Xiaoying Lü

The molecular mechanism of action of bactericidal gold nanoparticles on Escherichia coli.

This work examines the molecular mechanism of action of a class of bactericidal gold nanoparticles (NPs) which show potent antibacterial activities against multidrug-resistant Gram-negative bacteria by transcriptomic and proteomic approaches. Gold NPs exert their antibacterial activities mainly by two ways: one is to collapse membrane potential, inhibiting ATPase activities to decrease the ATP level; the other is to inhibit the subunit of ribosome from binding tRNA. Gold NPs enhance chemotaxis in the early-phase reaction. The action of gold NPs did not include reactive oxygen species (ROS)-related mechanism, the cause for cellular death induced by most bactericidal antibiotics and nanomaterials. Our investigation would allow the development of antibacterial agents that target the energy-metabolism and transcription of bacteria without triggering the ROS reaction, which may be at the same time harmful for the host when killing bacteria.

Mechanisms of cytotoxicity of nickel ions based on gene expression profiles

This study investigated cytotoxic effects of Ni(II) to mouse fibroblast cells (L-929) on the level of gene expression profiles with cDNA microarray. The gene expression profiles of L-929 were detected after the cells were cultured in the medium with 200 microm Ni(II) for 24, 48 and 72 h, respectively, and the cytotoxicity of Ni(II) was evaluated with methylthiazoltetrazolium (MTT) assay. 20 up-regulated genes and 19 down-regulated genes were differentially expressed in all three-culture periods. Gene ontology analysis showed that the L-929 cells which responded to Ni(II) covered a broad range of functional gene groups including cellular biological process, molecular function, and cellular component. Ni(II) has extensive effects on cells by inhibiting cell proliferation and differentiation through inducing cell apoptosis, affecting cell development and influencing cholesterol metabolism.

The molecular mechanism of mediation of adsorbed serum proteins to endothelial cells adhesion and growth on biomaterials

To explore molecular mechanism of mediation of adsorbed proteins to cell adhesion and growth on biomaterials, this study examined endothelial cell adhesion, morphology and viability on bare and titanium nitride (TiN) coated nickel titanium (NiTi) alloys and chitosan film firstly, and then identified the type and amount of serum proteins adsorbed on the three surfaces by proteomic technology. Subsequently, the mediation role of the identified proteins to cell adhesion and growth was investigated with bioinformatics analyses, and further confirmed by a series of cellular and molecular biological experiments. Results showed that the type and amount of adsorbed serum proteins associated with cell adhesion and growth was obviously higher on the alloys than on the chitosan film, and these proteins mediated endothelial cell adhesion and growth on the alloys via four ways. First, proteins such as adiponectin in the adsorbed protein layer bound with cell surface receptors to generate signal transduction, which activated cell surface integrins through increasing intracellular calcium level. Another way, thrombospondin 1 in the adsorbed protein layer promoted TGF-β signaling pathway activation and enhanced integrins expression. The third, RGD sequence containing proteins such as fibronectin 1, vitronectin and thrombospondin 1 in the adsorbed protein layer bound with activated integrins to activate focal adhesion pathway, increased focal adhesion formation and actin cytoskeleton organization and mediated cell adhesion and spreading. In addition, the activated focal adhesion pathway promoted the expression of cell growth related genes and resulted in cell proliferation. The fourth route, coagulation factor II (F2) and fibronectin 1 in the adsorbed protein layer bound with cell surface F2 receptor and integrin, activated regulation of actin cytoskeleton pathway and regulated actin cytoskeleton organization.

Genome-wide pathways analysis of nickel ion-induced differential genes expression in fibroblasts.

