Model For Translational Research Research Articles

Abstract 5204: Preclinical models for translational immuno-oncology research: Rare patient-derived xenografts on humanized mice

Abstract Background The preclinical evaluation of novel immune checkpoint modulators is dependent on models with functional human immune cells. In previous experiments, we have demonstrated, that we can use either peripheral blood mononuclear cells (PBMC), subset of immune cells like NK cells or hematopoietic stem cells (HSC) to establish a humanized immune system on highly immunodeficient 1st and 2nd generation NOG mice with functional T-, B-, and NK cells, as well as monocytes and dendritic cells. Furthermore, we determined PD-L1 expression as a predictive marker and target for immunotherapy on different patient-derived xenografts (PDX). By co-transplantation of rare PDX, like leukemia or Nuclear-protein-in-testis (NUT) carcinoma, we successfully generated a fully human tumor-immune-cell model in mice. Finally, we evaluated the functionality of the model by the treatment with checkpoint inhibitors like Ipilimumab (Ipi), Nivolumab (Nivo) or Pembrolizumab (Pembro). In parallel, we investigated the functionality of the human immune cells and evaluated concepts for combination therapies i.e. with chemotherapy or radiation. Methods HSC-humanized mice were generated by i.v. HSC transplantation and engraftment of immune cells was monitored by FACS analysis. For PBMC-humanized mice, immune cells were implanted i.v.. PDX models from different entities (leukemia or NUT carcinoma) were transplanted on HSC-humanized (HIS) mice and treated with Ipi, Nivo and Pembro alone or in combination with radiation. Blood and tumor samples were analysed by FACS for immune cell infiltration and activation. Results The transplanted HSC showed engraftment in mice with proliferation and differentiation. 14 weeks after HSC inoculation up to 20% of the human immune cells in the blood were T-cells, characterized by a high PD-1 expression. We have transplanted PDX from different tumor entities on HSC-humanized mice. Most of the investigated PDX (>70%) successfully engrafted on humanized mice and showed no significant difference in tumor growth compared to growth on non-humanized mice. However, for some PDX we observed a delayed tumor growth or a complete rejection. Engraftment delay seems to correlate with the PD-L1 expression of PDX (the higher PD-L1, then the higher growth delay). Furthermore, our results demonstrate the functionality of the engrafted human immune cells against some PDX. Treatment with Ipi, Nivo or Pembro led to a minor tumor growth delay. Response to checkpoint inhibitors showed a correlation to innate immune response and PD-L1 expression of PDX and could further be increased by combination with radiotherapy. Conclusions Our humanized immune-PDX models enable appropriate preclinical translational research on tumor immune biology and the evaluation of new therapies and combinations, as well as the identification and validation of biomarkers for immune therapy, especially in rare PDX models. Citation Format: Maria Stecklum, Annika Wulf-Goldenberg, Bernadette Brzezicha, Wolfgang Walther, Jens Hoffmann. Preclinical models for translational immuno-oncology research: Rare patient-derived xenografts on humanized mice. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5204.

Abstract 40: NCI patient derived models repository: PDX, organoid and cell lines from the same patient - bridging the translational pipeline

