The impact of long-course chemoradiotherapy on the myenteric plexus, neuromuscular functions and responses to prokinetic drugs in the human rectum.

Abstract

The long-term effects of chemoradiotherapy on human rectum are poorly understood. The aims were to investigate changes in inflammatory status, myenteric neuron numbers/phenotype, neuromuscular functions and prokinetic drug efficacy. Macroscopically normal proximal-to-mid rectum was obtained from 21 patients undergoing surgery for bowel cancer, 98days (range: 63-350) after concurrent capecitabine and pelvic radiotherapy, and 19 patients without chemoradiotherapy. Inflammatory status was measured by H&E, CD45 staining and qPCR. Myenteric neurons were examined by immunohistochemistry. Neuromuscular functions and drug efficacy were studied using exogenous agents and electrical field stimulation (EFS) to activate intrinsic nerves. Inflammation was not detected. Numbers of myenteric ganglia/neurons were unchanged (11.7±2.4 vs. 10.3±2.2 neurons/mm myenteric plexus with/without chemoradiotherapy) as were the numbers of cholinergic/nitrergic neurons. EFS stimulated cholinergic and nitrergic neurons so the contractile response of the muscle was the sum of both but dominated by cholinergic (causing contraction) or less often, nitrergic activity (relaxation), followed, after termination of EFS, by neuronally mediated contraction. Inhibition of nitric oxide synthase (by L-NAME 300μM) more clearly defined EFS-evoked contractions. The 5-HT4 agonist prucalopride 10μM and the cholinesterase inhibitor donepezil 1µM, respectively increased and greatly increased the composite contractile response to EFS (measured as 'area-under-the curve') and the contractions isolated by L-NAME (respectively, by 22±14% and 334±87%; n=11/8). After chemoradiotherapy, nitrergic-mediated muscle relaxations occurred more often during EFS (in 29.8±6.1% preparations vs. 12.6±5.1% without chemoradiotherapy, n=21/18). With L-NAME, the ability of prucalopride to facilitate EFS-evoked contraction was lost and that of donepezil approximately halved (contractions increased by 132±36%; n=8). Several months after chemoradiotherapy, the rectum was not inflamed and myenteric neuron numbers/phenotype unchanged. However, nitrergic activity was increased relative to cholinergic activity, and prokinetic-like drug activity was lost or greatly reduced. Thus, chemoradiotherapy causes long-term changes in neuromuscular functions and markedly reduces the efficacy of drugs for treating constipation.