多靶点全景数字病理：从原理到应用 -

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多靶点全景数字病理：从原理到应用

doi:10.37188/CO.2022-0091

1.
海南省生物医学工程重点实验室, 海南大学生物医学工程学院, 海南海口 570100
2.
海南大学计算机科学与技术学院, 海南海口 570100

基金项目:国家自然科学基金地区项目（No. 82160345）；海南省重点研发项目（No. ZDYF2021GXJS017）；海口市重点科技计划项目（No. 2021-016)；海南省自然科学基金（No. 620RC558）

详细信息

作者简介:
张新华（1995—），女，河南安阳人，博士研究生；2021年于海南大学获得硕士学位，主要从事图像处理方面的研究。E-mail：xinhuazhang@hainanu.edu.cn
史　冰（1991—），女，吉林白城人，博士，讲师；2014年于长春理工大学获得学士学位，2021年于中国科学院大学获得博士学位，主要从事微流控液滴分析及临床细胞、组织样本的快速检测分析研究。E-mail：shibing@hainanu.edu.cn

中图分类号:TP394.1;TH691.9
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出版历程
- 收稿日期:2022-05-09
- 修回日期:2022-05-31
- 网络出版日期:2022-07-02

Multi-target panoramic digital pathology: from principle to application

1.
Key Laboratory of Biomedical Engineering of Hainan Province, School of Biomedical Engineering, Hainan University, Haikou 570100, China
2.
School of Computer Science and Technology, Hainan University, Haikou 570100, China

Funds:Supported by Key National Natural Science Foundation of China (No. 82160345); Research and Development Program of Hainan Province (No. ZDYF2021GXJS017); Key Science andTechnology Plan Project of Haikou (No. 2021-016); Hainan Provincial Natural Science Foundation of China (No. 620RC558)

More Information

Corresponding author:shibing@hainanu.edu.cn

摘要

摘要:
数字病理凭借其便捷的存储、管理、浏览、传输等特点，为远程病理会诊及联合会诊带来了新契机。然而，显微镜的视场有限，在保证分辨率的前提下，无法兼顾全景成像。全景数字病理的提出弥补了这一缺陷，其在保证分辨率的同时可兼顾全景成像。但单张切片仅能实现单靶点检测，而疾病诊断需同时观测多个靶点的表达情况。近年来，多靶点全景数字病理技术发展迅速，因其在药物研发、临床科研以及基础科研等领域有巨大的应用潜力而广受关注。该系统凭借视场大、颜色多、通量高的特点，可在短时间内原位检测整张组织切片上的多种生物标记物的表达情况，借以识别组织上每个细胞表型、丰度、状态及其相互关系。本文首先梳理了数字病理、全景数字病理以及多靶点全景数字病理的发展过程，并简要介绍发展过程中技术的更新迭代，以及发展多靶点全景数字病理的重要性。然后，分别从生物样本准备、多色光学成像以及图像处理3个部分重点介绍多靶点全景数字病理。接下来，阐述了多靶点全景数字病理在肿瘤微环境与肿瘤分子分型等生物医学领域的应用情况。最后，对多靶点全景数字病理的技术优势、目前面临的挑战及其未来的发展趋势进行了总结。
- 多靶点全景数字病理/
- 生物标记物/
- 多色成像/
- 图像处理
Abstract:
Digital pathology has brought new opportunities for remote pathological consultation and joint consultation owing to its convenient storage, management, browsing and transmission. However, because of the limited field of view of a microscope, panoramic imaging cannot be achieved while ensuring a high resolution. The proposal of panoramic digital pathology makes up for this defect and achieves panoramic imaging while ensuring high resolution. However, a single slice can only detect a single target, and disease diagnosis needs to observe the expression of multi-target at the same time. In recent years, multi-target panoramic digital pathology technology has developed rapidly. It has attracted much attention because of its great application potential in drug research and development, clinical research and basic research. Owing to its large field of view, wide range of colors and high flux, the system can detect the expression of various biomarkers on a whole tissue section in situ in a short time to identify the phenotype, abundance, state, and relationship of each cell. Firstly, this paper reviews the development process of digital pathology, panoramic digital pathology and multi-target panoramic digital pathology, as well as the update and iteration of technology in the development process, and illustrates the importance of developing multi-target panoramic digital pathology. Then, the multi-target panoramic digital pathology is described in detail from three perspectives: biological sample preparation, multi-color imaging system and image processing. Next, the applications of multi-target panoramic digital pathology in biomedical fields, such as tumor microenvironments and tumor molecular typing are described. Finally, the advantages, challenges and future development of multi-target panoramic digital pathology are summarized.
- multi-target panoramic digital pathology/
- biomarkers/
- multi-color imaging/
- image processing

