Altered cortical synaptic lipid signaling leads to intermediate phenotypes of mental disorders.

Excitation/inhibition (E/I) balance plays important roles in mental disorders. Bioactive phospholipids like lysophosphatidic acid (LPA) are synthesized by the enzyme autotaxin (ATX) at cortical synapses and modulate glutamatergic transmission, and eventually alter E/I balance of cortical networks. Here, we analyzed functional consequences of altered E/I balance in 25 human subjects induced by genetic disruption of the synaptic lipid signaling modifier PRG-1, which were compared to 25 age and sex matched control subjects. Furthermore, we tested therapeutic options targeting ATX in a related mouse line. Using EEG combined with TMS in an instructed fear paradigm, neuropsychological analysis and an fMRI based episodic memory task, we found intermediate phenotypes of mental disorders in human carriers of a loss-of-function single nucleotide polymorphism of PRG-1 (PRG-1R345T/WT). Prg-1R346T/WT animals phenocopied human carriers showing increased anxiety, a depressive phenotype and lower stress resilience. Network analysis revealed that coherence and phase-amplitude coupling were altered by PRG-1 deficiency in memory related circuits in humans and mice alike. Brain oscillation phenotypes were restored by inhibtion of ATX in Prg-1 deficient mice indicating an interventional potential for mental disorders.

A 3D-Printed Dummy for Training Distal Phalanx Amputation in Mice.

The development of realistic dummies for training the distal phalanx amputation (DPA) technique in mouse pups is a promising alternative to reduce and replace animals in training for research and teaching. To test this, we obtained micro-CT data from postnatal day-five mouse pups, meticulously segmented them, and converted them into a 3D mesh format suitable for 3D printing. Once the dummy was printed, it was evaluated during actual training courses in two different groups: in the first group, users received no dummies to train the DPA, and in the second group, users were trained with three dummies. To assess the effectiveness of the dummy, we conducted a survey followed by an expert veterinarian evaluation. Our results showed that DPA is a complex procedure, and it is commonly poorly performed. When implementing the dummies, users who were not provided with dummies to practice only had an 8.3% success rate in DPA, while users provided with three dummies had a 45.5% success rate, respectively. Despite additional research being needed, our dummy offered improved practical training by providing a safe and effective alternative in line with ethical considerations while demonstrating the feasibility of using 3D printing technology to promote the 3Rs in experimental research.

Quantitative multi-energy micro-CT: A simulation and phantom study for simultaneous imaging of four different contrast materials using an energy integrating detector.

Emerging from the development of single-energy Computed Tomography (CT) and Dual-Energy Computed Tomography, Multi-Energy Computed Tomography (MECT) is a promising tool allowing advanced material and tissue decomposition and thereby enabling the use of multiple contrast materials in preclinical research. The scope of this work was to evaluate whether a usual preclinical micro-CT system is applicable for the decomposition of different materials using MECT together with a matrix-inversion method and how different changes of the measurement-environment affect the results. A matrix-inversion based algorithm to differentiate up to five materials (iodine, iron, barium, gadolinium, residual material) by applying four different acceleration voltages/energy levels was established. We carried out simulations using different ratios and concentrations (given in fractions of volume units, VU) of the four different materials (plus residual material) at different noise-levels for 30 keV, 40 keV, 50 keV, 60 keV, 80 keV and 100 keV (monochromatic). Our simulation results were then confirmed by using region of interest-based measurements in a phantom-study at corresponding acceleration voltages. Therefore, different mixtures of contrast materials were scanned using a micro-CT. Voxel wise evaluation of the phantom imaging data was conducted to confirm its usability for future imaging applications and to estimate the influence of varying noise-levels, scattering, artifacts and concentrations. The analysis of our simulations showed the smallest deviation of 0.01 (0.003-0.15) VU between given and calculated concentrations of the different contrast materials when using an energy-combination of 30 keV, 40 keV, 50 keV and 100 keV for MECT. Subsequent MECT phantom measurements, however, revealed a combination of acceleration voltages of 30 kV, 40 kV, 60 kV and 100 kV as most effective for performing material decomposition with a deviation of 0.28 (0-1.07) mg/ml. The feasibility of our voxelwise analyses using the proposed algorithm was then confirmed by the generation of phantom parameter-maps that matched the known contrast material concentrations. The results were mostly influenced by the noise-level and the concentrations used in the phantoms. MECT using a standard micro-CT combined with a matrix inversion method is feasible at four different imaging energies and allows the differentiation of mixtures of up to four contrast materials plus an additional residual material.

