Examination of intestinal ultrastructure, bowel wall apoptosis and tight junctions in the early phase of sepsis.

Gut hyperpermeability can be caused by either apoptosis of the intestinal epithelium or altered status, permeability or porosity of tight junctions. This project aims to elucidate these mechanisms in the early phase of sepsis. Eighteen male wild type mice were randomized to two groups. All mice received one single gavage of fluorescein isothiocyanate (FITC) dextran 30 min before intervention. One group (n = 10) underwent cecal ligation and puncture to induce sepsis. The other group (n = 8) was sham operated. Septic animals exhibited significantly increased permeability for FITC 8 h post-operatively. Significantly increased serum interleukin-6, tumor-necrosis-factor-alpha and interleukin-1-beta confirmed sepsis. Septic animals showed significant bowel wall inflammation of ileum and colon samples. PCR revealed significantly increased expression of claudin-2 and decreased expressions of claudin-4, tight-junction-protein-1 and occludin-1 resembling increased permeability of tight junctions. However, these alterations could not be confirmed at the protein level. Light microscopy revealed significant dilatation of intercellular spaces at the basal sections of intestinal epithelial cells (IEC) in septic animals confirmed by increased intercellular spaces at the level of tight junctions and adherens junctions in electron microscopy (TEM). In small angle X-ray scattering no increase in number or size of nanopores could be shown in the bowel wall. HOECHST staining and PCR of ileum samples for apoptosis markers proofed no relevant differences in intestinal epithelial cell apoptosis between the groups. Intestinal hyperpermeability in septic animals was most likely caused by alterations of the intercellular contacts and not by apoptosis or increased size/number of nanopores of intestinal epithelial cells in this murine model of early sepsis.

Bone-implant degradation and mechanical response of bone surrounding Mg-alloy implants.

In the present paper, first results of the influence of the degradation of biodegradable materials on the hardness of the bone are presented in detail. For this purpose, different materials (Mg, Ti and biopolymers) were implanted into the femora of growing rats and bone cross sections were examined for the micro-hardness (MH). The aim of the present paper was to examine the mechanical response of the bone areas surrounding the implant at defined sites and at specified periods after implantation. A special focus was set on Mg alloys. In earlier in-vitro and in-vivo studies, an accumulation of Magnesium in the vicinity of the implant was detected by using different techniques. Therefore, micro-hardness measurements were performed, and the mechanical strength of bone was correlated with the exchange of Magnesium and Calcium in Hydroxyapatite. After the operation and implantation, the micro-hardness values became temporarily lower, but after complete degradation of the implants, the values were identical with those of specimens containing no implants.

The potential of isotopically enriched magnesium to study bone implant degradation in vivo.

This pilot study highlights the substantial potential of using isotopically enriched (non-radioactive) metals to study the fate of biodegradable metal implants. It was possible to show that magnesium (Mg) release can be observed by combining isotopic mass spectrometry and isotopic pattern deconvolution for data reduction, even at low amounts of Mg released a from slowly degrading 26Mg enriched (>99%) Mg metal. Following implantation into rats, structural in vivo changes were monitored by µCT. Results showed that the applied Mg had an average degradation rate of 16±5µmyear-1, which corresponds with the degradation rate of pure Mg. Bone and tissue extraction was performed 4, 24, and 52weeks after implantation. Bone cross sections were analyzed by laser ablation inductively coupled plasma mass spectrometry (ICP-MS) to determine the lateral 26Mg distribution. The 26Mg/24Mg ratios in digested tissue and excretion samples were analyzed by multi collector ICP-MS. Isotope pattern deconvolution in combination with ICP-MS enabled detection of Mg pin material in amounts as low as 200ppm in bone tissues and 20ppm in tissues up to two fold increased Mg levels with a contribution of pin-derived Mg of up to 75% (4weeks) and 30% (24weeks) were found adjacent to the implant. After complete degradation, no visual bone disturbance or residual pin-Mg could be detected in cortical bone. In organs, increased Δ26Mg/24Mg values up to 16‰ were determined compared to control samples. Increased Δ26Mg/24Mg values were detected in serum samples at a constant total Mg level. In contrast to urine, feces did not show a shift in the 26Mg/24Mg ratios. This investigation showed that the organism is capable of handling excess Mg well and that bones fully recover after degradation. STATEMENT OF SIGNIFICANCE: Magnesium alloys as bone implants have faced increasing attention over the past years. In vivo degradation and metabolism studies of these implant materials have shown the promising application in orthopaedic trauma surgery. With advance in Mg research it has become increasingly important to monitor the fate of the implant material in the organism. For the first time, the indispensible potential of isotopically enriched materials is documented by applying 26Mg enriched Mg implants in an animal model. Therefore, the spatial distribution of pin-Mg in bone and the pin-Mg migration and excretion in the organism could be monitored to better understand metal degradation as well as Mg turn over and excretion.

µXRF Elemental Mapping of Bioresorbable Magnesium-Based Implants in Bone.

This study investigated the distribution of the elemental constituents of Mg-based implants at various stages of the degradation process in surrounding bone tissue, with a focus on magnesium (Mg), as the main component of the alloy, and yttrium (Y), due to its potential adverse health effects. The measurements were performed on the implant-bearing thin sections of rat bone in a time series of implant degradation between one and 18 months. Micro X-ray fluorescence analysis (µXRF) with a special spectrometer meeting the requirements for the measurements of low-Z elements was used. It was found that the migration and accumulation behaviour of implant degradation products is element-specific. A sharp decrease in Mg was observed in the immediate vicinity of the interface and no specific accumulation or aggregation of Mg in the adjacent bone tissue was detected. By contrast, Y was found to migrate further into the bone over time and to remain in the tissue even after the complete degradation of the implant. Although the nature of Y accumulations must still be clarified, its potential health impact should be considered.

Biodegradable Fe-based alloys for use in osteosynthesis: outcome of an in vivo study after 52 weeks.

This study investigates the degradation performance of three Fe-based materials in a growing rat skeleton over a period of 1 year. Pins of pure Fe and two Fe-based alloys (Fe-10 Mn-1Pd and Fe-21 Mn-0.7C-1Pd, in wt.%) were implanted transcortically into the femur of 38 Sprague-Dawley rats and inspected after 4, 12, 24 and 52 weeks. The assessment was performed by ex vivo microfocus computed tomography, weight-loss determination, surface analysis of the explanted pins and histological examination. The materials investigated showed signs of degradation; however, the degradation proceeded rather slowly and no significant differences between the materials were detected. We discuss these unexpected findings on the basis of fundamental considerations regarding iron corrosion. Dense layers of degradation products were formed on the implants' surfaces, and act as barriers against oxygen transport. For the degradation of iron, however, the presence of oxygen is an indispensable prerequisite. Its availability is generally a critical factor in bony tissue and rather limited there, i.e. in the vicinity of our implants. Because of the relatively slow degradation of both pure Fe and the Fe-based alloys, their suitability for bulk temporary implants such as those in osteosynthesis applications appears questionable.