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图 1常规免疫组织化学实验流程

Figure 1.Conventional immunohistochemistry staining procedures

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图 2基于TSA的多重荧光免疫组化流程

Figure 2.Multiplex fluorescent immunohistochemistry based on TSA

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图 3光谱成像示意图^[68]（F：滤光片；BS：分光镜；M1/M2/M3：反射镜；P：色散棱镜）

Figure 3.Schematic diagram of spectral imaging^[68](F: filter; BS: beam splitter; M1/M2/M3: mirror; P: dispersive prism)

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图 4面扫描和线扫描^[77]

Figure 4.Area scanning and line scan^[77]

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图 5Olympus VS200 研究级全玻片扫描系统^[84]

Figure 5.Olympus VS200 research-grade slide scanner^[84]

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图 6全景数字病理图像拼接流程

Figure 6.Mosaic process of panoramic digital pathological images

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图 7肿瘤微环境^[109]

Figure 7.Tumor micro environment^[109]

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图 8三级淋巴结构免疫标记荧光成像^[114]

Figure 8.Immunolabeling fluorescence imaging of tertiary lymphoid structures^[114]

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表 1靶点选择原则

Table 1.Target selection principles

疾病	靶点	功能	表达部位
非小细胞癌^[22]	CK	一种细胞角蛋白，是上皮来源的肿瘤标志物^[23]	主要在上皮细胞中表达的中间丝蛋白^[23]
	CD68	巨噬细胞标记物^[24]	存在于骨髓和神经的吞噬细胞^[25]
	Siglec-15	作为免疫调节靶点在肿瘤微环境能抑制抗原特异性T细胞反应^[22]	骨髓细胞，人类癌细胞和肿瘤浸润性骨髓细胞^[22]
肝癌组织^[26]	PD-L1	对抗PD-1/PD-L1治疗反应的预测性生物标志物^[27]	骨髓、淋巴、正常上皮细胞和癌症中组成型表达或诱导的共抑制受体^[27]
	CD68	同上	同上
	CD33	髓系细胞分化抗原^[28]	主要分布在髓系血细胞^[28]
	CD57	人类自然杀伤细胞标记物^[29]	人类自然杀伤细胞和T淋巴细胞^[29]
	CD11b	粘附分子和介导多种配体识别的膜受体^[30]	在吞噬细胞、B细胞和T细胞的次要亚群以及自然杀伤细胞上表达^[30]
	CD20	B细胞的表面抗原^[31]	在B淋巴细胞上表达^[32]
肺癌组织^[33]	PD-L1	同上	同上
	PD-1	在活化的T细胞中诱导的抑制性受体^[34]	活化的T、自然杀伤和B淋巴细胞、巨噬细胞、树突状细胞和单核细胞上表达^[35]
	CD8	细胞毒性T淋巴细胞标志物^[36]	所有细胞毒性T淋巴细胞或杀伤细胞上^[36]
	FoxP3	调节性T细胞的标志性分子^[37]	调节性T细胞和正常上皮细胞及多种肿瘤细胞中^[37]
	CD68	同上	同上
	CK	同上	同上
头颈癌组织^[38]	PD-1	同上	同上
	OX40	免疫调节蛋白，主要由T细胞表达^[39]	活化的CD4+和CD8+上表达、T细胞上表达以及许多其他淋巴和非淋巴细胞^[39]
	FoxP3	同上	同上
	CD3	T细胞标志物^[40]	存在于T细胞表面^[40]
乳腺癌组织^[41]	CD103	肿瘤浸润调节性T细胞的标志^[42]	肠道粘膜上皮内的T细胞群和肠固有层白细胞上表达^[42]
乳腺癌组织^[41]	CD8	同上	同上

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表 2全景数字病理设备产品主要参数

Table 2.The main parameters of panoramic digital pathology equipments

公司	型号	成像模式	切片容量	视场	成像速度
Zeiss	Axio Scan.Z1	明场，荧光，偏振（选配）	12或100片（选配）	15 mm × 15 mm	20×明场：240 s/片；荧光：NA
Zeiss	Axioscan 7	明场，荧光，偏振（选配）	100片	10 mm ×10 mm	20×明场：73 s/片 20×荧光：（4通道）：323 s/片
（ps: non-available, NA）

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