What did we learn from Riata™?

The Riata™ 8 French (Fr) and Riata ST™ 7 Fr families of silicone leads have experienced a specific form of insulation abrasion characterized by externalization of conductor cables outside the lead body. Design differences between the 8 Fr and 7 Fr leads make conductor externalization less likely in the smaller diameter lead. The Riata Lead Evaluation Study (RLES) reported on the prevalence of externalized conductors (EC) in patients implanted with Riata™ and Riata ST™ silicone leads and provided details on the incidence of electrical abnormalities in Riata™ and Riata ST™ leads with and without EC. The prevalence of EC was significantly lower for the Riata ST™ leads (7 Fr) as compared with Riata™ leads (8 Fr) (24/259 (9.3 %) vs. 125/517 (24.2 %), P < 0.001), and the majority of leads with EC were not associated with electrical malfunction. The aim of this manuscript is to describe the design of the Riata™ and Riata ST™ families of leads, provide details regarding the mechanisms and rates of lead failure, and discuss recommendations for patient management.

Adsorbed IgG: a potent adhesive substrate for human macrophages.

Previous reports from our laboratory have demonstrated qualitatively that preabsorbed IgG can enhance long-term macrophage adhesion in vitro. This investigation further characterizes and quantifies the biological effect of adsorbed human IgG on human macrophages and probes the potential mechanisms. Ten-day human monocyte/macrophage cultures on Plastek M (PM), a normally poor cellular substrate for macrophages, confirmed the ability of preabsorbed IgG to dramatically enhance long-term macrophage adhesion. An adsorption solution concentration of 200 microg/mL of IgG was necessary to provide a consistent, optimal cellular response. (125)I adsorption studies indicated Langmuir-style IgG adsorption at low concentrations; however, no adsorption maximum was observed. Additional adsorption analysis revealed that the IgG fragments Fab, F(ab')(2), and Fc adsorb at levels only 20-40% that of the whole molecule. Despite the lower adsorption levels, both preabsorbed Fab and F(ab')(2) were shown to be as effective as whole molecule IgG at enhancing long-term macrophage adhesion. Surprisingly, the preabsorbed Fc fragment demonstrated no IgG-like activity, thereby eliminating the possibility of an Fc receptor-based mechanism. Other possible mechanisms, such as macrophage lectins, novel macrophage Fab receptors, and complement activation by adsorbed IgG and IgG fragments, are discussed.

Adsorbed serum proteins responsible for surface dependent human macrophage behavior.

Substrate specific cellular responses are the result of a complex biological system that includes protein adsorption, receptor-ligand binding, and signal transduction. This investigation attempted to identify specific proteins adsorbed from human serum that may be responsible for the previously reported in vitro surface dependent behavior of human macrophages and foreign body giant cells (FBGCs). The adsorption of human albumin, alpha(2)-macroglobulin, complement factor 3b, fibronectin, IgG, thrombospondin, vitronectin (VN), and von Willebrand factor (vWF) from a 25% serum solution was quantified with (125)I-labeled protein. Adsorption substrates included clean glass, alkyl-silane modified glass, amino-silane modified glass, poly(ethylene oxide) (PEO)-coupled glass, and the reference biomaterials poly(etherurethane urea), Silastic(R), and poly(tetrafluoroethylene) (PTFE). Following quantification of 2-h adsorption, surfaces were treated with sodium dodecyl sulfate (SDS) and the level of adsorbed proteins remaining was quantified. The pre- and post-SDS adsorption were both compared to previously reported surface dependent in vitro macrophage and FBGC behavior on the same surfaces; however, no correlations could be made. Adsorption strength, defined as the percentage of initially adsorbed protein that remained adsorbed after SDS treatment, correlated well with previously reported in vitro cellular behavior indicating that adsorbed vWF, IgG, and VN may be involved in the modulation of adherent macrophage and FBGC behavior. Those surfaces that strongly adsorbed vWF also inhibited long-term macrophage adhesion, while those surfaces that strongly adsorbed IgG promoted long-term macrophage adhesion. In addition, the highest levels of FBGC formation had been observed only on those surfaces that strongly adsorbed VN. Subsequent human monocyte cultures on protein preadsorbed substrates confirmed the inhibitory effect of adsorbed vWF and the promoting effect of IgG on longterm macrophage adhesion as predicted by adsorption strength correlations. However, preadsorbed VN was not observed to modulate FBGC formation, which is in contrast to the conclusions of the adsorption correlations.