Abstract The National Cancer Institute’s Patient-Derived Models Repository (NCI PDMR; https://pdmr.cancer.gov) has developed a national repository of Patient-Derived Models (PDMs) comprised of patient-derived xenografts (PDXs), in vitro patient-derived tumor cell cultures (PDCs) and cancer associated fibroblasts (CAFs) as well as patient-derived organoids (PDOrg). These PDMs are clinically annotated with molecular information available in an easily accessible database for the extramural community. A key effort in developing these models is to develop matched models sets allowing for larger scale screening efforts using 2D or 3D models to prioritize selection of PDX models for preclinical translational research. To date, over 220 model sets with a PDX, PDOrg, and PDC from a single patient have been developed; 40 of these have matched CAF models allowing for exploration of research questions focused on tumor microenvironment. The largest model sets are in colorectal cancer (COADREAD, n=76), gynecologic cancers (n=33), pancreatic adenocarcinoma (PAAD, n=29), melanoma (MEL, n=19), and head and neck squamous cell carcinomas (HNSCC, n=17). Every model undergoes several quality control assessments that serve as go/no-go criteria including pathology assessment, STR validation, NGS concordance assessment and for PDXs, human:mouse DNA content assessment. It should be noted that not every model is successful in the development or QC phase so additional model sets with only one or two of the model types are also available for researcher requests, for example there are over 125 PDX/PDOrg matched model sets. The NCI is currently performing parallel preclinical screening of PDXs and PDCs or PDOrgs to determine the ability of the in vitro lines to predict in vivo activity. Genetic and histopathologic assessment of these matched model sets have demonstrated a high degree of stability by somatic mutation, copy number alteration (CNA) and gene expression data. Gene expression correlation analysis shows that mean of Spearman r between PDXs 0.89, between matched PDC/PDXs 0.79, and between matched PDOrg/PDXs 0.82. As expected, some variation at the gene expression level when comparing PDX to in vitro cultures by t-SNE can be observed, likely due to the differences in culture conditions. Funded by NCI Contract No. HHSN261200800001E Citation Format: Yvonne A. Evrard, Li Chen, Sergio Alcoser, Gareth Bliss, Carrie Bonomi, Suzanne Borgel, John Carter, Ting-Chia Chang, Alice Chen, Kevin Cooper, Biswajit Das, Kelly Dougherty, Lindsay Dutko, Marion Gibson, Michelle M. Ahalt-Gottholm, Tara Grinnage-Pulley, Keegan Kalmbach, Chris Karlovich, Kimberly Klarmann, Shahanawaz Jiwani, Tiffanie Miner, Michael Mullendore, Matthew Murphy, Kevin Plater, Gloryvee Rivera, Jessica Steed, Luke Stockwin, Cindy R. Timme, Dianne L. Newton, Paul Mickey Williams, Melinda G. Hollingshead, James H. Doroshow. NCI patient derived models repository: PDX, organoid and cell lines from the same patient - bridging the translational pipeline [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 40.

Translational Echocardiography: The Dog as a Clinical Research Model of Cardiac Dysfunction

Heart disease is a major contributor to mortality and disability on a global scale. Hence, there is a need for research to improve non-invasive diagnostic techniques. Diseases in dogs with characteristics very similar to those of human pathologies hold promise as a source of data for evaluating and developing echocardiographic techniques and devices. Methods: We conducted a structured literature search from June 2022 to January 2023 to evaluate the relevance of dogs as a translational model for echocardiographic clinical research. We searched various academic databases, including PubMed Central (PMC), Core, DIGITAL.CSIC, DOAB, DOAJ, EBSCO host, Elsevier B.V, Redib, Scopus, and Web of Science, available through the Academic Information System of the Autonomous University of Baja California. Results: Out of the 243 articles initially screened, we identified 119 relevant articles that met our inclusion criteria for further analysis. This review is an introduction to the canine model by analyzing the cardiovascular anatomical similarities between the two species, the pathophysiological overlaps in some diseases, the parallels in echocardiographic techniques in dogs compared to humans, and the suitability of dogs with a naturally occurring cardiac disease as a model for translational clinical research compared to other animal species. Conclusions: This review emphasizes the importance of canine patients as an ideal cardiac disease symmetrical clinical model since they share common heart diseases with humans. Furthermore, dogs have a shorter lifespan, leading to the relatively rapid evolution of these diseases, which makes studying these pathologies and developing echocardiographic techniques more feasible. The results strongly indicate the need for interdisciplinary collaboration and translational medical research to create innovative echocardiographic technologies and improve the connection between veterinary and human cardiac imaging research.

Experimental mouse models for translational human cancer research.

