Confocal laser endomicroscopy in gastro-intestinal endoscopy: technical aspects and clinical applications

Abstract

Confocal laser endomicroscopy (CLE) is an advanced endoscopic imaging technology that provides a magnified, cellular level view of gastrointestinal epithelia. In conjunction with topical or intravenous fluorescent dyes, CLE allows for an "optical biopsy" for real-time diagnosis. Two different CLE system have been used in clinical endoscopy, probe-based CLE (pCLE) and endoscope-based CLE (eCLE). Using pCLE, the device can be delivered: (I) into the luminal gastrointestinal tract through the working channel of standard endoscopes; (II) into extraluminal cystic and solid parenchymal lesions through an endoscopic ultrasound (EUS) needle; or (III) into the biliary system through an endoscopic retrograde cholangiopancreatography (ERCP) accessory channel. With eCLE, the probe is directly integrated into the tip of a conventional endoscope, however, these endoscopes are no longer commercially available. CLE has moderate to high diagnostic accuracy for neoplastic and inflammatory conditions through the gastrointestinal tract including: oesophageal, gastric and colonic neoplasia, pancreatic cysts and solid lesions, malignant pancreatobiliary strictures and inflammatory bowel disease. Some studies have demonstrated the diagnostic benefit of CLE imaging when combined with either conventional white light endoscopy or advanced imaging technologies. Therefore, optical biopsies using CLE can resolve diagnostic dilemmas in some cases where conventional imaging fails to achieve conclusive results. CLE could also reduce the requirement for extensive tissue sampling during surveillance procedures. In the future, CLE in combination with molecular probes, could allow for the molecular characterization of diseases and assess response to targeted therapy. However, the narrow field of view, high capital costs and specialized operator training requirements remain the main limitations. Future multi-center, randomized trials with a focus on conventional diagnostic applications, cost-effectiveness and standardized training will be required for definitive evidence. The objective of this review is to evaluate the technical aspects and current applications of CLE in patients with gastrointestinal and pancreatobiliary diseases and discuss future directions for this technique.

Keywords: Confocal laser endomicroscopy (CLE); colon cancer; esophageal cancer; fluorescent endoscopy; image enhanced endoscopy; inflammatory bowel disease (IBD); pancreatic cancer; stomach cancer.

Figure 1

Principles of confocal laser endomicroscopy. The light passing through a pinhole is reflected by the beam splitter and then focused by the objective lens. The tissue is illuminated and the fluorescence emitted is collected by the objective. A pin hole placed before the detector allows to reject the light signal from out-of-focus tissue layers. Magnification: 10×.

Figure 2

CLE technologies. (A) Probe-based confocal laser endomicroscopy system (pCLE) (Mauna Kea Technologies, Paris, France). (B) pCLE probe can be delivered in vivo through the standard 2.8 mm endoscope channel. (C) Endoscope-based confocal laser endomicroscopy (eCLE). The arrows point to the different components of the eCLE system. Adapted from De Palma GD, World J Gastroenterol, 2009. CLE, confocal laser endomicroscopy.

Figure 3

Normal squamous epithelium in oesophagus-dark, homogeneous epithelial cells with regular architecture and clearly visible borders, capillaries directed toward the epithelium within the lamina propria, without penetration of the contrast agent into the surrounding tissue. Adapted from Templeton A, et al., Clin Endosc, 2013.

Figure 4

pCLE criteria for low-grade dysplasia and high-grade dysplasia in Barrett’s oesophagus. (A) Dark non-round glands. (B) Variable degree of darkness with sharp cut-off. (C) Cellular stratification. (D) Poorly identifiable goblet cells. (E) Glands unequal in size and shape. (F) Non-equidistant and irregular cells. (G) “Saw-toothed” epithelial surface. (H) Non-equidistant glands. (I) Enlarged cells. pCLE, probe-based confocal laser endomicroscopy.

Figure 5

Oesophageal squamous carcinoma-squamous cells are irregularly arranged and differ in size and morphology. Capillary leakage of fluorescein is observed. Magnification: 1,000×. Adapted from Liu J, et al., PLoS One, 2014.

