Chi-Chun Fong

Gold nanoparticles promote osteogenic differentiation of mesenchymal stem cells through p38 MAPK pathway.

Understanding the interaction mechanisms between nanomaterials and biological cells is important for the control and manipulation of these interactions for biomedical applications. In this study, we investigated the cellular effects of gold nanoparticles (AuNPs) on the differentiation of mesenchymal stem cells (MSCs) and the associated molecular mechanisms. The results showed that AuNPs promoted the differentiation of MSCs toward osteoblast cells over adipocyte cells by inducing an enhanced osteogenic transcriptional profile and an attenuated adipogenic transcriptional profile. AuNPs exerted the effects by interacting with the cell membrane and binding with proteins in the cytoplasm, causing mechanical stress on the MSCs to activate p38 mitogen-activated protein kinase pathway (MAPK) signaling pathway, which regulates the expression of relevant genes to induce osteogenic differentiation and inhibit adipogenic differentiation.

Cell Adhesion and Spreading Behavior on Vertically Aligned Silicon Nanowire Arrays

In this report, we studied the interactions between biological cells and vertically aligned silicon nanowire (SiNW) arrays and focused on how SiNW arrays affected cellular behaviors such as cell adhesion and spreading. We observed that SiNW arrays could support cell adhesion and growth and guide cell adhesion and spreading behaviors. The results also showed that SiNW arrays could not only enhance the cell-substrate adhesion force but also restrict cell spreading. Combining the results from scanning electron microscopy images of cell morphology and the expression analysis of genes and proteins related to cell adhesion and spreading process, we proposed a mechanism on how cell adhesion and spreading was controlled by arrayed SiNWs. The effects of SiNW arrays in guiding cell adhesion and spreading behavior might be useful in the development of cell microarrays, tissue engineering scaffolds, and molecule delivery vehicles in which strong cell-substrate adhesion and reduced cell-cell communication were beneficial.

BIOSENSOR MEASUREMENT OF THE INTERACTION KINETICS BETWEEN INSULIN-LIKE GROWTH FACTORS AND THEIR BINDING PROTEINS

The binding kinetics of human insulin-like growth factor binding protein (IGFBP) 1-6 for recombinant human insulin-like growth factor (IGF) I and II were measured and compared in the present study using surface plasmon resonance biosensor technique. Different concentrations of IGFBPs (5-100 nM) were allowed to interact with the immobilized IGF-I or IGF-II on sensor chip surface. Both des(1-3)IGF-I and insulin are known to bind weakly to the IGFBPs and therefore are used as negative controls for the binding experiments. The resultant sensorgrams were analyzed by using simple 1:1 binding model to derive both the association rate (k(a)) and dissociation rate (k(d)) constants for IGFBP-IGF interactions. The k(a) values of IGFBPs are in the range of 1x10(4) to 9x10(5) M(-1) s(-1) for IGF-I and 7x10(3) to 1.7x10(6) M(-1) s(-1) for IGF-II, respectively. The orders of k(a) for both IGF-I and IGF-II are IGFBP-3>IGFBP-5>IGFBP-6>IGFBP-4>IGFBP-2>++ +IGFBP-1. The k(d) values of IGFBPs are in the range of 1.5x10(-5) to 2x10(-4) s(-1) for IGF-I and 3.6x10(-5) to 3.7x10(-4) s(-1) for IGF-II, respectively. The order of k(d) for IGF-I is IGFBP-6>IGFBP-5>IGFBP-4>IGFBP-3>IGFBP-2>++ +IGFBP-1 and that for IGF-II is IGFBP-5>IGFBP-6>IGFBP-2>IGFBP-4>IGFBP-3>++ +IGFBP-1, respectively. The equilibrium affinity constants (K(A)) were calculated based on the ratio of k(a)/k(d) and were more precise than the published literature values based on competitive radioligand binding assays. The systematic study enables a direct comparison on the IGF-binding properties among the various IGFBPs, and the kinetic data provide additional information to delineate the physiological role of different IGFBPs in vivo.

Hypoxia induces the activation of human hepatic stellate cells LX‐2 through TGF‐β signaling pathway

Hypoxia is a common environmental stress factor and is also associated with various physiological and pathological conditions such as fibrogenesis. The activation of hepatic stellate cells (HSCs) is the key event in the liver fibrogenesis. In this study, the behavior of human HSCs LX‐2 in low oxygen tension (1% O2) was analyzed. Upon hypoxia, the expression of HIF‐1α and VEGF gene was induced. The result of Western blotting showed that the expression of α‐SMA was increased by hypoxic stimulation. Furthermore, the expression of MMP‐2 and TIMP‐1 genes was increased. Hypoxia also elevated the protein expression of the collagen type I in LX‐2 cells. The analysis of TGF‐β/Smad signaling pathway showed that hypoxia potentiated the expression of TGF‐β1 and the phosphorylation status of Smad2. Gene expression profiles of LX‐2 cells induced by hypoxia were obtained by using cDNA microarray technique.

