Early clinical indicators of acute kidney injury caused by administering high-dose methotrexate therapy to juvenile pigs.

Introduction: Early identification of compromised renal clearance caused by high-dose methotrexate (HDMTX) is essential for initiating timely interventions that can reduce acute kidney injury and MTX-induced systemic toxicity. Methods: We induced acute kidney injury (AKI) by infusing 42 juvenile pigs with 4 g/kg (80 g/m2) of MTX over 4 hours without high-volume alkalinizing hydration therapy. Concentrations of serum creatinine and MTX were measured at 15 time points up to 148 hours, with 10 samples collected during the first 24 hours after the start of the HDMTX infusion. Results: During the first 28 hours, 81% of the pigs had increases in the concentrations of serum creatinine in one or more samples indicative of AKI (i.e., > 0.3g/dL increase). A rate of plasma MTX clearance of less than 90% during the initial 4 hours after the HDMTX infusion and a total serum creatinine increase at 6 and 8 hours after starting the infusion greater than 0.3 g/dL were predictive of AKI at 28 hours (p < 0.05 and p < 0.001, respectively). At conclusion of the infusion, pigs with a creatinine concentration more than 0.3 g/dL higher than baseline or serum MTX greater than 5,000 µmol/L had an increased risk of severe AKI. Conclusions: Our findings suggest that serum samples collected at conclusion and shortly after HDMTX infusion can be used to predict impending AKI. The pig model can be used to identify biological, environmental, and iatrogenic risk factors for HDMTX-induced AKI and to evaluate interventions to preserve renal functions, minimize acute kidney injury, and reduce systemic toxicity.

Report of an Experiment With a Fetal Ex-Utero Support System in Piglets.

Extreme prematurity remains one of the leading causes of neonatal death. An ex-utero treatment strategy that allows the fetus to develop beyond this period until capable of tolerating the transition to post-natal physiology would significantly impact the quality of care offered for this pre-viable patient population. In this study, we report our experience with an ex-utero support system for fetal pigs with the goal of support and survival for eight hours. Our experiment included two pigs at a gestational age equivalent to a 32-week human fetus. Following ultrasound assessment and delivery via hysterotomy, the fetuses were transferred to a 40 L glass aquarium filled with warmed lactated Ringer's solution and connected to an arteriovenous (AV) circuit that included a centrifugal pump and a pediatric oxygenator. Fetus 1 was successfully cannulated and survived for seven hours (expected maximum duration of eight hours). Fetus 2 died shortly after hysterotomy, secondary to failure at the cannulation stage. Our results suggest that ex-utero support of the premature fetal pig is feasible, contributing to a scarce body of evidence. However, further studies are needed before effectively translating an artificial placenta system into the clinical arena.

Selective and Concentrative Enteropancreatic Recirculation of Antibiotics by Pigs.

Antibiotics that are efficacious for infectious pancreatitis are present in pancreatic exocrine secretion (PES) after intravenous administration and above minimal inhibitory concentrations. We measured concentrations of four antibiotics by tandem liquid chromatography-mass spectroscopy in plasma and PES after enteral administration to juvenile pigs with jugular catheters and re-entrant pancreatic-duodenal catheters. Nystatin, which is not absorbed by the intestine nor used for infectious pancreatitis (negative control), was not detected in plasma or PES. Concentrations of amoxicillin increased in plasma after administration (p = 0.035), but not in PES (p = 0.51). Metronidazole and enrofloxacin that are used for infectious pancreatitis increased in plasma after enteral administration and even more so in PES, with concentrations in PES averaging 3.1 (±0.5)- and 2.3 (±0.6)-fold higher than in plasma, respectively (p's < 0.001). The increase in enrofloxacin in PES relative to plasma was lower after intramuscular administration (1.8 ± 0.5; p = 0.001). The present results demonstrate the presence of a selective and concentrative enteropancreatic pathway of secretion for some antibiotics. Unlike the regulated secretion of bile, the constitutive secretion of PES and intestinal reabsorption may provide a continuous exposure of pancreas tissue and the small intestine to recirculated antibiotics and potentially other therapeutic molecules. There is a need to better understand the enteropancreatic recirculation of antibiotics and the associated mechanisms.

Multiple Asparaginase Infusions Cause Increasingly Severe Acute Hyperammonemia.