Figure 6

The pCLE appearance of normal and pathological gastric mucosa. (A) Normal cardiac mucosa-regular pits with wide openings. (B) Dysplastic gastric-dark epithelium with irregular and varying thickness is observed. (C) Differentiated adenocarcinoma-disorganized epithelium with dark and irregular glands. (D) Undifferentiated adenocarcinoma-dark and irregular cells with no identifiable glandular structures are observed. Adapted from Kin MY, et al., World J Gastroenterol, 2016. pCLE, probe-based confocal laser endomicroscopy.

Figure 7

The eCLE appearance of normal and pathological gastric mucosa. (A) Normal gastric mucosa in the fundus - gastric pits are round (solid arrow), net-like subepithelial capillary network patterns surround the gastric pits (dash arrow). (B) Normal gastric mucosa in the body-gastric pits are round (solid arrow). Honeycomb-like subepithelial capillary network patterns surround the gastric pits (dash arrows). (C) Normal gastric mucosa in the antrum-gastric pits are the line type (solid arrow). Coil-shaped subepithelial capillary network patterns surround the gastric pits (dash arrow). (D) Low grade gastric intraepithelial neoplasia-different sizes of gastric pits, capillary network is thickening and circuitous. (E) High grade gastric intraepithelial neoplasia-abnormal arrangement of gastric pits. The thickening capillary network and the increasing branch present a mass shape. Magnification: 1,000×. Adapted from Shu-Fang W, et al., World J Gastroenterol, 2012. eCLE, endoscope-based confocal laser endomicroscopy.

Figure 8

The CLE appearance of normal and pathological small bowel mucosa. (A) Normal small bowel mucosa-normal epithelium border with regular capillary pattern. (B) Confocal image of celiac disease-Marsh type 3b. Magnification: 1,000×. Adapted from De Palma, GD, World J Gastroenterol, 2009; Venkatesh K, World J Gastroenterol, 2009. CLE, confocal laser endomicroscopy.

Figure 9

nCLE images of pancreas. (A) Intraductal papillary mucinous neoplasm: a single papilla with a central fibrovascular core and overlying epithelium. (B) Serous cystadenoma: branching and tortuous network of multiple blood vessels in a “fern like” pattern. (C) Pseudocyst: clusters of bright, floating particles with a background which is nondescript and lacks blood vessels. (D) Mucinous cystic neoplasm: solitary epithelial bands without formation of papillae. Adapted from Krishna G, et al., World J Gastroenterol, 2015. nCLE, needle-based confocal laser endomicroscopy.

Figure 10

CLE images of biliary ducts. (A) Normal bile duct-reticular network of thin dark branching bands ≤20 µm. (B-E) Features of malignant bile duct strictures: (B) thick white bands (>20 µm), (C) thick dark bands (>40 µm), (D) epithelium, (E) dark clumps. (F-J) features of inflammatory bile duct strictures on confocal laser endomicroscopy: (F) thickened reticular structures, (G) multiple white bands, (H) dark granular pattern in scales, (I) increased spaces between scales. Magnification: 1,000×. Adopted from Almadi et al., World J Gastroenterol, 2015. CLE, confocal laser endomicroscopy.

Figure 11

CLE images of colonic mucosa. (A) Normal colonic mucosa-round shaped crypts, dark goblet cells, narrow and regular blood vessels surrounding the crypts. (B) Adenomatous polyp-irregular or villiform structures and a dark, irregularly thickened epithelium with a decreased number of goblet cells. (C) Adenocarcinoma-disorganized mucosa, lack of structure, elongated crypts, irregularly thickened epithelium, dilated and distorted blood vessels. Adapted from De Palma GD, et al., World J Gastroenterol, 2009. CLE, confocal laser endomicroscopy.

Figure 12

CLE images in inflammatory bowel diseases. (A) Active ulcerative colitis (distal colitis-the switch from normal mucosa (top of the figure) to inflamed mucosa with irregular arrangement of crypts, crypt fusion and capillaries alterations. (B) Crohn’s disease-dilated and distorted capillaries, altered crypt architecture, increased distance between the crypts. Magnification: 1,000×. Adapted from De Palma GD, et al., World J Gastroenterol, 2013. CLE, confocal laser endomicroscopy.