title: Endocytic mechanism of internalization of dietary peptide lunasin into macrophages in inflammatory condition associated with cardiovascular disease.
abstract: Cardiovascular disease (CVD) is the leading cause of death in the United States. Diet influences risk factors associated with CVD and atherosclerosis, a major vascular disease that arises from inflammation. Lunasin, a peptide derived from plant foods such as soybeans, contains a unique Arg-Gly-Asp cell-adhesion motif and inhibits the pathways involved in the inflammatory cascade. The objective was to determine the mechanism by which lunasin is internalized into human THP-1 macrophages, investigate the expression of endocytic membrane proteins in inflammatory conditions and to identify the pathways involved. While lipopolysaccharide (10 nM), vitronectin (130 nM) and a combination of these two molecules enhanced lunasin uptake and increased basal αVβ3 integrin expression, lunasin reduced αVβ3 expression by 25.5, 26.8 and 49.2%, respectively. The pretreatment of cells with brefeldin A (71 µM), an inhibitor of protein trafficking, inhibited lunasin internalization by up to 99.8%. Lunasin increased caveolin-1 expression by up to 204.8%, but did not modulate clathrin. The pretreatment of macrophages with nystatin (54 µM), an inhibitor of caveolae-dependent endocytosis, reduced lunasin internalization. The presence of amantadine (1 mM) and amiloride (1 mM), inhibitors of clathrin-mediated endocytosis and macropinocytosis, abolished lunasin cell entry. Lunasin elicited a transient reduction in intracellular levels of Ca²⁺ in LPS-induced macrophages. The results suggest that internalization of lunasin into macrophages is amplified in inflammatory conditions and is primarily mediated by endocytic mechanisms that involve integrin signaling, clathrin-coated structures and macropinosomes. Lunasin may be responsible for attenuation of CVD risk factors by interacting with pathways involved in endocytosis and inflammation.
authors.affiliation: Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, Illinois, United States of America.
authors.forename: Anthony
authors.initials: A
authors.lastname: Cam
authors.forename: Mayandi
authors.initials: M
authors.lastname: Sivaguru
authors.forename: Elvira
authors.initials: E
authors.lastname: Gonzalez de Mejia
mesh_terms: D020126:Brefeldin A / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002318:Cardiovascular Diseases / Q000276:immunology*; D051242:Caveolin 1 / Q000378:metabolism; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002966:Clathrin / Q000378:metabolism; D022163:Clathrin-Coated Vesicles / Q000378:metabolism; D004705:Endocytosis; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000276:immunology / Q000378:metabolism*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D021381:Protein Transport / Q000187:drug effects; D030262:Soybean Proteins / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D051242:Caveolin 1; D002966:Clathrin; C120514:GM2S-1 protein, soybean; D039302:Integrin alphaVbeta3; D008070:Lipopolysaccharides; D011500:Protein Synthesis Inhibitors; D030262:Soybean Proteins; D020126:Brefeldin A; D002118:Calcium
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