Effects of surface-coupled polyethylene oxide on human macrophage adhesion and foreign body giant cell formation in vitro.

Surface immobilized polyethylene oxide (PEO) has been shown to efficiently reduce protein adsorption and cellular adhesion, resulting in a biologically passive surface. To explore the in vitro effects of surface immobilized PEO on the human inflammatory cells, macrophages, and foreign body giant cells (FBGCs), we developed a diisocyanate-based method for coupling PEO to amine-modified glass, a surface previously shown to enhance macrophage adhesion and FBGC formation. Contact angle analysis and X-ray photoelectron spectroscopy confirmed the presence of PEO molecules bound to the surface and revealed that PEO molecular weight significantly influenced the efficiency of PEO coupling. We used a 10-day human monocyte culture protocol to demonstrate that the presence of surface coupled PEO molecules does not significantly decrease initial monocyte density or monocyte-derived macrophage density after 3 days. However, PEO-coupled surfaces significantly reduced long-term monocyte-derived macrophage density and virtually eliminated interleukin-4-induced FBGC formation observed at day 10. The cellular response to these PEO-coupled surfaces was related to the molecular weight of the PEO chains, which was varied between 200 Da and 18.5 kDa. These results suggest that an optimized PEO surface treatment may be effective in reducing inflammatory cell adhesion and possible degradation during the inflammatory response to an implanted biomedical device.

Alkylsilane-modified surfaces: inhibition of human macrophage adhesion and foreign body giant cell formation.

A homologous set of alkylsilane-modified glass surfaces with chain lengths ranging from methyl to octadecyl was prepared in order to examine the influence of alkyl surface chemistry on macrophage adhesion and foreign body giant cell (FBGC) formation. Contact angle and X-ray photoelectron spectroscopy analysis confirmed our silanation technique and indicated a consistent alkyl chain density independent of chain length. Human peripheral blood monocytes were isolated and cultured on these alkylsilane surfaces for a period of 10 days. The initial density of human monocytes was similar on all surfaces. Beyond day 0 the clean glass, methyl (DM and C1), propyl (C3), and hexyl (C6) surfaces maintained a high cell density and supported macrophage development. In contrast, long-term macrophage density was extremely low on the tetradecyl (C14) and octadecyl (C18) surfaces. When interleukin-4 was added to induce FBGC formation in vitro, the DM, C1, C3, and C6 surfaces supported high levels of macrophage fusion while clean glass strongly inhibited fusion. The C14 and C18 surfaces did not contain sufficient macrophages to support FBGC formation. Cage implant studies revealed that in vivo macrophage density and FBGC formation on clean glass and C6 surfaces was similar to in vitro data. In contrast to the monocyte culture results, the C18 cage implant samples supported significant FBGC formation, possibly as a result of different conditions within each experimental system. Radiotracer adsorption studies of eight human serum proteins identified the high concentration and tenacious hold of adsorbed von Willebrand factor as being possibly involved in the poor long-term macrophage density observed on C14 and C18.

Disruption of filamentous actin inhibits human macrophage fusion.

The foreign body reaction to implanted biomaterials, characterized by the presence of macrophages and foreign body giant cells (FBGC), can result in structural and functional failure of the implant. Recently, we have shown that interleukin-4 and interleukin-13 can independently induce human macrophage fusion to form FBGC via a macrophage mannose receptor (MR) -mediated pathway. The MR is believed to mediate both endocytosis of glycoproteins and phagocytosis of microorganisms, which bear terminal mannose, fucose, N-acetylglucosamine, or glucose residues. Polarization of microfilaments to closely apposed macrophage membranes as observed with fluorescence confocal microscopy led us to ask whether MR-mediated fusion occurred via a filamentous actin-dependent pathway. Cytochalasins B and D and latrunculin-A, agents that disrupt microfilaments, inhibited macrophage fusion in a concentration-dependent manner. The concentrations of cytochalasins D and B that inhibited fusion did not significantly decrease macrophage adhesion, spreading, or motility but did inhibit internalization of Candida albicans during interleukin-13-enhanced, MR-mediated phagocytosis. Very low concentrations of cytochalasin B (< 2 microM) induced a slight enhancement of macrophage fusion. Taken together, the results of this study suggest that cytokine-induced, MR-mediated macrophage fusion requires an intact F-actin cytoskeleton and that the mechanism of fusion is similar to phagocytosis.--DeFife, K. M., Jenney, C. R., Colton, E., Anderson, J. M. Disruption of filamentous actin inhibits human macrophage fusion.

