To build a microfluidic device with various morphological features of the tumor vasculature for study of the effects of tumor vascular structures on the flow field and tumor cellular flow behaviors. The designed microfluidic device was able to approximatively simulate the in vivo structures of tumor vessels and the flow within it. In this models, the influences of the angle of bifurcation, the number of branches, and the narrow channels on the flow field and the influence of vorticity on the retention of HepG2 cells were significant. Additionally, shear stress below physiological conditions of blood circulation has considerable effect on the formation of the lumen-like structures (LLSs) of HepG2 cells. These results can provide some data and reference in the understanding of the interaction between hemorheological properties and tumor vascular structures in solid tumors.

OBJECTIVES:To build a three-dimensional co-culture model in a microfluidic device for cancer research and evaluate its feasibility by investigating cancer stem-like cells (SCs) induced migration of human umbilical vein endothelial cells (ECs).RESULTS:The microfluidic device provided two-dimensional and three-dimensional (2D/3D) culture and co-culture environments without affecting cell viability. The device also provided an effective concentration for the chemiotaxis of cells, and to support real-time monitoring of cell behavior. In this model, SCs significantly increased the migration area of ECs with a hepatocarcinoma cell line (MHCC97H; MCs). The presence of ECs also induced both MCs and SCs invasion into Matrigel. The migration area of MCs and SCs significantly increased when co-cultured with ECs.CONCLUSIONS:This 3D co-culture microfluidic model is a suitable model in cancer research. Compared with MCs, SCs had greater potential in inducing EC migration and interacting with ECs.

Cardiovascular diseases (CVDs) are considered the major cause of death worldwide, so more researchers pay more and more attention to the development of a non-invasive method to obtain as much cardiovascular information (CVI) as possible for early screening and diagnosing. It is known that considerable brain information could be probed by a variety of stimuli (such as video, light, and sound). Therefore, it is quite possible that much more CVI could be extracted via giving the human body some special interrelated stimulus. Based on this hypothesis, we designed a novel signal platform to acquire more CVI with a special stimulus, which is to give a gradual decrease and a different settable constant pressure to six air belts placed on two-side brachia, wrists, and ankles, respectively. During the stimulating process, the platform is able to collect 24-channel dynamic signals related with CVI synchronously. Moreover, to improve the measurement accuracy of signal acquisition, a high precision reference chip and a software correction are adopted in this platform. Additionally, we have also shown some collection instances and analysis results in this paper for its reliability. The results suggest that our platform can not only be applied on study in a deep-going way of relationship between collected signals and CVDs but can also serve as the basic tool for developing a new noninvasive cardiovascular function detection instrument and system that can be used both at home and in the hospital.

Microfluidic technology is an important research tool for investigating angiogenesis in vitro. Here, we fabricated a polydimethylsiloxane (PDMS) microfluidic device with five cross-shaped chambers using a coverslip molding method. Then, the perforated PDMS microhole arrays prepared by soft lithography were assembled in the device as barriers; a single microhole had a diameter of 100 μm. After injecting type I collagen into the middle gel chamber, we added a culture medium containing a vascular endothelial growth factor (VEGF) into the middle chamber. It would generate a linear concentration gradient of VEGF across the gel region from the middle chamber to the four peripheral chambers. Human umbilical vein endothelial cells (HUVECs) were then seeded on the microhole barrier. With VEGF stimulation, cells migrated along the inner walls of the microholes, formed annularly distributed cell clusters at the gel-barrier interface, and then three-dimensionally (3D) sprouted into the collagen scaffold. After 4 days of culture, we quantitatively analyzed the sprouting morphogenesis. HUVECs cultured on the microhole barrier had longer sprouts than HUVECs cultured without the barrier (controls). Furthermore, the initial distribution of sprouts was more regular and more connections of tube-like structures were generated when the microhole barrier was used. This study introduces a novel microfluidic device containing both microtopographic structures and 3D collagen. HUVECs cultured with the microhole barrier could form well-interconnected tube-like structures and are thus an ideal in vitro angiogenesis model.

