Injectable ultrasonic sensor for wireless monitoring of intracranial signals.

Direct and precise monitoring of intracranial physiology holds immense importance in delineating injuries, prognostication and averting disease1. Wired clinical instruments that use percutaneous leads are accurate but are susceptible to infection, patient mobility constraints and potential surgical complications during removal2. Wireless implantable devices provide greater operational freedom but include issues such as limited detection range, poor degradation and difficulty in size reduction in the human body3. Here we present an injectable, bioresorbable and wireless metastructured hydrogel (metagel) sensor for ultrasonic monitoring of intracranial signals. The metagel sensors are cubes 2 × 2 × 2 mm3 in size that encompass both biodegradable and stimulus-responsive hydrogels and periodically aligned air columns with a specific acoustic reflection spectrum. Implanted into intracranial space with a puncture needle, the metagel deforms in response to physiological environmental changes, causing peak frequency shifts of reflected ultrasound waves that can be wirelessly measured by an external ultrasound probe. The metagel sensor can independently detect intracranial pressure, temperature, pH and flow rate, realize a detection depth of 10 cm and almost fully degrade within 18 weeks. Animal experiments on rats and pigs indicate promising multiparametric sensing performances on a par with conventional non-resorbable wired clinical benchmarks.

Recovering intestinal redox homeostasis to resolve systemic inflammation for preventing remote myocardial injury by oral fullerenes.

The unbalanced immune state is the dominant feature of myocardial injury. However, the complicated pathology of cardiovascular diseases and the unique structure of cardiac tissue lead to challenges for effective immunoregulation therapy. Here, we exploited oral fullerene nanoscavenger (OFNS) to maintain intestinal redox homeostasis to resolve systemic inflammation for effectively preventing distal myocardial injury through bidirectional communication along the heart-gut immune axis. Observably, OFNS regulated redox microenvironment to repair cellular injury and reduce inflammation in vitro. Subsequently, OFNS prevented myocardial injury by regulating intestinal redox homeostasis and recovering epithelium barrier integrity in vivo. Based on the profiles of transcriptomics and proteomics, we demonstrated that OFNS balanced intestinal and systemic immune homeostasis for remote cardioprotection. Of note, we applied this principle to intervene myocardial infarction in mice and mini-pigs. These findings highlight that locally addressing intestinal redox to inhibit systemic inflammation could be a potent strategy for resolving remote tissue injury.

Integrated analysis of lncRNA, miRNA and mRNA expression profiles reveals regulatory pathways associated with pig testis function.

Long noncoding RNA (lncRNA) and microRNA (miRNA) are known to play pivotal roles in mammalian testicular function and spermatogenesis. However, their impact on porcine male reproduction has yet to be well unraveled. Here, we sequenced and identified lncRNA and miRNA expressed in the testes of Chinese indigenous Banna mini-pig inbred line (BMI) and introduced Western Duroc (DU) and Large White (LW) pigs. By pairwise comparison (BMI vs DU, BMI vs LW, and DU vs LW), we found the gene expression differences in the testes between Chinese local pigs and introduced Western commercial breeds were more striking than those between introduced commercial breeds. Furthermore, we found 1622 co-differentially expressed genes (co-DEGs), 122 co-differentially expressed lncRNAs (co-DELs), 39 co-differentially expressed miRNAs (co-DEMs) in BMI vs introduced commercial breeds (DU and LW). Functional analysis revealed that these co-DEGs and co-DELs/co-DEMs target genes were enriched in male sexual function pathways, including MAPK, AMPK, TGF-ß/Smad, Hippo, NF-kappa B, and PI3K/Akt signaling pathways. Additionally, we established 10,536 lncRNA-mRNA, 11,248 miRNA-mRNA pairs, and 62 ceRNA (lncRNA-miRNA-mRNA) networks. The ssc-miR-1343 had the most interactive factors in the ceRNA network, including 20 mRNAs and 3 lncRNAs, consisting of 56 ceRNA pairs. These factors played extremely important roles in the regulation of testis function as key nodes in the interactive regulatory network. Our results provide insight into the functional roles of lncRNAs and miRNAs in porcine testis and offer a valuable resource for understanding the differences between Chinese indigenous and introduced Western pigs.

