SS-OCTA對(duì)黑色素瘤皮膚結(jié)構(gòu)和血管的成像實(shí)驗(yàn)

劉敬璇，樊金宇，汪 權(quán)，史國華*

SS-OCTA對(duì)黑色素瘤皮膚結(jié)構(gòu)和血管的成像實(shí)驗(yàn)

劉敬璇1,2，樊金宇2，汪 權(quán)2，史國華2*

1中國科學(xué)技術(shù)大學(xué)，安徽 合肥 230000；2中國科學(xué)院蘇州生物醫(yī)學(xué)工程技術(shù)研究所，江蘇 蘇州 215000

掃頻源光學(xué)相干層析血管成像(SS-OCTA)是一種基于分頻幅去相關(guān)血管造影法(SSADA)的新型血管成像技術(shù)，在腫瘤等疾病的早期診斷方面擁有較大前景。本文在5.12 mm′5.12 mm成像視場、標(biāo)準(zhǔn)圖像最大信噪比34.3 dB的SS-OCTA成像平臺(tái)，對(duì)黑色素瘤C57BL6小鼠進(jìn)行皮膚結(jié)構(gòu)和血管成像采集。結(jié)果表明在皮膚科疾病的早期診斷方面，利用SS-OCTA系統(tǒng)進(jìn)行血管成像優(yōu)于結(jié)構(gòu)成像。

掃頻源光學(xué)相干層析血管成像；皮膚結(jié)構(gòu)；腫瘤血管；黑色素瘤；SS-OCTA

1 引 言

近年來，光學(xué)相干層析成像(Optical coherence tomography, OCT)迅速發(fā)展，成為一種熱門的新型成像技術(shù)[1]。OCT通過測量光學(xué)弱相干反射和背向散射，利用超外差探測技術(shù)提高信噪比獲取生物組織斷層圖像[2]。在保證一定探測深度的前提下，OCT兼具非侵入、高分辨和高速成像的優(yōu)點(diǎn)，非常適合生物醫(yī)學(xué)領(lǐng)域的應(yīng)用[2-3]。掃頻源光學(xué)相干層析血管成像(swept source optical coherence tomography angiography，SS-OCTA)是一種基于分頻幅去相關(guān)血管造影法(split spectrum amplitude deeorrelation angiography，SSADA)的高分辨率的新型非侵入性血管成像技術(shù)[4]，主要用于血管內(nèi)紅細(xì)胞運(yùn)動(dòng)的檢測和測量[5-6]。SS-OCTA依舊以邁克爾遜干涉儀作為基礎(chǔ)光路[7]，再通過測量組織后向散射光的低相干干涉信號(hào)進(jìn)行斷層成像，軸向分辨率可達(dá)1mm~15mm，能夠清晰顯示微細(xì)結(jié)構(gòu)，通過對(duì)截面圖進(jìn)行三維重構(gòu)，可以獲取生物組織結(jié)構(gòu)的3D圖像，在眼科、皮膚科成像、腫瘤檢測等領(lǐng)域有廣泛的應(yīng)用[2,8]。尤其在眼科領(lǐng)域的眼底血管成像方面，SS-OCTA除了擁有傳統(tǒng)OCT可獲得血管結(jié)構(gòu)信息的優(yōu)點(diǎn)以外，還能夠分層觀察視網(wǎng)膜脈絡(luò)膜血管形態(tài)及血流改變情況，使用偽彩可區(qū)分正常與異常的血管結(jié)構(gòu)，并且能夠?qū)ρ餍盘?hào)進(jìn)行探測和量化分析，將原始全頻譜圖像分裂為數(shù)個(gè)不同頻譜圖像并減少其噪聲，提高信噪比，然后再將其合并，從而達(dá)到視網(wǎng)膜、脈絡(luò)膜各層血管形態(tài)在橫斷面的清晰成像[5-8,15-17]。

