title: Retinoic acid morpholine amide (RAMA) inhibits expression of Fas ligand through EP1 receptor in colon cancer cells.
abstract: Among the members of tumour necrosis factor family Fas ligand on binding to its receptor strongly induces apoptosis of tumour-infiltrating lymphocytes (TIL). Thus, FasL acts as an inhibitor of anti-tumour immune response. The present study demonstrates that retinoic acid morpholine amide (RAMA) significantly suppresses FasL expression in colon cancer cells in a dose- and time-dependent manner. The suppression of FasL mRNA and proteins was significant at a concentration of 30 μM after 48 h in CLT85 and HT26 colon cancer cells. There was around 2.6- and 3.2-fold decrease in FasL mRNA after incubation with 30 μM of RAMA in CLT85 cells and HT26 cells, respectively. The results from Western blot showed a decrease in FasL mRNA and protein expression in both CLT85 and HT26 cells after suppression of cyclooxygenase (COX)-2 and COX-1 by RNAi. However, when COX-2-specific silencer RNA (siCOX-2)- and siCOX-1-treated CLT85 and HT26 cells were exposed to RAMA, inhibition of FasL expression was further suppressed. The siCOX-2-treated CLT85 and HT26 cells on exposure to RAMA showed ∼87 and ∼54 % reduction in FasL mRNA, respectively. Co-culture of Jurkat T cells with RAMA-treated HT26 and CLT85 cells decreased the viability of Jurkat T cells by only 2 and 4.3 %, respectively, compared to 19.5 and 37.3 % in control HT26 and CLT85 cells. The results from real-time reverse transcription polymerase chain reaction (RT-PCR) and immunoblotting showed that suppression of EP1 prevented RAMA-induced FasL suppression in CLT85 cells at both the mRNA and protein levels. Thus, RAMA can be a potent therapeutic agent for the treatment of colon tumours.
authors.affiliation: Department of Endoscopy Center, China-Japan Friendship Hospital, No. 2 Yinghua Dong Street, Chaoyang District, Beijing, 100029, China.
authors.forename: Shao-Xuan
authors.initials: SX
authors.lastname: Chen
authors.affiliation: Department of Gastroenterology, China-Japan Friendship Hospital, Beijing, 100029, China.
authors.forename: Shi-Yu
authors.initials: SY
authors.lastname: Du
authors.affiliation: Department of Orthopedics, China-Japan Friendship Hospital, Beijing, 100029, China.
authors.forename: Yun-Ting
authors.initials: YT
authors.lastname: Wang
authors.affiliation: Department of Gastroenterology, China-Japan Friendship Hospital, Beijing, 100029, China.
authors.forename: Hong-Chuan
authors.initials: HC
authors.lastname: Zhao
authors.affiliation: Department of Gastroenterology, China-Japan Friendship Hospital, Beijing, 100029, China.
authors.forename: Yan-Li
authors.initials: YL
authors.lastname: Zhang
authors.affiliation: Department of Endoscopy Center, China-Japan Friendship Hospital, No. 2 Yinghua Dong Street, Chaoyang District, Beijing, 100029, China. yaoli9886@gmail.com.
authors.forename: Li
authors.initials: L
authors.lastname: Yao
mesh_terms: D000577:Amides / Q000737:chemistry*; D045744:Cell Line, Tumor; D002470:Cell Survival; D018920:Coculture Techniques; D003110:Colonic Neoplasms / Q000378:metabolism*; D051546:Cyclooxygenase 2 / Q000378:metabolism; D053222:Fas Ligand Protein / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007107:Immune System; D019169:Jurkat Cells; D008856:Microscopy, Fluorescence; D009025:Morpholines / Q000737:chemistry*; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D058306:Receptors, Prostaglandin E, EP1 Subtype / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014212:Tretinoin / Q000737:chemistry*; D019014:fas Receptor / Q000378:metabolism
chemical_list: D000577:Amides; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D009025:Morpholines; D012333:RNA, Messenger; D058306:Receptors, Prostaglandin E, EP1 Subtype; D019014:fas Receptor; D014212:Tretinoin; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human
keywords: Cytotoxicity; Fas-sensitive; Immune response; Tumour necrosis