Receptor Activator of Nuclear Factor kappa-B Ligand is Not Regulated During Chronic Osteomyelitis in Pigs.

Bone loss is a major complication of osteomyelitis and from numerous in-vitro studies, it has been concluded that the bone lysis is caused by elevated expression of the receptor activator of nuclear factor κB ligand (RANKL), leading to increased osteoclast activity. However, we failed to find any relationship between bone loss and osseous RANKL expression in a porcine model of acute and chronic implant-associated osteomyelitis (IAO) due to Staphylococcus aureus or in chronic osteomyelitis lesions in slaughter pigs. Surprisingly, we found that the expression of RANKL was reduced during chronic bone infections. This is in line with the few studies conducted on human samples. A significant bone loss was observed in IAO lesions and in lesions from slaughter pigs, but with no indication of osteoclast involvement using histochemistry, immunohistochemistry for RANKL, receptor activator of nuclear factor kappa-B, osteoprotegerin and cathepsin K, and high-throughput quantitative real-time polymerase chain reaction on bone tissue from osteomyelitic lesions. A strong inflammatory response was seen in the infected animals and, therefore, we propose proteolytic enzymes induced by inflammation to be a major component of the bone loss. Furthermore, we found a significant upregulation of the IL26 gene in infected animals, which can inhibit RANKL-induced osteoclastogenesis, but has no homologue in mice. This finding emphasises that neither murine models nor in-vitro studies can mirror human disease development completely. The present study emphasises that the interactions between microorganisms, the immune system and bone cells in osteomyelitis are too complex to be accurately represented by an in-vitro model.

Desires for beverages and liking of skin care product odors in imaginative and immersive virtual reality beach contexts.

Immersive technologies simulating real-life environments allow repeated simulation of complex real-life situations at one location and, therefore, bear potential for consumer product evaluations and food behavior studies. This study aimed to assess whether a contextual exposure by immersive virtual reality (VR) or a photo-enhanced imaginative (PIC) condition could induce effects on desires and liking aligning with the situation. A total of 60 subjects (30 females) were presented to VR and PIC beach settings in a randomized balanced design performed in a laboratory one week apart. A follow-up test in the same room but without the contextual exposure was also performed (Neutral). Subjects rated their desires for hot and cold beverages during and just after the contextual exposure. Liking for skin care samples (sun screen and regular lotion) were assessed during exposure. The retention of the contextual exposure was further explored in a food choice behavior test. A similar test setup was followed in the Neutral condition. Engagement by presence dimensions (e.g., felt like being on a beach) and level of excitement was scored following the contextual exposure. The VR condition induced a significantly stronger engagement, compared with the PIC condition. During each beach exposure a significantly stronger desire for cold beverages compared to hot beverages was induced. The desire for cold versus hot beverages was, however, significantly larger in the VR simulation compared to the PIC context. Results from the Neutral condition provided no significant difference in the desires for hot versus cold beverages. A significant shift in desires was observed after the completion of the contextual beach exposures, as desire scores became aligned with scores assessed in the Neutral condition. The shift in desires was, however, more pronounced after VR. Variations in the contextual settings had little effect on odor liking and no retention effect on choice behavior was observed. Immersive VR induced a positive sensation of presence less dependent on the level of excitement and had a stronger effect on beverage-related desires. The VR technology seems to be a promising tool for evaluating contextual influences in food consumer research.

Comparison of innate and Th1-type host immune responses in Oesophagostomum dentatum and Trichuris suis infections in pigs.

This study investigated the details of the innate and Th1/Treg-type-associated host immune responses in Trichuris suis and Oesophagostomum dentatum mono- and co-infected pigs and in vitro in stimulated porcine dendritic cell cultures. Forty-eight pigs were allocated into a 2-factorial design with two groups trickle-inoculated with 10 T. suis eggs/kg/day (Group T) or 20 O. dentatum L3/kg/day (O). Another group (OT) was infected with both parasites. Group C remained uninfected. Expression of innate and Th1/Treg-cell-associated genes in gut mucosa and associated lymph nodes was determined by qPCR at necropsy day 35 and 71. Gene expression showed suppressed/inhibited Th1 and Treg-type immune reactions, in accordance with previous findings of a predominant Th2-type immune response to both nematodes. The in vitro part examined the production of TNF-α in porcine dendritic cells (DC) exposed to T. suis and/or O. dentatum excretory/secretory (E/S) products. Further, binding capacity and structure of E/S products were characterized. Glycan and lectin-binding capacity were generally lower in O. dentatum E/S products compared to T. suis which may explain the earlier found weaker Th2 response to the former. Surprisingly, O. dentatum E/S products induced a significant (P < 0·0001) increase in TNF-α DC production, potentially indicating a new mode of helminth-host immune response interaction.