Convergent dwarfism consequences of minipigs under independent artificial selections.

BACKGROUND: Currently, diverse minipigs have acquired a common dwarfism phenotype through independent artificial selections. Characterizing the population and genetic diversity in minipigs is important to unveil genetic mechanisms regulating their body sizes and effects of independent artificial selections on those genetic mechanisms. However, full understanding for the genetic mechanisms and phenotypic consequences in minipigs still lag behind. RESULTS: Here, using whole genome sequencing data of 41 pig breeds, including eight minipigs, we identified a large genomic diversity in a minipig population compared to other pig populations in terms of population structure, demographic signatures, and selective signatures. Selective signatures reveal diverse biological mechanisms related to body size in minipigs. We also found evidence for neural development mechanism as a minipig-specific body size regulator. Interestingly, selection signatures within those mechanisms containing neural development are also highly different among minipig breeds. Despite those large genetic variances, PLAG1, CHM, and ESR1 are candidate key genes regulating body size which experience different differentiation directions in different pig populations. CONCLUSIONS: These findings present large variances of genetic structures, demographic signatures, and selective signatures in the minipig population. They also highlight how different artificial selections with large genomic diversity have shaped the convergent dwarfism.

Genomic evidence for the suitability of Göttingen Minipigs with a rare seizure phenotype as a model for human epilepsy.

Epilepsy is a complex genetic disorder that affects about 2% of the global population. Although the frequency and severity of epileptic seizures can be reduced by a range of pharmacological interventions, there are no disease-modifying treatments for epilepsy. The development of new and more effective drugs is hindered by a lack of suitable animal models. Available rodent models may not recapitulate all key aspects of the disease. Spontaneous epileptic convulsions were observed in few Göttingen Minipigs (GMPs), which may provide a valuable alternative animal model for the characterisation of epilepsy-type diseases and for testing new treatments. We have characterised affected GMPs at the genome level and have taken advantage of primary fibroblast cultures to validate the functional impact of fixed genetic variants on the transcriptome level. We found numerous genes connected to calcium metabolism that have not been associated with epilepsy before, such as ADORA2B, CAMK1D, ITPKB, MCOLN2, MYLK, NFATC3, PDGFD, and PHKB. Our results have identified two transcription factor genes, EGR3 and HOXB6, as potential key regulators of CACNA1H, which was previously linked to epilepsy-type disorders in humans. Our findings provide the first set of conclusive results to support the use of affected subsets of GMPs as an alternative and more reliable model system to study human epilepsy. Further neurological and pharmacological validation of the suitability of GMPs as an epilepsy model is therefore warranted.

Biomechanical stress regulates mammalian tooth replacement via the integrin ß1-RUNX2-Wnt pathway.

Renewal of integumentary organs occurs cyclically throughout an organism's lifetime, but the mechanism that initiates each cycle remains largely unknown. In a miniature pig model of tooth development that resembles tooth development in humans, the permanent tooth did not begin transitioning from the resting to the initiation stage until the deciduous tooth began to erupt. This eruption released the accumulated mechanical stress inside the mandible. Mechanical stress prevented permanent tooth development by regulating expression and activity of the integrin ß1-ERK1-RUNX2 axis in the surrounding mesenchyme. We observed similar molecular expression patterns in human tooth germs. Importantly, the release of biomechanical stress induced downregulation of RUNX2-wingless/integrated (Wnt) signaling in the mesenchyme between the deciduous and permanent tooth and upregulation of Wnt signaling in the epithelium of the permanent tooth, triggering initiation of its development. Consequently, our findings identified biomechanical stress-associated Wnt modulation as a critical initiator of organ renewal, possibly shedding light on the mechanisms of integumentary organ regeneration.

