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Mechanism of Absorption and Transportation of Ovotransferrin in the Intestine
Author(s) -
Shirkhani Rebin Mohammad,
Lee Eun Joo,
Talukder Jamil
Publication year - 2018
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2018.32.1_supplement.747.17
Subject(s) - ovotransferrin , transcytosis , transferrin , lactoferrin , chemistry , transferrin receptor , internalization , absorption (acoustics) , paracellular transport , biochemistry , cell , microbiology and biotechnology , membrane , endocytosis , biology , materials science , composite material , permeability (electromagnetism)
Background Ovotransferrin (OTF) is an iron‐binding protein present in eggs. It belongs to the transferrin family and shows sequence‐homology with mammalian transferrin and lactoferrin. This protein has been demonstrated to possess bioactive characteristics such as delivering iron to cells and inhibiting bacterial multiplication. However, the absorption and transportation of OTF through the intestinal epithelial cells remained unknown. Aims The objectives of this study were to determine the absorption and transportation mechanism of OTF through rat intestinal epithelial cells (IEC‐6). Methods IEC‐6 monolayer cells were grown on 6‐well transwell membrane with DMEM, bovine fetal serum, β‐hydroxybutyric acid, antibiotics, and insulin in a humidified CO 2 incubator at 37°C. 10‐day post‐confluent cells were used for absorption and transport studies. OTF was loaded on the apical part and DPBS was collected from the bottom at 30 min at room temperature. Trans‐epithelial transport of OTF was initially detected by SDS‐PAGE with a molecular weight of 78 kDa. In order to carry out the real‐time uptake and transcytosis of OTF, 1 mg OTF/ml of DPBS was loaded and DPBS was collected from the bottom at different time intervals. Finally, the amount of OTF transported through the IEC‐6 cells following internalization was quantified by ELISA. Results and Conclusion The OTF destined to the cell membrane and internalized was visualized by immunofluorescent and laser confocal microscopy. Microscopic images clearly showed the accumulation of OTF on the cell membrane provided the evidence of OTF‐receptors' presence on the plasma membrane. Also, OTF was detected within the cytoplasm of IEC‐6 cells, revealed that it was internalized by receptor‐mediated endocytosis. ELISA results demonstrated that OTF can be taken up by IEC‐6 cells as an intact protein from the apical surface and transported to the basolateral surface through trans‐epithelial exocytosis mechanism. The maximum internalization and exocytosis were observed at 90 min. These processes were time and concentration dependent, as the transferrin and lactoferrin do. Therefore, it is concluded that absorption and trans‐epithelial transportation of OTF accomplished by receptor‐mediated transcytosis in intestinal epithelial cells.