NECAB2 is an endosomal protein important for striatal function.

Synaptic signaling depends on ATP generated by mitochondria. Dysfunctional mitochondria shift the redox balance towards a more oxidative environment. Due to extensive connectivity, the striatum is especially vulnerable to mitochondrial dysfunction. We found that neuronal calcium-binding protein 2 (NECAB2) plays a role in striatal function and mitochondrial homeostasis. NECAB2 is a predominantly endosomal striatal protein which partially colocalizes with mitochondria. This colocalization is enhanced by mild oxidative stress. Global knockout of Necab2 in the mouse results in increased superoxide levels, increased DNA oxidation and reduced levels of the antioxidant glutathione which correlates with an altered mitochondrial shape and function. Striatal mitochondria from Necab2 knockout mice are more abundant and smaller and characterized by a reduced spare capacity suggestive of intrinsic uncoupling respectively mitochondrial dysfunction. In line with this, we also found an altered stress-induced interaction of endosomes with mitochondria in Necab2 knockout striatal cultures. The predominance of dysfunctional mitochondria and the pro-oxidative redox milieu correlates with a loss of striatal synapses and behavioral changes characteristic of striatal dysfunction like reduced motivation and altered sensory gating. Together this suggests an involvement of NECAB2 in an endosomal pathway of mitochondrial stress response important for striatal function.

Mutations in plasticity-related-gene-1 (PRG-1) protein contribute to hippocampal seizure susceptibility and modify epileptic phenotype.

The Phospholipid Phosphatase Related 4 gene (PLPPR4,  *607813) encodes the Plasticity-Related-Gene-1 (PRG-1) protein. This cerebral synaptic transmembrane-protein modulates cortical excitatory transmission on glutamatergic neurons. In mice, homozygous Prg-1 deficiency causes juvenile epilepsy. Its epileptogenic potential in humans was unknown. Thus, we screened 18 patients with infantile epileptic spasms syndrome (IESS) and 98 patients with benign familial neonatal/infantile seizures (BFNS/BFIS) for the presence of PLPPR4 variants. A girl with IESS had inherited a PLPPR4-mutation (c.896C > G, NM_014839; p.T299S) from her father and an SCN1A-mutation from her mother (c.1622A > G, NM_006920; p.N541S). The PLPPR4-mutation was located in the third extracellular lysophosphatidic acid-interacting domain and in-utero electroporation (IUE) of the Prg-1p.T300S construct into neurons of Prg-1 knockout embryos demonstrated its inability to rescue the electrophysiological knockout phenotype. Electrophysiology on the recombinant SCN1Ap.N541S channel revealed partial loss-of-function. Another PLPPR4 variant (c.1034C > G, NM_014839; p.R345T) that was shown to result in a loss-of-function aggravated a BFNS/BFIS phenotype and also failed to suppress glutamatergic neurotransmission after IUE. The aggravating effect of Plppr4-haploinsufficiency on epileptogenesis was further verified using the kainate-model of epilepsy: double heterozygous Plppr4-/+|Scn1awt|p.R1648H mice exhibited higher seizure susceptibility than either wild-type, Plppr4-/+, or Scn1awt|p.R1648H littermates. Our study shows that a heterozygous PLPPR4 loss-of-function mutation may have a modifying effect on BFNS/BFIS and on SCN1A-related epilepsy in mice and humans.

EGFL7 loss correlates with increased VEGF-D expression, upregulating hippocampal adult neurogenesis and improving spatial learning and memory.

