Vastly Extended Drug Release from Poly(pro-17β-estradiol) Materials Facilitates in Vitro Neurotrophism and Neuroprotection

Central nervous system (CNS) injuries persist for years, and currently there are no therapeutics that can address the complex injury cascade that develops over this timescale. 17β-estradiol (E2) has broad tropism within the CNS, targeting and inducing beneficial phenotypic changes in myriad cells following injury. To address the unmet need for vastly prolonged E2 release, we report first-generation poly(pro-E2) biomaterial scaffolds that release E2 at nanomolar concentrations over the course of 1–10 years via slow hydrolysis in vitro. As a result of their finely tuned properties, these scaffolds demonstrate the ability to promote and guide neurite extension ex vivo and protect neurons from oxidative stress in vitro. The design and testing of these materials reported herein demonstrate the first step towards next-generation implantable biomaterials with prolonged release and excellent regenerative potential

Development of a Slow-Degrading Polymerized Curcumin Coating for Intracortical Microelectrodes

Intracortical microelectrodes are used with brain-computer interfaces to restore lost limb function following nervous system injury. While promising, recording ability of intracortical microelectrodes diminishes over time due, in part, to neuroinflammation. As curcumin has demonstrated neuroprotection through anti-inflammatory activity, we fabricated a 300 nm-thick intracortical microelectrode coating consisting of a polyurethane copolymer of curcumin and polyethylene glycol (PEG), denoted as poly (curcumin-PEG1000 carbamate) (PCPC). The uniform PCPC coating reduced silicon wafer hardness by two orders of magnitude and readily absorbed water within minutes, demonstrating that the coating is soft and hydrophilic in nature. Using an in vitro release model, curcumin eluted from the PCPC coating into the supernatant over 1 week; the majority of the coating was intact after an 8-week incubation in buffer, demonstrating potential for longer term curcumin release and softness. Assessing the efficacy of PCPC within a rat intracortical microelectrode model in vivo, there were no significant differences in tissue inflammation, scarring, neuron viability, and myelin damage between the uncoated and PCPC-coated probes. As the first study to implant nonfunctional probes with a polymerized curcumin coating, we have demonstrated the biocompatibility of a PCPC coating and presented a starting point in the design of poly(pro-curcumin) polymers as coating materials for intracortical electrodes

Characterization of Fibroblast Cells Derived from a Patient with Infantile Neuronal Ceroid Lipofuscinosis

Infantile Neuronal Ceroid Lipofuscinosis (INCL) is a pediatric neurodegenerative disorder characterized by progressive retinal and central nervous system deterioration during infancy. This lysosomal storage disorder results from deficiency in the Palmitoyl Protein Thioesterase 1 (PPT1) enzyme; a lysosomal hydrolase which cleaves fatty acid chains such as palmitate from lipid-modified proteins. In the absence of PPT1 activity, these proteins fail to be degraded, leading to the accumulation of autofluorescent storage material in the lysosome. The underlying molecular mechanisms leading to INCL pathology remain poorly understood. A role for oxidative stress has been postulated, yet little evidence has been reported to support this possibility. Here we report the findings of a comprehensive characterization of human PPT1- deficient fibroblast cells harboring Met1Ile and Tyr247His compound heterozygous mutations. Autofluorescent storage material, characteristic of INCL, was identified in human fibroblast cells. We observed distinct organellar abnormalities of the lysosomal and mitochondrial structures, which supports previous postulations about endoplasmic reticulum and mitochondrial-mediated pathologies. There was an abundance of lysosomal compartments in the body of INCL fibroblasts, which suggests an upregulation of lysosomal biogenesis, and is known to be associated with endoplasmic reticulum stress. The mitochondrial network displayed a morphology consistent with mitochondrial dysfunction, which supports a role for the mitochondria and oxidative stress in INCL cell death. We then showed, for the first time, that human INCL fibroblasts have a heightened susceptibility to exogenous reactive oxygen species (ROS)-induced cell death, which suggests elevated basal levels of endogenous ROS in the cell. These findings offer evidence in support of a promising direction for future work investigating the molecular mechanisms underlying INCL pathology

Human INCL fibroblasts display abnormal mitochondrial and lysosomal networks and heightened susceptibility to ROS-induced cell death

AbstractInfantile Neuronal Ceroid Lipofuscinosis (INCL) is a pediatric neurodegenerative disorder characterized by progressive retinal and central nervous system deterioration during infancy. This lysosomal storage disorder results from a deficiency in the Palmitoyl Protein Thioesterase 1 (PPT1) enzyme - a lysosomal hydrolase which cleaves fatty acid chains such as palmitate from lipid-modified proteins. In the absence of PPT1 activity, these proteins fail to be degraded, leading to the accumulation of autofluorescence storage material in the lysosome. The underlying molecular mechanisms leading to INCL pathology remain poorly understood. A role for oxidative stress has been postulated, yet little evidence has been reported to support this possibility. Here we present a comprehensive cellular characterization of human PPT1-deficient fibroblast cells harboring Met1Ile and Tyr247His compound heterozygous mutations. We detected autofluorescence storage material and observed distinct organellar abnormalities of the lysosomal and mitochondrial structures, which supported previous postulations about the role of ER, mitochondria and oxidative stress in INCL. An increase in the number of lysosomal structures was found in INCL patient fibroblasts, which suggested an upregulation of lysosomal biogenesis, and an association with endoplasmic reticulum stress response. The mitochondrial network also displayed abnormal spherical punctate morphology instead of normal elongated tubules with extensive branching, supporting the involvement of mitochondrial and oxidative stress in INCL cell death. Autofluorescence accumulation and lysosomal pathologies can be mitigated in the presence of conditioned wild type media suggesting that a partial restoration via passive introduction of the enzyme into the cellular environment may be possible. We also demonstrated, for the first time, that human INCL fibroblasts have a heightened susceptibility to exogenous reactive oxygen species (ROS)-induced cell death, which suggested an elevated basal level of endogenous ROS in the mutant cell. Collectively, these findings support the role of intracellular organellar networks in INCL pathology, possibly due to oxidative stress.</jats:p