摘要
通过各种力学装置对体外培养的活细胞施加机械力,是目前细胞力学研究的主要途径。在力学刺激下活细胞将发生多种适应性变化,不同的力学环境其变化的程度不尽相同。细胞形态学上的变化是最直观的变化,它预示并隐含着细胞功能上的变化。运用图象分析系统,定量研究细胞和亚细胞结构的运动和形态的动态变化特性,有助于理解力学刺激和细胞生长的关系,也有助于理解细胞病变的机理。但是,细胞力学实验中针对的是不能染色的生物活细胞图象,图象因细胞种类而异,并且图象性质随实验条件的变化而变化。迄今为止尚没有一个图象分析系统适用于所有的图象分析。鉴于此,本文针对交变应力环境下的细胞剪切力实验和细胞膜式张应力实验,开展了细胞力学实验图象处理平台和方法的研究。
针对剪切力环境下内皮细胞的形态变化,本文提出了两个反映细胞形态变化速率的指标,拉伸长度时间变化率和方向角时间变化率,可以综合表征细胞形态学变化的快慢。
图象分割是图象处理过程中最为困难也是最为关键的环节。本文深入分析了细胞力学实验中细胞图象的特点,针对细胞无交叠的图象,提出了基于微分边缘检测算子的图象分割技术路线,共有8个步骤。依次为图象增强、灰度化、消除光晕、Canny边缘检测、数学形态学膨胀、区域填充、平滑边缘、形态学重建。针对交叠现象严重的细胞图象,提出了基于watershed的的图象分割技术路线,共有7个步骤。依次为图象增强、灰度化、形态学光源校正分离细胞与背
景、形态学重建去噪、距离转换、提取marke。图、water、hed转换,初步实现
了对交叠细胞的分割。
本文使用ol帅pus IX70倒置相差显微镜观察细胞图象,采用两种采集方式
采集细胞数字图象,建立了细胞力学实验的图象获取与处理硬件平台。一种采
集方式是采用JvC一C68O一EC模拟CCO配合V工以g000V图象采集卡,另一种方式
采用Pixera 1 50ES专业数字显微CCD。并且建立了显微镜照明自动控制系统,可
以按照捕获设定,定时接通显微镜照明电源,减少显微镜照明灯泡的损耗。实
现了细胞图象采集的程控化,使用者可以根据实验需要设定捕获方案,如单帧
捕获、多帧捕获和视频图象流捕获,便于实现无人职守。
本文使用VisualC++ 6.0开发环境,建立了细胞图象处理的软件平台。
它是一个集成化的软件包,将图像采集、文件管理、图像处理、特征提取、参
数测量以及结果输出等多项功能有机地组织在一个集成环境中。由于C++集成了
面向对象技术,该平台功能模块很容易实现功能扩充和代码移植。
运用建立的图象处理硬件平台,采集多个实验的细胞图象。结果表明,模
拟CCD方案虽然分辨率不高,但是在采集细胞的视频图象方面具有优势:专用
数字显微CCD方案分辨率高,光灵敏度高,图象采集数据损耗小,比较适合细
胞静态图象采集。分别采用两种分割技术路线分割细胞图象,得到的细胞轮廓
线与细胞实际边界比较吻合。运用开发的图象处理系统,开展了大鼠主动脉血
管内皮细胞在病理脉动大剪切力环境下的形态学研究,实验结果表明,图象处
理系统可以胜任细胞的形态学参数的定量分析。
Because of the staggering complexity of the in vivo environment, systematic study of the cellular response to mechanical stimulations has relied heavily on the use of in vitro preparations. Living cells under mechanical stimulations can make adaptive changes and it must be recognized that different biomechanical stimulations can induce different cellular changes. Morphological change is the most instinctive one and it can also be an indicator for the functional changes in cells. The quantitative study on cellular and sub-cellular morphology and movement by image analyzing system is useful not only in understanding the relations between force simulation and cellular growth, but also in catching on cellular pathological mechanism. However, few image analysis systems are found to suitable for all kinds of images, since images, especially the biological living cellular images, which are sourced by different methods have different characteristics. According to this condition, a new image-processing platform to analyze the cellular changes induced by shear stress and tensile stress has been set up in our lab and the image-processing methods for cellular biomechanical experiments have been detailed studied in this dissertation.
The morphological parameters of endothelial cells exposed on shear stress are analyzed in detail and then two novel parameters are suggested. They are named VOA (velocity of oriental angle) and VL (velocity of major axis length) and can reflect velocity in cellular morphological changes.
Image segmentation plays a key role in image analysis and it is also the most hardly step in image processing pipeline. As for all the non-overlapping images, one method which has eight steps based on difference arithmetic operators is proposed. Image enhancing, image graying, halation removing, Canny edge detection, mathematical morphology dilation, edge smoothing, mathematical morphological reconstruction are implemented according to the set sequence. As for all the seriously
overlapping images, another method which has seven steps based on watershed transform is put forward. These steps in turn are image enhancing, image graying, morphological illumination, mathematical morphological reconstruction, distance transform, marker image extraction, watershed transform.
