Large Animal Studies Research Articles

In vivo characterization of 3D-printed polycaprolactone-hydroxyapatite scaffolds with Voronoi design to advance the concept of scaffold-guided bone regeneration.

Three-dimensional (3D)-printed medical-grade polycaprolactone (mPCL) composite scaffolds have been the first to enable the concept of scaffold-guided bone regeneration (SGBR) from bench to bedside. However, advances in 3D printing technologies now promise next-generation scaffolds such as those with Voronoi tessellation. We hypothesized that the combination of a Voronoi design, applied for the first time to 3D-printed mPCL and ceramic fillers (here hydroxyapatite, HA), would allow slow degradation and high osteogenicity needed to regenerate bone tissue and enhance regenerative properties when mixed with xenograft material. We tested this hypothesis in vitro and in vivo using 3D-printed composite mPCL-HA scaffolds (wt 96%:4%) with the Voronoi design using an ISO 13485 certified additive manufacturing platform. The resulting scaffold porosity was 73% and minimal in vitro degradation (mass loss <1%) was observed over the period of 6months. After loading the scaffolds with different types of fresh sheep xenograft and ectopic implantation in rats for 8weeks, highly vascularized tissue without extensive fibrous encapsulation was found in all mPCL-HA Voronoi scaffolds and endochondral bone formation was observed, with no adverse host-tissue reactions. This study supports the use of mPCL-HA Voronoi scaffolds for further testing in future large preclinical animal studies prior to clinical trials to ultimately successfully advance the SGBR concept.

Advancing autologous urothelial micrografting and composite tubular grafts for future single-staged urogenital reconstructions

Urogenital reconstructive surgery can be impeded by lack of tissue. Further developments within the discipline of tissue engineering may be part of a solution to improve clinical outcomes. In this study, we aimed to design an accessible and easily assembled tubular graft with autologous tissue, which could be constructed and implanted as a single-staged surgical procedure within the premises of an ordinary operating room. The ultimate goals would be to optimize current treatment-options for long-term urinary diversion. Therefore, we evaluated the optimal composition of a collagen-based scaffold with urothelial micrografts in vitro, and followingly implanted the construct in vivo as a bladder conduit. The scaffold was evaluated in relation to cell regeneration, permeability, and biomechanical properties. After establishing an optimized scaffold in vitro, consisting of high-density collagen with submerged autologous micrografts and reinforced with a mesh and stent, the construct was successfully implanted in an in vivo minipig model. The construct assemblance and surgical implantation proved feasible within the timeframe of a routine surgical intervention, and the animal quickly recovered postoperatively. Three weeks post-implantation, the conduit demonstrated good host-integration with a multilayered luminal urothelium. Our findings have encouraged us to support its use in more extensive preclinical large-animal studies.

Aerosolized, Gas-Phase, Intravitreal Methotrexate Reduces Proliferative Vitreoretinopathy in a Randomized Trial in a Porcine Model.

To determine the effectiveness of aerosol-delivered methotrexate (AD-MTx) in a large-animal (porcine) model of proliferative vitreoretinopathy (PVR). Prospective, randomized, interventional, double-masked, controlled, large-animal study with predetermined clinical and histopathologic outcome criteria. Half of the pigs were randomly assigned to receive an identical volume of aerosol-delivered normal saline (AD-NS) using identical delivery systems and treatment intervals. Proliferative vitreoretinopathy was surgically induced in 16 pigs (8 males and 8 females), randomly assigned to receive 2 doses (group A) or 3 doses (group B) of either AD-MTx (1.6 mg/0.4 ml) or normal saline (AD-NS). Group A pigs were euthanized at week 2 (n= 8), and group B pigs were euthanized at week 3 (n= 8). Masked clinical PVR scores (0-6) by a vitreoretinal surgeon and histopathology PVR scores (0-8) by a masked ophthalmic pathologist were used to determine outcomes. The mean, combined clinical and histopathology scores (both anterior and posterior) were used to determine the overall treatment effect between the groups. The mean masked score (± standard deviation) when all grading end points (clinical+ histopathology) were combined was a mean of 8.0 ± 2.3 in the AD-MTx group versus a higher 9.9 ± 2.0 in the AD-NS control group (P= 0.05). The clinical score was 3.88 ± 1.2 in the AD-MTx group versus 4.63 ± 1.6 in the AD-NS group (P= 0.16). The histopathology score for anterior PVR was 2.5 ± 0.8 in the AD-MTx group versus 2.5 ± 0.5 in the AD-NS group (P= 0.50), and the posterior PVR was 1.63 ± 1.6 in the AD-MTx group versus 2.75 ± 1.3 in the AD-NS group (P= 0.07). When the frequency of methotrexate dosing in group A (2 doses) was compared with that in group B (3 doses), the mean score was 8.75 versus 9.13 (P= 0.38), respectively, suggesting an insignificant difference. After surgical induction of PVR in an aggressive, high-risk, large-animal model, AD-MTx reduced posterior PVR formation compared with AD-NS. Additional dosing at week 3 did not improve the outcomes. No difference in anterior PVR formation was noted with intervention. This novel drug delivery system has implications for PVR reduction and warrants further investigation. Proprietary or commercial disclosure may be found after the references.

