First description of the baubellum in the spectacled bear Tremarctos ornatus (Mammalia: Ursidae).

The baubellum (os clitoridis) is a bone found in the clitoris of many female eutherian mammals and is homologous to the baculum in males. In contrast to the baculum, the baubellum has received very little attention regarding its morphological or interspecific diversity, or on hypotheses for its function. The presence of the baubellum in bears (Ursidae) has only been established and mentioned in the literature for the Ursus genus, and not for the other genera of bears. Moreover, no scaled photographs are available for baubella of this clade, and the sizes reported vary between sources. We hereby present and describe the baubellum of a spectacled bear (Tremarctos ornatus), providing a detailed account of baubella in a basal ursid species. The baubellum of Tremarctos is slightly bowed dorsally, with two small prominences at the distal apex. The length of the Tremarctos baubellum in this study is comparable to that of Ursus americanus (American black bear). We infer the specific shape, with longitudinal ridges, of the baubellum in Tremarctos could indicate a discrete function during copulation or sexual arousal. However, future studies, especially regarding the associated soft tissues, will be required to confirm whether this is indeed the case. Our study expands the understanding of baubella within Ursidae, providing new data (including a three-dimensional model) that can be used to further explore the morphological diversity and function of this enigmatic extra-skeletal bone.

Drug Disposition in Neonatal Göttingen Minipigs: Exploring Effects of Perinatal Asphyxia and Therapeutic Hypothermia.

Asphyxiated neonates often undergo therapeutic hypothermia (TH) to reduce morbidity and mortality. Since both perinatal asphyxia (PA) and TH influence physiology, altered pharmacokinetics (PK) and pharmacodynamics (PD) are expected. Given that TH is the standard of care for PA with moderate to severe hypoxic-ischemic encephalopathy, disentangling the effect of PA versus TH on PK/PD is not possible in clinical settings. However, animal models can provide insights into this matter. The (neonatal) Göttingen Minipig, the recommended strain for nonclinical drug development, was selected as translational model. Four drugs-midazolam (MDZ), fentanyl (FNT), phenobarbital (PHB), and topiramate (TPM)-were intravenously administered under four conditions: control (C), therapeutic hypothermia (TH), hypoxia (H), and hypoxia plus TH (H+TH). Each group included six healthy male neonatal Göttingen Minipigs anesthetized for 24 hours. Blood samples were drawn at 0 (predose) and 0.5, 2, 2.5, 3, 4, 4.5, 6, 8, 12, and 24 hours post drug administration. Drug plasma concentrations were determined using validated bioanalytical assays. The PK parameters were estimated through compartmental and noncompartmental PK analysis. The study showed a statistically significant decrease in FNT clearance (CL; 66% decrease), with an approximately threefold longer half-life (t1/2) in the TH group. The H+TH group showed a 17% reduction in FNT CL, with a 62% longer t1/2 compared with the C group; however, it was not statistically significant. Although not statistically significant, trends toward lower CL and longer t1/2 were observed in the TH and H+TH groups for MDZ and PHB. Additionally, TPM demonstrated a 28% decrease in CL in the H group compared with controls. SIGNIFICANCE STATEMENT: The overarching goal of this study using the neonatal Göttingen Minipig model was to disentangle the effects of systemic hypoxia and TH on PK using four model drugs. Such insights can subsequently be used to inform and develop a physiologically based pharmacokinetic model, which is useful for drug exposure prediction in human neonates.

Preweaning performance in intrauterine growth-restricted piglets: Characteristics and interventions.

Intrauterine growth restriction (IUGR) is frequently observed in pig production, especially when using highly prolific sows. IUGR piglets are born with low body weight and shape indicative of differences in organ growth. Insufficient uteroplacental nutrient transfer to the fetuses is the leading cause of growth restriction in the pig. Supplementing the sow's gestation diet with arginine and/or glutamine improves placenta growth and functionality and consequently is able to reduce IUGR incidence. IUGR piglets are at higher risk of dying preweaning and face higher morbidity than their normal-weight littermates. A high level of surveillance during farrowing and individual nutrient supplementation can reduce the mortality rates. Still, these do not reverse the long-term consequences of IUGR, which are induced by persistent structural deficits in different organs. Dietary interventions peri-weaning can optimize performance but these are less effective in combating the metabolic changes that occurred in IUGR, which affect reproductive performance later in life. IUGR piglets share many similarities with IUGR infants, such as a poorer outcome of males. Using the IUGR piglet as an animal model to further explore the structural and molecular basis of the long-term consequences of IUGR and the potential sex bias could aid in fully understanding the impact of prenatal undernutrition and finding solutions for both species and sexes.