Neural stem cells reside in the subgranular zone, a specialized neurogenic niche of the hippocampus. Throughout adulthood, these cells give rise to neurons in the dentate gyrus, playing an important role in learning and memory. Given that these core cognitive processes are disrupted in numerous disease states, understanding the underlying mechanisms of neural stem cell proliferation in the subgranular zone is of direct practical interest. Here, we report that mature neurons, neural stem cells and neural precursor cells each secrete the neurovascular protein epidermal growth factor-like protein 7 (EGFL7) to shape this hippocampal niche. We further demonstrate that EGFL7 knock-out in a Nestin-CreERT2-based mouse model produces a pronounced upregulation of neurogenesis within the subgranular zone. RNA sequencing identified that the increased expression of the cytokine VEGF-D correlates significantly with the ablation of EGFL7. We substantiate this finding with intraventricular infusion of VEGF-D upregulating neurogenesis in vivo and further show that VEGF-D knock-out produces a downregulation of neurogenesis. Finally, behavioral studies in EGFL7 knock-out mice demonstrate greater maintenance of spatial memory and improved memory consolidation in the hippocampus by modulation of pattern separation. Taken together, our findings demonstrate that both EGFL7 and VEGF-D affect neurogenesis in the adult hippocampus, with the ablation of EGFL7 upregulating neurogenesis, increasing spatial learning and memory, and correlating with increased VEGF-D expression.

Laparoscopic Electromyography and Electrostimulation of the Gastrointestinal Tract Before Placement of Theranostic Devices.

NEED: Electrical stimulation (ES) is a promising therapy for multisegmental gastrointestinal (GI) motility disorders such as gastroparesis with slow-transit constipation or chronic intestinal pseudo-obstruction. Wireless communicating GI devices for smart sensing and ES-based motility modulation will soon be available. Before placement, a potential benefit for each GI segment must be intraoperatively assessed. TECHNICAL SOLUTION: A minimally invasive multisegmental electromyography (EMG) analysis with ES of the GI tract is required. PROOF OF CONCEPT: Two porcine experiments were performed with a laparoscopic setup. Multiple hook-needle electrodes were subserosally applied in the stomach, duodenum, jejunum, ileum, and colon. EMG signals were acquired for computer-assisted motility analysis. Gastric ES, duodenal ES, jejunal ES, ileal ES, and colonic ES were applied. NEXT STEPS: Further technological and rapid regulatory solutions are desired to initialize a clinical trial of the next generation devices in the near future. CONCLUSION: We demonstrate a laparoscopic strategy with EMG analysis and ES of multiple GI segments. Thus, GI function may be evaluated before theranostic devices are placed. Extended GI resection or organ transplantation may be delayed or even avoided in affected patients.

The Current Status and Work of Three Rs Centres and Platforms in Europe.

The adoption of Directive 2010/63/EU on the protection of animals used for scientific purposes has given a major push to the formation of Three Rs initiatives in the form of centres and platforms. These centres and platforms are dedicated to the so-called Three Rs, which are the Replacement, Reduction and Refinement of animal use in experiments. ATLA's 50th Anniversary year has seen the publication of two articles on European Three Rs centres and platforms. The first of these was about the progressive rise in their numbers and about their founding history; this second part focuses on their current status and activities. This article takes a closer look at their financial and organisational structures, describes their Three Rs focus and core activities (dissemination, education, implementation, scientific quality/translatability, ethics), and presents their areas of responsibility and projects in detail. This overview of the work and diverse structures of the Three Rs centres and platforms is not only intended to bring them closer to the reader, but also to provide role models and show examples of how such Three Rs centres and platforms could be made sustainable. The Three Rs centres and platforms are very important focal points and play an immense role as facilitators of Directive 2010/63/EU 'on the ground' in their respective countries. They are also invaluable for the wide dissemination of information and for promoting the implementation of the Three Rs in general.

Clicker Training Mice for Improved Compliance in the Catwalk Test.

