Biology subjects

Paraskevas, S.

Publications and source records attributed to Paraskevas, S..

5 recordsLinked to original sources

bioRxiv2026

Novel retinoic acid receptor-related orphan receptor gamma inverse agonist enhances checkpoint blockade in hepatocellular carcinoma: a preclinical mouse study

Background and Aims: Neutrophil-associated inflammation is associated with poor outcomes in hepatocellular carcinoma (HCC), but tumour-intrinsic regulators of neutrophil recruitment and checkpoint response remain unclear. We evaluated whether inhibiting retinoic acid receptor-related orphan receptor gamma (RORgamma) alters neutrophil recruitment, programmed death-ligand 1 (PD-L1) expression, and response to PD-L1 blockade. Approach and Results: Survival associations were analysed in The Cancer Genome Atlas HCC cohort. Neutrophil migration toward HCC cells and PD-L1 expression were assessed after RORC silencing or TF-S10 treatment. Immunocompetent mice bearing orthotopic Hep-55.1c tumours received vehicle, TF-S10, anti-PD-L1, or combination treatment. Higher S100A9 and CXCL8 expression was associated with shorter survival (P = 0.000459 and P = 0.0004413, respectively), and tumour-bearing mice had higher neutrophil-to-lymphocyte ratios (P < 0.0001). RORgamma; targeting reduced neutrophil migration and intratumoural Ly6G; infiltration while increasing tumour-cell PD-L1 expression in vitro. Combination treatment reduced endpoint liver tumour volume versus anti-PD-L1 monotherapy (P < 0.01). Half of combination-treated mice had tumour volumes less than or equal to 0.10 cm^3 and none exceeded 1.00 cm^3, compared with 62% of controls exceeding 1.00 cm^3. Histologically, 85% of combination-treated mice had less than 10% liver tumour involvement, versus no controls. Combination treatment also reduced tumour-associated neutrophil populations and peripheral neutrophil-to-lymphocyte ratios. All active treatments reduced pulmonary metastatic burden versus controls, without significant differences among treatment groups; 53% of combination-treated mice had no detectable lung metastases. Conclusions: RORgamma inhibition reduces neutrophil recruitment and increases tumour-cell PD-L1 expression. Combined RORgamma inhibition and PD-L1 blockade improves hepatic tumour control in preclinical HCC and warrants further investigation.

cancer biology↗

bioRxiv2026

Bridging scale-up to transplantation: pluripotent stem cell-derived pancreatic islet encapsulation in emulsion-generated high concentration alginate beads

Pluripotent stem cell-derived pancreatic islet-like cell clusters (SC-islets) have emerged as potential cellular therapy for type 1 diabetes. While SC-islets can theoretically be produced in bioreactors to meet transplantation needs of many recipients per batch, biomanufacturing and translational challenges remain. SC-islet can be cultured in suspension using stirred tank or vertical wheel bioreactors, but these can impart hydrodynamic damage and lead to cellular agglomeration, particularly upon scale-up. We previously described a robust emulsion-based process to encapsulate murine pancreatic beta cells in high-concentration alginate beads which improved graft survival in allogeneic recipients. Here, we present a full pipeline for scalable microencapsulated SC-islet biomanufacturing with extended in vitro bioreactor culture leading into transplantation. We hypothesized that encapsulation would prevent cellular agglomeration, reduce mechanical stress, and preserve differentiation potential during scale-up. Encapsulation of Stage 6 SC-islets prevented cellular agglomeration during extended suspension culture (25 days) and increased cell recovery (91 +/- 3%) compared with non-encapsulated aggregates (60 +/- 10%). No significant differences in glucose-stimulated insulin secretion were observed with vs without microencapsulation. Stage 7 SC-islets matured in the bioreactor were transplanted either as encapsulated islets via the intraperitoneal route or as free SC-islets under the kidney capsule, where they displayed glucose-responsive human C-peptide secretion and remained functional in vivo for up to 98 days. Overall, this work establishes a scalable, robust, transplantation-ready encapsulation platform that supports SC-islet maturation and delivery, providing a generalizable strategy for scaling and transplanting encapsulated organoid systems.

bioengineering↗

bioRxiv2026

β-cell NCK1 is reduced in type 2 diabetes, leading to inefficient β-cell UPR and insulin secretion and revealing sex-specific adaptation during metabolic stress

