One-Year Outcome of an Ongoing Pre-Clinical Growing Animal Model for a Tissue-Engineered Valved Pulmonary Conduit.

Objectives: A self-constructed valved pulmonary conduit made out of a de-cellularized porcine small intestinal submucosal extracellular matrix biological scaffold was tested in a chronic growing lamb model. Methods: The conduit was implanted in pulmonary valve position in 19 lambs. We monitored clinical, laboratory, and echocardiographic findings until 12 months after surgery. In two animals, euthanasia was planned at nine and twelve months. Pre-mortem chest computed tomography and post-mortem pathologic work up were performed. Data are presented as frequency and percentage, median and range, or mean and standard deviation. Results: Twelve (63.2%) animals survived the perioperative period. Three unexpected deaths occurred during the follow-up period: one due to aspiration pneumonia at 23 days after surgery, and two due to early and late infective endocarditis of the conduit at 18 and 256 days. In the two animals with planned scarification, the pre-mortem CT scan revealed mild or no calcification within the conduit or valve leaflets. In the echocardiographic examination at 12 months, peak and mean systolic pressure gradients across the conduit valve were 6.5 (3-21) mmHg and 3 (2-12) mmHg, while valve regurgitation was none (n = 2), trivial (n = 5), moderate (n = 1), or severe (n = 1). No clinical or laboratory signs of hemolysis were seen. After 12 months of follow-up, the animals' body weights had increased from 33 (27-38) kg to 53 (38-66) kg (p = 0.010). Conclusions: Implantation of a valved pulmonary conduit in our growing lamb model was feasible. Infective endocarditis of the implanted valved conduit remained a significant complication.

Vitamin C as a Potential Prophylactic Measure Against Frozen Shoulder in an In Vivo Shoulder Contracture Animal Model.

BACKGROUND: Frozen shoulder is a common, painful, and movement-restricting condition. Although primary frozen shoulder is idiopathic, secondary frozen shoulder can occur after trauma or surgery. Prophylactic and therapeutic options are often unsatisfactory. Vitamin C (ascorbic acid) is a potent physiological antioxidant and likely inhibits the activation of nuclear factor κB, which plays a decisive role in inflammatory reactions. HYPOTHESIS: Because of its anti-inflammatory effects, vitamin C may be valuable in the prevention of secondary frozen shoulder. STUDY DESIGN: Controlled laboratory study. METHODS: An in vivo shoulder contracture model was conducted by fixation of the right proximal limb of Sprague-Dawley rats. A treatment group (n = 8) receiving vitamin C orally was compared with a control group (n = 9) without vitamin C. The primary outcome was capsular thickness at the shoulder joint measured on magnetic resonance imaging (MRI) examination. Further histological examination was performed but was not statistically analyzed because of variability of the cutting plane through the glenoid. RESULTS: Vitamin C treatment resulted in less thickening of the axillary fold of the operated shoulder at 2 of the 3 locations measured on MRI compared with untreated controls (insertion to the glenoid, P = .074; insertion to the humerus, P = .006; middle of the axillary recess, P = .008). The observed structural changes in histological examination corroborated the significant changes obtained from the MRI measurements. CONCLUSION: Prophylactic vitamin C seemed to reduce the thickening of the axillary recess in secondary frozen shoulder in this preclinical study. CLINICAL RELEVANCE: Vitamin C may be helpful as a noninvasive therapeutic measure to prevent secondary frozen shoulder (eg, within the context of surgery in the shoulder region or immobilization) or to treat primary frozen shoulder at an early stage. Further studies are required to evaluate the effect of this treatment in humans and the necessary dosage in humans.

Platelet-rich plasma as a potential prophylactic measure against frozen shoulder in an in vivo shoulder contracture model.

