Advanced material modelling in numerical simulation of primary acetabular press-fit cup stability

Primary stability of artificial acetabular cups, used for total hip arthroplasty, is required for the subsequent osteointegration and good long-term clinical results of the implant. Although closed-cell polymer foams represent an adequate bone substitute in experimental studies investigating primary stability, correct numerical modelling of this material depends on the parameter selection. Material parameters necessary for crushable foam plasticity behaviour were originated from numerical simulations matched with experimental tests of the polymethacrylimide raw material. Experimental primary stability tests of acetabular press-fit cups consisting of static shell assembly with consecutively pull-out and lever-out testing were subsequently simulated using finite element analysis. Identified and optimised parameters allowed the accurate numerical reproduction of the raw material tests. Correlation between experimental tests and the numerical simulation of primary implant stability depended on the value of interference fit. However, the validated material model provides the opportunity for subsequent parametric numerical studies.     

Establishment of a Novel In Vitro Test Setup for Electric and Magnetic Stimulation of Human Osteoblasts

When large defects occur, bone regeneration can be supported by bone grafting and biophysical stimuli like electric and magnetic stimulation (EMS). Clinically established EMS modes are external coils and surgical implants like an electroinductive screw system, which combines a magnetic and electric field, e.g., for the treatment of avascular bone necrosis or pseudarthrosis. For optimization of this implant system, an in vitro test setup was designed to investigate effects of EMS on human osteoblasts on different 3D scaffolds (based on calcium phosphate and collagen). Prior to the cell experiments, numerical simulations of the setup, as well as experimental validation, via measurements of the electric parameters induced by EMS were conducted. Human osteoblasts (3 × 10⁵ cells) were seeded onto the scaffolds and cultivated. After 24 h, screw implants (Stryker ASNIS III s-series) were centered in the scaffolds, and EMS was applied (3 × 45 min per day at 20 Hz) for 3 days. Cell viability and collagen type 1 (Col1) synthesis were determined subsequently. Numerical simulation and validation showed an adequate distribution of the electric field within the scaffolds. Experimental measurements of the electric potential revealed only minimal deviation from the simulation. Cell response to stimulation varied with scaffold material and mode of stimulation. EMS-stimulated cells exhibited a significant decrease of metabolic activity in particular on collagen scaffolds. In contrast, the Col1/metabolic activity ratio was significantly increased on collagen and non-sintered calcium phosphate scaffolds after 3 days. Exclusive magnetic stimulation showed similar but nonsignificant tendencies in metabolic activity and Col1 synthesis. The cell tests demonstrate that the new test setup is a valuable tool for in vitro testing and parameter optimization of the clinically used electroinductive screw system. It combines magnetic and electric stimulation, allowing in vitro investigations of its influence on human osteoblasts.     

Effect of propionate on the utilization of nitrogen from 15NH4Cl for urea synthesis in hepatocytes isolated from sheep liver

1. The effect of ornithine (2.0 mM) and propionate (5.0 mM) on the utilization of N from 15NH4Cl (5.0 mM) for urea synthesis in hepatocytes isolated from sheep liver was investigated. 2. The capacity of sheep hepatocytes to utilize [15N]ammonia in the absence of the other exogenous substrates was very low and amounted 132 +/- 37.3 mumol/hr per 1 g dry wt. 3. Ornithine failed to affect the total [15N]ammonia uptake and total urea synthesis, but at the same time it markedly increased the utilization of [15N]ammonia for ureagenesis and diminished the rate of urea synthesis from endogenous sources. 4. Propionate markedly increased total [15N]ammonia utilization and total urea formation; this increase resulted from the rise of ammonia utilization for urea synthesis and it was similar in the presence or absence of ornithine. 5. The capacity of sheep liver cells to utilize ammonia in the presence of propionate (in the presence or absence of ornithine) amounted to 256 mumol/hr per 1 g dry wt, thus being similar to the values in vivo. 6. It is concluded that in sheep hepatocytes both ornithine and propionate stimulate the utilization of ammonia for urea synthesis and these effects take place independently and occur by different mechanisms.     