Low birth weight female piglets show altered intestinal development, gene expression, and epigenetic changes at key developmental loci.

Intestinal development is compromised in low birth weight (LBW) pigs, negatively impacting their growth, health, and resilience. We investigated the molecular mechanisms of the altered intestinal maturation observed in neonatal and juvenile LBW female piglets by comparing the changes in intestinal morphology, gene expression, and methylation in LBW versus normal birth weight (NBW) female piglets. A total of 16 LBW/NBW sibling pairs were sacrificed at 0 hours, 8 hours, 10 days, and 8 weeks of age. The gastrointestinal tract was weighed, measured, and the small intestine was sampled for histomorphology, gene expression, and methylation analyses. Impaired intestinal development, with shorter villi and shallower crypts, was observed in LBW female piglets. The expression of intestinal development markers (ALPI and OLFM) rapidly peaked after birth in NBW but not in LBW female piglets. The lower expression of genes involved in nutrient digestion (ANPEP and SI) and barrier function (OCLN and CLDN4) in LBW, together with their delayed development of intestinal villi and crypts could help to explain the compromised health and growth potential of LBW female piglets. The changes in methylation observed in LBW in key regulators of intestinal development (OLFM4 and FZD5) suggest long-term effects of BW on intestinal gene expression, development, and function. Accordingly, experimental demethylation induced in IPEC-J2 cells led to increased expression of intestinal genes (MGA, DPP4, and GLUT2). Overall, we have identified the alterations in transcription or epigenetic marking at a number of genes critical to intestinal development, which may contribute to both the short- and long-term failure of LBW female piglets to thrive.

Developmental Toxicity and Biotransformation of Two Anti-Epileptics in Zebrafish Embryos and Early Larvae.

The zebrafish (Danio rerio) embryo is gaining interest as a bridging tool between in-vitro and in-vivo developmental toxicity studies. However, cytochrome P450 (CYP)-mediated drug metabolism in this model is still under debate. Therefore, we investigated the potential of zebrafish embryos and larvae to bioactivate two known anti-epileptics, carbamazepine (CBZ) and phenytoin (PHE), to carbamazepine-10,11-epoxide (E-CBZ) and 5-(4-hydroxyphenyl)-5-phenylhydantoin (HPPH), respectively. First, zebrafish were exposed to CBZ, PHE, E-CBZ and HPPH from 5»- to 120-h post fertilization (hpf) and morphologically evaluated. Second, the formations of E-CBZ and HPPH were assessed in culture medium and in whole-embryo extracts at different time points by targeted LC-MS. Finally, E-CBZ and HPPH formation was also assessed in adult zebrafish liver microsomes and compared with those of human, rat, and rabbit. The present study showed teratogenic effects for CBZ and PHE, but not for E-CBZ and HPPH. No HPPH was detected during organogenesis and E-CBZ was only formed at the end of organogenesis. E-CBZ and HPPH formation was also very low-to-negligible in adult zebrafish compared with the mammalian species. As such, other metabolic pathways than those of mammals are involved in the bioactivation of CBZ and PHE, or, these anti-epileptics are teratogens and do not require bioactivation in the zebrafish.

A Medium-Throughput System for In Vitro Oxidative Stress Assessment in IPEC-J2 Cells.

