人工智能技术在计算机辅助诊断领域的发展新趋势

计算机辅助诊断是医学影像分析发展的重要方向之一.该技术具有精确定量分析、可重复处理,降低影像医师工作负担等优点.本文对近年来国内外基于人工智能技术的计算机辅助诊断技术的研究进展进行了综述和讨论,介绍了当前针对医学影像的计算机辅助诊断系统所涉及的人工智能算法、流程及相关技术,分析了目前研究所存在的不足和挑战,并提出了有可...     

VITEK2 Compact全自动微生物分析系统对牛乳中葡萄球菌的鉴定效果评价

【目的】评价VITEK2 Compact全自动微生物分析系统对牛乳中葡萄球菌属细菌鉴定效果。【方法】对从西宁、白银、西安三地乳样中分离的52株葡萄球菌通过VITEK2 Compact全自动微生物分析系统进行鉴定,并与革兰氏染色、触酶试验、血浆凝固酶试验以及16S r RNA基因序列分析结果进行比对;通过对凝固酶阴性葡萄球菌和凝固酶阳性葡萄球菌在不同麦氏浓度的鉴定,评价该系统的稳定性。【结果】VITEK2 Compact全自动微生物分析系统对葡萄球菌在\"属\"水平的鉴定结果符合率达96.15%,与血浆凝固酶结果对比,凝固酶阴性葡萄球菌属细菌鉴定结果符合率为92.86%,对凝固酶阳性葡萄球菌属细菌鉴定结果符合率为50%,但对具体种鉴定时,结果符合率仅为44.23%。在稳定性检测的试验中,鉴定符合率为100%,鉴定结果可靠性不随菌液浓度变化而变化。【结论】该系统可用于牛乳中葡萄球菌在\"属\"水平的鉴定。     

自动感应照相系统在大熊猫以及同域分布的野生动物研究中的应用

鉴于野生大熊猫种群的濒危现状,已经不允许对其生境进行破坏性或干扰其行为活动较多的调查活动。例如,野生大熊猫个体数量稀少、其栖息地地形复杂或植被茂密,野外直接观察和调查野生大熊猫极为困难。自动感应照相系统是一种非损伤性野生动物调查工具,在很大程度上弥补了传统调查方法的不足,为野生动物的调查和研究提供了新的有效途径。本研究利用自行研究和开发的自动感应照相系统,获得了野生大熊猫及与其同域分布的其它物种的重要生态信息,显示了自动感应照相系统在物种鉴定、区系调查、个体识别、种群监测、性别确定和行为生态学研究等多方面的应用价值[动物学报51(3)：495—500,2005]。     

华南虎、东北虎、孟加拉虎的D-loop和ND5序列及其在系统进化分析中的应用

本文采用PCR和质粒克隆测序方法,获得了华南虎线粒体D-loop区的480 bp序列和东北虎、孟加拉虎线粒体D-loop区的503 bp序列;同时还获得了这三个虎亚种和金钱豹线粒体ND5基因5'端309 bp的部分序列.根据D-loop序列分析,华南虎与孟加拉虎、东北虎的平均距离(p-distance)分别为0.110 88和0.110 87,而东北虎与孟加拉虎间的平均距离为0.009 94;根据ND5序列分析,华南虎与孟加拉虎、东北虎的平均距离分别为0.114 34和0.117 58,而东北虎与孟加拉虎间的平均距离为0.003 24.三个虎亚种的mtDNA D-loop和ND5序列比较表明,华南虎是这三个虎亚种中最为古老的亚种.     

SELEX技术在胰腺癌分子诊断及靶向治疗中应用的研究进展

胰腺癌由于起病隐匿,早期诊断率较低,临床治疗效果差,是目前预后最差的恶性肿瘤之一。目前,临床上尚缺乏有效的非创伤早期筛查胰腺癌的手段,因而胰腺癌的早期诊断和治疗显得尤为重要。近年来,指数富集配基的系统进化(SELEX)技术以其在其他疾病中所表现的应用价值为疾病的诊治提供了一个新的途径。对于缺乏有效确诊手段,发病隐匿且病死率高的胰腺癌而言,SELEX技术基于胰腺癌发病的分子机制,可以筛选出特异结合于胰腺癌分子靶标的适配体,对筛选所得适配体进一步化学修饰,可以实现分子水平成像及靶向治疗,进而达到胰腺癌早期诊治的目的,具有重要的临床意义。本文就SELEX技术在胰腺癌分子诊断及靶向治疗中的应用研究进展进行了综述。     

