Immunohistochemical and molecular expression of CB1 receptors in human colonic segment. Preliminary results

Abstract

Recent studies document that CB1 cannabinoid receptors, considered since its first identification as “brain specific”, could be expressed by peripheral tissues targets (adipose organs, enteric nervous system, striated muscle, epatocytes). This receptors were found in the GI tract of different species, including mice, rats, guinea pigs, pigs and humans. Endocannabinoids play a role in the pathophysiology of the gastrointestinal tract. They play a role in emesis, acid-related disorders, motility-related disorders, irritable bowel syndrome and diarrhea, and in inflammatory bowel disease. Endocannabinoid system might be involved in cancer differentiation, growth and cell migration. Distribution of receptors in different colonic segment is not known. We studied CB1 receptors distribution in 10 patients colonic segments; the present report shows preliminary results. Fragments of sigma, transverse and ascending human colon were obtained during endoscopic screening procedure. Materials and Methods The specimens, for immunohistochemistry (IHC), were fixed in buffered formalin and embedded in paraffin; obtained sections were processed with anti CB1 (Biosource Europe S.A.) by EnVision+System HRP (AEC) and have been studied with photomicroscope Leica DMLB. The specimens for the Reverse Transcription Polymerase Chain Reaction (RT-PCR) were frozen in liquid nitrogen and stored at -80° until use. Total RNA extraction was performed using the “illustra RNA spin Mini Kit”. RT reaction was performed using the “Enhanced avian HS RT PCR kit” The PCR was performed using the “PCR enzyme Selection Kit-High specificity”. Results and Discussion Following the endoscopic procedure was performed anatomopathological examination of specimens. The results have revealed that four of the 10 patients had a polyp the others were not suffering from any disease of the colon. Our immunohistochemical evaluation has revealed the presence of CB1 receptors expressed by regional differences in patients who had a polyp. This reactivity appeared to be lower than that shown in non-pathological patients. The PCR results instead of doubtful interpretation. In fact, despite being detected in some cases also marked a positive control, in two cases, the PCR is a totally negative compared with a positive IHC. Since the number of cases studied is not statistically significant, it is our intention to continue to evaluate additional patients both normal and suffering from polyposis in order to clarify the difference beetween immunohistochemical and molecular biology results.