Chondroitin Sulfate Proteoglycan 4,6 sulfation regulates sympathetic nerve regeneration after myocardial infarction

Abstract

Sympathetic denervation of the heart following myocardial infarction (MI) is sustained by chondroitin sulfate proteoglycans (CSPGs) in the cardiac scar, and denervation predicts risk of sudden cardiac death. Blocking CSPG signaling restores sympathetic axon outgrowth into the cardiac scar, decreasing arrhythmia susceptibility. Axon growth inhibition by CSPGs is thought to depend on the sulfation status of the glycosaminoglycans (CS-GAGs) attached to the core protein. Tandem sulfation of CS-GAGs at the 4th (4S) and 6th (6S) positions of n-acetyl-galactosamine inhibits outgrowth in several types of neurons within the central nervous system, but it is not known if sulfation is similarly critical during peripheral nerve regeneration. We asked if CSPG sulfation prevented sympathetic axon outgrowth. Sympathetic neurite outgrowth across purified CSPGs is restored in vitro by reducing 4S with the 4-sulfatase enzyme Arylsulfatase-B (ARSB). Additionally, we co-cultured cardiac scar tissue with sympathetic ganglia ex vivo and found that reducing 4S with ARSB restored axon outgrowth to control levels. We examined levels of the enzymes responsible for adding and removing sulfation to CS-GAGs by western blot to determine if they were altered in the left ventricle after MI. We found that CHST15 (4S dependent 6-sulfotransferase) was upregulated, and ARSB was downregulated after MI. Increased CHST15 combined with decreased ARSB suggests a mechanism for production and maintenance of sulfated CSPGs in the cardiac scar. We altered tandem sulfated 4S,6S CS-GAGs in vivo by transient siRNA knockdown of Chst15 and found that reducing 4S,6S restored Tyrosine Hydroxylase (TH) positive sympathetic nerve fibers in the cardiac scar after MI and reduced arrhythmias. Overall, our results suggest that modulating CSPG-sulfation after MI may be a therapeutic target to promote sympathetic nerve regeneration in the cardiac scar and reduce post-MI cardiac arrhythmias.