猫中缝-脊髓5-羟色胺能和阿片肽能下行抑制系统在抗内脏伤害性反应中作用的比较

在无麻醉的麻痹猫,以伤害性电刺激内脏大神经传入纤维诱发的连续内脏躯体反射放电为指标,分别刺激腓总神经、延髓中缝大核区以及包括导水管周围灰质、中央中核在内的脑区以产生相应的抑制效应。分别全身与脊髓硬膜内给予纳洛酮以及对氯苯丙胺与羟甲丙基甲基麦角酰胺(Methysergide),观察各种抑制效应的变化。发现:(1)静脉注射纳洛酮虽在不少的动物上能暂时地逆转上述效应,但逆转的程度都是不完全的,硬膜内给药也有类似作用;(2)静脉或腹腔内注射对氯苯丙胺一般能较完全而持久地阻断这些抑制效应,但对腓总神经的抑制效应影响较小,硬膜内给药也有相似的效果;(3)羟甲丙基甲基麦角酰胺(静脉或硬膜内注射)主要阻断腓总神经的抑制效应,对中缝大核的抑制效应影响较小。上述结果显示,阿片肽能和5-羟色胺能下行系统均直接参与内脏痛觉传递的延髓中缝-脊髓调节机制,但后者所起的作用可能更大。     

循环肿瘤细胞在肺癌诊断中的应用及临床意义

为了探讨循环肿瘤细胞(circulating tumor cells,CTCs)在肺癌诊断中的应用及临床意义,本研究收集了我院2014年10月至2017年12月收治并确诊的肺癌患者87例、健康体检者40名以及肺部良性疾病患者50例作为研究对象,采用人循环肿瘤细胞试剂盒测定外周血CTC水平,两组间比较采用Mann-Whitney U检验,多组间比较采用Kruskal-Wallis检验,利用ROC曲线评价CTC在肺癌诊断上的灵敏度以及特异度。实验发现肺癌患者CTC水平(M=12.67 Unints/3 mL)显著高于肺部良性病患者(M=4.76 Unints/3 mL)和健康者(M=4.48 Unints/3 mL),差异存在统计学意义(p<0.001)。在不同性别、不同年龄段以及不同病理类型之间肺癌患者的CTC水平比较差异无统计学意义(p>0.05)。Ⅲ+Ⅳ期肺癌患者CTC水平显著高于Ⅰ+Ⅱ期患者,差异存在统计学意义(p<0.001)。有远处转移的肺癌患者CTC水平显著高于无远处转移的肺癌患者,差异存在统计学意义(p<0.001)。ROC曲线下面积为0.878(95%CI 0.820~0.936),临界值为6.34 Unints/3 mL,对应的灵敏度为0.77,特异度为0.989。本研究初步认为,CTC检测对肺癌诊断具有较高的灵敏度以及特异度,可能存在重要的临床应用价值。     

Detection of Circulating Tumor Cells and Circulating Tumor Microemboli in Gastric Cancer

PURPOSE: Gastric cancer studies indicated a potential correlation between circulating tumor cells (CTCs) in peripheral blood and tumor relapse/metastasis. The prevalence and significance of circulating tumor microemboli (CTM) in gastric cancer remain unknown. We investigated the prevalence and prognostic value of CTCs and CTM for progression-free survival (PFS) and overall survival (OS) in gastric cancer patients. METHODS:Eighty-one gastric cancer patients consented to provide 5 ml of peripheral blood before systematic therapy. CTCs and CTM were isolated using isolation by size of epithelial tumor cells and characterized by cytopathologists. For 41 stage IV gastric cancer patients, CTM was investigated as a potential biomarker to predict prognosis. RESULTS:CTCs were detected in 51 patients; the average count was 1.81. In clinical stage I, II, III, and IV patients, the average CTC counts were 1.40, 0.67, 1.24, and 2.71, respectively. CTM were detected in 3 of 33 clinical stage I to IIIb patients, at an average of 0.12 (0-2). CTM were detected in 13 of 53 clinical stage IIIc to IV patients, at an average of 1.26 (0-22). In stage IV patients, CTM positivity correlated with the CA125 level. PFS and OS in CTM-positive patients were significantly lower than in CTM-negative patients (P < .001). CTM positivity was an independent factor for determining the PFS (P = .016) and OS (P = .003) of stage IV patients in multivariate analysis. Using markers of the epithelial-mesenchymal transition, single CTCs were divided into three phenotypes including epithelial CTCs, biphenotypic epithelial/mesenchymal CTCs, and mesenchymal CTCs. For CTM, CK?/Vimentin+/CD45? and CK+/Vimentin+/CD45? phenotypes were observed, but the CK+/Vimentin?/CD45? CTM phenotype was not. CA125 was detected in gastric cancer cell lines BGC823 and MGC803. CONCLUSIONS: In stage IV patients, CTM positivity was correlated with serum CA125 level. CTM were an independent predictor of shorter PFS and OS in stage IV patients. Thus, CTM detection may be a useful tool to predict prognosis in stage IV patients.     

Mechanical Aspects of Microtubule Bundling in Taxane-Treated Circulating Tumor Cells

Microtubules play an important role in many cellular processes, including mitotic spindle formation and cell division. Taxane-based anticancer treatments lead to the stabilization of microtubules, thus preventing the uncontrolled proliferation of tumor cells. One of the striking physical features of taxane-treated cells is the localization of their microtubules, which can be observed via fluorescent microscopy as an intense fluorescent band and are referred to as a microtubule bundle. With the recent advances in capturing and analyzing tumor cells circulating in a patient’s blood system, there is increasing interest in using these cells to examine a patient’s response to treatment. This includes taxanes that are used routinely in clinics to treat prostate, breast, lung, and other cancers. Here, we have used a computational model of microtubule mechanics to investigate self-arrangement patterns of stabilized microtubules, which allowed for the identification of specific combinations of three physical parameters: microtubule stiffness, intracellular viscosity, and cell shape, that can prevent the formation of microtubule bundles in cells with stabilized microtubules, such as taxane-treated cells. We also developed a method to quantify bundling in the whole microtubule aster structure and a way to compare the simulated results to fluorescent images from experimental data. Moreover, we investigated microtubule rearrangement in both suspended and attached cells and showed that the observed final microtubule patterns depend on the experimental protocol. The results from our computational studies can explain the heterogeneous bundling phenomena observed via fluorescent immunostaining from a mechanical point of view without relying on heterogeneous cellular responses to the microtubule-stabilizing drug.     

EpCAM-Independent Enrichment of Circulating Tumor Cells in Metastatic Breast Cancer

Circulating tumor cells (CTCs) are the potential precursors of metastatic disease. Most assays established for the enumeration of CTCs so far–including the gold standard CellSearch—rely on the expression of the cell surface marker epithelial cell adhesion molecule (EpCAM). But, these approaches may not detect CTCs that express no/low levels of EpCAM, e.g. by undergoing epithelial-to-mesenchymal transition (EMT). Here we present an enrichment strategy combining different antibodies specific for surface proteins and extracellular matrix (ECM) components to capture an EpCAM^low/neg cell line and EpCAM^neg CTCs from blood samples of breast cancer patients depleted for EpCAM-positive cells. The expression of respective proteins (Trop2, CD49f, c-Met, CK8, CD44, ADAM8, CD146, TEM8, CD47) was verified by immunofluorescence on EpCAM^pos (e.g. MCF7, SKBR3) and EpCAM^low/neg (MDA-MB-231) breast cancer cell lines. To test antibodies and ECM proteins (e.g. hyaluronic acid (HA), collagen I, laminin) for capturing EpCAM^neg cells, the capture molecules were first spotted in a single- and multi-array format onto aldehyde-coated glass slides. Tumor cell adhesion of EpCAM^pos/neg cell lines was then determined and visualized by Coomassie/MitoTracker staining. In consequence, marginal binding of EpCAM^low/neg MDA-MB-231 cells to EpCAM-antibodies could be observed. However, efficient adhesion/capturing of EpCAM^low/neg cells could be achieved via HA and immobilized antibodies against CD49f and Trop2. Optimal capture conditions were then applied to immunomagnetic beads to detect EpCAM^neg CTCs from clinical samples. Captured CTCs were verified/quantified by immunofluorescence staining for anti-pan-Cytokeratin (CK)-FITC/anti-CD45 AF647/DAPI. In total, in 20 out of 29 EpCAM-depleted fractions (69%) from 25 metastatic breast cancer patients additional EpCAM^neg CTCs could be identified [range of 1–24 CTCs per sample] applying Trop2, CD49f, c-Met, CK8 and/or HA magnetic enrichment. EpCAM^neg dual-positive (CK^pos/CD45^pos) cells could be traced in 28 out of 29 samples [range 1–480]. By single-cell array-based comparative genomic hybridization we were able to demonstrate the malignant nature of one EpCAM^neg subpopulation. In conclusion, we established a novel enhanced CTC enrichment strategy to capture EpCAM^neg CTCs from clinical blood samples by targeting various cell surface antigens with antibody mixtures and ECM components.     