The feed industry continuously seeks new molecules with antioxidant capacity since oxidative stress plays a key role in intestinal health. To improve screening of new antioxidants, this study aims to set up an assay to assess oxidative stress in the porcine small intestinal epithelial cell line IPEC-J2 using plate-reader-based analysis of fluorescence. Two oxidants, H2O2 and menadione, were tested at 1, 2 and 3 mM and 100, 200 and 300 µM, respectively. Trolox (2 mM) was used as the reference antioxidant and the probe CM-H2DCFDA was used to indicate intracellular oxidative stress. Cell culture, reactive oxygen species (ROS) production and assessment conditions were optimized to detect a significant ROS accumulation that could be counteracted by pre-incubation with trolox. Menadione (200 µM) reproducibly increased ROS levels, H2O2 failed to do so. Trolox significantly decreased intracellular ROS levels in menadione (200 µM)-exposed cells in a consistent way. The system was further used to screen different concentrations of the commercially available antioxidant ELIFE®. Concentrations between 100 and 200 ppm protected best against intracellular ROS accumulation. In conclusion, the combination of CM-H2DCFDA fluorescence analysis by a plate-reader, trolox as a reference antioxidant and 200 µM of menadione as a stressor agent, provides a replicable and reliable medium-throughput setup for the evaluation of intracellular oxidative stress in IPEC-J2 cells.

Rapid Postnatal Adaptation of Neurodevelopment in Pigs Born Late Preterm.

Preterm birth interrupts intrauterine brain growth and maturation and may induce a delay in postnatal neurodevelopment. Such developmental delays can result from the reduced fetal age at birth, together with the clinical compli-cations of preterm birth (e.g., hypoxia, ischemia, and inflammation). We hypothesized that late preterm birth, inducing only mild clinical complications, has minimal effects on brain-related outcomes such as motor function and behavior. Using the pig as a model for late preterm infants, piglets were cesarean delivered preterm (90%, 106 days gestation) or at full term, reared by identical procedures, and euthanized for tissue collection at birth or after 11 days (e.g., term-corrected age for preterm pigs). Clinical variables and both structural and functional brain endpoints were assessed. The preterm pigs were slow to get on their feet, gained less weight (-30%), and had a higher cerebral hydration level and blood-to-cerebrospinal fluid permeability than the term pigs. At term-corrected age (11 days), the absolute weight of the brain and the weights of its regions were similar between 11-day-old preterm and newborn term pigs, and both were lower than in 11-day-old term pigs. Postnatally, physical activity and movements in an open field were similar, except that preterm pigs showed a reduced normalized stride length and increased normalized maximum stride height. Perinatal brain growth is closely associated with advancing postconceptional age in pigs, and late preterm birth is initially associated with impaired brain growth and physical activity. Postnatally, neuromuscular functions mature rapidly and become similar to those in term pigs, even before term-corrected age. Neuromuscular functions and behavior may show rapid postnatal adaptation to late preterm birth in both pigs and infants.

From mRNA Expression of Drug Disposition Genes to In Vivo Assessment of CYP-Mediated Biotransformation during Zebrafish Embryonic and Larval Development.

The zebrafish (Danio rerio) embryo is currently explored as an alternative for developmental toxicity testing. As maternal metabolism is lacking in this model, knowledge of the disposition of xenobiotics during zebrafish organogenesis is pivotal in order to correctly interpret the outcome of teratogenicity assays. Therefore, the aim of this study was to assess cytochrome P450 (CYP) activity in zebrafish embryos and larvae until 14 d post-fertilization (dpf) by using a non-specific CYP substrate, i.e., benzyloxy-methyl-resorufin (BOMR) and a CYP1-specific substrate, i.e., 7-ethoxyresorufin (ER). Moreover, the constitutive mRNA expression of CYP1A, CYP1B1, CYP1C1, CYP1C2, CYP2K6, CYP3A65, CYP3C1, phase II enzymes uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1) and sulfotransferase 1st1 (SULT1ST1), and an ATP-binding cassette (ABC) drug transporter, i.e., abcb4, was assessed during zebrafish development until 32 dpf by means of quantitative PCR (qPCR). The present study showed that trancripts and/or the activity of these proteins involved in disposition of xenobiotics are generally low to undetectable before 72 h post-fertilization (hpf), which has to be taken into account in teratogenicity testing. Full capacity appears to be reached by the end of organogenesis (i.e., 120 hpf), although CYP1-except CYP1A-and SULT1ST1 were shown to be already mature in early embryonic development.

Intrauterine growth restriction in neonatal piglets affects small intestinal mucosal permeability and mRNA expression of redox-sensitive genes.

