title: Endocytosis of a functionally enhanced GFP-tagged transferrin receptor in CHO cells.
abstract: The endocytosis of transferrin receptor (TfR) has served as a model to study the receptor-targeted cargo delivery system for cancer therapy for many years. To accurately evaluate and optically measure this TfR targeting delivery in vitro, a CHO cell line with enhanced green fluorescent protein (EGFP)-tagged human TfR was established. A chimera of the hTfR and EGFP was engineered by fusing EGFP to the amino terminus of hTfR. Data were provided to demonstrate that hTfR-EGFP chimera was predominantly localized on the plasma membrane with some intracellular fluorescent structures on CHO cells and the EGFP moiety did not affect the endocytosis property of hTfR. Receptor internalization occurred similarly to that of HepG2 cells expressing wild-type hTfR. The internalization percentage of this chimeric receptor was about 81 ± 3% of wild type. Time-dependent co-localization of hTfR-EGFP and PE-conjugated anti-hTfR mAb in living cells demonstrated the trafficking of mAb-receptor complexes through the endosomes followed by segregation of part of the mAb and receptor at the late stages of endocytosis. The CHO-hTfR cells preferentially took up anti-hTfR mAb conjugated nanoparticles. This CHO-hTfR cell line makes it feasible for accurate evaluation and visualization of intracellular trafficking of therapeutic agents conjugated with transferrin or Abs targeting the hTfRs.
authors: Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors: Qi
authors: Q
authors: He
authors: Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China; Department of Clinical Laboratory, the First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, China.
authors: Xiaoxu
authors: X
authors: Sun
authors: Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors: Chong
authors: C
authors: Chu
authors: Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors: Qing
authors: Q
authors: Jiang
authors: Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors: Huifen
authors: H
authors: Zhu
authors: Department of Nuclear Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, Hubei, China.
authors: Yong
authors: Y
authors: He
authors: Beijing Pushikang Pharmaceutical Technology Co., Ltd, Beijing, China.
authors: Tingting
authors: T
authors: Yue
authors: Beijing Pushikang Pharmaceutical Technology Co., Ltd, Beijing, China.
authors: Ruibo
authors: R
authors: Wang
authors: Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors: Ping
authors: P
authors: Lei
authors: Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
authors: Guanxin
authors: G
authors: Shen
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D015153:Blotting, Western; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D017931:DNA Primers / Q000235:genetics; D004705:Endocytosis / Q000502:physiology*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D053758:Nanoparticles / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011990:Receptors, Transferrin / Q000378:metabolism* / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D018709:Statistics, Nonparametric
chemical_list: D000911:Antibodies, Monoclonal; D017931:DNA Primers; D011990:Receptors, Transferrin; D011993:Recombinant Fusion Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins