Macrophage depletion in stellate ganglia alleviates cardiac sympathetic overactivation and ventricular arrhythmogenesis by attenuating neuroinflammation in heart failure.

Cardiac sympathetic overactivation is involved in arrhythmogenesis in patients with chronic heart failure (CHF). Inflammatory infiltration in the stellate ganglion (SG) is a critical factor for cardiac sympathoexcitation in patients with ventricular arrhythmias. This study aims to investigate if macrophage depletion in SGs decreases cardiac sympathetic overactivation and ventricular arrhythmogenesis in CHF. Surgical ligation of the coronary artery was used for induction of CHF. Clodronate liposomes were microinjected into bilateral SGs of CHF rats for macrophage depletion. Using cytokine array, immunofluorescence staining, and Western blot analysis, we found that macrophage expansion and expression of TNFα and IL-1ß in SGs were markedly increased in CHF rats. Flow cytometry data confirmed that the percentage of macrophages in SGs was higher in CHF rats than that in sham rats. Clodronate liposomes significantly reduced CHF-elevated proinflammatory cytokine levels and macrophage expansion in SGs. Clodronate liposomes also reduced CHF-increased N-type Ca2+ currents and excitability of cardiac sympathetic postganglionic neurons and inhibited CHF-enhanced cardiac sympathetic nerve activity. ECG data from 24-h, continuous telemetry recording in conscious rats demonstrated that clodronate liposomes not only restored CHF-induced heterogeneity of ventricular electrical activities, but also decreased the incidence and duration of ventricular tachycardia/fibrillation in CHF. Macrophage depletion with clodronate liposomes attenuated CHF-induced cardiac sympathetic overactivation and ventricular arrhythmias through reduction of macrophage expansion and neuroinflammation in SGs.

Red blood cell membrane-camouflaged poly(lactic-co-glycolic acid) microparticles as a potential controlled release drug delivery system for local stellate ganglion microinjection.

The stellate ganglion (SG) is a part of the sympathetic nervous system that has important regulatory effects on several human tissues and organs in the upper body. SG block and intervention have been clinically and preclinically implemented to manage chronic pain in the upper extremities, neck, head, and upper chest as well as chronic heart failure. However, there has been very limited effort to develop and investigate polymer-based drug delivery systems for local delivery to the SG. In this study, we fabricated red blood cell (RBC) membrane-camouflaged poly(lactic-co-glycolic acid) (PLGA) (PLGAM) microparticles for use as a potential long-term controlled release system for local drug delivery. The structure, size, and surface zeta potential results indicated that the spherical PLGAM microparticles were successfully fabricated. Both PLGA and PLGAM microparticles exhibited biocompatibility with human adipose mesenchymal stem cells (ADMSC) and satellite glial cells and showed hemocompatibility. In addition, both PLGA and PLGAM displayed no significant effects on the secretion of proinflammatory cytokines by human monocyte derived macrophages in vitro. We microinjected microparticles into rat SGs and evaluated the retention time of microparticles and the effects of the microparticles on inflammation in vivo over 21 days. Subsequently, we fabricated drug-loaded PLGAM microparticles by using GW2580, a colony stimulating factor-1 receptor inhibitor, as a model drug and assessed its encapsulation efficiency, drug release profiles, biocompatibility, and anti-inflammatory effects in vitro. Our results demonstrated the potential of PLGAM microparticles for long-term controlled local drug release in the SG. STATEMENT OF SIGNIFICANCE: SG block by locally injecting therapeutics to inhibit the activity of the sympathetic nerves provides a valuable benefit to manage chronic pain and chronic heart failure. We describe the fabrication of RBC membrane-camouflaged PLGA microparticles with cytocompatibility, hemocompatibility, and low immunogenicity, and demonstrate that they can be successfully and safely microinjected into rat SGs. The microparticle retention time within SG is over 21 days without eliciting detectable inflammation. Furthermore, we incorporate a CSF-1R inhibitor as a model drug and demonstrate the capacities of long-term drug release and regulation of macrophage functions. The strategies demonstrate the feasibility to locally microinject therapeutics loaded microparticles into SGs and pave the way for further efficacy and disease treatment evaluation.

