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Current methods of staging liver fibrosis have notable limitations. We investigated the utility of PET in staging liver fibrosis by correlating liver uptake of (68)Ga-labeled fibroblast activation protein inhibitor (FAPI) with histology in a human-sized swine model. Methods: Five pigs underwent baseline (68)Ga-FAPI-46 ((68)Ga-FAPI) PET/MRI and liver biopsy, followed by liver parenchymal embolization, 8 wk of oral alcohol intake, endpoint (68)Ga-FAPI PET/MRI, and necropsy. Regions of interest were drawn on baseline and endpoint PET images, and SUV(mean) was recorded. At the endpoint, liver sections corresponding to regions of interest were identified and cut out. Fibrosis was histologically evaluated using a modified METAVIR score for swine liver and quantitatively using collagen proportionate area (CPA). Box-and-whisker plots and linear regression were used to correlate SUV(mean) with METAVIR score and CPA, respectively. Results: Liver (68)Ga-FAPI uptake strongly correlated with CPA (r = 0.89, P < 0.001). (68)Ga-FAPI uptake was significantly and progressively higher across F2 and F3/F4 fibrosis stages, with a respective median SUV(mean) of 2.9 (interquartile range [IQR], 2.7–3.8) and 7.6 (IQR, 6.7–10.2) (P < 0.001). There was no significant difference between (68)Ga-FAPI uptake of baseline liver and endpoint liver sections staged as F0/F1, with a respective median SUV(mean) of 1.7 (IQR, 1.3–2.0) and 1.7 (IQR, 1.5–1.8) (P = 0.338). Conclusion: The strong correlation between liver (68)Ga-FAPI uptake and the histologic stage of liver fibrosis suggests that (68)Ga-FAPI PET can play an impactful role in noninvasive staging of liver fibrosis, pending validation in patients.

DNA-methylation profiles have been used successfully to develop highly accurate biomarkers of age, epigenetic clocks, for many species. Using a custom methylation array, we generated DNA methylation data from n = 238 porcine tissues including blood, bladder, frontal cortex, kidney, liver, and lung, from domestic pigs (Sus scrofa domesticus) and minipigs (Wisconsin Miniature Swine™). Samples used in this study originated from Large White X Landrace crossbred pigs, Large White X Minnesota minipig crossbred pigs, and Wisconsin Miniature Swine™. We present 4 epigenetic clocks for pigs that are distinguished by their compatibility with tissue type (pan-tissue and blood clock) and species (pig and human). Two dual-species human-pig pan-tissue clocks accurately measure chronological age and relative age, respectively. We also characterized CpGs that differ between minipigs and domestic pigs. Strikingly, several genes implicated by our epigenetic studies of minipig status overlap with genes (ADCY3, TFAP2B, SKOR1, and GPR61) implicated by genetic studies of body mass index in humans. In addition, CpGs with different levels of methylation between the two pig breeds were identified proximal to genes involved in blood LDL levels and cholesterol synthesis, of particular interest given the minipig’s increased susceptibility to cardiovascular disease compared to domestic pigs. Thus, breed-specific differences of domestic and minipigs may potentially help to identify biological mechanisms underlying weight gain and aging-associated diseases. Our porcine clocks are expected to be useful for elucidating the role of epigenetics in aging and obesity, and the testing of anti-aging interventions. Supplementary Information The online version contains supplementary material available at 10.1007/s11357-021-00439-6.

Brachial plexus injury (BPI) occurs when the brachial plexus is compressed, stretched, or avulsed. Although rodents are commonly used to study BPI, these models poorly mimic human BPI due to the discrepancy in size. The objective of this study was to compare the brachial plexus between human and Wisconsin Miniature SwineTM (WMSTM ), which are approximately the weight of an average human (68-91 kg), to determine if swine would be a suitable model for studying BPI mechanisms and treatments. To analyze the gross anatomy, WMS brachial plexuses were dissected both anteriorly and posteriorly. For histological analysis, sections from various nerves of human and WMS brachial plexuses were fixed in 2.5% glutaraldehyde, and postfixed with 2% osmium tetroxide. Subsequently paraffin sections were counter-stained with Masson's Trichrome. Gross anatomy revealed that the separation into three trunks and three cords is significantly less developed in the swine than in human. In swine, it takes the form of upper, middle, and lower systems with ventral and dorsal components. Histological evaluation of selected nerves revealed differences in nerve trunk diameters and the number of myelinated axons in the two species. The WMS had significantly fewer myelinated axons than humans in median (p = 0.0049), ulnar (p = 0.0002), and musculocutaneous nerves (p = 0.0454). The higher number of myelinated axons in these nerves for humans is expected because there is a high demand of fine motor and sensory functions in the human hand. Due to the stronger shoulder girdle muscles in WMS, the WMS suprascapular and axillary nerves were larger than in human. Overall, the WMS brachial plexus is similar in size and origin to human making them a very good model to study BPI. Future studies analyzing the effects of BPI in WMS should be conducted.

