Exhalomics as a noninvasive method for assessing rumen fermentation in dairy cows: Can exhaled-breath metabolomics replace rumen sampling?

Previously, we used secondary electrospray ionization-mass spectrometry (SESI-MS) to investigate the diurnal patterns and signal intensities of exhaled (EX) volatile fatty acids (VFA) of dairy cows. The current study aimed to validate the potential of an exhalomics approach for evaluating rumen fermentation. The experiment was conducted in a switchback design, with 3 periods of 9 d each, including 7 d for adaptation and 2 d for sampling. Four rumen-cannulated original Swiss Brown (Braunvieh) cows were randomly assigned to 1 of 2 diet sequences (ABA or BAB): (A) low starch (LS; 6.31% starch on a dry matter basis) and (B) high starch (HS; 16.2% starch on a dry matter basis). Feeding was once per day at 0830 h. Exhalome (with the GreenFeed System), and rumen samples were collected 8 times to represent every 3 h of a day, and EX-VFA and ruminal (RM)-VFA were analyzed using SESI-MS and HPLC, respectively. Furthermore, the VFA concentration in the gas phase (HR-VFA) was predicted based on RM-VFA and Henry's Law (HR) constants. No interactions were identified between the types of diets (HS vs. LS) and the measurement methods on daily average VFA profiles (RM vs. EX or HR vs. EX), suggesting a consistent performance among the methods. Additionally, when the 3-h interval VFA data from HS and LS diets were analyzed separately, no interactions were observed between methods and time of day, indicating that the relative daily pattern of VFA molar proportions was similar regardless of the VFA measurement method used. The results revealed that the levels of acetate sharply increased immediately after feeding, trailed by an increase in the acetate:propionate ratio and a steady increase for propionate (2 h after feeding the HS diet, 4 h for LS), and butyrate. This change was more pronounced for the HS diet than the LS diet. However, there was no overall diet effect on the VFA molar proportions, although the measurement methods affected the molar proportions. Furthermore, we observed a strong positive correlation between the levels of RM and EX acetate for both diets (HS: r = 0.84; LS: r = 0.85), RM and EX propionate (r = 0.74), and RM and EX acetate:propionate ratio (r = 0.80). Both EX-VFA and RM-VFA exhibited similar responses to feeding and dietary treatments, suggesting that EX-VFA could serve as a useful proxy for characterizing RM-VFA molar proportions to evaluate rumen fermentation. Similar relationships were observed between RM-VFA and HR-VFA. In conclusion, this study underscores the potential of exhalomics as a reliable approach for assessing rumen fermentation. Moving forward, research should further explore the depth of exhalomics in ruminant studies to provide a comprehensive insight into rumen fermentation metabolites, especially across diverse dietary conditions.

Effects of graded levels of dietary pomegranate peel on methane and nitrogen losses, and metabolic and health indicators in dairy cows.