Effect of hydrogel matrix on binding kinetics of protein–protein interactions on sensor surface

Abstract Surface plasmon resonance (SPR) biosensor has become a standard technology for measuring kinetics of bimolecular interactions without the need for labeling. Sensor chips coated with a carboxymethylated dextran (CMD) hydrogel matrix are commonly used for immobilizing a protein binding partner in kinetic studies. The sensor chip provides a biocompatible surface with low non-specific binding, but it also presents some problems, such as steric effect and re-binding which may bias the kinetic measurement. In the present study, the effect of hydrogel matrix on protein–protein interaction was investigated. The insulin-like growth factors (IGFs) and their binding proteins were used as the model system. Kinetic parameters obtained with either of the binding partners immobilized on the matrix were compared to evaluate the effects of the matrix on the binding kinetics. The surface capacity and sensitivity of the hydrogel-modified sensor chip (CM5) and those of a sensor chip modified with a self-assembled monolayer (SAM) were measured and the performance of both chips was discussed.

Effects of Silicon Nanowires on HepG2 Cell Adhesion and Spreading

The unique capabilities of nanomaterials make them good candidates for catalysts, biosensors, and even drug carriers. However, interactions of nanomaterials with biological systems and the environment could lead to toxicity. 2] While there have been reports on the cytotoxicity of carbon nanotubes, quantum dots, 9] gold nanoparticles, and the biocompatibilities of carbon nanotubes, 12] no study has yet appeared on the biological effects of silicon nanowires, which are becoming increasingly important as nanomaterial. 14] Silicon nanowires (SiNWs) are one-dimensional nanomaterials that are typically composed of a single crystalline silicon core and an amorphous SiOx sheath. Their outstanding properties, such as quantum size effects, diameter-dependent thermal conductivity, and large piezoresistance coefficient have attracted a lot of research interest, including for application in biological materials and devices. 14] However, before SiNWs can be incorporated into new and existing biomedical devices, their cytotoxicity and potential adverse effects on biological systems should be thoroughly investigated. Here, we report the first study on the cytotoxicity of SiNWs on the human hepatocellular carcinoma cell line, HepG2, and their effect on cell adhesion and spreading. When the concentration of SiNWs was more than 100 mg mL , the nanowires did not form suspensions and precipitated in the cell-culture plates. Therefore, 100 mg mL 1 was chosen as the highest concentration for the experiments. The incubation time of 48 h has been used in other cytotoxicity studies with nanomaterials, 16] and was also found to be optimal for our experiments. Cytotoxicity of SiNWs was evaluated by using the Alamar blue assay. Cell viability was determined by exposing HepG2 cells to SiNWs suspensions of various concentrations (0.1, 1, 25, 50, 100 mg mL ) for 48 h at 37 8C. Cell viability was normalized with respect to the untreated control sample. The results of the Alamar blue assay showed that ACHTUNGTRENNUNGviability of HepG2 cells relative to the untreated control was affected by the SiNWs suspensions in a dose-dependent manner: the higher the concentration of SiNWs suspension, the lower the viability. It seems that SiNWs exerted a degree of cytotoxicity towards HepG2 cells under these experimental concentrations (Figure 1).

Gold nanoparticles stimulate differentiation and mineralization of primary osteoblasts through the ERK/MAPK signaling pathway.

Gold nanoparticles (AuNPs) have shown great promise for a variety of applications, including chemistry, biology, and medicine. Recently, AuNPs have found promising applications in cartilage and bone repair. However, to realize the above promised applications, more work needs to be carried out to clarify the interactions between biological systems and AuNPs. In the present study, primary osteoblasts were used to evaluate the biocompatibility of 20-nm and 40-nm AuNPs, including morphology, proliferation, differentiation, gene and protein expression, and the underlying mechanisms. The results demonstrated that AuNPs were taken up by osteoblasts and aggregated in perinuclear compartment and vescular structures, but no morphological changes were observed. AuNPs could significantly promote the proliferation of osteoblasts, enhance the ALP activities, and increase the number of bone nodules and calcium content in vitro. In addition, the expression of BMP-2, Runx-2, OCN and Col-1 was remarkably up-regulated in the presence of AuNPs. It is noteworthy that 20-nm AuNPs are more potent than 40-nm AuNPs in regulating osteoblast activities. Besides, AuNPs increased the level of ERK phosphorylation/total ERK, suggesting the activation of ERK/MAPK pathway is involved in above activities. In conclusion, AuNPs exhibited great biocompatibility with osteoblasts, and have tremendous potential to be used as drug and/or gene delivery carrier for bone and tissue engineering in the future.

