HD tissue specimens (n = 10) were collected at the time of pull-through surgery, while colonic control samples were obtained at the time of colostomy closure in patients with imperforate anus (n = 10). Immunolabeling of RB1 was visualized using confocal microscopy to assess protein distribution, while western blot analysis was undertaken to quantify RB1 protein expression.
Immunohistochemistry revealed RB1 co-localized with platelet derived growth factor receptor alpha-positive (PDGFRα+) cells, nitrergic neurons and glia in controls and the ganglionic region of HD, with a marked reduction in the aganglionic HD specimens. Western blotting revealed a marked decrease in RB1 protein expression in the aganglionic region of HD colon compared to ganglionic and normal controls.
We provide evidence of the presence of RB1 expression in the human colon in HD. As RB1 is known to colocalize with nitrergic neurons, the decreased expression of RB1 in the aganglionic bowel is most likely a secondary phenomenon because of the deficient nitrergic innervation in HD.