Calreticulin attenuates TGF-beta extracellular matrix for tissue regeneration
Miguel A. Manzanares, Ana Tellechea, Leslie I. Gold.
New York University School of Medicine, New York, NY, USA.
Background: Calreticulin (CRT), an ER chaperone, has been shown to have extracellular functions via outside-in signaling. We show that both intracellular CRT and extracellular CRT are required for the synthesis and release of the extracellular matrix (ECM) proteins, collagen, fibronectin, and elastin. Using diabetic mice, topically applied CRT for 4 days post-wounding healed full thickness excisional wounds by tissue regeneration characterized by abundant neo-dermis, epidermal appendage neogenesis, and no scarring. By day 14 post-wounding upon epidermal closure, hair follicles evaginate from the epidermis into the dermis, which show pigmentation by day 21 post-wounding. In vitro, we have shown that induction of ECM proteins by CRT in fibroblasts [to fill in the wound defect] is via TGF-beta signaling. However, as TGF-beta critically induces scarring and CRT heals by tissue regeneration, we interrogated potential mechanisms that might be involved in CRT modulation of TGF-beta-induced fibrosis/scarring using human dermal fibroblasts. Results: Inhibiting protein translation with cycloheximide, we show that CRT induces a faster turn-over rate of ECM proteins than TGF-beta. On a molar basis, TGF-beta induces between 1.7-3.4 higher levels ECM proteins than CRT. Over time, the production of ECM proteins by CRT decreases earlier than by TGF-beta. The connection between TGF-β and Wnt pathways plays a role in fibrosis by inducing ECM proteins. As a mechanism involved in fibrosis, TGF-beta was shown to downregulate DKK-1, an inhibitor of Wnt signaling. In contrast, CRT increases DKK-1 as a potential mechanism for decreasing Wnt-related beta-catenin activation. Furthermore, at 3h, CRT induces miR29, known to block collagen and fibronectin transcription. Conclusion: Revealing mechanisms involved in CRT attenuation of TGF-beta induced scarring should implicate new therapeutic targets for tissue regeneration of chronic wounds.
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