The development and growth of tumors remains an important and ongoing threat to human life around the world. While advanced therapeutic strategies such as immune checkpoint therapy and CAR-T have achieved astonishing progress in the treatment of both solid and hematological malignancies, the malignant initiation and progression of cancer remains a controversial issue, and further research is urgently required. The experimental animal model not only has great advantages in simulating the occurrence, development, and malignant transformation mechanisms of tumors, but also can be used to evaluate the therapeutic effects of a diverse array of clinical interventions, gradually becoming an indispensable method for cancer research. In this paper, we have reviewed recent research progress in relation to mouse and rat models, focusing on spontaneous, induced, transgenic, and transplantable tumor models, to help guide the future study of malignant mechanisms and tumor prevention.

A Narrative Review of the Applications of Ex-vivo Human Liver Perfusion.

Ex-vivo perfusion describes the extra-corporeal delivery of fluid to an organ or tissue. Although it has been widely studied in the context of organ preservation and transplantation, it has also proven to be an invaluable tool in the development of novel models for translational pre-clinical research. Here, we review the literature reporting ex-vivo human liver perfusion experiments to further understand current perfusion techniques and protocols together with their applications. A computerised search was made of Ovid, MEDLINE, and Embase using the search words "ex-vivo liver or hepatic perfusion". All relevant studies in English describing experiments using ex-vivo perfusion of human livers between 2016 and 2021, inclusive, were included. Of 21 reviewed studies, 19 used ex-vivo human liver perfusion in the context of allogeneic liver transplantation. The quality and size of the studies varied considerably. Human liver perfusion was almost exclusively limited to whole organs and "split" livers, although one study did describe the successful perfusion of tissue sections following a partial hepatectomy. This review of recent literature involving ex-vivo human liver perfusion demonstrates that the technique is not limited to whole liver perfusion. Split-liver perfusion is extremely valuable allowing one lobe to act as a control and increasing the number available for research. This review also highlights the present lack of any reports of segmental liver perfusion.The discarded donor liver is a scarce resource, and the successful use of segmental perfusion has the potential to expand the available experimental models to facilitate pre-clinical experimentation.

Intubation, Central Venous Catheter, and Arterial Line Placement in Swine for Translational Research in Abdominal Transplantation Surgery.

Translational surgical research models in swine are crucial for developing safe preclinical protocols. However, the success of the experimental surgeries does not solely rely on the research team's surgical skills; perioperative care and management procedures, like intubation, central venous line, and arterial line placement, are necessary and of the utmost importance for favorable experiment results. As it is uncommon for research teams to have anesthesiologists or any other staff other than the surgical team, the surgical team involved in translational research must acquire and/or develop the skills to perform the perioperative care. The purpose of this paper is to show the techniques of intubation, central venous catheter, and arterial line placement used and perfected at the Toronto Organ Preservation Laboratory over the last 10 years, to be used as a reference for future researchers joining either this team or any other lab performing translational research protocols in swine and/or abdominal transplantation.

Testing spatial working memory in pigs using an automated T-maze.

Pigs are an important large animal model for translational clinical research but underutilized in behavioral neuroscience. This is due, in part, to a lack of rigorous neurocognitive assessments for pigs. Here, we developed a new automated T-maze for pigs that takes advantage of their natural tendency to alternate. The T-maze has obvious cross-species value having served as a foundation for cognitive theories across species. The maze (17' × 13') was constructed typically and automated with flanking corridors, guillotine doors, cameras, and reward dispensers. We ran nine pigs in (1) a simple alternation task and (2) a delayed spatial alternation task. Our assessment focused on the delayed spatial alternation task which forced pigs to wait for random delays (5, 60, 120, and 240s) and burdened spatial working memory. We also looked at self-paced trial latencies, error types, and coordinate-based video tracking. We found pigs naturally alternated but performance declined steeply across delays (R2 = 0.84). Self-paced delays had no effect on performance suggestive of an active interference model of working memory. Positional and head direction data could differentiate subsequent turns on short but not long delays. Performance levels were stable over weeks in diverse strains and sexes, and thus provide a benchmark for future neurocognitive assessments in pigs.