Beta-adrenergic signals regulate adipogenesis of mouse mesenchymal stem cells via cAMP/PKA pathway

The adipogenic capacity of mesenchymal stem cells (MSCs) and the involvement of beta-adrenergic signals in lipolysis and thermogenesis have been well established. However, little is known about the development of beta-adrenergic receptor (beta-AR) systems and the role of beta-adrenergic signals in adipogenic differentiation of MSCs. In this study, we demonstrated that both the mRNA and protein levels of beta2- and beta3-AR were up-regulated following adipogenesis of mouse bone marrow derived MSCs. We also established that beta-AR agonists negatively while antagonists positively affected MSC adipogenesis. Both the beta2- and beta3-AR were involved in MSC adipogenesis, with beta3-AR being the predominant subtype. The effect of beta-ARs on MSC adipogenesis was at least partly mediated via the cAMP/PKA signaling pathway. These findings suggested that MSC is also a target for beta-adrenergic regulation, and beta-adrenergic signaling (major beta3-signaling) plays a role in MSC adipogenesis.

AU nanoparticles decorated TiO2 nanotube arrays as a recyclable sensor for photoenhanced electrochemical detection of Bisphenol A

A photorefreshable and photoenhanced electrochemical sensing platform for bisphenol A (BPA) detection based on Au nanoparticles (NPs) decorated carbon doped TiO2 nanotube arrays (TiO2/Au NTAs) is described. The TiO2/Au NTAs were prepared by quick annealing of anodized nanotubes in argon, followed by controllable electrodeposition of Au NPs. The decoration of Au NPs not only improved photoelectrochemical behavior but also enhanced electrocatalytic activities of the resulted hybrid NTAs. Meanwhile, the high photocatalytic activity of the NTAs allowed the electrode to be readily renewed without damaging the microstructures and surface states after a short UV treatment. The electrochemical detection of BPA on TiO2/Au NTAs electrode was significantly improved under UV irradiation as the electrode could provide fresh reaction surface continuously and the further increased photocurrent resulting from the improved separation efficiency of the photogenerated electron-hole pairs derived from the consumption of holes by BPA. The results showed that the refreshable TiO2/Au NTAs electrode is a promising sensor for long-term BPA monitoring with the detection limit (S/N = 3) of 6.2 nM and the sensitivity of 2.8 μA·μM(-1)·cm(-2).

Herba epimedii flavonoids suppress osteoclastic differentiation and bone resorption by inducing G2/M arrest and apoptosis

Accumulating evidences suggest that Herba epimedii has the potential benefits against osteoporosis. However, previous studies were focused on the crude extract, total flavonoids (TF) and icariin (ICA), and the detailed molecular mechanisms of action and structure-activity relationship (SAR) remain unclear. Herein we aimed to systematically investigate the effects of Herba epimedii flavonoids (HEF) on the activity of osteoclasts, and explore the potential SAR. Both ICA and baohuoside-1 (BS) significantly inhibited the proliferation of RAW 264.7 cells (IC(50) 25 μM and 67 μM, respectively). Treatment of ICA resulted in G2/M arrest and apoptosis in RAW 264.7 cells as early as 12 h. Besides, HEF remarkably suppressed vitamin D-induced differentiation of osteoclasts in rabbit bone marrow cells and the bone resorption of rabbit mature osteoclasts in vitro. It is notable that the inhibitory effect of 100 μM ICA and BS on osteoclast formation is almost 90%; and the inhibition rate on bone resorption is 50% and 80%, respectively. Besides, RANKL-induced osteoclast formation from RAW 264.7 cells and the expression of TRAP, CA II, CTSK and MMP-9 was significantly reduced by the treatment of 25 μM HEF and 17β-estradiol (ES), and the inhibitory strength increases in the order TF < ES < ICA < BS, which was blocked by ICI182780 suggesting that the regulation of osteoclast activity might be ER dependent. Furthermore, the free hydroxyl group at C-7 of BS played an important role in the SAR for anti-osteoclast action. To conclude, HEF could regulate the formation and activity of osteoclasts by inhibiting the proliferation and differentiation, inducing apoptosis and cell cycle arrest and suppressing bone resorption of osteoclasts. Changes in osteoclast activity are probably mediated predominantly by interaction with nuclear estrogen receptors and via mitochondrial pathway. HEF, especially BS, has great potential for the prevention and treatment of osteoporosis.