Purpose: To analyze radiation induced changes in Hounsfield units and determine their correlation with changes in perfusion and ventilation. Additionally, to compare the post-RT changes in human subjects to those measured in a swine model used to quantify perfusion changes, and validate their use as a preclinical model. Methods: A cohort of 5 Wisconsin Miniature Swine (WMS™) were studied. Additionally, 19 human subjects were recruited as part of an IRB approved clinical trial studying functional avoidance radiation therapy for lung cancer and were treated with SBRT. Imaging (a contrast enhanced dynamic perfusion CT in the swine and 4DCT in the humans) was performed prior to and post-RT. Jacobian elasticity maps were calculated on all 4DCT images. Contours were created from the isodose lines to discretize analysis into 10 Gy dose bins. B-spline deformable image registration allowed for voxel-by-voxel comparative analysis in these contours between timepoints. The WMS™ underwent a research course of 60 Gy in 5 fractions delivered locally to a target in the lung using an MRI-LINAC system. In the WMS™ subjects, the dose-bin contours were copied onto the contralateral lung, which received < 5 Gy for comparison. Changes in HU and changes in Jacobian were analyzed in these contours.Results: Statistically significant (p < 0.05) changes in the mean HU value post-RT compared to pre-RT were observed in both the human and WMS™ groups at all timepoints analyzed. The HU increased linearly with dose for both groups. Strong linear correlation was observed between the changes seen in the swine and humans (Pearson coefficient > 0.97, p< 0.05) at all timepoints. Changes seen in the swine closely modeled the changes seen in the humans at 12 months post RT (slope=0.95). Jacobian analysis showed between 30-60% of voxels were damaged post-RT. Perfusion analysis in the swine showed a statistically significant (p < 0.05) reduction in contrast inside the vasculature 3 months post-RT compared to pre-RT. The increases in contrast outside the vasculature was strongly correlated (Pearson Correlation 0.88) with the reduction in HU inside the vasculature but were not correlated with the changes in Jacobians. Conclusions: Radiation induces changes in pulmonary anatomy at 3 months post-RT, with a strong linear correlation with dose. The change in HU seen in the non-vessel lung parenchyma suggests this metric is a potential biomarker for change in perfusion. Finally, this work suggests that the WMS™ swine model is a promising pre-clinical model for analyzing radiation-induced changes in humans and poses several benefits over conventional swine models.

Candida auris proliferates and persists on the skin of patients, often leading to health care-associated infections with high mortality. Here, we describe 2 clinically relevant skin models and show that C. auris grows similarly on human and porcine skin. Additionally, we demonstrate that other Candida spp., including those with phylogenetic similarity to C. auris, do not display high growth in the skin microenvironment. These studies highlight the utility of 2 ex vivo models of C. auris colonization that allow reproducible differentiation among Candida spp., which should be a useful tool for comparison of C. auris clinical isolates and genetically mutated strains.

Elevated levels of dietary fats in westernized diets, associated with increased risk of obesity and other chronic diseases, are increasingly consumed by children in the United States. Cooking practices such as high heat frying and increased use of oxidizable sources of fats have introduced high levels of lipid oxidation products (LOPs) into these diets. The effects of these highly reactive dietary compounds on human biology are largely unstudied, especially in the gut where these compounds are likely present at higher concentrations. Given that the gut microbiome can be influenced by dietary components and then in turn have a systemic impact, we investigated the effects of consuming LOPs on gut bacterial and fungal communities and on growth and body composition during the prepubertal period in a porcine model. The presence of LOPs in the high fat diet reduced growth and body fat gain in the model. The gut microbiome was uniquely altered by both high fat and the presence of LOPs, with notable changes in the abundances of Turicibacterales, Spriochaetales, RF39, Lactobacillales and Erysipelotrichales. The mycobiome was dominated by Kazachstania, a porcine specific yeast, which was only minimally influenced by the dietary regimen. Application of machine learning identified dietary fat and LOPs as strong predictors of body fat. The genus Methanobrevibacter was the key microbial predictor of body fat. This study highlights the need for further studies on the biological effects of LOPs which have become ubiquitous in human, livestock and pet diets in developed countries.

