|Year : 2019 | Volume
| Issue : 4 | Page : 77-82
Laser confocal refractive microscopy and early detection of oral cancer: A narrative review
RJ Sameen, V Bindushree, Vijeev Vasudevan, Nimi Susan Mathew, D Devaraju, TG Shrihari
Department of Oral Medicine and Radiology, Krishnadevaraya College of Dental Sciences, Bengaluru, Karnataka, India
|Date of Submission||24-Sep-2019|
|Date of Acceptance||28-Nov-2019|
|Date of Web Publication||12-Feb-2020|
R J Sameen
Krishnadevaraya College of Dental Sciences, Bengaluru, Karnataka
Source of Support: None, Conflict of Interest: None
Laser confocal refractive microscopy is a modern technique that aids in the detection of early malignancies in recent years. However, it is inappropriately explored in oral oncology. To date, the gold standard for diagnosis of oral malignancies is still a biopsy for histopathological examination as this is an invasive method, and it causes discomfort for the patients. In this concern, the focus of this present article is to understand how this laser confocal microscopy could be used in the diagnosis of oral malignancies without the actual need for biopsy. Thus, it can be an effective noninvasive diagnostic aid for detecting oral malignancies.
Keywords: Early detection, laser confocal refractive microscopy, noninvasive diagnostic aid, oral malignancies
|How to cite this article:|
Sameen R J, Bindushree V, Vasudevan V, Mathew NS, Devaraju D, Shrihari T G. Laser confocal refractive microscopy and early detection of oral cancer: A narrative review. J Dent Res Rev 2019;6:77-82
|How to cite this URL:|
Sameen R J, Bindushree V, Vasudevan V, Mathew NS, Devaraju D, Shrihari T G. Laser confocal refractive microscopy and early detection of oral cancer: A narrative review. J Dent Res Rev [serial online] 2019 [cited 2022 May 18];6:77-82. Available from: https://www.jdrr.org/text.asp?2019/6/4/77/278220
| Introduction|| |
Oral cancer incidence and mortality are high. All over the globe, there are about 350,000 new cases of oral cancers with estimated deaths of 177,000 every year. Almost 50% of all cancers are seen in late stages. The most prevalent malignant tumor of the oral mucosa is oral squamous cell carcinoma (OSCC) which is frequently arising from precancerous lesions. Malignancies show a greater prognosis when it is diagnosed at its initial stages. This necessitates the diagnosis of oral malignancies at the earliest to improve the quality of life and patient survival., In detecting the skin lesions, the technique of laser confocal refractive microscopy, which is a noninvasive method, showed satisfactory evidence for the diagnosis of a review of skin malignancies.
With this background, the use of the same technique has been evolved in the detection of oral malignancies which showed acceptable results and has been proved as a satisfactory method for the detection of oral cancer at its early stages without the actual need for biopsy.,
| The Course of Working|| |
Laser confocal refractive microscopy consists of low-power laser with the wavelength of 488 nm which is a point light source reflected by a dichroic mirror and illuminated at only one point in a defined field of view (FOV) of a microscope, and the same lens acts as folding optical path for both condenser and object as seen in [Figure 1].,,,,,, Thus, the point of illumination coincides with the point of detection. The light is focused through a pinhole to the specimen, then the specimen starts emitting light which is detected by the detector and light which is emitting outside the illuminated spot gets rejected. The scanned region of the specimen can be constructed and digitized by measuring the light coming to a detector from a particular plane to be measured as seen in [Figure 2]. The confocal lens which is responsible for the cluster of images at the cellular and subcellular levels and three-dimensional images can be reproduced.
According to recent research studies, this process requires contrast agents to make objects visible, thus fluorescent dyes are used that will be locally injected into the lesion site or topically applied on site of interest. Fluorescein sodium (10%) and topical application acriflavine (2%) are safer and compatible to use in humans.,,,,,,,,,,,,,,,,,,,, The picture of the specimen in the selected site is obtained by scanning the focused beam by high-speed oscillating mirrors which are driven with galvanometer motors. Then, the laser light is illuminated to an area of interest in a single focal plane.