INTRODUCTION: Frozen shoulder (adhesive capsulitis) is a common painful and functionally-limiting disease affecting around 2% of the population. So far, therapeutic options are limited and often unsatisfactory. Platelet-rich plasma (PRP) has been used as a treatment option in other orthopedic diseases since it contains growth factors that stimulate tissue repair. So far, the effect of PRP on frozen shoulder lacks evidence. We hypothesized that PRP may be valuable in the prophylaxis and treatment of secondary frozen shoulder due to capsular remodeling. MATERIALS AND METHODS: An experimental study of an in vivo frozen shoulder model was conducted. Twenty Sprague-Dawley rats underwent surgery in which the body of the scapula was connected to the humerus with a high-strength suture. Two groups of 8 weeks survival time were allocated; a treatment group with one intraoperative injection of PRP into the glenohumeral joint (n = 10) and a control group without PRP (n = 10). The primary outcome was the structural change in the posterior synovial membrane of the posterior and inferior part of the glenohumeral joint using a semi-quantitative grading from 0 (lowest) to 3 (highest). RESULTS: The posterior synovial membrane structural changes were significantly lower in the PRP group (median = 1 [interquartile range (IQR) = 0-1]) compared to controls (median = 2 [IQR = 1-3]) (p = 0.028). There were no differences for the remaining synovial membrane changes and fibrous capsule responses between groups. CONCLUSIONS: In this in vivo shoulder contracture model, PRP injections seem to reduce the histological severity grade of some parts (i.e., posterior synovial membrane changes) of the secondary frozen shoulder without causing any side effects. It may be considered to investigate this effect further in future studies as a potential prophylaxis of secondary frozen shoulder (e.g., in operated or immobilized shoulders) or as a treatment option for patients with frozen shoulder in the early stage.

Cardiovascular magnetic resonance imaging of functional and microstructural changes of the heart in a longitudinal pig model of acute to chronic myocardial infarction.

BACKGROUND: We examined the dynamic response of the myocardium to infarction in a longitudinal porcine study using relaxometry, functional as well as diffusion cardiovascular magnetic resonance (CMR). We sought to compare non contrast CMR methods like relaxometry and in-vivo diffusion to contrast enhanced imaging and investigate the link of microstructural and functional changes in the acute and chronically infarcted heart. METHODS: CMR was performed on five myocardial infarction pigs and four healthy controls. In the infarction group, measurements were obtained 2 weeks before 90 min occlusion of the left circumflex artery, 6 days after ischemia and at 5 as well as 9 weeks as chronic follow-up. The timing of measurements was replicated in the control cohort. Imaging consisted of functional cine imaging, 3D tagging, T2 mapping, native as well as gadolinium enhanced T1 mapping, cardiac diffusion tensor imaging, and late gadolinium enhancement imaging. RESULTS: Native T1, extracellular volume (ECV) and mean diffusivity (MD) were significantly elevated in the infarcted region while fractional anisotropy (FA) was significantly reduced. During the transition from acute to chronic stages, native T1 presented minor changes (< 3%). ECV as well as MD increased from acute to the chronic stages compared to baseline: ECV: 125 ± 24% (day 6) 157 ± 24% (week 5) 146 ± 60% (week 9), MD: 17 ± 7% (day 6) 33 ± 14% (week 5) 29 ± 15% (week 9) and FA was further reduced: - 31 ± 10% (day 6) - 38 ± 8% (week 5) - 36 ± 14% (week 9). T2 as marker for myocardial edema was significantly increased in the ischemic area only during the acute stage (83 ± 3 ms infarction vs. 58 ± 2 ms control p < 0.001 and 61 ± 2 ms in the remote area p < 0.001). The analysis of functional imaging revealed reduced left ventricular ejection fraction, global longitudinal strain and torsion in the infarct group. At the same time the transmural helix angle (HA) gradient was steeper in the chronic follow-up and a correlation between longitudinal strain and transmural HA gradient was detected (r = 0.59 with p < 0.05). Comparing non-gadolinium enhanced data T2 mapping showed the largest relative change between infarct and remote during the acute stage (+ 33 ± 4% day 6, with p = 0.013 T2 vs. MD, p = 0.009 T2 vs. FA and p = 0.01 T2 vs. T1) while FA exhibited the largest relative change between infarct and remote during the chronic follow-up (+ 31 ± 2% week 5, with p = N.S. FA vs. MD, p = 0.03 FA vs. T2 and p = 0.003 FA vs. T1). Overall, diffusion parameters provided a higher contrast (> 23% for MD and > 27% for FA) during follow-up compared to relaxometry (T1 17-18%/T2 10-20%). CONCLUSION: During chronic follow-up after myocardial infarction, cardiac diffusion tensor imaging provides a higher sensitivity for mapping microstructural alterations when compared to non-contrast enhanced relaxometry with the added benefit of providing directional tensor information to assess remodelling of myocyte aggregate orientations, which cannot be otherwise assessed.

