Deletion of the nuclear localization sequence and C-terminus of parathyroid hormone-related protein decreases osteogenesis and chondrogenesis but increases adipogenesis and myogenesis in murine bone marrow stromal cells

Blake E Hildreth 3rd; Krista M Hernon; Wessel P Dirksen; John Leong; Wachiraphan Supsavhad; Prosper N Boyaka; Thomas J Rosol; Ramiro E Toribio

doi:10.1177/2041731415609298

Deletion of the nuclear localization sequence and C-terminus of parathyroid hormone-related protein decreases osteogenesis and chondrogenesis but increases adipogenesis and myogenesis in murine bone marrow stromal cells

J Tissue Eng. 2015 Oct 12:6:2041731415609298. doi: 10.1177/2041731415609298. eCollection 2015 Jan-Dec.

Authors

Blake E Hildreth 3rd¹, Krista M Hernon², Wessel P Dirksen¹, John Leong¹, Wachiraphan Supsavhad¹, Prosper N Boyaka¹, Thomas J Rosol¹, Ramiro E Toribio²

Affiliations

¹ Department of Veterinary Biosciences, College of Veterinary Medicine, The Ohio State University, Columbus, OH, USA.
² Department of Veterinary Clinical Sciences, College of Veterinary Medicine, The Ohio State University, Columbus, OH, USA.

Abstract

The N-terminus of parathyroid hormone-related protein regulates bone marrow stromal cell differentiation. We hypothesized that the nuclear localization sequence and C-terminus are involved. MicroRNA and gene expression analyses were performed on bone marrow stromal cells from mice lacking the nuclear localization sequence and C-terminus (Pthrp^Δ/Δ ) and age-matched controls. Differentiation assays with microRNA, cytochemical/histologic/morphologic, protein, and gene expression analyses were performed. Pthrp^Δ/Δ bone marrow stromal cells are anti-osteochondrogenic, pro-adipogenic, and pro-myogenic, expressing more Klf4, Gsk-3β, Lif, Ct-1, and microRNA-434 but less β-catenin, Igf-1, Taz, Osm, and microRNA-22 (p ⩽ 0.024). Pthrp^Δ/Δ osteoblasts had less mineralization, osteocalcin, Runx2, Osx, Igf-1, and leptin (p ⩽ 0.029). Pthrp^Δ/Δ produced more adipocytes, Pparγ, and aP2, but less Lpl (p ⩽ 0.042). Pthrp^Δ/Δ cartilage pellets were smaller with less Sox9 and Pth1r, but greater Col2a1 (p ⩽ 0.024). Pthrp^Δ/Δ produced more myocytes, Des, and Myog (p ⩽ 0.021). MicroRNA changes supported these findings. In conclusion, the nuclear localization sequence and C-terminus are pro-osteochondrogenic, anti-adipogenic, and anti-myogenic.

Keywords: C-terminus; Parathyroid hormone–related protein; bone marrow stromal cell; differentiation; nuclear localization sequence.

Abstract

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