Rationale of the PAPAartis trial.

Surgical and interventional repair of thoracoabdominal aortic aneurysms improve survival significantly compared to the natural history of the disease. However, both strategies are associated with a substantial risk of spinal cord ischemia, which has been reported to occur-even in contemporary series by expert centers-in up to 12% of patients, depending on the extent of the disease. Following improved neurological outcomes after staged approaches in extensive clinical and long-term large animal studies, and the description of the "collateral network", the concept of "Minimally Invasive Staged Segmental Artery Coil Embolization" (MIS2ACE) was introduced by Etz et al. This concept of priming the collateral network in order to improve spinal cord blood supply showed promising experimental and early clinical outcomes, and consequently led to the initiation of the randomized controlled multicenter PAPAartis trial (Paraplegia Prevention in Aortic Aneurysm Repair by Thoracoabdominal Staging). This Keynote Lecture describes the background and rationale for this trial and gives an update on the current status.

BSLD-01 DYNAMIC CONTROL OF CSF FOR PERSONALIZED CNS DRUG DELIVERY FOR LEPTOMINGEAL DISEASE

Abstract BACKGROUND Appropriate drug distribution is needed for effective treatment of brain metastases and leptomeningeal disease. Lack of proper drug bioavailability in the CNS contributes to the poor outcome in these patients. EnClear Therapies has developed a device delivering intrathecal therapeutics utilizing the dynamic control of CSF flow through an external pump. Herein, we look at intrathecal drug delivery of methotrexate with EnClear’s system and its impact of drug concentration in CNS tissue in sheep and non-human primates. METHODS EnClear’s system utilizes two implantable catheters (lateral ventricle and lumbar thecal sac) and an extracorporeal pump that controls the speed and direction of CSF flow. Software continuously monitors and reports the output from sensor arrays for pressure, respiration, and heart rate. Imaging studies were used to track the CSF flow characteristics during the use of EnClear’s system. Data from administration of methotrexate followed by the use of EnClear’s system was compared to the current standard of delivery, intracerebroventricular or lumbar intrathecal administration. Methotrexate levels were measured with LC/MS in CSF, blood, and multiple CNS and peripheral tissues. RESULTS There was a two-fold increase in methotrexate levels in multiple regions of the brain including deeper brain structures including the striatum using EnClear's system, compared to traditional lumbar intrathecal and intracerebroventricular delivery. Higher peak levels of methotrexate were reached in the CSF with the use of EnClear’s system, with simultaneous reduction in the peripheral nerves and systemic tissues – a potential source of toxicity. CONCLUSIONS In large animal studies, EnClear’s system has for the first time, demonstrated increased brain parenchymal drug levels via intrathecal delivery. Drug delivery with this novel device has shown improved distribution throughout the CNS and leptomeninges as compared to traditional intrathecal drug delivery and decreased peripheral distribution.

Non-invasive monitoring of pulmonary blood flow, functional residual capacity, and shunt index in a porcine model