Neonates with intrauterine growth restriction (IUGR) show lower efficiency of nutrient utilization compared to normal birth weight (NBW) newborns. This study was conducted using neonatal piglets as a model to test the hypothesis that IUGR affects the intestinal barrier function, intestinal structure, and antioxidant system development during the suckling period. The small intestinal mucosae were obtained from IUGR and NBW littermates in the suckling period (d 0, 3, 8, and 19 postnatal). The epithelial barrier function was assessed by FITC-dextran 4 (FD4) and horseradish peroxidase (HRP) fluxes across the epithelium, histomorphologic measurements, and expression of tight-junction proteins. Redox status represented by the glutathione disulfide/glutathione ratio and malondialdehyde concentrations was determined, whereas mRNA expressions of some redox-sensitive proteins were quantified. Results showed that IUGR piglets exhibited a 2-fold higher intestinal permeability in the proximal small intestine on d 0 (P < 0.05), and this difference between IUGR and NBW piglets was widened to 3 and 4 times for FD4 and HRP, respectively (P < 0.05), on d 3. In accordance, expression of occludin was down-regulated at the transcriptional level in IUGR piglets at d 0 and 19 (P < 0.01). Furthermore, the transcription of heme oxygenase 1, catalase, and thioredoxin reductase genes was down-regulated in IUGR piglets, mainly on postnatal d 0 and 19 (P < 0.01). It appears that IUGR subjects have a lower capacity to mount an antioxidant response in the early postnatal period. Collectively, these results add to our understanding of the mechanisms responsible for intestinal dysfunction in IUGR neonates.

How innate is locomotion in precocial animals? A study on the early development of spatio-temporal gait variables and gait symmetry in piglets.

Locomotion is one of the most important ecological functions in animals. Precocial animals, such as pigs, are capable of independent locomotion shortly after birth. This raises the question whether coordinated movement patterns and the underlying muscular control in these animals is fully innate or whether there still exists a rapid maturation. We addressed this question by studying gait development in neonatal pigs through the analysis of spatio-temporal gait characteristics during locomotion at self-selected speed. To this end, we made video recordings of piglets walking along a corridor at several time points (from 0 h to 96 h). After digitization of the footfalls, we analysed self-selected speed and spatio-temporal characteristics (e.g. stride and step lengths, stride frequency and duty factor) to study dynamic similarity, intralimb coordination and interlimb coordination. To assess the variability of the gait pattern, left-right asymmetry was studied. To distinguish neuromotor maturation from effects caused by growth, both absolute and normalized data (according to the dynamic similarity concept) were included in the analysis. All normalized spatio-temporal variables reached stable values within 4 h of birth, with most of them showing little change after the age of 2 h. Most asymmetry indices showed stable values, hovering around 10%, within 8 h of birth. These results indicate that coordinated movement patterns are not entirely innate, but that a rapid neuromotor maturation, potentially also the result of the rearrangement or recombination of existing motor modules, takes place in these precocial animals.

In vitro Phase I- and Phase II-Drug Metabolism in The Liver of Juvenile and Adult Göttingen Minipigs.

PURPOSE: In view of pediatric drug development, juvenile animal studies are gaining importance. However, data on drug metabolizing capacities of juvenile animals are scarce, especially in non-rodent species. Therefore, we aimed to characterize the in vitro biotransformation of four human CYP450 substrates and one UGT substrate in the livers of developing Göttingen minipigs. METHODS: Liver microsomes from late fetal, Day 1, Day 3, Day 7, Day 28, and adult male and female Göttingen minipigs were incubated with a cocktail of CYP450 substrates, including phenacetin, tolbutamide, dextromethorphan, and midazolam. The latter are probe substrates for human CYP1A2, CYP2C9, CYP2D6, and CYP3A4, respectively. In addition, the UGT multienzyme substrate (from the UGT-GloTM assay), which is glucuronidated by several human UGT1A and UGT2B enzymes, was also incubated with the porcine liver microsomes. RESULTS: For all tested substrates, drug metabolism significantly rose postnatally. At one month of age, 60.5 and 75.4% of adult activities were observed for acetaminophen and dextrorphan formations, respectively, while 35.4 and 43.2% of adult activities were present for 4-OH-tolbutamide and 1'-OH-midazolam formations. Biotransformation of phenacetin was significantly higher in 28-day-old and adult females compared with males. CONCLUSIONS: Maturation of metabolizing capacities occurred postnatally, as described in man.