RAAS and stress markers in acute ischemic stroke: preliminary findings.

OBJECTIVES: Angiotensin II type 1 receptor blockade has neuroprotective effects in animal stroke models, but no effects in clinical stroke trials. We evaluated cerebral and peripheral changes in the renin angiotensin aldosterone system (RAAS) and stress responses in acute ischemic stroke patients. MATERIALS AND METHODS: Blood from a jugular and cubital vein was collected within 48 h of stroke onset, after 24 and 48 h, and renin, angiotensin I, angiotensin II, aldosterone, norepinephrine, epinephrine, and cortisol were measured. Post-stroke cubital vein samples were collected after 8 (4.7-10) months. RESULTS: The acute systolic blood pressure was significantly increased, 148 (141-168) vs 140 (130-147) mmHg post-stroke. Angiotensin I, renin and aldosterone levels were significantly lower, angiotensin II was unchanged, and ACE activity was higher in the acute phase compared to post-stroke. No differences in RAAS were detected between jugular and cubital plasma levels. Jugular venous plasma levels of epinephrine and cortisol were elevated in the acute phase compared to cubital levels (P < 0.05). CONCLUSION: Increased epinephrine and cortisol levels in the jugular vein blood may reflect a higher peripheral turnover. The observed changes in RAAS in the acute stroke phase are consistent with responses to increased blood pressure.

Extensive changes in innate immune gene expression in obese Göttingen minipigs do not lead to changes in concentrations of circulating cytokines and acute phase proteins.

The usefulness of Göttingen minipigs as models for obesity and obesity-related pathologies is well established. The low-grade inflammation associated with obesity involves a range of innate immune factors; however, to our knowledge, the impact of obesity on innate immune factor expression has not been studied in Göttingen minipigs. Therefore, we studied the expression of innate immune genes in liver and adipose tissues as well as serum concentrations of cytokines and acute phase proteins in obese vs. lean Göttingen minipigs. In the liver, of 35 investigated genes, the expression of nine was significantly different in obese pigs (three up-regulated, six down-regulated). Of 33 genes in adipose tissues, obesity was associated with changed expression of 12 genes in the visceral adipose tissue (VAT) (three up-regulated), 11 in the abdominal retroperitoneal adipose tissue (RPAT) (seven of these up-regulated) and eight in the subcutaneous adipose tissue (SAT) from the neck (five of which were up-regulated). Obesity-associated expression changes were observed for three genes in all adipose tissues, namely chemokine (C-C motif) ligand 3-like 1 (up-regulated), CD200 molecule (down-regulated) and interleukin 1 receptor antagonist (up-regulated) with interleukin 1 receptor antagonist being the most highly regulated gene in both VAT and RPAT. Looking at patterns of expression across the three types of adipose tissues, obesity was associated with an increased number of acute phase proteins differentially expressed between adipose tissues and a decreased tissue-specific expression of cytokines and chemokines. In contrast to obese humans, no changes in serum concentrations of haptoglobin, C-reactive protein, serum amyloid A, tumor necrosis factor-α and interleukin 6 were found in obese Göttingen minipigs.

Expression studies of six human obesity-related genes in seven tissues from divergent pig breeds.

