Untreated Pigs Research Articles

Risk Factors for Salmonella Infection in Fattening Pigs – An Evaluation of Blood and Meat Juice Samples

The main objective of this study was to analyse potential herd-level factors associated with the detection of Salmonella antibodies in fattening pigs. Two independent datasets, consisting of blood and meat juice samples respectively, were used. Additional information about husbandry, management and hygiene conditions was collected by questionnaire for both datasets. The serological analysis showed that 13.8% of the blood samples and 15.7% of the meat juice samples had to be classified as Salmonella-positive. Logistic-regression models were used to assess statistically significant risk factors associated with a positive sample result. The results of the statistical blood sample analysis showed that the application of antibiotics increased the odds ratio (OR) by a factor of 5.21 (P < 0.001) compared to untreated pigs. A fully slatted floor decreased the prevalence of Salmonella as well as the use of protective clothing or the cleaning of the feed tube (ORs 0.35-0.54, P < 0.001). It was shown that a distance of less than 2 km to other swine herds increased the chance of a positive Salmonella result (OR = 3.76, P < 0.001). The statistical analysis of the meat juice samples revealed the importance of feed aspects. The chance of obtaining a positive meat juice sample increased by a factor of 3.52 (P < 0.001) by using granulated feed instead of flour. It also became clear that liquid feeding should be preferred to dry feeding (OR = 0.33, P < 0.001). A comparison of the blood sample analysis to the meat juice model revealed that the latter was less powerful because data structure was less detailed. The expansion of data acquisition might solve these problems and improve the suitability of QS monitoring data for risk factor analyses.

Inhaled 1,8-Cineole Reduces Inflammatory Parameters in Airways of Ovalbumin-Challenged Guinea Pigs

Eucalyptol, also known as 1,8-cineole, is a monoterpene traditionally used to treat respiratory disorders due to its secretolytic properties. In addition to its myorelaxant effects, it also has anti-inflammatory actions in vitro. In this study, we aimed to evaluate the efficacy of acute treatment with 1,8-cineole on reducing airway inflammatory parameters. Ovalbumin (OVA)-sensitized guinea pigs were submitted to antigenic challenge (OVA) with or without pre-treatment with a single dose of 1,8-cineole administered by inhalation. Airway inflammatory parameters were reduced or absent in 1,8-cineole-treated animals as compared with untreated guinea pigs. Acute treatment with 1,8-cineole impaired the development of airway hyperresponsiveness to carbachol in isolated tracheal rings. Levels of the pro-inflammatory cytokines TNFα and IL-1β was lower in bronchoalveolar lavage fluid (BALF) of 1,8-cineol-treated guinea pigs than in untreated animals. 1,8-Cineole impaired the OVA-induced increase of the myeloperoxidase activity in BALF. 1,8-Cineole also prevented the reduction of the mucociliary clearance induced by the antigen presentation. The present investigation provides evidence that inhaled 1,8-cineole prevents hyperresponsiveness and inhibits inflammation in airways of ovalbumin-challenged guinea pigs.

Comparative evaluation of PRRS virus infection in vaccinated and naïve pigs

The purpose of this study was to evaluate the time-course of the immune response to a field Porcine Respiratory and Reproductive Syndrome virus (PRRSV) strain in PRRS-naïve, untreated pigs, as well as in four groups of age and breed-matched pigs injected with a live attenuated PRRS vaccine, its adjuvant, an inactivated PRRS vaccine and an irrelevant, inactivated Porcine Circovirus type 2 (PCV2) vaccine, respectively. PRRSV infection was confirmed in all groups by PCR and antibody assays. The antibody response measured by ELISA took place earlier in pigs injected with the live attenuated vaccine, which also developed a much stronger serum-neutralizing antibody response to the vaccine strain. Yet, no clear protection was evidenced in terms of viremia against the field virus strain, which showed 11.1% nucleotide divergence in ORF7 from the vaccine strain. In vitro, the interferon (IFN)-γ response to PRRSV was almost absent on PVD 60 in all groups under study, whereas the prevalence of interleukin (IL)-10 responses to PRRSV was fairly high in PCV2-vaccinated animals, only. Results indicate that distinct patterns of immune response to a field PRRSV strain can be recognized in PRRS-vaccinated and naïve pigs, which probably underlies fundamental differences in the development and differentiation of PRRSV-specific immune effector cells.

