Fine needle aspiration biopsy technique and following histological examination show its effectiveness and safety but its performing takes several time. However, the problem of real-time analysis of pathological changes in tissues remains relevant. We demonstrate optical fine-needle biopsy method, combining a fine needle (17.5G) and a fiber-optic probe (1 mm diameter) for minimally invasive interrogation of tissue in vivo. During rat tumor experiment, we collected spectrally-resolved diffuse reflectance and fluorescence. Quantified differences between tumor and normal tissues were demonstrated and approved with morphological analysis. The proposed methodology seems promising for developing new diagnostic criteria for clinical practice

Alekseyev, Alexander

Dremin, Viktor

Dunaev, Andrey

Kandurova, Ksenia

Kozlov, Igor

Kuznetsov, Sergey

Mamoshin, Andrian

Piavchenko, Gennadii

Potapova, Elena

Seryogina, Evgeniya

Zherebtsov, Evgeny

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PROCEEDINGS OF SPIESPIEDigitalLibrary.org/conference-proceedings-of-spieOptical fine-needle aspiration biopsyin a rat modelDremin, Viktor, Potapova, Elena, Zherebtsov, Evgeny,Kozlov, Igor, Seryogina, Evgeniya, et al.Viktor Dremin, Elena Potapova, Evgeny Zherebtsov, Igor Kozlov, EvgeniyaSeryogina, Ksenia Kandurova, Alexander Alekseyev, Gennadii Piavchenko,Sergey Kuznetsov, Andrian Mamoshin, Andrey Dunaev, "Optical fine-needleaspiration biopsy in a rat model," Proc. SPIE 10877, Dynamics andFluctuations in Biomedical Photonics XVI, 108770K (1 March 2019); doi:10.1117/12.2509255Event: SPIE BiOS, 2019, San Francisco, California, United StatesDownloaded From: https://www.spiedigitallibrary.org/conference-proceedings-of-spie on 18 Dec 2020  Terms of Use: https://www.spiedigitallibrary.org/terms-of-useOptical fine-needle aspiration biopsy in a rat modelViktor Dremina, Elena Potapovaa, Evgeny Zherebtsova, Igor Kozlova, Evgeniya Seryoginaa,Ksenia Kandurovaa, Alexander Alekseyevb, Gennadii Piavchenkoc,d, Sergey Kuznetsovc,Andrian Mamoshina,e, and Andrey DunaevaaResearch Development Center of Biomedical Photonics, Orel State University,Komsomolskaya Str. 95, Orel, Russia, 302026bDepartment of Anatomy, Operative Surgery and Emergency Medicine, Medical Institute, OrelState University, 95 Komsomolskaya St., Orel, Russia, 302026cHistology, Cytology and Embryology Department, I.M. Sechenov First Moscow State MedicalUniversity (Sechenov University), Mokhovaya Str. 11, Moscow, Russia, 125009dCentre of Preclinical Research, JSC “Retinoids ”, Plekhanova 2/46-5, Moscow, Russia, 111123eOrel Regional Clinical Hospital, Orel, Russia, 302028ABSTRACTFine needle aspiration biopsy technique and following histological examination show its effectiveness and safetybut its performing takes several time. However, the problem of real-time analysis of pathological changes intissues remains relevant. We demonstrate optical fine-needle biopsy method, combining a fine needle (17.5G)and a fiber-optic probe (1 mm diameter) for minimally invasive interrogation of tissue in vivo. During rattumor experiment, we collected spectrally-resolved diffuse reflectance and fluorescence. Quantified differencesbetween tumor and normal tissues were demonstrated and approved with morphological analysis. The proposedmethodology seems promising for developing new diagnostic criteria for clinical practice.Keywords: optical biopsy, fluorescence spectroscopy, diffuse reflectance spectroscopy, fine-needle aspirationbiopsy, tumor, minimally invasive surgery1. INTRODUCTIONIn recent decades, there has been an increase in patients with malignant tumors of the abdominal organs. Inparticular, hepatocellular carcinoma remains one of the most common malignant tumors and leading cause ofcancer-related deaths worldwide.1 However, there are no sufficiently accurate diagnostic methods that allow forclarifying the nature of pathological formation at the stage of obtaining a biopsy material in vivo and in situ.The biopsy taken at the time of surgery operation demands immediate analysis. Otherwise, it necessitates therepeated surgical intervention which usually leads to a worsening