Physiological Concentration of Exogenous Lactate Reduces Antimycin A Triggered Oxidative Stress in Intestinal Epithelial Cell Line IPEC-1 and IPEC-J2 In Vitro

Weaning triggers an adaptation of the gut function including luminal lactate generation by lactobacilli, depending on gastrointestinal site. We hypothesized that both lactobacilli and lactate influence porcine intestinal epithelial cells. In vivo experiments showed that concentration of lactate was significantly higher in gastric, duodenal and jejunal chyme of suckling piglets compared to their weaned counterparts. In an in vitro study we investigated the impact of physiological lactate concentration as derived from the in vivo study on the porcine intestinal epithelial cells IPEC-1 and IPEC-J2. We detected direct adherence of lactobacilli on the apical epithelial surface and a modulated F-actin structure. Application of lactobacilli culture supernatant alone or lactate (25 mM) at low pH (pH 4) changed the F-actin structure in a similar manner. Treatment of IPEC cultures with lactate at near neutral pH resulted in a significantly reduced superoxide-generation in Antimycin A-challenged cells. This protective effect was nearly completely reversed by inhibition of cellular lactate uptake via monocarboxylate transporter. Lactate treatment enhanced NADH autofluorescence ratio (F_cytosol/F_nucleus) in non-challenged cells, indicating an increased availability of reduced nucleotides, but did not change the overall ATP content of the cells. Lactobacilli-derived physiological lactate concentration in intestine is relevant for alleviation of redox stress in intestinal epithelial cells.     

The fate of nitrogen from 15N-labeled nitrate after single intravenous administration of Na15NO3 in sheep

The metabolic fate of nitrogen from 15N-labeled sodium nitrate has been investigated in four healthy Polish Merino ewes. 15N-labeled sodium nitrate was administered intravenously at the dosage of 400 micromol.kg(-1) body weight. Blood plasma and urine concentrations of nitrate, ammonia, and urea and 15N enrichment of ammonia and urea were estimated over a 50-h period following 15N-nitrate administration. Nitrate (NO3-) was slowly eliminated from the blood plasma, and the presence of NO3(-) in the blood plasma above the nitrate "background" was observed for 50 h. 15N enrichment of blood plasma urea already appeared at 15 min and reached the maximum 6 h after 15N-nitrate administration. The urinary excretion of nitrate occured during 50 h after 15N-nitrate injection; the total urine excretion of NO3(-) was 23.63+/-2.39% of the administered dose. The mean urinary recoveries of nitrogen as 15N-urea and 15N-ammonia were 14.76+/-1.32% and 0.096+/-0.015% of the administered 15N-nitrate dose, respectively. It should be pointed out that in total only 38.49% of the administered nitrate-N was excreted in urine (as nitrate, ammonia and urea nitrogen) during 50 h. The results obtained indicate that sheep are able to store nitrate nitrogen in their body. The fate of the remaining approximately 60% of the 15NO3(-) administered dose is unknown. The results obtained do not allow one to conclude what fraction of the unrecovered approximately 60% of the 15NO3(-) dose was utilized by gastrointestinal microorganisms, and (or) metabolized, or stored in sheep tissues.     

Advanced material modelling in numerical simulation of primary acetabular press-fit cup stability

Primary stability of artificial acetabular cups, used for total hip arthroplasty, is required for the subsequent osteointegration and good long-term clinical results of the implant. Although closed-cell polymer foams represent an adequate bone substitute in experimental studies investigating primary stability, correct numerical modelling of this material depends on the parameter selection.

Material parameters necessary for crushable foam plasticity behaviour were originated from numerical simulations matched with experimental tests of the polymethacrylimide raw material. Experimental primary stability tests of acetabular press-fit cups consisting of static shell assembly with consecutively pull-out and lever-out testing were subsequently simulated using finite element analysis.

Identified and optimised parameters allowed the accurate numerical reproduction of the raw material tests. Correlation between experimental tests and the numerical simulation of primary implant stability depended on the value of interference fit. However, the validated material model provides the opportunity for subsequent parametric numerical studies.     