Adapted Morris Water Maze protocol to prevent interference from confounding motor deficits on cognitive functioning.

Purpose/aim of the study: Cognitive functioning in the Morris Water Maze (MWM) is assumed to be reflected by path length. In this study, the interference of motor deficits, as a confounding factor on cognitive functioning, was assessed by means of a lateralization study with hemicerebellectomized (HCX) mice. This model is characterized by motor deficits restricted to the lesion side, allowing comparison within the model itself (left vs. right), rather than the effect of the manipulation on this measure (experimental vs. control). MATERIALS AND METHODS: Spatial learning was assessed after left or right hemicerebellectomy in adult mice by means of two MWM designs in which the location of the starting positions was altered for one condition in the adapted (Adap) MWM experiment, hypothesizing that motor impairments ipsilateral to the lesion side result in a difference in path length. RESULTS: When the starting positions were equal for both conditions in the traditional (Trad) MWM experiment, path length during the acquisition phase and spatial memory were more affected for the left HCX, while these effects disappeared after mirroring the starting positions in the Adap MWM, implying that motor phenotype and corresponding increase in task difficulty are responsible for the contradictory results in the Trad MWM experiment. CONCLUSION: The differences found in the latter experiment were circumvented in the adapted MWM protocol, and therefore, excluding the motor deficit as a confounding factor on cognitive MWM parameters.

In Vitro Biotransformation of Two Human CYP3A Probe Substrates and Their Inhibition during Early Zebrafish Development.

At present, the zebrafish embryo is increasingly used as an alternative animal model to screen for developmental toxicity after exposure to xenobiotics. Since zebrafish embryos depend on their own drug-metabolizing capacity, knowledge of their intrinsic biotransformation is pivotal in order to correctly interpret the outcome of teratogenicity assays. Therefore, the aim of this in vitro study was to assess the activity of cytochrome P450 (CYP)-a group of drug-metabolizing enzymes-in microsomes from whole zebrafish embryos (ZEM) of 5, 24, 48, 72, 96 and 120 h post-fertilization (hpf) by means of a mammalian CYP substrate, i.e., benzyloxy-methyl-resorufin (BOMR). The same CYP activity assays were performed in adult zebrafish liver microsomes (ZLM) to serve as a reference for the embryos. In addition, activity assays with the human CYP3A4-specific Luciferin isopropyl acetal (Luciferin-IPA) as well as inhibition studies with ketoconazole and CYP3cide were carried out to identify CYP activity in ZLM. In the present study, biotransformation of BOMR was detected at 72 and 96 hpf; however, metabolite formation was low compared with ZLM. Furthermore, Luciferin-IPA was not metabolized by the zebrafish. In conclusion, the capacity of intrinsic biotransformation in zebrafish embryos appears to be lacking during a major part of organogenesis.

Organ data from the developing Göttingen minipig: first steps towards a juvenile PBPK model.

The Göttingen minipig is the most commonly used pig breed in preclinical drug development in Europe and has recently also been explored for physiologically based pharmacokinetic modelling. To develop such a model, not only physiological data from adult animals but also data from juvenile animals are required, especially when using this model for paediatric drug development. Therefore, the aim of our study was to document body and organ weights (brain, heart, lungs, liver, gastrointestinal tract, spleen and kidney), lengths of the small and large intestines and pH values of the gastrointestinal tract in Göttingen minipigs from the foetal stage until the age of 5 months. Postnatal organ and body weights were fitted to regression models to find suitable equations that could be used to estimate organ weights as a function of body weight in the neonatal and juvenile Göttingen minipig. Most organs followed a non-linear growth curve during the first 5 months of life. In general, relative organ weights were the highest during the first week of life, during which the gastric pH was more alkaline than at 28 days of age.

Physical activity level is impaired and diet dependent in preterm newborn pigs.