THERAPEUTIC INJECTION OF PARP INHIBITOR INO-1001 PRESERVES CARDIAC FUNCTION IN PORCINE MYOCARDIAL ISCHEMIA AND REPERFUSION WITHOUT REDUCING INFARCT SIZE

Pharmacological protection from myocardial reperfusion injury, despite plenty of approaches, has still not been realized in humans. We studied the putative infarct size (IS)-sparing capacity of poly(ADP-ribose)polymerase inhibitor, INO-1001, and focused on cardiac functional recovery during reperfusion. Male farm-bred Landrace pigs were subjected to 1-h left anterior descending coronary artery occlusion followed by 3 h of reperfusion (control). Infarct size was determined by triphenyltetrazolium chloride/Evans blue staining. Plasma markers of myocardial injury (troponin T, creatine kinase, lactate dehydrogenase) were determined upon protocol completion. Cardiac function was continuously assessed via pulmonary and femoral artery catheters. INO-1001 (1 mg/kg) was administered upon reperfusion in the treatment group. As a positive control, untreated pigs were subjected to ischemic preconditioning (10-min left anterior descending coronary artery occlusion followed by 15-min reperfusion before the intervention). Ischemic preconditioning reduced myocardial damage reflected by a smaller IS and lower plasma markers of myocardial injury. INO-1001 did not reduce IS but significantly improved functional recovery (increased stroke volume, cardiac index, and mixed venous oxygen saturation) during reperfusion compared with vehicle-treated control and ischemic preconditioning. Although we could not confirm the IS-sparing capacities of poly(ADP-ribose)polymerase inhibitor, INO-1001, the drug holds the potential of hemodynamic improvement during reperfusion.

Endothelial Progenitor Cells Homing and Renal Repair in Experimental Renovascular Disease

Tissue injury triggers reparative processes that often involve endothelial progenitor cells (EPCs) recruitment. We hypothesized that atherosclerotic renal artery stenosis (ARAS) activates homing signals that would be detectable in both the kidney and EPCs, and attenuated on renal repair using selective cell-based therapy. Pigs were treated with intrarenal autologous EPC after 6 weeks of ARAS. Four weeks later, expression of homing-related signals in EPC and kidney, single kidney function, microvascular (MV) density, and morphology were compared with untreated ARAS and normal control pigs (n = 7 each). Compared with normal EPC, EPC from ARAS pigs showed increased stromal cell-derived factor (SDF)-1, angiopoietin-1, Tie-2, and c-kit expression, but downregulation of erythropoietin (EPO) and its receptor. The ARAS kidney released the c-kit-ligand stem cell factor, uric acid, and EPO, and upregulated integrin beta2, suggesting activation of corresponding homing signaling. However, angiopoietin-1 and SDF-1/CXCR4 were not elevated. Administration of EPC into the stenotic kidney restored angiogenic activity, improved MV density, renal hemodynamics and function, decreased fibrosis and oxidative stress, and attenuated endogenous injury signals. The ARAS kidney releases specific homing signals corresponding to cognate receptors expressed by EPC. EPC show plasticity for organ-specific recruitment strategies, which are upregulated in early atherosclerosis. EPC are renoprotective as they attenuated renal dysfunction and damage in chronic ARAS, and consequently decreased the injury signals. Importantly, manipulation of homing signals may potentially allow therapeutic opportunities to increase endogenous EPC recruitment.

Pilot studies of pressure-immobilization bandages for rattlesnake envenomations

Study objective. Pressure-immobilization bandages sequester venom in extremities and are recommended for snakebites without local toxicity. Pilot studies were performed to determine the time of onset of toxicity and efficacy of pressure-immobilizations bandages in a porcine model of rattlesnake envenomation. Methods. After IACUC approval, anesthetized pigs were injected subcutaneously in a distal hind leg with 200 mg of Crotalus atrox venom. After 1 min, pigs received either a pressure-immobilization bandage (N = 3) or no treatment (N = 3). At 24 h, surviving pigs received antivenin and then the pressure-immobilization bandages were removed. Surviving subjects were followed for 1 week. Chi-square analysis and paired t-test were used. Results. Pigs with pressure-immobilization bandages survived for 24 h, whereas untreated pigs died at 13.68 ± 3.42 h (p = 0.014). Surviving pigs walked on the extremity at 7 days. Potassium rose from 4.033 ± 0.252 at baseline to 17.767 ± 5.218 mEq/L (p < 0.0001) at time of death in untreated pigs but was normal at 24 h in treated subjects. Widespread tissue necrosis was seen in the untreated group but only local necrosis in the treatment group. Conclusions. Pressure-immobilization bandages prevented death from severe C. atrox envenomations with a 24 h delay to treatment. Surviving pigs had recovery of limb use at 1 week.