實(shí)體瘤的生長對(duì)受到誘導(dǎo)所形成的血管網(wǎng)有較強(qiáng)的依賴性[9]。無論是直接研究，還是間接研究，都是腫瘤生長依賴血管的有力證據(jù)[9-10]。絕大多數(shù)的腫瘤在出現(xiàn)癌變現(xiàn)象之前處于靜息期，血管不會(huì)生長；一旦進(jìn)入血管期，新生血管將迅速生長，為腫瘤代謝提供了支撐，對(duì)腫瘤增殖發(fā)揮了非常關(guān)鍵的作用[9]。對(duì)于血管成像有不同的成像方式，如X線血管成像、計(jì)算機(jī)斷層血管成像、磁共振血管成像、光學(xué)相干斷層成像和血管內(nèi)超聲[11]。但是對(duì)于皮膚腫瘤，并不像冠狀動(dòng)脈疾病、眼底疾病那樣具有公認(rèn)的關(guān)于血管成像評(píng)估標(biāo)準(zhǔn)[8, 12]。近年來，SS-OCTA的應(yīng)用領(lǐng)域不再局限于眼底成像，皮膚淺層血管成像也越來越得到人們的關(guān)注。由于皮膚病，如炎癥、角化不全等均會(huì)引起皮膚組織微觀結(jié)構(gòu)的病變，而SS-OCTA技術(shù)的成像深度正好可以覆蓋皮膚的全層組織結(jié)構(gòu)，滿足臨床上的需要，更重要的是可以避免傳統(tǒng)的組織活檢所帶來的損傷。同時(shí)偏轉(zhuǎn)OCT還可用于組織表層的燒傷程度的檢測[20]。任杰等[13]利用激光散斑成像和光學(xué)相干層析成像技術(shù)，對(duì)兔耳皮膚真皮層微血管的相關(guān)參數(shù)進(jìn)行無創(chuàng)測量。對(duì)皮膚血管的無創(chuàng)實(shí)時(shí)成像，意味著將SS-OCTA技術(shù)應(yīng)用在腫瘤、皮膚科等疾病上進(jìn)行輔助診斷成為可能，對(duì)于一些腫瘤的早期診斷有很大應(yīng)用前景。

所以，本文在掃頻源光學(xué)相干層析血管成像(SS-OCTA)系統(tǒng)的基礎(chǔ)上，對(duì)接種黑色素瘤的C57BL6小鼠進(jìn)行圖像采集，并作出對(duì)比分析，發(fā)現(xiàn)了腫瘤早期生長時(shí)血管的發(fā)育的異常變化。在早期診斷方面，利用SS-OCTA系統(tǒng)進(jìn)行血管成像優(yōu)于結(jié)構(gòu)成像，表明SS-OCTA系統(tǒng)在皮膚科疾病的臨床診斷方面具有潛在價(jià)值。

2 理論推導(dǎo)

2.1 SS-OCTA系統(tǒng)搭建

SS-OCTA手術(shù)導(dǎo)航系統(tǒng)平臺(tái)的基本設(shè)計(jì)搭建如圖1所示。

掃頻光源發(fā)出近紅外波段的光，5%的光進(jìn)入馬赫-曾德爾干涉儀，其干涉信號(hào)由平衡探測器獲取，用于對(duì)OCT干涉信號(hào)進(jìn)行校準(zhǔn)；95%的光經(jīng)耦合器與環(huán)形器分別進(jìn)入到參考臂和樣品臂中，二維掃描振鏡[14]將樣品臂的光偏轉(zhuǎn)后照射至小鼠皮膚成像區(qū)域，其部分光從樣品內(nèi)部各層組織原路返回，并分別與參考臂返回的光發(fā)生干涉，最終由光電平衡探測器與數(shù)字采集卡記錄，經(jīng)過OCT深度重構(gòu)處理形成包含組織內(nèi)微觀結(jié)構(gòu)的圖像。其橫向掃描可以快速獲取非侵入二維和三維分辨率優(yōu)于10 μm的圖像。光束照射到標(biāo)本上，形成顯微結(jié)構(gòu)反射，對(duì)反射光時(shí)間延遲起到測量作用，可對(duì)組織的縱向內(nèi)部結(jié)構(gòu)進(jìn)行無創(chuàng)檢測。

2.2 分頻幅去相關(guān)血管造影法

本文中SS-OCTA系統(tǒng)的算法采用分頻幅去相關(guān)血管造影法(split spectrum amplitude deeorrelation angiography，SSADA)[15]，能在最大程度上降低噪聲、偽影等的影響，使得信噪比得到明顯提升[16-17]。