BackgroundManagement of mechanically ventilated patients can be improved with monitoring of key pulmonary function parameters that facilitate individualization and optimization. The VQm Pulmonary Health Monitor™ (PHM) (Rostrum Medical Innovations Inc., Vancouver, Canada) is a new monitor that continuously measures pulmonary blood flow (PBF), functional residual capacity (FRC) and a novel parameter: shunt index (Qsi). The goal of this study was to provide an initial assessment of the performance of the VQm PHM™ when compared with reference measures of PBF, FRC and intra-pulmonary shunt.MethodsThis was a prospective, experimental, large animal (porcine) study. After baseline measurements, three interventions were performed: increased cardiac output (CO) using dobutamine infusion, increased PEEP (from 4 to 12 cmH2O), and experimental shunt induced by an extracorporeal circuit. PBF, FRC and shunt were measured by the VQm Pulmonary Health Monitor™ (PHM) before and after each intervention. The PHM™ uses sequential gas delivery to deliver targeted alveolar concentrations of CO2 or N2O. PBF and FRC were calculated using CO2 bolus delivery and the modified differential Fick equation. Shunt was estimated from the number of breaths required to eliminate N2O after a 25-breath N2O bolus and expressed as Qsi. PHM™-derived PBF and Qsi were compared to thermodilution CO and calculated Berggren shunt, respectively.ResultsStudies were completed in 19 animals. Measurements of PBF, FRC and Qsi obtained from the PHM™ trended as expected following each intervention. The mean difference between paired values of PBF was − 0.2 ± 0.9 L/min and the 95% limits of agreement were 1.5 and − 1.9 L/min. Concordance was 94.1%. The mean baseline FRC was 1.7 pm 0.4 L and increased to 2.0 pm0.6 L, following the increase in PEEP (p = 0.0078). For shunt (Qsi), the mean values during low (1 L/min) and high (50% of baseline CO) shunt value were 40 pm 4 and 27 pm 5, respectively, p = 0.002.ConclusionsPBF obtained through the modified differential Fick equation and Qsi obtained through N2O uptake and decay dynamics by the VQm PHM™ provide comparable results to reference standards. FRC measurements trended as expected following interventions.

90-OR: 3D Bioprinting of a Bionic Pancreas with a Vascular System—Results of Transplantation in Large Animals

The transplantation of the pancreas is recommended in diabetes with complications. The combination of cell biology and 3D-bioprinting can create organs with vasculature which should be functional. There are issues to be solved: leakage, thrombosis, enhancement against high pressure, connecting organ to the recipient. The purpose of this study was to demonstrate 3D-bioprinting of bionic organs with use of dECM-derived biomaterials with stable flow through the organ in vitro and in large animal studies. Materials and methods: Mathematical analysis in the designed bionic pancreas was performed. The bionic pancreas was 3D bioprinted with 2 types of bioinks. The first tests were carried out in a bioreactor then organs were subjected to magnetic resonance and X-ray with contrast. Pressure endurance tests were performed. 14 pigs received 3D-bioprinted bionic pancreas transplantation. DCL-VESS-Group (n=5) received decellularized vessels as anastomosis. 9 pigs received bionic pancreas with vascular prosthesis as anastomosis - Prosthesis-Group. Pigs were observed up to 14 days. Hist-pat analysis of removed pancreas was performed. Results: The tests carried out in the bioreactor showed the stability of the bionic organ and vascular system in MRI and X-ray examination for 5 days. No leaks beyond the bionic pancreas were observed up to 400 mmHg. In all 14 cases of transplantation there was stable flow throughout the organ after release of vascular clamp. DCL-VESS-Group - In all cases vascular cloth was observed. Prosthesis-Group - In 5 cases there were no complications influencing blood flow through the organ. 4 died to postoperative complications. Stable flow within a transplanted bionic organ was observed till 2 weeks post - surgery within ultrasound examination and radiology intervention. Histopatological showed the presence of CD31 / PECAM-1 (new blood formation). Conclusion: It is feasible to 3Dbioprint and transplant bionic organ in big animal model. Disclosure M.Wszola: Board Member; Polbionica Ltd., Stock/Shareholder; Polbionica Ltd.. K.I.Roszkowicz-ostrowska: None. L.Kownacki: None. J.Wolinski: None. A.Dobrzyn: None. A.Kaminski: None. A.Berman: Board Member; Polbionica Ltd., Stock/Shareholder; Polbionica Ltd. M.Klak: Employee; Polbionica Sp. z o.o., Stock/Shareholder; Polbionica Sp. z o.o. T.Dobrzanski: Employee; Polbionica. O.Janowska: Employee; POLBIONICA Sp. z o.o. S.Domanski: Employee; Polbionica. D.Piatek: None. J.Wejman: None. D.Szkopek: None.

