In vivo histologic imaging of the muscularis propria and myenteric neurons with probe-based confocal laser endomicroscopy in porcine models (with videos).

BACKGROUND The submucosal tunneling technique enables us to endoscopically access deeper tissue layers. Use of probe-based confocal laser endomicroscopy (pCLE) provides optical histologic imaging on the site. OBJECTIVE To determine the technical feasibility of ex vivo and in vivo pCLE imaging of the muscularis propria and myenteric neurons by using submucosal endoscopy with a mucosal flap safety valve (SEMF). DESIGN Acute porcine model study. SETTING Animal laboratory. INTERVENTION Two ex vivo and 6 in vivo porcine models were used. A submucosal space was created with SEMF, and a neuronal molecular probe was topically applied onto the muscularis. Confocal imaging of the stained muscularis was performed by using pCLE. The selected sites were sampled, and the histopathology of the sites was analyzed. MAIN OUTCOME MEASUREMENTS The two main outcome measures were the procedural success rate of submucosal access and the correlation between pCLE and histologic images. RESULTS Submucosal access to the pCLE study site was successful in all attempts (100%; 17/17 sites). The muscularis propria was visualized with pCLE in the ex vivo and in vivo porcine models in 83.3% of sites (20/24), and the neuron-like cells were identified in 41.7% of sites (10/24). LIMITATIONS Animal experiment. CONCLUSION The muscularis propria and myenteric neurons could be selectively visualized with pCLE in vivo.

[1]  Pavel V. Belichenko,et al.  Confocal Laser Scanning Microscopy and 3-D Reconstructions of Neuronal Structures in Human Brain Cortex , 1995, NeuroImage.

[2]  Barry Campbell,et al.  Confocal laser endomicroscopy is a new imaging modality for recognition of intramucosal bacteria in inflammatory bowel disease in vivo , 2010, Gut.

[3]  R. Kiesslich,et al.  Confocal laser endoscopy: new approach to the early diagnosis of tumors of the esophagus and stomach. , 2006, Future oncology.

[4]  M. Wallace,et al.  Optimal fluorescein dose for intravenous application in miniprobe‐based confocal laser scanning microscopy in pigs , 2011, Journal of biophotonics.

[5]  K. Ikeda,et al.  Pilot study for in vivo cellular imaging of the muscularis propria and ex vivo molecular imaging of myenteric neurons (with video). , 2009, Gastrointestinal endoscopy.

[6]  G. Locke,et al.  Evaluation of endoscopic approaches for deep gastric-muscle-wall biopsies: what works? , 2008, Gastrointestinal endoscopy.

[7]  C. Cirstoiu,et al.  Interstitial cells of Cajal in the gut--a gastroenterologist's point of view. , 2008, World journal of gastroenterology.

[8]  V. Annese,et al.  An Immunohistochemical Study of the Myenteric Plexus in Idiopathic Achalasia , 2009, Journal of clinical gastroenterology.

[9]  R. Kiesslich,et al.  Feasibility of confocal endomicroscopy in the diagnosis of pediatric gastrointestinal disorders. , 2009, World journal of gastroenterology.

[10]  S. Pal,et al.  Dynamics in the DNA recognition by DAPI: exploration of the various binding modes. , 2008, The journal of physical chemistry. B.

[11]  C. Gostout,et al.  Chemically assisted endoscopic mechanical submucosal dissection by using mesna. , 2008, Gastrointestinal endoscopy.

[12]  V. Herlea,et al.  The evaluation of interstitial Cajal cells distribution in non-tumoral colon disorders. , 2008, Romanian journal of morphology and embryology = Revue roumaine de morphologie et embryologie.

[13]  J. Cooper,et al.  Accurate counting of neurons in frozen sections: some necessary precautions. , 1988, Journal of Anatomy.

[14]  G. Locke,et al.  Endoscopic "no hole" full-thickness biopsy of the stomach to detect myenteric ganglia. , 2008, Gastrointestinal endoscopy.

[15]  R. Abalo,et al.  Age-related changes in the gastrointestinal tract: a functional and immunohistochemical study in guinea-pig ileum. , 2007, Life sciences.

[16]  K. Ikeda,et al.  Chemically assisted submucosal injection facilitates endoscopic submucosal dissection of gastric neoplasms. , 2010, Endoscopy.

[17]  R. Kiesslich,et al.  Simultaneous confocal laser endomicroscopy and chromoendoscopy with topical cresyl violet. , 2009, Gastrointestinal endoscopy.

[18]  Giovanni Domenico De Palma,et al.  Confocal laser endomicroscopy in the "in vivo" histological diagnosis of the gastrointestinal tract. , 2009 .

[19]  H. Wagner,et al.  A fluorescent double stain for visualization of neural tissue by confocal laser scanning microscopy , 1998, Journal of Neuroscience Methods.

[20]  B. Chazotte Labeling nuclear DNA with hoechst 33342. , 2011, Cold Spring Harbor protocols.

[21]  Steven Mark Miller,et al.  Distribution of interstitial cells of Cajal and nitrergic neurons in normal and diabetic human appendix , 2008, Neurogastroenterology and motility : the official journal of the European Gastrointestinal Motility Society.

[22]  R. Kiesslich,et al.  Confocal laser endomicroscopy: technical status and current indications , 2006, Endoscopy.

[23]  C. Gostout,et al.  Submucosal endoscopy with mucosal flap safety valve. , 2007, Gastrointestinal endoscopy.

[24]  H. Tajiri,et al.  Examination of normal intestine using confocal endomicroscopy , 2007, Journal of gastroenterology and hepatology.

[25]  H. Tajiri,et al.  Submucosal endoscopy with mucosal flap safety valve (SEMF) technique: A safe access method into the peritoneal cavity and mediastinum , 2008, Minimally invasive therapy & allied technologies : MITAT : official journal of the Society for Minimally Invasive Therapy.

[26]  S. Staibano,et al.  In vivo characterisation of superficial colorectal neoplastic lesions with high-resolution probe-based confocal laser endomicroscopy in combination with video-mosaicing: a feasibility study to enhance routine endoscopy. , 2010, Digestive and liver disease : official journal of the Italian Society of Gastroenterology and the Italian Association for the Study of the Liver.

[27]  J. Changeux,et al.  Live imaging of neural structure and function by fibred fluorescence microscopy , 2006, EMBO reports.

[28]  Alexander Meining,et al.  Preliminary accuracy and interobserver agreement for the detection of intraepithelial neoplasia in Barrett's esophagus with probe-based confocal laser endomicroscopy. , 2010, Gastrointestinal endoscopy.

[29]  J. Furness,et al.  Morphology and Neurochemistry of Descending and Ascending Myenteric Plexus Neurons of Sheep Ileum , 2007, Anatomical record.