Optical endomicroscopy and the road to real-time, in vivo pathology: present and future

Abstract

Epithelial cancers account for substantial mortality and are an important public health concern. With the need for earlier detection and treatment of these malignancies, the ability to accurately detect precancerous lesions has an increasingly important role in controlling cancer incidence and mortality. New optical technologies are capable of identifying early pathology in tissues or organs in which cancer is known to develop through stages of dysplasia, including the esophagus, colon, pancreas, liver, bladder, and cervix. These diagnostic imaging advances, together as a field known as optical endomicroscopy, are based on confocal microscopy, spectroscopy-based imaging, and optical coherence tomography (OCT), and function as "optical biopsies," enabling tissue pathology to be imaged in situ and in real time without the need to excise and process specimens as in conventional biopsy and histopathology. Optical biopsy techniques can acquire high-resolution, cross-sectional images of tissue structure on the micron scale through the use of endoscopes, catheters, laparoscopes, and needles. Since the inception of these technologies, dramatic technological advances in accuracy, speed, and functionality have been realized. The current paradigm of optical biopsy, or single-area, point-based images, is slowly shifting to more comprehensive microscopy of larger tracts of mucosa. With the development of Fourier-domain OCT, also known as optical frequency domain imaging or, more recently, volumetric laser endomicroscopy, comprehensive surveillance of the entire distal esophagus is now achievable at speeds that were not possible with conventional OCT technologies. Optical diagnostic technologies are emerging as clinically useful tools with the potential to set a new standard for real-time diagnosis. New imaging techniques enable visualization of high-resolution, cross-sectional images and offer the opportunity to guide biopsy, allowing maximal diagnostic yields and appropriate staging without the limitations and risks inherent with current random biopsy protocols. However, the ability of these techniques to achieve widespread adoption in clinical practice depends on future research designed to improve accuracy and allow real-time data transmission and storage, thereby linking pathology to the treating physician. These imaging advances are expected to eventually offer a see-and-treat paradigm, leading to improved patient care and potential cost reduction.

Virtual slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/5372548637202968.

Figure 1

Two types of confocal endomicroscopy systems are currently available in the United States. A mini-probe based system (MaunaKeaTechnology, France; upper panel) can be used through the working channel of most conventional endoscopes. In the confocal laser endomicroscope (Pentax, Japan; lower panel), the laser scanner is integrated into the endoscope. Reprinted with permission from Goetz M, Kiesslich R [33].

Figure 2

Barrett’s mucosa with early mucosal adenocarcinoma recorded with in vivo miniprobe confocal laser microscopy. Neoplastic characteristics include irregular epithelial lining with variable width (white arrows), increased cell density seen as dark areas with variable fluorescein uptake (white triangle), fusion of glands (black arrow), and irregular dilated blood vessels (arrowheads). Reprinted with permission from Pohl H, et al. [37].

Figure 3

Neutrophils and microabscesses of H. pylori-positive gastric mucosa. (A) Neutrophils were identified by their nuclear features. White arrow shows the mononuclear cell. (B) Microabscesses appeared in superficial epithelium and foveola. Reprinted with permission from Ji R, et al. [43].

Figure 4

SECM and histopathological images of BE stained with 0.6% acetic acid. (A) Large-area SECM image shows columnar epithelium (arrowhead) and squamous epithelium (arrow). (B) Histopathologic image demonstrates squamoglandular junctional mucosa. (C) High-magnification SECM image shows the presence of goblet cells (arrow). (D) High-magnification histopathological image shows BE with the presence of goblet cells (arrow). Scale bars represent 250 μm. Reprinted with permission from Kang D, et al. [50].

Figure 5

Comparison of depth, area, and images achieved with a/LCI and confocal microscopy. Typical a/LCI data. (A) Angle-resolved depth scan of light scattered from tissue. Lighter shades of gray indicate increased amount of scattered light. (B) Amplitude scan indicating depth increments used for processing. Tissue layers are labeled, and gray bar indicates basal layer (optical coherence tomography). Example angular scans for 3 tissue types pictured (solid line) with best-fit Mie theory solutions (dashed line) and size indicated. Reprinted with permission from Terry NG, et al. [26].