Synovial Sarcoma Preclinical Modeling: Integrating Transgenic Mouse Models and Patient-Derived Models for Translational Research.

Synovial sarcomas (SyS) are rare malignant tumors predominantly affecting children, adolescents, and young adults. The genetic hallmark of SyS is the t(X;18) translocation encoding the SS18-SSX fusion gene. The fusion protein interacts with both the BAF enhancer and polycomb repressor complexes, and either activates or represses target gene transcription, resulting in genome-wide epigenetic perturbations and altered gene expression. Several experimental in in vivo models, including conditional transgenic mouse models expressing the SS18-SSX fusion protein and spontaneously developing SyS, are available. In addition, patient-derived xenografts have been estab-lished in immunodeficient mice, faithfully reproducing the complex clinical heterogeneity. This review focuses on the main molecular features of SyS and the related preclinical in vivo and in vitro models. We will analyze the different conditional SyS mouse models that, after combination with some of the few other recurrent alterations, such as gains in BCL2, Wnt-β-catenin signaling, FGFR family, or loss of PTEN and SMARCB1, have provided additional insight into the mechanisms of synovial sarcomagenesis. The recent advancements in the understanding of SyS biology and improvements in preclinical modeling pave the way to the development of new epigenetic drugs and immunotherapeutic approaches conducive to new treatment options.

Establishment of a diverse head and neck squamous cancer cell bank using conditional reprogramming culture methods.

Most laboratory models of head and neck squamous cell cancer (HNSCC) rely on established immortalized cell lines, which carry inherent bias due to selection and clonality. We established a robust panel of HNSCC tumor cultures using a "conditional reprogramming" (CR) method, which utilizes a rho kinase inhibitor (Y-27632) and co-culture with irradiated fibroblast (J2 strain) feeder cells to support indefinite tumor cell survival. Sixteen CR cultures were successfully generated from 19 consecutively enrolled ethnically and racially diverse patients with HNSCC at a tertiary care center in the Bronx, NY. Of the 16 CR cultures, 9/16 were derived from the oral cavity, 4/16 were derived from the oropharynx, and 3/16 were from laryngeal carcinomas. Short tandem repeat (STR) profiling was used to validate culture against patient tumor tissue DNA. All CR cultures expressed ΔNp63 and cytokeratin 5/6, which are markers of squamous identity. Human papillomavirus (HPV) testing was assessed utilizing clinical p16 staining on primary tumors, reverse transcription polymerase chain reaction (RT-PCR) of HPV16/18-specific viral oncogenes E6 and E7 in RNA extracted from tumor samples, and HPV DNA sequencing. Three of four oropharyngeal tumors were p16 and HPV-positive and maintained HPV in culture. CR cultures were able to establish three-dimensional spheroid and murine flank and orthotopic tongue models. CR methods can be readily applied to all HNSCC tumors regardless of patient characteristics, disease site, and molecular background, providing a translational research model that properly includes patient and tumor diversity.

Multifocal demyelinating leukoencephalopathy and oligodendroglial lineage cell loss with immune effector cell-associated neurotoxicity syndrome (ICANS) following CD19 CAR T-cell therapy for mantle cell lymphoma.

Immune effector cell-associated neurotoxicity syndrome (ICANS) is a prevalent condition seen after treatment with chimeric antigen receptor T-cell (CAR T) therapy and other cancer cell therapies. The underlying pathophysiology and neuropathology of the clinical syndrome are incompletely understood due to the limited availability of brain tissue evaluation from patient cases, and a lack of high-fidelity preclinical animal models for translational research. Here, we present the cellular and tissue neuropathologic analysis of a patient who experienced grade 4 ICANS after treatment with anti-CD19 CAR T therapy for mantle cell lymphoma. Our pathologic evaluation reveals a pattern of multifocal demyelinating leukoencephalopathy associated with a clinical course of severe ICANS. A focused analysis of glial subtypes further suggests region-specific oligodendrocyte lineage cell loss as a potential cellular and pathophysiologic correlate in severe ICANS. We propose a framework for the continuum of neuropathologic changes thus far reported across ICANS cases. Future elucidation of the mechanistic processes underlying ICANS will be critical in minimizing neurotoxicity following CAR T-cell and related immunotherapy treatments across oncologic and autoimmune diseases.