Candida auris spreads person to person in hospitals and other healthcare facilities. The heightened capacity for C. auris to colonize skin contributes to the difficulty in eradicating this drug-resistant and deadly pathogen in nosocomial settings. Models for the study of C. auris skin colonization are critical for understanding this virulence trait. In light of the similarities between the skin properties of humans and pigs, pigs represent an ideal model for the investigation of skin-C. auris interactions. Here, we describe how to utilize porcine skin for ex vivo studies of C. auris colonization.

Purpose. To investigate indirect radiation-induced changes in airways as precursors to atelectasis post radiation therapy (RT). Methods. Three Wisconsin Miniature Swine (WMS TM ) underwent a research course of 60 Gy in 5 fractions delivered to a targeted airway/vessel in the inferior left lung. The right lung received a max point dose WA ) for each airway were investigated. Changes in ventilation were assessed using the Jacobian ratio and were measured in three different regions: the inferior left lung Results. Airways (n = 25) in the right lung for all swine showed no significant changes (p = 0.48) in Ai post-RT compared to pre-RT. Airways (n = 28) in the left lung of all swine were found to have a significant decrease (p < 0.001) in Ai post-RT compared to pre-RT, correlated (Pearson R = −0.97) with airway dose. Additionally, WA decreased significantly (p < 0.001) with airway dose. Lastly, the Jacobian ratio of the ILL (0.883) was lower than that of the SLL (0.932) and the RL (0.955). Conclusions. This work shows that for the swine analyzed, there were significant correlations between Ai and WA change with radiation dose. Additionally, there was a decrease in lung function in the regions of the lung supplied by the irradiated airways compared to the regions supplied by unirradiated airways. These results support the hypothesis that airway dose should be considered during treatment planning in order to potentially preserve functional lung and reduce lung toxicities.

Recent functional lung imaging studies have presented evidence of an “indirect effect” on perfusion damage, where regions that are unirradiated or lowly irradiated but that are supplied by highly irradiated regions observe perfusion damage post-radiation therapy (RT). The purpose of this work was to investigate this effect using a contrast-enhanced dynamic CT protocol to measure perfusion change in five novel swine subjects. A cohort of five Wisconsin Miniature Swine (WMS) were given a research course of 60 Gy in five fractions delivered locally to a vessel in the lung using an Accuray Radixact tomotherapy system with Synchrony motion tracking to increase delivery accuracy. Imaging was performed prior to delivering RT and 3 months post-RT to yield a 28–36 frame image series showing contrast flowing in and out of the vasculature. Using MIM software, contours were placed in six vessels on each animal to yield a contrast flow curve for each vessel. The contours were placed as follows: one at the point of max dose, one low-irradiated (5–20 Gy) branching from the max dose vessel, one low-irradiated (5–20 Gy) not branching from the max dose vessel, one unirradiated (

Purpose/objective(s) Daily variations in abdominal anatomy introduce significant uncertainty in estimating the maximum tolerated dose (MTD). We investigated the gastroduodenal toxicity using stereotactic MR-guided online adaptive radiotherapy (SMART) to precisely control the dose to a novel swine model that closely matches human physiology and can be maintained at 80 kg. We hypothesized for a cumulative SMART course of 50 Gy in 5 fractions to the pancreas, the duodenum MTD is significantly > 37 Gy to 0.5 cc (D0.5cc). Materials/methods A total of 18 WMSTM were included at 3 duodenal dose levels: 37 Gy (n = 12), 43.5 Gy (n = 3), and 50 Gy (n = 3) in 5 fractions. We defined MTD as: > 10% (≥2 of 12) WMSTM present with dose limiting toxicity (DLT). A DLT was defined as ≥ Grade 3 non-hematologic toxicity due to RT per NCI CTCAE v4.0. All WMSTM underwent SMART on a clinical MR-guided radiotherapy (MRgRT) LINAC, under ventilation at breath hold (BH). Plans were IMRT and prescribed 50 Gy/5 fractions to pancreatic head, while maintaining uniform dose level (37, 43.5, 50 Gy) to duodenal circumference at 3 cm inferior to pylorus sphincter, typical uniform irradiated range 3-9 cc based on duodenal diameter. Circumference approach was used to eliminate underestimation of cumulative D0.5cc. Duodenal hotspot was 1.03% of the dose level (37, 43.5, 50 Gy) to 2 cc, 1.05% to 0.1 cc. Gated BH delivery was performed with sagittal plane tracking of the duodenum at 4 frames per second. Toxicity monitoring occurred for 30 days post-RT. Fecal occult blood test (FOBT) was done during RT and toxicity monitoring. Endoscopy occurred 14 days post-RT. Gross pathology was obtained at time of necropsy, 30 days post-RT. Results For the 37 Gy level, 1 of 12 WMSTM presented with DLT-pinpoint ulcer with surface damage on gross pathology (30 days post-RT). No DLT for 2 of 3 WMSTM at the 43.5 Gy level. One WMSTM at the 43.5 Gy level perforated on endoscopy, potentially due duodenal wall degradation and unknown RT-induced DLT on gross exam of emergency necropsy. Endoscopy was eliminated for the 50 Gy level. DLT was found for 2 of 3 WMSTM at the 50 Gy level upon gross pathology at necropsy, with each subject having 2-3 visible ulcerations at level of RT. One WMSTM at the 50 Gy level had duodenal pre-ulcer erosion at necropsy. All three WMSTM at 50 Gy level had visibly damaged papilla at necropsy. Another WMSTM at the 50 Gy level had common bile duct blockage, gall bladder obstruction, and jaundice at necropsy. FOBT were negative for all 18 WMSTM at 14 days post-RT. Conclusion A duodenum D0.5cc of 37 Gy in 5 fractions was found to be safe in a swine model for a 50 Gy SMART course to the pancreas, showing the duodenal MTD is > 37 Gy - an increase from the RTOG 1112 limit (i.e., 30 Gy). Our results translate to a recommended MTD in human subjects and can potentially allow for greater tumor dose escalation and survival outcomes in future pancreatic patients. We are currently pursuing an expansion cohort to evaluate the MTD at 43.5 Gy.