Type 2 diabetes is characterized by failure of pancreatic {beta} cells to adapt insulin secretion to metabolic demand, due to impaired {beta}-cell function and/or reduced {beta}-cell mass. The unfolded protein response (UPR) is central to this adaptation by maintaining endoplasmic reticulum homeostasis and supporting insulin biosynthesis, secretion, proliferation, and survival. NCK1, is an adaptor protein that regulates diverse cellular processes, including insulin biosynthesis and UPR activation, positioning it at the crossroads of several processes essential for {beta}-cell function. Moreover its silencing is reported to enhance adaptive PERK signaling and {beta}-cell survival in vitro, suggesting that it could represent an important regulator of {beta}-cell adaptation. Here, we explored this potential role for NCK1 using {beta}-cell-specific knockout mice (NCK1{beta}KO) and human islets of both sexes. NCK1 expression was positively regulated by glucose yet reduced in islets from individuals living with type 2 diabetes. Loss of {beta}-cell NCK1 impaired insulin gene expression, insulin content, and glucose-stimulated insulin secretion in vitro, and disrupted UPR activation. In vivo, {beta}-cell NCK1 deletion led to sex-dependent adaptation to maintain glucose homeostasis. Under high-fat/high-sucrose diet, both NCK1{beta}KO male and female mice increased pancreatic insulin content, but only males showed improved insulin secretion associated with islet expansion and {beta}-cell proliferation. Females, in contrast, exhibited impaired insulin secretion despite preserved insulin stores, associated with increased numbers of small islets and altered PERK pathway activation. These findings identify NCK1 as a regulator of {beta}-cell insulin synthesis, secretion, and UPR signaling, and reveal sex-specific adaptive mechanisms to {beta}-cell stress. Reduced NCK1 in islets from people living type 2 diabetes may disrupt {beta}-cell adaptation to metabolic stress and contribute to diabetes.

physiology↗

bioRxiv2026

Extended perfused culture of cm-scale endocrine pancreatic tissues created through sacrificial embedded printing into alginate

Pluripotent stem cells represent a potentially unlimited cell source for the fabrication of human bioartificial tissues to study and treat degenerative conditions such as type 1 diabetes. Alginate is widely used for mammalian cell immobilization and the primary hydrogel studied for pancreatic islet encapsulation. Rheological properties of alginate solutions or fully gelled forms are unsuitable as support matrix for embedded 3D printing. We describe partially gelled self-healing alginate formulations tuned for embedded 3D printing. Perfusable multi-plane hierarchical networks branching into 10 parallel channels, obtained by 3D printing of Pluronic F127 into the alginate support, show high fidelity to computer-assisted models. Therapeutic {beta}-cell doses (40x106 cells/mL) within centimeter-thick perfusable constructs remained viable for at least 1 week of culture under flow, with rapid insulin secretion detected upon glucose challenges. Stem cell-derived islet clusters cultured in 5-channel contructs for 25 days differentiated towards functional insulin-expressing cells. We describe a novel approach to generate cm-scale perfusable endocrine pancreatic constructs using sacrificial embedded 3D printing into alginate. This approach offers an adaptable platform to engineer perfusable cm-scale functional endocrine pancreatic tissues and potentially other vascularized bioartificial tissues.

bioengineering↗

bioRxiv2024

Iterative sacrificial 3D printing and polymer casting to create complex vascular grafts and multi-compartment bioartificial organs

Several emerging strategies to engineer artificial organs employ 3D printing to create vascular templates to provide nutrients and oxygen to immobilized cells. Significant challenges emerge when considering clinical implementation such as immune rejection of allogeneic cell sources, as well as achieving adequate perfusion and integration with endogenous vasculature. We propose a method by which cell-laden hydrogels are molded around ready-made polymeric vascular templates created via 3D printing to create human-scale artificial organs with internal vasculature. We applied this technique to create bioartificial pancreas systems with up to 9 internal flow channels via sacrificial carbohydrate glass 3D printing, porogen-loaded polycarbonate polyurethane dip-coating, followed by casting cell-laden hydrogels around the vascular templates. We optimized porogen size and concentration to maximise the porosity of our scaffolds without compromising mechanical properties, resulting in suture retention strength and compliance respectively matching commercial vascular grafts and native vessels. Bioreactor perfusion studies showed survival and maturation of stem cell derived pancreatic islets without significant differences to traditional suspension culture protocols. Insulin response dynamics were rapid in response to a glucose challenge at the perfusion inlet. Transplantation of the devices as iliac arteriovenous shunts in nondiabetic pigs confirmed safety and patency. These results show promise for the development of an implantable vascularized pancreas for the treatment of type 1 diabetes and demonstrate how bioartificial organs with engineered vascular geometries can be designed for translational applications.

bioengineering↗