Publications by authors named "E R Balmayor"

45 Publications

Gene therapy for bone healing: lessons learned and new approaches.

Transl Res 2021 May 5. Epub 2021 May 5.

Rehabilitation Medicine Research Center, Mayo Clinic, Rochester, Minnesota; Musculoskeletal Gene Therapy Research Laboratory, Mayo Clinic, Rochester, Minnesota; IBE, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, Maastricht, Netherlands.

Although gene therapy has its conceptual origins in the treatment of Mendelian disorders, it has potential applications in regenerative medicine, including bone healing. Research into the use of gene therapy for bone healing began in the 1990s. Prior to this period, the highly osteogenic proteins bone morphogenetic protein (BMP)-2 and -7 were cloned, produced in their recombinant forms and approved for clinical use. Despite their promising osteogenic properties, the clinical usefulness of recombinant BMPs is hindered by delivery problems that necessitate their application in vastly supraphysiological amounts. This generates adverse side effects, some of them severe, and raises costs; moreover, the clinical efficacy of the recombinant proteins is modest. Gene delivery offers a potential strategy for overcoming these limitations. Our research has focused on delivering a cDNA encoding human BMP-2, because the recombinant protein is Food and Drug Administration approved and there is a large body of data on its effects in people with broken bones. However, there is also a sizeable literature describing experimental results obtained with other transgenes that may directly or indirectly promote bone formation. Data from experiments in small animal models confirm that intralesional delivery of BMP-2 cDNA is able to heal defects efficiently and safely while generating transient, local BMP-2 concentrations 2-3 log orders less than those needed by recombinant BMP-2. The next challenge is to translate this information into a clinically expedient technology for bone healing. Our present research focuses on the use of genetically modified, allografted cells and chemically modified messenger RNA.
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http://dx.doi.org/10.1016/j.trsl.2021.04.009DOI Listing
May 2021

Transcript-Activated Coatings on Titanium Mediate Cellular Osteogenesis for Enhanced Osteointegration.

Mol Pharm 2021 03 25;18(3):1121-1137. Epub 2021 Jan 25.

IBE, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, 6200 MD Maastricht, The Netherlands.

Osteointegration is one of the most important factors for implant success. Several biomolecules have been used as part of drug delivery systems to improve implant integration into the surrounding bone tissue. Chemically modified mRNA (cmRNA) is a new form of therapeutic that has been used to induce bone healing. Combined with biomaterials, cmRNA can be used to develop transcript-activated matrices for local protein production with osteoinductive potential. In this study, we aimed to utilize this technology to create bone morphogenetic protein 2 (BMP2) transcript-activated coatings for titanium (Ti) implants. Therefore, different coating methodologies as well as cmRNA incorporation strategies were evaluated. Three different biocompatible biomaterials were used for the coating of Ti, namely, poly-d,l-lactic acid (PDLLA), fibrin, and fibrinogen. cmRNA-coated Ti disks were assayed for transfection efficiency, cmRNA release, cell viability and proliferation, and osteogenic activity . We found that cmRNA release was significantly delayed in Ti surfaces previously coated with biomaterials. Consequently, the transfection efficiency was greatly improved. PDLLA coating improved the transfection efficiency in a concentration-dependent manner. Lower PDLLA concentration used for the coating of Ti resulted in higher transfection efficiency. Fibrin and fibrinogen coatings showed even higher transfection efficiencies compared to all PDLLA concentrations. In those disks, not only the expression was up to 24-fold higher but also the peak of maximal expression was delayed from 24 h to 5 days, and the duration of expression was also extended until 7 days post-transfection. For fibrin, higher transfection efficiencies were obtained in the coatings with the lowest thrombin amounts. Accordingly, fibrinogen coatings gave the best results in terms of cmRNA transfection. All biomaterial-coated Ti surfaces showed improved cell viability and proliferation, though this was more noticeable in the fibrinogen-coated disks. The latter was also the only coating to support significant amounts of BMP2 produced by C2C12 cells . Osteogenesis was confirmed using BMP2 cmRNA fibrinogen-coated Ti disks, and it was dependent of the cmRNA amount present. Alkaline phosphatase (ALP) activity of C2C12 increased when using fibrinogen coatings containing 250 ng of cmRNA or more. Similarly, mineralization was also observed that increased with increasing cmRNA concentration. Overall, our results support fibrinogen as an optimal material to deliver cmRNA from titanium-coated surfaces.
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http://dx.doi.org/10.1021/acs.molpharmaceut.0c01042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7927143PMC
March 2021

A Graded, Porous Composite of Natural Biopolymers and Octacalcium Phosphate Guides Osteochondral Differentiation of Stem Cells.