Desflurane induces airway contraction mainly by activating transient receptor potential A1 of sensory C-fibers

We previously reported that desflurane induced airway contraction via antidromic tachykinin release from sensory C-fibers. Here, we investigated the effect of desflurane on airway lung resistance (R(L)) using specific receptor antagonists in C-fibers. Young guinea pigs were anesthetized and their tracheas were cannulated with an endotracheal tube via a tracheotomy. A Fleisch pneumotachograph and a differential transducer were used to monitor respiratory flow rate, intrapleural pressure, and airway pressure, and R(L) was calculated and recorded. A transient receptor potential A1 (TRPA1) or a transient receptor potential V1 (TRPV1) selective antagonist of sensory C-fibers, i.e., HC030031 or BCTC, was administered before the exposure to desflurane. In an additional experiment, tachykinin receptor of airway smooth muscles was antagonized only by the neurokinin-2 receptor antagonist MEN-10376 before the exposure to desflurane. HC030031 completely inhibited both the first and the second contractile responses induced by desflurane, whereas BCTC had little effect. MEN-10376 also significantly and substantially diminished the contractile response. Desflurane contracts the airway in untreated guinea pigs mainly by activating irritant gas receptor TRPA1 of afferent C-fibers, resulting in the release of contractile tachykinins such as neurokinin A.

Magnetic resonance imaging reveals that galantamine prevents structural brain damage induced by an acute exposure of guinea pigs to soman

Galantamine, a drug used to treat Alzheimer's disease, has recently emerged as a potential medical countermeasure against the toxicity of organophosphorus (OP) compounds, including the nerve agent soman. Here, magnetic resonance imaging (MRI) was used to characterize the neurotoxic effects of soman and the ability of galantamine to prevent these effects in guinea pigs, the best non-primate model to predict the effectiveness of antidotes against OP toxicity in humans. The brains of treated and untreated guinea pigs were imaged using a clinical 3.0 Tesla MRI scanner at 48 h before and 6–7 h, 48 h and 7 days after their challenge with 1.0xLD50 soman (26.6 μg/kg, sc). Significant brain atrophy was observed among all untreated animals at 7 days after their challenge with soman. In mildly intoxicated animals, significant shortening of spin–spin relaxation times (T2) was observed in the thalamus and amygdala at 7 h after the challenge. In severely intoxicated animals, T2 values and T2-weighted signal intensities increased significantly in the piriform cortex, hippocampus, thalamus and amygdala; in most regions, changes were long-lasting. Voxel-based morphometric analysis of the images revealed that other brain regions were also damaged in these animals. Neuronal loss was confirmed histopathologically. In animals that were treated with galantamine (8 mg/kg, im) 30 min prior to the exposure to soman, T2, T2-weighted signal intensities, and CSF volumes were largely unaffected. It is, therefore, concluded that galantamine can effectively prevent the structural brain damage induced by an acute exposure to soman.

Use of tylvalosin-medicated feed to control porcine proliferative enteropathy

The effect of an oral treatment with the tartrate salt of tylvalosin on the development of proliferative enteropathy in 60 experimentally challenged pigs was studied. Thirty of the pigs were...

Alteration of the Ileal Microbiota of Weanling Piglets by the Growth-Promoting Antibiotic Chlortetracycline