#5615 VALIDITY AND ADEQUACY OF ANIMAL MODELS IN HAEMODIALYSIS RESEARCH: A SYSTEMATIC REVIEW

Abstract Background and Aims with the worldwide dialysis population growing rapidly, improving haemodialysis (HD) outcomes is crucial and HD innovations are urgently needed. Prior to clinical application, novel HD technologies must undergo extensive preclinical testing. However, to date there is no consensus on the induction method of kidney failure in animal models nor which animal species is most suitable for research on HD innovations. Moreover, there is no consensus on which parameters should be used to validate adequate induction of kidney failure and functioning of novel HD devices. Using a systematic review approach, we summarized available literature of HD in kidney failure animal models. Method we performed a systematic search on PubMed and Embase for relevant studies up to February 4th 2022. After removal of duplicates, 5723 abstracts were screened for eligibility by three independent reviewers (JdV, KW and MK). Inclusion was based on publication of a HD intervention in an animal model with adequate (acute or chronic) kidney failure (independent of induction method). Detailed in- and exclusion criteria were registered in PROSPERO 2022 CRD42022307144. Data from individual reports, including animal species and sex, kidney injury parameters and HD details, were extracted in a pre-set data extraction form. Future efforts will focus on risk of bias and key quality indicators. Results of the 5723 abstracts screened, 195 records were included as full text and 41 full text articles were included for data-extraction (Figure 1). Current data extraction is completed for 31 articles (75%). Publication years range from 1973 to 2021 with over half (16/31) published after 2000. HD experiments were most frequently performed on dogs (45%), followed by rats (29%), goats (13%), pigs (10%), and sheep (3%) respectively. Parameters such as weight (81% reported), sex (58%) and age (29%) were not systematically reported. Most studies (67% of records reporting sex) used male animals only. Dog models were found in older studies, with no records after 2009. Other large animal studies were almost exclusively (88%) performed from the year 2007 onwards. Independent of animal species, studies mainly used acute kidney injury (AKI) models (84%), induced by bilateral nephrectomy (48%) or bilateral ureteral ligation (29%). The majority confirmed kidney failure by an increase in BUN and/or plasma creatinine. In AKI models, dialysis was on average initiated on AKI day 2 ± 0.8, and generally either HD (61%) or veno-venous hemofiltration as a single session (35%) was applied. Except for one clearance study, dialysis efficiency was reported as reduction ratios of BUN and/or plasma creatinine. Conclusion both large and small animal models have been used in HD research and preclinical validation of dialysis innovations. The use of dogs to test HD innovations has seen a sharp decline over the past decades, possibly resulting from a shift in public ethical perception. However, large animal models such as goat, sheep and pigs remain popular. A pressing issue appears the overall lack of adequate ARRIVE guideline adherence for reporting of animal parameters. We found no apparent improvement in reporting over time, emphasizing the need for clear guidelines to drive innovative HD research. Further (quantitative) analysis and meta-analysis are ongoing.

Maternal Preconception Glucose Homeostasis and Insulin Resistance Are Associated with Singleton and Twin Birthweight of Neonates Conceived by PCOS Women Undergoing IVF/ICSI Cycles.

Polycystic ovary syndrome (PCOS) can induce fertility and metabolism disorders, which may increase the prevalence of glucose metabolism disorders and cause health hazards to women and their offspring. We aim to evaluate the effect of maternal preconception glucose metabolism on neonatal birthweight in PCOS women undergoing IVF/ICSI cycles. We retrospectively analyzed 269 PCOS women who delivered 190 singletons and 79 twins via IVF/ICSI at a reproductive center. The effects of maternal preconception glucose metabolism indicators on singleton and twin birthweight were evaluated using generalized linear models and generalized estimate equations, respectively. The potential nonlinear associations were evaluated using generalized additive models. The analyses were further stratified by maternal preconception BMI and delivery mode to evaluate the possible interaction effects. Among PCOS women, maternal preconception fasting plasma glucose (FPG) and glycohemoglobin (HbA1c) had significant negative associations with singleton birthweight (all p for trends = 0.04). We also found an overweight-specific association between elevated maternal preconception 2 h plasma insulin (2hPI) and twin birthweight (p for interactions = 0.05) and a caesarean-specific association between maternal preconception HbA1c and singleton birthweight (p for interactions = 0.02) in PCOS women. Maternal preconception glucose metabolism may affect neonatal birthweight, suggesting the importance of preconception glucose and insulin management for PCOS women. Further large prospective cohorts and animal studies are needed to confirm these findings and investigate the potential mechanisms.

Cardiopulmonary haemodynamic effects and gas exchange during apnoeic oxygenation with high-flow and low-flow nasal oxygen: A large animal study.