Adv Healthc Mater 2021 03 14;10(6):e2001692. Epub 2021 Jan 14.

Experimental Trauma Surgery, Klinikum rechts der Isar, Technical University of Munich, Munich, 81675, Germany.

Lesions involving the osteochondral unit are difficult to treat. Biomimetic scaffolds are previously shown as promising alternative. Such devices often lack multiple functional layers that mimic bone, cartilage, and the interface. In this study, multilayered scaffolds are developed based on the use of natural extracellular matrix (ECM)-like biopolymers. Particular attention is paid to obtain a complex matrix that mimics the native osteochondral transition. Porous, sponge-like chitosan-collagen-octacalcium phosphate (OCP) scaffolds are obtained. Collagen content increases while the amount of OCP particles decreases toward the cartilage layer. The scaffolds are bioactive as a mineral layer is deposited containing hydroxyapatite at the bony side. The scaffolds stimulate proliferation of human adipose-derived mesenchymal stem cells, but the degree of proliferation depends on the cell seeding density. The scaffolds give rise to a zone-specific gene expression. RUNX2, COL1A1, BGLAP, and SPP1 are upregulated in the bony layer of the scaffold. SOX9 is upregulated concomitant with COL2A1 expression in the cartilage zone. Mineralization in presence of the cells is prominent in the bone area with Ca and P steadily increasing over time. These results are encouraging for the fabrication of biomimetic scaffolds using ECM-like materials and featuring gradients that mimic native tissues and their interface.
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http://dx.doi.org/10.1002/adhm.202001692DOI Listing
March 2021

Author Correction: Effect of donor age and 3D-cultivation on osteogenic differentiation capacity of adipose-derived mesenchymal stem cells.

Sci Rep 2020 Jul 31;10(1):13184. Epub 2020 Jul 31.

Department of Experimental Trauma Surgery, Klinikum rechts der Isar, Technical University Munich, Munich, Germany.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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http://dx.doi.org/10.1038/s41598-020-69876-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7395092PMC
July 2020

A New Non-invasive Technique for Measuring 3D-Oxygen Gradients in Wells During Mammalian Cell Culture.

Front Bioeng Biotechnol 2020 17;8:595. Epub 2020 Jun 17.

cBITE, MERLN Institute for Technology-Inspired Regenerative Medicine, Maastricht University, Maastricht, Netherlands.

Oxygen tension plays an important role in overall cell function and fate, regulating gene expression, and cell differentiation. Although there is extensive literature available that supports the previous statement, little information is to be found about accurate O measurements during culture. In fact, O concentration at the cell layer during culture is commonly assumed to be equal to that of the incubator atmosphere. This assumption does not consider oxygen diffusion properties, cell type, cell density, media composition, time in culture nor height of the cell culture medium column. In this study, we developed a non-invasive, optical sensor foil-based technique suitable for measuring the 3D oxygen gradient that is formed during cell culture as a result of normal cell respiration. For this propose, we created a 3D printed ramp to which surface an oxygen optode sensor foil was attached. The ramps were positioned inside the culture wells of 24 well plate prior cell seeding. This set up in conjunction with the VisiSens TD camera system allows to investigate the oxygen gradient formation during culture. Cultivation was performed with three different initial cell densities of the cell line A549 that were seeded on the plate containing the ramps with the oxygen sensors. The O gradient obtained after 96 h of culture showed significantly lower O concentrations closer to the bottom of the well in high cell density cultures compared to that of lower cell density cultures. Furthermore, it was very interesting to observe that even with low cell density culture, oxygen concentration near the cell layer was lower than that of the incubator atmosphere. The obtained oxygen gradient after 96 h was used to calculate the oxygen consumption rate (OCR) of the A549 cells, and the obtained value of ~100 fmol/h/cell matches the OCR value already reported in the literature for this cell line. Moreover, we found our set up to be unique in its ability to measure oxygen gradient formation in several wells of a cell culture plate simultaneously and in a non-invasive manner.
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http://dx.doi.org/10.3389/fbioe.2020.00595DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7313265PMC
June 2020