BACKGROUND: Preterm infants show delayed development of motor function after birth. This may relate to functional immaturity of many organs, including the gut and brain. Using pigs as model for preterm infants, we hypothesized that early initiation of enteral feeding stimulates both gut growth and neonatal physical activity. METHODS: In experiment 1, preterm and term pigs were fed parenteral nutrition (PN) or PN plus bovine colostrum (BC, 16-64 ml/kg/d enterally) for 5 d. In experiment 2, preterm pigs were fed PN+BC or PN+formula for 5 d. In experiment 3, preterm pigs were fed BC, formula, or human milk (HM) for 10 d. Incubator home cage activity (HCA) was quantified by continuous camera recordings. RESULTS: Preterm birth was associated with reduced intestinal weight and HCA (experiment 1), and BC or formula supplementation increased intestinal weights and HCA (experiments 1+2). Enteral BC and HM feeding increased HCA, intestinal weights, and necrotizing enteritis resistance, relative to formula (experiment 3). CONCLUSION: Preterm pigs show decreased physical activity, and the first enteral feeds diet dependently stimulate both gut growth and physical activity. The effects may arise from maturation of digestive, metabolic, and neurological functions, including gut serotonin production, by the first enteral feeds and milk bioactive factors.

Welfare Assessment in Pigs Using the Salivary Proteome.

Identifying the potential presence of stress at the pig farm is fundamental since it affects pig welfare. As a result, a reliable and straightforward tool to monitor stress could record the welfare status of the animals. Although numerous methods to assess the welfare of pigs have been developed in the past, no gold standard has been established yet. Recently, the value of saliva as a tool to identify chronic stress in piglets was explored, as it can be collected fast and non-invasively. Since the protein composition, i.e., the proteome of porcine saliva, responds to stress, the affected proteins could be used as salivary stress biomarkers. The present review first defines stress and its relationship with welfare. Next, the porcine gland-specific salivary proteome is characterized. Finally, six potential salivary biomarkers for stress are proposed, i.e., odorant-binding protein, vomeromodulin-like protein, chitinase, lipocalin-1, long palate lung and nasal epithelium protein, and alpha-2-HS-glycoprotein.

A workflow for automatic, high precision livestock diagnostic screening of locomotor kinematics.

Locomotor kinematics have been challenging inputs for automated diagnostic screening of livestock. Locomotion is a highly variable behavior, and influenced by subject characteristics (e.g., body mass, size, age, disease). We assemble a set of methods from different scientific disciplines, composing an automatic, high through-put workflow which can disentangle behavioral complexity and generate precise individual indicators of non-normal behavior for application in diagnostics and research. For this study, piglets (Sus domesticus) were filmed from lateral perspective during their first 10 h of life, an age at which maturation is quick and body mass and size have major consequences for survival. We then apply deep learning methods for point digitization, calculate joint angle profiles, and apply information-preserving transformations to retrieve a multivariate kinematic data set. We train probabilistic models to infer subject characteristics from kinematics. Model accuracy was validated for strides from piglets of normal birth weight (i.e., the category it was trained on), but the models infer the body mass and size of low birth weight (LBW) piglets (which were left out of training, out-of-sample inference) to be "normal." The age of some (but not all) low birth weight individuals was underestimated, indicating developmental delay. Such individuals could be identified automatically, inspected, and treated accordingly. This workflow has potential for automatic, precise screening in livestock management.

Ontogeny of CYP3A and UGT activity in preterm piglets: a translational model for drug metabolism in preterm newborns.