Adverse reactions during and shortly after infusing asparaginase for the treatment of acute lymphoblastic leukemia can increase in severity with later doses, limiting further use and increasing relapse risk. Although asparaginase is associated with hyperammonemia, the magnitude of the increase in serum ammonia immediately after the infusion and in response to multiple infusions has not been examined. The concurrence of hyperammonemia and infusion reactions was studied using weaned juvenile pigs that received 12 infusions of Erwinia asparaginase (Erwinase; 1250 U/kg) over 28 days, with two 5-day recovery periods without asparaginase after the eighth and eleventh doses. Infusion reactions and prolonged hyperammonemia (>50 µM ammonia 48 h after the infusion) began after the fourth dose and increased with later doses. Dense sampling for 60 min revealed an acute phase of hyperammonemia that peaked within 20 min after starting the first infusion (298 + 62 µM) and lasted less than 1 h, without apparent symptoms. A pronounced acute hyperammonemia after the final infusion (1260 + 250 µM) coincided with severe symptoms and one mortality during the infusion. The previously unrecognized acute phase of hyperammonemia associated with asparaginase infusion coincides with infusion reactions. The juvenile pig is a translational animal model for understanding the causes of acute and chronic hyperammonemia, differentiating from hypersensitivity reactions, and for improving infusion protocols to reduce acute hyperammonemia and to allow the continued use of asparaginase.

Organ Growth and Intestinal Functions of Preterm Pigs Fed Low and High Protein Formulas With or Without Supplemental Leucine or Hydroxymethylbutyrate as Growth Promoters.

The goal of enteral nutritional support for infants born preterm or small for gestational age (SGA) is to achieve normal growth and development. Yet, this is difficult to achieve because of intestinal immaturity. Our objective was to determine if birth weight, protein intake, and the growth promoters leucine (10 g/L) or calcium-ß-hydroxy-ß-methylbutryate (HMB; 1.1 g/L) would affect trajectories of intestinal growth and functions and weights of other organs. Preterm pigs were delivered at gestational day 105 (91% of term) and fed for 6 or 7 days isocaloric formulas that differed in protein content (50 g or 100 g protein/L), with and without the growth promoters leucine or HMB. For comparative purposes organ weights were measured within 12 h after delivery for six term pigs of low and six of average birth weights. The responses of intestinal growth and total intestinal brush border membrane carbohydrases to protein level and supplemental leucine were of greater magnitude for preterm pigs of lower birth weight. Forskolin stimulated chloride secretion in the proximal small intestine was lower for pigs fed the low protein milk replacers. Capacities of the entire small intestine to transport glucose (mmol/kg-day) were not responsive to protein level, leucine, or HMB, and did not differ between small and large pigs. Relative organ weights of the small and average weight term pigs were similar, but some differed from those of the preterm pigs suggesting preterm birth and the standards of care used for this study altered the trajectories of development for the intestine and other organs. Although leucine is an effective generalized growth promoter that enhances gut development of small preterm pigs, it does not mitigate compromised neurodevelopment. Our findings using preterm pigs as a relevant preclinical model indicate nutrition support strategies can influence development of some gastrointestinal tract characteristics and the growth of other organs.

Nutrient Intake and Physical Exercise Significantly Impact Physical Performance, Body Composition, Blood Lipids, Oxidative Stress, and Inflammation in Male Rats.

BACKGROUND: Humans consuming a purified vegan diet known as the "Daniel Fast" realize favorable changes in blood lipids, oxidative stress, and inflammatory biomarkers, with subjective reports of improved physical capacity. OBJECTIVE: We sought to determine if this purified vegan diet was synergistic with exercise in male rats. METHODS: Long⁻Evans rats (n = 56) were assigned to be exercise trained (+E) by running on a treadmill three days per week at a moderate intensity or to act as sedentary controls with normal activity. After the baseline physical performance was evaluated by recording run time to exhaustion, half of the animals in each group were fed ad libitum for three months a purified diet formulated to mimic the Daniel Fast (DF) or a Western Diet (WD). Physical performance was evaluated again at the end of month 3, and body composition was assessed using dual-energy x-ray absorptiometry. Blood was collected for measurements of lipids, oxidative stress, and inflammatory biomarkers. RESULTS: Physical performance at the end of month 3 was higher compared to baseline for both exercise groups (p < 0.05), with a greater percent increase in the DF + E group (99%) than in the WD + E group (51%). Body fat was lower in DF than in WD groups at the end of month 3 (p < 0.05). Blood triglycerides, cholesterol, malondialdehyde, and advanced oxidation protein products were significantly lower in the DF groups than in the WD groups (p < 0.05). No significant differences were noted in cytokines levels between the groups (p > 0.05), although IL-1ß and IL-10 were elevated three-fold and two-fold in the rats fed the WD compared to the DF rats, respectively. CONCLUSIONS: Compared to a WD, a purified diet that mimics the vegan Daniel Fast provides significant anthropometric and metabolic benefits to rats, while possibly acting synergistically with exercise training to improve physical performance. These findings highlight the importance of macronutrient composition and quality in the presence of ad libitum food intake.