Antibiotics such as chlortetracycline (CTC) have been used to promote growth of pigs for decades, but concerns over increased antibiotic-resistant infections in humans have prompted the development of alternative strategies. Developing alternatives to antibiotic growth promoters (AGPs) could be informed by information on the mechanisms of growth promotion, notably, how AGPs affect the microbial populations of the gastrointestinal tract. Pigs from three sows were aseptically delivered by cesarean section. Six piglets were distributed to each of two foster mothers until weaning, when piglets were fed a diet with or without 50 mg/kg CTC for 2 weeks. The ileal bacterial microbiota was characterized by using a cultivation-independent approach based on DNA extraction, PCR amplification, cloning, and sequencing of the 16S rRNA gene pool. The ileal and mucosal communities of these growing pigs were dominated by Lactobacillus bacteria, various members of the family Clostridiaceae, and members of the poorly known genus Turicibacter. Overall, CTC treatment resulted in three shifts: a decrease in Lactobacillus johnsonii, an increase in L. amylovorus, and a decrease in Turicibacter phylotypes. The composition of the microbiota varied considerably between individual pigs, as revealed by shared operational taxonomic units (OTUs) and similarity (SONS) analysis (theta(YC) values). While the observed variation between untreated pigs obscured the possible effect of CTC, integral-LIBSHUFF and SONS analyses of pooled libraries indicated a significant shift due to CTC in both the lumen and the mucosa, with some OTUs unique to either treated or control ileum. DOTUR analysis revealed little overlap between control and treated communities at the 3% difference level, indicating unique ileal communities in the presence of CTC.

The reduction of CSFV transmission to untreated pigs by the pestivirus inhibitor BPIP: A proof of concept

5-[(4-Bromophenyl)methyl]-2-phenyl-5H-imidazo[4,5-c]pyridine (BPIP) is a representative molecule of a novel class of highly active in vitro inhibitors of the replication of Classical swine fever virus (CSFV). We recently demonstrated in a proof of concept study that the molecule has a marked effect on viral replication in CSFV-infected pigs. Here, the effect of antiviral treatment on virus transmission to untreated sentinel pigs was studied. Therefore, BPIP-treated pigs (n=4), intra-muscularly infected with CSFV, were placed into contact with untreated sentinel pigs (n=4). Efficient transmission of CSFV from four untreated seeder pigs to four untreated sentinels was observed. In contrast, only two out of four sentinel animals in contact with BPIP-treated seeder animals developed a short transient infection, of which one was likely the result of sentinel to sentinel transmission. A significant lower viral genome load was measured in tonsils of sentinels in contact with BPIP-treated seeder animals compared to the positive control group (p=0.015). Although no significant difference (p=0.126) in the time of onset of viraemia could be detected between the groups of contact animals, a tendency towards the reduction of virus transmission was observed. Since sentinel animals were left untreated in this exploratory trial, the study can be regarded as a worst case scenario and gives therefore an underestimation of the potential efficacy of the activity of BPIP on virus transmission.

Effect of Immunostimulation by Detoxified E. coli Lipopolysaccharide Combined with Inactivated Propionibacterium granulosum Cells on Porcine Immunity

The aim of this experiment was to evaluate the immunomodulating activities of inactivated Propionibacterium granulosum cell walls and E. coli lipopolysaccharide (PG/LPS) on porcine immunity. Piglets were intramuscularly administered PG/LPS (1 ml/10 kg body weight) once or twice. The function of natural killer cells, lymphocytes and neutrophils and the adjuvant effect on antibody induction by attenuated classical swine fever virus (CSFV) and inactivated Mycoplasma hyopneumoniae vaccination were evaluated. The results showed that the cytotoxicity of natural killer cells and proliferation of lymphocytes in response to mitogen stimulation were significantly enhanced (P<0.05) in those pigs receiving PG/LPS injection compared with the controls. However, there was no significant effect on the phagocytic activity of neutrophils (P>0.05). PG/LPS also displayed adjuvant effects with CSFV and Mycoplasma hyopneumoniae vaccines. Moreover, pigs receiving two injections of PG/LPS showed a 20.8% growth enhancement compared with untreated pigs. Thus, PG/LPS caused positive immunoregulation of porcine innate immune system effectors, non-specific activation of lymphocytes and antibody production.

