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1.
Various single-cell retention structures (SCRSs) were reported for analysis of single cells within microfluidic devices. Undesirable flow behaviors within micro-environments not only influence single-cell manipulation and retention significantly but also lead to cell damage, biochemical heterogeneity among different individual cells (e.g., different cell signaling pathways induced by shear stress). However, the fundamentals in flow behaviors for single-cell manipulation and shear stress reduction, especially comparison of these behaviors in different microstructures, were not fully investigated in previous reports. Herein, flow distribution and induced shear stress in two different single-cell retention structures (SCRS I and SCRS II) were investigated in detail to study their effects on single-cell trapping using computational fluid dynamics (CFD) methods. The results were successfully verified by experimental results. Comparison between these two SCRS shows that the wasp-waisted configuration of SCRS II has a better performance in trapping and manipulating long cylinder-shaped cardiac myocytes and provides a safer “harbor” for fragile cells to prevent cell damage due to the shear stress induced from strong flows. The simulation results have not only explained flow phenomena observed in experiments but also predict new flow phenomena, providing guidelines for new chip design and optimization, and a better understanding of the cell micro-environment and fundamentals of microfluidic flows in single-cell manipulation and analysis. 相似文献
2.
Interest in single-cell analysis has increased because it allows to understand cell metabolism and characterize disease states, cellular adaptation to environmental changes, cell cycles, etc. Here, the authors propose a device to electrically trap and lyse single-bacterial cells in an array format for high-throughput single-cell analysis. The applied electric field is highly deformed and concentrated toward the inside of the microwell structures patterned on the planar electrode. This configuration effectively generates dielectrophoretic force to attract a single cell per well. The microwell has a comparable size to the target bacterial cell making it possible to trap single cells by physically excluding additional cells. Inducing highly concentrated electric potential on the cell membrane can also effectively lyse the trapped single-bacterial cells. The feasibility of the authors' approach was demonstrated by trapping and lysing Escherichia coli cells at the single-cell level. The present microwell array can be used as a basic tool for individual bacterial cell analysis. 相似文献
3.
Circulating tumor cells (CTCs) are the principal vehicle for the spread of non-hematologic cancer disease from a primary tumor, involving extravasation of CTCs across blood vessel walls, to form secondary tumors in remote organs. Herein, a polydimethylsiloxane-based microfluidic system is developed and characterized for in vitro systematic studies of organ-specific extravasation of CTCs. The system recapitulates the two major aspects of the in vivo extravasation microenvironment: local signaling chemokine gradients in a vessel with an endothelial monolayer. The parameters controlling the locally stable chemokine gradients, flow rate, and initial chemokine concentration are investigated experimentally and numerically. The microchannel surface treatment effect on the confluency and adhesion of the endothelial monolayer under applied shear flow has also been characterized experimentally. Further, the conditions for driving a suspension of CTCs through the microfluidic system are discussed while simultaneously maintaining both the local chemokine gradients and the confluent endothelial monolayer. Finally, the microfluidic system is utilized to demonstrate extravasation of MDA-MB-231 cancer cells in the presence of CXCL12 chemokine gradients. Consistent with the hypothesis of organ-specific extravasation, control experiments are presented to substantiate the observation that the MDA-MB-231 cell migration is attributed to chemotaxis rather than a random process. 相似文献
4.
This review presents an overview of literature that describes the applications of microfluidics to assay individual cells. We quantify the content of an individual mammalian cell, so that we can understand what criteria a single-cell assay must satisfy to be successful. We put in context the justification for single-cell assays and identify the characteristics that are relevant to single-cell assays. We review the literature from the past 24 months that describe the methods that use microfabrication-conventional or otherwise-and microfluidics in particular to study individual cells, and we present our views on how an increasing emphasis on three-dimensional cell culture and the demonstration of the first chemically defined cell might impact single-cell assays. 相似文献
5.
Xiaowei Xie Mengyao Liu Yawen Zhang Bingrui Wang Caiying Zhu Chenchen Wang Qing Li Yingying Huo Jiaojiao Guo Changlu Xu Linping Hu Aiming Pang Shihui Ma Lina Wang Wenbin Cao Shulian Chen Qiuling Li Sudong Zhang Xueying Zhao Wen Zhou Hongbo Luo Guoguang Zheng Erlie Jiang Sizhou Feng Lixiang Chen Lihong Shi Hui Cheng Sha Hao Ping Zhu Tao Cheng 《国家科学评论(英文版)》2021,8(3)
6.