This study aimed to quantify the effects of dietary inclusion of tannin-rich pomegranate peel (PP) on intake, methane and nitrogen (N) losses, and metabolic and health indicators in dairy cows. Four multiparous, late-lactating Brown Swiss dairy cows (796 kg body weight; 29 kg/d of energy corrected milk yield) were randomly allocated to 3 treatments in a randomized cyclic change-over design with 3 periods, each comprising 14 d of adaptation, 7 d of milk, urine, and feces collection, and 2 d of methane measurements. Treatments were formulated using PP that replaced on a dry matter (DM) basis 0% (control), 5%, and 10% of the basal mixed ration (BMR) consisting of corn and grass silage, alfalfa, and concentrate. Gaseous exchange of the cows was determined in open-circuit respiration chambers. Blood samples were collected on d 15 of each period. Individual feed intake as well as feces and urine excretion were quantified, and representative samples were collected for analyses of nutrients and phenol composition. Milk was analyzed for concentrations of fat, protein, lactose, milk urea N, and fatty acids. Total phenols and antioxidant capacity in milk and plasma were determined. In serum, the concentrations of urea and bilirubin as well as the activities of alanine aminotransferase (ALT), aspartate aminotransferase, glutamate dehydrogenase, alkaline phosphatase, and γ-glutamyl transferase were measured. The data were subjected to ANOVA with the Mixed procedure of SAS, with treatment and period as fixed and animal as random effects. The PP and BMR contained 218 and 3.5 g of total extractable tannins per kg DM, respectively, and thereof 203 and 3.3 g of hydrolyzable tannins. Total DM intake, energy corrected milk, and methane emission (total, yield, and intensity) were not affected by PP supplementation. The proportions of C18:2n-6 and C18:3n-3 in milk increased linearly as the amount of PP was increased in the diet. Milk urea N, blood urea N, and urinary N excretion decreased linearly with the increase in dietary PP content. Total phenols and antioxidant capacity in milk and plasma were not affected by the inclusion of PP. The activity of ALT increased in a linear manner with the inclusion of PP. In conclusion, replacing up to 10% of BMR with PP improved milk fatty acid composition and alleviated metabolic and environmental N load. However, the elevated serum ALT activity indicates an onset of liver stress even at 5% PP, requiring the development of adaptation protocols for safe inclusion of PP in ruminant diets.

The application of Gadopentate-Dimeneglumin has no impact on progression free and overall survival as well as renal function in patients with monoclonal plasma cell disorders if general precautions are taken.

OBJECTIVES: The current analysis investigated the prognostic significance of gadopentetate dimeglumine on survival and renal function in patients with monoclonal plasma cell disorders. METHODS: In this study 263 patients who had received gadopentetate dimeglumine within a prospective trial investigating dynamic contrast-enhanced magnetic resonance imaging (MRI) were compared with 335 patients who had undergone routine, unenhanced MRI. RESULTS: We found no significant prognostic impact of the application of contrast agent on progression-free survival in patients with either monoclonal gammopathy of undetermined significance, smouldering or symptomatic myeloma and no significant prognostic impact on overall survival in patients with symptomatic myeloma. Since renal impairment is a frequent complication of myeloma, and decreased renal function is associated with a higher risk of complications in patients receiving contrast agents, we evaluated the impact of contrast agent on renal function after 1 year. In the present analysis the only significant adverse impact on kidney function occurred in symptomatic myeloma patients who already had impaired renal parameters at baseline. Here, the renal function did not recover during therapy, whereas it did so in patients with normal or only slightly impaired renal function. CONCLUSION: If general recommendations are adhered to, gadopentetate dimeglumine can be safely applied in patients with monoclonal plasma cell disease.

Emergence of tick-borne encephalitis in new endemic areas in Austria: 42 years of surveillance.

Human infections with tick-borne encephalitis (TBE)virus are a public health concern in certain regions of Europe, central and eastern Asia. Expansions of endemic areas and increased incidences have been associated with different factors including ecological changes supporting tick reproduction, socioeconomic changes increasing human outdoor activities and climatic changes favouring virus circulation in natural foci. Austria is among the most strongly affected countries in Central Europe, but the annual number of cases has strongly declined due to vaccination. Here,we have analysed changes of the incidence of TBE in the unvaccinated population of all federal states of Austria over a period of 42 years. The overall incidence in Austria has remained constant, but new strongly affected endemic regions have emerged in alpine valleys in the west of Austria. In parallel, the incidence in low-land regions in the north-east of the country is decreasing. There is no evidence for a shift to higher altitudes of infection sites in the traditional TBE zones,but the average altitudes of some newly established endemic areas in the west are significantly higher. Our analyses underscore the focal nature of TBE endemic areas and the potential of TBE virus to emerge in previously unaffected regions.

Tumour-site-dependent expression profile of angiogenic factors in tumour-associated stroma of primary colorectal cancer and metastases.