The CatWalk test relies on the run of mice across the platform to measure a constant speed with low variation. Mice usually require a stimulus to walk to the end of the catwalk. However, such stimuli are usually aversive and can impair welfare. Positive reinforcement training of laboratory animals is a thriving tool for refinement and contributes to meeting the demands instituted by Directive 2010/63/EU. We have already demonstrated the positive effects of clicker training. In this study, we trained male and female mice to complete the CatWalk protocol while assessing the effects of training on their well-being (Open Filed and Elevated Plus Maze). In the CatWalk test, we observed that clicker training improved the running speed of the mice. In addition, clicker training reduced the number of runs required by mice, which was more pronounced in males. Clicker training lowered anxiety-like behaviors in our mice, especially in females, where a significant difference was observed between trained and untrained ones. Based on our findings, we hypothesize that clicker training is an effective tool to motivate mice and increase performance on the CatWalk test without potentially impairing their welfare (e.g., by puffing them).

Development of a 3D simulator for training the mouse in utero electroporation.

In utero electroporation (IUE) requires high-level training in microinjection through the mouse uterine wall into the lateral ventricle of the mouse brain. Training for IUE is currently being performed in live mice as no artificial models allow simulations yet. This study aimed to develop an anatomically realistic 3D printed simulator to train IUE in mice. To this end, we created embryo models containing lateral ventricles. We coupled them to uterus models in six steps: (1) computed tomography imaging, (2) 3D model segmentation, (3) 3D model refinement, (4) mold creation to cast the actual model, (5) 3D mold printing, and (6) mold casting the molds with a mix of soft silicones to ensure the hardness and consistency of the uterus and embryo. The results showed that the simulator assembly successfully recreated the IUE. The compression test did not differ in the mechanical properties of the real embryo or in the required load for uterus displacement. Furthermore, more than 90% of the users approved the simulator as an introduction to IUE and considered that the simulator could help reduce the number of animals for training. Despite current limitations, our 3D simulator enabled a realistic experience for initial approximations to the IUE and is a real alternative for implementing the 3Rs. We are currently working on refining the model.

AgRP neurons control feeding behaviour at cortical synapses via peripherally derived lysophospholipids.

Phospholipid levels are influenced by peripheral metabolism. Within the central nervous system, synaptic phospholipids regulate glutamatergic transmission and cortical excitability. Whether changes in peripheral metabolism affect brain lipid levels and cortical excitability remains unknown. Here, we show that levels of lysophosphatidic acid (LPA) species in the blood and cerebrospinal fluid are elevated after overnight fasting and lead to higher cortical excitability. LPA-related cortical excitability increases fasting-induced hyperphagia, and is decreased following inhibition of LPA synthesis. Mice expressing a human mutation (Prg-1R346T) leading to higher synaptic lipid-mediated cortical excitability display increased fasting-induced hyperphagia. Accordingly, human subjects with this mutation have higher body mass index and prevalence of type 2 diabetes. We further show that the effects of LPA following fasting are under the control of hypothalamic agouti-related peptide (AgRP) neurons. Depletion of AgRP-expressing cells in adult mice decreases fasting-induced elevation of circulating LPAs, as well as cortical excitability, while blunting hyperphagia. These findings reveal a direct influence of circulating LPAs under the control of hypothalamic AgRP neurons on cortical excitability, unmasking an alternative non-neuronal route by which the hypothalamus can exert a robust impact on the cortex and thereby affect food intake.

The Rise of Three Rs Centres and Platforms in Europe.

