An oxygen-delivering Hyaluronic acid-based matrix reduces inflammation and promotes human islet survival in macroencapsulation devices
Heide Brandhorst1, Daniel Brandhorst1, Daniel A Domingo-Lopez2, Eoin O'Cearbhaill3, Liam McDonough4, Fergal B Coulter3, Stefan Deotti3, Helena Kelly4, Garry Duffy2, Paul RV Johnson1.
1Nuffield Department of Surgical Sciences, University of Oxford, Oxford, United Kingdom; 2Anatomy and Regenerative Medicine Institute (REMEDI), National University of Ireland, Galway, Ireland; 3School of Mechanical Engineering, University College of Dublin (UCD), Dublin, Ireland; 4School of Pharmacy & Biomolecular Sciences, Royal College of Surgeons in Ireland (RCSI), Dublin, Ireland
Introduction: Encapsulation of isolated islets has the potential to enable transplantation without the requirement for life-long immunosuppression. However, to date, insufficient oxygen supply limits graft survival of macroencapsulated islets. The aim of this study was to develop a bio-compatible matrix that can deliver oxygen to facilitate early graft survival and providing a replacement for the collagenase-degraded extracellular matrix.
Methods: Isolated human islets (n = 7) were subjected to quality assessment comprising yield, counted as islet particle number (IN) and islet equivalents (IEQ); viability determined by FDA-PI; early apoptosis measured by Annexin-V and reactive oxygen species (ROS) production evaluated by DCFH-DA. Afterwards, islets were mixed with different matrices: (A) supplemented CMRL (sham-treated control); (B) native Hyaluronic-acid (HA)-based Gel; (C) Beta (β)-Gel (HA-Gel+Perfluorodecalin-emulsion) or (D) pre-oxygenated ß-Gel (β-Gel+O2) and subsequently infused into silicone-based 3D-printed macrodevices charged with 600 matrix-immersed IEQ and cultured for 4–5 d in normoxia. After culture, islet characterization was performed as described above. Parameters were corrected for IEQ and normalized to preculture (PC) data (mean ± SEM).
Results: Post culture, a massive loss of islets was noted when embedded in CMRL (11 ± 3% recovery, p<0.001 vs PC, vs β-Gel+O2 [81 ± 7%]; p<0.01 vs β-Gel [63 ± 4%]) or HA-Gel (35 ± 8%, p<0.01). Reduction of islet yield correlated with substantially enhanced fragmentation (ratio of IN over IEQ) (421 ± 70%, p<0.001 vs PC; 283 ± 80%, p<0.01; 158 ± 11%, p<0.05; 152 ± 22%, NS) in CMRL, HA-Gel, β-Gel or β-Gel+O2, respectively. Accumulation of cell fragments was associated with increased islet ROS production (733 ± 403%; 497 ± 293%, p<0.05 vs CMRL; 177 ± 89%, p<0.01; 140 ± 64%, p<0.01) after encapsulation in CMRL, HA-Gel, β-Gel and β-Gel+O2, respectively. Compared with CMRL (75 ± 4%, p<0.001 vs PC, vs β-Gel+O2) and HA-Gel (84 ± 2%, p<0.05 vs PC, vs β-Gel+O2) viability was significantly better preserved in β-Gel (84 ± 3%, p<0.05 vs PC, vs CMRL) and β-Gel+O2 (92 ± 5%).
Early apoptosis was highest in CMRL (1095 ± 228%, p<0.001 vs PC, vs β-Gel+O2) and HA-Gel (716 ± 193%, p<0.01 vs PC, p<0.05 vs CMRL) but lower in β-Gel (345 ± 62%, p<0.05 vs PC, vs CMRL) or β-Gel+O2 (337 ± 63%).
Overall survival, defined as survival of viable cells only, was marginal in CMRL (8 ± 2%, p<0.001 vs PC, vs β-Gel+O2), low in HA-Gel (29 ± 7%, p<0.001 vs PC) but higher in β-Gel (54 ± 6.3%, p<0.05 vs PC, p<0.01 vs CMRL) and β-Gel+O2 (75 ± 7%, p<0.01 vs HA-Gel).
Conclusions: Our findings demonstrate that the use of a suitable bio-compatible matrix is essential to reduce hypoxia-induced inflammation and to protect the integrity of macro-encapsulated human islets. As hypoxia is the most decisive factor for islet survival, the efficient delivery of oxygen, even for a limited time, helps to promote islet survival within macrodevices.
Juvenile Diabetes Research Foundation (31-2008-617). Diabetes Research and Wellness Foundation. Oxford NIHR Biomedical Research Centre Theme. European Union’s Horizon 2020 (645991).