To reveal molecular mechanisms of the interaction between Ni2+ and cells, cDNA microarray technology and GenMAPP analysis were utilized to investigate changes of gene expression profile and identify significant biological pathways in mouse fibroblast cells (L-929) treated by 100 microm Ni2+ for 12, 24, 48 and 72 h, respectively. The microarray data was validated by real-time PCR. Methylthiazoltetrazolium (MTT) analysis and flow cytometry experiment were used to assess the cellular response of L-929 cells to Ni2+. It was found that six main biological pathways were affected by Ni2+ with 118 differentially expressed genes involved. Further analysis illuminated that the exposure of cells to Ni2+ may evoke series of cellular responses to hypoxia by regulating hypoxia-inducible gene expression and cause irreversible DNA damage. Cell cycle pathway analysis results showed DNA replication in S phase could be inhibited by Ni2+ which was consistent with the data gained from flow cytometry experiment. Compared to previous researches based on conventional molecular biology experiments, the present work has not only indirectly validated the findings of other groups but also obtained several discoveries related to cell-Ni2+ interaction, such as inhibition of electron transport chain and accumulation of extracellular matrix (ECM) collagens. The routine of the present study not only can analyze gene expression profile but also may provide a more convenient and efficient approach to explain molecular mechanisms of cell-biomaterial interaction.

Preparation and Characterization of Natural Hydroxyapatite from Animal Hard Tissues

The purpose of this study is to find out an effective method to prepare natural hydroxyapatite (HA) from biological source, i.e., pig bones, pig teeth, and extracted human teeth. For the preparation of natural HA a calcining method with different temperatures was used, in combination with the thermal gravimetric analysis (TGA). Three analysis methods, i.e., Fourier transform infrared spectroscope (FTIR), X-ray diffraction (XRD), and inductively coupled plasma (ICP) have been used to investigate the characteristics of the prepared materials. The spectrum of the prepared material, obtained by means of FTIR and XRD, are consistent with the standard FTIR spectrum and JCPDS index of XRD of hydroxyapitite. It confirms that the material prepared is hydroxyapitite indeed. The natural HA obtained by calcining at 850°C shows a desired quality.

Competitive protein adsorption on biomaterial surface studied with reflectometric interference spectroscopy

Reflectometry interference spectroscopy (RIfS) is known as a highly sensitive and robust technique for direct, label-free detection of the interaction of biomacromolecules in real time and in situ. The aim of the present study was to investigate the competitive protein adsorption on the surface of fluorocarbon end-capped poly(carbonate) urethane (PCUF) and polystyrene (PS) based on the RIfS method. The surface energy and microstructures of PCUF and PS were characterized by contact angle measurement and atomic force microscopy. Interfacial energies between these surfaces and the proteins were then calculated. The protein adsorption experiments were carried out with both single solution and ternary solutions composed of albumin, fibrinogen and immunoglobulin-G (IgG). The results of surface characterization showed that PCUF was more hydrophilic than PS with a smaller surface energy, and micro-phases separation of PCUF was observed. RIfS analysis results revealed that more albumins, less fibrinogen and IgG were detected on the PCUF surface compared with PS after simplex and competitive protein adsorption, which indicated that PCUF had a preferential adsorption for albumin. The special morphology, smaller surface energy and calculated interfacial energies between PCUF and proteins may be responsible for the better blood compatibility of PCUF compared to PS. The results suggest that RIfS could serve as a novel, effective method for studying the competitive protein adsorption on biomaterial surfaces.

Aqueous synthesis of gold nanoparticles and their cytotoxicity in human dermal fibroblasts–fetal

The unique physicochemical properties of nanoparticles make them promising substrates for application in the medical area. As there are no safety regulations yet, concerns about future health problems are rising. This study was conducted to prepare approximately 20 nm gold nanoparticles (GNPs) by a chemical reduction method and evaluate their cytotoxicity by MTT assay using human dermal fibroblasts-fetal (HDF-f). 10-50 nm GNPs could be obtained in redistilled water by varying the amount of sodium citrate. MTT results showed that approximately 20 nm GNPs did not cause cell death at a maximum concentration of 300 microM but affected the morphology of HDF-f when their concentration increased.

