Treatment Depth Effects of Combined Magnetic Field Technology using a Commercial Bone Growth Stimulator

Lumbar spinal fusion is one of the more common spinal surgeries, and its use is on the rise. If the bone fails to fuse properly, then a pseudarthrosis or “false joint” develops and results in pain, instability, and disability. Since1974, three types of electrical stimulation technologies have been approved for clinical use to enhance spinal fusions. One such technology is inductive coupling, which includes combined magnetic fields (CMFs). The purpose of this study was to evaluate the effects of a CMF device known as the Donjoy (SpinaLogic®) on MG-63 (ATCC® CRL1427TM) human osteosarcoma cells at treatment depths ranging from 0.5” to 6.0”. The cells were grown to confluence on 4-well chamber slides that were kept in a nickel-alloy chamber within an incubator to shield the cells from unwanted environmental electromagnetic fields. During treatment, a specially designed apparatus held both the treatment device and the chamber slide. Briefly, the chamber slide was placed inside an acrylic tube at a specific distance from the transducer housing, and the device was turned on for 30 minutes. The chamber slides were then returned to the incubator to be evaluated at 7, 14, and 21 days post treatment for cell viability and bone nodule formation. Our results showed that compared to control cells, the cells located at 3” from the source had the greatest increase in bone nodule formation 7 days post treatment which is the depth at this consistent with manufacturer recommendations.

Growth and Osteogenic Differentiation of CD117+ Dental Pulp and Periodontal Ligament Cells

BACKGROUND: Dental pulp stem cell (DPSC) and periodontal ligament stem cell (PDLSC) have been suggested as valuable seed cells for bone engineering, suggesting that both stem cells are potential osteogenic sources. Since DPSC and PDLSC seem like to have similar potential in bone formation, we conducted a study to compare morphology, immunophenotype and cell growth of DPSC and PDLSC isolated from the same teeth.METHODS: Human dental pulps and periodontal ligaments were obtained from freshly extracted partial impacted third molar teeth. Collected samples were digested with type I collagenase. Resulted cell suspension was washed and cultured. For biomarker identification, the cells were fixed and bound with anti-fluorescein isothiocyanate (FITC)-cluster of differentiation (CD)117 antibody. For cell growth quantification, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used. Meanwhile for osteogenic differentiation, the cells were cultured in osteogenic medium for 1-3 weeks, fixed and stained with alizarin red.RESULTS: Morphology of dental pulps cell (DPC) and periodontal ligament cell (PDLC) in passage 5 was similar. Clear CD117 green fluorescence of DPC and PDLC in passage 5 was observed. Cell growth rate of PDLC was higher than the one of DPC, 0.3858 and 0.3848 respectively. DPC formed bone nodule on the third week culture in osteogenic medium, while PDLC showed bone nodule formation on the second week culture.CONCLUSION: We suggest that DPC and PDLC are potential seed cells for osteogenic regeneration, since they had cell growth capacity and osteogenic differentiation, particularly PDLC that had faster osteogenic differentiation.KEYWORDS: dental pulp, periodontal ligament, cell, growth, osteogenic differentiation

Cytokines TNF-α, IL-6, IL-17F, and IL-4 Differentially Affect Osteogenic Differentiation of Human Adipose Stem Cells

During the initial stages of bone repair, proinflammatory cytokines are released within the injury site, quickly followed by a shift to anti-inflammatory cytokines. The effect of pro- and anti-inflammatory cytokines on osteogenic differentiation of mesenchymal stem cells is controversial. Here, we investigated the effect of the proinflammatory cytokines TNF-α, IL-6, IL-8, and IL-17F and the anti-inflammatory cytokine IL-4 on proliferation and osteogenic differentiation of human adipose stem cells (hASCs). hASCs were treated with TNF-α, IL-6, IL-8, IL-17F, or IL-4 (10 ng/mL) for 72 h mimicking bone repair. TNF-αreduced collagen type I gene expression but increased hASC proliferation and ALP activity. IL-6 also strongly enhanced ALP activity (18-fold), as well as bone nodule formation by hASCs. IL-8 did not affect proliferation or osteogenic gene expression but reduced bone nodule formation. IL-17F decreased hASC proliferation but enhanced ALP activity. IL-4 enhanced osteocalcin gene expression and ALP activity but reduced RUNX2 gene expression and bone nodule formation. In conclusion, all cytokines studied have both enhancing and reducing effects on osteogenic differentiation of hASCs, even when applied for 72 h only. Some cytokines, specifically IL-6, may be suitable to induce osteogenic differentiation of mesenchymal stem cells as a strategy for enhancing bone repair.

Continuous and intermittent exposure of neonatal rat calvarial cells to PTHrP (1-36) inhibits bone nodule mineralization in vitro by downregulating bone sialoprotein expression via the cAMP signaling pathway

The development and growth of the skeleton in the absence of parathyroid-hormone-related protein (PTHrP) is abnormal. The shortening of appendicular bones in PTHrP gene null mice is explained by an effect of PTHrP on endochondral bone growth. Whether or not PTHrP influences intramembranous ossification is less clear. The purpose of this study was to determine the effect of exogenous PTHrP on intramembranous ossificationin vitro. Neonatal rat calvarial cells maintained in primary cell culture conditions that permit spontaneous formation of woven bone nodules by intramembranous ossification were studied. The expression of PTHrP, parathyroid hormone 1 receptor (PTH1R), and alkaline phosphatase (AP) by osteogenic cells in developing nodules and the effects of PTHrP (1-36) on nodule development was determined over 3-18 days. PTHrP and PTH1R were detected colonies of osteogenic cells on culture day three, and AP was detected on day six. PTHrP and its receptor were localized in pre-osteoblasts, osteoblasts, and osteocytes, and AP activity was detected in pre-osteoblasts and osteoblasts but not osteocytes. Continuous and intermittent exposure to PTHrP (1-36) decreased the number of mineralized bone nodules and bone sialoprotein (BSP) mRNA and protein, but had no effect on the number of AP-positive osteogenic cell colonies, cell proliferation, apoptosis, or osteopontin (OPN) mRNA. These results demonstrate that osteogenic cells that participate in the formation of woven bone nodulesin vitroexhibit PTHrP and PTH1R before they demonstrate AP activity. Exogenous PTHrP (1-36) inhibits the mineralization of woven bone deposited during bone nodule formationin vitro,possibly by reducing the expression of BSP.

Antiosteoporotic Activity ofDioscorea alataL. cv. Phyto through Driving Mesenchymal Stem Cells Differentiation for Bone Formation

The aim of this study was to evaluate the effect of an ethanol extract of the rhizomes ofDioscorea alataL. cv. Phyto, Dispo85E, on bone formation and to investigate the mechanisms involved. Our results showed that Dispo85E increased the activity of alkaline phosphatase (ALP) and bone nodule formation in primary bone marrow cultures. In addition, Dispo85E stimulated pluripotent C3H10T1/2 stem cells to differentiate into osteoblasts rather than adipocytes. Ourin vivodata indicated that Dispo85E promotes osteoblastogenesis by increasing ALP activity and bone nodule formation in both intact and ovariectomized (OVX) mice. Microcomputed tomography (μCT) analysis also showed that Dispo85E ameliorates the deterioration of trabecular bone mineral density (tBMD), trabecular bone volume/total volume (BV/TV), and trabecular bone number (Tb.N) in OVX mice. Our results suggested that Dispo85E is a botanical drug with a novel mechanism that drives the lineage-specific differentiation of bone marrow stromal cells and is a candidate drug for osteoporosis therapy.