Investigation of confocal microscopy for differentiation of renal cell carcinoma versus benign tissue.Can an optical biopsy be performed?

Objective:Novel optical imaging modalities are under development with the goal of obtaining an“optical biopsy”to efficiently provide pathologic details.One such modality is confocal microscopy which allows in situ visualization of cells within a layer of tissue and imaging of cellular-level structures.The goal of this study is to validate the ability of confocal microscopy to quickly and accurately differentiate between normal renal tissue and cancer.Methods:Specimens were obtained from patients who underwent robotic partial nephrectomy for renal mass.Samples of suspected normal and tumor tissue were extracted from the excised portion of the kidney and stained with acridine orange.The stained samples were imaged on a Nikon E600 C1 Confocal Microscope.The samples were then submitted for hematoxylin and eosin processing and read by an expert pathologist to provide a gold-standard diagnosis that can later be compared to the confocal images.Results:This study included 11 patients,17 tissue samples,and 118 confocal images.Of the 17 tissue samples,10 had a gold-standard diagnosis of cancer and seven were benign.Of 118 confocal images,66 had a gold-standard diagnosis of cancer and 52 were benign.Six confocal images were used as a training set to train eight observers.The observers were asked to rate the test images on a six point scale and the results were analyzed using a web based receiver operating characteristic curve calculator.The average accuracy,sensitivity,specificity,and area under the empirical receiver operating characteristic curve for this study were 91%,98%,81%,and 0.94 respectively.Conclusion:This preliminary study suggest that confocal microscopy can be used to distinguish cancer from normal tissue with high sensitivity and specificity.The observers in this study were trained quickly and on only six images.We expect even higher performance as observers become more familiar with the confocal images.

Cervical inlet patch-optical coherence tomography imaging and clinical significance

AIM:To demonstrate the feasibility of optical coherence tomography(OCT)imaging in differentiating cervical inlet patch(CIP)from normal esophagus,Barrett'sesophagus(BE),normal stomach and duodenum. METHODS:This study was conducted at the Veterans Affairs Boston Healthcare System(VABHS).Patients undergoing standard esophagogastroduodenoscopy at VABHS,including one patient with CIP,one representative patient with BE and three representative normal subjects were included.White light video endoscopy was performed and endoscopic 3D-OCT images were obtained in each patient using a prototype OCT system.The OCT imaging probe passes through the working channel of the endoscope to enable simultaneous video endoscopy and 3D-OCT examination of the human gastrointestinal(GI)tract.Standard hematoxylin and eosin(H and E)histology was performed on biopsy or endoscopic mucosal resection specimens in order to compare and validate the 3D-OCT data. RESULTS:CIP was observed from a 68-year old male with gastroesophageal reflux disease.The CIP region appeared as a pink circular lesion in the upper esophagus under white light endoscopy.OCT imaging over the CIP region showed columnar epithelium structure,which clearly contrasted the squamous epithelium structure from adjacent normal esophagus.3D-OCT images obtained from other representative patients demonstrated distinctive patterns of the normal esophagus,BE,normal stomach,and normal duodenum bulb.Microstructures,such as squamous epithelium,lamina propria, muscularis mucosa,muscularis propria,esophageal glands,Barrett's glands,gastric mucosa,gastric glands, and intestinal mucosal villi were clearly observed with OCT and matched with H and E histology.These results demonstrated the feasibility of using OCT to evaluate GI tissue morphology in situ and in real-time. CONCLUSION:We demonstrate in situ evaluation of CIP microstructures using 3D-OCT,which may be a useful tool for future diagnosis and follow-up of patients with CIP.

A Confocal Endoscope for Cellular Imaging

Since its inception, endoscopy has aimed to establish an immediate diagnosis that is virtually consistent with a histologic diagnosis. In the past decade, confocal laser scanning microscopy has been brought into endoscopy, thus enabling in vivo microscopic tissue visualization with a magnification and resolution comparable to that obtained with the ex vivo microscopy of histological specimens. The major challenge in the development of instrumentation lies in the miniaturization of a fiber-optic probe for microscopic imaging with micron-scale resolution. Here, we present the design and construction of a confocal endoscope based on a fiber bundle with 1.4-μm lateral resolution and 8-frames per second(fps) imaging speed. The fiber-optic probe has a diameter of 2.6 mm that is compatible with the biopsy channel of a conventional endoscope. The prototype of a confocal endoscope has been used to observe epithelial cells of the gastrointestinal tracts of mice and will be further demonstrated in clinical trials. In addition, the confocal endoscope can be used for translational studies of epithelial function in order to monitor how molecules work and how cells interact in their natural environment.