Apnoeic oxygenation with high-flow nasal oxygen prolongs the safe apnoeic period during induction of general anaesthesia. However, central haemodynamic effects and the characteristics of central gaseous exchange remain unexplored. To describe mean pulmonary arterial pressure along with arterial and mixed venous blood gases and central haemodynamic parameters during apnoeic oxygenation with low-flow and high-flow nasal oxygen in pigs. Experimental crossover study. Animal study of 10 healthy Swedish landrace pigs at Karolinska Institutet, Sweden, April-May 2021. The pigs were anaesthetised, their tracheas intubated and their pulmonary arteries catheterised. The animals were preoxygenated and paralysed before apnoea. Apnoeic periods between 45 and 60 min were implemented with either 70 or 10 l min -1 100% O 2 delivered via nasal catheters. In addition, seven animals underwent an apnoea without fresh gas flow. Cardiopulmonary parameters and blood gases were measured repeatedly. Mean pulmonary arterial pressure during apnoeic oxygenation with high-flow and low-flow oxygen. Nine pigs completed two apnoeic periods of at least 45 min with a Pa O 2 not lower than 13 kPa. The mean pulmonary arterial pressure increased during 45 min of apnoea, from 18 ± 1 to 33 ± 2 mmHg and 18 ± 1 to 35 ± 2 mmHg, at 70 and 10 l min -1 O 2 , respectively ( P < 0.001); there was no difference between the groups ( P = 0.87). The Pa CO 2 increased by 0.48 ± 0.07 and 0.52 ± 0.04 kPa min -1 , at 70 and 10 l min -1 O 2 , respectively; there was no difference between the groups ( P = 0.22). During apnoea without fresh gas flow, the SpO 2 declined to less than 85% after 155 ± 11 s. During apnoeic oxygenation in pigs, the mean pulmonary arterial pressure increased two-fold and Pa CO 2 five-fold after 45 min, while the arterial oxygen levels were maintained over 13 kPa, irrespective of high-flow or low-flow oxygen.

(506) Histology Appears to Reveal Good Sealing of the Human, Porcine and Dog Vasectomy Specimens using the Vasectomy Sealing Procedure

Abstract Introduction The Sealed Vasectomy Procedure (SVP) is a new bipolar radiofrequency sealing tool for transdermal vasectomy in hopes of making the procedure a faster, simpler, and safer office procedure. Testing has been successful in rabbits with excellent wound healing, complete vas occlusion on histology, and no sperm in post-procedural semen analysis. The study data was presented last year as a video submission and subsequently won the AUA 2022’s “Best Video” of the andrology / infertility / sexual dysfunction session. Further studies in larger animals were indicated and FDA approval is needed before its application in humans. The issue of “does the SVP effectively seal the vas deferens in animals larger than a rabbit and in humans?” is a critical one for treating physicians. Objective Evaluate the effectiveness of the SVP in sealing the vas deferens in animals larger than a rabbit and in humans. Methods Animal (rabbit) studies have been previously described. We performed microscopic evaluation of the vas deferens tissues from canines treated with the SVP at either 40 or 60 watts, euthanized at 14 days post-treatment under the conditions of this study. Human vas deferens specimens after removal from the patient were then immediately treated with SVP tool on benchtop. Porcine vas deferens samples were procured and underwent benchtop testing. Specimens were sent for pathologic evaluation by a third-party professional vendor [Crisp County Regional Medical Center in Cordele, Georgia]. Results 4 canine, 6 human, and 26 porcine vas deferens were included in SVP testing. Human, porcine, and canine specimens demonstrated tissue coagulation and fibrosis in the treated vasa deferentia. (See Figure 1 Human and 2 Canine) Conclusions SVP is a potentially exciting novel vasectomy procedure that appears to provide good sealing of the vas deferens in the canine animal model and on benchtop for the human and porcine vas deferens. FDA approval is still needed before use in humans. Disclosure Yes, this is sponsored by industry/sponsor: Signati Medical Clarification Industry initiated, executed and funded study Any of the authors act as a consultant, employee or shareholder of an industry for: Signati Medical.

Chronic Periodontal Infection and Not Iatrogenic Interference Is the Trigger of Medication-Related Osteonecrosis of the Jaw: Insights from a Large Animal Study (PerioBRONJ Pig Model).