Gait analysis in swine, sheep, and goats after neurologic injury: a literature review.

Medical research on neurologic ailments requires representative animal models to validate treatments before they are translated to human clinical trials. Rodents are the predominant animal model used in neurological research despite limited anatomic and physiologic similarities to humans. As a result, functional testing designed to assess locomotor recovery after neurologic impairment is well established in rodent models. Comparatively, larger, more clinically relevant models have not been as well studied. To achieve similar locomotor testing standardization in larger animals, the models must be accessible to a wide array of researchers. Non-human primates are the most relevant animal model for translational research, however ethical and financial barriers limit their accessibility. This review focuses on swine, sheep, and goats as large animal alternatives for transitional studies between rodents and non-human primates. The objective of this review is to compare motor testing and data collection methods used in swine, sheep, and goats to encourage testing standardization in these larger animal models. The PubMed database was analyzed by searching combinations of swine, sheep, and goats, neurologic injuries, and functional assessments. Findings were categorized by animal model, data collection method, and assessment design. Swine and sheep were used in the majority of the studies, while only two studies were found using goats. The functional assessments included open pen analysis, treadmill walking, and guided free walking. Data collection methods included subjective behavioral rating scales and objective tools such as pressure-sensitive mats and image-based analysis software. Overall, swine and sheep were well-suited for a variety of assessment designs, with treadmill walking and guided free walking offering the most consistency across multiple trials. Data collection methods varied, but image-based gait analysis software provided the most robust analysis. Future studies should be conducted to standardize functional testing methods after neurologic impairment in large animals.

Morbidity and mortality in elderly dogs – a model for human aging

Over the last decades, canines have experienced a marked increase in their lifespan, mirroring human populations. Several authors have pointed out the domestic dog as a suitable animal model for geropathology translational research. The aim of this study is to assess age-related morbidities and mortality in a population of 269 elderly canines (130 males and 139 females) submitted to necropsy. The organic systems exhibiting the higher number of age-related morbidities were the reproductive, cardiovascular and urinary systems and, in females, also the mammary gland. The prevalence of cardiovascular and urinary disease was significantly higher in males and mammary lesions were exclusively found in females. Urinary disease was more frequent in small breeds dogs, while peritoneum and male genital morbidities were significantly higher in larger breeds. Hyperplastic and degenerative lesions were common morbidities found in this elderly dog population. The main cause of death was neoplasia, which accounted for almost half of the deaths. Cardiovascular and urinary pathology also emerged as a frequent cause of mortality. These findings partially parallel data obtained for human species, displaying cancer and cardiovascular pathology as major causes of disease and death in elderlies. Our data reinforce the potential of the domestic dog for further translational investigations on gerontology, meeting the concept of One Health.

Patient-derived xenograft model in colorectal cancer basic and translational research.

Colorectal cancer (CRC) is one of the most popular malignancies globally, with 930 000 deaths in 2020. The evaluation of CRC-related pathogenesis and the discovery of potential therapeutic targets will be meaningful and helpful for improving CRC treatment. With huge efforts made in past decades, the systematic treatment regimens have been applied to improve the prognosis of CRC patients. However, the sensitivity of CRC to chemotherapy and targeted therapy is different from person to person, which is an important cause of treatment failure. The emergence of patient-derived xenograft (PDX) models shows great potential to alleviate the straits. PDX models possess similar genetic and pathological characteristics as the features of primary tumors. Moreover, PDX has the ability to mimic the tumor microenvironment of the original tumor. Thus, the PDX model is an important tool to screen precise drugs for individualized treatment, seek predictive biomarkers for prognosis supervision, and evaluate the unknown mechanism in basic research. This paper reviews the recent advances in constructed methods and applications of the CRC PDX model, aiming to provide new knowledge for CRC basic research and therapeutics.