Public awareness and discussion about animal experiments and replacement methods has greatly increased in recent years. The term 'the Three Rs', which stands for the Replacement, Reduction and Refinement of animal experiments, is inseparably linked in this context. A common goal within the Three Rs scientific community is to develop predictive non-animal models and to better integrate all available data from in vitro, in silico and omics technologies into regulatory decision-making processes regarding, for example, the toxicity of chemicals, drugs or food ingredients. In addition, it is a general concern to implement (human) non-animal methods in basic research. Toward these efforts, there has been an ever-increasing number of Three Rs centres and platforms established over recent years - not only to develop novel methods, but also to disseminate knowledge and help to implement the Three Rs principles in policies and education. The adoption of Directive 2010/63/EU on the protection of animals used for scientific purposes gave a strong impetus to the creation of Three Rs initiatives, in the form of centres and platforms. As the first of a series of papers, this article gives an overview of the European Three Rs centres and platforms, and their historical development. The subsequent articles, to be published over the course of ATLA's 50th Anniversary year, will summarise the current focus and tasks as well as the future and the plans of the Three Rs centres and platforms. The Three Rs centres and platforms are very important points of contact and play an immense role in their respective countries as 'on the ground' facilitators of Directive 2010/63/EU. They are also invaluable for the widespread dissemination of information and for promoting implementation of the Three Rs in general.

Comparing Aachen Minipigs and Pietrain Piglets as Models of Experimental Pediatric Urology to Human Reference Data.

INTRODUCTION: Swine had special roles in the development of minimally invasive procedures to treat vesicoureteral reflux, and minipigs have been gaining ground in recent years in experimental pediatric urology as they combine small size with less vulnerable adult physiology, but their suitability as a model has never been assessed. We therefore compared a landrace piglet with a juvenile minipig to elucidate comparability. METHODS: We evaluated five 3-week old Pietrain piglets and five 3-month old Aachen Minipigs as representatives of landrace and minipig models based on their expected bodyweight being similar to a newborn human. We compared renal weight, volume - via the ellipsoid formula - and ureteral length. In addition, we calculated porcine renal function via Gasthuys' formula. In order to compare the groups with previously published values for infants, we used resampling techniques to allow comparison to humans. RESULTS: Renal weight was higher in humans than in Pietrain piglets (ΔL = 7.6 g; ΔR = 5.4 g) and Aachen Minipigs (ΔL = 11 g; ΔR = 9.4 g). Renal volumes in humans were higher than in both Pietrain piglets (ΔL = 5.6 mL, p < 0.001; ΔR = 3.7 mL, p = 0.004) and Aachen Minipigs (ΔL = 8.1 mL; ΔR = 6.6 mL; both p < 0.001). Ureteral lengths in humans and both pig breeds were comparable as were estimated renal functions between both pig breeds. DISCUSSION AND CONCLUSION: Both landrace piglets and juvenile minipigs are suitable models for experimental pediatric urology as parameters did not differ between them. In addition, the anatomic parameters are comparable or smaller than in infants. This might facilitate translational research as technical failure is less likely in larger organs. Additional research is necessary to cover higher age ranges than those included in the present pilot study.

Monitoring of tumor growth and vascularization with repetitive ultrasonography in the chicken chorioallantoic-membrane-assay.

The chorioallantoic-membrane (CAM)-assay is an established model for in vivo tumor research. Contrary to rodent-xenograft-models, the CAM-assay does not require breeding of immunodeficient strains due to native immunodeficiency. This allows xenografts to grow on the non-innervated CAM without pain or impairment for the embryo. Considering multidirectional tumor growth, limited monitoring capability of tumor size is the main methodological limitation of the CAM-assay for tumor research. Enclosure of the tumor by the radiopaque eggshell and the small structural size only allows monitoring from above and challenges established imaging techniques. We report the eligibility of ultrasonography for repetitive visualization of tumor growth and vascularization in the CAM-assay. After tumor ingrowth, ultrasonography was repetitively performed in ovo using a commercial ultrasonographic scanner. Finally, the tumor was excised and histologically analyzed. Tumor growth and angiogenesis were successfully monitored and findings in ultrasonographic imaging significantly correlated with results obtained in histological analysis. Ultrasonography is cost efficient and widely available. Tumor imaging in ovo enables the longitudinal monitoring of tumoral development, yet allowing high quantitative output due to the CAM-assays simple and cheap methodology. Thus, this methodological novelty improves reproducibility in the field of in vivo tumor experimentation emphasizing the CAM-assay as an alternative to rodent-xenograft-models.