单细胞内组分的分析和测定对疾病的治疗和药物筛选具有重要的意义。本文比较了两种单细胞组分分析的方法即毛细管电泳和微流控芯片,并叙述了微流控芯片在单细胞研究方面的进展。 相似文献
7.
Vibha Singh Neena Srivastava Priyanka Srivastava Rama Devi Mittal 《Indian journal of clinical biochemistry : IJCB》2013,28(3):259-264
Chemokine are small, inducible pro-inflammatory cytokines involved in many biological processes, such as migration of leukocytes, atherosclerosis, angiogenesis, tumor growth, and metastasis. Chemokine are also known to influence tumor cell’s activity. Specifically, tumor cells express chemokine receptors in a non random manner suggesting a role of chemokine in metastatic destination of tumor cells. The present study was conducted to determine distribution of (Chemokine receptor 2) CCR2 V64I, Chemokine ligand 2 CCL2 I/D, and CCL2 2518 A>G gene polymorphisms in North Indian population and compare with different populations globally. Polymerase chain reaction (PCR)-based analysis was conducted in 200 normal healthy individuals of similar ethnicity. Allelic frequencies in wild type (GG) of CCR2 V64I G>A were 63 % G; CCL2 I/D 42 % II; CCL2 2518 A>G 40.5 % A. The minor variant allele frequency in our population was as follows: 19.5 % for CCR2 V64I, 35.5 % for CCL2 I/D, 35.3 % for CCL2 2518 A>G. We further compared frequency distribution for these genes with various published studies in different ethnicity. Our results suggested that frequency in chemokine genes exhibit distinctive pattern in India that could be attributed to ethnicity variation. This could assist in high-risk screening of human exposed to environmental carcinogens and cancer predisposition in different ethnic groups. Thus, they signify an impact of ethnicity and provide a basis for future epidemiological and clinical studies. 相似文献
8.
Mei-Zhen Zou Wen-Long Liu Han-Shi Chen Xue-Feng Bai Fan Gao Jing-Jie Ye Han Cheng Xian-Zheng Zhang 《国家科学评论(英文版)》2021,8(2)
The hypoxic tumor microenvironment is characterized by disordered vasculature and rapid proliferation of tumors, resulting from tumor invasion, progression and metastasis. The hypoxic conditions restrict efficiency of tumor therapies, such as chemotherapy, radiotherapy, phototherapy and immunotherapy, leading to serious results of tumor recurrence and high mortality. Recently, research has concentrated on developing functional nanomaterials to treat hypoxic tumors. In this review, we categorize such nanomaterials into (i) nanomaterials that elevate oxygen levels in tumors for enhanced oxygen-dependent tumor therapy and (ii) nanomaterials with diminished oxygen dependence for hypoxic tumor therapy. To elevate oxygen levels in tumors, oxygen-carrying nanomaterials, oxygen-generating nanomaterials and oxygen-economizing nanomaterials can be used. To diminish oxygen dependence of nanomaterials for hypoxic tumor therapy, therapeutic gas-generating nanomaterials and radical-generating nanomaterials can be used. The biocompatibility and therapeutic efficacy of these nanomaterials are discussed. 相似文献
9.
K. Hockemeyer C. Janetopoulos A. Terekhov W. Hofmeister A. Vilgelm Lino Costa J. P. Wikswo A. Richmond 《Biomicrofluidics》2014,8(4)
Stromal cells in the tumor microenvironment play a key role in the metastatic properties of a tumor. It is recognized that cancer-associated fibroblasts (CAFs) and endothelial cells secrete factors capable of influencing tumor cell migration into the blood or lymphatic vessels. We developed a microfluidic device that can be used to image the interactions between stromal cells and tumor cell spheroids in a three dimensional (3D) microenvironment while enabling external control of interstitial flow at an interface, which supports endothelial cells. The apparatus couples a 200-μm channel with a semicircular well to mimic the interface of a blood vessel with the stroma, and the design allows for visualization of the interactions of interstitial flow, endothelial cells, leukocytes, and fibroblasts with the tumor cells. We observed that normal tissue-associated fibroblasts (NAFs) contribute to the “single file” pattern of migration of tumor cells from the spheroid in the 3D microenvironment. In contrast, CAFs induce a rapid dispersion of tumor cells out of the spheroid with migration into the 3D matrix. Moreover, treatment of tumor spheroid cultures with the chemokine CXCL12 mimics the effect of the CAFs, resulting in similar patterns of dispersal of the tumor cells from the spheroid. Conversely, addition of CXCL12 to co-cultures of NAFs with tumor spheroids did not mimic the effects observed with CAF co-cultures, suggesting that NAFs produce factors that stabilize the tumor spheroids to reduce their migration in response to CXCL12. 相似文献
10.