Despite considerable progress in understanding drug metabolism in the human pediatric population, data remains scarce in preterm neonates. Improving our knowledge of the ADME properties in this vulnerable age group is of utmost importance to avoid suboptimal dosing, which may lead to adverse drug reactions. The juvenile (mini)pig is a representative model for hepatic drug metabolism in human neonates and infants, especially phase I reactions. However, the effect of prematurity on the onset of hepatic phase I and phase II enzyme activity has yet to be investigated in this animal model. Therefore, the aim of this study was to assess the ontogeny of CYP3A and UGT enzyme activity in the liver of preterm (gestational day 105-107) and term-born (gestational day 115-117) domestic piglets. In addition, the ontogeny pattern between the preterm and term group was compared to examine whether postconceptional or postnatal age affects the onset of enzyme activity. The following age groups were included: preterm postnatal day (PND) 0 (n = 10), PND 5 (n = 10), PND 11 (n = 8), PND 26 (n = 10) and term PND 0 (n = 10), PND 5 (n = 10), PND 11 (n = 8), PND 19 (n = 18) and PND 26 (n = 10). Liver microsomes were extracted, and the metabolism of CYP3A and UGT-specific substrates assessed enzyme activity. Preterm CYP3A activity was only detectable at PND 26, whereas term CYP3A activity showed a gradual postnatal increase from PND 11 onwards. UGT activity gradually increased between PND 0 and PND 26 in preterm and term-born piglets, albeit, being systematically lower in the preterm group. Thus, postconceptional age is suggested as the main driver affecting porcine CYP3A and UGT enzyme ontogeny. These data are a valuable step forward in the characterization of the preterm piglet as a translational model for hepatic drug metabolism in the preterm human neonate.

Early Development of Locomotion in the Term Piglet Model: Does Size Matter?

Intrauterine undernutrition in humans typically results in low birth weight ([small for gestational age] SGA) and delayed postnatal neuromotor maturation. Since SGA and intrauterine growth retardation are also common in domestic pigs, piglets are premised as models to study delayed motor development. Applied to the locomotor paradigm, however, questions emerge: (i) how to map the developmental time scale of the precocial model onto the altricial target species and (ii) how to distinguish size from maturation effects? Gait data were collected at self-selected voluntary walking speed during early development (0-96 hours postpartum; pp) for SGA- and normal ([appropriate for gestational age] AGA) piglets. Dimensionless spatiotemporal gait characteristics (according to dynamic similarity) become invariant already after 4 hours pp, suggesting rapid postnatal neuromotor maturation. Moreover, dimensionless gait data are largely identical for SGA- and AGA-siblings, indicating that primarily size effects explain absolute locomotor differences. This is further supported by (i) normalized force-generating capacity of limb muscles, (ii) joint kinematics (<10 hours pp), and (iii) normalized ground reaction forces (<5 days pp) being indifferent between SGA- and AGA- piglets. Furthermore, predictive modeling based on limb joint kinematics is unable to discern the majority of SGA- from AGA-piglets (<10 hours pp). All this leads to the conclusion that, although smaller than the AGA piglets in absolute terms, SGA-piglets mature (neuromechanically speaking) just like, and equally fast as their AGA littermates. Yet, it remains a fact that early SGA piglets are reported to be less mobile, less vital, and less competitive than their AGA siblings (even often die before day 3 pp). This conspicuous difference likely results from the energy level (blood glucose and glycogen) and its mobilization being considerably different between the piglet categories during early development.

Platelet Activation by Antisense Oligonucleotides (ASOs) in the Göttingen Minipig, including an Evaluation of Glycoprotein VI (GPVI) and Platelet Factor 4 (PF4) Ontogeny.

Antisense oligonucleotide (ASO) is a therapeutic modality that enables selective modulation of undruggable protein targets. However, dose- and sequence-dependent platelet count reductions have been reported in nonclinical studies and clinical trials. The adult Göttingen minipig is an acknowledged nonclinical model for ASO safety testing, and the juvenile Göttingen minipig has been recently proposed for the safety testing of pediatric medicines. This study assessed the effects of various ASO sequences and modifications on Göttingen minipig platelets using in vitro platelet activation and aggregometry assays. The underlying mechanism was investigated further to characterize this animal model for ASO safety testing. In addition, the protein abundance of glycoprotein VI (GPVI) and platelet factor 4 (PF4) was investigated in the adult and juvenile minipigs. Our data on direct platelet activation and aggregation by ASOs in adult minipigs are remarkably comparable to human data. Additionally, PS ASOs bind to platelet collagen receptor GPVI and directly activate minipig platelets in vitro, mirroring the findings in human blood samples. This further corroborates the use of the Göttingen minipig for ASO safety testing. Moreover, the differential abundance of GPVI and PF4 in minipigs provides insight into the influence of ontogeny in potential ASO-induced thrombocytopenia in pediatric patients.