Hard tissue compatibility of natural hydroxyapatite/chitosan composite

The natural hydroxyapatite/chitosan (NHC) composite is a new synthesized material. The aim of this experiment was to assess the bone tissue compatibility of this NHC composite in vivo. Twenty-four healthy New Zealand rabbits were included in this study. Of those, 20 were used as the experimental group and four as the control group. In the experimental group, animals receive a cranium defect procedure and NHC composite repair. In the control group, animals underwent the cranium defect procedure without NHC composite repair. At 1, 4, 12, 24, and 40 weeks after surgery, the animals were sacrificed and samples were taken and assessed by gross observation, three-dimensional (3D) computerized tomographic (CT) reconstruction, histology and scanning electron microscope. Our results showed that at 1 week after repairing the bone defect with the NHC composite in the experimental group, new bone appeared around the composite and matured gradually. At 24 weeks after surgery, there were little collagenous tissues present between the material and surrounding bones. At 40 weeks after surgery, new bone had grown into the mature bone and total osseointegration had occurred. In the control group, however, no bone defect healing was observed at 40 weeks after surgery. All these results of the present in vivo work suggest that the NHC composite has a good hard tissue biocompatibility and an excellent osteoconductivity. It is suitable for artificial bone implants and frame materials of tissue engineering.

Gene expression profile study on osteoinductive effect of natural hydroxyapatite

The aim of this study was to investigate the osteoinductive effect of natural hydroxyapatite (NHA). NHA was extracted from pig bones and prepared into disk-like samples. Then, proliferation of mouse bone mesenchymal stem cells (MSCs) cultured on NHA was assessed by the methylthiazoltetrazolium (MTT) assay. Furthermore, microarray technology was applied to obtain the gene expression profiles of MSCs cultured on NHA at 24, 48, and 72 h. The gene expression profile was then comprehensively analyzed by clustering, Gene Ontology (GO), Gene Microarray Pathway Profiler (GenMAPP) and Ingenuity Pathway Analysis (IPA). According to the results of microarray experiment, 8992 differentially expressed genes were obtained. 90 differential expressed genes related to HA osteogenic differentiation were determined by GO analysis. These genes included not only 6 genes related to HA osteogenic differentiation as mentioned in the literatures but also newly discovered 84 genes. Some important signaling pathways (TGF-β, MAPK, Wnt, etc.) were influenced by these genes. Gene interaction networks were obtained by IPA software, in which the scoring values of two networks were highest, and their main functions were related to cell development. The comprehensive analysis of these results indicate that NHA regulate some crucial genes (e.g., Bmp2, Spp1) and then activate some pathways such as TGF-β signaling pathway, and ultimately osteogenic differentiation was induced.

MicroRNA sequencing and molecular mechanisms analysis of the effects of gold nanoparticles on human dermal fibroblasts.

The aim of this study is to investigate the mechanism of the effects of gold nanoparticles (GNPs) on human dermal fibroblasts (HDFs) at the microRNA level. First, 20-nm GNPs were synthesized and their effect on HDF proliferation was assayed. SOLiD sequencing technology was then utilized to obtain the microRNA expression profile after GNP treatment. The microRNA expression data were compared with previously obtained mRNA and protein expression data to identify the microRNA target mRNAs/proteins. Moreover, bioinformatics analyses and validation experiments were conducted. Lastly, the roles of GNPs and silver nanoparticle (SNPs) on HDFs were compared at the microRNA level. The results showed that GNPs were not cytotoxic as 202 microRNAs were differentially expressed after treatment with 200 μm GNPs for 1, 4 and 8 h. Bioinformatics analyses revealed that these dysregulated miRNAs mainly functioned in metabolic processes and participated in 71 biological pathways, including two key pathways in which the differentially expressed miRNA, target mRNAs and proteins were simultaneously joined, the mRNA processing pathway and MAPK signaling pathway. Biological experiments in cells confirmed that GNPs affected energy metabolism but did not induce apoptosis, destroy the cytoskeleton or induce reactive oxygen species (ROS) production. Comparing the mechanism of the effects of GNPs and SNPs on HDFs at the microRNA level, it was found that, unlike SNPs, GNPs impacted the cell cycle, weakened the ATP synthesis inhibition and cytoskeleton damage, suppressed apoptosis, and did not lead to cytotoxicity. The difference in ROS production by these two nanoparticles might partially explain the fact that GNPs showed no cytotoxic effects on HDFs, unlike SNPs.