Determination of the efficacy of terbinafine hydrochloride nail solution in the topical treatment of dermatophytosis in a guinea pig model

Currently available topical antifungals are often not satisfactory for the treatment of nail infections, because of the inability to penetrate the nail plate. Terbinafine HCl nail solution is a novel antifungal formulation containing a nail penetration enhancer dodecyl-2-N,N-dimethylaminopropionate hydrochloride (DDAIP HCl, trade name NexACT-88). In this study, we used a guinea pig model of Trichophyton mentagrophytes dermatophytosis and evaluated the clinical and mycological efficacy of different terbinafine HCl nail solutions (TNS) formulated with or without DDAIP HCl. Ciclopirox (8%) nail lacquer (Penlac), the only Food and Drug Administration approved topical treatment for onychomycosis, was used as a comparator. Following the IACUC Guidelines, the skin of male albino guinea pigs was abraded under anaesthesia. Each animal was infected with T. mentagrophytes ATCC 24953 (cell suspension containing 1 x 10(7) conidia). The experimental animals were divided into 11 groups (five animals per group) and tested with the following formulations: vehicle control, 0.5% DDAIP HCl, 1%, 5% and 10% TNS (without DDAIP HCl), 1% TNS with 0.5%, 2.5% and 5.0% DDAIP HCl, 5% and 10% TNS with 0.5% DDAIP HCl, 8% ciclopirox nail lacquer and an untreated control group. Evaluation of clinical and mycological efficacy was performed 72 h after completion of a 7-day treatment regimen. Skin biopsy samples were processed for histopathological examination. The infected untreated control guinea pigs showed patches of hair loss and ulcerated or scaly skin and fungal invasion of hair roots. The vehicle and 0.5% DDAIP HCl treated groups showed minimal clinical efficacy (only 11% and 5%, respectively). In contrast, all three concentrations of TNS (1%, 5% and 10% terbinafine HCl) formulated with or without 0.5% DDAIP HCl showed 100% mycological efficacy by the hair root invasion test. Clinical efficacy of the 5% and 10% TNS improved with addition of 0.5% DDAIP HCl (47.4% and 73.8% vs. 68.4% and 89.5%, respectively). In addition, no fungal elements were detected in the treated guinea pig skin. All formulations of TNS resulted in a higher clinical and mycological efficacy compared with the 8% ciclopirox nail lacquer (P = 0.0444). In conclusion, TNS containing 1%, 5% and 10% terbinafine HCl formulated with and without DDAIP HCl demonstrated high antifungal efficacy in experimental dermatophytosis. Addition of 0.5% DDAIP HCl to 5% and 10% TNS significantly enhanced the clinical and mycological efficacy of these formulations which were superior compared with the 8% ciclopirox nail lacquer. Evaluation of the 1%, 5% and 10% TNS in clinical trials for the treatment of dermatophytosis and onychomycosis is warranted.

Using Platelet Proteomics as a Marker of Cardiac Failure in a Pacing-Induced Pig Heart Failure Model

Cardiac failure is a leading cause of morbidity and mortality in the United States today. There is also a well-described gender bias to this disease, with the occurrence of cardiac failure higher in men than women. Anti-platelet drugs are the mainstay for prevention and therapy of cardiovascular disease in humans, and we have therefore conjectured that blood platelets could function as essential biomarkers of these processes. Thus, by studying the protein expression of platelets using proteomic tools, we anticipated that a proteomic profile of the “cardiac platelet” could be defined that would permit mechanistic discrimination of the development of this process. For this study, we hypothesized that signal transduction pathways, specifically including inflammatory or pro-coagulatory pathways, are key mediators of platelet contributions to the cardiac failure process. To test this hypothesis, ventricular dysfunction was induced in male and female swine by the placement of an atrial active-fixation pacing lead and active pacing initiated at 200 bpm. Ventricular performance was defined by Fractional Shortening (Fractional Shortening = [(left ventricular end-diastolic diameter - left ventricular end-systolic diameter)]/left ventricular end-diastolic diameter). When the fractional shortening (normal 32%) dropped below 16%, the animals were considered to be in terminal cardiac failure, and were euthanized. Blood samples were collected throughout the experiment and platelets isolated for analysis. Proteomic analysis of platelet lysates was performed using antibody microarrays (Clontech 507 AB arrays; n = 2 males and 2 females, replicates = 2), reverse capture microarrays (replicates = 6) and Western blotting (n ≥2). Platelets samples were compared as pairs of platelet lysates from initial and terminal bleeds. Samples of ventricular muscle tissue collected following termination of the experiments were run in parallel studies and were compared against gender matched untreated pigs. Results of the analysis demonstrated that the expression of many proteins was altered both in platelets and heart tissue in a gender-dependent manner following cardiac stress. The top 5 proteins up-regulated in platelets isolated from males were p57, integrin b1, GCIP, K channel a and NF-ATc1. Three of these proteins, p57, GCIP and NF-ATc1, are associated with cell proliferation and may reflect increased megakaryocyte production of platelets. In a recent study with Sanqi (Yao et al., Phytomedicine 2008), a traditional Chinese medicine with cardiovascular effects, k channel subfamily V was also shown to be increased in platelets. In contrast, in female platelets, the top 5 proteins were GAP-43 and IL1beta which are proteins associated with inflammation, beta-catenin which is a protein with wide-spread effects mediated through the cytoskeleton, STAT3 and COMT (catechol-O-methyl transferase). None of these proteins were in the top 5 up-regulated proteins in heart tissue. The top 5 down-regulated proteins in the platelets were, for the males: GIT1, Bog, IL1beta, TIAR and Ku70 and for the females: SCAR-1, crp-2, NTF, SMN and MGMT. GIT1 is involved in cytoskeletal changes and is normally phosphorylated by src kinase following stimulation by platelet activation. TIAR, an mRNA-binding protein, regulates expression of diverse proteins such as MYC, IL-6 and Fas. It has been implicated in cellular processes ranging from proliferation to apoptosis. In the female samples, SCAR-1 interacts with GPVI to form the bridge to collagen binding. Down regulation of this protein may inhibit platelet binding to extracellular substrates. The proteins SMN and MGMT in the hematopoietic system are commonly associated with stem cell differentiation and may also relate to levels or rates of differentiation of megakaryocytes into platelets. Confirmatory experiments with reverse capture methodology supported our findings for Bog, GIT, catenin, SCAR-1 and crp2 as well as caspase 6 and TRAF2, proteins that were not in the top 5 group. Western blots confirmed changes in caspases 6 and PKCa. In conclusion, our preliminary data support the hypothesis that that signal transduction pathways in platelets may be key mediators of platelet contributions to cardiac failure. Network analysis of the many proteins altered during this process will be necessary to understand the varied and subtle changes that may be occurring during this process.