Establishing a pre-clinical growing animal model to test a tissue engineered valved pulmonary conduit.

BACKGROUND: Many valvular pathologies of the heart may be only sufficiently treated by replacement of the valve if a reconstruction is not feasible. However, structural deterioration, thrombosis with thromboembolic events and infective endocarditis are commonly encountered complications over time and often demand a re-operation. In congenital heart disease the lack of small diameter valves with the potential to grow poses additional challenges and limits treatment options to homo- or xenograft implants. METHODS: In this study, a chronic sheep model (24 months follow-up), a self-constructed valved conduit was created out of a tissue engineered (TE) patch (CorMatrix® Cardiovascular, Inc, USA) and implanted in orthotopic right ventricular (RV)-pulmonary artery (PA) position. Thereafter, the sheep were regularly monitored by clinical, laboratory and echocardiographic examinations to evaluate cardiac function and the implanted RV-PA-conduit. DISCUSSION: Here, we summarize the study protocol and our experiences during the perioperative phase and the follow up period and explain how we constructed a valved conduit out of a commercially available TE patch. TRIAL REGISTRATION: License number: ZH 284/14.

Computed Tomography-based evaluation of porcine cardiac dimensions to assist in pre-study planning and optimized model selection for pre-clinical research.

The pig (Sus Scrofa Domestica) is an accepted model for preclinical evaluation of prosthetic heart valves and trans-catheter implantation techniques. Understanding porcine cardiac dimensions through three-dimensional computed tomography (CT), increases preclinical study success, leading to higher cost efficiency and to the observance of the obligation to the 3 R principles. Cardiac CT images of twenty-four Swiss large white pigs were segmented; aortic root, mitral valve, pulmonary trunk, tricuspid valve, as well as the aorto-mitral angle and left atrial height were analyzed. Correlation coefficient (r) was calculated in relation to body weight. In Swiss large white pigs, valvular dimensions, length of the pulmonary artery and ascending aorta as well as left atrial height correlate with body weight. Coronary ostia heights and aorto-mitral angle size can be neglected in animal size selection; no changes were found for either of the two parameters with increasing body weight.

Transplantation of Autologous Dermo-Epidermal Skin Substitutes in a Pig Model.

Due to its similarity of skin anatomy and physiology, the pig appears to be a well-suited animal model for preclinical studies of skin analog transplantations. The choice of the location of the skin defect and appropriate postoperative measures are essential for the protection of the transplanted graft. This protocol describes in detail a porcine skin transplantation model including peri- and postoperative measures taken to improve and refine the study outcome.

Left ventricular blood flow patterns at rest and under dobutamine stress in healthy pigs.

Intracardiac blood flow patterns are affected by the morphology of cardiac structures and are set up to support the heart's pump function. Exercise affects contractility and chamber size as well as pre- and afterload. The aim of this study was to test the feasibility of four-dimensional phase contrast cardiovascular MRI under pharmacological stress and to study left ventricular blood flow under stress. 4D flow data were successfully acquired and analysed in 12 animals. During dobutamine infusion, heart rate and ejection fraction increased (82 ± 5 bpm versus 124 ± 3 bpm/46 ± 9% versus 65 ± 7%; both p < 0.05). A decrease in left ventricular end-diastolic volume (72 ± 14 mL versus 55 ± 8 mL; p < 0.05) and end-systolic volume (40 ± 15 mL versus 19 ± 6 mL; p < 0.05) but no change in stroke volume were observed. Trans-mitral diastolic inflow velocity increased under dobutamine and the trajectory of inflowing blood was directed towards the anterior septum with increased inflow angle (26 ± 5°) when compared with controls (15 ± 2°). In 5/6 animals undergoing stress diastolic vortices developed later, and in 3/6 animals vortices collapsed earlier with significantly smaller cross-sectional area during diastole. The vorticity index was not affected. Under the stress condition direct flow (% ejection within the next heart beat) increased from 43 ± 6% to 53 ± 8%. 4D MRI blood flow acquisition and analysis are feasible in pig hearts under dobutamine-induced stress. Flow patterns characterized by high blood velocity and antero-septally oriented diastolic inflow as well as decreased ventricular volumes are unfavourable conditions for diastolic vortex development under pharmacological stress, and cardiac output is increased by a rise in heart rate and directly ejected left ventricular blood volume.