Growth Responses of Preterm Pigs Fed Formulas with Different Protein Levels and Supplemented with Leucine or ß-Hydroxyl ß-Methylbutyrate.

Growth after preterm birth is an important determinant of long-term outcomes. Yet, many preterm infants suffer ex utero growth retardation. We evaluated effects of leucine and the metabolite, ß-hydroxy ß-methylbutyrate (HMB) on growth of preterm pigs, a previously-validated translational model for preterm infants. After 48 h of parenteral nutrition preterm pigs were fed for 6 to 7 days isocaloric formulas with different levels of protein (50 or 100 g/L) with leucine (10 g/L, 76 mM) or HMB (at 1.1 g/L, 4 mM) added to stimulate protein synthesis or with alanine (6.8 g/L; 76 mM) as the control. Rates of growth of pigs fed the low protein formula with alanine (3.4 ± 0.2% gain per day) or leucine (3.7 ± 0.2) exceeded that of pigs fed the high protein formula (2.8 ± 0.2, p = 0.02 for comparison with both low protein formulas; p = 0.01 compared with low protein + leucine). Supplementing the high protein formula with leucine or HMB did not increase growth relative to alanine (2.72 ± 0.20, 2.74 ± 0.27, and 2.52 ± 0.20, respectively). Small pigs (.

A Phosphatidylserine Source of Docosahexanoic Acid Improves Neurodevelopment and Survival of Preterm Pigs.

The amount, composition, and sources of nutrition support provided to preterm infants is critical for normal growth and development, and particularly for structural and functional neurodevelopment. Although omega-3 long chain polyunsaturated fatty acids (LC-PUFA), and particularly docosahexanoic acid (DHA), are considered of particular importance, results from clinical trials with preterm infants have been inconclusive because of ethical limitations and confounding variables. A translational large animal model is needed to understand the structural and functional responses to DHA. Neurodevelopment of preterm pigs was evaluated in response to feeding formulas to term-equivalent age supplemented with DHA attached to phosphatidylserine (PS-DHA) or sunflower oil as the placebo. Newborn term pigs were used as a control for normal in utero neurodevelopment. Supplementing formula with PS-DHA increased weight of the brain, and particularly the cerebellum, at term-equivalent age compared with placebo preterm pigs (P's < 0.10 and 0.05 respectively), with a higher degree of myelination in all regions of the brain examined (all p < 0.06). Brains of pigs provided PS-DHA were similar in weight to newborn term pigs. Event-related brain potentials and performance in a novel object recognition test indicated the PS-DHA supplement accelerated development of sensory pathways and recognition memory compared with placebo preterm pigs. The PS-DHA did not increase weight gain, but was associated with higher survival. The benefits of PS-DHA include improving neurodevelopment and possibly improvement of survival, and justify further studies to define dose-response relations, compare benefits associated with other sources of DHA, and understand the mechanisms underlying the benefits and influences on the development of other tissues and organ systems.

Localized Delivery of Cl-Amidine From Electrospun Polydioxanone Templates to Regulate Acute Neutrophil NETosis: A Preliminary Evaluation of the PAD4 Inhibitor for Tissue Engineering.

