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Novel Dose Dependent Positive Feedback Loop in Endothelial Cells: A Response to rhBMP2 Treatment
Author(s) -
Hussein Khaled,
Helwa Inas,
Zakhary Ibrhaim,
Elrefai Rami,
Al-Shabrawey Mohamed,
Sharawy Mohamed
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.728.3
Subject(s) - bone morphogenetic protein 2 , downregulation and upregulation , secretion , in vivo , umbilical vein , endogeny , chemistry , andrology , smad , bone morphogenetic protein , medicine , in vitro , biology , transforming growth factor , biochemistry , gene , microbiology and biotechnology
Recently, we reported that Endothelial Cells (ECs) could be a potential source of endogenous BMP2 which is essential for successful bone regeneration in vivo. The aim of this study was to investigate the capability of ECs to respond to exogenous BMP2 by secretion of endogenous BMP2. Methods Human Umbilical Vein Endothelial Cells were treated with serial concentrations of rhBMP2 (25, 50 and 100ng/ml of rhBMP2) for only 3 hours and the cultured cells were maintained for 12 hours. Cell lysates were collected at 5 minutes, 3hours, 6 hours and 12 hours time points after rhBMP2 treatment. ELISA for BMP2 was done to detect its secretion in supernatant at the 6 and 12 hours time points. Real time PCR was done to detect mRNA of BMP2, BMP2R1B and SMAD1/5. Western blot analysis was done to detect the phosphorylation of SMAD1/5/8. Results Exogenous rhBMP2 significantly induced BMP2 secretion only in the 100ng rhBMP2 treatment at the 6 and 12hours time points consistent with a gradual upregulation of BMP2 mRNA. BMP2R1B, SMAD1 and SMAD5 gene expression were also significantly upregulated. SMAD1/5/8 was consistently phospholyated in the 5 minutes and 3 hours time points. Conclusion ECs respond to 100ng/ml rhBMP2 treatment by secreting BMP2 via a SMAD dependent pathway. These results suggest that ECs treated in‐vitro with rhBMP2 could be used as a source of BMP2 in‐vivo to optimize bone regeneration in challenging clinical situations. Grant Funding Source : Ministry of Higher Education, Egypt