Phf21b imprints the spatiotemporal epigenetic switch essential for neural stem cell differentiation.

Cerebral cortical development in mammals involves a highly complex and organized set of events including the transition of neural stem and progenitor cells (NSCs) from proliferative to differentiative divisions to generate neurons. Despite progress, the spatiotemporal regulation of this proliferation-differentiation switch during neurogenesis and the upstream epigenetic triggers remain poorly known. Here we report a cortex-specific PHD finger protein, Phf21b, which is highly expressed in the neurogenic phase of cortical development and gets induced as NSCs begin to differentiate. Depletion of Phf21b in vivo inhibited neuronal differentiation as cortical progenitors lacking Phf21b were retained in the proliferative zones and underwent faster cell cycles. Mechanistically, Phf21b targets the regulatory regions of cell cycle promoting genes by virtue of its high affinity for monomethylated H3K4. Subsequently, Phf21b recruits the lysine-specific demethylase Lsd1 and histone deacetylase Hdac2, resulting in the simultaneous removal of monomethylation from H3K4 and acetylation from H3K27, respectively. Intriguingly, mutations in the Phf21b locus associate with depression and mental retardation in humans. Taken together, these findings establish how a precisely timed spatiotemporal expression of Phf21b creates an epigenetic program that triggers neural stem cell differentiation during cortical development.

Robotic Setup Promises Consistent Effects of Multilocular Gastrointestinal Electrical Stimulation: First Results of a Porcine Study.

BACKGROUND: Electrical stimulation (ES) of several gastrointestinal (GI) segments is a promising therapeutic option for multilocular GI dysmotility, but conventional surgical access by laparotomy involves a high degree of tissue trauma. We evaluated a minimally invasive surgical approach using a robotic surgical system to perform electromyographic (EMG) recordings and ES of several porcine GI segments, comparing these data to an open surgical approach by laparotomy. MATERIALS AND METHODS: In 5 acute porcine experiments, we placed multiple electrodes on the stomach, duodenum, jejunum, ileum, and colon. Three experiments were performed with a median laparotomy and 2 others using a robotic platform. Multichannel EMGs were recorded, and ES was sequentially delivered with 4 ES parameters to the 5 target segments. We calculated pre- and poststimulatory spikes per minute (Spm) and performed a statistical Poisson analysis. RESULTS: Electrode placement was achieved in all cases without complications. Increased technical and implantation time were required to achieve the robotic electrode placement, but invasiveness was markedly reduced in comparison to the conventional approach. The highest calculated (c)Spm values were found in the poststimulatory period of the small bowel with both the conventional and robotic approaches. Six of the 20 Poisson test results in the open setup reached statistical significance and 12 were significant in the robotic experiments. CONCLUSIONS: The robotic setup was less invasive, revealed more consistent effects of multilocular ES in several GI segments, and is a promising option for future preclinical and clinical studies of GI motility disorders.

Bax inhibitor-1 deficiency leads to obesity by increasing Ca2+-dependent insulin secretion.