Quantitative myocardial first-pass cardiovascular magnetic resonance perfusion imaging using hyperpolarized [1-13C] pyruvate.

BACKGROUND: The feasibility of absolute myocardial blood flow quantification and suitability of hyperpolarized [1-13C] pyruvate as contrast agent for first-pass cardiovascular magnetic resonance (CMR) perfusion measurements are investigated with simulations and demonstrated in vivo in a swine model. METHODS: A versatile simulation framework for hyperpolarized CMR subject to physical, physiological and technical constraints was developed and applied to investigate experimental conditions for accurate perfusion CMR with hyperpolarized [1-13C] pyruvate. Absolute and semi-quantitative perfusion indices were analyzed with respect to experimental parameter variations and different signal-to-noise ratio (SNR) levels. Absolute myocardial blood flow quantification was implemented with an iterative deconvolution approach based on Fermi functions. To demonstrate in vivo feasibility, velocity-selective excitation with an echo-planar imaging readout was used to acquire dynamic myocardial stress perfusion images in four healthy swine. Arterial input functions were extracted from an additional image slice with conventional excitation that was acquired within the same heartbeat. RESULTS: Simulations suggest that obtainable SNR and B0 inhomogeneity in vivo are sufficient for the determination of absolute and semi-quantitative perfusion with ≤25% error. It is shown that for expected metabolic conversion rates, metabolic conversion of pyruvate can be neglected over the short duration of acquisition in first-pass perfusion CMR. In vivo measurements suggest that absolute myocardial blood flow quantification using hyperpolarized [1-13C] pyruvate is feasible with an intra-myocardial variability comparable to semi-quantitative perfusion indices. CONCLUSION: The feasibility of quantitative hyperpolarized first-pass perfusion CMR using [1-13C] pyruvate has been investigated in simulations and demonstrated in swine. Using an approved and metabolically active compound is envisioned to increase the value of hyperpolarized perfusion CMR in patients.

Direct comparison of in vivo versus postmortem second-order motion-compensated cardiac diffusion tensor imaging.

PURPOSE: To directly compare in vivo versus postmortem second-order motion-compensated spin-echo diffusion tensor imaging of the porcine heart. METHODS: Second-order motion-compensated spin-echo cardiac diffusion tensor imaging was performed during systolic contraction in vivo and repeated upon cardiac arrest by bariumchloride without repositioning of the study animal or replaning of imaging slices. In vivo and postmortem reproducibility was assessed by repeat measurements. Comparison of helix, transverse, and sheet (E2A) angulation as well as mean diffusivity and fractional anisotropy was performed. RESULTS: Intraclass correlation coefficients for repeated measurements (postmortem/in vivo) were 0.95/0.96 for helix, 0.70/0.66 for transverse, and 0.79/0.72 for E2A angulation; 0.83/0.72 for mean diffusivity; and 0.78/0.76 for fractional anisotropy. The corresponding 95% levels of agreement across the left ventricle were: helix 14 to 18°/12 to 15°, transverse 9 to 10°/10 to 11°, E2A 15 to 20°/16 to 18°. The 95% levels of agreement across the left ventricle for the comparison of postmortem versus in vivo were 20 to 22° for helix, 13 to 19° for transverse, and 24 to 31° for E2A angulation. CONCLUSIONS: Parameters derived from in vivo second-order motion-compensated spin-echo diffusion tensor imaging agreed well with postmortem imaging, indicating sufficient suppression of motion-induced signal distortions of in vivo cardiac diffusion tensor imaging. Magn Reson Med 79:2265-2276, 2018. © 2017 International Society for Magnetic Resonance in Medicine.