光在樣品內(nèi)部散射，然后通過處理形成高分辨率、高深度的圖像來分析內(nèi)在的微觀結(jié)構(gòu)。光束照射到標(biāo)本上，形成顯微結(jié)構(gòu)反射，對(duì)反射光時(shí)間延遲起到測量作用，可對(duì)組織的縱向內(nèi)部結(jié)構(gòu)進(jìn)行無創(chuàng)檢測。一個(gè)光帶所提供的血流信息都是獨(dú)立的，對(duì)多幀掃描圖像所對(duì)應(yīng)的去相干圖像進(jìn)行計(jì)算，將不同光帶去相干圖像進(jìn)行合并處理，就能獲得平均值，使得血流信號(hào)變強(qiáng)，具有重要的臨床研究意義[16]。

SS: 掃頻光源(swept-source)；MZI: 馬赫-曾德爾干涉儀(Mach-Zehnder interferometer)；BD: 平衡探測器(balance detector)；PC: 偏振控制器(polarize controller)；M: 反射鏡(mirror)；G: 振鏡(galvanometer)

因?yàn)镾SADA只對(duì)信號(hào)強(qiáng)度信息進(jìn)行干涉，所以其算法就能對(duì)動(dòng)態(tài)物質(zhì)背景進(jìn)行分離，使得抖動(dòng)影響大大變小，對(duì)流場檢測的信噪比明顯增強(qiáng)，然后利用幅值去相關(guān)技術(shù)提取流動(dòng)信息，如下式[16]：

式中：為相同位置實(shí)現(xiàn)多幀重復(fù)掃描的幀數(shù)索引；為分光譜幀數(shù)索引用；1、分別為分光譜數(shù)和多幀重復(fù)掃描數(shù)。當(dāng)采集時(shí)間間隔不增加時(shí)，增加1就可導(dǎo)致信噪比大大提升。

本文利用SSADA算法對(duì)SS-OCTA系統(tǒng)進(jìn)行成像檢測，結(jié)果如圖2所示。圖中顯示的噪聲基底為70.3 dB，最大信號(hào)強(qiáng)度為104.6 dB，標(biāo)準(zhǔn)圖像的最大信噪比為34.3 dB。

3 成像實(shí)驗(yàn)

本次實(shí)驗(yàn)使用的實(shí)驗(yàn)動(dòng)物為C57BL6小鼠。小鼠經(jīng)麻醉、脫毛后，左背部后肢部位皮下注射0.2 ml B16F10單細(xì)胞懸液，注射細(xì)胞量為105個(gè)，接種5天后進(jìn)行成像。

圖2 利用SSADA算法對(duì)SS-OCTA系統(tǒng)進(jìn)行成像檢測的結(jié)果

本次成像實(shí)驗(yàn)設(shè)定SS-OCTA實(shí)驗(yàn)平臺(tái)的采集視場為5.12 mm′5.12 mm，黑色素瘤因體積大于采集視場，無法單次采集整個(gè)腫瘤的血管圖像。為此，本次實(shí)驗(yàn)以不同方向與角度進(jìn)行采集，將采集到的結(jié)構(gòu)、血管圖像進(jìn)行拼接，形成完整的皮膚黑色素瘤的結(jié)構(gòu)圖與血管圖。圖3分別展示了小鼠腫瘤處的實(shí)物圖、SS-OCTA掃描的結(jié)構(gòu)圖和血管圖。

圖3(a)可以看到，接種后，小鼠腫瘤處皮膚呈黑色，與粉色的正常皮膚有明顯差異。圖3(b)為SS-OCTA掃描采集的結(jié)構(gòu)圖經(jīng)過拼接后的完整皮膚表面。由圖可知腫瘤皮膚表面不平整，存在一些凹凸不平的現(xiàn)象，猜測應(yīng)該是皮膚表面的粗糙程度，并沒有明顯的異?，F(xiàn)象。圖3(c)為SS-OCTA掃描采集的血管圖像經(jīng)過拼接后的黑色素瘤處完整血管圖像，由圖紅色虛線區(qū)域可知，腫瘤部位血管呈現(xiàn)不均勻分布，并且血管分支復(fù)雜、紊亂，很多微血管結(jié)構(gòu)不完整，這些特征跟周圍正常皮膚血管相比尤為突出(紅色箭頭)。并且腫瘤新生的毛細(xì)血管主要分布在腫瘤生長活躍的邊緣，呈現(xiàn)由外向內(nèi)的生長趨勢。