Upon interaction, neutrophils can potentially release neutrophil extracellular traps (NETs) on the surface of an implanted electrospun template, which may be a significant preconditioning event for implantable biomaterials of yet unknown consequences. In this study, we investigated the potential of polydioxanone templates as a delivery vehicle for Cl-amidine, an inhibitor of peptidyl arginase deiminase 4 (PAD4), and if drug elution could attenuate PAD4-mediated NETosis in the vicinity of implanted templates. Electrospun polydioxanone templates were fabricated with distinct architectures, small diameter (0.4 µm) or large diameter (1.8 µm) fibers, and incorporated with 0-5 mg/mL Cl-amidine to examine dose-dependent effects. Acute neutrophil-template interactions were evaluated in vitro with freshly isolated human neutrophils and in vivo with a rat subcutaneous implant model. The in vitro results suggest large diameter templates with 0 mg/mL Cl-amidine significantly attenuate NETosis compared to small diameter templates. As the drug concentration increased, NETosis was significantly decreased on small diameter templates in a dose-dependent manner. The opposite was observed for large diameter templates, indicating multiple mechanisms of NETosis may be regulating neutrophil template preconditioning. Similar results were observed in vivo, verifying local NETosis inhibition by Cl-amidine eluting templates in a physiological environment. Importantly, large diameter templates with Cl-amidine enhanced neutrophil invasion and survival, supporting the potential for long-term modulation of tissue integration and regeneration. This preliminary study demonstrates a novel delivery vehicle for Cl-amidine that can be used to regulate acute NETosis as the potential critical link between the innate immune response, inflammation, and template-guided tissue regeneration.

Maturation of the Coordination Between Respiration and Deglutition with and Without Recurrent Laryngeal Nerve Lesion in an Animal Model.

The timing of the occurrence of a swallow in a respiratory cycle is critical for safe swallowing, and changes with infant development. Infants with damage to the recurrent laryngeal nerve, which receives sensory information from the larynx and supplies the intrinsic muscles of the larynx, experience a significant incidence of dysphagia. Using our validated infant pig model, we determined the interaction between this nerve damage and the coordination between respiration and swallowing during postnatal development. We recorded 23 infant pigs at two ages (neonatal and older, pre-weaning) feeding on milk with barium using simultaneous high-speed videofluoroscopy and measurements of thoracic movement. With a complete linear model, we tested for changes with maturation, and whether these changes are the same in control and lesioned individuals. We found (1) the timing of swallowing and respiration coordination changes with maturation; (2) no overall effect of RLN lesion on the timing of coordination, but (3) a greater magnitude of maturational change occurs with RLN injury. We also determined that animals with no surgical intervention did not differ from animals that had surgery for marker placement and a sham procedure for nerve lesion. The coordination between respiration and swallowing changes in normal, intact individuals to provide increased airway protection prior to weaning. Further, in animals with an RLN lesion, the maturation process has a larger effect. Finally, these results suggest a high level of brainstem sensorimotor interactions with respect to these two functions.

The Risk of Necrotizing Enterocolitis Differs Among Preterm Pigs Fed Formulas With Either Lactose or Maltodextrin.

OBJECTIVES: When breast milk is unavailable for preterm infants, formulas are needed that won't increase the risk of necrotizing enterocolitis (NEC). Adding novel ingredients to formula to reduce NEC has not been effective clinically. Instead, we tested the prediction that NEC can be reduced by removing the maltodextrin now included in preterm formulas. METHODS: The preterm pig model of spontaneous NEC was used to evaluate growth, health, and intestinal responses to 6 to 7 days of feeding formulas that were identical except for the source of carbohydrate; either 100% lactose or maltodextrin; colostrum was used as the control. RESULTS: Formula with maltodextrin resulted in a 50% incidence of NEC with 30% mortality. The lactose formula and colostrum resulted in a 0% incidence of NEC. Growth was highest for pigs fed the formula with lactose, intermediate with maltodextrin, and minimal when bovine colostrum was fed (P < 0.05). Although the small intestine was larger when colostrum was fed (P < 0.05), because rates of glucose uptake were lower (P < 0.05), total small intestine capacities to transport glucose were similar for healthy pigs in all 3 groups. CONCLUSIONS: If lactose-based formulas reduce NEC clinically, the transition of preterm infants to enteral feeding can be accelerated, improving growth and development, and shortening reliance on parenteral nutrition. Although colostrum protects against NEC, chronic feeding does not promote body weight gain after preterm birth. The preterm pig can be used for preclinical studies that evaluate the mechanisms by which carbohydrates and other ingredients influence growth, development, health, and risk of NEC.

Intestinal adaptations to a combination of different diets with and without endurance exercise.