Hyperpolarized 13C urea myocardial first-pass perfusion imaging using velocity-selective excitation.

BACKGROUND: A velocity-selective binomial excitation scheme for myocardial first-pass perfusion measurements with hyperpolarized 13C substrates, which preserves bolus magnetization inside the blood pool, is presented. The proposed method is evaluated against gadolinium-enhanced 1H measurements in-vivo. METHODS: The proposed excitation with an echo-planar imaging readout was implemented on a clinical CMR system. Dynamic myocardial stress perfusion images were acquired in six healthy pigs after bolus injection of hyperpolarized 13C urea with the velocity-selective vs. conventional excitation, as well as standard 1H gadolinium-enhanced images. Signal-to-noise, contrast-to-noise (CNR) and homogeneity of semi-quantitative perfusion measures were compared between methods based on first-pass signal-intensity time curves extracted from a mid-ventricular slice. Diagnostic feasibility is demonstrated in a case of septal infarction. RESULTS: Velocity-selective excitation provides over three-fold reduction in blood pool signal with a two-fold increase in myocardial CNR. Extracted first-pass perfusion curves reveal a significantly reduced variability of semi-quantitative first-pass perfusion measures (12-20%) for velocity-selective excitation compared to conventional excitation (28-93%), comparable to that of reference 1H gadolinium data (9-15%). Overall image quality appears comparable between the velocity-selective hyperpolarized and gadolinium-enhanced imaging. CONCLUSION: The feasibility of hyperpolarized 13C first-pass perfusion CMR has been demonstrated in swine. Comparison with reference 1H gadolinium data revealed sufficient data quality and indicates the potential of hyperpolarized perfusion imaging for human applications.

Voluntary intake of paracetamol-enriched drinking water and its influence on the success of embryo transfer in mice.

Embryo transfer (ET) in mice is a key technique in biomedical research, and is carried out mostly via surgery by transferring founder embryos into pseudo-pregnant recipient females. To cover post-operative analgesic requirements in surrogate mothers, oral self-administration of painkillers has several advantages, but its effectiveness has also been criticized as voluntary ingestion of the drug can be uncertain. Additionally, concerns about potential negative side effects of analgesics on embryo viability and development have been raised. In this regard, we investigated the impact of orally administered analgesia by comparing the outcome of ET with and without paracetamol in the drinking water (3.5mg/ml) of surrogate mothers. Water intake increased significantly when paracetamol, as a sweet-tasting formulation (children's syrup), was added to the drinking water. Measurements of paracetamol concentrations in blood serum confirmed reasonable drug uptake. Success rate of ETs and the body weight of newborn offspring were not different whether paracetamol was administered for two days after surgery or not. In conclusion, paracetamol in drinking water was consumed voluntarily in substantial doses, without detectable side-effects, by freshly operated surrogate mothers, and can therefore be recommended as a feasible method for providing analgesic treatment for surgical ET in mice.

Injection anaesthesia with fentanyl-midazolam-medetomidine in adult female mice: importance of antagonization and perioperative care.