DNA-methylation profiles have been used successfully to develop highly accurate biomarkers of age, epigenetic clocks, for many species. Using a custom methylation array, we generated DNA methylation data from n=238 porcine tissues including blood, bladder, frontal cortex, kidney, liver and lung, from domestic pigs (Sus scrofa domesticus) and minipigs (Wisconsin Miniature Swine™). We present 4 epigenetic clocks for pigs that are distinguished by their compatibility with tissue type (pan-tissue and blood clock) and species (pig and human). Two dual-species human-pig pan-tissue clocks accurately measure chronological age and relative age, respectively. We also characterized CpGs that differ between minipigs and domestic pigs. Strikingly, several genes implicated by our epigenetic studies of minipig status overlap with genes (ADCY3, TFAP2B, SKOR1, and GPR61) implicated by genetic studies of body mass index in humans. In addition, CpGs with different levels of methylation between the two pig breeds were identified proximal to genes involved in blood LDL levels and cholesterol synthesis, of particular interest given the minipig’s increased susceptibility to cardiovascular disease compared to domestic pigs. Thus, inbred differences of domestic and minipigs may potentially help to identify biological mechanisms underlying weight gain and aging-associated diseases. Our porcine clocks are expected to be useful for elucidating the role of epigenetics in aging and obesity, and the testing of anti-aging interventions.

Background: Brachial plexus injury (BPI) occurs when the brachial plexus is compressed, stretched, or avulsed. Although rodents are commonly used to study BPI, these models poorly mimic human BPI due to the discrepancy in size. The objective of this study was to compare the brachial plexus between human and Wisconsin Miniature SwineTM (WMSTM), which are approximately the weight of an average human (68–91 kg), to determine if swine would be a suitable model for studying BPI mechanisms and treatments. Methods: To analyze the gross anatomy, WMS brachial plexi were dissected both anteriorly and posteriorly. For histological analysis, sections from various nerves of human and WMS brachial plexi were fixed in 2.5% glutaraldehyde and then myelinated axons were labeled using 2% osmium tetroxide before being counter-stained with Masson’s Trichrome. Results: Gross anatomy revealed that the separation into 3 trunks and 3 cords is significantly less developed in the swine than in human. In swine, it takes the form of upper, middle, and lower systems with ventral and dorsal components. Histological results showed that despite the similarity in body size between the miniature swine model and humans, there was some discrepancy in nerve size and the number of the myelinated axons. The WMS had significantly fewer myelinated axons than humans in median (p = 0.0009), ulnar (p = 0.0001), and musculocutaneous nerves (p = 0.0451). The higher number of myelinated axons in these nerves for humans is expected, because there is a high demand of fine motor function in the human hand, with more motor units. Due to the stronger shoulder girdle muscles in WMS, the WMS suprascapular nerves were larger than in human. Conclusion: Overall, the WMS brachial plexus is similar in size and origin to human making them an excellent model to study BPI. Future studies analyzing the effects of BPI in WMS should be conducted.