Nerve growth factor (NGF)/nerve growth factor receptors (NGFRs) axis and canonical WNT/β-catenin pathway have shown to play crucial roles in tumor initiation, progression and prognosis. But little did we know the relationship between them in modulation of tumor progress. In this report, we found that NGF/NGFRs and β-catenin were coexpression in ovarian cancer cell lines, and NGF can decrease the expression level of β-catenin and affect its activities, which may be related to the NGF-induced down-regulation of B-cell CLL/lymphoma 9-like (BCL9L, BCL9-2). Furthermore, NGF can also increase or decrease the downstream target gene expression levels of WNT/β-catenin depending on the cell types. Especially, we created a novel in vitro cell growth model based on a microfluidic device to intuitively observe the effects of NGF/NGFRs on the motility behaviors of ovarian cancer cells. The results showed that the migration area and maximum distance into three dimensional (3D) matrigel were decreased in CAOV3 and OVCAR3 cells, but increased in SKOV3 cells following the stimulation with NGF. In addition, we found that the cell colony area was down-regulated in CAOV3 cells, however, it was augmented in OVCAR3 cells after treatment with NGF. The inhibitors of NGF/NGFRs, such as Ro 08-2750, K252a and LM11A-31,can all block NGF-stimulated changes of gene expression or migratory behavior on ovarian cancer cells. The different results among ovarian cancer cells illustrated the heterogeneity and complexity of ovarian cancer. Collectively, our results suggested for the first time that NGF is functionally linked to β-catenin in the migration of human ovarian cancer cells, which may be a novel therapeutic perspective to prevent the spread of ovarian carcinomas by studying the interaction between NGF/NGFRs and canonical WNT/β-catenin signaling.

We present a new economical microfluidic viscometer to measure the viscosity of biological fluids, using sample volumes of less than 200 μl. It is fabricated using a microwire-molding technique, making it easier and cheaper to produce than existing viscometers. The viscometer is based on laminar flow inside a polydimethylsiloxane microchip. The velocity of the sample flow inside the capillary was monitored with a camera, and the movement of the liquid column was determined by a Matlab video-processing program. The device was calibrated using deionized water, which is a Newtonian fluid, at 20 °C. The viscometer provides accurate measurements of viscosity for values as small as 0.69 mPa s. The viscosity of water at different temperatures was measured, showing more than 98% agreement with the values provided by the National Institute of Standards and Technology. Various samples including a series of glycerol solutions, phosphate-buffered saline, alcohol, and cell media were also tested, and the measured viscosities were compared with those from a traditional glass capillary viscometer. The results show good agreement between the two methods, with an average relative error of less than 1%. Furthermore, the viscosities of several cell suspensions were measured, showing a relative standard deviation of less than 1.5%. The microchip viscometer is economical and is shown to be accurate, which is very important for the simulation and control of lab-on-a-chip experiments.

在肿瘤及相伴血管生长过程中,微环境中的多种理化因素协同地发挥着重要的作用.传统体外实验多借助于Transwell等模型,在单一因素下考察细胞生物学效应,并不能反映在体的多因素微环境.基于微流控技术,本文构建了一种新的多细胞共培养模型,整合了多环境维度(二维/三维)、细胞与细胞及细胞与胞外基质相互作用、不同生化因子的浓度梯度、细胞区域性等多个重要因素,形成微环境,并能实时监测细胞的迁移和侵袭等响应.为评价该模型的可行性和功能上的独特优势,我们模拟了肿瘤细胞(HepG2,CAOV-3)和人脐静脉内皮细胞(HUVECs)共存的三维微环境,考察了它们共培养时相互诱导向三维基质材料中的迁移情况.结果表明,在三维共培养模型中细胞能够相互影响并出现明显形态差异;2种肿瘤细胞的诱导均使HUVECs迁移能力显著提高;同时2种不同肿瘤细胞出现了与其病理特质(HepG2低浸润,CAOV-3高浸润)相对应的迁移能力差异.以上结果表明,该模型可望为研究肿瘤微环境下的相关问题提供一个相对简便且更具整合价值的研究平台.

Stromal cell-derived factor 1 (SDF-1) is a critical regulator of endothelial progenitor cells (EPCs) mediated physiological and pathologic angiogenesis. It was considered to act via its unique receptor CXCR4 for a long time. CXCR7 is a second, recently identified receptor for SDF-1, and its role in human EPCs is unclear. In present study, CXCR7 was found to be scarcely expressed on the surface of human EPCs derived from cord blood, but considerable intracellular CXCR7 was detected, which differs from that on EPCs derived from rat bone marrow. CXCR7 failed to support SDF-1 induced human EPCs migration, proliferation, or nitric oxide (NO) production, but mediated human EPCs survival exclusively. Besides that, CXCR7 mediated EPCs tube formation along with CXCR4. Blocking CXCR7 with its antagonist CCX733 impaired SDF-1/CXCR4 induced EPCs adhesion to active HUVECs and trans-endothelial migration. Those results suggested that CXCR7 plays an important role in human cord blood derived EPCs in response to SDF-1.