Background and Objectives: Antiresorptive drugs are widely used in osteology and oncology. An important adverse effect of these drugs is medication-induced osteonecrosis of the jaw (MRONJ). There is scientific uncertainty about the underlying pathomechanism of MRONJ. A promising theory suspects infectious stimuli and local acidification with adverse effects on osteoclastic activity as crucial steps of MRONJ etiology. Clinical evidence showing a direct association between MRONJ and oral infections, such as periodontitis, without preceding surgical interventions is limited. Large animal models investigating the relationship between periodontitis and MRONJ have not been implemented. It is unclear whether the presence of infectious processes without surgical manipulation can trigger MRONJ. The following research question was formulated: is there a link between chronic oral infectious processes (periodontitis) and the occurrence of MRONJ in the absence of oral surgical procedures? Materials and Methods: A minipig large animal model for bisphosphonate-related ONJ (BRONJ) using 16 Göttingen minipigs divided into 2 groups (intervention/control) was designed and implemented. The intervention group included animals receiving i.v. bisphosphonates (zoledronate, n = 8, 0.05 mg/kg/week: ZOL group). The control group received no antiresorptive drug (n = 8: NON-ZOL group). Periodontitis lesions were induced by established procedures after 3 months of pretreatment (for the maxilla: the creation of an artificial gingival crevice and placement of a periodontal silk suture; for the mandible: the placement of a periodontal silk suture only). The outcomes were evaluated clinically and radiologically for 3 months postoperatively. After euthanasia a detailed histological evaluation was performed. Results: Periodontitis lesions could be induced successfully in all animals (both ZOL and NON-ZOL animals). MRONJ lesions of various stages developed around all periodontitis induction sites in the ZOL animals. The presence of MRONJ and periodontitis was proven clinically, radiologically and histologically. Conclusions: The results of this study provide further evidence that the infectious processes without prior dentoalveolar surgical interventions can trigger MRONJ. Therefore, iatrogenic disruption of the oral mucosa cannot be the decisive step in the pathogenesis of MRONJ.

Deep Penetrating and Sensitive Targeted Magnetic Particle Imaging and Photothermal Therapy of Early-Stage Glioblastoma Based on a Biomimetic Nanoplatform.

Early diagnosis can effectively improve the survival of glioblastoma multiforme (GBM). A specific imaging technique that is simultaneously deep penetrating and sensitive to small tissue changes is desired to identify GBM. Due to its excellent features in signal contrast, detection sensitivity, and none or little attenuation in tissue, magnetic particle imaging (MPI) possesses great potential in cancer diagnosis, especially when the imaging modality is equipped with specifically targeted nanoprobes. However, when gliomas are small, the blood-brain barrier (BBB) is complete and prevents nanoprobes from entering the brain, which negates the theranostic effect. This study proposes a biomimetic nanoplatform that assist the MPI tracers in breaking through the BBB and then demonstrate a targeted and sensitive diagnosis of GBM. Afterward, the photothermal therapy and immune regulationshow an excellent therapeutic effect on the GBM. It is experimentally confirmed that the MPI signal does not decay with tissue depthand shows excellent sensitivity for thousands-cells. Only small animals are conducted in this study due to the limitations of the current commercial MPI scanner, however, this research theoretically enables large animal and human studies, which encourages a promising pathway toward the noninvasive diagnosis of early-stage GBM in clinics.

PO-02-244 AAV6-MEDIATED GFP GENE TRANSFER INDUCES TACHYARRHYTHMIAS IN A PORCINE MODEL OF AV BLOCK

Green fluorescent protein (GFP) is a reporter widely utilized to evaluate gene transfer efficiency and used as control in therapeutic overexpression strategies. Although it is frequently considered to be an innocuous protein, there is also evidence indicating interference with contractile function and electrophysiological properties, potentially leading to cardiac dilation and functional deterioration. Cognizant of these possible limitations, we considered GFP gene transfer to provide a valuable starting point to evaluate local gene transfer and its potential deleterious effects. To evaluate gene transfer efficiency and potential pro-arrhythmic effects in a porcine model of complete atrioventricular block (CAVB). A total of eight pigs were implanted with a pacemaker, after which a radiofrequency ablation CAVB was generated. Four weeks after induction of the model the animals received catheter-based intramyocardial injections of saline, AAV6-CMV-GFP or were not injected at all. All animals were then observed for four more weeks. Monitoring included weekly pacemaker interrogations. Ventricular arrhythmias were assessed by means of increase in maximum heart rates and pacemaker dependency. Four weeks post-ablation all pigs displayed maximal heart rates of 75 ± 16,9 bpm which notably increased in AAV6-CMV-GFP-injected animals two weeks post-treatment (240 ± 10 bpm; Figure A), but not in the control animals (n = 5, 78 ± 29,5 bpm; Figure A). Electronically paced beats showed a remarkable reduction in transduced animals two weeks post-injection (40 ± 37,36 %; Figure B). Although GFP is commonly used in gene transfer studies, our experiments reveal the occurrence of ventricular arrhythmias after treatment. Mechanistically, these arrhythmias could be triggered by local tissue damage stemming from a possible inflammatory response exacerbated by a reaction to the capsid and transgene. As a result, locally injected AAV6-CMV-GFP should not be used as a control transgene in large animal studies to validate cardiac gene therapies.

