Jian-Lin Yu

Adhesion of coagulase-negative staphylococci and adsorption of plasma proteins to heparinized polymer surfaces

Protease treatment of cells of coagulase-negative staphylococci reduced the adhesion of bacteria to heparinized polyethylene preadsorbed with serum. Fibronectin (Fn), fibrinogen (Fg), vitronectin, complement factor C3c, plasminogen, laminin and to a low extent albumin were detected on tridodecylmethylammonium chloride (TDMAC)-heparinized polyvinyl chloride (PVC) catheters extirpated from the circulation of patients. Using a perfusion model we show that during the first hours of perfusion with human plasma, Fn and Vn dominate, whereas after 22 h of perfusion Fg is the dominant protein. Field emission scanning electron microscopy and atomic force microscopy studies on TDMAC-heparinized catheters as well as on end-point attached heparinized PVC catheters indicate that quantitatively more Fg than Fn is exposed on these surfaces after prolonged exposure (> 22 h) to human plasma.

Fibronectin exposes different domains after adsorption to a heparinized and an unheparinized poly(vinyl chloride) surface

The adsorption of fibronectin to poly(vinyl chloride) catheters with end-point attached (EPA) heparin and tridodecylmethylammonium chloride-heparinized poly(vinyl chloride) was compared to that of unheparinized poly(vinyl chloride) using antibodies directed against four different domains of the protein. After perfusion of human plasma on the EPA-heparinized surface, the exposure of the N-terminal 29-kD fragment increased during the first 5 h of perfusion. Also, the exposure of the 30-kD gelatin-binding and 65-kD cell-binding fragments increased with time, but at a lower level. On the unheparinized catheter, low levels of antibodies bound to the different domains, and the binding showed little variation during the 5 h of plasma perfusion, indicating that the fibronectin molecule does not change configuration to a significant extent on this surface after the initial adsorption. When the EPA-heparinized surface was preabsorbed with human fibrinogen before incubation with fibronectin, significantly less of the 29-kD (fibrin-binding) domain was exposed, and the 30-kD domain was not exposed. Exposure of the 31- and 65-kD domains increased after preadsorption of fibrinogen to the surface. Since fibronectin has heparin-binding domains, it adsorbs differently to a heparinized versus an unheparinized surface. This will influence subsequent binding of other proteins to the surface, as well as potential binding of microbes. The use of antibodies to defined domains of the fibronectin molecule provides a powerful tool in studies of configurational changes of fibronectin after adsorption to different surfaces.

A Bacteriologic and Scanning Electron Microscope Study after Implantation of Foreign Bodies in the Biliary Tract in Rats

BACKGROUND Bacterial adherence to the stent surfaces, concomitant colonization, and possible stent blockage are the main complications after the use of biliary stents. The present study was assigned to investigate bacteriologic and morphologic changes in the biliary tract after the implantation of biliary drain materials. METHODS Rubber and silicone pieces with a surface area of 1 cm2 were implanted into the biliary tract in rats after temporary obstruction of the common bile duct by the use of a mini-occluder. The animals were killed at 4, 8 and 14 weeks, respectively, after implantation, and the implants were retrieved, cultured, and examined by scanning electron microscopy (SEM). Bacterial culture and SEM were also performed on tissue samples obtained from the mucosal surface of the biliary tract. RESULTS Bacterial colonization and biofilm formation were found on the surfaces of the implanted materials and on the mucosal surface of the biliary tract in animals with implants but not on the biliary tract mucosa in rats without implants. CONCLUSION Foreign bodies implanted in the biliary tract facilitate bacterial adherence not only to the surface of the implants but also to the mucosal surface in the biliary tract.

Experimental foreign-body infection in the biliary tract in rats.