Choline-deficient, high-fat diet-induced MASH in Göttingen Minipigs: characterization and effects of a chow reversal period.

The prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) and metabolic dysfunction-associated steatohepatitis (MASH) is increasing, and translational animal models are needed to develop novel treatments for this disease. The physiology and metabolism of pigs have a relatively high resemblance to humans, and the present study aimed to characterize choline-deficient and high-fat diet (CDAHFD)-fed Göttingen Minipigs as a novel animal model of MASLD/MASH. Göttingen Minipigs were fed CDAHFD for up to 5 mo, and the phenotype was investigated by the analysis of plasma parameters and repeated collection of liver biopsies. Furthermore, changes in hepatic gene expression during the experiment were explored by RNA sequencing. For a subset of the minipigs, the diet was changed from CDAHFD back to chow to investigate whether the liver pathology was reversible. Göttingen Minipigs on CDAHFD gained body weight, and plasma levels of cholesterol, AST, ALT, ALP, and GGT were increased. CDAHFD-fed minipigs developed hepatic steatosis, inflammation, and fibrosis, which in 5 of 16 animals progressed to cirrhosis. During an 11-wk chow reversal period, steatosis regressed, while fibrosis persisted. Regarding inflammation, the findings were less clear, depending on the type of readout. MASH Human Proximity Scoring (combined evaluation of transcriptional, phenotypic, and histopathological parameters) showed that CDAHFD-fed Göttingen Minipigs resemble human MASLD/MASH better than most rodent models. In conclusion, CDAHFD-fed minipigs develop a MASH-like phenotype, which, in several aspects, resembles the changes observed in human patients with MASLD/MASH. Furthermore, repeated collection of liver biopsies allows detailed characterization of histopathological changes over time in individual animals.NEW & NOTEWORTHY The physiology and metabolism of pigs have a relatively high resemblance to humans. This study characterizes a new animal model of MASLD/MASH using CDAHFD-fed Göttingen Minipigs. Göttingen Minipigs fed CDAHFD gained weight and developed hepatic steatosis, inflammation, fibrosis, and cirrhosis. After an 11-wk chow-reversal period, hepatic steatosis and some inflammatory parameters reversed. Combined evaluation of phenotypic, transcriptional, and histological parameters revealed the minipig model showed a higher resemblance to human disease than many rodent models.

Differences in vasomotor function of mesenteric arteries between Ossabaw minipigs with predisposition to metabolic syndrome and Göttingen minipigs.

Metabolic syndrome predisposes and contributes to the development and progression of atherosclerosis. The minipig strain "Ossabaw" is characterized by a predisposition to develop metabolic syndrome. We compared vasomotor function in Ossabaw minipigs before they developed their diseased phenotype to that of Göttingen minipigs without such genetic predisposition. Mesenteric arteries of adult Ossabaw and Göttingen minipigs were dissected postmortem and mounted on a myograph for isometric force measurements. Maximal vasoconstriction to potassium chloride (KClmax) was induced. Cumulative concentration-response curves were determined in response to norepinephrine. Endothelium-dependent (with carbachol) and endothelium-independent (with nitroprusside) vasodilation were analyzed after preconstriction by norepinephrine. In a bioinformatic analysis, variants/altered base pairs within genes associated with cardiovascular disease were analyzed. KClmax was similar between the minipig strains (15.6 ± 6.7 vs. 14.1 ± 3.4 ΔmN). Vasoconstriction in response to norepinephrine was more pronounced in Ossabaw than in Göttingen minipigs (increase of force to 143 ± 48 vs. 108 ± 38% of KClmax). Endothelium-dependent and endothelium-independent vasodilation were less pronounced in Ossabaw than in Göttingen minipigs (decrease of force to 46.4 ± 29.6 vs. 16.0 ± 18.4% and to 36.7 ± 25.2 vs. 2.3 ± 3.7% of norepinephrine-induced preconstriction). Vasomotor function was not different between the sexes. More altered base pairs/variants were identified in Ossabaw than in Göttingen minipigs for the exon encoding adrenoceptor-α1A. Vasomotor function in lean Ossabaw minipigs is shifted toward vasoconstriction and away from vasodilation in comparison with Göttingen minipigs, suggesting a genetic predisposition for vascular dysfunction and atherosclerosis in Ossabaw minipigs. Thus, Ossabaw minipigs may be a better model for human cardiovascular disease than Göttingen minipigs.NEW & NOTEWORTHY Animal models with a predisposition to metabolic syndrome and atherosclerosis are attracting growing interest for translational research, as they may better mimic the variability of patients with cardiovascular disease. In Ossabaw minipigs, with a polygenic predisposition to metabolic syndrome, but without the diseased phenotype, vasoconstriction is more and vasodilation is less pronounced in mesenteric arteries than in Göttingen minipigs. Ossabaw minipigs may be a more suitable model of human cardiovascular disease.