of the prognosis of the disease. Developmentof efficient technology for optical biopsy may offer a solution of the problem.2,3 The ability to recognize thesuspect tissue in a short time without interruption of the operation is able to increase the effectiveness of theendoscopic surgery of the cancerous tumors.Currently, many researchers apply the method of fluorescence spectroscopy (FS) for studies of tissue or cellularmetabolism.4,5 The FS application in oncology is based on differences in the intensity and spectral compositionof the fluorescence of healthy and malignant tissues when excited by laser radiation in the UV or visible rangeof the spectrum.6 Another technique named diffuse reflectance spectroscopy (DRS) provides information aboutlight absorption in tissues. DRS is used separately or in combination with FS. This method allows to differentiatemalignant and benign tumors as well, as neoplastic tissue undergo significant architectural changes at the cellularand intracellular levels.7 The combination of several complementary optical modalities can give more valuableinformation for diagnosis and treatment.Send correspondence to Viktor DreminE-mail: dremin viktor@mail.ru,Telephone: +7 953 612 7713Dynamics and Fluctuations in Biomedical Photonics XVI, edited by Valery V. Tuchin, Martin J. Leahy, Ruikang K. WangProc. of SPIE Vol. 10877, 108770K · © 2019 SPIE · CCC code: 1605-7422/19/$18 · doi: 10.1117/12.2509255Proc. of SPIE Vol. 10877  108770K-1Downloaded From: https://www.spiedigitallibrary.org/conference-proceedings-of-spie on 18 Dec 2020Terms of Use: https://www.spiedigitallibrary.org/terms-of-use2. MATERIAL AND METHODSIn this study, we present experimental studies with a fine-needle fiber optic probe comprising emitting andcollecting fibers for measurements of fluorescence intensity (with excitation at 365 nm and 450 nm) and diffusereflectance (in the range 400-900 nm).The choice of wavelengths of FS channel is caused by NADH and FAD coenzymes and collagen fluorescenceexcitation. The DRS channel includes HL-2000-FHSA Tungsten Halogen Light Source (“ Ocean Optics ”, USA)with a wavelength range of 360-2400 nm. The light collected from the tissue is filtered by optical filter toattenuate backscattered radiation and then it is analyzed by a CCD Flame Spectrometer (“ Ocean Optics ”,USA) in the range of 350-1000 nm (Fig.1a). System control and further data processing is performed using apersonal computer with custom developed software in Matlab program environment.The optic probe has been developed to be compatible with the 17.5G biopsy needle standard and has thediameter of 1 mm. The probe has 10 optical fibers. The nine transmitting ones (100 µm each) include three fibersconnected to a halogen light source, three fibers connected to a laser diode and three fibers connected to an LED.The fibers are located around the central one (200 µm), which delivers the collected light to spectrometer. Theamount and orientation of optical fibers inside the fiber-optic probe provides uniform and bright illuminationof the diagnostic volume and allows reaching a high signal-to-noise ratio. The probe has an bevel angle of 20degrees, which ensures reliable contact of the probe with dense tissues.3. RESULTS AND DISCUSSIONWith the probe, measurements of pathologically altered rat tissues of the in vivo were carried out. The animalwas stored in the Center of Preclinical Research in accordance with GLP principles. The studies have beencombined with the standard procedure of the biopsy sampling (Fig.1b). After the spectroscopy analysis thetumor tissue was fixed in the neutral formalin and embedded in parafin. The 5 µm sections were stained byhaematoxylin and eosin method with a standard operational procedures.Figure 1. (a) The multimodal FS/DRS fine needle setup; (b) rat tumor experiment; (c) fragment of a tumor node up to2.5 cm in a diameter; histological examination of tumor tissue with haematoxylin and eosin staining at (d) x100 and (e)x400.The morphological picture (Fig.1d,e) of tumor corresponds to low-grade myxofibrosarcoma 8811/3. Fig.2a,bshow that for rat tissues measured in vivo, there are