BACKGROUND: Tumour-associated stroma has a critical role in tumour proliferation. Our aim was to determine a specific protein expression profile of stromal angiogenic cytokines and matrix metalloproteinases (MMPs) to identify potential biomarkers or new therapy targets. METHODS: Frozen tissue of primary colorectal cancer (n=25), liver (n=25) and lung metastases (n=23) was laser-microdissected to obtain tumour epithelial cells and adjacent tumour-associated stroma. Protein expression of nine angiogenic cytokines and eight MMPs was analysed using a multiplex-based protein assay. RESULTS: We found a differential expression of several MMPs and angiogenic cytokines in tumour cells compared with adjacent tumour stroma. Cluster analysis displayed a tumour-site-dependent stromal expression of MMPs and angiogenic cytokines. Univariate analysis identified stromal MMP-2 and MMP-3 in primary colorectal cancer, stromal MMP-1, -2, -3 and Angiopoietin-2 in lung metastases and stromal MMP-12 and VEGF in liver metastases as prognostic markers (P>0.05, respectively). Furthermore, stroma-derived Angiopoietin-2 proved to be an independent prognostic marker in colorectal lung metastases. CONCLUSION: Expression of MMPs and angiogenic cytokines in tumour cells and adjacent tumour stroma is dependent on the tumour site. Stroma-derived MMPs and angiogenic cytokines may be useful prognostic biomarkers. These data can be helpful to identify new agents for a targeted therapy in patients with colorectal cancer.

Autologous stem cell transplantation for elderly patients with newly diagnosed multiple myeloma in the era of novel agents.

BACKGROUND: High-dose therapy (HDT) with autologous stem cell transplantation (ASCT) is considered the standard of care for multiple myeloma (MM) patients <65 years. Safety and outcome of ASCT for patients >65 years is currently uncertain, especially since the introduction of novel agents for induction and maintenance therapy. Furthermore, there are no conclusive data available on risk assessment in elderly patients treated with HDT. PATIENTS AND METHODS: We retrospectively analyzed 202 patients ≥60 years with newly diagnosed MM who underwent ASCT at our institution. Patients were stratified by age into three groups (60-64, 65-69 and 70-75 years). For safety assessment, we compared data about hospitalization, hematopoetic reconstitution and early mortality. Remission before and after ASCT was analyzed according to age and application of novel agents. Event-free (EFS) and overall survival (OS) were analyzed to identify impact of age, remission before/after ASCT and maintenance therapy as well as ISS score and cytogenetic aberrations on outcome in elderly patients. RESULTS: The assessment of safety, remission before/after ASCT as well as EFS and OS showed no significant differences between the three groups (median EFS: 60-64 years: 27 months; 65-69 years: 23 months; 70-75 years: 23 months; median OS: not reached). Patients receiving novel agents as part of induction therapy achieved significantly higher nCR + CR rates than patients treated without novel agents. In Cox regression analysis, ISS and cytogenetics as well as remission after ASCT had the highest prognostic impact on EFS and OS. Maintenance therapy was associated with longer EFS in uni- and multivariate analyses. CONCLUSION: ASCT is feasible for selected patients >65 and >70 years without increased mortality. Age at transplantation has no prognostic significance on outcome after ASCT. Novel agents during induction therapy and maintenance therapy improves outcome of older patients eligible for ASCT. ISS and cytogenetic analysis should be carried out routinely for risk assessment.

[The ABC's of medical statistics. Reading and understanding clinical trials]. / Das ABC der medizinischen Statistik. Klinische Studien lesen und verstehen.

Clinical trials test hypotheses that are accepted or rejected according to a predetermined probability of error (level of significance). Significance does not however mean relevance. Good parameters of relevance are absolute risk reduction and based on this the calculation of the number of patients who need to be treated for one additional patient to benefit. The randomized controlled trial is the gold standard for comparative evaluation of effects. In the ideal scenario it is designed so that a difference established by statistical methods becomes probable. In non-inferiority studies care should be taken that no equivalence is shown but rather that the difference is not greater than a predefined margin of error for differences. Meta-analyses of studies with similar endpoints have the potential to improve the level of evidence. Since the findings of meta-analyses depend on the studies included, critical assessment of the results is essential.