BACKGROUND: Endurance athletes search for diet regimens that will improve performance and decrease gastrointestinal disturbances during training and events. Although the intestine can adapt to changes in the amount and composition of dietary inputs, the responses to the combination of endurance exercise and diet are poorly understood. METHODS: We evaluated small intestinal dimensions and mucosal architecture and calculated the capacities of the entire small intestine to digest maltose and maltodextrin and absorb glucose in response to two different diet types; a western human diet and the Daniel Fast, a vegan style diet, and with moderate intensity endurance training or a no-exercise sedentary lifestyle for a 13 week period (n = 7 per group). The influences of diet and exercise, alone and in combination, were analyzed by analysis of variation. RESULTS: Rats fed the western diet gained more weight (P < 0.05) due to more fat mass (P < 0.05), with a similar response for the sedentary compared with the exercised rats in each diet group (P < 0.05). The Daniel Fast rats had longer and heavier intestines with deeper crypts with villi that were wider (P < 0.05), but not taller. Despite increased energetic demands, the exercised rats had shorter and lighter intestines with shorter villi (P < 0.05). Yet, the percentage of mucosa did not differ among groups. Total small intestinal activities for maltase and α-glucoamylase, and capacities for glucose absorption were similar regardless of diet or exercise. CONCLUSIONS: These findings indicate the structural responses of the small intestine to a vegan style diet are modified by exercise, but without altering the capacities of the brush border membrane to digest and absorb carbohydrates.

Responses of Preterm Pigs to an Oral Fluid Supplement During Parenteral Nutrition.

BACKGROUND: Nutrients and electrolytes in amniotic fluid swallowed by fetuses are important for growth and development. Yet, preterm infants requiring parenteral nutrition (PN) receive minimal or no oral inputs. With the limited availability of amniotic fluid, we evaluated the responses of preterm pigs receiving PN to an oral fluid supplement (OFS) based on the electrolyte and nutrient composition of amniotic fluid. MATERIALS AND METHODS: Preterm pigs (92% of term) received a combination of PN (6 mL/kg-h) and 4 mL/kg-h of supplemental fluid as an experimental OFS (n = 9), lactated Ringer's either enterally (n = 10) or intravenously (n = 8). Outcome measures after 96 hours were weight gain, blood chemistry, organ weights, and small intestine mass and brush-border membrane carbohydrases. RESULTS: The OFS did not improve weight gain compared with providing lactated Ringer's orally or intravenously, or increase serum urea nitrogen values, but resulted in higher serum total and low-density lipoprotein cholesterol, as well as improved glucoregulation and heavier intestines, livers, kidneys, and brains and lighter lungs. CONCLUSIONS: Providing supplemental fluid and electrolytes during PN either intravenously or orally increases weight gain after preterm birth. An oral fluid supplement based on amniotic fluid may accelerate development and maturation of organs critical for extrauterine life after preterm birth and may enhance neurodevelopment.

A preterm pig model of lung immaturity and spontaneous infant respiratory distress syndrome.

Respiratory distress syndrome (RDS) and bronchopulmonary dysplasia remain the leading causes of preterm infant morbidity, mortality, and lifelong disability. Research to improve outcomes requires translational large animal models for RDS. Preterm pigs delivered by caesarian section at gestation days (GD) 98, 100, 102, and 104 were provided 24 h of neonatal intensive care, monitoring (pulse oximetry, blood gases, serum biomarkers, radiography), and nutritional support, with or without intubation and mechanical ventilation (MV; pressure control ventilation with volume guarantee). Spontaneous development of RDS and mortality without MV are inversely related with GD at delivery and correspond with inadequacy of tidal volume and gas exchange. GD 98 and 100 pigs have consolidated lungs, immature alveolar architecture, and minimal surfactant protein-B expression, and MV is essential at GD 98. Although GD 102 pigs had some alveoli lined by pneumocytes and surfactant was released in response to MV, blood gases and radiography revealed limited recruitment 1-2 h after delivery, and mortality at 24 h was 66% (35/53) with supplemental oxygen provided by a mask and 69% (9/13) with bubble continuous positive airway pressure (8-9 cmH2O). The lungs at GD 104 had higher densities of thin-walled alveoli that secreted surfactant, and MV was not essential. Between GD 98 and 102, preterm pigs have ventilation inadequacies and risks of RDS that mimic those of preterm infants born during the saccular phase of lung development, are compatible with standards of neonatal intensive care, and are alternative to fetal nonhuman primates and lambs.

Parenteral nutrition compromises neurodevelopment of preterm pigs.