Transmembrane BAX inhibitor motif containing 6 (TMBIM6), also known as Bax inhibitor-1, is an evolutionarily conserved protein involved in endoplasmic reticulum (ER) function. TMBIM6 is an ER Ca2+ leak channel and its deficiency enhances susceptibility to ER stress due to inhibition of the ER stress sensor IRE1α. It was previously shown that TMBIM6 overexpression improves glucose metabolism and that TMBIM6 knockout mice develop obesity. We here examined the metabolic alterations underlying the obese phenotype and subjected TMBIM6 knockout mice to indirect calorimetry and euglycemic-hyperinsulinemic tests with stable isotope dilution to gauge tissue-specific insulin sensitivity. This demonstrated no changes in heat production, food intake, activity or hepatic and peripheral insulin sensitivity. TMBIM6 knockout mice, however, featured a higher glucose-stimulated insulin secretion in vivo as assessed by the hyperglycemic clamp test and hepatic steatosis. This coincided with profound changes in glucose-mediated Ca2+ regulation in isolated pancreatic ß cells and increased levels of IRE1α levels but no differences in downstream effects of IRE1α like increased Xbp1 mRNA splicing or Ire1-dependent decay of insulin mRNA in the pancreas. We therefore conclude that lack of TMBIM6 does not affect insulin sensitivity but leads to hyperinsulinemia, which serves to explain the weight gain. TMBIM6-mediated metabolic alterations are mainly caused by its role as a Ca2+ release channel in the ER. KEY MESSAGES: TMBIM6-/- leads to obesity and hepatic steatosis. Food intake and energy expenditure are not changed in TMBIM6-/- mice. No changes in insulin resistance in TMBIM6-/- mice. Increased insulin secretion caused by altered calcium dynamics in ß cells.

Translational development and pre-clinical evaluation of prototype gastrointestinal mock-up devices: only robotic placement of plastic?

Background: The aim of this study was to address the vision of wireless theranostic devices distributed along the gastrointestinal (GI) tract by defining design requirements, developing prototype mock-ups, and establishing a minimally invasive surgical approach for the implantation process.Methods: Questionnaires for contextual analysis and use case scenarios addressing the technical issues of an implantable GI device, a possible scenario for implantation, preparation and calibration of a device, and therapeutic usage by professionals and patients were completed and discussed by an interdisciplinary team of surgeons, engineers, and product designers. Two acute porcine experiments were conducted with a robotic surgical system under general anaesthesia.Results: A variety of requirements for the design and implantation of implantable devices for modulating GI motility were defined. Five prototype implant mock-ups were three-dimensional (3D)-printed from black polymer material (width 22.32 mm, height 7.66 mm) and successfully implanted on the stomach, duodenum, jejunum, ileum, and colon using the robotic surgical system, without any complications.Conclusions: Our study shows the development and successful pre-clinical evaluation of a reliable device design with a minimally invasive implantation approach. Several stages of device development, including pre-clinical tests, characterisation of clinical requirements, regulatory affairs, and marketing issues should be managed side by side.

Using multidimensional scaling in model choice for congenital oesophageal atresia: similarity analysis of human autopsy organ weights with those from a comparative assessment of Aachen Minipig and Pietrain piglets.

Swine models had been popular in paediatric oesophageal surgery in the past. Although being largely replaced by rodent models, swine experienced a revival with the establishment of minipig models. However, none of them has ever been investigated for similarity to humans. We conducted a pilot study to determine whether three-week old Pietrain piglets and three-month old Aachen Minipigs are suitable for experimental paediatric oesophageal atresia surgery. We tested the operation's feasibility, performed a necropsy, weighed organs, measured organ length and calculated relative weights and lengths, and measured laboratory parameters. We used multidimensional scaling to assess the similarity of the swine breeds with previously published human data. Pietrain piglets had a higher a priori bodyweight than Aachen Minipigs (Δ = 1.31 kg, 95% confidence interval (CI): 0.37-2.23, p = 0.015), while snout-to-tail length was similar. Pietrain piglets had higher absolute and relative oesophageal lengths (Δ = 5.43 cm, 95% CI: 2.2-8.6; p = 0.0062, q1* = 0.0083 and Δ = 11.4%, 95% CI: 5.1-17.6; p = 0.0025, q3* = 0.0053). Likewise, absolute and relative small intestinal lengths were higher in Pietrains, but all other parameters did not differ, with the exception of minor differences in laboratory parameters. Multidimensional scaling revealed three-week old Pietrain piglets to be similar to two-month old humans based on their thoracoabdominal organ weights. This result indicates three-week old Pietrain piglets are a suitable model of paediatric oesophageal atresia surgery, because clinically many procedures are performed at around eight weeks age. Three-month old Aachen Minipigs were more dissimilar to eight-week old humans than three-week old Pietrain piglets.