Peng Zhao1, Bo Chen2, Lushen Li3, Hao Wu4, Yan Li1, Baxter Shaneen3, Xi Zhan5, Ning Gu6. 1. State Key Laboratory of Bioelectronics, Jiangsu Key Laboratory for Biomaterials and Devices, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, PR China; Collaborative Innovation Center of Suzhou Nano Science and Technology, Suzhou 215123, PR China. 2. State Key Laboratory of Bioelectronics, Jiangsu Key Laboratory for Biomaterials and Devices, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, PR China; Materials Science and Devices Institute, Suzhou University of Science and Technology, 1 Kerui Road, Suzhou, Jiangsu 215009, PR China. 3. Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, Baltimore, MD 21201, USA. 4. School of Biomedical Engineering and Informatics, Nanjing Medical University, Nanjing 210029, PR China. 5. Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, Baltimore, MD 21201, USA. Electronic address: xzhan@som.umaryland.edu. 6. State Key Laboratory of Bioelectronics, Jiangsu Key Laboratory for Biomaterials and Devices, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, PR China; Collaborative Innovation Center of Suzhou Nano Science and Technology, Suzhou 215123, PR China. Electronic address: guning@seu.edu.cn.
Abstract
BACKGROUND: Magnetic nanoparticles (MNPs) have been widely used in biomedical applications. Proper control of the duration of MNPs in circulation promises to improve further their applications, in particularly drug delivery. It is known that the uptake of tissue-associated MNPs is mainly carried out by macrophages. Yet, the molecular mechanism to control MNPs internalization in macrophages remains to be elusive. Missing-in-metastasis (MIM) is a scaffolding protein that is highly expressed in macrophages and regulates receptor-mediated endocytosis. We hypothesize that uptake of MNPs may also involve the function of MIM. METHODS: We investigated the effect of MIM expression on the intracellular trafficking of MNPs by transmission electronic microscopy, flow cytometry, o-phenanthroline photometric analysis, Perl's staining, immunofluorescence microscopy and co-immunoprecipitation. To explore the molecular events in MIM-mediated MNPs uptake, we examined the effect of MNPs on the interaction of MIM with clathrin, Rab5 and Rab7. RESULTS: Uptake of MNPs was significantly enhanced in cells overexpressing MIM. Upon exposure to MNPs, MIM was associated with clathrin light chain in endocytic vesicles and Rab7, a protein that regulates late endosomes. However, MNPs caused dissociation of MIM with Rab5, an early endosome-associated protein. CONCLUSIONS: MIM regulates internalization of MNPs via promoting their trafficking from plasma membrane to late endosomes. GENERAL SIGNIFICANCE: Our data unveiled a novel pathway which MNPs internalization and intracellular trafficking in macrophages. This new pathway may allow us to control the uptake of MNPs within cells by targeting MIM, thereby improving their medical applications.
BACKGROUND: Magnetic nanoparticles (MNPs) have been widely used in biomedical applications. Proper control of the duration of MNPs in circulation promises to improve further their applications, in particularly drug delivery. It is known that the uptake of tissue-associated MNPs is mainly carried out by macrophages. Yet, the molecular mechanism to control MNPs internalization in macrophages remains to be elusive. Missing-in-metastasis (MIM) is a scaffolding protein that is highly expressed in macrophages and regulates receptor-mediated endocytosis. We hypothesize that uptake of MNPs may also involve the function of MIM. METHODS: We investigated the effect of MIM expression on the intracellular trafficking of MNPs by transmission electronic microscopy, flow cytometry, o-phenanthroline photometric analysis, Perl's staining, immunofluorescence microscopy and co-immunoprecipitation. To explore the molecular events in MIM-mediated MNPs uptake, we examined the effect of MNPs on the interaction of MIM with clathrin, Rab5 and Rab7. RESULTS: Uptake of MNPs was significantly enhanced in cells overexpressing MIM. Upon exposure to MNPs, MIM was associated with clathrin light chain in endocytic vesicles and Rab7, a protein that regulates late endosomes. However, MNPs caused dissociation of MIM with Rab5, an early endosome-associated protein. CONCLUSIONS:MIM regulates internalization of MNPs via promoting their trafficking from plasma membrane to late endosomes. GENERAL SIGNIFICANCE: Our data unveiled a novel pathway which MNPs internalization and intracellular trafficking in macrophages. This new pathway may allow us to control the uptake of MNPs within cells by targeting MIM, thereby improving their medical applications.
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