BACKGROUND: Despite advances in nutritional support and intensive care, preterm infants are at higher risk of compromised neurodevelopment. OBJECTIVE: This study evaluated the contribution of total parenteral nutrition (PN) to compromised neurodevelopment after preterm birth. METHODS: Preterm pigs were provided PN or enteral nutrition (EN) for 10 d. Neurodevelopment was assessed by observations of motor activity and evaluation of sensory/motor reflexes, brain weight, MRI, and cerebellar histology. RESULTS: Despite similar gains in body weight, PN pigs had smaller brains (32 ± 0.4 vs. 35 ± 0.6 g; P = 0.0002) including the cerebellum, as well as reduced motor activity (P = 0.005), which corresponded to underdeveloped myelination (P = 0.004) measured by diffusion tensor imaging. PN resulted in lower serum triglycerides (17 ± 5.9 vs. 27 ± 3.1 mg/dL; P = 0.05), total cholesterol (31 ± 9.6 vs. 85 ± 8.1 mg/dL; P = 0.04), VLDL cholesterol (3.7 ± 1.2 vs. 5.7 ± 0.7 mg/dL; P = 0.04), and HDL cholesterol (16 ± 4.6 vs. 57 ± 7.3 mg/dL; P = 0.03) and nonsignificantly lower LDL cholesterol (10.7 ± 4.4 vs. 22.7 ± 2.9 mg/dL; P = 0.09). CONCLUSIONS: The compromised neurodevelopment caused by total PN is a novel finding, was independent of confounding variables (disease, inconsistent gestational ages, diverse genetics, extrauterine growth retardation, and inconsistent neonatal intensive care unit protocols), and highlights a need to improve current PN solutions. The preterm pig is a translational animal model for improving nutrition support to enhance neurodevelopment of preterm infants requiring PN.

The weaned pig as a model for Doxorubicin-induced mucositis.

BACKGROUND: Chemotherapy-induced mucositis (CIM) complicates cancer therapy and limits maximum tolerated doses and efficacy. Rodent models do not reproducibly mimic clinical CIM, so alternative models are needed. METHODS: CIM severity was assessed after weaned pigs were treated with doxorubicin (5 and 3.75 mg/kg) using clinical observations, laboratory parameters and gastrointestinal structure and functions. Bovine colostrum was provided as an experimental intervention to the pigs treated receiving the 3.75 mg/kg dose. RESULTS: Doxorubin at 3.75 mg/kg decreased food intake and weight gain (p < 0.05) and caused diarrhea and vomiting that coincided with damage to the small intestine mucosa based on histological scoring (p < 0.05). It resulted in higher serum TNF-α concentrations, increased chloride secretion and reduced brush border membrane disaccharidase activities and carrier-mediated glucose uptake (all p < 0.05). The gastrointestinal damage and dysfunction resemble the clinical and laboratory features of CIM in humans; these can be partially prevented by providing cow colostrum. CONCLUSION: The weaned pig is a relevant large animal for studying CIM and evaluating existing and experimental interventions for mucositis.

Maternal-to-infant transmission of probiotics: concept validation in mice, rats, and pigs.

BACKGROUND: Postnatal introduction of probiotics results in a low incidence of colonization, whereas maternal fecal and vaginal bacteria colonize the gastrointestinal tract (GIT) of vaginally delivered infants. OBJECTIVE: We tested if probiotic bacteria, fed to three pregnant animal models, would colonize the GIT of offspring delivered vaginally. METHODS: Probiotic strains of Lactobacillus acidophilus and Bifidobacterium lactis were fed to pregnant mice, rats, and sows for at least 7 days prior to vaginal delivery. Cultural approaches and genotyping were used to determine if the probiotic bacteria colonized the GIT after birth. RESULTS: The probiotic bacteria were detected in the feces and vagina of maternal mice, rats, and sows after, but not before, administration. L. acidophilus was detected at postnatal day 14 in 22, 33, and 75% of the mice, rats, and pigs, respectively, and after weaning in 35% of the mice and 1 of 5 pigs. B. lactis was present at postnatal day 14 in 30 and 80% of the mice and pigs. Bacterial assemblages in the GIT of the colonized young differed from those in which the probiotics were not detected. CONCLUSIONS: Probiotic bacteria administered to mothers during late gestation are transferred to infants born vaginally and influence the assemblages of GIT bacteria. However, colonization of the neonatal GIT and persistence past weaning does not occur in all offspring and varies among probiotics and animal models.

Composite chitosan/nano-hydroxyapatite scaffolds induce osteocalcin production by osteoblasts in vitro and support bone formation in vivo.

