Elastic Membrane That Undergoes Mechanical Deformation Enhances Osteoblast Cellular Attachment and Proliferation

The main objective of this paper was to investigate the effect of transmission of force on bone cells that were attached to a deformable membrane. We functionalized a silastic membrane that measured 0.005 inches thickness and coated it with an extra cellular matrix (ECM) protein, fibronectin (FN). MC3T3-E1 osteoblast-like cells were cultured on the functionalized FN-coated membrane after which cell attachment and proliferation were evaluated. We observed an immediate attachment and proliferation of the bone cells on the functionalized membrane coated with FN, after 24 hours. Upon application of a mechanical force to cells cultured on the functionalized silicone membrane in the form of a dynamic equibiaxial strain, 2% magnitude; at 1-Hz frequency for 2 h, the osteoblast cells elicited slightly elevated phalloidin fluorescence, suggesting that there was reorganization of the cytoskeleton. We concluded from this preliminary data obtained that the engineered surface transduced applied mechanical forces directly to the adherent osteoblast cells via integrin binding tripeptide receptors, present in the FN molecules, resulting in the enhanced cellular attachment and proliferation

Fibronectin adsorption on surface-activated poly(dimethylsiloxane) and its effect on cellular function

This article reports that surface modification of poly(dimethylsiloxane) (PDMS) influences fibronectin (Fn) adsorption and enhances cell attachment. Controlled adsorption of Fn on chemically activated polymer substrates is known to influence cellular function. Thin films of PDMS were spun cast on silicon wafers to obtain homogeneous and molecularly smooth surfaces. The films were made hydrophilic by exposure to ultraviolet ozone activation (PDMS*). The films then were characterized by contact angle goniometry, ellipsometry, atomic force microscopy (AFM). Rutherford backscattering spectrometry and X-ray photoelectron spectroscopy. Contact angle measurements indicated higher hydrophobicity of the nonactivated PDMS substrates than PDMS*. AFM scans of the substrates indicated higher surface roughness of PDMS* (Ra = 0.55 nm) than PDMS (Ra = 0.25 nm). Although Fn surface density (Γ) was slightly higher on PDMS than on PDMS*, due to hydrophobic interactions between substrate and Fn, cell function was greatly enhanced on the Fn-coated PDMS* (PDMS*-Fn) than on PDMS (PDMS-Fn). Higher attachment of MC3T3-E1 osteoblast-like cells was observed on PDMS*-Fn than on PDMS-Fn. Moreover, cell spreading and cytoskeleton organization after 72 h was clearly favored on the Fn-coated PDMS* surfaces. © 2004 Wiley Periodicals, Inc

Initial attachment of osteoblast-like cells on functionalized surfaces coated with calcium phosphate

Tissue engineering techniques which combine synthetic grafts with molecules and cells are considered as viable long term solutions for bone tissue repair and reconstruction procedures. The self SAM technology enables modeling of surface functional groups on biomaterials, while the concept of ion-selective precipitation reaction causes formation of Ca-P coating on these functionalized surfaces. Hydrophilic surfaces such as -COOH or -OH end groups have very powerful induction capability for the heterogeneous nucleation of hydroxyapatite-like layer, while nucleation could be prohibited on an - NH 2-terminated surface. Using the SAM technique we grafted three different organosilanes onto silicon wafers to yield -NH2, -COOH and -OH functionalized surfaces respectively. The surfaces were characterized by contact angle measurements, ellipsometry, FTIR, SEM/EDX and RBS. Ca-P coating was formed on the SAMs by immersion in a simulated physiological fluid (pH 7.4, at 37°C). FTIR showed dependence of the morphology of the Ca-P coating on both the type of surface functionality and on the duration of coating. The spectrum exhibited P-O and C-O absorption bands characteristic of a carbonated calcium hydroxyapatite with stronger P-O bands on -OH surface. MC3T3-E1 osteoblast-like cells were used in bioassays to study the effect of the end groups on the initial cellular attachment and alkaline phosphatase expression on Ca-P coated surfaces. Cell assays showed increased cellular attachment on the Ca-P coated-OH SAM, after cells were seeded for 1h, compared to the -NH 2 and -COOH-terminated surfaces, while up to 5x alkaline phosphatase activity was shown on Ca-P coated surfaces. At a low temperature therefore, the end groups of SAMs act as effective nucleation sites to induce formation of a biomimetic apatite coating, in a one-step biomineralization process. The most potent apatite forming surface, the -OH SAM surface, demonstrates that surface -OH groups are important to induce bone bioactive behavior at surfaces