significant differences between fluorescence spectra of intacttissue and myxofibrosarcoma. This may be due to changes in NADH, FAD and collagen content. Fig.2c showsthat for rat tissues, there are significant differences between the diffuse reflectance spectra of normal and tumortissue, notably in the wavelength range 400-600 nm. Reflectance measurements clearly show the pronouncedreflectance peak in region 450-500 nm. Interesting is the absence of peaks of oxyhemoglobin absorption on thetumor spectra, which indicates tissue ischemia.Proc. of SPIE Vol. 10877  108770K-2Downloaded From: https://www.spiedigitallibrary.org/conference-proceedings-of-spie on 18 Dec 2020Terms of Use: https://www.spiedigitallibrary.org/terms-of-useFigure 2. Fluorescence spectra at excitation wavelengths (a) 365 nm, (b) 450 nm and (c) diffuse reflectance spectra ofnormal (blue line) and tumor rat tissue (red line).4. CONCLUSIONFrom optical measurements acquired during rat tumor experiment, we verified the ability to collect spectrally-resolved diffuse reflectance and fluorescence rapidly in vivo. Overall, these results suggest that optical fine-needleaspiration biopsy is a promising method for the improved diagnosis of tumor in vivo.Simultaneous registration of fluorescence and diffuse reflectance spectra carries the information about thecorresponding parameters of metabolic processes and morphological structure of tissues, which is of interestfor clinical practice. As the biopsy remains a valuable diagnostic method required to ensure the diagnosis, thenecessity of real-time analysis determine the relevance of optical biopsy research. The proposed methodologyof acquiring the optical parameters data and its correspondence with biopsy samples from almost the samediagnostic volume will provide the basis for development of automatic classifier using the neural network. Thiswill make diagnostics during minimally invasive surgical procedures faster and accessible, will increase its accuracyand reliability.ACKNOWLEDGMENTSThis study was supported by the Russian Science Foundation under project No.18-15-00201.REFERENCES[1] Clark, T., Maximin, S., Meier, J., Pokharel, S., and Bhargava, P., “Hepatocellular carcinoma: Reviewof epidemiology, screening, imaging diagnosis, response assessment, and treatment,” Current Problems inDiagnostic Radiology 44(6), 479–486 (2015).[2] Kandurova, K., Dremin, V., Zherebtsov, E., Potapova, E., Dunaev, A., Mamoshin, A., Alyanov, A., andMuradyan, V., “Application of the fluorescence spectroscopy for the analysis of the state of abdominal cavityorgans tissues in mini-invasive surgery,” Proc. SPIE 10685, 106854P (2018).[3] Kandurova, K., Dremin, V., Zherebtsov, E., Potapova, E., Alyanov, A., Mamoshin, A., Ivanov, Y., Borsukov,A., and Dunaev, A., “Fiber-optic system for intraoperative study of abdominal organs during minimallyinvasive surgical interventions,” Applied Sciences 9(2) (2019).[4] Walsh, A., Cook, R. S., Rexer, B., Arteaga, C. L., and Skala, M. C., “Optical imaging of metabolism in her2overexpressing breast cancer cells,” Biomed. Opt. Express 3(1), 75–85 (2012).[5] Bartolome, F. and Abramov, A. Y., “Measurement of mitochondrial nadh and fad autofluorescence in livecells.,” Methods in molecular biology 1264, 263–70 (2015).[6] Rafailov, I., Palmer, S., Litvinova, K., Dremin, V., Dunaev, A., and Nabi, G., “A novel excitation-emissionwavelength model to facilitate the diagnosis of urinary bladder diseases,” Proc. SPIE 9303, 93030W (2015).[7] Lloyd, W. R., Wilson, R. H., Lee, S. Y., Chandra, M., McKenna, B., Simeone, D., Scheiman, J., and Mycek,M.-A., “In vivo optical spectroscopy for improved detection of pancreatic adenocarcinoma: a feasibilitystudy,” Biomed. Opt. Express 5(1), 9–15 (2014).Proc. of SPIE Vol. 10877  108770K-3Downloaded From: https://www.spiedigitallibrary.org/conference-proceedings-of-spie on 18 Dec 2020Terms of Use: https://www.spiedigitallibrary.org/terms-of-use

Optical fine-needle aspiration biopsy in a rat model

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Optical fine-needle aspiration biopsy in a rat model

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