Introduction: Acute liver failure (ALF) and Acute-on-chronic liver failure (ACLF) are treated with liver transplantation. Metabolic liver diseases such as phenylketonuria (PKU) and alpha-1-antitrypsin deficiency (A1ATD) can also be treated with liver transplantation. Due to the limited number of donor organs available, many patients are unable to receive a transplant. Transplant recipients are required to spend a lifetime on immune-suppressing drugs. Pediatric ALF has previously been treated via transplantation of isolated and encapsulated primary human hepatocytes (PHHs) with promising results. Herein we use Bioprinted Tissue Therapeutics (BTTs) containing PHHs and MSCs to rescue immune-competent mice with ALF and to explore the use of hepatic BTTs to treat chronic liver disease. Methods: Hepatic BTTs were created using Aspect Biosystems’ proprietary 3D bioprinting technology. BTTs are comprised of co-aggregated PHHs and MSCs suspended in a combination of solidified semi-permeable, immune-protective hydrogel biomaterials. ALF was induced via injection of CCl4 into immunocompetent mice. BTTs were implanted into either the intraperitoneal (IP) or subcutaneous (SubQ) space. Survival was recorded over 7 days. Healthy NSG mice were also implanted with BTTs to examine release of soluble A1AT. Blood was collected at regular intervals and livers were embedded for histology upon the completion of the study. Results: BTT treatment resulted in a survival rate of 78% (n=15) and 85% (n=7) in IP and SubQ implanted mice respectively, compared to 46% (IP, n=15) and 50% (SubQ, n=8) for mice receiving cell-free controls. SubQ implantation results were confirmed using PHHs from an alternate donor, wherein the survival rate for hepatic BTT-treated mice was 50% (n=8) compared to 13% for mice receiving cell-free controls. Combined the survival rate for hepatic BTT-treated mice was 68% vs 35% for mice receiving cell-free controls (P=0.003). Additionally, BTTs metabolized phenylalanine and produced A1AT. Conclusion: CCl4-induced ALF in immunocompetent mice can be successfully treated with hepatic BTTs containing primary human hepatocytes. Additionally, there is promise that this treatment could be used for chronic metabolic liver diseases. The high level of function of these devices suggests that they could be used to support liver function on an ongoing basis or to act as a bridge to transplantation or recovery. Our next step is to test a scaled up BTT for testing in rats for acute and chronic liver disease.

Transplantation of cadaveric islets can reverse type 1 diabetes (T1D) but requires life long immunosuppression. Cell encapsulation has potential to overcome this challenge by blocking immune cell access to islet grafts while allowing nutrient exchange, glucose sensing and insulin release. In this study, 3D tissues consisting of patterned, multi-shell fibres with an islet-containing core and immunoprotective shell were manufactured using Aspect Biosystems’ microfluidic 3D bioprinting technology for implantation in diabetic rats. Blood glucose and animal body weight were monitored for several months following surgery. Upon retrieval, bioprinted tissues were assayed for islet cell viability and function by glucose-stimulated insulin secretion (GSIS) , followed by histology and immunohistochemistry (IHC) to analyze foreign body response (FBR) and immune cell infiltration. 3D bioprinted implants containing reaggregated allogeneic rat islets re-established normoglycemia for > 90 days in streptozotocin-treated diabetic immunodeficient rats, and > 30 days in diabetic immunocompetent rats. Post-retrieval, tissue implants demonstrate high viability and functionality, with absence of immune cell penetration through the shell, confirming its immune protective properties. Modification of the outer shell biomaterial to reduce FBR was shown to extend islet tissue implant functionality. This is the first study describing a 3D bioprinted implant composed of a core/shell micro-fibre used to deliver therapeutic islets into diabetic animals. The promising results obtained in rodent models open the door to further testing in larger animal models and, eventually, T1D patients. Disclosure V.Russo: None. S.Getsios: None. T.J.Kieffer: Consultant; Fractyl Health, Inc., Sigilon Therapeutics, Inc., Employee; ViaCyte, Inc., Research Support; Aspect Biosystems. S.Wadsworth: None. R.Jalili: None. Y.Yu: None. R.Agarwal: None. S.Pan: None. N.Hakimi: None. K.Jahan: None. E.M.Wilts: None. S.Ida: None. Funding National Research Council Canada - Industrial Research Assistance Program (#935249) , Canadian International Innovation Program (#938664) , Genome British Columbia (#GEN009) , Stem Cell Network (#FY20/FBP-7 Kieffer) , Mitacs (#IT19764) ,

Introduction: Type 1 diabetes (T1D) is a disease characterized by elevated blood glucose due to insufficient insulin release from pancreatic β-cells. Transplantation of cadaveric islets demonstrates that cell therapy can fully reverse hyperglycemia. However, limited cell supply, immune rejection of implanted allogeneic cells, and cell survival represent major challenges. Cell encapsulation has great potential to overcome these challenges by blocking immune cell access to the grafts while allowing nutrient exchange and secreted products from the implanted cells to be delivered to the body. In this study, we use a unique microfluidic bioprinting technology to precisely control the placement of cells and biomaterials within 3D tissues with micro-architectures optimized for cellular fitness and immune protection. Methods: Living tissues consisting of fibres with a cell-containing core and immune-protective alginate-based shell were generated using Aspect Biosystems’ RX1 bioprinter technology. Core-shell fibres with reaggregated primary human pancreatic islets or embryonic stem cell-derived β-cells were tested in vitro using viability and functional (glucose-stimulated insulin secretion, GSIS) assays. Bioprinted tissues were then implanted into the IP space or omentum of streptozotocin (STZ)-induced diabetic mice and rats, respectively. Glucose homeostasis, body weight, and human C-peptide secretion were monitored for up to 3 months following implantion. Retrieved grafts were fixed and analyzed by histology (H&E, Masson’s trichrome stain) and immunohistochemistry (α-SMA, CD45) to quantify fibrotic encapsulation and immune cell infiltration. Results: Bioprinted tissues supported viability and dynamic insulin secretion of cells in vitro up to 28 days. When transplanted into immunodeficient and immunocompetent diabetic rodents, bioprinted tissues containing reaggregated human islets successfully regulated blood glucose for up to 3 months (Figure 1), although normoglycemia was only sustained in a subset of immunocompetent animals and associated with variable fibrosis. Post-retrieval viability stain, assessment of GSIS, and histology revealed high viability and functionality of implanted cells, and the absence of leukocyte infiltration through the shell. Discussion: This is the first study showing a fully 3D bioprinted tissue composed of a core/shell fibre can successfully deliver a therapeutic dose of xenogeneic cells into a diabetic animal (Figure 2). The unique features of the microfluidic technology were leveraged to bioprint an implantable and retrievable tissue patch that merges the benefits of a single fibre (cell fitness, access to nutrients, immune-protection, perm-selectivity) with those provided by a 3D structure (retrievability, ease of implant, structural integrity). The promising results obtained in rodent studies warrant further investigation to minimize fibrosis and explore the scaling-up of bioprinted tissues in large animal models of T1D.NRC-IRAP. NSERC. Stem Cell Network. Genome BC.