A Photo-induced Cross-Linking Enhanced A and B Combined Multi-Functional Spray Hydrogel Instantly Protects and Promotes of Irregular Dynamic Wound Healing.

Wounds in harsh environments can face long-term inflammation and persistent infection, which can slow healing. Wound spray is a product that can be rapidly applied to large and irregularly dynamic wounds, and can quickly form a protective film in situ to inhibit external environmental infection. In this study, a biodegradable A and B combined multi-functional spray hydrogel is developed with methacrylate-modified chitosan (CSMA1st) and ferulic acid (FA) as type A raw materials and oxidized Bletilla striata polysaccharide (OBSP) as type B raw materials. The precursor CSMA1st-FA/OBSP (CSOB-FA1st) hydrogel is formed by the self-cross-linking of dynamic Schiff base bonds, the CSMA-FA/OBSP (CSOB-FA) hydrogel is formed quickly after UV-vis light, so that the hydrogel fits with the wound. Rapid spraying and curing provide sufficient flexibility and rapidity for wounds and the hydrogel has good injectability, adhesive, and mechanical strength. In rats and miniature pigs, the A and B combined spray hydrogel can shrink wounds and promote healing of infected wounds, and promote the enrichment of fibrocyte populations. Therefore, the multifunctional spray hydrogel combined with A and B can protect irregular dynamic wounds, prevent wound infection and secondary injury, and be used for safe and effective wound treatment, which has a good prospect for development.

The porcine skin microbiome exhibits broad fungal antagonism.

The skin and its microbiome function to protect the host from pathogen colonization and environmental stressors. In this study, using the Wisconsin Miniature Swine™ model, we characterize the porcine skin fungal and bacterial microbiomes, identify bacterial isolates displaying antifungal activity, and use whole-genome sequencing to identify biosynthetic gene clusters encoding for secondary metabolites that may be responsible for the antagonistic effects on fungi. Through this comprehensive approach of paired microbiome sequencing with culturomics, we report the discovery of novel species of Corynebacterium and Rothia. Further, this study represents the first comprehensive evaluation of the porcine skin mycobiome and the evaluation of bacterial-fungal interactions on this surface. Several diverse bacterial isolates exhibit potent antifungal properties against opportunistic fungal pathogens in vitro. Genomic analysis of inhibitory species revealed a diverse repertoire of uncharacterized biosynthetic gene clusters suggesting a reservoir of novel chemical and biological diversity. Collectively, the porcine skin microbiome represents a potential unique source of novel antifungals.

Isolation of an Ecotropic Porcine Endogenous Retrovirus PERV-C from a Yucatan SLAD/D Inbred Miniature Swine.

