Effects of epidermal growth factor on total cellular RNA,

• Main Authors: Hsieh, Watson H.-S., 謝華昇
• Other Authors: Jane C.-J. Chao Ph.D
• Format: Others
• Language: zh-TW
• Published: 1997
• Online Access: http://ndltd.ncl.edu.tw/handle/15494960968179242647

碩士 === 台北醫學院 === 保健營養學研究所 === 85 === Epidermal growth factor (EGF) composed of 53 amino acids (~ 6 kDa) is a single acid polypeptide and presents in various body fluids and tissues. The differentiation of various cells stimulated by EGF via the EGF receptor has been reported in vivo and in vitro. EGF in human milk has shown to modulate the growth and development of fetal intestinal cells. Therefore, the purpose of this study was to investigate the role of EGF in human milk on growth and the protein digestion enzyme - leucine aminopeptidase (LAP) in human colon adenocarcinoma (Caco-2) cells. The total cellular RNA and protein, and the activity and content of LAP were examined after the addition of different concentrations of EGF. The Caco-2 cells were grown in 80% minimum essential medium (MEM) supplemented with non-essential amino acids, 20% fetal bovine serum, 50 U/mL penicillin and 50 mg/mL strep- tomycin. The cells were incubated at 37℃, in an atmosphere of 95% air -5% CO2. Upon 80% confluency, the cells were switched to serum-free MEM without antibiotics for 24 h. The cells were then incubated with EGF at 0, 5(close to EGF concentration in human milk), 50 (close to EGF concen- tration in colostrum) or 250 nM for another 24 h. The cells were trypsinized and frozen for further analyses. The results showed that the total cellular RNA of the EGF treatment groups was 1.6- to 3.1-fold higher than the control group, but no significant differences were observed among the control, 5 nM and 50 nM EGF treatment groups. The total cellular RNA of 250 nM EGF treatment group was significantly elevated compared to the control and 5 nM EGF treatment groups. Although the cell numbers of the EGF treatment groups were similar to the control group, the total cellular protein of 5 nM EGF treatment group was higher than the control and 250 nM EGF treatment groups. The LAP activity of the treatment groups became 78~86% of the control group. The LAP protein (53 kDa) was separated by 10% SDS-PAGE and analyzed by Western blotting. It showed that the LAP content of the EGF treatment groups was 86~93% of the control group. In conclusion, 250 nM EGF treatment increased total cellular RNA and 5 nM EGF treatment increased total cellular protein content, however, EGF treatments (5 to 250 nM) slightly decreased the activity and content of LAP on the cell membranes of Caco-2 cells.