title: Insulin-induced ectodomain shedding of heparin-binding epidermal growth factor-like growth factor in adipocytes in vitro.
abstract: Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is synthesized as a type I transmembrane protein, which is proteolytically cleaved to release a soluble form via members of the a disintegrin and metalloproteinase (ADAM) family of proteolytic enzymes. This study was designed to elucidate the molecular mechanism underlying insulin-induced HB-EGF shedding in adipocytes in vitro. The 3T3-L1 adipocytes with stable expression of alkaline phosphatase (AP)-tagged proHB-EGF (3T3-L1/HB-EGF-AP adipocytes) were developed and AP activities of conditioned media were determined. Using 3T3-L1/HB-EGF-AP adipocytes, we demonstrated that insulin induces HB-EGF shedding in differentiated 3T3-L1 adipocytes in a dose- and time-dependent manner. There is no significant increase in insulin-induced HB-EGF shedding in undifferentiated 3T3-L1 preadipocytes. Studies with metalloprotease inhibitors suggested that insulin-induced HB-EGF shedding in adipocytes is mediated at least in part via ADAM17. Treatment with recombinant HB-EGF results in a dose- and time-dependent increase in HB-EGF shedding in adipocytes, which is significantly suppressed by pharmacologic blockade of ADAM17 (P < 0.01). Moreover, insulin-induced HB-EGF shedding in adipocytes is significantly inhibited by AG1478, an EGF receptor antagonist (P < 0.01). This study provides in vitro evidence that insulin induces HB-EGF shedding in 3T3-L1 adipocytes. Our data also suggest the role of ADAM17 in insulin-induced HB-EGF shedding in adipocytes.
authors.affiliation: Department of Molecular Medicine and Metabolism, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Takanobu
authors.initials: T
authors.lastname: Yamamoto
authors.forename: Takayoshi
authors.initials: T
authors.lastname: Suganami
authors.forename: Minako
authors.initials: M
authors.lastname: Kiso-Narita
authors.forename: Peggy A
authors.initials: PA
authors.lastname: Scherle
authors.forename: Yasutomi
authors.initials: Y
authors.lastname: Kamei
authors.forename: Mitsuaki
authors.initials: M
authors.lastname: Isobe
authors.forename: Shigeki
authors.initials: S
authors.lastname: Higashiyama
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Ogawa
mesh_terms: D041721:3T3-L1 Cells; D051722:ADAM Proteins / Q000378:metabolism*; D000072198:ADAM17 Protein; D017667:Adipocytes / Q000187:drug effects / Q000378:metabolism*; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D002454:Cell Differentiation; D066246:ErbB Receptors / Q000037:antagonists & inhibitors; D066257:Heparin-binding EGF-like Growth Factor; D007328:Insulin / Q000378:metabolism* / Q000494:pharmacology; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D051379:Mice; D011799:Quinazolines; D011994:Recombinant Proteins; D020032:Tyrphostins / Q000494:pharmacology
chemical_list: C587884:Hbegf protein, mouse; D066257:Heparin-binding EGF-like Growth Factor; D007328:Insulin; D036341:Intercellular Signaling Peptides and Proteins; D011799:Quinazolines; D011994:Recombinant Proteins; D020032:Tyrphostins; C101044:RTKI cpd; D066246:ErbB Receptors; D000469:Alkaline Phosphatase; D051722:ADAM Proteins; D000072198:ADAM17 Protein; C000606463:Adam17 protein, mouse