Xenotransplantation may compensate the limited number of human allografts for transplantation using pigs as organ donors. Porcine endogenous retroviruses inherit infectious potential if pig cells, tissues, or organs were transplanted to immunosuppressed human recipients. Particularly, ecotropic PERV-C that could recombine with PERV-A to highly replication-competent human-tropic PERV-A/C should be excluded from pig breeds designed for xenotransplantation. Because of their low proviral background, SLAD/D (SLA, swine leukocyte antigen) haplotype pigs are potential candidates as organ donors as they do not bear replication-competent PERV-A and -B, even if they carry PERV-C. In this work, we characterized their PERV-C background isolating a full-length PERV-C proviral clone number 561 from a SLAD/D haplotype pig genome displayed in a bacteriophage lambda library. The provirus truncated in env due to cloning in lambda was complemented by PCR, and the recombinants were functionally characterized, confirming an increased infectivity in vitro compared to other PERV-C. Recombinant clone PERV-C(561) was chromosomally mapped by its 5'-proviral flanking sequences. Full-length PCR using 5'-and 3'-flanking primers specific to the PERV-C(561) locus verified that this specific SLAD/D haplotype pig harbors at least one full-length PERV-C provirus. The chromosomal location is different from that of the previously described PERV-C(1312) provirus, which was derived from the porcine cell-line MAX-T. The sequence data presented here provide further knowledge about PERV-C infectivity and contribute to targeted knockout in order to generate PERV-C-free founder animals. IMPORTANCE Yucatan SLAD/D haplotype miniature swine are candidates as organ donors for xenotransplantation. A full-length replication-competent PERV-C provirus was characterized. The provirus was chromosomally mapped in the pig genome. In vitro, the virus showed increased infectivity compared to other functional PERV-C isolates. Data may be used for targeted knockout to generate PERV-C free founder animals.

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.

Protective role of arachidonic acid against diabetic myocardial ischemic injury: a translational study of pigs, rats, and humans.

AIM: Patients with diabetes mellitus have poor prognosis after myocardial ischemic injury. However, the mechanism is unclear and there are no related therapies. We aimed to identify regulators of diabetic myocardial ischemic injury. METHODS AND RESULTS: Mass spectrometry-based, non-targeted metabolomic approach was used to profile coronary sinus blood from diabetic and non-diabetic Bama-mini pigs at 0.5-h post coronary artery ligation. Six metabolites had a |log2 (Fold Change)|> 1.3. Among them, the most changed is arachidonic acid (AA), levels of which were 32 times lower in diabetic pigs than in non-diabetic pigs. The AA-derived products, PGI2 and 6-keto-PGF1α, were also significantly reduced. AA treatment of cultured cardiomyocytes protected against cell death by 30% at 48 h of high glucose and oxygen deprivation, which coincided with increased mitophagic activity (as indicated by increased LC3II/LC3I, decreased p62 and increased parkin & PINK1), improved mitochondrial renewal (upregulation of Drp1 and FIS1), reduced ROS generation and increased ATP production. These cardioprotective effects were abolished by PINK1(a crucial mitophagy protein) knockdown or the autophagy inhibitor 3-Methyladenine. The protective effect of AA was also inhibited by indomethacin and Cay10441, a prostacyclin receptor antagonist. Furthermore, diabetic Sprague Dawley rats were subjected to coronary ligation for 40 min and AA treatment (10 mg/day per animal gavaged) decreased myocardial infarct size, cell apoptosis index, inflammatory cytokines and improved heart function. Scanning electron microscopy showed more intact mitochondria in the border zone of infarcted myocardium in AA treated rats. Lastly, diabetic patients after myocardial infarction had lower plasma levels of AA and 6-keto-PGF1α and reduced cardiac ejection fraction, compared with non-diabetic patients after myocardial infarction. Plasma AA level was inversely correlated with fasting blood glucose. CONCLUSIONS: AA protects against diabetic ischemic myocardial damage by promoting mitochondrial autophagy and renewal, which is related to AA derived PGI2 signaling. AA may represent a new strategy to treat diabetic myocardial ischemic injury.

Evaluation of dermal toxicity and toxicokinetics of povidone­iodine in Göttingen minipigs.