Simvastatin abates development of renal fibrosis in experimental renovascular disease

Epithelial-to-mesenchymal transition contributes to renal fibrogenesis, which is regulated by profibrogenic and antifibrogenic mediators. 3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors can prevent epithelial-to-mesenchymal transition in some models. Therefore, we tested the hypothesis that epithelial-to-mesenchymal transition participates in renal injury in porcine atherosclerotic renovascular disease and can be attenuated by simvastatin. Renal hemodynamics, function, and endothelial function were quantified in vivo in pigs after 12 weeks of combined hypercholesterolemia + renal artery stenosis without (n = 8) or with oral simvastatin supplementation (1.2 mg/kg, n = 6), and in controls (n = 8). Ex-vivo studies assessed renal immunoreactivity to fibrogenic factors and renal histology. Blood pressure, cholesterol levels, and basal renal function were similar in treated and untreated pigs with hypercholesterolemia + renal artery stenosis. Hypercholesterolemia + renal artery stenosis significantly upregulated renal transforming growth factor-beta signaling and elicited epithelial-to-mesenchymal transition, accompanied by glomerulosclerosis and renal fibrosis. Simvastatin did not affect smad 2/3 expression but upregulated expression of hepatocyte growth factor, bone morphogenetic factor-7, and smad 7 and prevented most of these renal structural and functional alterations. Furthermore, simvastatin improved renal blood flow response to endothelium-dependent challenge (+111.3 +/- 35.5 vs. -30.4 +/- 18.7 ml/min in untreated pigs, P < 0.05). Simvastatin upregulates inhibitors of transforming growth factor-beta signaling, attenuates epithelial-to-mesenchymal transition, and decreases renal fibrosis in hypercholesterolemia + renal artery stenosis. These lipid-lowering-independent effects result in improvement of renal function, suggesting clinically valuable potential for statins in preserving the stenotic kidney and limiting deterioration of renal function in atherosclerotic renovascular disease.