Moreover, after passing through dichromatic mirrors and pass through the pinhole aperture [Figure 2], fluorescence emission is converted into an analog signal having a change in alteration of voltage by the photomultiplier as seen in [Figure 2].,,,,,, With the help of an analog-to-digital converter which is present in the scanning unit, the analog signal is periodically converted into pixels and displayed in the monitor.
To increase the FOV, a newer video-mosaicking approach in recent studies made a successful application in early cancer detection [Figure 3] and [Figure 4]. The stable multiwavelength laser system that gives improved coverage of the ultraviolet (UV) light, visible light, and infrared spectral regions, improved interference filters, sensitive low-noise wideband detectors, and powerful computers. These powerful computers are available with relatively low-cost memory arrays, image analysis software packages, high-resolution video displays, and high-quality image printers.
|Figure 3: (a and b) Computer-aided drawing rendering of the handheld confocal microscope adapted with a rigid (relay telescope) probe for intraoral imaging|
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|Figure 4: (a-g): Imaging on a patient with oral squamous cell carcinoma on the right lateral tongue. (a) Clinical image showing cancer area (blue arrow) and surrounding margin (normal tissue) area (yellow arrow) where imaging and biopsy was performed. (b) Reflectance confocal microscopy video-mosaic of the tumor area, when imaging with the superficial cap, displays what appears to be large dark vesicles in multiple areas (red arrow, within and also to theleft of the yellow-dotted box). Also seen is a dark area of cancer cells (yellow arrow) (Color figure can be viewed at wileyonlinelibrary.com)|
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| Instrumentation|| |
To achieve images, the handheld confocal microscope will be used [Figure 3]. A rigid probe is designed for intraoral imaging with a relatively smaller objective lens. For oral and maxillofacial surgeons, the specified size of the instrument is about 150 nm in length and not more than 12 mm in diameter as seen in [Figure 3].
The probe which has a 1:1 relay telescope is made up of four doublet lenses with a diameter of 10 mm and a focal length of 60 mm. This complete setup is housed in a 6061-T6 aluminum body. The probe with a relay telescope is positioned between the dual galvanometric scanners and the objective lens. The working diameter should be a minimum of 0.6 mm.
The objective lens needs a slender coverslip of 0.2 mm thick. The coverslip contacts oral mucosal tissue and helps in the imaging site still. This helps in minimizing the motion blur artifacts. The objective lens is covered with epoxy resin at its both ends. This facilitates sterilization with ethylene oxide of the probe between uses on patients. Two cap and coverslip windows, one of length 8.6 mm and the other of length 8.7 mm, are designed for imaging at two particular depths. The longer cap is termed as “superficial cap” that allows for superficial imaging, and the shorter cap is termed as “deep cap” that allows for relatively deeper imaging.
| Imaging Protocol|| |
Cap and coverslip window to tissue contact should be consistent and stable for continuous capturing of the videos. The scanning body (proximal end) of the microscope should rest on the first digit, whereas the distal end of the probe is held between the first and second digits to avoid sudden movements and to have consistent and stable contact. This allows the probe to move smoothly over the mucosa for capturing videos [Figure 3] and [Figure 4].
In recent studies, using confocal refractive microscopy which is used in imaging the oral cavity at certain anatomical sites such as buccal mucosa, hard palate, dorsal, ventral, and lateral surface of the tongue in oral squamous cell carcinoma patients undergoing surgery, imaging and videos are recorded compared with the biopsies stained pathology given successive descriptions about cellular and subcellular structures involved in this disease. There is a disposable cap, and window coverslip is a single use for a patient.