There is a significant clinical need to develop alternatives to autografts and allografts for bone grafting procedures. Porous, biodegradable scaffolds based on the biopolymer chitosan have been investigated as bone graft substitutes, and the addition of calcium phosphate to these scaffolds has been shown to improve the mechanical properties of the scaffold and may increase osteoconductivity. In this study, in vitro mineralization was examined for osteoblasts seeded in a porous scaffold composed of fused chitosan/nano-hydroxyapatite microspheres. Human fetal osteoblasts were cultured on composite and chitosan scaffolds for 21 days. On days 1, 4, 7, 14, and 21, total dsDNA, alkaline phosphatase, type I collagen, and osteocalcin production were measured. Total cellularity (measured by dsDNA), alkaline phosphatase, and type I collagen production were similar between the two scaffold groups. However, osteocalcin production occurred significantly earlier (day 7 vs. day 21) and was more than three times greater (0.0022 vs. 0.0068 ng/mL/ng DNA) on day 21 when osteoblasts were cultured on composite scaffolds. Osteocalcin is a marker of late osteoblastic differentiation and mineralized bone matrix formation. Therefore, the increase in osteocalcin production seen when cells were cultured on composite scaffolds may indicate that these scaffolds were superior to chitosan-only scaffolds in facilitating osteoblast mineralization. Composite scaffolds were also shown to be biocompatible and osteoconductive in a preliminary critical size rat calvarial defect study. These results demonstrate the potential of composite chitosan/nano-hydroxyapatite scaffolds to be used in bone tissue engineering.

Oral administration of sucrose solutions and measurement of serum sucrose concentrations to evaluate gastric permeability in adult bottlenose dolphins (Tursiops truncatus).

OBJECTIVE: To measure concentrations of sucrose in the serum of captive dolphins after oral administration of a sucrose solution and determine the suitability of this method for use as a test to detect gastric ulcers. ANIMALS: 8 adult captive bottlenose dolphins (Tursiops truncatus). PROCEDURES: Blood samples were collected from the ventral fluke vein of dolphins before and 45 minutes after oral administration of 500 mL of solution containing 25 or 50 g of sucrose; oral administration was achieved by use of gastric intubation. Serum was separated, diluted in a solution of 90% acetonitrile-to 10% water that contained 10 ng of an internal standard (trichlormethiazide)/microL, mixed, and centrifuged. Supernatant was analyzed by use of liquid chromatography-mass spectrometry-mass spectrometry (LC-MS-MS). RESULTS: Serum sucrose concentrations of dolphins were at or less than the limits of detection before oral administration. Values after administration of sucrose solution varied among dolphins and were higher and more variable after administration of 50 g, compared with concentrations after administration of 25 g. CONCLUSIONS AND CLINICAL RELEVANCE: Serum sucrose concentrations in samples collected during routine health evaluations of captive dolphins can be reliably measured by use of LC-MS-MS. Correlating serum sucrose concentrations with endoscopic observations of the gastric mucosa of dolphins will validate this approach for use in screening for the prevalence and severity of gastric ulcers and determining the efficacy of treatment regimens.

Absorption of alpha-ketoglutarate by the gastrointestinal tract of pigs.

Only a small percentage of alpha-ketoglutarate (AKG) administered lumenally to pigs appears in the portal circulation. This has been attributed to mucosal metabolism, and possibly by limited absorption. Although transporters for di- and tricarboxylic acids, which includes the sodium-dependent transporter NaDC-1, have been detected in the small intestine, correlations with functional assays are lacking. Therefore, intact tissues from three regions of the small intestine, stomach, and colon of weaned pigs were used to measure rates of AKG absorption. Western analysis was used to detect NaDC-1 in the three regions of small intestine. Rates of AKG absorption were highest in the small intestine, lowest in the colon, and intermediate in the stomach. Immunoreactive NaDC-1 was detected in the small intestine and this coincided with a component of AKG absorption that was inhibited by AKG and succinate. In contrast, absorption of AKG was inhibitable by unlabeled AKG, but not succinate, in the stomach, and by neither in the colon. Feeding studies indicated that the amounts of AKG that might be included in practical diets for pigs would not (1) upregulate rates of AKG absorption or (2) exceed estimated capacities of the small intestine to absorb AKG. The present findings indicate that the efficacy of AKG as an alternative metabolic fuel for enterocytes to spare dietary amino acids is not limited by absorption.