4 分析討論

將小鼠腫瘤處的皮膚結(jié)構(gòu)圖和血管圖進(jìn)行疊加，得到圖4。在圖4中，可以更清晰看到，小鼠黑色素瘤處的皮膚與其皮下異常生長的血管叢并無太大關(guān)系。單單看皮膚結(jié)構(gòu)圖，很難判別皮膚是否異常、是否存在腫瘤。而在看血管圖片時(shí)，則很容易看出血管生長、分布和結(jié)構(gòu)的異常。這說明本文設(shè)計(jì)搭建的SS-OCTA成像系統(tǒng)在對(duì)皮膚黑色素瘤成像時(shí)，可以從血管特征方面對(duì)早期的黑色素瘤進(jìn)行簡要判定，而在皮膚結(jié)構(gòu)界面則很難起到早期判定的作用。相比較而言，SS-OCTA對(duì)于血管特征的分辨要高于對(duì)皮膚組織結(jié)構(gòu)的分辨。

與內(nèi)窺式OCT相比較，在實(shí)時(shí)無創(chuàng)成像的同時(shí)，還可以基本給出腫瘤血管與正常血管的邊界線，這將有助于在手術(shù)切除腫瘤的同時(shí)更少地切除周邊正常組織[18]。相比較對(duì)黑色素瘤進(jìn)行早期輔助診斷的普遍性手段皮膚鏡而言，SS-OCTA成像不僅可以進(jìn)行皮膚表面結(jié)構(gòu)成像，而且還可以進(jìn)行更深層次的皮下血管成像，并且同樣可以做到實(shí)時(shí)、無創(chuàng)成像[19]。同時(shí)，本實(shí)驗(yàn)也證明SS-OCTA可以對(duì)生物組織結(jié)構(gòu)、血管進(jìn)行無創(chuàng)成像，對(duì)一些腫瘤的早期診斷有很大應(yīng)用前景。

圖3 小鼠腫瘤處的皮膚結(jié)構(gòu)圖、血管圖。(a) 小鼠皮膚實(shí)物圖；(b) 小鼠皮膚結(jié)構(gòu)圖；(c) 小鼠皮膚血管圖

圖4 結(jié)構(gòu)圖與血管圖的疊加

5 結(jié) 論

本文利用掃頻源光學(xué)相干層析血管成像(SS-OCTA)系統(tǒng)完成了對(duì)皮膚的結(jié)構(gòu)成像，并對(duì)黑色素瘤小鼠進(jìn)行腫瘤部位的皮膚結(jié)構(gòu)和血管成像。證明了本文的SS-OCTA系統(tǒng)能發(fā)現(xiàn)腫瘤早期生長時(shí)血管發(fā)育的異常變化。同時(shí)，在早期診斷方面，利用SS-OCTA系統(tǒng)進(jìn)行血管成像優(yōu)于結(jié)構(gòu)成像，表明SS-OCTA系統(tǒng)在皮膚科疾病的臨床診斷方面具有潛在價(jià)值。

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Imaging of skin structure and vessels in melanoma by swept source optical coherence tomography angiography

Liu Jingxuan1,2, Fan Jinyu2, Wang Quan2, Shi Guohua2*

1University of Science and Technology of China, Hefei, Anhui 230000, China;2Suzhou Institute of Biomedical Engineering Technology, Chinese Academy of Sciences, Suzhou, Jiangsu 215000, China