BACKGROUND: Povidone­iodine (PVP-I) is an effective and commonly used broad-spectrum antiseptic; limited information exists around its long-term safety and impact on endocrine disruption. We assessed the dermal toxicity and toxicokinetics following a once-daily application of 7.5% (w/v) and 10% (w/v) PVP-I in Göttingen Minipigs® for up to 39 weeks. METHODS: An in vivo study was conducted in male (n = 27) and female (n = 27) minipigs. Animals were randomized into untreated control, 7.5% and 10% PVP-I, and matching vehicle treatment groups. Animals were assessed for general in-life measurements, including skin irritation and organ weights. Serum samples were analyzed for PVP, total iodine, triiodothyronine [T3], thyroxine [T4], thyroid stimulating hormone [TSH], and toxicokinetic parameters. RESULTS: Neither 7.5% nor 10% PVP-I affected general in-life measurements. Increased mean thyroid gland absolute weights were noted with 7.5% and 10% PVP-I. Serum levels of PVP, T3, T4, and TSH in the 7.5% and 10% PVP-I treatment group animals were similar to those in vehicle treatment group animals. Mean total serum iodine concentration was 52- and 13-fold higher with 7.5% and 10% PVP-I, respectively, vs respective vehicle treatments. There was no dose-dependent increase in mean maximum serum concentration and area under the curve from 0 to 24 h for PVP, T3, T4, and TSH, nor accumulation of PVP, T3, T4, or TSH in the study. CONCLUSION: Once-daily dermal application of 7.5% and 10% PVP-I for up to 39 weeks was safe and well tolerated in Göttingen Minipigs® and was not associated with skin irritation, thyroid dysfunction, or endocrine disruption. As the anatomy and physiology of the minipig skin closely resembles that of human skin, the findings of this study suggest that 7.5% and 10% PVP-I may be translated into antimicrobial benefits for humans without the risk of endocrine disruption.

Mono- and multimeric PSMA-targeting small molecule-thorium-227 conjugates for optimized efficacy and biodistribution in preclinical models.

PURPOSE: PSMA (prostate-specific membrane antigen) is highly expressed on prostate cancer (PrCa) cells and extensively used as a homing target for PrCa treatment. Most prominently, PSMA-targeting conjugate PSMA-617, carrying a DOTA chelator and labeled with therapeutic radionuclides like beta-emitting lutetium-177 or alpha-emitting actinium-225, has shown clinical activity in PrCa patients. We sought to develop PSMA-targeting small molecule (SMOL) conjugates that show high uptake in PSMA-expressing tumors and fast clearance, and can easily be labeled with the alpha emitter thorium-227 (half-life 18.7 days). METHODS: A novel linker motif with improved competition against 3H-PSMA-617 on PSMA-expressing LNCaP cells was identified. A 2,3-hydroxypyridinone chelator modified with carboxyl groups (carboxy-HOPO) with increased hydrophilicity and robust labeling with thorium-227 was developed and allowed the synthesis of mono-, di-, tri-, and tetrameric conjugates. The resulting monomeric and multimeric PSMA SMOL-TTCs (targeted thorium conjugate) were evaluated for cellular binding, internalization, and antiproliferative activity. The in vivo antitumor efficacy of the PSMA SMOL-TTCs was determined in ST1273 and KUCaP-1 PrCa models in mice, and their biodistribution was assessed in cynomolgus monkeys, minipigs, and mice. RESULTS: The monomeric and multimeric PSMA SMOL conjugates were readily labeled with thorium-227 at room temperature and possessed high stability and good binding, internalization, and antiproliferative activity in vitro. In vivo, the monomeric, dimeric, and trimeric PSMA SMOL-TTCs showed fast clearance, potent antitumor efficacy, and high uptake and retention in prostate tumors in mice. No major uptake or retention in other organs was observed beyond kidneys. Low uptake of free thorium-227 into bone confirmed high complex stability in vivo. Salivary gland uptake remained inconclusive as mini pigs were devalidated as a relevant model and imaging controls failed in cynomolgus monkeys. CONCLUSION: Monomeric and multimeric PSMA SMOL-TTCs show high tumor uptake and fast clearance in preclinical models and warrant further therapeutic exploration.