DNA Repair in mammalian cells: Mismatched repair: variations on a theme.

Complementary base pairing underlies the genetic template function of the DNA double helix. Therefore, to assure faithful DNA transactions, cells must adhere to a strict application of the Watson-Crick base pairing principle.Yet, mispairing does arise in DNA, most frequently as a result of DNA polymerase errors or base damage. These mismatches need be rectified to avoid mutation. Sometimes, however, mispairing is actively induced to trigger mutagenesis. This happens in activated B-lymphocytes, where the targeted generation and processing of G.U mismatches contributes to somatic hypermutation and antibody diversification. Non-mutagenic mismatches arise in heteroduplex intermediates of homologous recombination, and their processing helps restrict homeologous recombination. Depending on the type of mismatch and the biological context of its occurrence, cells must apply appropriate strategies of repair to properly control mutagenesis. This review will illustrate conceptual and functional challenges of cellular mismatch correction on typical examples of mutagenic base-base mismatches. (Part of a Multi-author Review).

[The ABC's of medical statistics. Reading and understanding clinical trials]. / Das ABC der medizinischen Statistik. Klinische Studien lesen und verstehen.

Clinical trials test hypotheses that are accepted or rejected according to a predetermined probability of error (level of significance). Significance does not however mean relevance. Good parameters of relevance are absolute risk reduction and based on this the calculation of the number of patients who need to be treated for one additional patient to benefit. The randomized controlled trial is the gold standard for comparative evaluation of effects. In the ideal scenario it is designed so that a difference established by statistical methods becomes probable. In non-inferiority studies care should be taken that no equivalence is shown but rather that the difference is not greater than a predefined margin of error for differences. Meta-analyses of studies with similar endpoints have the potential to improve the level of evidence. Since the findings of meta-analyses depend on the studies included, critical assessment of the results is essential.

Xenon-133: ambient activity from nuclear power stations.

The average activity of xenon-133 within and at approximately 100 kilometers from Albany, New York, from April to July 1975 was 2.6 picocuries per cubic meter of air. The source was gaseous effluents from boiling water reactors located in the northeastern United States. Its 5.29-day half-life makes xenon-133 an appropriate isotope to observe for the study of regional and hemispheric dispersion of pollutants.

Radioactive plume from the Three Mile Island accident: xenon-133 in air at a distance of 375 kilometers.

The transit of an air mass containing radioactive gas released from the Three Mile Island reactor was recorded in Albany, New York, by measuring xenon-133. These measurements provide an evaluation of Three Mile Island effluents to distances greater than 100 kilometers. Two independent techniques identified xenon-133 in ambient air at concentrations as high as 3900 picocuries per cubic meter. The local gamma-ray whole-body dose from the passing radioactivity amounted to 0.004 millirem, or 0.004 percent of the annual dose from natural sources.

Transfer of total phenols from a grapeseed-supplemented diet to dairy sheep and goat milk, and effects on performance and milk quality.