By combined use of traditional Chinese acupuncture Fe needle electrode and in vivo electrochemistry, we achieved in vivo H2 generation in tumors in a controllable manner and exploited it for effective and green therapy of tumors for the first time. The cathodic acupuncture electrodes working under an applied voltage of ∼3 V (with minimal damage to the living body) undergo effective electrochemical reactions in the acidic tumor area that produce sufficient H2 locally to cause cancer cells to burst and die. Due to puncture positioning, the acidic tumor microenvironment and gas diffusion effect, the developed H2 generation electrochemotherapy (H2-ECT) strategy enables precise and large-scale tumor therapy, as demonstrated by in vivo treatment of diseased mice (glioma and breast cancers). Such green H2-ECT is simple, highly efficient and minimally invasive, requiring no expensive medical equipment or nano materials and medication, and is therefore very promising for potential clinical applications. 相似文献
11.
A sequential and high-throughput single-cell manipulation system for a large volume of cells was developed and the successive manipulation for single cell involving single-cell isolation, individual labeling, and individual rupture was realized in a microhydrodynamic flow channel fabricated by using two-dimensional simple flow channels. This microfluidic system consisted of the successive single-cell handlings of single-cell isolation from a large number of cells in cell suspension, labeling each isolated single cell and the lysate extraction from each labeled single cell. This microfluidic system was composed of main channels, cell-trapping pockets, drain channels, and single-cell content collection channels which were fabricated by polydimethylsiloxane. We demonstrated two kinds of prototypes for sequential single-cell manipulations, one was equipped with 16 single-cell isolation pockets in microchannel and the other was constructed of 512 single-cell isolation pockets. In this study, we demonstrated high-throughput and high-volume single-cell isolation with 512 pocket type device. The total number of isolated single cells in each isolation pocket from the cell suspension at a time was 426 for the cell line of African green monkey kidney, COS-1, and 360 for the rat primary brown preadipocytes, BAT. All isolated cells were stained with fluorescence dye injected into the same microchannel successfully. In addition, the extraction and collection of the cell contents was demonstrated using isolated stained COS-1 cells. The cell contents extracted from each captured cell were individually collected within each collection channel by local hydrodynamic flow. The sequential trapping, labeling, and content extraction with 512 pocket type devices realized high-throughput single-cell manipulations for innovative single-cell handling, feasible staining, and accurate cell rupture. 相似文献
12.
A. Khamenehfar T. V. Beischlag P. J. Russell M. T. P. Ling C. Nelson P. C. H. Li 《Biomicrofluidics》2015,9(6)
Circulating tumor cells (CTCs) are found in the blood of patients with cancer. Although these cells are rare, they can provide useful information for chemotherapy. However, isolation of these rare cells from blood is technically challenging because they are small in numbers. An integrated microfluidic chip, dubbed CTC chip, was designed and fabricated for conducting tumor cell isolation. As CTCs usually show multidrug resistance (MDR), the effect of MDR inhibitors on chemotherapeutic drug accumulation in the isolated single tumor cell is measured. As a model of CTC isolation, human prostate cancer cells were mixed with mouse blood cells and the label-free isolation of the tumor cells was conducted based on cell size difference. The major advantages of the CTC chip are the ability for fast cell isolation, followed by multiple rounds of single-cell measurements, suggesting a potential assay for detecting the drug responses based on the liquid biopsy of cancer patients. 相似文献
13.