Three main nitrate reservoirs in the mammalian body

Nitric oxide (NO) is a free radical with multiple functions in virtually all cells. In mammals, NO is generated by nitric oxide synthase from arginine and by reduction of nitrate/nitrite by tissue xanthine oxidoreductase (XOR) or microbiome. Previously, we reported in several mammalian species, including human, that skeletal muscle is a major nitrate reservoir in the body and that nitrate from this reservoir is systemically available and used to generate nitrite and NO. In our recent large animal study, we found that there are similar levels of nitrate stored in bone and skin. In our prior study, skeletal muscle showed a high level of nitrate at 57±39 nmol/g, relative to other tissues, such as liver and plasma at 15±5 nmol/g and 34±19 nmol/g, respectively. Here, we found that the average level of nitrate in the bone is 45±17 nmol/g and in the skin 41±19 nmol/g.Considering that the skeletal muscle, skin and skeleton are three largest organs in mammals, forming about 35-50%, 21% and 14% of body weight, respectively, these results highlight the importance of skeletal muscle as a main nitrate reservoir and the role of skin and bone as new auxiliary nitrate reservoirs. Along with skeletal muscle, the bones and skin likely play an important role in maintaining homeostasis of the nitrate-nitrite-nitric oxide pathway.Also, important from therapeutic point of view, all three organs are well responding to dietary nitrate interventions by increasing their respective nitrate levels, which leads to general increase of systemic and local NO availability and perhaps could be used as one of the ways to increase NO availability in some diseases. This work was supported by intramural NIDDK/NIH grant ZIA DK 025104-15 to Alan N. Schechter. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Bone Marrow Stem Cells with Tissue-Engineered Scaffolds for Large Bone Segmental Defects. A Systematic Review.

Critical-sized bone defects (CSBDs) represent a significant clinical challenge, stimulating researchers to seek new methods for successful bone reconstruction. The aim of this systematic review is to assess whether bone marrow stem cells (BMSCs) combined with tissue-engineered scaffolds have demonstrated improved bone regeneration in the treatment of CSBD in large preclinical animal models. A search of electronic databases (PubMed, Embase, Web of Science, and Cochrane Library) focused on in vivo large animal studies identified 10 articles according to the following inclusion criteria: (1) in vivo large animal models with segmental bone defects; (2) treatment with tissue-engineered scaffolds combined with BMSCs; (3) the presence of a control group; and (4) a minimum of a histological analysis outcome. Animal research: reporting of in Vivo Experiments guidelines were used for quality assessment, and Systematic Review Center for Laboratory animal Experimentation's risk of bias tool was used to define internal validity. The results demonstrated that tissue-engineered scaffolds, either from autografts or allografts, when combined with BMSCs provide improved bone mineralization and bone formation, including a critical role in the remodeling phase of bone healing. BMSC-seeded scaffolds showed improved biomechanical properties and microarchitecture properties of the regenerated bone when compared with untreated and scaffold-alone groups. This review highlights the efficacy of tissue engineering strategies for the repair of extensive bone defects in preclinical large-animal models. In particular, the use of mesenchymal stem cells, combined with bioscaffolds, seems to be a successful method in comparison to cell-free scaffolds.

MP01-12 HISTOLOGY APPEARS TO REVEAL GOOD SEALING OF THE HUMAN, PORCINE AND DOG VASECTOMY SPECIMENS USING THE VASECTOMY SEALING PROCEDURE