BREAKING GROUND IN TRANSLATIONAL GEROSCIENCE: FROM BIOMARKERS TO CLINICAL TRIALS

Abstract The geroscience hypothesis posits that common biological mechanisms of aging drive susceptibility of aged individuals to functional decline, multi-morbidity, and death. The promise of geroscience is that some of these mechanisms may be intervenable, thereby preventing or delaying declines, and providing new therapeutic opportunities for hard-to-treat chronic diseases. This is supported by specific examples of translational research models, and interventions that are at the point of entering human clinical trials. This award presentation will review how we are reimagining existing resources and creating new translational frameworks to test the geroscience hypothesis in humans.

Genetic and Diet-Induced Animal Models for Non-Alcoholic Fatty Liver Disease (NAFLD) Research.

A rapidly increasing incidence of non-alcoholic fatty liver disease (NAFLD) is noted worldwide due to the adoption of western-type lifestyles and eating habits. This makes the understanding of the molecular mechanisms that drive the pathogenesis of this chronic disease and the development of newly approved treatments of utmost necessity. Animal models are indispensable tools for achieving these ends. Although the ideal mouse model for human NAFLD does not exist yet, several models have arisen with the combination of dietary interventions, genetic manipulations and/or administration of chemical substances. Herein, we present the most common mouse models used in the research of NAFLD, either for the whole disease spectrum or for a particular disease stage (e.g., non-alcoholic steatohepatitis). We also discuss the advantages and disadvantages of each model, along with the challenges facing the researchers who aim to develop and use animal models for translational research in NAFLD. Based on these characteristics and the specific study aims/needs, researchers should select the most appropriate model with caution when translating results from animal to human.

A O03 Ex vivo perfusion of isolated human liver segments: the development of a novel model for ethical, translational research

