Myenteric plexuses atrophy in the vicinity of colorectal cancer tissue is not caused by apoptosis or necrosis

Abstract

Introduction. The previously performed studies showed that the presence of colorectal cancer (CRC) tumor is associated with the atrophy of myenteric plexuses in the vicinity of cancer invasion; however, the possible mechanisms of this phenomenon are not known. The aim of the present study was to determine whether the atrophic changes of the enteric nervous system (ENS) within an intestine wall of the CRC patients were caused by apoptosis or necrosis and whether they were associated with changes in the number of galanin-immunoreactive (GAL-Ir) neurons. Material and methods. Samples of the large intestine wall located close to the CRC invasion and control, distally-located part of the colon, were collected from 9 CRC patients. The size of ENS plexuses and the number of neurons were compared. Triple immunofluorescent staining was used to visualize the co-expression of caspase 3 (CASP3) or caspase 8 (CASP8) with GAL and protein gene-product 9.5 (PGP 9.5, panneuronal marker) in the submucosal and myenteric ENS plexuses. The cells expressing myeloperoxidase (MPO, marker of neutrophils) and CD68 (marker of macrophages) were detected by immunohistochemistry around/in myenteric plexuses (MPs) and in the muscularis externa of the colon wall in the vicinity of tumor invasion. Results. Myenteric plexuses in the vicinity of the CRC tissue were significantly smaller and had lower number of neurons per plexus than distantly located plexuses. The number of CASP8-and CASP3-Ir neurons in the ENS plexuses was similar in the colon wall both close to and distally from tumor invasion. The number of CASP8-Ir neurons within MPs located close to CRC invasion was higher than of CASP3-Ir neurons. The percentage of neurons co-expressing CASP8 and GAL in myenteric plexuses close and distantly from tumor was three-fold lower than of those co-expressing CASP3 and GAL. The mean number of neutrophils and macrophages inside and around myenteric plexuses located close to tumor invasion was higher or similar, respectively, as compared with adjacent muscularis externa. Conclusions. The atrophy of myenteric plexuses in the vicinity of CRC invasion is not caused by apoptosis or necrosis. The differences in the proportions of neurons expressing galanin and the studied caspases suggest as yet unknown role of this neuropeptide in the mechanisms of neuron's atrophy in MPs located close to CRC tumor.

Ultrastructural characteristics of myenteric plexus in patients with colorectal cancer

“…They found significant reduction (by approximately 50%) of the area occupied by galanin-immunoreactive neurons in the MPs located in the vicinity of CRC invasion in comparison to distally located plexuses. The authors suggested that neither apoptosis nor necrosis has been involved in the observed phenomena [15,16]. These finding have been confirmed by the results of our ultrastructural study, which showed neither neurons' apoptosis nor necrosis to occur in myenteric plexuses located close to the CRC tumor.…”

“…Our study presents an attempt to explain the observations by Kwiatkowski et al [15] and Kozlowska et al [16] who applied morphometry to measure the area of MPs close to and distant from CRC invasion. They found significant reduction (by approximately 50%) of the area occupied by galanin-immunoreactive neurons in the MPs located in the vicinity of CRC invasion in comparison to distally located plexuses.…”

“…Since the mentioned histomorphometric studies clearly demonstrated atrophy of myenteric plexuses in this location we suggest that the atrophic changes must have taken place earlier during the CRC progression. Since in the present study the patients represented an advanced phase of the disease (stages II and III) it was not possible to determine at the TEM level (this study) or at the light microscopy level, which type of cell death (apoptosis, autophagy or necrosis) was responsible for the atrophy of MPs close to the CRC invasion [16]. Although, we found occasionally presence of apoptotic cells close to the bundles of nerve fibers, the morphology of these cells does not allow defining their type due to cell condensation and shrinkage.…”

Co-expression of caspase-3 or caspase-8 with galanin in the human stomach section affected by carcinoma

“…Latest studies showed that ENS may undergo severe decomposition under various pathological conditions. For example, it was reported that MPs in the vicinity of the colorectal cancer (CRC) tissue were significantly smaller and had lower number of neurons per plexus compared to distally located plexuses [ 5 , 6 ]. Similar observations were also reported in the colon wall of patients with diabetes [ 7 ] and severe constipation [ 8 ].…”

“…These stimuli are transmitted by caspases (CASP) which can be divided into initiator (CASP8; CASP9) and executioner endoproteases (CASP3; CASP6; CASP7). Caspases can be activated via extrinsic and intrinsic pathways and both coincide at a common point, which is activation of executioner caspase that leads to protein proteolysis and DNA damage [ 5 , 11 ]. Galanin (GAL) can be another candidate which may play important role in this process.…”