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The small peptide OGP(10-14) acts through Src kinases and RhoA pathways in Mo-7e cells: Morphologic and immunologic evaluation

Department of Human Morphology and Applied Biology, Section of Histology and General Embryology, University of Pisa, Via Roma 67, Pisa, Italy.
Medical science monitor: international medical journal of experimental and clinical research (Impact Factor: 1.22). 07/2008; 14(6):BR103-108.
Source: PubMed

ABSTRACT Osteogenic growth peptide (OGP) is an endogenous tetradecapeptide present in micromolar concentrations in mammalian serum; its carboxy-terminal pentapeptide, OGP(10-14), represents its physiologically active fragment. OGP(10-14) induces proliferation and differentiation in fibroblast and osteoblast cell lines, and it enhances hematopoiesis in vitro and in vivo. The signaling pathways triggered by OGP(10-14) are not yet fully known. In the present report, we evaluated the effect of OGP(10-14) on differentiation of a cancer megakaryoblast cell line and its involvement on RhoA and Src family kinases signaling pathway.
Cell proliferation of the Mo-7e line was evaluated using the MTT test. Mo-7e differentiation was evaluated by microscopic observation of cell morphology and by expression of the factor VIII-related antigen. Involvement of RhoA and Src kinases on signaling pathways triggered by OGP(10-14) was analyzed using RhoA and Src family kinase (SFK) inhibitors (C3 and PP2) and an immunoperoxidase technique.
OGP(10-14) induces expression of the factor VIII-related antigen, morphologic changes indicative of megakaryocytic differentiation, and a down-regulation of the Fyn Src kinase. These OGP(10-14) effects were prevented by C3 and enhanced by PP2.
The anti-proliferative and pro-differentiating activities of OGP(10-14) on thrombopoietin (TPO)-primed Mo-7e cells are mediated by RhoA and Src kinase pathways as demonstrated by the use of C3 and PP2.

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    • "The physiologically active form of OGP, which corresponds to its C-terminal pentapeptide sequence YGFGG (OGP10–14), is generated from full-length OGP by proteolytic cleavage [5]. This fragment directly interacts with cell membrane receptors, activating the MAP kinase, Src and RhoA signaling pathways [9] [10] [11]. Upon intravenous administration, synthetic OGP and OGP10–14 were shown to promote increased bone mass and fracture healing in vivo [12] [13]. "
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    • "PP2 SFK inhibitor was dissolved in DMSO and added to cell cultures 40 minutes before adding OGP(10-14). DMSO was used at a concentration that does not interfere with M07-e cell proliferation [9]. Experiments were performed at 1, 72 and 144 hours on the basis of the previous results, showing in M07-e cells RhoA activation and signs of differentiation at 1h and 144h, respectively [9]. "
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    ABSTRACT: Osteogenic growth peptide (OGP) is a 14-mer peptide found in relevant concentration in blood, and its carboxy-terminal fragment [OGP(10-14)] represents the active portion of the full-length peptide. In addition to stimulating bone formation, OGP(10-14) shows hematological activity. In fact, it highly enhances hematopoiesis-affecting stem progenitors. Moreover, OGP(10-14) reduces the growth and induces the differentiation of the hematological tumour cell line trombophoietin(TPO)-primed M07-e by interfering with RhoA and Src kinase pathways. In the present report, we went deeper into this mechanism and evaluated the possible interference of the OGP(10-14) signal pathway with TGFβ1 and TPO receptor Mpl. In OGP(10-14)-treated M07-e cells cultured with or without RhoA and Src kinases inhibitors (C3 and PP2), expression of TGFβ1, Mpl, and Src kinases was analyzed by immunoperoxidase technique. Activated RhoA expression was studied using the G-LISA™ quantitative test. In M07-e cells, both OGP(10-14) and PP2 activate RhoA, inhibit Src kinases, reduce Mpl expression and increase TGFβ1 expression. OGP(10-14) and PP2 show the same behavior, causing an additive effect when associated. OGP(10-14) induces TPO-primed M07-e cells differentiation through RhoA/TGFβ1/SFKs signalling pathway. In particular OGP(10-14) acts as a Src inhibitor, showing the same effects of PP2.
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