The emerging fungal pathogen Candida auris causes invasive infections and is spreading in hospitals worldwide. Why this species exhibits the capacity to transfer efficiently among patients is unknown. Our findings reveal that C. auris forms high-burden biofilms in conditions mimicking sweat on the skin surface. These adherent biofilm communities persist in environmental conditions expected in the hospital setting. Using a pig skin model, we show that C. auris also forms high-burden biofilm structures on the skin surface. Identification of this mode of growth sheds light on how this recently described pathogen persists in hospital settings and spreads among patients., Emerging pathogen Candida auris causes nosocomial outbreaks of life-threatening invasive candidiasis. It is unclear how this species colonizes skin and spreads in health care facilities. Here, we analyzed C. auris growth in synthetic sweat medium designed to mimic axillary skin conditions. We show that C. auris demonstrates a high capacity for biofilm formation in this milieu, well beyond that observed for the most commonly isolated Candida sp., Candida albicans. The C. auris biofilms persist in environmental conditions expected in the hospital setting. To model C. auris skin colonization, we designed an ex vivo porcine skin model. We show that C. auris proliferates on porcine skin in multilayer biofilms. This capacity to thrive in skin niche conditions helps explain the propensity of C. auris to colonize skin, persist on medical devices, and rapidly spread in hospitals. These studies provide clinically relevant tools to further characterize this important growth modality. IMPORTANCE The emerging fungal pathogen Candida auris causes invasive infections and is spreading in hospitals worldwide. Why this species exhibits the capacity to transfer efficiently among patients is unknown. Our findings reveal that C. auris forms high-burden biofilms in conditions mimicking sweat on the skin surface. These adherent biofilm communities persist in environmental conditions expected in the hospital setting. Using a pig skin model, we show that C. auris also forms high-burden biofilm structures on the skin surface. Identification of this mode of growth sheds light on how this recently described pathogen persists in hospital settings and spreads among patients.

Cardiac troponin T (cTnT) regulates the Ca2+-mediated interaction between myosin thick filaments and actin thin filaments during cardiac contraction and relaxation. cTnT is released into the blood following injury, and increased serum levels of the protein are used clinically as a biomarker for myocardial infarction. Moreover, mutations in cTnT are causative in a number of familial cardiomyopathies. With the increasing use of large animal (swine) model to recapitulate human diseases, it is essential to characterize species-dependent protein sequence variants, alternative RNA splicing, and post-translational modifications (PTMs), but challenges remain due to the incomplete database and lack of validation of the predicted splicing isoforms. Herein, we integrated top-down mass spectrometry (MS) with online liquid chromatography (LC) and immunoaffinity purification to comprehensively characterize miniature swine cTnT proteoforms, including those arising from alternative RNA splicing and PTMs. A total of seven alternative splicing isoforms of cTnT were identified by LC/MS from swine left ventricular tissue, with each isoform containing un-phosphorylated and mono-phosphorylated proteoforms. The phosphorylation site was localized to Ser1 for the mono-phosphorylated proteoforms of cTnT1, 3, 4, and 6 by online MS/MS combining collisionally activated dissociation (CAD) and electron transfer dissociation (ETD). Offline MS/MS on Fourier-transform ion cyclotron resonance (FT-ICR) mass spectrometer with CAD and electron capture dissociation (ECD) was then utilized to achieve deep sequencing of mono-phosphorylated cTnT1 (35.2 kDa) with a high sequence coverage of 87%. Taken together, this study demonstrated the unique advantage of top-down MS in the comprehensive characterization of protein alternative splicing isoforms together with PTMs. Graphical Abstract ᅟ.

The relationship among diet, human health, and disease is an area of growing interest in biomarker research. Previous studies suggest that the consumption of cranberries (Vaccinium macrocarpon) could beneficially influence urinary and digestive health. The present study sought to determine if daily consumption of sweetened dried cranberries (SDC) changes the urinary proteome and fecal microbiome, as determined in a prospective sample of 10 healthy individuals. Baseline urine and fecal samples were collected from the subjects in the fasted (8–12 h) state. The subjects then consumed one serving (42 g) of SDC daily with lunch for 2 weeks. Urine and fecal samples were collected again the day after 2 weeks of SDC consumption. Orbitrap Q-Exactive mass spectrometry of urinary proteins showed that consumption of SDC resulted in changes to 22 urinary proteins. Multiplex sequencing of 16S ribosomal RNA genes in fecal samples indicated changes in relative abundance of several bacterial taxonomic units after consumption of SDC. There was a shift in the Firmicutes:Bacteroidetes ratio, increases in commensal bacteria, and decreases or the absence of bacteria associated with negative health effects. A decrease in uromodulin in all subjects and an increase in Akkermansia bacteria in most subjects were observed and warrant further investigation. Future larger clinical studies with multiomics and multitissue sampling designs are required to determine the effects of SDC consumption on nutrition and health.