| Noninvasive Diagnostic Aid in Early Cancer Detection|| |
Laser confocal refractive microscopy can be used in the detection of precancerous lesions and potentially malignant disorders [Figure 4]. It is capable of capturing images in the FOV of 0.75 mm × 0.75 mm, and the video displays from 2 mm × 2 mm up to 4 mm × 2 mm with the probable extension of the FOV. This extended FOV for larger areas is compared with standard 2–4 mm punch biopsies. Imaging is at the rate of 7–8 frames/s with a maximum coverage rate of approximately 150 mm square/min. It can also be used as a noninvasive diagnosis without the need for biopsy and also in accessing in a biopsy of maximally affected areas to be excised. This noninvasive aid is patient-friendly but needs more time, cost-effective. It is technique sensitive and needs more training in handling the devices. However, certain artifacts and image distortion are also hindering this noninvasive method.,,,,,,
During the sequence of images captured in the epithelial structures they are observed as polygonal in the superficial layer, whereas circular in deeper layers and nuclei appear light at upper layers and dark at lower layers, but the advantage of this method in detecting the severity of the lesion and showing whether the lesion is benign or malignant. It also helps in preventive and conservative approach and plays a vital role in diagnostics as well as treatment planning. The patients can be imaged in a normal clinical setting, and staining free techniques also make this method more comfortable and biocompatible for patients. At the site of malignancy, we can observe large nuclei with hyperkeratosis, and epithelial dysplastic features are evident and displayed in the monitor screen.,,,,, The main advantage can see the cellular and morphological patterns of the tumor and the invasion of tumor margins over larger areas of the lesion and can be zoomed for clear variations of particular changes undergoing in cancer cells at involved sites.,,,,,,,,
| Currently Available Noninvasive Diagnostic Tools in Early Detection of Oral Cancer|| |
Colposcopy, a medical diagnostic technique, is used to examine the vaginal, vulval, and cervix tissues under illuminated light with a magnified view of the area of interest. Three-dimensional images of the tissue surfaces are viewed on a monitor screen and scanned with a portable video. The use of a green/blue filter enables the assessment of changes in the vascularity and color quality, as unfiltered white or yellow light diminishes the dissimilarity concerning the adjoining tissue and the arterioles. A 70%–98% accuracy is reported for the recognition of oral mucosal alterations in oral premalignant lesions.,,,,,,,
Recently, a novel bio-nanochip (BNC) sensor which is a fast oral cytology test that amalgamates the power of cytological morphometric examination with quantification of neoplastic biomarkers was documented. In general, microfluidics technology (lab-on-a-chip) is the adjustment, miniaturization, amalgamation, and automation of analytical laboratory procedures into a solitary chip. The conducted study on the quantitative BNC method to oral cytology effectively revealed cancerous and precancerous conditions in short time duration (<45 min). The recognition of cancerous cells in the BNC sensor utilized membrane-related cell proteins that are especially present on the cellular membrane structure of neoplastic cells.
Tissue fluorescence spectroscopy
Illumination of the oral cavity tissues with the use of UV-visible light region results in the absorption of photons by fluorophores. It results in the excitation of fluorophores that cause emission of lower energy photons which are perceived as fluorescence from the mucosal surface. A study revealed that 405 nm wavelength excitation best differentiates normal oral mucosa with oral premalignant lesions.
Narrow emission tissue fluorescence
When tissues are exposed to the light of a particular wavelength, there is autofluorescence of cellular fluorophores after excitation (fluorescence imaging). A visual examination of variation in colors is observed due to cellular changes that modulate fluorophores' concentrations affecting the absorption of light in the cells.,
Visually Enhanced Lesion Scope (VELscope system; LED Dental Inc., White Rock, B.C.) comprises a light source (wavelength: 400–460 nm) and a component (manual) to assist in detailed examination or inspection. Typically, oral mucosal tissues emanate an autofluorescence light of green color, but anomalous oral mucosal lesions absorb the autofluorescent light and emerge as darker areas. However, its routine usage is not corroborated since there is an increased specificity, expense, and the absence of scientific verification.
| Conclusion|| |
In the field of dentistry, laser confocal refractive microscopy will play a prevital role in early detection and diagnosis of oral cancer. It also helps oral and maxillofacial surgeons in accessing and treating oral cancer. This method has improved the viability of cells with scanning into deeper tissues with better spatial resolution and pixel properties. Further researches will help to improve the efficiency and advancements should be made in this method, and the needs of clinician and surgeons at various clinical setting will make this a noninvasive diagnostic aid in upcoming future.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4]