Abstract Background Ex vivo perfusion techniques for human organs, in particular for the human liver, have been extensively studied for decades. Although ex vivo perfusion of human organs has been widely studied in the context of organ preservation and transplantation, it has also proven to be an invaluable tool in the development of novel models for translational pre-clinical research. Not only do these models allow more accurate study of human organ response to noxious external stimuli, but they also represent a far more ethical alternative to live animal experimentation.Although split-liver perfusion of the left or right hepatic lobe is well described in literature, ex vivo perfusion of isolated liver segments has not been previously attempted. A successful hepatic segmental perfusion model would provide a unique opportunity to study inflammation, response to infection and novel therapeutic approaches.The overall aim of this study was to establish an experimental ex vivo hepatic perfusion model on surgically resected human liver segments as a platform to evaluate and study organ preservation and function. The development of an ex vivo perfusion model of human liver segments would produce the ideal platform to study treatment effects without needing to sacrifice animals. Methods Patients were recruited as part of the TIMOLD (Tissue Models for Liver Disease) clinical trial (ClinicalTrials.gov Identifier: NCT05255042; 8/9/2021). Patients recruited were those aged eighteen or over undergoing elective liver resections at a single hepatobiliary unit. Nine human liver segments were retrieved following hemi-hepatectomy for colorectal liver metastases or hepatocellular carcinoma. A healthy segment was resected from the diseased specimen in theatre and a segmental hepatic artery and portal vein was cannulated immediately and flushed with ice cold heparinised preservation solution. Four segments were perfused with 600ml O negative expired red blood cells (HL-RBC) and five segments were perfused with 250ml Oxyglobin diluted with 250ml Volplex (HL-OXY). All segments were administered several drugs to provide metabolic support and optimise perfusion owing to their anticoagulant, antioxidant, and anti-inflammatory properties. Segments were perfused on a bespoke normothermic machine perfusion circuit for 4–6 hours. Perfusion parameters were monitored throughout perfusion and hourly hepatic venous blood gases were taken to monitor glucose and lactate levels. Hourly core biopsies were also taken for H&E staining and interpreted by a consultant histopathologist. Data is reported as the median (range) and statistical analysis has been performed using the Mann-Whitney U test on Graphpad Prism. Results HL-RBC (n=4) had a median pre-perfusion weight of 216 (146–1330) grams whilst HL-OXY (n=5) had a median weight of 269(128–367) grams (p>0.05). HL-RBC segments had a median warm ischaemic (WI) time of 76.5(68–81) minutes and a median cold ischaemic (CI) time of 97 (35–119) minutes. HL-OXY segments had a median WI time of 5 (3–80) minutes and a median CI time of 38 (28–63) minutes (p>0.05). Median portal venous resistance was lower for the HL-OXY group (0.076mmHg/ml/min) in comparison to the HL-RBC group (0.62mmHg/ml/min); p<0.05. There was no significant difference in hepatic arterial resistance between the two groups (HL-RBC 3 mmHg/ml/min; HL-OXY 2.38 mmHg/ml/min; p>0.05). HL-OXY had a lower median lactate of 4.54mmol/L in comparison to HL-RBC (14.7mmol/L) throughout perfusion (p<0.05). HL-OXY segments also had a lower median glucose of 5.9mmol/L compared to 9.65mmol/L for HL-RBC (p<0.05). 1/4 HL-RBC segments had normal histology after 6 hours of perfusion. 2/4 segments showed evidence of necrosis after 2 hours (one with established cirrhosis), and 1/4 segment demonstrated necrosis after 3 hours of perfusion. 4/5 HL-OXY segments demonstrated normal histology after 6 hours of perfusion. One segment, with known cirrhosis, demonstrated necrosis after two hours of perfusion. Conclusions We present here, for the first time, the development of a successful ex vivo isolated hepatic segmental perfusion model. We demonstrate, in the development of this model, that bovine haemoglobin glutamer-200 (Oxyglobin) is a superior oxygen carrier to expired O negative red blood cells in the ex vivo perfusion of isolated human hepatic segments. The HL-OXY group demonstrated a significantly lower portal venous resistance in comparison to HL-RBC, although there was no difference in arterial resistance. Furthermore, HL-OXY segments had lower median glucose and lactate levels during perfusion compared to HL-RBC segments. 4 of 5 segments perfused with Oxyglobin demonstrated viable hepatocytes on histopathological analysis after 6 hours of perfusion. The segment demonstrating early necrosis was one with established cirrhosis on pre-perfusion histology. We describe a protocol for the successful ex vivo perfusion of non-cirrhotic human hepatic segments. Human liver is routinely resected in tertiary hepatobiliary units and much of the specimen discarded. The paradigm presented here identifies a readily available ethical source of human liver with minimal warm ischaemia. This facilitates optimal conditions for translational research, is cost-effective and avoids animal experimentation.

Immunologic and pathologic characterization of a novel swine biomedical research model for eosinophilic esophagitis.

Eosinophilic esophagitis (EoE) is a chronic allergy-mediated condition with an increasing incidence in both children and adults. Despite EoE's strong impact on human health and welfare, there is a large unmet need for treatments with only one recently FDA-approved medication for EoE. The goal of this study was to establish swine as a relevant large animal model for translational biomedical research in EoE with the potential to facilitate development of therapeutics. We recently showed that after intraperitoneal sensitization and oral challenge with the food allergen hen egg white protein (HEWP), swine develop esophageal eosinophilia-a hallmark of human EoE. Herein, we used a similar sensitization and challenge treatment and evaluated immunological and pathological markers associated with human EoE. Our data demonstrate that the incorporated sensitization and challenge treatment induces (i) a systemic T-helper 2 and IgE response, (ii) a local expression of eotaxin-1 and other allergy-related immune markers, (iii) esophageal eosinophilia (>15 eosinophils/0.24 mm2), and (iv) esophageal endoscopic findings including linear furrows and white exudates. Thereby, we demonstrate that our sensitization and oral challenge protocol not only induces the underlying immune markers but also the micro- and macro-pathological hallmarks of human EoE. This swine model for EoE represents a novel relevant large animal model that can drive translational biomedical research to develop urgently needed treatment strategies for EoE.