Background/Aims Spine and spinal cord pathologies and associated neuropathic pain are among the most complex medical disorders to treat. While rodent models are widely used in spine and spinal cord research and have provided valuable insight into pathophysiological mechanisms, these models offer limited translatability. Thus, studies in rodent models have not led to the development of clinically effective therapies. More recently, swine has become a favored model for spine research because of the high congruency of the species to humans with respect to spine and spinal cord anatomy, vasculature, and immune responses. However, conventional breeds of swine commonly used in these studies present practical and translational hurdles due to their rapid growth toward weights well above those of humans. Methods In the current study, we evaluated the suitability of a human-sized breed of swine developed at the University of Wisconsin-Madison, the Wisconsin Miniature SwineTM (WMSTM), in the context of thoracic spine morphometry for use in research to overcome limitations of conventional swine breeds. The morphometry of thoracic vertebrae (T1-T15) of 5-6 months-old WMS was analyzed and compared to published values of human and conventional swine spines. Results The key finding of this study is that WMS spine more closely models the human spine for many of the measured vertebrae parameters, while being similar to conventional swine in respect to the other parameters. Conclusion WMS provides an improvement over conventional swine for use in translational spinal cord injury studies, particularly long-term ones, because of its slower rate of growth and its maximum growth being limited to human weight and size.

Candida auris readily colonizes skin and efficiently spreads among patients in healthcare settings worldwide. Given the capacity of this drug-resistant fungal pathogen to cause invasive disease with high mortality, hospitals frequently employ chlorhexidine bathing to reduce skin colonization. Using an ex vivo skin model, we show only a mild reduction in C. auris following chlorhexidine application. This finding helps explain why chlorhexidine bathing may have failures clinically, despite potent in vitro activity. We further show that isopropanol augments the activity of chlorhexidine against C. auris on skin. Additionally, we find both tea tree (Melaleuca alternifolia) oil and lemongrass (Cymbopogon flexuosus) oil to further enhance the activity of chlorhexidine/isopropanol for decolonization. We link this antifungal activity to individual oil components and show how some of these components act synergistically with chlorhexidine/isopropanol. Together, the studies provide strategies to improve C. auris skin decolonization through the incorporation of commonly used topical compounds.

Spinal cord injury (SCI) is a physically and psychologically devastating clinical condition. The typical treatment regimens of decompressive surgery and rehabilitation therapy still leave many patients with permanent disability. The development of new therapies and devices can be accelerated if relevant translational animal models are more effectively used in pre-clinical stages. Swine is a highly relevant model for SCI research, especially with respect to spine and spinal cord anatomy, spine vasculature, immune responses to injury, and functional assessments. Several spine injury models have recently been developed for swine and are beginning to be used to evaluate new therapies. Swine models of SCI offer tremendous advantages for efficient translation of pre-clinical discoveries and the development of new therapies and devices. Future swine models will also be enhanced by advances in gene-editing technology to further elucidate the complex pathophysiology associated with SCI and provide a means to engineer specific spinal pathologies.

Atypical teratoid/rhabdoid tumor (AT/RT) is the most common malignant CNS tumor of children below 6 months of age. The majority of AT/RT demonstrate genomic alterations in the SMARCB1 gene. There are two major hurdles in the development of safe and effective treatments for AT/RT: first, the mouse models do not fully recapitulate the disease seen in patients and their predictivity of clinical efficacy is still unproven. Second, due to a small patient population, the ability to recruit enough patients for clinical trials is challenging. Genetic studies have demonstrated that germline deletion of SMARCB1 exons 4 and 5 predisposes to AT/RT at an early age. Comparison of human, swine, and mouse SMARCB1 genes show similarities in gene and protein structure, with 100% amino acid identity between swine and human SMARCB1 isoforms. Thus, we hypothesized that germline deletion of exons 4 and 5 will predispose heterozygote swine to AT/RT development. SMARCB1+/- founder pigs are obtained using a CRISPR/Cas9 mediated gene-editing of conventional crossbred swine embryos, followed by embryo transfer into female swine surrogates. They are evaluated for clinical criteria used to diagnose AT/RT and by MRI at 6, 12, and 24 months of age, followed by histopathology and molecular analysis of the tumors as they are detected. Generating a large animal model of AT/RT would represent a breakthrough in the field from a genomic, pathophysiologic, pre-clinical and therapeutic perspective.