抗癌药物在进行动物和临床试验以前,需要用体外肿瘤组织模型评估药效.由于三维(3D)多细胞球体(multicellular tumor spheroids MCTSs)在抗药性和组织结构等方面与体内肿瘤组织相似,常被用作体外肿瘤组织模型.为监测MCTSs在形成过程中,肿瘤细胞之间和肿瘤细胞与基质之间的相互作用,基于微流控技术基础上自行设计和构建MCTSs模型.该肿瘤MCTSs模型实验结果表明,在3D微环境下,血清能够诱导MDA-MB-231形成直径为289 μm的MCTSs,肿瘤细胞MCTSs之间有相互靠近的趋势,并且发现凋亡细胞多分布在MCTSs之间.肿瘤坏死因子(tumor necrosis factor.-α,TNF-α)诱导MDA-MB-231形成MCTSs之间没有相互靠近的趋势,并且MCTSs直径的长度很难达到100 μm.以上结果表明,该模型有望为研究肿瘤形成MCTSs机制和药物筛选提供有用的体外肿瘤模型.

Stromal cell-derived factor 1(SDF-1) is a principal regulator of tumor invasion and metastasis.SDF-1 had been considered to mediate biological process through its unique receptor CXC chemokine receptor 4(CXCR4) for many years.Recent studies reported that SDF-1 was also a ligand of a novel chemokine receptor-CXC chemokine receptor 7(CXCR7).It was confirmed that CXCR7 plays an important role in invasion and metastasis of several types of tumor induced by SDF-1.However,its role in cervical cancer is still unclear.In the present study,the expressions of CXCR4 and CXCR7 on HeLa cells were detected by Western blotting and the effects of CXCR4 and CXCR7 on cell behaviors were tested by blocking with their antagonists,respectively.Cell proliferation,cell invasion and cell adhesion were evaluated by MTT,Transwell assay,adhesion assay,respectively.Results reveal that both CXCR4 and CXCR7 are expressed on HeLa cells.The proliferation,invasion and adhesion of HeLa cells induced by SDF-1 are inhibited by CXCR4 blockage or CXCR7 blockage.These results suggest that CXCR7 can mediate proliferation,invasion and adhesion of HeLa cells induced by SDF-1,which indicates that CXCR7 is a potential target for cervical cancer therapy.

Objective: To develop a microfluidic device with the adjustable concentration and pressure gradient for 3D cell culture in hydrogel and set up an in vitro model with the capability to closely simulate in vivo microenvironment for cell growth. Methods: The microfluidic chip, with a middle channel for 3D cell culture and two-side channels for delivering cell culture medium, was designed and fabricated using standard soft lithography and replica molding techniques. Its capability to generate concentration gradient, interstitial flow and image cell in situ was demonstrated. Results: A simple microfluidic chip for 3D cell culture in hydrogel with the capability to generate the concentration and pressure gradient was obtained. At a flow rate of 2 μL·min -1 in each side channel, the concentration gradients remained constant after 3 h. The interstitial flow across the gel scaffold was generated by a 100 Pa pressure difference between two-side channels with the pressure gradient of 0.11 Pa/μm. Human adult dermal microvascular endothelial cells (HMVEC) were maintained in 3D culture with collagen type I and observed with confocal microscopy. Conclusions: The microfluidic chip is simple and easy to operate and it can simulate the complicated microenvironment in vivo. The chip also allows the multiparameter control of microenvironment, facilitating the better understanding of interaction between cells and microenvironment.

