Expression of PGP 9.5 by Enteric Neurons in Horses and Donkeys with and without Intestinal Disease

Intestinal motility disorders are an important problem in horses and donkeys and this study was carried out in order to evaluate the enteric neurons in animals with and without intestinal disease. Surplus intestinal tissue samples were collected from 28 horses undergoing exploratory laparotomy for colic. In addition, surplus intestinal samples from 17 control horses were collected immediately following humane destruction for clinical conditions not relating to the intestinal tract. Similar samples were also collected during routine post-mortem examinations from 12 aged donkeys; six animals were humanely destroyed for conditions related to the intestinal tract, while the remaining six were humanely destroyed for other reasons including dental and orthopaedic diseases. Tissue samples were fixed in formalin and immunohistochemical labelling was performed targeting the enteric neurons using a polyclonal antibody specific for the neuronal marker PGP 9.5. The distribution and density of neuronal networks were assessed qualitatively and semiquantitatively. There was strong PGP 9.5 expression in both the horse and donkey samples and labelling was detected throughout the tissue sections. In both species, PGP 9.5-immunoreactive nerve fibres were detected in all layers of the intestinal tract, both in large and small intestinal samples. Networks of enteric neurons were present in the donkey with a similar distribution to that seen in the horse. There was no demonstrable difference in enteric neuronal density and distribution in the groups of animals with intestinal disease compared with those without, apart from two (out of 28) horses with intestinal disease that showed a marked reduction in PGP 9.5 immunoreactivity. Apart from these two animals, this total cohort analysis differs from some previously observed findings in horses with intestinal disease and may therefore reflect the different pathophysiological processes occurring in varying intestinal conditions resulting in colic both in the donkey and the horse. © 2013 Elsevier Ltd

Comparative analysis of c-kit gene expression and c-Kit immunoreactivity in horses with and without obstructive intestinal disease

Previous immunohistochemical studies targeting the receptor tyrosine kinase (c-Kit) have demonstrated an apparent reduction in the number of gastrointestinal pacemaker cells--the interstitial cells of Cajal (ICC)--in horses with intestinal motility disorders. This study compared the level of transcription of the c-kit gene encoding this receptor in horses with and without such motility disorders. Transcription levels of this gene were also compared to the density of ICC immunohistochemically positive for the c-Kit antigen. Intestinal samples were collected from 18 horses with intestinal disease and from 15 control animals. Following gene extraction and identification, real-time quantitative analysis of c-kit and a control gene, ACTB (β-actin), was carried out on all samples and the density of the c-Kit-positive ICC compared. There was a significant reduction in c-Kit immunoreactivity in the ICC of horses with large intestinal obstructive disorders relative to controls but no significant difference in the transcription of the c-kit gene between normal and affected animals. Further studies will be required to elucidate the mechanisms regulating c-Kit expression and to assess the pathophysiological significance of these findings

Expression of PGP 9.5 by enteric neurons in horses and donkeys with and without intestinal disease

Intestinal motility disorders are an important problem in horses and donkeys and this study was carried out in order to evaluate the enteric neurons in animals with and without intestinal disease. Surplus intestinal tissue samples were collected from 28 horses undergoing exploratory laparotomy for colic. In addition, surplus intestinal samples from 17 control horses were collected immediately following humane destruction for clinical conditions not relating to the intestinal tract. Similar samples were also collected during routine post-mortem examinations from 12 aged donkeys; six animals were humanely destroyed for conditions related to the intestinal tract, while the remaining six were humanely destroyed for other reasons including dental and orthopaedic diseases. Tissue samples were fixed in formalin and immunohistochemical labelling was performed targeting the enteric neurons using a polyclonal antibody specific for the neuronal marker PGP 9.5. The distribution and density of neuronal networks were assessed qualitatively and semiquantitatively. There was strong PGP 9.5 expression in both the horse and donkey samples and labelling was detected throughout the tissue sections. In both species, PGP 9.5-immunoreactive nerve fibres were detected in all layers of the intestinal tract, both in large and small intestinal samples. Networks of enteric neurons were present in the donkey with a similar distribution to that seen in the horse. There was no demonstrable difference in enteric neuronal density and distribution in the groups of animals with intestinal disease compared with those without, apart from two (out of 28) horses with intestinal disease that showed a marked reduction in PGP 9.5 immunoreactivity. Apart from these two animals, this total cohort analysis differs from some previously observed findings in horses with intestinal disease and may therefore reflect the different pathophysiological processes occurring in varying intestinal conditions resulting in colic both in the donkey and the horse. © 2013 Elsevier Ltd

Interstitial cells of Cajal (ICC) in equine colic: an immunohistochemical study of horses with obstructive disorders of the small and large intestines.

REASONS FOR PERFORMING STUDY: The gastrointestinal pacemaker cells, the interstitial cells of Cajal (ICC), have been implicated in several human gastrointestinal dysmotility syndromes. Recently, the involvement of these cells in equine gastrointestinal diseases has been investigated in cases of equine grass sickness where a significant reduction in ICC density was observed. OBJECTIVE: To investigate ICC density in equine obstructive gastrointestinal disorders using immunohistochemical labelling methods. METHODS: Intestinal samples were analysed from 44 horses undergoing exploratory surgery for colic and from 11 control animals subjected to euthanasia for conditions not related to the gastrointestinal tract. Immunohistochemical labelling of ICC was carried out using an anti-c-Kit antibody. Two independent observers assessed ICC density using a semiquantitative grading system. RESULTS: There was a significant reduction in ICC density in horses with large colon disorders compared to the controls (P<0.01). Horses with strangulating lesions of the small intestine showed no difference when compared to the controls. CONCLUSIONS: There was a reduction in ICC density in horses with large intestinal disorders. POTENTIAL RELEVANCE: The reduction in ICC density may be associated with the clinical findings as well as recurrent colic episodes observed in a number of these cases. This immunohistochemical study provides a basis for future functional electrophysiological investigations to determine the precise effect of ICC reduction on equine intestinal motility

Comparative analysis of c-kit gene expression and c-Kit immunoreactivity in horses with and without obstructive intestinal disease.

Previous immunohistochemical studies targeting the receptor tyrosine kinase (c-Kit) have demonstrated an apparent reduction in the number of gastrointestinal pacemaker cells--the interstitial cells of Cajal (ICC)--in horses with intestinal motility disorders. This study compared the level of transcription of the c-kit gene encoding this receptor in horses with and without such motility disorders. Transcription levels of this gene were also compared to the density of ICC immunohistochemically positive for the c-Kit antigen. Intestinal samples were collected from 18 horses with intestinal disease and from 15 control animals. Following gene extraction and identification, real-time quantitative analysis of c-kit and a control gene, ACTB (β-actin), was carried out on all samples and the density of the c-Kit-positive ICC compared. There was a significant reduction in c-Kit immunoreactivity in the ICC of horses with large intestinal obstructive disorders relative to controls but no significant difference in the transcription of the c-kit gene between normal and affected animals. Further studies will be required to elucidate the mechanisms regulating c-Kit expression and to assess the pathophysiological significance of these findings