Multifunctional Microgel-Based Cream Hydrogels for Postoperative Abdominal Adhesion Prevention.

Postoperative abdominal adhesions are a common problem after surgery and can produce serious complications. Current antiadhesive strategies focus mostly on physical barriers and are unsatisfactory and inefficient. In this study, we designed and synthesized advanced injectable cream-like hydrogels with multiple functionalities, including rapid gelation, self-healing, antioxidation, anti-inflammation, and anti-cell adhesion. The multifunctional hydrogels were facilely formed by the conjugation reaction of epigallocatechin-3-gallate (EGCG) and hyaluronic acid (HA)-based microgels and poly(vinyl alcohol) (PVA) based on the dynamic boronic ester bond. The physicochemical properties of the hydrogels including antioxidative and anti-inflammatory activities were systematically characterized. A mouse cecum-abdominal wall adhesion model was implemented to investigate the efficacy of our microgel-based hydrogels in preventing postoperative abdominal adhesions. The hydrogels, with a high molecular weight HA, significantly decreased the inflammation, oxidative stress, and fibrosis and reduced the abdominal adhesion formation, compared to the commercial Seprafilm group or Injury-only group. Label-free quantitative proteomics analysis demonstrated that S100A8 and S100A9 expressions were associated with adhesion formation; the microgel-containing hydrogels inhibited these expressions. The microgel-containing hydrogels with multifunctionality decreased the formation of postoperative intra-abdominal adhesions in a murine model, demonstrating promise for clinical applications.

Morphological Regeneration and Functional Recovery of Neuromuscular Junctions after Tourniquet-Induced Injuries in Mouse Hindlimb.

Tourniquet application and its subsequent release cause serious injuries to the skeletal muscle, nerve, and neuromuscular junction (NMJ) due to mechanical compression and ischemia-reperfusion (IR). Monitoring structural and functional repair of the NMJ, nerve, and skeletal muscle after tourniquet-induced injuries is beneficial in exploring potential cellular and molecular mechanisms responsible for tourniquet-induced injuries, and for establishing effective therapeutic interventions. Here, we observed long-term morphological and functional changes of the NMJ in a murine model of tourniquet-induced hindlimb injuries. Unilateral hindlimbs of C57/BL6 mice were subjected to 3 h of tourniquet by placing an orthodontic rubber band, followed by varied periods of tourniquet release (1 day, 3 days, 1 week, 2 weeks, 4 weeks, and 6 weeks). NMJ morphology in the gastrocnemius muscle was imaged, and the endplate potential (EPP) was recorded to evaluate NMJ function. In NMJs, nicotinic acetylcholine receptor (nAChR) clusters normally displayed an intact, pretzel-like shape, and all nAChR clusters were innervated (100%) by motor nerve terminals. During 3 h of tourniquet application and varied periods of tourniquet release, NMJs in the gastrocnemius muscle were characterized by morphological and functional changes. At 1 day and 3 days of tourniquet release, nAChR clusters retained normal, pretzel-like shapes, whereas motor nerve terminals were completely destroyed and no EPPs recorded. From 1 to 6 weeks of tourniquet release, motor nerve terminals gradually regenerated, even reaching that seen in sham mice, whereas nAChR clusters were gradually fragmented with prolongation of tourniquet release. Additionally, the amplitude of EPPs gradually increased with prolongation of tourniquet release. However, even at 6 weeks after tourniquet release, the amplitude of EPPs did not restore to the level seen in sham mice (13.9 ± 1.1 mV, p < 0.05 vs. sham mice, 29.8 ± 1.0 mV). The data suggest that tourniquet application and subsequent release impair the structure and function of NMJs. Morphological change in motor nerve terminals is faster than in nAChR clusters in NMJs. Slow restoration of fragmented nAChR clusters possibly dampens neuromuscular transmission during the long phase following tourniquet release.