The 4-(dimethylamino)cinnamaldehyde (DMAC) assay is currently used to quantify proanthocyanidin (PAC) content in cranberry products. In a multi-operator/multi-day study design, a cranberry proanthocyanidin (c-PAC) standard was compared to procyanidin A2 (ProA2) dimer for accurate quantification of PAC in commercial cranberry juices, lab generated cranberry blends and cranberry powders. The c-PAC standard reflects the structural heterogeneity of cranberry PAC degree of polymerization, hydroxylation pattern and ratios of ‘A-type’ to ‘B-type’ interflavanyl bonds. Use of the c-PAC standard to quantify PAC content in cranberry samples resulted in values that were 3.6 times higher than those determined by ProA2. Overall, there was no effect (P > 0.05) of operator or day on estimation of PAC concentration. The adoption of c-PAC standard should be considered as an improvement over the use of ProA2 for accurate quantification of cranberry PAC. Improved standardization of bioactive PAC components in functional cranberry foods will aid in establishment of dosage guidelines.

Studies examining the effects of feeding lipid oxidation products (LOPs) to pigs on pork quality and storage stability have mostly focused on refrigerated storage and produced mixed results. We investigated the effects of adding yellow grease, containing commercially relevant levels of LOPs, to swine diets on quality and storage stability of ground salted pork. Twenty-four domestic pigs were divided into three study groups and fed the following diet regimens for five months: (1) Standard Diet (STD), (2) STD + yellow grease (YG, high LOPs), or (3) STD + corn oil (CO, negligible LOPs). Post-harvest carcass characteristics and the effects of frozen and refrigerated storage on color and lipid oxidation of salted pork patties were studied. While feeding of yellow grease had no impact on color, it increased the susceptibility of pork patties to lipid oxidation during storage (186% and 73% higher accumulation of LOPs in patties from pigs fed STD + YG when compared to those fed STD and STD + CO, respectively).

The Rapacz familial hypercholesterolemic (FH) swine model is well-characterized and used for studies of both spontaneous and inducible atherosclerosis but has not been used for studies of metabolic dysfunction to date. We examined whether parameters of metabolic syndrome including weight and adiposity, serum cholesterol, and glucoregulatory function could be modulated by restriction of caloric intake in the FH swine. Three groups of FH swine (n = 6 per group) were fed without restriction (AL), 80% of AL caloric intake, or 60% of AL caloric intake for 8.8 ± 0.5 mo beginning 2 wk after weaning. Caloric intake influenced the rate and magnitude of body weight gain and change in adiposity, as determined by dual-emission X-ray absorptiometry. At the conclusion of the study, pigs in the AL group reached a total least-square mean body weight of 94.2 kg and fat mass of 31.1%, whereas those fed 80% AL were 71.6 kg and 24.3% fat, and swine fed 60% AL were 46.1 kg and 14.1% fat. Serum cholesterol was greater in AL than 60% AL pigs at the end of the study. At 10 mo of age, intravenous glucose tolerance testing, performed to assess glucoregulatory function, indicated significant differences in serum glucose clearance profiles and insulin sensitivity between the AL- and 60% AL-fed swine. The AL-fed animals showed almost 5-fold lower insulin sensitivity when compared with animals fed 60% AL caloric intake. These results highlight the value of the FH swine model to study metabolic dysfunction due to changes in caloric intake.

Metabolic syndrome is linked with obesity and is often first identified clinically by elevated BMI and elevated levels of fasting blood glucose that are generally secondary to insulin resistance. Using the highly translatable rhesus monkey (Macaca mulatta) model, we asked if metabolic syndrome risk could be identified earlier. The study involved 16 overweight but healthy, euglycemic monkeys, one-half of which spontaneously developed metabolic syndrome over the course of 2 years while the other half remained healthy. We conducted a series of biometric and plasma measures focusing on adiposity, lipid metabolism, and adipose tissue-derived hormones, which led to a diagnosis of metabolic syndrome in the insulin-resistant animals. Plasma fatty acid composition was determined by gas chromatography for cholesteryl ester, FFA, diacylglycerol (DAG), phospholipid, and triacylglycerol lipid classes; plasma lipoprotein profiles were generated by NMR; and circulating levels of adipose-derived signaling peptides were determined by ELISA. We identified biomarker models including a DAG model, two lipoprotein models, and a multiterm model that includes the adipose-derived peptide adiponectin. Correlations among circulating lipids and lipoproteins revealed shifts in lipid metabolism during disease development. We propose that lipid profiling may be valuable for early metabolic syndrome detection in a clinical setting.