Colonic polyps: Is it useful to characterize them with advanced endoscopy?

There have been major developments in endoscopic imaging techniques in recent years.Endoscopes with high definition and magnification can provide high quality images that allow for the histological estimation of lesions in vivo and in situ when combined with ancillary enhancement techniques such as chromoendoscopy(CE)and virtual CE(narrow band imaging fujinon intelligent chromoendoscopy,or i-Scan).Despite the enormous potential for these advanced techniques,their value and feasibility in the clinic are still doubted,particularly in cases of colonic polyps that are slated for removal,where in vivo characterization may be deemed unnecessary.However,there are several advantages offered by such advanced endoscopic imaging.CE with or without magnification demonstrates highly accurate histology and invasion depth prediction,and virtual CE is a feasible and less cumbersome alternative to CE in terms of histological estimation,though not sufficiently accurate for depth invasion prediction.Furthermore,the supplementary information provided by advanced imaging systems can assist the endoscopist in the selection of a strategic approach,such as in deciding whether a colonic lesion should be resected,left in situ,or requires more intensive surgical treatment.Lastly,advanced high-resolution imaging techniques may be more cost effective,such that histopathology of lowrisk lesions following resection can be eliminated.The results of these evaluations and comparisons with traditional CE are presented and discussed.Taken together,the benefits provided by these advanced capabilities justify their development,and advocates their use for the treatment and management of colonic polyps.

Multiphoton microscopy:An introduction to gastroenterologists

Multiphoton microscopy, relying on the simultaneous absorption of two or more photons by a fluorophore, has come to occupy a prominent place in modern biomedical research with its ability to allow real-time observation of a single cell and molecules in intact tissues. Multiphoton microscopy exhibits nonlinear optical contrast properties, which can make it possible to provide an exceptionally large depth penetration with less phototoxicity. This system becomes more and more an inspiring tool for a non-invasive imaging system to realize "optical biopsy" and to examine the functions of living cells. In this review, we briefly present the physical principles and properties of multiphoton microscopy as well as the current applications in biological fields. In addition, we address what we see as the future potential of multiphoton microscopy for gastroenterologic research.

Real-time in vivo distal margin selection using confocal laser endomicroscopy in transanal total mesorectal excision for rectal cancer

BACKGROUND Transanal total mesorectal excision(TaTME)allows patients with ultralow rectal cancer to be treated with sphincter-saving surgery.However,accurate delineation of the distal resection margin(DRM),which is essential to achieve R0 resection for low rectal cancer in TaTME,is technically demanding.AIM To assess the feasibility of optical biopsy using probe-based confocal laser endomicroscopy(pCLE)to select the DRM during TaTME for low rectal cancer.METHODS A total of 43 consecutive patients who were diagnosed with low rectal cancer and scheduled for TaTME were prospectively enrolled from January 2019 to January 2021.pCLE was used to determine the distal edge of the tumor as well as the DRM during surgery.The final pathological report was used as the gold standard.The diagnostic accuracy of pCLE examination was calculated.RESULTS A total of 86 pCLE videos of 43 patients were included in the analyses.The sensitivity,specificity and accuracy of real-time pCLE examination were 90.00%[95%confidence interval(CI):76.34%-97.21%],86.96%(95%CI:73.74%-95.06%)and 88.37%(95%CI:79.65%-94.28%),respectively.The accuracy of blinded pCLE reinterpretation was 86.05%(95%CI:76.89%-92.58%).Furthermore,our results show satisfactory interobserver agreement(κ=0.767,standard error=0.069)for the detection of cancer tissue by pCLE.There were no positive DRMs(≤1 mm)in this study.The median DRM was 7 mm[interquartile range(IQR)=5-10 mm].The median Wexner score was 5(IQR=3-6)at 6 mo after stoma closure.CONCLUSION Real-time in vivo pCLE examination is feasible and safe for selecting the DRM during TaTME for low rectal cancer(clinical trial registration number:NCT04016948).