Polyphenols are known to affect digestion of ruminants, whereas there is little information about their metabolic effects. In a 2 × 2-factorial experiment the effects of supplementing a phenolic grapeseed extract were compared in 11 East Friesian dairy sheep and 9 Saanen goats. The concentrate supplemented with 7.4 g/100 g DM grapeseed extract had contents of 3.5 g additional phenols/100 g DM and was compared with a low phenolic control concentrate. Performance, total phenols in blood, milk, urine and feces, antioxidant capacity of the blood, and saliva properties were examined. The experiment lasted for 11 wk from parturition to late lactation, with an initial adaptation phase of 1 wk. Milk yield was measured daily after weaning at about 7 wk after parturition. Blood, milk, saliva, feces and urine were sampled 4, 3, 2, 2 and 2 times per animal, respectively. The phenolic diet increased phenol concentrations in blood (+10% and 17% in wk 5 and wk 11, respectively) and in milk (+32% in wk 5) on some of the sampling weeks. There were no clear species differences in phenol concentrations in blood plasma, milk, urine and feces. However, at the end of the experiment the supplemented goats had a higher (48%) urinary phenol concentrations than the non-supplemented goats. A weak relationship (P < 0.05) was found between phenol intake and phenol excretion with milk for sheep but not goats. The phenolic diet did not influence blood antioxidant capacity and tannin binding capacity of the saliva. The saliva of the goats had a higher tannin binding capacity than sheep saliva. The effects of the extract on milk yield were inconsistent between sheep and goats. In general, goats had higher feed and nutrient intakes, were heavier, and yielded more milk. Additionally, milk protein and lactose contents were lower and milk urea content was higher in goats than sheep. In conclusion, supplementing grapeseed extract to sheep and goats elevated phenol concentrations in milk and blood to a certain extent, but most of the phenols were lost via urine. The study gave another indication that goats seem to have developed coping mechanisms like a higher salivary tannin binding capacity, mechanisms which are less pronounced in sheep.

Potential anti-inflammatory and anti-infectious effects of human milk oligosaccharides.

There is increasing evidence of the local effects within the gastro intestinal tract and the systemic functions of human milk oligosaccharides (HMO). In addition to the vast majority of in vitro data, animal studies underline the high potential of HMO to influence very different processes. HMO probably influence the composition of the gut microflora through effects on the growth of bifidus bacteria. Whether the concomitant low number of pathogenic microorganisms in breastfed infants is also caused by HMO is an intriguing question that still has yet to be proven. Due to the similarity of HMO to epithelial cell surface carbohydrates, an inhibitory effect on the adhesion of pathogens to the cell surface is most likely. If this could be shown in humans, HMO would provide a new way to prevent or treat certain infections. It would also indicate supplementing infant formula based on cow's milk with HMO, as those oligosaccharides are either not detectable or present only in low numbers in bovine milk. As some HMO can be absorbed and circulate in blood, systemic effects may also be influenced. Due to their similarities to selectin ligands, HMO have been tested in in vitro studies demonstrating their anti-inflammatory abilities. For example, it has been shown that sialic acid-containing oligosaccharides reduce the adhesion of leukocytes to endothelial cells, an indication for an immune regulatory effect of certain HMO. We cover these topics after a short introduction on the structures of HMO, with a particular emphasis on their blood group and secretor specificity.

A comprehensive classification of mandibular fractures: a preliminary agreement validation study.

This study evaluates a comprehensive classification system for mandibular fractures based on imaging analysis. The AO/ASIF scheme, defining three fracture types (A, B, C), three groups within each type (e.g. A1, A2, A3) and three subgroups within each group (e.g. A1.1, A1.2, A1.3) with increasing severity from A1.1 (lowest) to C3.3 (highest) was used. The mandible is divided into two vertical units (I and V), two lateral horizontal units (II and IV) and one central unit (III) comprising the symphyseal and parasymphyseal region. Type A fractures are non-displaced, type B are displaced and type C are multifragmentary/defect injuries. Groups and subgroups are further defined in the classification system. Two classification sessions using semi-automatic software with 7 and 9 surgeons were performed to evaluate 100 fracture cases in the first session and 50 in the second. Inter-observer reliability and individual rater's accuracy were evaluated by kappa coefficient and latent class analysis, respectively. The analysis of inter-observer agreement for the detailed coding showed kappa coefficients around 0.50 with higher agreement among raters in the vertical units. This system allows standardization of documentation of mandibular fractures, although improvement in the definition of categories and their application is required.