BACKGROUND/AIMS: Spine and spinal cord pathologies and associated neuropathic pain are among the most complex medical disorders to treat. While rodent models are widely used in spine and spinal cord research and have provided valuable insight into pathophysiological mechanisms, these models offer limited translatability. Thus, studies in rodent models have not led to the development of clinically effective therapies. More recently, swine has become a favored model for spine research because of the high congruency of the species to humans with respect to spine and spinal cord anatomy, vasculature, and immune responses. However, conventional breeds of swine commonly used in these studies present practical and translational hurdles due to their rapid growth toward weights well above those of humans. METHODS: In the current study, we evaluated the suitability of a human-sized breed of swine developed at the University of Wisconsin-Madison, the Wisconsin Miniature Swine(TM) (WMS(TM)), in the context of thoracic spine morphometry for use in research to overcome limitations of conventional swine breeds. The morphometry of thoracic vertebrae (T1–T15) of 5–6 months-old WMS was analyzed and compared to published values of human and conventional swine spines. RESULTS: The key finding of this study is that WMS spine more closely models the human spine for many of the measured vertebrae parameters, while being similar to conventional swine in respect to the other parameters. CONCLUSION: WMS provides an improvement over conventional swine for use in translational spinal cord injury studies, particularly long-term ones, because of its slower rate of growth and its maximum growth being limited to human weight and size.

Supercritical fluid extraction (SFE) removed lipophilic compounds and low molecular weight flavonoids from cranberries. However, SFE did not extract proanthocyanidins (PAC). The SFE PAC-enriched residue was submitted to fractionation on Sephadex LH-20 using ethanol, ethanol/methanol, and 80% acetone. PAC degree of polymerization (DP) and ratios of "A-type" to "B-type" interflavan bonds were compared with those of PAC fractions without SFE. Mass spectrometry showed that when SFE was used, PAC distribution was shifted toward higher DP and contained higher amounts of two and three "A-type" bonds compared to PAC fractions without SFE. The 80% acetone fraction with SFE had significantly greater extraintestinal pathogenic Escherichia coli (ExPEC) agglutination and significantly lower ExPEC invasion of enterocytes than the fraction without SFE. Cranberry PAC with higher numbers of "A-type" interflavan bonds are more bioactive in agglutinating ExPEC and inhibiting ExPEC enterocyte invasion.

Purpose To assess measurements of pulse wave velocity (PWV) and wall shear stress (WSS) in a swine model of atherosclerosis. Materials and Methods Nine familial hypercholesterolemic (FH) swine with angioplasty balloon catheter-induced atherosclerotic lesions to the abdominal aorta (injured group) and 10 uninjured FH swine were evaluated with a 4D phase contrast (PC) magnetic resonance imaging (MRI) acquisition, as well as with radial and Cartesian 2D PC acquisitions, on a 3T MR scanner. PWV values were computed from the 2D and 4D PC techniques, compared between the injured and uninjured swine, and validated against reference standard pressure probe-based PWV measurements. WSS values were also computed from the 4D PC MRI technique and compared between injured and uninjured groups. Results PWV values were significantly greater in the injured than in the uninjured groups with the 4D PC MRI technique (P = 0.03) and pressure probes (P = 0.02). No significant differences were found in PWV between groups using the 2D PC techniques (P = 0.75–0.83). No significant differences were found for WSS values between the injured and uninjured groups. Conclusion The 4D PC MRI technique provides a promising means of evaluating PWV and WSS in a swine model of atherosclerosis, providing a potential platform for developing the technique for the early detection of atherosclerosis. J. Magn. Reson. Imaging 2015;41:1475–1485. © 2014 Wiley Periodicals, Inc.

Many publications describe separation and quantification techniques for anthocyanins. However, chromatographic conditions to optimize the separation were seldom reported. We developed a sensitive and reliable HPLC method and validated the method for anthocyanin separation, identification, and quantification. Separation was performed on a Zorbax SB C18 column. The conditions including ion pair reagent, mobile phase pH, and mobile phase flow rate were optimized to improve the separation efficiency. Tandem MS was used to identify the total 6 anthocyanins in black currant. Validation of the developed method showed good linearity, precision, repeatability, and excellent recovery. The validated method was used to quantify anthocyanins from black currant samples.

Rapacz familial hypercholesterolemic (RFH) swine is a well-established model of human FH, a highly prevalent hereditary disease associated with increased risk of coronary artery disease and calcific aortic valve disease (CAVD). However, while these animals have been used extensively for the study of atherosclerosis, the heart valves from RFH swine have not previously been examined. We report the analysis of valvular interstitial cell gene expression in adult (two year old) and juvenile (three months old) RFH and WT swine by microarray analysis via the Affymetrix Porcine Genome Array (GEO #: GSE53997). Principal component and hierarchical clustering analysis revealed grouping and almost no variability between the RFH juvenile and WT juvenile groups. Additionally, only 21 genes were found differentially expressed between these two experimental groups whereas over 900 genes were differentially expressed when comparing either RFH or WT juvenile swine to RFH adults.