Injection anaesthesia is commonly used in laboratory mice; however, a disadvantage is that post-anaesthesia recovery phases are long. Here, we investigated the potential for shortening the recovery phase after injection anaesthesia with fentanyl-midazolam-medetomidine by antagonization with naloxone-flumazenil-atipamezole. In order to monitor side-effects, the depth of anaesthesia, heart rate (HR), core body temperature (BT) and concentration of blood gases, as well as reflex responses, were assessed during a 50 min anaesthesia. Mice were allowed to recover from the anaesthesia in their home cages either with or without antagonization, while HR, core BT and spontaneous home cage behaviours were recorded for 24 h. Mice lost righting reflex at 330 ± 47 s after intraperitoneal injection of fentanyl-midazolam-medetomidine. During anaesthesia, HR averaged 225 ± 23 beats/min, respiratory rate and core BT reached steady state at 131 ± 15 breaths/min and 34.3 ± 0.25℃, respectively. Positive pedal withdrawal reflex, movement triggered by tail pinch and by toe pinch, still occurred in 25%, 31.2% and 100% of animals, respectively. Arterial blood gas analysis revealed acidosis, hypoxia, hypercapnia and a marked increase in glucose concentration. After anaesthesia reversal by injection with naloxone-flumazenil-atipamezole, animals regained consciousness after 110 ± 18 s and swiftly returned to physiological baseline values, yet they displayed diminished levels of locomotion and disrupted circadian rhythm. Without antagonization, mice showed marked hypothermia (22 ± 1.9℃) and bradycardia (119 ± 69 beats/min) for several hours. Fentanyl-midazolam-medetomidine provided reliable anaesthesia in mice with reasonable intra-anaesthetic side-effects. Post-anaesthetic period and related adverse effects were both reduced substantially by antagonization with naloxone-flumazenil-atipamezole.

Transcatheter direct mitral annuloplasty with Cardioband: feasibility and efficacy trial in an acute preclinical model.

AIMS: The aim of the study was to report preclinical safety and feasibility of a new transcatheter direct mitral annuloplasty intervention in an acute animal model. METHODS AND RESULTS: Twenty healthy pigs underwent Cardioband (Valtech Cardio, Or Yehuda, Israel) transcatheter implantation under intracardiac echocardiographic and fluoroscopic guidance. Through a neo inferior vena cava approach, transseptal access was arranged. The device was delivered into the left atrium using a multi-steerable catheter and fixed to the mitral annulus with multiple helix anchors. Following device cinching, reduction of annular size was evaluated. In all animals the device could be successfully implanted and displayed 100% function, with the average procedure duration and fluoroscopy times being 78±23 minutes and 27±9 minutes, respectively. In total, 246 anchors (average 12.3 per device) were delivered and optimal anchor placement was achieved in 95.1%, while inadequate anchor position (4%) and injury of the coronary sinus or atrium (0.8%) occurred in the rest. Following maximal cinching, diastolic transmitral flow velocity and coaptation lengths were markedly increased (p<0.001), whereas septolateral and intercommissural distances were significantly decreased (p<0.001), when compared to pre-contraction baseline, demonstrating efficient annular reduction by the device. CONCLUSIONS: Transcatheter direct annuloplasty with a surgical-like adjustable device is feasible in the porcine animal model. The humanised porcine model has been instrumental in demonstrating feasibility and in establishing the procedural steps.

Assessment of postsurgical distress and pain in laboratory mice by nest complexity scoring.

Preliminary studies have suggested a correlation between postsurgical pain and nest building behaviour in laboratory mice. However, there is no standardized measure for estimating pain by means of nest building performance. Here, we investigated nest building under various conditions, and scored nest complexity to assess postsurgical pain. Mice of both sexes, different strains [C57BL/6J, DBA/2J, and B6D2-Tg(Pr-mSMalphaActin)V5rCLR-25], and kept under different housing conditions, showed no differences in their latency to use the offered nest material. Healthy female C57BL/6J mice were engaged 4.3% of the day with nest building and showed three peaks of this behaviour: in the beginning and middle of the light phase, and in the second half of the dark phase. For assessment of postsurgical pain, female C57BL/6J mice underwent a sham embryo transfer +/− different doses of the analgesic carprofen or control treatment. Nest complexity scoring at 9 h after the experimental treatments (i.e. at the end of the light phase) resulted in less than 10% of animals with noticeably manipulated nest material (nestlet) after surgery and more than 75% of healthy mice having built identifiable-to-complex nests or had noticeably manipulated nestlets, while animals after anaesthesia-only showed intermediate nest complexity. Carprofen analgesia resulted in no (5 mg/kg) or only slight (50 mg/kg) improvement of nest complexity after surgery. Thus, nest complexity scoring can be incorporated into daily laboratory routine and can be used in mice as a sensitive tool for detecting reduced wellbeing and general condition, but probably not for determining the efficacy of pain treatment.