BACKGROUND Biomaterials used for biliary drainage may potentially result in biomaterial-associated infections. METHODS Foreign-body infection in the biliary tract was investigated in rats. Rubber drain pieces were implanted in the biliary tract in rats for 1-4 weeks, followed by challenges with 10(2) to 10(4) colony-forming units (cfu) Escherichia coli injected into the common bile duct. The rate of infection was calculated, the bacterial growth in the biliary tract was observed over 72 h after challenges, and the opsonic activity in bile and in sera was assessed. RESULTS In the group with drain material, inocula as small as 10(2) cfu produced persisting infection in the common bile duct in 90% of animals, whereas the same number of E. coli infected only 30% of rats in the control and sham implantation groups (p < 0.01, chi-square test). Complement-mediated opsonic activity in bile of animals with implanted drain pieces gradually decreased with time, whereas opsonic activity in sera from the same animals remained unchanged. CONCLUSIONS Implants in the biliary tract impair local host defense, resulting in an increased susceptibility to microbial infection.

Reduction of E. coli adherence to rubber slices treated with phospholipids

The present study aimed at modifying the surface of biliary drain material to reduce bacterial adherence. The adherence of cells of seven E. coli strains to rubber slices treated with phosphatidylcholin (PC) or phosphatidylinositol (PI) and the adherence of cells of E. coli strain NG7C to PC‐ or PI‐treated rubber slices implanted in the common bile duct in rats were studied in vitro. The rubber slices were incubated with 1 times 107 cfu radiolabeled E. coli cells/ml at 37°C for 60 min and then drained and washed thrice in 2 ml PBS, and adherent E. coli cells were quantified by radioactivity counting. The results show that both PC and PI absorbed on the surface of slices reduced the adherence of E. coli cells in at least two ways, i.e. by changing surface properties in vitro and by reducing deposition of host‐derived molecules on phospholipid‐treated surfaces in vivo. The results may be of use for modification of the biomaterial surface in the clinical situation.

Studying Bacterial Adhesion to Biliary Stents

Endoscopie biliary stenting has become an effective treatment for obstructive jaundice since its introduction by Soehendra and Reijinders-Fredrix.66 Randomized trials have shown it to be the preferred approach for the palliation of malignant biliary obstruction when surgical process is not possible.1,62,65,69 Stenting provides relief of jaundice with low morbidity, and it significantly improves patients’ quality of life.3 In benign conditions, endoscopie stenting offers effective short term treatment.24,30 The major limitation to long term biliary stenting is the problem of late stent occlusion.20 Once a stent is placed in the bile duct, an encrustation of amorphous material and bacteria starts to accumulate on its surfaces.27 Given sufficient time, the lumen becomes occluded, bile flow ceases, and the patient develops symptoms of recurrent biliary obstruction, complicated by cholangitis and sepsis. Late clogging is clearly the most important complication of long term treatment with stents. Numerous studies of bacterial adhesion to bile stents have been conducted to explore mechanisms behind stent clogging since the late 1980s, a decade after endoscopie biliary stenting was introduced. In this chapter, bacteriology and defense mechanisms in the biliary tract are briefly reviewed and the methodology for the study of bacterial adhesion to biliary stents described.

Binding of immobilized fibronectin by biliary drain isolates

Occlusion of biliary stents, as the result of bacterial adhesion and colonization onto biliary stents, still remains a major problem. Biliary proteins, such as fibronectin (Fn) and vitronectin (Vn), have been presumed to be involved in the process of bacterial adhesion to biliary biomaterial. In the present study, Fn binding by 5 strains of E. coli isolated from biliary drains or from bile was studied. All strains did not bind detectable amounts of soluble Fn but bound to immobilized plasma Fn. Adhesion of four strains of E. coli to ovalbumin was reduced by periodate treatment of ovalbumin, but adhesion to Fn was unaffected. Adhesion was inhibited by mannose-containing saccharides, trypsin treatment of the protein, and protease treatment of the bacterial cells. Autoradiography showed that components of cell extracts from three E. coli strains bind 125I-Fn but not a 150 kD Fn fragment. The findings indicate that the adhesion of these bacteria to Fn is a protein-protein interaction, inhibited by D-mannose, and possibly mediated by fimbrial components.