In this research project, rats were made into animal models of acute focal cerebral ischemia and reperfusion (IR) by occlusion of their middle cerebral artery (MCAO). We observed the effect of endogenous endothelial progenitor cells (EPCs) and serum cytokines on cerebral ischemia rats treated by electro-acupuncture(EA). The results showed: MCAO model had high stability after EA treatment which was delivered via the acupuncture needles inserted into "quchi" and "zusanli" points, the nervous functions of cerebral IR rats recovered faster than those of rats not treated; EPCs in rats' blood increased after acute focal cerebral ischemia and reperfusion; and the growth rate was obvious in IR group. This phenomenon might be related to the inflammation elicited by injury of ischemia and self-repair. Besides, EA treatment could decrease induced nitric oxide synthase (iNOS) activity, alleviate injury after cerebral ischemia, and regulate the quantity of EPCs in blood. The quantity of EPCs in blood increased in IR-24hr. In IR-48 hr, the rise of EPCs quantity was significant (P < 0.01). The level of vascular endothelium growth factor (VEGF) in serum of rats after cerebral ischemia was escalated, which indicated to a certain extent that cerebral ischemia could stimulate stress reaction. EA treatment could raise VEGF level, which suggested that high expression of VEGF could accelerate mobilization, chemotaxis and homing of EPCs. At the same time, the levels of matrix metalloproteinase-9 (MMP-9) and basic fibroblast growth factor (bFGF) also changed. In conclusion, EA treatment could promote neovascularization after cerebral ischemia by mobilizing EPCs, decreasing iNOS activity and increasing VEGF level. This may be one of the ways by which EA could treat cerebral ischemia.

Objective:To observe the effects of electro-acupuncture on the quantity of endothelial progenitor cells(EPCs) in peripheral blood and on the expression of Vessel Endothelium Growth Factor Receptor-2(VEGFR-2) and Platelet Endothelial Cell Adhesion Molecule-1(PECAM-1) in ischemic cerebral cortex of cerebral ischemia-reperfusion rats,discussing the mechanism of angiogenesis after cerebral ischemia with the effect of electro-acupuncture.Methods:Male rats were randomly and evenly assigned to normal group,sham-operation group,model group and EA group.quchi and housanli were selected.EPCs quantity was displayed using flow cytometry.VEGFR-2 and PECAM-1 expression are observed with immunohistochemistry.Results:EPCs quantity in blood achieved peak in model group at 24hrs,and began to decreased after 48hrs;while in EA group EPCs quantity in blood started to increase at 24hrs,and reached peak in 48hrs,then decreased at 72hrs.VEGFR-2 and PECAM-1 expression began to increase at 24hrs in model group and EA group,and the expression increased with the time prolonged.The expression of electro-acupuncture group was more than that in model group.Conclusion:The electro-acupuncture could increase EPCs quantity in peripheral blood,and raise VEGFR-2 and PECAM-1 expression in ischemic cerebral cortex.Those can contribute to promote the formation of blood vessels in cerebral ischemia region.

OBJECTIVE:To observe the effect of electroacupuncture (EA) on peripheral blood endothelial progenitor cell (EPC) counts, vascular endothelial growth factor (VEGF) level and total nitric oxide synthase (TNOS) and inducible nitric oxide synthase (iNOS) activity in cerebral ischemia-reperfusion injury (CI/RI) rats. METHODS:A total of 72 male rats were randomly and evenly assigned to normal control, sham-operation (sham), model and EA groups which were further divided into 24 h, 48 h and 72 h subgroups, with 6 cases in each. Acute focal cerebral ischemia model was established by occlusion of the middle cerebral artery (MCAO, 120 min) and reperfusion. EA (2/15 Hz, 1 mA) was applied to "Quchi" (LI 11) and "Zusanli" (ST 36) for 30 min, once daily. Peripheral blood was collected from abdominal aorta for detecting EPC count by using flow cytometry, serum VEGF level by using enzyme-linked immunosorbent assay (ELISA), and serum TNOS and iNOS activity by spectrophotometry, respectively. RESULTS:Compared with the corresponding normal control subgroups, blood EPC counts, serum TNOS and INOS activity and serum VEGF content at 24 h, INOS activity and VEGF level at 48 h, and EPCs and INOS at 72 h in model subgroups all increased significantly (P < 0.01, P < 0.05). In comparison with the corresponding model subgroups, EPC count at 24 h and 72 h, and TNOS activity at 24 h in EA subgroups decreased considerably (P < 0.01, P < 0.05); while EPC and VEGF levels at 48 h in EA subgroup increased evidently (P < 0.05, P < 0.01). No significant differences were found among normal, sham, model and EA subgroups in serum TNOS activity at 48 h and 72 h (P > 0.05). CONCLUSION:EA of LI 11 and ST 36 can suppress CI/ RI induced increase of blood EPC count and serum TNOS activity, and upregulate serum VEGF level, which may contribute to its effect in relieving CI/RI.