in puromycin-containing media to establish stable cell lines. The mRNA expression was assessed using RT-PCR, while protein levels were evaluated via western blot analysis. The over-expression cell lines underwent characterization through oncogenic phenotypic assays, including assessments of migration, invasion, proliferation, and colony formation. Additionally, Xcelligence was utilized to measure the impedance and proliferation dynamics of the recombinant cell lines.
Results: Our TCGA analysis highlighted the significant up-regulation of YBX1 in HCC patient tumors compared to normal tissue. This elevated YBX1 expression strongly correlates with the progression and metastasis of HCC, indicating its role as an indicator of poor survival among HCC patients. Successfully achieving YBX1 over-expression via transient transfection, we enriched GFP-expressing cells and subsequently developed stable SK-Hep1 cell lines with both puromycin resistance and GFP expression, ensuring sustained YBX1 over-expression. In parallel, control cell lines were developed using an empty vector with GFP and puromycin resistance. The observed over-expression of YBX1 led to notable enhancements in proliferation, migration, invasion, and colony formation capabilities of the cell lines. This increase in YBX1 expression was confirmed at both mRNA and protein levels, establishing its impact on various cellular functions in the context of HCC progression.
Conclusions and Future Directions: We've successfully established stable SK-Hep1 cell lines with YBX1 over-expression and thoroughly characterized them. Our next step involves delving deeper into the associated proteins by conducting comprehensive proteomic studies.
Academic/Professional Position
Undergraduate
Academic/Professional Position (Other)
Student Volunteer
Mentor/PI Department
Immunology and Microbiology
Recommended Citation
Abuchard, Yamile; Leslie, Sophia; Kwabiah, Dennis; Pequeno Bracho, Ricardo; Ayala Pazzi, Ana; Hussain, Mohammad; and Tripathi, Manish, "Unveiling the Influence of Transcription Factor YB1 in Hepatocellular Carcinoma Cell Lines" (2024). Research Symposium. 70.
https://scholarworks.utrgv.edu/somrs/2024/posters/70
Included in
Unveiling the Influence of Transcription Factor YB1 in Hepatocellular Carcinoma Cell Lines
Background: In the Rio Grande Valley (RGV), Hispanics constitute the predominant ethnic group, accounting for over 94% of the population. Projections for 2023 indicate liver cancer to rank fifth in male and seventh in female mortality. Hepatocellular carcinoma (HCC) manifests as a significant concern within the Hispanic community, influenced by a spectrum of factors—biogenetic, social, and cultural—that intertwine with genetic predispositions and dietary practices. The incidence of HCC is intertwined with various illnesses like hepatitis, liver cirrhosis, drug-induced liver injury (DILI), adding to the burden of cancer. Over the years, HCC prevalence in the RGV has surged by over 35%. A critical factor contributing to the high mortality rates is the development of drug resistance in the primary treatments. Notably, Y-Box Binding Protein 1 (YBX1) exhibits over-expression in HCC based on TCGA data, belonging to a protein superfamily governing mRNA translation. Further exploration of this protein might unveil insights into the mechanisms underpinning treatment resistance in HCC. Understanding its interactions with other pathways and genes could offer pivotal revelations for advancing therapeutic approaches.
Methods: SKHEP-1 HCC cells were obtained and cultured following the guidelines provided by ATCC. An over-expression plasmid for YBX1 (Lentiviral), featuring a Puromycin selection marker and GFP, was custom-designed and acquired from abmgood. The control and over-expression plasmids were introduced into the cells using Lipofectamine 2000. Transfected cells expressing GFP were isolated through FACS enrichment and subsequently maintained in puromycin-containing media to establish stable cell lines. The mRNA expression was assessed using RT-PCR, while protein levels were evaluated via western blot analysis. The over-expression cell lines underwent characterization through oncogenic phenotypic assays, including assessments of migration, invasion, proliferation, and colony formation. Additionally, Xcelligence was utilized to measure the impedance and proliferation dynamics of the recombinant cell lines.
Results: Our TCGA analysis highlighted the significant up-regulation of YBX1 in HCC patient tumors compared to normal tissue. This elevated YBX1 expression strongly correlates with the progression and metastasis of HCC, indicating its role as an indicator of poor survival among HCC patients. Successfully achieving YBX1 over-expression via transient transfection, we enriched GFP-expressing cells and subsequently developed stable SK-Hep1 cell lines with both puromycin resistance and GFP expression, ensuring sustained YBX1 over-expression. In parallel, control cell lines were developed using an empty vector with GFP and puromycin resistance. The observed over-expression of YBX1 led to notable enhancements in proliferation, migration, invasion, and colony formation capabilities of the cell lines. This increase in YBX1 expression was confirmed at both mRNA and protein levels, establishing its impact on various cellular functions in the context of HCC progression.
Conclusions and Future Directions: We've successfully established stable SK-Hep1 cell lines with YBX1 over-expression and thoroughly characterized them. Our next step involves delving deeper into the associated proteins by conducting comprehensive proteomic studies.