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:: Volume 11, Issue 2 (Int J Mol Cell Med (In Press) 2022) ::
Int J Mol Cell Med 2022, 11(2): 0-0 Back to browse issues page
A Decrease in CD44 on Cell Surfaces (MKN-45 Cell Line) After RELA Knockout Using CRISPR/Cas9
Saeid Karimi1, Sima Salmani1, Akram Alizadeh2, Leila Rezakhani3, Zohreh Saltanatpour4, Sorayya Ghasemi5
1- Cellular and Molecular Research Center, Basic Health Sciences Institute, Shahrekord University of Medical Sciences, Shahrekord, Iran.
2- Department of Tissue Engineering and Applied Cell Sciences, Faculty of Medicine, Semnan University of Medical Sciences, Semnan, Iran.
3- Fertility and Infertility Research Center, Health Technology Institute, Kermanshah University of Medical Sciences, Kermanshah, Iran.
4- Pediatric Cell and Gene Therapy Research Center, Gene, Cell & Tissue Research Institute, Tehran University of Medical Sciences, Tehran, Iran.
5- Cellular and Molecular Research Center, Basic Health Sciences Institute, Shahrekord University of Medical Sciences, Shahrekord, Iran. , sorayya.ghasemi@gmail.com
Abstract:   (39 Views)

The NF-kB signaling pathway was introduced as a key pathway in carcinogenesis that is induced by inflammation in gastrointestinal malignancies. The RelA transcription factor is an important component of this signaling pathway. Furthermore, CD44 is implicated in the tumorigenesis and metastasis of gastric cancer. The aim of this study was to assay the effect of RELA knockout on CD44 expression in MKN45 cells. CRISPR/Cas9 was used to knock out RELA in MKN-45. The median fluorescence intensity (MFI) of CD44 before and after RELA knockout is analyzed in MKN45. The CRISPR/Cas9 vector pSpCas9 (BB)-2A-Puro (PX459) was used for gRNA cloning (two guides). The MKN-45 cell line was co-transfected. The purified co-transfected cells with puromycin were cultured and used for the RELA gene expression assay by real-time PCR. Flow cytometry was used for the analysis of the MFI of CD44+ in MKN45. The results showed that 180 nucleotide sequences between exon 2 and exon 3 of RELA were deleted in MKN45. RELA expression was significantly (P<0.001) decreased after CRISPR/Cas9 knockout. Compared to the control group, the MFI of CD44 in transfected cells was significantly decreased (P <0.001). Knockout of RELA significantly decreased CD44 expression in MKN45 cells. It can be concluded that the NF-kB signaling pathway via RELA is related to CD44 expression and consequently the tumorigenesis of gastric cancer. More studies about this relationship are recommended.
 

Keywords: RELA, Knockout, CRISPR/Cas9, CD44, MKN-45 cell line
     
Type of Study: Original Article | Subject: Cancer
Received: 2022/01/16 | Accepted: 2023/01/21 | Published: 2023/01/22
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Karimi S, Salmani S, Alizadeh A, Rezakhani L, Saltanatpour Z, Ghasemi S. A Decrease in CD44 on Cell Surfaces (MKN-45 Cell Line) After RELA Knockout Using CRISPR/Cas9. Int J Mol Cell Med 2022; 11 (2)
URL: http://ijmcmed.org/article-1-1844-en.html


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Volume 11, Issue 2 (Int J Mol Cell Med (In Press) 2022) Back to browse issues page
International Journal of Molecular and Cellular Medicine (IJMCM) International Journal of Molecular and Cellular Medicine (IJMCM)
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