You have accessJournal of UrologyCME1 Apr 2023MP01-12 HISTOLOGY APPEARS TO REVEAL GOOD SEALING OF THE HUMAN, PORCINE AND DOG VASECTOMY SPECIMENS USING THE VASECTOMY SEALING PROCEDURE Gerard Henry, Tariq Hakky, Ryan Griggs, Warren Nighan, Alex Fuselier, and Leroy Jones Gerard HenryGerard Henry More articles by this author , Tariq HakkyTariq Hakky More articles by this author , Ryan GriggsRyan Griggs More articles by this author , Warren NighanWarren Nighan More articles by this author , Alex FuselierAlex Fuselier More articles by this author , and Leroy JonesLeroy Jones More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003212.12AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: The Sealed Vasectomy Procedure (SVP) is a new bipolar radiofrequency sealing tool for transdermal vasectomy in hopes of making the procedure a faster, simpler, and safer office procedure. Testing has been successful in rabbits with excellent wound healing, complete vas occlusion on histology, and no sperm in post procedural semen analysis. The study data was presented last year as a video submission and subsequently won the AUA 2022’s “Best Video” of the andrology/infertility/sexual dysfunction session. Further studies in larger animals were indicated and FDA-approval are needed before its application in humans. The issue of “does the SVP effectively seal the vas deferens in animals larger than a rabbit and in humans?” is a critical one for treating physicians. Evaluate the effectiveness of the SVP in sealing the vas deferens in animals larger than a rabbit and in humans. METHODS: Animal (rabbit) studies have been previously described. We preformed microscopic evaluation of the vas deferens tissues from canines treated with the SVP at either 40 or 60 watts, euthanized at 14 days post-treatment under the conditions of this study. Human vas deferens specimens after removal from the patient were then immediately treated with SVP tool on benchtop. Porcine vas deferens samples were procured and underwent benchtop testing. Specimens were sent for pathologic evaluation by a third-party professional vendor [Crisp County Regional Medical Center in Cordele, Georgia]. RESULTS: 4 canine, 6 human, and 26 porcine vas deferens were included in SVP testing. Human, porcine, and canine specimens demonstrated tissue coagulation and fibrosis in the treated vasa deferentia. (See Figure 1 Human and Figure 2 Canine) CONCLUSIONS: SVP is a potentially exciting novel vasectomy procedure that appears to provide good sealing of the vas deferens in the canine animal model and on benchtop for the human and porcine vas deferens. FDA-approval is still needed before use in humans. Source of Funding: Signati © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e6 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Gerard Henry More articles by this author Tariq Hakky More articles by this author Ryan Griggs More articles by this author Warren Nighan More articles by this author Alex Fuselier More articles by this author Leroy Jones More articles by this author Expand All Advertisement PDF downloadLoading ...

A Rabbit Aortic Valve Stenosis Model Induced by Direct Balloon Injury

Animal models are emerging as an important tool to understand the pathologic mechanisms underlying aortic valve stenosis (AVS) because of the lack of access to reliable sources of diseased human aortic valves. Among the various animal models, AVS rabbit models are one of the most commonly used in large animal studies. However, traditional AVS rabbit models require a long-term period of dietary supplementation and genetic manipulation to induce significant stenosis in the aortic valve, limiting their use in experimental studies. To address these limitations, a new AVS rabbit model is proposed, in which stenosis is induced by a direct balloon injury to the aortic valve. The present protocol describes a successful technique for inducing AVS in New Zealand white (NZW) rabbits, with step-by-step procedures for the preparation, the surgical procedure, and the post-operative care. This simple and reproducible model offers a promising approach for studying the initiation and progression of AVS and provides a valuable tool for investigating the underlying pathological mechanisms of the disease.

Technological considerations for delivering focused ultrasound exposures to the spinal cord

Focused ultrasound opening of the blood-brain barrier (BBB) has reached clinical investigations and seems poised to revolutionize the treatment of brain disorders. Far less attention has been paid to the spinal cord and functionally equivalent blood-spinal cord barrier (BSCB). Small and large animal studies demonstrate that ultrasound and microbubbles can be used to modulate the BSCB in a similar manner to the BBB. However, the complex bony geometry of the vertebral column poses a challenge to the non-invasive delivery of therapeutic ultrasound exposures to the spinal canal. This talk will review work developing spine-specific ultrasound pulse trains and identifying optimal apertures for maximizing sound transmission through the vertebral arch.

Chelating the valley of death: Deferoxamine's path from bench to wound clinic.

There is undisputable benefit in translating basic science research concretely into clinical practice, and yet, the vast majority of therapies and treatments fail to achieve approval. The rift between basic research and approved treatment continues to grow, and in cases where a drug is granted approval, the average time from initiation of human trials to regulatory marketing authorization spans almost a decade. Albeit with these hurdles, recent research with deferoxamine (DFO) bodes significant promise as a potential treatment for chronic, radiation-induced soft tissue injury. DFO was originally approved by the Food and Drug Administration (FDA) in 1968 for the treatment of iron overload. However, investigators more recently have posited that its angiogenic and antioxidant properties could be beneficial in treating the hypovascular and reactive-oxygen species-rich tissues seen in chronic wounds and radiation-induced fibrosis (RIF). Small animal experiments of various chronic wound and RIF models confirmed that treatment with DFO improved blood flow and collagen ultrastructure. With a well-established safety profile, and now a strong foundation of basic scientific research that supports its potential use in chronic wounds and RIF, we believe that the next steps required for DFO to achieve FDA marketing approval will include large animal studies and, if those prove successful, human clinical trials. Though these milestones remain, the extensive research thus far leaves hope for DFO to bridge the gap between bench and wound clinic in the near future.