Fluid shear stress (FSS) plays a critical role in regulating endothelium function and maintaining vascular homeostasis. Current microfluidic devices for studying FSS effects on cells either separate high shear stress zone and low shear stress zone into different culturing chambers, or arranging the zones serially along the flow direction, which complicates subsequent data interpretation. In this paper, we report a diamond shaped microfluidic shear device where the high shear stress zone and the low shear stress zone are arranged in parallel within one culturing chamber. Since the zones with different shear stress magnitudes are aligned normal to the flow direction, the cells in one stress group are not substantially affected by the flow-induced cytokine/chemokine releases by cells in the other group. Cell loading experiments using human umbilical vein endothelial cells show that the device is able to reveal stress magnitude-dependent and loading duration-dependent cell responses. The co-existence of shear stress zones with varied magnitudes within the same culturing chamber not only ensures that all the cells are subject to the identical culturing conditions, but also allows the resemblance of the differential shear stress pattern in natural arterial conditions. The device is expected to provide a new solution for studying the effects of heterogeneous hemodynamic patterns in the onset and progression of various vascular diseases. 相似文献
14.
近年来,基因组测序、编辑与合成技术日新月异,推动了基因型"设计"和"合成"能力的突飞猛进,同时也使人工细胞的表型检测成为合成生物学发展的瓶颈环节之一。对于细胞功能的快速测试与评价,单细胞分析技术具有重要意义与前景,但理想的解决方案需要具备活体无损、非标记式、提供全景式表型、能分辨复杂功能、快速高通量且低成本、能与组学分析联动等特征。以此为出发点,文章重点介绍了基于非标记式分子光谱学的单细胞功能表征、分选与组学技术体系的进展,并讨论了该领域的关键问题与发展方向。多种光谱技术之间扬长避短的运用与多模态成像,结合高通量的光谱激活细胞分选技术及下游单细胞组学技术,正构建与拓展着一条连接光谱学与遗传学的广阔桥梁。这一桥梁不仅为细胞工厂的高通量、全景式表型检测与筛选提供全新的解决方案,还将推动"单细胞精度的光谱表型组-功能基因组"作为一种新的生物大数据类型,服务于"数据科学"驱动下的合成生物技术。 相似文献
15.
Circulating tumor cells (CTCs) shed from the primary tumor undergo significant fragmentation in the microvasculature, and very few escape to instigate metastases. Inspired by this in vivo behavior of CTCs, we report a microfluidic method to phenotype cancer cells based on their ability to arrest and fragment at a micropillar-based bifurcation. We find that in addition to cancer cell size, mechanical properties determine fragmentability. We observe that highly metastatic prostate cancer cells are more resistant to fragmentation than weakly metastatic cells, providing the first indication that metastatic CTCs can escape rupture and potentially initiate secondary tumors. Our method may thus be useful in identifying phenotypes that succumb to or escape mechanical trauma in microcirculation. 相似文献
16.
Xue Wang Yuetong Wang Zhaoyuan Fang Hua Wang Jian Zhang Longfu Zhang Hsinyi Huang Zhonglin Jiang Yujuan Jin Xiangkun Han Shenda Hou Bin Zhou Feilong Meng Luonan Chen Kwok-Kin Wong Jinfeng Liu Zhiqi Zhang Xin Zhang Haiquan Chen Yihua Sun Liang Hu Hongbin Ji 《国家科学评论(英文版)》2021,8(10)
Somatic mutations of the chromatin remodeling gene ARID2 are observed in ∼7% of human lung adenocarcinomas (LUADs). However, the role of ARID2 in the pathogenesis of LUADs remains largely unknown. Here we find that ARID2 expression is decreased during the malignant progression of both human and mice LUADs. Using two KrasG12D-based genetically engineered murine models, we demonstrate that ARID2 knockout significantly promotes lung cancer malignant progression and shortens overall survival. Consistently, ARID2 knockdown significantly promotes cell proliferation in human and mice lung cancer cells. Through integrative analyses of ChIP-Seq and RNA-Seq data, we find that Hspa1a is up-regulated by Arid2 loss. Knockdown of Hspa1a specifically inhibits malignant progression of Arid2-deficient but not Arid2-wt lung cancers in both cell lines as well as animal models. Treatment with an HSPA1A inhibitor could significantly inhibit the malignant progression of lung cancer with ARID2 deficiency. Together, our findings establish ARID2 as an important tumor suppressor in LUADs with novel mechanistic insights, and further identify HSPA1A as a potential therapeutic target in ARID2-deficient LUADs. 相似文献
17.