Cold Preservation of Human Hepatocytes with High Viability.

Freshly isolated human hepatocytes are an important model for translational research, validation of experiments done in animals, and preclinical studies. Human hepatocyte isolation often cannot be carried out easily on demand in common research laboratories, and researchers often collaborate to share hepatocytes or outsource hepatocyte isolations. As a prerequisite for such a strategy, hepatocytes have to maintain their phenotypes after transport. Therefore, this study aimed to determine if overnight storage or shipment of hepatocytes affects their quality when viability, adherence, and cytochrome P450 (CYP) activities are considered. Hepatocytes were stored overnight or shipped to a collaborator in a cold storage solution on wet ice. On the next day, viability of hepatocytes was assessed before plating the cells to determine adherence. Hepatocytes were also cultured in a sandwich culture to determine CYP activities and inducibility. The results showed that although viability (79% ± 0.7% on isolation) was significantly decreased by overnight storage or shipment by 11% (p < 0.001) or 15% (p < 0.001), respectively, the viability of hepatocytes the next day at above 64% ± 2.2% remained sufficiently high for further experiments. In addition, hepatocytes stored for 18 or 24 hours were adherent the next day, and a high confluence of 81% ± 10% to 91% ± 4% was achieved after 48 hours in culture when hepatocytes were adhered on collagen-coated plates. Furthermore, CYP enzyme activities were inducible and not affected by variables such as fibrosis, age, type of operation, steatosis, and body mass index. However, our data would suggest that the type of cancer (primary/secondary), sex (male/female), hypertension, glutamic oxaloacetic transaminase activity, partial thromboplastin time, and size of perfused liver had significant effects (p < 0.05) on induction of some CYP enzymes. In conclusion, human hepatocyte isolation can be carried out at a centralized site and shared between multiple researchers, increasing flexibility and access to a representative human liver in vitro model.

Translational research and service model development based on smart care space for people with severely disability

Translational research and service model development based on smart care space for people with severely disability

Cathepsin S Inhibition Suppresses Experimental Systemic Lupus Erythematosus-Associated Pulmonary Arterial Remodeling.

Patients with systemic lupus erythematosus (SLE) associated with pulmonary arterial hypnertension (PAH) receive targeted therapy for PAH to decrease pulmonary arterial systolic pressure and significantly prolong their survival. Cysteine cathepsin proteases play critical roles in the progression of cardiovascular disease. Inhibition of cathepsin S (Cat S) has been shown to improve SLE and lupus nephritis. However, the effect of Cat S inhibitors on SLE-associated PAH (SLE-PAH) remains unclear, and there is no animal model for translational research on SLE-PAH. We hypothesized that the inhibition of Cat S may affect PAH development and arterial remodeling associated with SLE. A female animal model of SLE-PAH, female MRL/lpr (Lupus), was used to evaluate the role of pulmonary arterial remodeling in SLE. The key finding of the research work is the establishment of an animal model of SLE associated with PAH in female MRL/lpr mice that is able to evaluate pulmonary arterial remodeling starting from the age of 11 weeks to 15 weeks. Cat S protein level was identified as a marker of experimental SLE. Pulmonary hypertension in female MRL/lpr (Lupus) mice was treated by administering the selective Cat S inhibitor Millipore-219393, which stimulated peroxisome proliferator-activated receptor-gamma (PPARγ) in the lungs to inhibit Cat S expression and pulmonary arterial remodeling. Studies provide an animal model of female MRL/lpr (Lupus) associated with PAH and a deeper understanding of the pathogenesis of SLE-PAH. The results may define the role of cathepsin S in preventing progressive and fatal SLE-PAH and provide approaches for therapeutic interventions in SLE-PAH.