title: A truncated analogue of CCL2 mediates anti-fibrotic effects on murine fibroblasts independently of CCR2.
abstract: The truncated [1+9-76] CCL2 analogue, also known as 7ND, has been described in numerous reports as an anti-inflammatory and anti-fibrotic agent in a wide spectrum of animal models, e.g. models of cardiovascular disease, graft versus host disease and bleomycin-induced pulmonary fibrosis. 7ND has been reported to function as a competitive inhibitor of CCL2 signaling via CCR2 in human in vitro systems. In contrast, the mechanistic basis of 7ND action in animal models has not been previously reported. Here we have studied how 7ND interacts with CCL2 and CCR2 of murine origin. Surprisingly, 7ND was shown to be a weak inhibitor of murine CCL2/CCR2 signaling and displaced murine CCL2 (JE) from the receptor with a K(i)>1 μM. Using surface plasmon resonance, we found that 7ND binds murine CCL2 with a K(d) of 670 nM, which may indicate that 7ND inhibits murine CCL2/CCR2 signaling by a dominant negative mechanism rather than by competitive binding to the CCR2 receptor. In addition we observed that sub-nanomolar levels of 7ND mediate anti-fibrotic effects in CCR2 negative fibroblasts cultured from fibrotic lung of bleomycin-induced mice. Basal levels of extracellular matrix proteins were reduced (collagen type 1 and fibronectin) as well as expression levels of α-smooth muscle actin and CCL2. Our conclusion from these data is that the previously reported effects of 7ND in murine disease models most probably are mediated via mechanisms independent of CCR2.
authors.affiliation: Swedish Orphan Biovitrum AB, Stockholm, Sweden.
authors.forename: Christina
authors.initials: C
authors.lastname: Kalderén
authors.forename: Margareta
authors.initials: M
authors.lastname: Forsgren
authors.forename: Ulla
authors.initials: U
authors.lastname: Karlström
authors.forename: Karin
authors.initials: K
authors.lastname: Stefansson
authors.forename: Robert
authors.initials: R
authors.lastname: Svensson
authors.forename: Magnus M
authors.initials: MM
authors.lastname: Berglund
authors.forename: Gunnar
authors.initials: G
authors.lastname: Palm
authors.forename: Martin
authors.initials: M
authors.lastname: Selander
authors.forename: Maj
authors.initials: M
authors.lastname: Sundbom
authors.forename: Joakim
authors.initials: J
authors.lastname: Nilsson
authors.forename: Annelie
authors.initials: A
authors.lastname: Sjögren
authors.forename: Kristina
authors.initials: K
authors.lastname: Zachrisson
authors.forename: Stefan Svensson
authors.initials: SS
authors.lastname: Gelius
mesh_terms: D000199:Actins / Q000235:genetics / Q000378:metabolism; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000633:toxicity; D001761:Bleomycin / Q000633:toxicity; D002460:Cell Line; D018932:Chemokine CCL2 / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D003001:Cloning, Molecular; D006224:Cricetinae; D005260:Female; D005347:Fibroblasts / Q000187:drug effects*; D005355:Fibrosis / Q000139:chemically induced*; D005786:Gene Expression Regulation / Q000502:physiology; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D054390:Receptors, CCR2 / Q000235:genetics / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism
chemical_list: D000199:Actins; D000903:Antibiotics, Antineoplastic; C491768:Ccl2 protein, mouse; C515335:Ccr2 protein, mouse; D018932:Chemokine CCL2; D012333:RNA, Messenger; D054390:Receptors, CCR2; D016212:Transforming Growth Factor beta; C544013:alpha-smooth muscle actin, mouse; D001761:Bleomycin