Effect of functional end groups of silane self-assembled monolayer surfaces on apatite formation, fibronectin adsorption and osteoblast cell function

Bioactive glass (BG) can directly bond to living bone without fibrous tissue encapsulation. Key mechanistic steps of BG\u27s activity are attributed to calcium phosphate formation, surface hydroxylation and fibronectin (FN) adsorption. In the present study, self-assembled monolayers (SAMs) of alkanesilanes with different surface chemistry (OH, NH2 and COOH) were used as a model system to mimic BG\u27s surface activity. Calcium phosphate (Ca-P) was formed on SAMs by immersion in a solution that simulates the electrolyte content of physiological fluids. FN adsorption kinetics and monolayer coverage was determined on SAMs with or without Ca-P coating. The surface roughness was also examined on these substrates before and after FN adsorption. The effects of FN-adsorbed, Ca-P-coated SAMs on the function of MC3T3-E1 were evaluated by cell growth, expression of alkaline phosphatase activity and actin cytoskeleton formation. We demonstrate that, although the FN monolayer coverage and the root mean square (rms) roughness are similar on - OH and - COOH terminated SAMs with or without Ca-P coating, higher levels of ALP activity, more actin cytoskeleton formation and more cell growth are obtained on - OH- and - COOH-terminated SAMs with Ca-P coating. In addition, although the FN monolayer coverage is higher on Ca-P-coated - NH2-terminated SAMs and SiOx surfaces, higher levels of ALP activity and more cell growth are obtained on Ca-P-coated - OH- and - COOH-terminated SAMs. Thus, with the same Ca-P coatings, different surface functional groups have different effects on the function of osteoblastic cells. These findings represent new insights into the mechanism of bioactivity of BG and thereby may lead to designing superior constructs for bone grafting. Copyright © 2008 John Wiley & Sons, Ltd

Evaluation of cell behaviour on atmospheric plasma deposited siloxane and fluorosiloxane coatings

For developing functional biomaterials, an understanding of the biological response at material surfaces is of key importance. In particular, surface chemistry, roughness and cell type influence this response. Many previous reports in the literature have involved the study of single cell types and their adhesion to surfaces with a limited range of water contact angles. The objective of this study was to investigate the adhesion of five cell lines on surfaces with contact angles in the range of 20 to 115 . This range of water contact angles was obtained using siloxane and fluorosiloxane coatings deposited using atmospheric plasma deposition. These nm thick coatings were deposited by nebulizing liquid precursors consisting of poly(dimethylsiloxane) (PDMS) and a mixture of perfluorodecyl acrylate/ tetraethylorthosilicate (PPFDA/TEOS) into the atmospheric plasmas. Cell adhesion studies were carried out with the following cell types: Osteoblast, Human Embryonic Kidney (HEK), Chinese hamster ovary (CHO), Hepatocytes (HepZ) and THP1 leukemic cells. The study demonstrated that cell adhesion was significantly influenced by the type of cell line, water contact angle and coating chemistry. For example the sensitivity of cell lines to changes in contact angle was found to decrease in the following order: Osteoblasts >Hepatocytes> CHO. The HEK and THP-1 inflammatory cells in contrast were not found to be sensitive to changes in water contact angle.Science Foundation IrelandAuthor has checked copyrightAD 22/01/201

Biomimetic hydroxyapatite coating on glass coverslips for the assay of osteoclast activity in vitro

The osteoclast (OC) is the cell type responsible for the resorption of bone. The activity of this cell is important in the aetiology of a large number of skeletal pathologies, and also for the biocompatibility and osseointegration of orthopaedic implant materials. OC mediated acid hydrolysis of calcium phosphate from the bone matrix offers a prime means of studying the biology and activity of this cell type. We have developed a method of coating glass coverslips with a hydroxyapatite (HA)-like mineral, using a biomimetic approach. Hydroxylation followed by formation of a self assembled monolayer (SAM) using the surfactant triethoxysilylpropyl succinic anhydride (TESPSA), allowed biomimetic deposition of HA-like mineral from a simulated body fluid (SBF). The biocompatibility of the TESPSA SAM-HA coated glass coverslips was tested by culturing human mature OC present in samples of giant cell tumour of bone (GCT). Parameters of OC activity were assayed, including F-actin ring formation, release of calcium and formation of osteoclastic resorption pits, confirming that OC were able to attach to and resorb the coated surface. This approach for the preparation of HA coatings on glass coverslips could have wide applicability for the study of osteoclast behaviour in vitro.Asiri K. A. R. Wijenayak, Christopher B. Colby, Gerald J. Atkins and Peter Majewsk