18.
The capture and subsequent analysis of rare cells, such as circulating tumor cells from a peripheral blood sample, has the potential to advance our understanding and treatment of a wide range of diseases. There is a particular need for high purity (i.e., high specificity) techniques to isolate these cells, reducing the time and cost required for single-cell genetic analyses by decreasing the number of contaminating cells analyzed. Previous work has shown that antibody-based immunocapture can be combined with dielectrophoresis (DEP) to differentially isolate cancer cells from leukocytes in a characterization device. Here, we build on that work by developing numerical simulations that identify microfluidic obstacle array geometries where DEP–immunocapture can be used to maximize the capture of target rare cells, while minimizing the capture of contaminating cells. We consider geometries with electrodes offset from the array and parallel to the fluid flow, maximizing the magnitude of the resulting electric field at the obstacles'' leading and trailing edges, and minimizing it at the obstacles'' shoulders. This configuration attracts cells with a positive DEP (pDEP) response to the leading edge, where the shear stress is low and residence time is long, resulting in a high capture probability; although these cells are also repelled from the shoulder region, the high local fluid velocity at the shoulder minimizes the impact on the overall transport and capture. Likewise, cells undergoing negative DEP (nDEP) are repelled from regions of high capture probability and attracted to regions where capture is unlikely. These simulations predict that DEP can be used to reduce the probability of capturing contaminating peripheral blood mononuclear cells (using nDEP) from 0.16 to 0.01 while simultaneously increasing the capture of several pancreatic cancer cell lines from 0.03–0.10 to 0.14–0.55, laying the groundwork for the experimental study of hybrid DEP–immunocapture obstacle array microdevices. 相似文献
19.
Jang K Xu Y Tanaka Y Sato K Mawatari K Konno T Ishihara K Kitamori T 《Biomicrofluidics》2010,4(3):32208
Recently, interest in single cell analysis has increased because of its potential for improving our understanding of cellular processes. Single cell operation and attachment is indispensable to realize this task. In this paper, we employed a simple and direct method for single-cell attachment and culture in a closed microchannel. The microchannel surface was modified by applying a nonbiofouling polymer, 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer, and a nitrobenzyl photocleavable linker. Using ultraviolet (UV) light irradiation, the MPC polymer was selectively removed by a photochemical reaction that adjusted the cell adherence inside the microchannel. To obtain the desired single endothelial cell patterning in the microchannel, cell-adhesive regions were controlled by use of round photomasks with diameters of 10, 20, 30, or 50 μm. Single-cell adherence patterns were formed after 12 h of incubation, only when 20 and 30 μm photomasks were used, and the proportions of adherent and nonadherent cells among the entire UV-illuminated areas were 21.3%±0.3% and 7.9%±0.3%, respectively. The frequency of single-cell adherence in the case of the 20 μm photomask was 2.7 times greater than that in the case of the 30 μm photomask. We found that the 20 μm photomask was optimal for the formation of single-cell adherence patterns in the microchannel. This technique can be a powerful tool for analyzing environmental factors like cell-surface and cell-extracellular matrix contact. 相似文献
20.
Despite being invasive within surrounding brain tissues and the central nervous system, little is known about the mechanical properties of brain tumor cells in comparison with benign cells. Here, we present the first measurements of the peak pressure drop due to the passage of benign and cancerous brain cells through confined microchannels in a “microfluidic cell squeezer” device, as well as the elongation, speed, and entry time of the cells in confined channels. We find that cancerous and benign brain cells cannot be differentiated based on speeds or elongation. We have found that the entry time into a narrow constriction is a more sensitive indicator of the differences between malignant and healthy glial cells than pressure drops. Importantly, we also find that brain tumor cells take a longer time to squeeze through a constriction and migrate more slowly than benign cells in two dimensional wound healing assays. Based on these observations, we arrive at the surprising conclusion that the prevailing notion of extraneural cancer cells being more mechanically compliant than benign cells may not apply to brain cancer cells. 相似文献