Cytoskeletal and adhesive structural polarizations accompany IL-13-induced human macrophage fusion.

During the inflammatory response to an implanted biomaterial, monocytes undergo a striking phenotypic progression of differentiation into macrophages, which may subsequently fuse to form foreign body giant cells (FBGCs). Taking advantage of an in vitro system of cytokine-induced FBGC formation together with the optical slicing capabilities of a confocal microscope, we investigated the cytoskeletal reorganization and adhesive structure development during this dramatic morphological progression. Human monocytes demonstrated diffuse cytoplasmic staining of adhesive structural proteins. Punctate filamentous (F)-actin structures appeared along the ventral cell membrane of macrophages and were identified as the core of podosome adhesive structures by the distinctive ring staining of vinculin, talin, and paxillin around the F-actin. Cytokine-induced FBGCs were characterized by a restriction of podosomes to the extreme periphery of the ventral cell surface. Although macrophages and FBGC contained equivalent amounts of F-actin, significantly more F-actin was located within 1 micron of the ventral plasma membrane in FBGCs compared to macrophages. Taken together, these results provide new information on the dynamic cytoskeletal reorganization and adhesive structure development that occur during phenotypic progression from human monocytes to macrophages to FBGC. Furthermore, they suggest the acquisition of functional specializations on FBGC formation, which may enhance our understanding of chronic inflammatory processes.

Monocyte, macrophage and foreign body giant cell interactions with molecularly engineered surfaces.

To elucidate the mechanisms involved in monocyte/macrophage adhesion and fusion to form foreign body giant cells on molecularly engineered surfaces, we have utilized our in vitro culture system to examine surface chemistry effects, cytoskeletal reorganization and adhesive structure development, and cell receptor-ligand interactions in in vitro foreign body giant cell formation. Utilizing silane-modified surfaces, monocyte/macrophage adhesion was essentially unaffected by surface chemistry, however the density of foreign body giant cells (FBGCs) was correlated with surface carbon content. An exception to the surface-independent macrophage adhesion were the alkyl-silane modified surfaces which exhibited reduced adhesion and FBGC formation. Utilizing confocal immunofluorescent techniques, cytoskeletal reorganization and adhesive structure development in in vitro FBGC formation was studied. Podosomes were identified as the adhesive structures in macrophages and FBGCs based on the presence of characteristic cytoplasmic proteins and F-actin at the ventral cell surface. Focal adhesion kinase (FAK) and focal adhesions were not identified as the adhesive structures in macrophages and FBGCs. In studying the effect of preadsorbed proteins on FBGC formation, fibronectin or vitronectin do not play major roles in initial monocyte/macrophage adhesion, whereas polystyrene surfaces modified with RGD exhibited significant FBGC formation. These studies identify the potential importance of surface chemistry-dependent conformational alterations which may occur in proteins adsorbed to surfaces and their potential involvement in receptor-ligand interactions. Significantly, preadsorption of alpha2-macroglobulin facilitated macrophage fusion and FBGC formation readily on the RGD surface in the absence of any additional serum proteins. As alpha2-macroglobulin receptors are not found on blood monocytes but are expressed only with macrophage development, these results point to a potential interaction between adsorbed 2-macroglobulin and its receptors on macrophages during macrophage development and fusion. These studies identify important surface independent and dependent effects in foreign body reaction development that may be important in the identification of biological design criteria for molecularly engineered surfaces and tissue engineered devices.

Human monocyte/macrophage adhesion, macrophage motility, and IL-4-induced foreign body giant cell formation on silane-modified surfaces in vitro. Student Research Award in the Master's Degree Candidate Category, 24th Annual Meeting of the Society for Biomaterials, San Diego, CA, April 22-26, 1998.

