Low molecular weight heparin (LMWH) is widely used in anticoagulation therapies and for the prevention of thrombosis. LMWH is administered by subcutaneous injection usually once or twice per day. This frequent and invasive delivery modality leads to compliance issues for individuals on prolonged therapeutic courses, particularly pediatric patients. Here, we report a long-term delivery method for LMWH via subcutaneous injection of long-lasting hydrogels. LMWH is modified with reactive maleimide groups so that it can be crosslinked into continuous networks with four-arm thiolated polyethylene glycol (PEG-SH). Maleimide-modified LMWH (Mal-LMWH) retains bioactivity as indicated by prolonged coagulation time. Hydrogels comprising PEG-SH and Mal-LMWH degrade via hydrolysis, releasing bioactive LMWH by first-order kinetics with little initial burst release. Separately dissolved Mal-LMWH and PEG-SH solutions were co-injected subcutaneously in New Zealand White rabbits. The injected solutions successfully formed hydrogels in situ and released LMWH as measured via chromogenic assays on plasma samples, with accumulation of LMWH occurring at day two and rising to near-therapeutic dose equivalency by day 5. These results demonstrate the feasibility of using LMWH-containing, crosslinked hydrogels for sustained and controlled release of anticoagulants.

Cerebral palsy (CP) is a static encephalopathy occurring when a lesion to the developing brain results in disordered movement and posture. Patients present with sometimes overlapping spastic, athetoid/dyskinetic, and ataxic symptoms. Spastic CP, which is characterized by stiff muscles, weakness, and poor motor control, accounts for ∼80% of cases. The detailed mechanisms leading to disordered movement in spastic CP are not completely understood, but clinical experience and recent studies suggest involvement of peripheral motor synapses. For example, it is recognized that CP patients have altered sensitivities to drugs that target neuromuscular junctions (NMJs), and protein localization studies suggest that NMJ microanatomy is disrupted in CP. Since CP originates during maturation, we hypothesized that NMJ disruption in spastic CP is associated with retention of an immature neuromotor phenotype later in life. Scoliosis patients with spastic CP or idiopathic disease were enrolled in a prospective, partially-blinded study to evaluate NMJ organization and neuromotor maturation. The localization of synaptic acetylcholine esterase (AChE) relative to postsynaptic acetylcholine receptor (AChR), synaptic laminin β2, and presynaptic vesicle protein 2 (SV2) appeared mismatched in the CP samples; whereas, no significant disruption was found between AChR and SV2. These data suggest that pre- and postsynaptic NMJ components in CP children were appropriately distributed even though AChE and laminin β2 within the synaptic basal lamina appeared disrupted. Follow up electron microscopy indicated that NMJs from CP patients appeared generally mature and similar to controls with some differences present, including deeper postsynaptic folds and reduced presynaptic mitochondria. Analysis of maturational markers, including myosin, syntrophin, myogenin, and AChR subunit expression, and telomere lengths, all indicated similar levels of motor maturation in the two groups. Thus, NMJ disruption in CP was found to principally involve components of the synaptic basal lamina and subtle ultra-structural modifications but appeared unrelated to neuromotor maturational status.