Obesity has reached epidemic proportions globally and has become the cause of several major health risks worldwide. Presently, more than 100 loci have been related to obesity and metabolic traits in humans by genome-wide association studies. The complex genetic architecture behind obesity has triggered a need for the development of better animal models than rodents. The pig has emerged as a very promising biomedical model to study human obesity traits. In this study, we have characterized the expression patterns of six obesity-related genes, leptin (LEP), leptin receptor (LEPR), melanocortin 4 receptor (MC4R), fat mass and obesity associated (FTO), neuronal growth regulator 1 (NEGR)1 and adiponectin (ADIPOQ), in seven obesity-relevant tissues (liver; muscle; pancreas; hypothalamus; and retroperitoneal, subcutaneous and mesenteric adipose tissues) in two pig breeds (production pigs and Göttingen minipigs) that deviate phenotypically and genetically from each other with respect to obesity traits. We observe significant differential expression for LEP, LEPR and ADIPOQ in muscle and in all three adipose tissues. Interestingly, in pancreas, LEP expression is only detected in the fat minipigs. FTO shows significant differential expression in all tissues analyzed, and NEGR1 shows significant differential expression in muscle, pancreas, hypothalamus and subcutaneous adipose tissue. The MC4R transcript can be detected only in hypothalamus. In general, the expression profiles of the investigated genes are in accordance with those observed in human studies. Our study shows that both the differences between the investigated breeds and the phenotypic state with respect to obesity/leanness play a large role for differential expression of the obesity-related genes.

Ampicillin-improved glucose tolerance in diet-induced obese C57BL/6NTac mice is age dependent.

Ampicillin has been shown to improve glucose tolerance in mice. We hypothesized that this effect is present only if treatment is initiated prior to weaning and that it disappears when treatment is terminated. High-fat fed C57BL/6NTac mice were divided into groups that received Ampicillin at different ages or not at all. We found that both diet and Ampicillin significantly changed the gut microbiota composition in the animals. Furthermore, there was a significant improvement in glucose tolerance in Ampicillin-treated, five-week-old mice compared to nontreated mice in the control group. At study termination, expressions of mRNA coding for tumor necrosis factor, serum amyloid A, and lactase were upregulated, while the expression of tumor necrosis factor (ligand) superfamily member 15 was downregulated in the ileum of Ampicillin-treated mice. Higher dendritic cell percentages were found systemically in high-fat diet mice, and a lower tolerogenic dendritic cell percentage was found both in relation to high-fat diet and late Ampicillin treatment. The results support our hypothesis that a "window" exists early in life in which an alteration of the gut microbiota affects glucose tolerance as well as development of gut immunity and that this window may disappear after weaning.

Disseminated intravascular coagulation in a novel porcine model of severe Staphylococcus aureus sepsis fulfills human clinical criteria.

Sepsis is a common and often fatal complication in human patients in intensive care units. Relevant and well characterized animal models of sepsis may provide valuable information on pathophysiological mechanisms and be a mean of testing new therapeutic strategies. Large animal models of Staphylococcus aureus sepsis are rare, even though S. aureus increasingly affects human patients. Sepsis changes the haemostatic balance and leads to endothelial cell (EC) activation, coagulopathy and, in severe cases, disseminated intravascular coagulation (DIC). The aim of this study was to characterize the haemostatic and vascular alterations in a novel porcine model of severe S. aureus sepsis, investigating whether the changes fulfill the human clinical criteria for DIC. Five pigs were inoculated intravenously with S. aureus and two control animals were sham-inoculated. Blood samples were collected for thromboelastography (TEG) and assessment of plasma-based haemostatic parameters. Tissue was collected for histopathology and reverse transcriptase quantitative real-time polymerase chain reaction for measurement of mRNA encoding EC markers. All infected animals developed DIC; including procoagulant activation represented by hypercoagulable TEG profiles and prolonged clotting time. Histologically, numerous pulmonary thrombi were present in one pig. Inhibitor consumption was represented by decreasing antithrombin levels in infected pigs. Hyaline globules were found in three infected pigs, confirming fibrinolytic activation. EC activation was identified by expression of von Willebrand factor in small vessels together with elevated mRNA encoding activated EC markers. Severe haemostatic and vascular changes fulfilling the human criteria for DIC were therefore seen in all infected pigs. A tendency towards uncompensated DIC was seen in two animals.

The impact of Staphylococcus aureus concentration on the development of pulmonary lesions and cytokine expression after intravenous inoculation of pigs.