Metabolic response to an acute jump in cardiac workload: effects on malonyl-CoA, mechanical efficiency, and fatty acid oxidation

Inhibition of myocardial fatty acid oxidation can improve left ventricular (LV) mechanical efficiency by increasing LV power for a given rate of myocardial energy expenditure. This phenomenon has not been assessed at high workloads in nonischemic myocardium; therefore, we subjected in vivo pig hearts to a high workload for 5 min and assessed whether blocking mitochondrial fatty acid oxidation with the carnitine palmitoyltransferase-I inhibitor oxfenicine would improve LV mechanical efficiency. In addition, the cardiac content of malonyl-CoA (an endogenous inhibitor of carnitine palmitoyltransferase-I) and activity of acetyl-CoA carboxylase (which synthesizes malonyl-CoA) were assessed. Increased workload was induced by aortic constriction and dobutamine infusion, and LV efficiency was calculated from the LV pressure-volume loop and LV energy expenditure. In untreated pigs, the increase in LV power resulted in a 2.5-fold increase in fatty acid oxidation and cardiac malonyl-CoA content but did not affect the activation state of acetyl-CoA carboxylase. The activation state of the acetyl-CoA carboxylase inhibitory kinase AMP-activated protein kinase decreased by 40% with increased cardiac workload. Pretreatment with oxfenicine inhibited fatty acid oxidation by 75% and had no effect on cardiac energy expenditure but significantly increased LV power and LV efficiency (37 +/- 5% vs. 26 +/- 5%, P < 0.05) at high workload. In conclusion, 1) myocardial fatty acid oxidation increases with a short-term increase in cardiac workload, despite an increase in malonyl-CoA concentration, and 2) inhibition of fatty acid oxidation improves LV mechanical efficiency by increasing LV power without affecting cardiac energy expenditure.

Functional and bioenergetic modulations in the infarct border zone following autologous mesenchymal stem cell transplantation

Preclinical and clinical studies have demonstrated that stem cell transplantation can improve the left ventricular (LV) contractile performance, yet the underlying mechanisms remain unknown. We examined whether mesenchymal stem cell (MSC) transplantation-induced beneficial effects are secondary to paracrine-associated improvements in LV contractile performance, wall stress, and myocardial bioenergetics in hearts with postinfarction LV remodeling. Myocardial contractile function and bioenergetics were compared 4 wk after acute myocardial infarction in normal pigs (n = 6), untreated pigs with myocardial infarction (MI group; n = 6), and pigs receiving autologous MSC transplantation (MI + MSC group; n = 5). A distal occlusion of the left anterior descending coronary artery instigated significant myocardial hypertrophy. Ejection fraction decreased from 55.3 +/- 3.1% (normal) to 30.4 +/- 2.3% (MI group; P < 0.01) and to 45.4 +/- 3.1% (MI + MSC group; P < 0.01 vs. MI). Hearts in the MI group developed severe contractile dyskinesis in the infarct zone and border zone (BZ). MSC transplantation significantly improved contractile performance from dyskinesis to active contraction (P < 0.01 vs. MI). BZ systolic wall stress was severely increased in MI hearts but significantly improved after MSC transplantation (P < 0.01 vs. MI). The BZ demonstrated profound bioenergetic abnormalities in MI pigs; this was significantly improved after MSC transplantation (P < 0.01 vs. MI). Patchy spared myocytes were found in the infarct zone of hearts receiving MSC transplantation but not in control hearts. These data demonstrate that MSC transplantation into the BZ causes significant improvements in myocardial contractile performance and reduction in wall stress, which ultimately results in significant bioenergetic improvements. Low cell engraftment indicates that MSCs did not provide a structural contribution to the damaged heart and that the observed beneficial effects likely resulted from paracrine repair mechanisms.

The PPARα Activator Fenofibrate Slows Down the Progression of the Left Ventricular Dysfunction in Porcine Tachycardia-Induced Cardiomyopathy

It has been reported that high intramyocardial peroxisome proliferator-activated receptor alpha (PPARalpha) stimulation or overexpression altered cardiac contractile function in mouse models of cardiac hypertrophy and heart failure. Nevertheless, it has never been demonstrated that clinically relevant doses of drugs stimulating PPARalpha activity such as fenofibrate increase the risk to develop heart failure in humans. To determine if fenofibrate accelerates the development of heart failure in large mammals, we have tested its effects on the progression of left ventricular dysfunction in pacing-induced heart failure in pigs. Fenofibrate treatment blunted reduction in left ventricular ejection fraction, reduced cardiac hypertrophy, and attenuated clinical signs of heart failure. Fenofibrate impeded the increase in atrial natriuretic peptide, brain natriuretic peptide, and endothelin-1 plasma levels. The expression of PPARalpha, fatty acyl-CoA-oxidase, and carnitine palmitoyltransferase-Ibeta was reduced at mRNA levels in the left ventricle from untreated heart failure pigs but maintained near normal values with fenofibrate. Fenofibrate prevented heart failure-induced overexpression of TNFalpha mRNA and enhanced catalase activity in left ventricle compared to placebo. These data suggest that a clinically relevant dose of fenofibrate does not accelerate but slows down heart failure development in the model of pacing-induced heart failure in large mammals.