A cytokine-based, in vitro model of foreign body giant cell (FBGC) formation was utilized to examine the effect of biomaterial surface chemistry on the adhesion, motility, and fusion of monocytes and macrophages. Human monocytes were cultured for 10 days on 14 different silane-modified glass surfaces, during which time the cells assumed the macrophage phenotype. The adhesion of monocytes and macrophages during the culture period decreased by an average of approximately 50%, with the majority of cell loss observed during days 1-3. Most important, the adhesion of monocytes and macrophages was surface independent except for two surfaces containing terminal methyl groups, which decreased adhesion levels. Interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) were added to the medium to induce FBGC formation and enhance macrophage adhesion, respectively. Surprisingly, GM-CSF decreased long-term monocyte/macrophage adhesion. IL-4-induced FBGC density was strongly influenced by the surface carbon content, as determined by X-ray photoelectron spectroscopy (XPS). In contrast, contact angle and surface energy displayed no correlation with FBGC formation. The motility of adherent macrophages, as measured by time-lapse confocal microscopy, was not affected significantly by differences in surface chemistry or the addition of cytokines. The surface dependence of FBGC formation is hypothesized to be the result of varying levels of silane-derived surface carbon.

Interleukin-13 induces human monocyte/macrophage fusion and macrophage mannose receptor expression.

Inasmuch as we recently demonstrated that IL-4 is a strong inducer of monocyte/macrophage fusion and IL-13 has been observed to mimic many of the biologic effects of IL-4, the ability of IL-13 to promote human macrophage fusion in vitro was tested and compared with IL-4-mediated fusion. IL-13 induced the fusion of monocyte-derived macrophages as potently as IL-4 under identical culture conditions, and resulted in foreign body-type giant cell formation. At optimal concentrations of cytokine added, statistically equal numbers of macrophages participated in IL-13- and IL-4-induced fusion (66.1 +/- 4.6% and 63.9 +/- 4.4%, respectively). However, the effects of IL-13 and IL-4 were not additive or synergistic, and the maximum fusion obtained when both IL-4 and IL-13 were added was 63.8 +/- 3.6%. Only anti-human IL-13 Abs inhibited IL-13-induced foreign body giant cell formation; the fusion-inducing effects of IL-13 continued to be observed in the presence of neutralizing Abs to IL-4 and several other anti-cytokine Abs, including Abs against IFN-gamma, granulocyte-macrophage CSF, IL-3, and TNF-alpha. IL-13 also significantly enhanced the fluorescence intensity detected by anti-human macrophage mannose receptor Abs, indicating that IL-13, like IL-4, up-regulates expression of the receptor that may be an essential participant in macrophage fusion. The results of this study demonstrate that IL-13, like IL-4, is a potent human macrophage fusion factor, and suggest that although IL-13 acts independently of IL-4 to promote foreign body giant cell formation, it may trigger a common mechanism for macrophage fusion.

Protein adsorption on chemically modified surfaces.

Following the implantation of a biomaterial, the first event to occur at the tissue-material interface is protein adsorption. Once proteins have adsorbed to the surface of a material, cells no longer see the material, but only the protein coated surface layer. This adsorbed protein layer can mediate the type of cells that adhere to the surface, which ultimately can determine the type of tissue that develops. In this experiment, glass and polystyrene surfaces were chemically modified forming ionic, non-ionic, hydrophobic, and hydrophilic surfaces. The modified surfaces were incubated in a RPMI diluted serum solution. After incubation, a radioimmunoassay was used to quantify the exposed proteins adsorbed onto the surface of the material. In general, albumin, complement C3 (C3), fibronectin (FN) and vitronectin (VN) competitively adsorbed to the modified surface in a similar fashion, whereas IgG adsorption was the opposite. The hydrophobic surfaces had higher adsorbance of the adhesion proteins (C3, FN and VN) compared to higher adsorption of albumin and IgG onto the hydrophilic surfaces. The surface mobility of the silane modified surfaces also affected the adhesion of proteins. The differences seen in protein adsorption did not directly correlate to monocyte adhesion and Foreign Body Giant Cell (FBGC) development on these surfaces.