Mechanical Aspects of Microtubule Bundling in Taxane-Treated Circulating Tumor Cells

Microtubules play an important role in many cellular processes, including mitotic spindle formation and cell division. Taxane-based anticancer treatments lead to the stabilization of microtubules, thus preventing the uncontrolled proliferation of tumor cells. One of the striking physical features of taxane-treated cells is the localization of their microtubules, which can be observed via fluorescent microscopy as an intense fluorescent band and are referred to as a microtubule bundle. With the recent advances in capturing and analyzing tumor cells circulating in a patient’s blood system, there is increasing interest in using these cells to examine a patient’s response to treatment. This includes taxanes that are used routinely in clinics to treat prostate, breast, lung, and other cancers. Here, we have used a computational model of microtubule mechanics to investigate self-arrangement patterns of stabilized microtubules, which allowed for the identification of specific combinations of three physical parameters: microtubule stiffness, intracellular viscosity, and cell shape, that can prevent the formation of microtubule bundles in cells with stabilized microtubules, such as taxane-treated cells. We also developed a method to quantify bundling in the whole microtubule aster structure and a way to compare the simulated results to fluorescent images from experimental data. Moreover, we investigated microtubule rearrangement in both suspended and attached cells and showed that the observed final microtubule patterns depend on the experimental protocol. The results from our computational studies can explain the heterogeneous bundling phenomena observed via fluorescent immunostaining from a mechanical point of view without relying on heterogeneous cellular responses to the microtubule-stabilizing drug.     

Size-Based Isolation of Circulating Tumor Cells in Lung Cancer Patients Using a Microcavity Array System

Background

Epithelial cell adhesion molecule (EpCAM)-based enumeration of circulating tumor cells (CTC) has prognostic value in patients with solid tumors, such as advanced breast, colon, and prostate cancer. However, poor sensitivity has been reported for non-small cell lung cancer (NSCLC). To address this problem, we developed a microcavity array (MCA) system integrated with a miniaturized device for CTC isolation without relying on EpCAM expression. Here, we report the results of a clinical study on CTCs of advanced lung cancer patients in which we compared the MCA system with the CellSearch system, which employs the conventional EpCAM-based method.

Methods

Paired peripheral blood samples were collected from 43 metastatic lung cancer patients to enumerate CTCs using the CellSearch system according to the manufacturer’s protocol and the MCA system by immunolabeling and cytomorphological analysis. The presence of CTCs was assessed blindly and independently by both systems.

Results

CTCs were detected in 17 of 22 NSCLC patients using the MCA system versus 7 of 22 patients using the CellSearch system. On the other hand, CTCs were detected in 20 of 21 small cell lung cancer (SCLC) patients using the MCA system versus 12 of 21 patients using the CellSearch system. Significantly more CTCs in NSCLC patients were detected by the MCA system (median 13, range 0–291 cells/7.5 mL) than by the CellSearch system (median 0, range 0–37 cells/7.5 ml) demonstrating statistical superiority (p = 0.0015). Statistical significance was not reached in SCLC though the trend favoring the MCA system over the CellSearch system was observed (p = 0.2888). The MCA system also isolated CTC clusters from patients who had been identified as CTC negative using the CellSearch system.

Conclusions

The MCA system has a potential to isolate significantly more CTCs and CTC clusters in advanced lung cancer patients compared to the CellSearch system.     

Classification of Circulating Tumor Cells by Epithelial-Mesenchymal Transition Markers