Acute respiratory distress syndrome is a common complication in severe sepsis. In pigs, the lungs play an important role in clearing systemic bacterial infections due to pulmonary intravascular macrophages found specifically in pigs. However, this increases the exposure of the porcine lungs to pathogens and potential injury. The authors propose that increasing the concentration of the inoculum without changing the bacterial dose will lead to severe sepsis with pronounced pulmonary lesions. This could potentially create a risk of cytokine spillover to the circulation, leading to an increased systemic response. Eight Danish Landrace pigs, approximately 10 weeks old, were inoculated twice with a low or once with a high concentration of Staphylococcus aureus. Three pigs were sham-inoculated. The animals were grouped based on macro- and microscopic lung lesions. The mRNA expression of local pulmonary inflammatory markers was compared to protein levels of systemic inflammatory markers. The most severe pulmonary lesions were observed in animals receiving the high S. aureus concentration, indicating that severity of lesions is dependent on inoculum concentration rather than total numbers of bacteria. Furthermore, local mRNA expression of inflammatory cytokines appeared to be dependent on the magnitude and severity of tissue destruction, including the ability to confine the lesions. Increasing mRNA levels of serum amyloid A could be a confident marker of severity of pulmonary lesions. Since no correlation was observed between local and systemic levels of inflammatory cytokines, this finding could indicate an ability of the porcine lung to compartmentalize the local inflammatory response and thus restrict systemic contribution.

Molecular characterization of the porcine surfactant, pulmonary-associated protein C gene.

The surfactant, pulmonary-associated protein C (SFTPC) is a peptide secreted by the alveolar type II pneumocytes of the lung. We have characterized the porcine SFTPC gene at genomic, transcriptional, and protein levels. The porcine SFTPC is a single-copy gene on pig chromosome 14. Two transcripts were found in a newborn pig lung cDNA library: a full-length clone and a clone missing exon 5. cDNA sequence comparison revealed four synonymous and two nonsynonymous substitutions and in-frame insertions at the beginning of exon 5. Comparison of the SFTPC coding region between several mammals showed high levels of conservation. Northern blot studies showed lung-specific expression of the full-length SFTPC transcript, appearing in 50-day-old fetus and increasing during lung development. Both SFTPC transcripts were detected mainly in lung by real-time RT-PCR and they were significantly down-regulated in necrotic lungs of pigs infected with Actinobacillus pleuropneumoniae. Additionally, the protein levels were also decreased or absent in the necrotic tissue.

Evolution of Fusarium oxysporum f. sp. vasinfectum Races Inferred from Multigene Genealogies.

ABSTRACT Fusarium wilt of cotton is a serious fungal disease responsible for significant yield losses throughout the world. Evolution of the causal organism Fusarium oxysporum f. sp. vasinfectum, including the eight races described for this specialized form, was studied using multigene genealogies. Partial sequences of translation elongation factor (EF-1alpha), nitrate reductase (NIR), phosphate permase (PHO), and the mitochondrial small subunit (mtSSU) rDNA were sequenced in 28 isolates of F. oxysporum f. sp. vasinfectum selected to represent the global genetic diversity of this forma specialis. Results of a Wilcoxon Signed-Ranks Templeton test indicated that sequences of the four genes could be combined. In addition, using combined data from EF-1alpha and mtSSU rDNA, the phylogenetic origin of F. oxysporum f. sp. vasinfectum within the F. oxysporum complex was evaluated by the Kishino-Hasegawa likelihood test. Results of this test indicated the eight races of F. oxysporum f. sp. vasinfectum appeared to be nonmonophyletic, having at least two independent, or polyphyletic, evolutionary origins. Races 3 and 5 formed a strongly supported clade separate from the other six races. The combined EF-1alpha, NIR, PHO, and mtSSU rDNA sequence data from the 28 isolates of F. oxysporum f. sp. vasinfectum recovered four lineages that correlated with differences in virulence and geographic origin: lineage I contained race 3, mostly from Egypt, and race 5 from Sudan; lineage II contained races 1, 2, and 6 from North and South America and Africa; lineage III contained race 8 from China; and lineage IV contained isolates of races 4 and 7 from India and China, respectively.