Transcatheter based electromechanical mapping guided intramyocardial transplantation and in vivo tracking of human stem cell based three dimensional microtissues in the porcine heart.

Stem cells have been repeatedly suggested for cardiac regeneration after myocardial infarction (MI). However, the low retention rate of single cell suspensions limits the efficacy of current therapy concepts so far. Taking advantage of three dimensional (3D) cellular self-assembly prior to transplantation may be beneficial to overcome these limitations. In this pilot study we investigate the principal feasibility of intramyocardial delivery of in-vitro generated stem cell-based 3D microtissues (3D-MTs) in a porcine model. 3D-MTs were generated from iron-oxide (MPIO) labeled human adipose-tissue derived mesenchymal stem cells (ATMSCs) using a modified hanging-drop method. Nine pigs (33 ± 2 kg) comprising seven healthy ones and two with chronic MI in the left ventricle (LV) anterior wall were included. The pigs underwent intramyocardial transplantation of 16 × 10(3) 3D-MTs (1250 cells/MT; accounting for 2 × 10(7) single ATMSCs) into the anterior wall of the healthy pigs (n = 7)/the MI border zone of the infarcted (n = 2) of the LV using a 3D NOGA electromechanical mapping guided, transcatheter based approach. Clinical follow-up (FU) was performed for up to five weeks and in-vivo cell-tracking was performed using serial magnet resonance imaging (MRI). Thereafter, the hearts were harvested and assessed by PCR and immunohistochemistry. Intramyocardial transplantation of human ATMSC based 3D-MTs was successful in eight animals (88.8%) while one pig (without MI) died during the electromechanical mapping due to sudden cardiac-arrest. During FU, no arrhythmogenic, embolic or neurological events occurred in the treated pigs. Serial MRI confirmed the intramyocardial presence of the 3D-MTs by detection of the intracellular iron-oxide MPIOs during FU. Intramyocardial retention of 3D-MTs was confirmed by PCR analysis and was further verified on histology and immunohistochemical analysis. The 3D-MTs appeared to be viable, integrated and showed an intact micro architecture. We demonstrate the principal feasibility and safety of intramyocardial transplantation of in-vitro generated stem cell-based 3D-MTs. Multimodal cell-tracking strategies comprising advanced imaging and in-vitro tools allow for in-vivo monitoring and post-mortem analysis of transplanted 3D-MTs. The concept of 3D cellular self-assembly represents a promising application format as a next generation technology for cell-based myocardial regeneration.

Individual housing of female mice: influence on postsurgical behaviour and recovery.

Individual housing of laboratory mice may increase vulnerability to surgical stress, and interfere with postsurgical recovery. To analyse the effect of housing conditions on recovery, pair- and single-housed female C57BL/6J mice underwent a minor laparotomy +/- analgesia, anaesthesia only or no treatment. Animals were monitored using non-invasive methods during the immediate postsurgical period to assess pain and general impairment. While no appearance or posture abnormalities were observed postexperiment, home cage behaviours were affected distinctly. Discriminant analysis identified self-grooming, locomotion, climbing and resting as mainly responsible for experimental group separation. Behavioural rhythmicity was disrupted, and behaviours related to wellbeing, such as nest building, climbing and burrowing, decreased. Behavioural pain signs (e.g. press) increased. Most behavioural alterations showed a gradation between treatments, e.g. burrowing latency ranged from an intermediate level following anaesthesia only and surgery with analgesia, to pronounced prolongation after surgery without analgesia. Significantly lower burrowing performance after surgery without analgesia in individually-housed animals indicates better recovery in pairs. Social interaction in pairs--an important component of normal behaviour (64%) and a potential indicator for direct social support--was nearly absent (0.3-0.5%). While anaesthesia and surgery resulted in clear changes in behaviour, differences between housing conditions were minor. Hence, despite a tendency towards better recovery in pairs, we found no distinct negative effect of individual housing. In conclusion, both housing conditions are acceptable during the period immediately following minor surgery, though social housing is always preferable in female mice.