Figure 6

OCT image of colon adenoma (2-o’clock position). A well-organized linear crypt pattern is not present and image is darker because of altered light scattering compared with the nondysplastic mucosa as seen in the normal mucosa running horizontally in the 6-o’clock position. The marks of the vertical and horizontal axes are 1 mm apart. Reprinted with permission from Pfau PR, et al. [74].

Figure 7

Optical coherence tomography (OCT) images. (A) OCT image of normal cervical tissue, showing a well-organized, three-layer architecture (optical structure) with sharp borders. The thin basement membrane (BM) could not be resolved by OCT. However, because the basement membrane separates the epithelium (EP) from stroma (ST), a sharp interface could be visualized (length of the white bar: 1 mm). (B) OCT image showing a cervical intraepithelial neoplasia (CIN-3) lesion. The intensity of the stromal layer increases with less-organized layer architecture. The stroma seemed to push its way towards the surface as vertical columns. (C) OCT image showing invasive carcinoma. The tissue surface is an unstructured homogeneous highly backscattering region with a complete lack of layer architecture (optical structure). The basement membrane is no longer intact or defined and the tissue microstructure is no longer organized. Reprinted with permission from Gallwas J, et al. [76].

Figure 8

High-resolution images from VLE. (A) A comprehensive vascular map derived from the structural image set. (B–D) Cross-sectional images at the indicated locations. Arrows indicate corresponding vessels in the vascular map and cross-sectional images. Reprinted with permission from Vakoc BJ, et al. [70].

Figure 9

High-resolution images from VLE. (A) A transverse cross-sectional image showing all architectural layers of the squamous mucosa, including the epithelium (e), lamina propria (lp), muscularis mucosa (mm), submucosa (sm), and muscularis propria (mp); because of the large change in esophageal circumference during imaging (56 mm) and after resection (~22 mm), the cross-sectional image is displayed over a proportionately larger width. (B) Representative histology from the same swine (H&E, orig. mag. x2). Reprinted with permission from Vakoc BJ, et al. [70].

Figure 10

OFDI images obtained from patients with a normal-appearing stomach and esophagus by endoscopy. (A) OFDI image of squamous mucosa. (B) Expanded view of A demonstrates a layered appearance, including the epithelium (e), lamina propria (lp), muscularis mucosa (mm), submucosa (sm), and muscularis propria (mp). Vessels are clearly identified in the submucosa (arrows). (C) OFDI image of gastric cardia. (D) Expanded view of C demonstrates vertical pit and crypts, regular, broad architecture, high surface backscattering, and diminished image penetration. Tick marks in A and C and scale bars in B and D represent 1 mm. Reprinted with permission from Suter MJ, et al. [68].

Figure 11

Barrett’s esophagus with dysplasia. (A) Videoendoscopic image reveals a patchy mucosa consistent with SIM. (B) Histopathologic image of the biopsy specimen taken from the SCJ demonstrates intestinal metaplasia and low-grade dysplasia (H&E, orig. mag. °–2). (C) Cross-sectional OFDI image demonstrating regions consistent with SIM without dysplasia (blue arrow) and specialized intestinal metaplasia with high grade dysplasia (black arrow). (D) Expanded view of C taken from the region denoted by the blue arrow in C, demonstrating good surface maturation (arrowheads), which is consistent with SIM without dysplasia. (E) Expanded view of C taken from the region denoted by the black arrow in C, demonstrating features consistent with high grade dysplasia, including poor surface maturation (black arrowheads) and the presence of dilated glands (red arrowheads) in the mucosa. (F) A longitudinal slice highlights the transition from gastric cardia, through a 9-mm segment of specialized intestinal metaplasia and finally into squamous mucosa. Scale bars and tick marks represent 1 mm. Reprinted with permission from Suter MJ, et al. [68].