The superimposition of the structure map and the vascular map

Overview:In recent years, optical coherence tomography (OCT) has developed rapidly and become a new imaging technology. OCT weakens coherent reflection and backscattering. Super heterodyne detection technique was used to improve the signal-to-noise ratio of biological tissue tomography. OCT has the advantages of non-invasive, high resolution, and high-speed imaging, and thus it is very suitable for biomedical applications. Scanning source optical coherence tomography (SS-OCTA) is a frequency-domain OCT technology and can support a high resolution in vivo angiography. As a new angiography technique, SS-OCTA still uses the Michelson interferometer's basic optical path and can achieve axial resolution of 15 microns by measuring the back scattering of light from low-coherent interference signals in tissue. Cross sectional images of 3D reconstruction of 3D images of biological tissues can be obtained, which are widely used in ophthalmology, dermatology imaging, tumor detection, and other fields. In addition to imaging biological tissue, SS-OCTA can also image surface blood vessels such as fundus and skin. SS-OCTA can observe the changes of retinal blood vessel morphology and blood flow in the choroid retina in the field of ophthalmology such as retinal angiography. Furthermore, it can also use pseudo-color to distinguish normal and abnormal vascular structures, blood flow signal detection and quantitative analysis, split different spectral images of the original full-spectrum image, reduce noise, improve signal-to-noise ratio, and then merge, so as to achieve retinal, choroidal vascular formation of any layer of significant cross-sectional imaging. Finally, we use laser speckle imaging and optical coherence tomography to noninvasive measurement of animal skin irritation and obtain dermal microvascular parameters. Angiography provides a possibility for the applications of SS-OCTA in the diagnosis of tumors, skin diseases, and other diseases. In fact, solid tumor growth is strongly dependent on the induced vascular network. Direct and indirect studies can support a strong evidence that tumor growth depends on blood vessels. Most tumors remain inactive until they become cancerous, and blood vessels no longer grow. Once entering the vascular phase, new blood vessels will grow rapidly to support tumor metabolism and play an important role in tumor proliferation. SS-OCTA can perform noninvasive imaging of biological tissues and blood vessels. This is of great significance for the early diagnosis of some tumors. Therefore, skin structure and angiography of melanoma C57BL6 mice were collected and compared with the SS-OCTA system. To observe the changes of the vascular development and biological tissue structure in the early stage of tumor growth, SS-OCTA is better at distinguishing vascular functional structures than the structural imaging.

Citation: Liu J X, Fan J Y, Wang Q,Imaging of skin structure and vessels in melanoma by swept source optical coherence tomography angiography[J]., 2020, 47(2): 190239

Imaging of skin structure and vessels in melanoma by swept source optical coherence tomography angiography

Liu Jingxuan1,2, Fan Jinyu2, Wang Quan2, Shi Guohua2*

1University of Science and Technology of China, Hefei, Anhui 230000, China;2Suzhou Institute of Biomedical Engineering Technology, Chinese Academy of Sciences, Suzhou, Jiangsu 215000, China

Sweep source optical coherence tomographic angiography (SS-OCTA) is a kind of angiography technologies based on split spectrum amplitude deeorrelation angiography (SSADA). It has a great prospect in the early diagnosis of tumors and other diseases. In this paper, skin structure and angiography of melanoma C57BL6 mice were collected on the basis of the SS-OCTA imaging platform with an imaging field of 5.12 mm′5.12 mm and a standard image maximum signal-to-noise ratio of 34.3 dB. The results show that the SS-OCTA system is superior to the structural imaging in early diagnosis of dermatological diseases.

scanning frequency source optical coherence tomography; skin structure; tumor vessels; melanoma; SS-OCTA

Supported by Key Research and Development Plan of the Ministry of Science and Technology - Digital Diagnosis and Treatment Equipment (2017YFC0108201, 2017YFC0108200, 2017YFB0403700), Key Research Project of Frontier Science of Chinese Academy of Sciences (QYZDB-SSW-JSC03), and Key Projects for Inter-governmental International Scientific and Technological Innovation Cooperation (2016YFE0107700)

A

10.12086/oee.2020.190239

: Liu J X, Fan J Y, Wang Q,. Imaging of skin structure and vessels in melanoma by swept source optical coherence tomography angiography[J]., 2020,47(2): 190239

2019-05-13；

2019-11-23

科技部重點(diǎn)研發(fā)計(jì)劃-數(shù)字診療裝備(2017YFC0108201, 2017YFC0108200, 2017YFB0403700)；中國科學(xué)院前沿科學(xué)重點(diǎn)研究項(xiàng)目(QYZDB-SSW-JSC03)；政府間國際科技創(chuàng)新合作重點(diǎn)專項(xiàng)(2016YFE0107700)

劉敬璇(1993-)，女，碩士研究生，主要從事光學(xué)生物成像的研究。E-mail：ljx1216@126.com

史國華(1981-)，男，博士，研究員，主要從事新型在體光學(xué)成像與檢測的研究。E-mail：ghshi_lab@126.com

劉敬璇，樊金宇，汪權(quán)，等. SS-OCTA對(duì)黑色素瘤皮膚結(jié)構(gòu)和血管的成像實(shí)驗(yàn)[J]. 光電工程，2020，47(2): 190239

R732.2；TP391

* E-mail: ghshi_lab@126.com