Establishment of intestinal microbiota with focus on yeasts of unweaned and weaned piglets kept under different farm conditions

This study aimed to characterize the intestinal yeasts in weaning piglets and to establish their possible relationships with main bacterial groups. German Landrace piglets were weaned (WP, n=32) at 28 days of age or kept with the dams until day 39 without creep feed (UP, n=32). The experiment was performed at an experimental and a commercial farm (CF). Faeces were collected from the piglets, sows and pen floors on days 28, 33 and 39 for isolation of DNA and cultivation for enumeration of yeasts, enterobacteria, enterococci and lactobacilli. Fragments of the D1 domain of 26S rRNA gene were amplified and separated by denaturing gradient gel electrophoresis (DGGE). No yeasts could be cultured from water and feed samples. No or only low numbers of yeasts were detected among all UP. In WP at CF, yeasts correlated with lactobacilli (r=0.456; P=0.009) and enterobacteria (r=-0.407; P=0.021). Kazachstania slooffiae dominated among the cultured yeasts. It was the only yeast species detected by PCR-DGGE. Yeasts, especially K. slooffiae, established in the porcine gastrointestinal tract after consumption of grain-based feed and may interrelate with the intestinal microbiota. The study provides data indicating importance of K. slooffiae for the development of balanced porcine gut microbiota.     

Changes in the diversity of pig ileal lactobacilli around weaning determined by means of 16S rRNA gene amplification and denaturing gradient gel electrophoresis

Our study aimed to provide a comprehensive characterization of changes in porcine intestinal Lactobacillus populations around the time of weaning based on 16S rRNA gene amplification and denaturing gradient gel electrophoresis (DGGE). DNA was extracted from the ileal contents of piglets at weaning (28 days of age) and after 1, 2, 5 and 11 days. PCR amplicons (V2-V3 fragments of 16S rRNA genes) were separated using DGGE. Predominant bands were excised and sequenced after reamplification. A band corresponding to Lactobacillus salivarius was present 1 and 2 days post-weaning (pw), while Lactobacillus crispatus was detected only 1 and 11 days pw. Lactobacillus sobrius gave the most dominant band in all animals. The number of bands decreased from 13+/-3 at weaning to 9+/-1 at 5 days pw, but the species richness had recovered by 11 days pw. The similarity of profiles between sampling days was high for 1 and 2 days pw (>91%), but was low for 5 and 11 days pw (<59%). The diversity of the profiles was lower 5 days pw, based on the Shannon diversity index (0.83+/-0.076 vs. 1.02+/-0.127 at weaning, P=0.042), but had recovered to preweaning values by 11 days pw. The application of group-specific DGGE showed that the Lactobacillus community within the porcine ileum undergoes dramatic, partly reversible changes as a consequence of weaning.     

Utilization of nitrogen from 15NH4Cl and [15N]alanine for urea synthesis in hepatocytes from fed and starved rats

Utilization of N from 15NH4Cl and [15N]alanine for urea synthesis in hepatocytes isolated from fed and 24 hr starved rats was investigated. In hepatocytes isolated from fed rats, 54 and 65% of the added [15N]ammonia was utilized for urea synthesis in the presence of 0.5 and 2.0 mM NH4Cl, respectively. This utilization of [15N]ammonia in hepatocytes from starved rats was 2-fold lower. The amount of urea synthetized from endogenous sources was, in the presence of 0.5 and 2.0 mM NH4Cl, about 44 and 60% higher than in the control conditions (without NH4Cl). The considerable amount of added ammonia (30-44%) was utilized in processes other than urea synthesis. Alanine markedly diminished the utilization of 15N from NH4Cl in hepatocytes from both fed and starved rats. In these conditions (NH4Cl present), alanine significantly increased the urea formation in hepatocytes from starved rats and failed to affect the urea production in hepatocytes from fed rats. On the basis of 15N determination, it was concluded that both NH4Cl and alanine caused an increase in the utilization of nitrogen from endogenous sources in rat hepatocytes. This conclusion is in contrast with the results based only on the changes in ammonia and urea concentrations.