In Vitro Release of Bioactive Bone Morphogenetic Proteins (GDF5, BB-1, and BMP-2) from a PLGA Fiber-Reinforced, Brushite-Forming Calcium Phosphate Cement

Francesca Gunnella; Elke Kunisch; Victoria Horbert; Stefan Maenz; Jörg Bossert; Klaus D Jandt; Frank Plöger; Raimund W Kinne

doi:10.3390/pharmaceutics11090455

In Vitro Release of Bioactive Bone Morphogenetic Proteins (GDF5, BB-1, and BMP-2) from a PLGA Fiber-Reinforced, Brushite-Forming Calcium Phosphate Cement

Pharmaceutics. 2019 Sep 3;11(9):455. doi: 10.3390/pharmaceutics11090455.

Authors

Francesca Gunnella¹, Elke Kunisch¹, Victoria Horbert¹, Stefan Maenz^{2

3}, Jörg Bossert², Klaus D Jandt^{2

3

4}, Frank Plöger⁵, Raimund W Kinne⁶

Affiliations

¹ Experimental Rheumatology Unit, Department of Orthopedics, Jena University Hospital, Waldkrankenhaus "Rudolf Elle", Klosterlausnitzer Str. 81, 07607 Eisenberg, Germany.
² Chair of Materials Science, Otto Schott Institute of Materials Research, Friedrich Schiller University Jena, 07743 Jena, Germany.
³ Jena Center for Soft Matter (JCSM), Friedrich Schiller University Jena, 07743 Jena, Germany.
⁴ Jena School for Microbial Communication (JSMC), Friedrich Schiller University Jena, 07743 Jena, Germany.
⁵ BIOPHARM GmbH, 69115 Heidelberg, Germany.
⁶ Experimental Rheumatology Unit, Department of Orthopedics, Jena University Hospital, Waldkrankenhaus "Rudolf Elle", Klosterlausnitzer Str. 81, 07607 Eisenberg, Germany. raimund.w.kinne@med.uni-jena.de.

Abstract

Bone regeneration of sheep lumbar osteopenia is promoted by targeted delivery of bone morphogenetic proteins (BMPs) via a biodegradable, brushite-forming calcium-phosphate-cement (CPC) with stabilizing poly(l-lactide-co-glycolide) acid (PLGA) fibers. The present study sought to quantify the release and bioactivity of BMPs from a specific own CPC formulation successfully used in previous in vivo studies. CPC solid bodies with PLGA fibers (0%, 5%, 10%) containing increasing dosages of GDF5, BB-1, and BMP-2 (2 to 1000 µg/mL) were ground and extracted in phosphate-buffered saline (PBS) or pure sheep serum/cell culture medium containing 10% fetal calf serum (FCS; up to 30/31 days). Released BMPs were quantified by ELISA, bioactivity was determined via alkaline phosphatase (ALP) activity after 3-day exposure of different osteogenic cell lines (C2C12; C2C12BRlb with overexpressed BMP-receptor-1b; MCHT-1/26; ATDC-5) and via the influence of the extracts on the expression of osteogenic/chondrogenic genes and proteins in human adipose tissue-derived mesenchymal stem cells (hASCs). There was hardly any BMP release in PBS, whereas in medium + FCS or sheep serum the cumulative release over 30/31 days was 11-34% for GDF5 and 6-17% for BB-1; the release of BMP-2 over 14 days was 25.7%. Addition of 10% PLGA fibers significantly augmented the 14-day release of GDF5 and BMP-2 (to 22.6% and 43.7%, respectively), but not of BB-1 (13.2%). All BMPs proved to be bioactive, as demonstrated by increased ALP activity in several cell lines, with partial enhancement by 10% PLGA fibers, and by a specific, early regulation of osteogenic/chondrogenic genes and proteins in hASCs. Between 10% and 45% of bioactive BMPs were released in vitro from CPC + PLGA fibers over a time period of 14 days, providing a basis for estimating and tailoring therapeutically effective doses for experimental and human in vivo studies.

Keywords: BB-1; BMP-2; GDF5; PLGA fiber-reinforced; bioactive bone morphogenetic proteins; brushite-forming calcium phosphate cement; in vitro release.