Recombinant bmp6

Manufactured by R&D Systems

Recombinant BMP6 is a laboratory reagent produced by R&D Systems. It is a member of the bone morphogenetic protein (BMP) family, which are secreted signaling proteins involved in the regulation of bone and cartilage development. The recombinant form of BMP6 is intended for use in research applications.

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3 protocols using recombinant bmp6

Hepcidin Promoter Activity Assay

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A pGL2-basic reporter vector (Promega, Madison, WI, USA), harbouring a 2.9 kb fragment of human hepcidin promoter (Hep-Luc), was used to analyse the hepcidin promoter activity by luciferase assay in Hep3B cells cotransfected with wild-type or mutant HJV expressing vectors, as already reported 19 (link) and as detailed in the Supporting Information section. When indicated, transfected cells were incubated for 3 hrs with 10 ng/ml of recombinant BMP6 (R&D System, Minneapolis, MN). Exogenous HJV levels were measured by qRT-PCR on transfected Hep3B cells (see Data S1). Experiments were performed in triplicate. Two-tail Student's t-test was used for statistical analysis.

Rausa M., Ghitti M., Pagani A., Nai A., Campanella A., Musco G., Camaschella C, & Silvestri L. (2015). Identification of TMPRSS6 cleavage sites of hemojuvelin. Journal of Cellular and Molecular Medicine, 19(4), 879-888.

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Regulation of Osteogenesis by BMP6 in BMSC

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The conditioned medium of BMSC culture was collected 10 and 21 days after induction of osteogenesis and then stored in a −20°C freezer before analysis. To collect serum from mice, blood was drawn from the submandibular vein using 4‐mm lancets (Fisher Scientific, Hampton, NH) and centrifuged in Vacutainer blood collection tubes (BD Biosciences) at 2,000g for 10 minutes. The supernatant was then stored in a −20°C freezer before analysis. Levels of BMP6 in BMSC culture or mouse serum were measured by ELISA kits containing human‐ (Raybiotech, Norcross, GA) or mouse‐ (MyBioSource, San Diego, CA) specific antibodies following the manufacturer's instructions.
To determine if BMP6 is a target molecule of interest involved in the regulation of osteogenesis of T1DM‐ or non‐T1DM‐BMSCs, the cell in osteogenic culture was supplemented with or without 10 ng/ml of recombinant BMP6 or neutralized with or without 2 μg/ml of BMP6 antibody (R&D, Minneapolis, MN).

Wang J.F., Lee M., Tsai T., Leiferman E.M., Trask D.J., Squire M.W, & Li W. (2019). Bone Morphogenetic Protein‐6 Attenuates Type 1 Diabetes Mellitus‐Associated Bone Loss. Stem Cells Translational Medicine, 8(6), 522-534.

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Neuronal Cell Culture Protocol

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Earle’s Balanced Salt Solution (EBSS), Hank’s Balanced Salt Solution (HBSS), Dulbecco’s Modified Eagle’s Media (DMEM), 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), Modified Eagle’s Media (MEM), Neurobasal Media, GlutaMAX, B27 supplement, Trypsin-EDTA and Penicillin-Streptomycin were obtained from Invitrogen. Papain enzyme, poly-L-lysine, cytosine β-D-arabinofuranoside (Ara-C), losartan potassium, bovine serum albumin (BSA, minimum 98%), fluorescein isothio-cyanate conjugate bovine albumin, sodium deoxycholate (DOC, minimum 97%) and Evans blue (EB) were all obtained from Sigma Aldrich. Serum Extender was obtained from BD Biosciences. TGF-β type 1 receptor ALK4, 5 and 7 selective inhibitor SB431542 and the ALK5 selective inhibitor SJN2511 were obtained from Tocris Biosciences. Recombinant TGF-β1 was obtained from Peprotech and recombinant BMP6 was obtained from R&D Biosystems.

Bar-Klein G., Cacheaux L.P., Kamintsky L., Prager O., Weissberg I., Schoknecht K., Cheng P., Kim S.Y., Wood L., Heinemann U., Kaufer D, & Friedman A. (2014). Losartan prevents acquired epilepsy via TGF-β signaling suppression. Annals of neurology, 75(6), 864-875.

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