In cancer, epithelial-mesenchymal transition (EMT) is associated with metastasis. Characterizing EMT phenotypes in circulating tumor cells (CTCs) has been challenging because epithelial marker-based methods have typically been used for the isolation and detection of CTCs from blood samples. The aim of this study was to use the optimized CanPatrol CTC enrichment technique to classify CTCs using EMT markers in different types of cancers. The first step of this technique was to isolate CTCs via a filter-based method; then, an RNA in situ hybridization (RNA-ISH) method based on the branched DNA signal amplification technology was used to classify the CTCs according to EMT markers. Our results indicated that the efficiency of tumor cell recovery with this technique was at least 80%. When compared with the non-optimized method, the new method was more sensitive and more CTCs were detected in the 5-ml blood samples. To further validate the new method, 164 blood samples from patients with liver, nasopharyngeal, breast, colon, gastric cancer, or non-small-cell lung cancer (NSCLC) were collected for CTC isolation and characterization. CTCs were detected in 107(65%) of 164 blood samples, and three CTC subpopulations were identified using EMT markers, including epithelial CTCs, biophenotypic epithelial/mesenchymal CTCs, and mesenchymal CTCs. Compared with the earlier stages of cancer, mesenchymal CTCs were more commonly found in patients in the metastatic stages of the disease in different types of cancers. Circulating tumor microemboli (CTM) with a mesenchymal phenotype were also detected in the metastatic stages of cancer. Classifying CTCs by EMT markers helps to identify the more aggressive CTC subpopulation and provides useful evidence for determining an appropriate clinical approach. This method is suitable for a broad range of carcinomas.     

Biomolecular Surfaces for the Capture and Reprogramming of Circulating Tumor Cells

Circulating Tumor Cells(CTC)have the potential to be used clinically as a diagnostic tool and a treatment tool in the fieldof oncology.As a diagnostic tool,CTC may be used to indicate the presence of a tumor before it is large enough to cause noticeablesymptoms.As a treatment tool,CTC isolated from patients may be used to test the efficacy of chemotherapy options topersonalize patient treatment.One way for tumors to spread is through metastasis via the circulatory system.CTC are able toexploit the natural leukocyte recruitment process that is initially mediated by rolling on transient selectin bonds.Our capturedevices take advantage of this naturally occurring recruitment step to isolate CTC from whole blood by flowing samples throughselectin and antibody-coated microtubes.Whole blood was spiked with a known concentration of labeled cancer cells and thenperfused through pre-coated microtubes.Microtubes were then rinsed to remove unbound cells and the number of labeled cellscaptured on the lumen was assessed.CTC were successfully captured from whole blood at a clinically relevant level on the orderof 10 cells per mL.Combination tubes with selectin and antibody coated surface exhibited higher capture rate than tubes coatedwith selectin alone or antibody alone.Additionally,CTC capture was demonstrated with the KG 1 a hematopoietic cell line andthe DU 145 epithelial cell line.Thus,the in vivo process of selectin-mediated CTC recruitment to distant vessel walls can be usedin vitro to target CTC to a tube lumen.The biomolecular coatings can also be used to capture CTC of hematopoietic andepithelial tumor origin and is demonstrated to sensitivities down to the order of 10 CTC per mL.In a related study aimed at reducing the blood borne metastatic cancer load,we have shown that cells captured to a surfacecan be neutralized by a receptor-mediated biochemical signal.In the proposed method we have shown that using a combinedselectin and TRAIL(TNF Related Apoptosis Inducing Ligand or Apo 2L)functionalized surface we are abl     

Mutational Analysis of Circulating Tumor Cells from Colorectal Cancer Patients and Correlation with Primary Tumor Tissue

Circulating tumor cells (CTCs) provide a non-invasive accessible source of tumor material from patients with cancer. The cellular heterogeneity within CTC populations is of great clinical importance regarding the increasing number of adjuvant treatment options for patients with metastatic carcinomas, in order to eliminate residual disease. Moreover, the molecular profiling of these rare cells might lead to insight on disease progression and therapeutic strategies than simple CTCs counting. In the present study we investigated the feasibility to detect KRAS, BRAF, CD133 and Plastin3 (PLS3) mutations in an enriched CTCs cell suspension from patients with colorectal cancer, with the hypothesis that these genes` mutations are of great importance regarding the generation of CTCs subpopulations. Subsequently, we compared CTCs mutational status with that of the corresponding primary tumor, in order to access the possibility of tumor cells characterization without biopsy. CTCs were detected and isolated from blood drawn from 52 colorectal cancer (CRC) patients using a quantum-dot-labelled magnetic immunoassay method. Mutations were detected by PCR-RFLP or allele-specific PCR and confirmed by direct sequencing. In 52 patients, discordance between primary tumor and CTCs was 5.77% for KRAS, 3.85% for BRAF, 11.54% for CD133 rs3130, 7.69% for CD133 rs2286455 and 11.54% for PLS3 rs6643869 mutations. Our results support that DNA mutational analysis of CTCs may enable non-invasive, specific biomarker diagnostics and expand the scope of personalized medicine for cancer patients.