Survival and persistence of Lactobacillus plantarum 4.1 and Lactobacillus reuteri 3S7 in the gastrointestinal tract of pigs

Lactobacillus sp. are important inhabitants of the intestines of animals. They are also largely used as probiotics for both humans and animals. To exert beneficial effects, lactobacilli have to survive through the gastrointestinal transit. Based on bile-salt resistance, pH tolerance, antimicrobial activity and heat resistance, Lactobacillus plantarum 4.1 and Lactobacillus reuteri 3S7 were previously selected and used as probiotic additives in pelleted feeding trials. Both strains were fed to pigs (sows and piglets) at a cell number of ca. 10 10 viable cells per day. A polyphasic approach, comprising growth on selective media, Biolog System analysis, 16S rRNA gene sequencing and RAPD-PCR typing, was used to identify and differentiate L. plantarum 4.1 and L. reuteri 3S7 from other faecal Lactobacillus sp., L. plantarum 4.1 and L. reuteri 3S7 had the capacity to survive during the gastrointestinal transit and were found in the feaces at a cell number of 6–8 log cfu/g. Their persistence was shown after 6 days from the administration. Compared to untreated pigs, the administration of L. plantarum 4.1 and L. reuteri 3S7 significantly ( P < 0.05) decreased the population of Enterobacteriaceae. Besides, the β-glucuronidase activity of all pigs decreased of ca. 23.0% during administration. The findings of this study showed that L. plantarum 4.1 and L. reuteri 3S7 have the potential to be used as probiotic additives in pelleted feed for pigs.

Chito-oligosaccharides as an Alternative to Antimicrobials in Improving Performance, Digestibility and Microbial Ecology of the Gut in Weanling Pigs

A total of 126 crossbred weanling pigs (average body weight of 6.3±0.3 kg) were used to investigate the effect of chito-oligosaccharide (COS) on growth performance, nutrient digestibility, pH of gastro-intestinal tract (GI), intestinal and fecal microflora of young piglets. Pigs were allocated to three dietary treatments based on body weight and gender in a single factorial arrangement. Treatments were control (No COS), T1 (0.2% COS during starter (6-13 kg) and 0.1% COS during grower (13-30 kg) phases, and T2 (0.4% COS during starter (6-13 kg) and 0.3% COS during grower (13-30 kg) phases, respectively. Each treatment had 3 replicates and 14 pigs were raised in each pen. COS is a low molecular weight water-soluble chitosan that can be obtained from chitin of the crab shell after deacetylation with concentrated sodium hydroxide at high temperature and then further decomposition by chitosanase enzyme in the presence of ascorbic acid. For the starter and grower periods, there were no significant differences (p>0.05) in average daily gain (ADG) and feed to gain ratio among treatments. However, during the overall period (6-30 kg), T2 showed better (p<0.05) feed to gain ratio than other treatments. A digestibility study was conducted at the end of grower phase which showed improvement (p<0.05) in DM and crude fat digestibility in T2 over the control. At 25 kg body weight, 6 pigs per treatment (2 per replicate) were sacrificed to determine the effect of diets on pH and microbial count at different sections of the GI tract. The pH of the cecal contents in pigs fed 0.1% COS was higher (p<0.05) than in the other treatments. Total anaerobic bacterial number increased from cecum to rectum in all treatments. The weekly total bacterial counts showed higher (p<0.05) in feces of pigs fed COS than that of untreated pigs at the 8 th week. The number of fecal E. coli in untreated pigs at 4 th wk was 7.35 log CFU/g compared to 6.71 and 6.54 log CFU/g in 0.1 and 0.3% COS-treated pigs, respectively. Similarly, at 8 th wk, fecal clostridium spp. were lower in pigs fed 0.3% COS (5.43 log CFU/g) than in untreated pigs (6.26 log CFU/g). In conclusion, these results indicated that chito-oligosaccharide could improve feed efficiency in young pigs and inhibited the growth of harmful bacteria.