Stimulation of Odontogenesis and Angiogenesis via Bioactive Nanocomposite Calcium Phosphate Cements Through Integrin and VEGF Signaling Pathways.

Formulating self-setting calcium phosphate cements (CPCs) with secondary phases particularly in the nanoscale order holds great promise to improve biological properties. Here, we focus on the effect that bioactive glass nanoparticles (BGN) incorporated in CPC compositions can have on the proliferation, odontogenic differentiation, and angiogenic stimulation of stem cells derived from human dental pulp (HDPSCs). These odontogenic and angiogenic events are of special importance in the dentin-pulp regeneration processes. In comparison to pure CPCs, nanocomposite cements exhibit a significantly improved proliferation of HDPSCs, and the improvement is more significant as the BGN content increases. The nanocomposite cements substantially enhance the adhesion of cells, and significantly up-regulate odontogenic differentiation, including alkaline phosphatase (ALP) activity and the expressions of odontogenic genes (sialophosphoprotein, dentin matrix protein I, ALP, osteopontin and osteocalcin). Furthermore, the use of nanocomposite cements result in stimulation of angiogenic gene expression (VEGF, FGF-2, VEGFRs, PECAM-1, and VE-cadherin) and protein production (VEGF, VEGFR-1). The angiogenic stimulation by the HDPSCs significantly affects the endothelial cell behaviors, that is, the endothelial cell migration and the tubular network formation are substantially improved when treated with HDPSC-conditioned medium, particularly with the help of nanocomposite cements. The integrin and VEGF signaling pathways are reasoned for the stimulation of the odontogenesis and angiogenesis of cells, where the nanocomposite cements up-regulate the integrin subsets α1, α2, α3, and ß1, and activate the integrin downstream signal pathways, such as p-FAK, p-Akt, p-paxillin, JNK, EK, and NF-κB, as well as other nuclear transcriptional factors, including CREB, STAT-3, and ELK-1. The current results indicate that the new formulation of the nanocomposite self-setting cements might provide some beneficial microenvironments for the regenerative processes of dentin-pulp complex tissues.

Magnetic Nanocomposite Scaffold-Induced Stimulation of Migration and Odontogenesis of Human Dental Pulp Cells through Integrin Signaling Pathways.

Magnetism is an intriguing physical cue that can alter the behaviors of a broad range of cells. Nanocomposite scaffolds that exhibit magnetic properties are thus considered useful 3D matrix for culture of cells and their fate control in repair and regeneration processes. Here we produced magnetic nanocomposite scaffolds made of magnetite nanoparticles (MNPs) and polycaprolactone (PCL), and the effects of the scaffolds on the adhesion, growth, migration and odontogenic differentiation of human dental pulp cells (HDPCs) were investigated. Furthermore, the associated signaling pathways were examined in order to elucidate the molecular mechanisms in the cellular events. The magnetic scaffolds incorporated with MNPs at varying concentrations (up to 10%wt) supported cellular adhesion and multiplication over 2 weeks, showing good viability. The cellular constructs in the nanocomposite scaffolds played significant roles in the stimulation of adhesion, migration and odontogenesis of HDPCs. Cells were shown to adhere to substantially higher number when affected by the magnetic scaffolds. Cell migration tested by in vitro wound closure model was significantly enhanced by the magnetic scaffolds. Furthermore, odontogenic differentiation of HDPCs, as assessed by the alkaline phosphatase activity, mRNA expressions of odontogenic markers (DMP-1, DSPP,osteocalcin, and ostepontin), and alizarin red staining, was significantly stimulated by the magnetic scaffolds. Signal transduction was analyzed by RT-PCR, Western blotting, and confocal microscopy. The magnetic scaffolds upregulated the integrin subunits (α1, α2, ß1 and ß3) and activated downstream pathways, such as FAK, paxillin, p38, ERK MAPK, and NF-κB. The current study reports for the first time the significant impact of magnetic scaffolds in stimulating HDPC behaviors, including cell migration and odontogenesis, implying the potential usefulness of the magnetic scaffolds for dentin-pulp tissue engineering.

Preparation of recombined milk using modified butterfats containing α-linolenic acid.

UNLABELLED: Modified butterfats (MBFs) were produced by lipase-catalyzed interesterification with 2 substrate blends (6:6:8 and 4:6:10, by weight) of anhydrous butterfat (ABF), palm stearin, and flaxseed oil in a stirred-batch type reactor after short path distillation. The 6:6:8 and 4:6:10 MBF contained 21.7% and 26.5%α-linolenic acid, respectively. Total saturated fatty acids of the MBFs ranged from 41.4% to 47.4%. The cholesterol contents of the 6:6:8 and 4:6:10 MBFs were 21.0 and 12.1 mg/100 g, respectively. In addition, the melting points of the 6:6:8 and 4:6:10 MBFs were 32 °C and 31 °C, respectively. After preparation of recombined milks (oil-in-water emulsions) with MBFs, the stability of emulsions prepared with the MBFs (6:6:8 and 4:6:10) was compared to those with ABF during 10-d storage at 30 °C. Skim milk powder (containing 1% protein) was added to prepare emulsions as an emulsifier. Microstructures of emulsions freshly prepared with the ABF and the MBFs consisted of uniform fat globules with no flocculation during 10-d storage. With respect to fat globule size distribution, the volume-surface mean droplet diameter (d(32)) of the 6:6:8 and 4:6:10 MBF emulsions ranged between 0.33 and 0.34 µm, which was similar to the distribution in ABF emulsion. PRACTICAL APPLICATION: Milk, an expensive dairy food, has been widely used in various milk-derived food products. Modified butterfats (MBFs) contain α-linolenic acid as an essential fatty acid. Emulsion stability of recombined milks (oil-in-water emulsions) with MBFs was similar to that in anhydrous butterfat emulsion during 10-d storage. They may be a promising alternative for reconstituted milks to use in processed milk-based products.

Role of SIRT1 in heat stress- and lipopolysaccharide-induced immune and defense gene expression in human dental pulp cells.

INTRODUCTION: Although bacterial infection and heat stress are common causes of injury in human dental pulp cells (HDPCs), little is known about the potential defense mechanisms mediating their effects. This study examined the role of SIRT1 in mediating heat stress and lipopolysaccharide (LPS)-induced immune and defense gene expression in HDPCs. METHODS: HDPCs were exposed to heat stress (42°C) for 30 minutes after stimulation with LPS (1 µg/mL) for 48 hours. The expression of defense genes was evaluated by reverse-transcriptase polymerase chain reaction, Western blotting, and enzyme-linked immunosorbent assay. RESULTS: LPS and heat stress synergistically increased the expression of SIRT1 and immune and defense genes such as interleukin (IL)-8, hemeoxygenase-1 (HO-1), and human ß-defensin 2 (hBD-2). Resveratrol enhanced LPS- and heat stress-induced expression of HO-1 and hBD-2 but reduced IL-8 messenger RNA levels. The stimulation of HO-1 and hBD-2 messenger RNA expression by LPS and heat stress was inhibited by sirtinol; SIRT1 small interfering RNA; and inhibitors of p38, ERK, JNK, and nuclear factor κB. CONCLUSIONS: These results show for the first time that SIRT1 mediates the induction of immune and defense gene expression in HDPCs by LPS and heat stress. SIRT1 may play a pivotal role in host immune defense system in HDPCS.