Antimicrobial Silver Multilayer Coating for Prevention of Bacterial Colonization of Orthopedic Implants.

Due to increasing rates of periprosthetic joint infections (PJI), new approaches are needed to minimize the infection risk. The first goal of this study was to modify a well-established infection model to test surface-active antimicrobial systems. The second goal was to evaluate the antimicrobial activity of a silver multilayer (SML) coating. In vitro tests with SML items showed a >4 Log reduction in a proliferation assay and a 2.2 Log reduction in an agar immersion test (7 d). In the in vivo model blank and SML coated K-wires were seeded with ~2 × 104 CFU of a methicillin-sensitive Staphylococcus epidermidis (MSSE) and inserted into the intramedullary tibial canal of rabbits. After 7 days, the animals were sacrificed and a clinical, microbiological and histological analysis was performed. Microbiology showed a 1.6 Log pathogen reduction on the surface of SML items (p = 0.022) and in loosely attached tissue (p = 0.012). In the SML group 7 of 12 SML items were completely free of pathogens (cure rate = 58%, p = 0.002), while only 1 of 12 blank items were free of pathogens (cure rate = 8%, p = 0.110). No silver was detected in the blood or urine of the SML treated animals and only scarcely in the liver or adjacent lymph nodes. In summary, an in vivo infection model to test implants with bacterial pre-incubation was established and the antimicrobial activity of the SML coating was successfully proven.

Standard Biocompatibility Studies Do Not Predict All Effects of PVA/CMC Anti-Adhesive Gel in vivo.

PURPOSE: PVA/CMC (polyvinyl alcohol/carboxymethyl cellulose) hydrogel fulfills various physiochemical properties required for an adhesion barrier and has shown good anti-adhesion properties in previous in vivo studies. In this investigation, we assessed the in vitro and in vivo biocompatibility of PVA/CMC gel and compared this to the functionality and promotion of wound healing for two surgical indications. METHODS: Standardized ISO10993 in vitro and in vivo biocompatibility studies, comprising cytotoxicity, genotoxicity, acute systemic toxicity, delayed contact and maximization sensitization test, intracutaneous reactivity and local muscle implantation, were performed on PVA/CMC gel. In the functional studies, PVA/CMC gel was applied - on the one hand - to a rabbit abdominal wall model enforced with a polypropylene mesh for testing the anti-adhesion properties and - on the other hand - to an end- to-end anastomosis model that was selected for surveying potential influences of different dosages of PVA/CMC gel on anastomotic wound healing. RESULTS: The ISO10993 methods indicated generally good biocompatibility properties, such as the absence of cytotoxic and mutagenic effects as well as no signs of systemic toxicity and sensitization potentials. No irritation effects were observed after the intracutaneous injection of lipophilic PVA/CMC sesame oil extract. However, the injection of hydrophilic PVA/CMC physiologic saline extract induced slight irritation. Following rabbit muscle implantation of the PVA membrane for 2, 4, 12, 26 and 52 weeks, a slight irritant effect was observed at 12 weeks due to the peak of phagocytosis. In the functionality tests, PVA/CMC gel showed good anti-adhesive effects in the abdominal wall model enforced with the mesh, with significantly lower and less tense adhesions compared to the untreated control. However, moderate signs of inflammation, especially in the spleen were observed after the intra-abdominal implantation of 3.3 ml PVA/CMC gel per kg body weight. In the end-to-end anastomosis model, PVA/CMC gel had no influence on wound healing. For dosages of 1-6 ml gel per treatment, no signs of intestinal leaks were detected, and tensile strength was equal to that of the untreated control, but again more moderate signs of inflammation in the spleen were observed at a dosage >3 ml. CONCLUSION: Comparing the standardized ISO10993 methods, anti-adhesive PVA/CMC gel displays good biocompatibility. However, those methods do not seem to be sensitive enough because the rabbit abdominal wall and the end-to-end anastomosis models display more effects with respect to the dosage and routes of the intra-abdominal resorption of PVA/CMC gel - with the recommended <2 ml PVA/CMC gel per kg body weight as a secure dosage.

Triple fluorescence labelling of neuronal, glial and vascular markers revealing pathological alterations in various animal models.

The simultaneous detection of glia, vessels and neurons facilitates insights into the complex chemoarchitecture of the central nervous system. Here, we present a simple, robust and versatile approach for the carbocyanine triple fluorescence labelling of neuronal, vascular and glial markers. The usefulness of this procedure is shown for rat brain tissue under physiological conditions, after traumatic brain injury caused by controlled cortical impact injury, and after stroke following middle cerebral artery occlusion. Moreover, the versatility of the method is verified by its application to sections from old triple transgenic mice with age-dependent beta-amyloidosis and tau hyperphosphorylation in the hippocampus, modelling neuropathological alterations in Alzheimer's disease. To exemplify the usefulness of the approach for analysis of the enteric nervous system, it was applied to whole mounts from the horse intestine. The biotinylated lectin from potato (Solanum tuberosum) is presented as an excellent tool to detect both vessels and microglia. Furthermore, this lectin revealed macrophages after experimental insults, and senile plaques in aged triple transgenic mice. A large portion of astroglia was demonstrated by immunolabelling of glial fibrillary acidic protein. Neurons were detected by monoclonal antibodies directed against neuronal nuclei and, in horse tissues, mouse-anti-HuC/D recognizing a conserved nuclear protein. Confocal laser-scanning microscopy elucidated spatial relationships of the relevant markers and their pathological alterations after experimental insults and in transgenic mice with Alzheimer-like lesions.

Immunohistochemical characterization and quantitative analysis of neurons in the myenteric plexus of the equine intestine.

The present study was performed on whole-mount preparations to investigate the chemical neuroanatomy of the equine myenteric plexus throughout its distribution in the intestinal wall. The objective was to quantify neurons of the myenteric plexus, especially the predominant cholinergic and nitrergic subpopulations. Furthermore, we investigated the distribution of vasoactive intestinal polypeptide and the calcium-binding protein calretinin. Samples from different defined areas of the small intestine and the flexura pelvina were taken from 15 adult horses. After fixation and preparation of the tissue, immunofluorescence labeling was performed on free floating whole-mounts. Additionally, samples used for neuropeptide staining were incubated with colchicine to reveal the neuropeptide distribution within the neuronal soma. The evaluation was routinely accomplished using confocal laser-scanning microscopy. For quantitative and qualitative analysis, the pan-neuronal marker anti-HuC/D was applied in combination with the detection of the marker enzymes for cholinergic neurons and nitrergic nerve cells. Quantitative data revealed that the cholinergic subpopulation is larger than the nitrergic one in several different locations of the small intestine. On the contrary, the nitrergic neurons outnumber the cholinergic neurons in the flexura pelvina of the large colon. Furthermore, ganglia are more numerous in the small intestine compared with the large colon, but ganglion sizes are bigger in the large colon. However, comparison of the entire population of neurons in the different locations of the gut showed no difference. The present study adds further data on the chemoarchitecture of the myenteric plexus which might facilitate the understanding of several gastrointestinal disorders in the horse.