The msh2 gene of Schizosaccharomyces pombe is involved in mismatch repair, mating-type switching, and meiotic chromosome organization.

We have identified in the fission yeast Schizosaccharomyces pombe a MutS homolog that shows highest homology to the Msh2 subgroup. msh2 disruption gives rise to increased mitotic mutation rates and increased levels of postmeiotic segregation of genetic markers. In bandshift assays performed with msh2Delta cell extracts, a general mismatch-binding activity is absent. By complementation assays, we showed that S. pombe msh2 is allelic with the previously identified swi8 and mut3 genes, which are involved in mating-type switching. The swi8-137 mutant has a mutation in the msh2 gene which causes a truncated Msh2 peptide lacking a putative DNA-binding domain. Cytological analysis revealed that during meiotic prophase of msh2-defective cells, chromosomal structures were frequently formed; such structures are rarely found in the wild type. Our data show that besides having a function in mismatch repair, S. pombe msh2 is required for correct termination of copy synthesis during mating-type switching as well as for proper organization of chromosomes during meiosis.

Reconstruction of orbital wall defects: critical review of 72 patients.

Between January 1996 and December 2001, 72 out of 354 patients were included in a retrospective study analysing the outcome of repaired orbital wall defects. Selection was dependent on the availability of pre and postoperative CT scans and on ophthalmologic examination. In particular, orthoptical assessment was performed up to 1 year after operation. In 72 patients, 83 orbital wall defects were analysed and allocated to one of five categories. Accuracy and type of reconstruction were assessed in unilateral orbital wall defects (n=61) and compared with functional outcome. Reconstruction was performed by using PDS membrane (39%), calvarian bone (13%), titanium mesh (7%) or a combination of these materials (37%). Postoperatively, 91% of the patients had normal vision without double images within 20 degrees at every gaze. Accuracy of reconstruction correlated with severity of orbital injury and functional outcome. Functional outcome between category II and III fractures showed no significant difference. The medial margin of the lateral infraorbital fissure being preserved (category II fracture) facilitates reconstruction technically. Accuracy of orbital reconstruction is one important factor to obtain best functional outcome, but other determinants like displacement and/or atrophy of intramuscular cone fat should be considered.

Mineralization and mechanical properties of the canine mandible distraction wound following acute molding.

To investigate the influence of an acute single step callus manipulation immediately after distraction on mechanical properties and mineralization of the regenerate, custom made distraction devices were bilaterally placed in the mandibular angle of 15 beagle dogs, allowing to simultaneously compress and stretch the regenerate after completed linear distraction. The animals were divided in three groups (n=5): Group 1 and 2 underwent manipulation of the regenerate, group 3 remained in a linear position. After 42 (group1) and 90 (group 2 and 3) days of consolidation the animals were sacrificed. The mechanical properties were assessed in an Instron testframe and bone density quantified by quantitative computed tomography and three- dimensionally assessed (Scion Image processing and analysis software). After 6 weeks of consolidation 25% of the specimens reached a stiffness which was >/=90% of the mean values of the unoperated reference hemi-mandibles. After a 13 week consolidation period, 62.5% were as stiff as the referenced specimens. Manipulated regenerates, allowed to heal under stable conditions for 13 weeks, had the same mechanical properties as specimens that underwent pure linear distraction. A temporary but not significant delay of osseous healing had to be postulated for the stretched zone after 6 weeks, indicating this area to be more critical than the compressed area.

A rat histone H2B pseudogene is closely associated with the histone H1d gene.

A 9 kb EcoRI restriction fragment was isolated from a recombinant phage out of a rat genomic library. This DNA fragment contains a rat H1d histone gene, its flanking sequences and a H2B histone pseudogene closely associated with the H1d gene. A comparison of the H2B pseudogene with human H2B genes flanking regions reveals sequence homologies to a human H2B histone gene (Albig, W. et al. (1991) Genomics 10, 940-948).