Protein Ingredient Quality within Infant Formulas Impacts Plasma Amino Acid Concentrations in Neonatal Minipiglets.

BACKGROUND: Infant formulas (IFs), the only adequate substitute to human milk, are complex matrices that require numerous ingredients and processing steps that may impact protein digestion and subsequent amino acid (AA) absorption. OBJECTIVES: The objective was to understand the impact of the protein ingredient quality within IFs on postprandial plasma AA profiles. METHODS: Four isonitrogenous and isocaloric IFs were produced at a semi-industrial scale using whey proteins from different origins (cheese compared with ideal whey) and denaturation levels (IF-A, -B, -C), and caseins with different supramolecular organizations (IF-C, -D). Ten Yucatan minipiglets (12- to 27-d-old) were used as a human infant model and received each IF for 3 d according to a Williams Latin square followed by a 2-d wash-out period. Jugular plasma was regularly sampled from 10 min preprandial to 4 h postprandial on the third day to measure free AAs, urea, insulin, and glucose concentrations. Data were statistically analyzed using a mixed linear model with diet (IFs), time, and sex as fixed factors and piglet as random factor. RESULTS: IFs made with cheese whey (IF-A and -B) elicited significantly higher plasma total and essential AA concentrations than IFs made with ideal whey (IF-C and -D), regardless of the pre- and postprandial times. Most of the differences observed postprandially were explained by AA homeostasis modifications. IFs based on cheese whey induced an increased plasma concentration of Thr due to both a higher Thr content in these IFs and a Thr-limiting degrading capability in piglets. The use of a nonmicellar casein ingredient led to reduced plasma content of AA catabolism markers (IF-D compared with IF-C). CONCLUSIONS: Overall, our results highlight the importance of the protein ingredient quality (composition and structure) within IFs on neonatal plasma AA profiles, which may further impact infant protein metabolism.

Dietary Methionine Enhances Portal Appearance of Guanidinoacetate and Synthesis of Creatine in Yucatan Miniature Piglets.

BACKGROUND: Creatine plays a significant role in energy metabolism and positively impacts anaerobic energy capacity, muscle mass, and physical performance. Endogenous creatine synthesis requires guanidinoacetic acid (GAA) and methionine. GAA can be an alternative to creatine supplements and has been tested as a beneficial feed additive in the animal industry. When pigs are fed GAA with excess methionine, creatine is synthesized without feedback regulation. In contrast, when dietary methionine is limited, creatine synthesis is limited, yet, GAA does not accumulate in plasma, urine, or liver. OBJECTIVE: We hypothesized that portal GAA appearance requires adequate dietary methionine. METHODS: Yucatan miniature piglets (17-21 d old; n = 20) were given a 4 h duodenal infusion of complete elemental diets with supplemental GAA plus 1 of 4 methionine concentrations representing either 20%, 80%, 140%, or 200% of the dietary methionine requirement. Arterial and portal blood metabolites were measured along with blood flow to determine mass balance across the gut. [3H-methyl] methionine was infused to measure the methionine incorporation rate into creatine. RESULTS: GAA balance across the gut was highest in the 200% methionine group, indicating excess dietary methionine enhanced GAA absorption. Creatine synthesis in the liver and jejunum was higher with higher concentrations of methionine, emphasizing that the transmethylation of GAA to creatine depends on sufficient dietary methionine. Hepatic GAA concentration was higher in the 20% methionine group, suggesting low dietary methionine limited GAA conversion to creatine, which led to GAA accumulation in the liver. CONCLUSIONS: GAA absorption and conversion to creatine require a sufficient amount of methionine, and the supplementation strategies should accommodate this interaction.

Cross-species transmission and histopathological variation in specific-pathogen-free minipigs infected with different hepatitis E virus strains.

