Expression and Functionality of Connexin-Based Channels in Human Liver Cancer Cell Lines
Abstract
:1. Introduction
2. Results
2.1. Selection of Liver Cancer Cell Lines
2.2. Connexin Gene Expression in Liver Cancer Cell Lines
2.3. Connexin Protein Expression in Liver Cancer Cell Lines
2.4. Connexin Protein Subcellular Localization in Liver Cancer Cell Lines
2.5. Gap Junctional Intercellular Communication in Liver Cancer Cell Lines
3. Discussion
4. Materials and Methods
4.1. Reagents
4.2. Liver Cancer Cell Lines and Primary Human Hepatocytes
4.3. Phase-Contrast Microscopy
4.4. Real-Time Quantitative Reverse Transcription Polymerase Chain Reaction Analysis
4.5. Immunoblot Analysis
4.6. Immunocytochemistry Analysis
4.7. Scrape Loading/Dye Transfer Assay
4.8. Statistical Analysis
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Cell Line | Morphology | Disease | Age | Gender | Ethnicity | HBV | Medium |
---|---|---|---|---|---|---|---|
SK-HEP-1 | Epithelial | Adenocarcinoma | 52 | Male | Caucasian | Not detected | EMEM + FBS |
C3A | Epithelial | HCC | 15 | Male | Caucasian | Not detected | EMEM + FBS |
SNU-449 | Epithelial | Grade II-III/IV, HCC | 52 | Male | Asian | DNA detected | RPMI-1640 + heat-inactivated FBS |
SNU-423 | Epithelial | Grade III/IV, pleomorphic HCC | 40 | Male | Asian | DNA detected | RPMI-1640 + heat-inactivated FBS |
SNU-387 | Epithelial | Grade IV/V, pleomorphic HCC | 41 | Female | Asian | DNA detected | RPMI-1640 + FBS |
SNU-475 | Epithelial | Grade II-IV/V, HCC | 43 | Male | Asian | DNA detected | RPMI-1640 + heat-inactivated FBS |
PLC/PRF/5 | Epithelial | HCC | 24 | Male | African | Secreted HbsAg | EMEM + FBS |
PHH | Epithelial | Liver metastasis of colorectal adenocarcinoma | 74 | Female | Caucasian | Not detected | CHRM + CHPM |
Gene Symbol | Assay ID | Accession Number | Assay Location | Amplicon Size (Base Pairs) | Exon Boundary |
---|---|---|---|---|---|
GJA1 (Cx43) | Hs00748445-s1 | NM_000165.4 | 1031 | 142 | 2 |
GJB1 (Cx32) | Hs00939759-s1 | NM_000166.5 NM_001097642.2 | 1547 1496 | 63 | 2 |
GJB2 (Cx26) | Hs00269615-s1 | NM_004004.5 | 715 | 123 | 2 |
ACTB | Hs01060665-g1 | NM_001101.3 | 208 | 63 | 2–3 |
B2M | Hs00187842-m1 | NM_004048.2 | 134 | 64 | 1–2 |
GAPDH | Hs02786624-g1 | NM_001256799.2 NM_001289745.1 NM_001289746.1 NM_002046.5 | 870 928 822 836 | 157 | 7 8 7 8 |
HMBS | Hs00609296-g1 | NM_000190.3 NM_001024382.1 NM_001258208.1 NM_001258209.1 | 1070 972 950 1041 | 69 | 13–14 13–14 12–13 12–13 |
UBC | Hs01871556-s1 | M26880.1 | 2173 | 135 | – |
Target | Product Code | Dilution Primary Antibody | Supplier | |
---|---|---|---|---|
Immunoblot | Immunocytochemistry | |||
Cx26 | 51–2800 | 1:250 | 1:250 | Invitrogen, Waltham, MA, USA |
Cx32 | C3470 | 1:600 | 1:500 | Sigma-Aldrich, St. Louis, MO, USA |
Cx43 | C6219 | 1:4000 | 1:1000 | Sigma-Aldrich, St. Louis, MO, USA |
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Leroy, K.; Silva Costa, C.J.; Pieters, A.; dos Santos Rodrigues, B.; Van Campenhout, R.; Cooreman, A.; Tabernilla, A.; Cogliati, B.; Vinken, M. Expression and Functionality of Connexin-Based Channels in Human Liver Cancer Cell Lines. Int. J. Mol. Sci. 2021, 22, 12187. https://doi.org/10.3390/ijms222212187
Leroy K, Silva Costa CJ, Pieters A, dos Santos Rodrigues B, Van Campenhout R, Cooreman A, Tabernilla A, Cogliati B, Vinken M. Expression and Functionality of Connexin-Based Channels in Human Liver Cancer Cell Lines. International Journal of Molecular Sciences. 2021; 22(22):12187. https://doi.org/10.3390/ijms222212187
Chicago/Turabian StyleLeroy, Kaat, Cícero Júlio Silva Costa, Alanah Pieters, Bruna dos Santos Rodrigues, Raf Van Campenhout, Axelle Cooreman, Andrés Tabernilla, Bruno Cogliati, and Mathieu Vinken. 2021. "Expression and Functionality of Connexin-Based Channels in Human Liver Cancer Cell Lines" International Journal of Molecular Sciences 22, no. 22: 12187. https://doi.org/10.3390/ijms222212187