Hydrogel-based delivery of Tat-fused protein Hsp70 protects dopaminergic cells in vitro and in a mouse model of Parkinson’s disease

Abstract

Neurodegenerative disorders such as Parkinson's disease (PD) have no effective therapies. However, many promising drugs are precluded from clinical trials because of their poor brain availability. The chaperone protein Hsp70 has been reported to be effective in PD models, but its brain targeting is challenging. We developed a novel brain Hsp70 delivery system using injectable, biocompatible, and biodegradable semi-interpenetrating polymer networks of collagen (COLL) and low-molecular-weight hyaluronic acid (LMW HA) structured with gelatin particles. We produced human recombinant Hsp70-1A fused with the cell-penetrating peptide Tat (Tat-Hsp70) that was neuroprotective in vitro against the dopaminergic toxin 6-hydroxydopamine (6-OHDA). We assessed Tat-Hsp70 release from the selected COLL-LMW HA composites in vitro, observing a 95% release of loaded protein after 96 h. The release kinetics FITTED the Korsmeyer-Peppas model (regression coefficient 0.98) and the released Tat-Hsp70 remained neuroprotective for SH-SY5Y cells. Magnetic resonance imaging revealed that COLL-LMW HA composites lasted at least 96 h at the brain level, and in vivo Tat-Hsp70 release studies indicated that hydrogel presence is pivotal for a spatially focused neuroprotective effect. In an in vivo model of dopaminergic degeneration, Tat-Hsp70-loaded composites conveyed neuroprotection at both the behavioral and dopaminergic neuronal levels against the striatal injection of 6-OHDA. After the injection of Tat-Hsp70-loaded composites, mice showed a transient inflammatory response, with a decrease in GFAP and CD11b immunostaining after 7 days. Our delivery system enabled the effective brain release of Tat-Hsp70 and is ready for further improvements.

Advances in Tissue Engineering and Innovative Fabrication Techniques for 3-D-Structures: Translational Applications in Neurodegenerative Diseases

“…Systems have been developed for the administration of neurotrophic factors, already partially studied as treatment of the diseases, such as GDNF-loaded microspheres stereotaxically implanted in brain of PD-affected animals [ 158 , 191 , 192 ]. Hydrogels have also been used for the delivery of less canonical therapeutic agents, such as Tat-fused protein Heat Shock Protein 70 (Hsp70) [ 193 ], activin-B [ 194 ], or even the secretome of mesenchymal stem cells [ 195 ]. In comparison to canonical drug delivery, cell therapy is also a promising approach for PD therapy [ 196 , 197 ].…”

A miniaturized hydrogel-based in vitro model for dynamic culturing of human cells overexpressing beta-amyloid precursor protein

“…To obtain a 3D model of brain cells to be hosted in the device and able to produce high levels of amyloid fragments, thus favoring their accumulation, we exploited COLL-based semi-IPNs already investigated in our lab 15,16 and studied their scaling down from 5.10 to 1.25 mm thick. Because of its design, the thinnest condition is the most suitable to be hosted in the device, also to ensure an optimal optical accessibility during perfusion.…”

“…Both hydrogels were previously developed and tested for biocompatibility with brain cells. 15 , 16 To obtain finely tunable matrices with enhanced controllability and reproducibility, 15 , 16 both hydrogels were conceived as semi-interpenetrating polymer networks (semi-IPNs).…”

Recent Advances in Natural Gum-Based Biomaterials for Tissue Engineering and Regenerative Medicine: A Review

“…Regeneration of nerves is a complex phenomenon of the biological system, where small injuries can be regenerated on their own, but large injuries need proper surgical treatment [206][207][208][209]. For large nerve repair, several approaches have been investigated for peripheral nerve regeneration and repairing spinal cord [206].…”