Hepatitis E virus (HEV) is a major cause of viral hepatitis worldwide. Pigs are the natural host of HEV genotype 3 and the main reservoir of HEV. As the host range of HEV genotype 3 expands, the possibility that HEV from various species can be transmitted to humans via pigs is increasing. We investigated the potential cross-species transmission of HEV by infecting minipigs with swine HEV (swHEV), rabbit HEV (rbHEV), and human HEV (huHEV) and examining their histopathological characteristics and distribution in various organs. Fifteen specific-pathogen-free Yucatan minipigs were infected with swHEV, rbHEV, huHEV, or a mock control. In the present study, we analysed faecal shedding, viremia, and serological parameters over a seven-week period. Our results indicated that swHEV exhibited more robust shedding and viremia than non-swHEVs. Only swHEV affected the serological parameters, suggesting strain-specific differences. Histopathological examination revealed distinct patterns in the liver, pancreas, intestine, and lymphoid tissues after infection with each HEV strain. Notably, all three HEVs induced histopathological changes in the pancreas, supporting the association of HEVs with acute pancreatitis. Our results also identified skeletal muscle as a site of HEV antigen presence, suggesting a potential link to myositis. In conclusion, this study provides valuable insights into the infection dynamics of different HEV strains in minipigs, emphasizing the strain-specific variations in virological, serological, and histological parameters. The observed differences in infection kinetics and tissue tropism will contribute to our understanding of HEV pathogenesis and the potential for cross-species transmission.

Cardiovascular magnetic resonance characterization of myocardial tissue injury in a miniature swine model of cancer therapy-related cardiovascular toxicity.

BACKGROUND: Left ventricular ejection fraction (LVEF) is the most commonly clinically used imaging parameter for assessing cancer therapy-related cardiac dysfunction (CTRCD). However, LVEF declines may occur late, after substantial injury. This study sought to investigate cardiovascular magnetic resonance (CMR) imaging markers of subclinical cardiac injury in a miniature swine model. METHODS: Female Yucatan miniature swine (n = 14) received doxorubicin (2 mg/kg) every 3 weeks for 4 cycles. CMR, including cine, tissue characterization via T1 and T2 mapping, and late gadolinium enhancement (LGE) were performed on the same day as doxorubicin administration and 3 weeks after the final chemotherapy cycle. In addition, magnetic resonance spectroscopy (MRS) was performed during the 3 weeks after the final chemotherapy in 7 pigs. A single CMR and MRS exam were also performed in 3 Yucatan miniature swine that were age- and weight-matched to the final imaging exam of the doxorubicin-treated swine to serve as controls. CTRCD was defined as histological early morphologic changes, including cytoplasmic vacuolization and myofibrillar loss of myocytes, based on post-mortem analysis of humanely euthanized pigs after the final CMR exam. RESULTS: Of 13 swine completing 5 serial CMR scans, 10 (77%) had histological evidence of CTRCD. Three animals had neither histological evidence nor changes in LVEF from baseline. No absolute LVEF <40% or LGE was observed. Native T1, extracellular volume (ECV), and T2 at 12 weeks were significantly higher in swine with CTRCD than those without CTRCD (1178 ms vs. 1134 ms, p = 0.002, 27.4% vs. 24.5%, p = 0.03, and 38.1 ms vs. 36.4 ms, p = 0.02, respectively). There were no significant changes in strain parameters. The temporal trajectories in native T1, ECV, and T2 in swine with CTRCD showed similar and statistically significant increases. At the same time, there were no differences in their temporal changes between those with and without CTRCD. MRS myocardial triglyceride content substantially differed among controls, swine with and without CTRCD (0.89%, 0.30%, 0.54%, respectively, analysis of variance, p = 0.01), and associated with the severity of histological findings and incidence of vacuolated cardiomyocytes. CONCLUSION: Serial CMR imaging alone has a limited ability to detect histologic CTRCD beyond LVEF. Integrating MRS myocardial triglyceride content may be useful for detection of early potential CTRCD.