Olympus 1X70 phase contrast inverted microscope is used to observe cellular images and two methods are applied to capture digital images in the construction of image processing hardware. One is through video CCD (JVOC680-EC) equipped with image capture board (VIGA 9000V), the other is through professional digital CCD for microscopy (Pixera 150ES). A system for automatic control illumination of microscope is set up, which can put through the illumination power of microscope according to the set capture schedule and will help decrease the loss of the microscope's bulb. And the images can be captured in a manner of single frame, frame-by-frame or video stream.
The cellular mechanical experimental image-processing platform is developed by using Visual C++ 6.0 (Microsoft) programming language. It is an integrated software package, which integrates image acquiring, files managing, image processing, feature extracting, parameter measuring and result outputting together. Because Oriental Object Programming (OOP) skills are applied to this system, the modules in this package are easy to accomplish functional expanding and code implanting.
Enormous cellular images are captured through this image-processing platform and the results suggest that video CCD schedule has advantages in capturing video stream with the disadvantages in the image resolution, and digital CCD is very useful in capturing still cellular image because of its relatively high resolution, high sensitivity and less data loss. The superposition of the processed image by either segmentation strategy on the original cell image indicates that the two methods are suitable. By using developed image analysis system, the morphological parameters of rat aortic endothelial cells (RAECs) expo
引文
1 陶祖莱,孟庆国,关于我国生物力学发展的几点意见。力学进展,2000,30(3) : 472-5
2 Editorial. Cell Mechanics. Journal of Biomechanies. 2000;33:1-2
3 Kataoka N, Ujita S, Sato M. Effect of flow direction on the morphological responses of cul tured bovine aortic endothelial cells. Medical & Biological Engineering & Computing. 1998:36(1) :122-8.
4 Helmlinger G, Geiger RV, Schreck S, Nerem RM. Effects of pulsatile flow on cultured vascular endothelial cell morphology. Journal of Biomechanical Engineering. 1991;113(2) :123-31.
5 Chiu JJ, Wang DL, Chien S, et al. Effects of disturbed flow on endothelial cells. Journal of Biomechanical Engineering. 1998:120(1) :2-8.
6 Kapur R, Rudolph AS. Cellular and cytoskeleton morphology and strength of adhesion of cells on self-assembled monolayers of organosilanes. Experimental Cell Research. 1998;244(1) :275-85.
7 Tardy Y, Resnick N, Nagel T, Gimbrone MA Jr, Dewey CF Jr. Shear stress gradients remodel endothelial monolayers in vitro via a cell proliferation-migration-loss cycle. Arterioscler Thromb Vasc Biol. 1997;17(11) :3102-6.
8 DePaola N, Gimbrone MA Jr, Davies PF, Dewey CF Jr. Vascular endothelium responds to fluid shear stress gradients. Arterioscler Thromb. 1992; 12(11) : 1254-7.
9 Zhao S, SuciuA, ZieglerT, et al. Synergistic effects of fluid shear stress and cyclic circumferential stretch on vascular endothelial cell morphology and cytoskeleton. Arterioscler Thromb Vasc Biol. 1995; 15(10) : 1781-6.
10 Divies PF, Dewey CF Jr.Bussplari SR,et al. Influence of hemodynamicforce on vascular endothelial function: In vitro studies of shear stress and pinocytosis in bovine aortic cells. J. Clin. Invest. 1984; 73: 112-19
11 Sprague EA, AreinbachBL, Nerem RM. Influence of a laminar steady state fluid-imposed wall shear stress on the binding, internalization, and degradation of low density lipoproteins by cultured arterial endothelium. Lab. Invest. 1987;76:648-56
12 Grabowski EF, JaffeEA, Weksler BB. Prostacyclin production by cultured endothelial cell monolayers exposed to step increases in shear stress. J. lab. Clin Med. 1985:105:36-43
13 Geiger RV, Berk BC, Alexander EW, et al. Flow-induced calcium transients in single endothelial cells: spatial and temporal analysis. Am. J. Physiol, 1992:262(Cell Physiol. 31) :C14111-7
14 Evans EA, Skalak R. Mechanical and Thermodynamics of Biomembrane. CRC press, Boca Taton, Fla. , 1990
15 Fung YC, Liu SQ, Elementary mechanics of endothelium of blood vessels. ASME of J. Biomech. Engng. 1993;115:1-12
16 Sato M, Levesque MJ, Nerem RM, An application of the micropipet technique to the measurementof the mechanical properties of cultured bovine aortic endothelial cells. ASME J. Biomech. Engng.,109(1987) :27-34
17 Dong C, Skalak R, Sung K-L P, et al, Passive deformation analysis of human leukocytes. ASME J. Biomech. Engng., 1988:110:27-36
18 冯元桢,生物力学,科学出版社。北京,1983
19 Cheng LY, Deformation analysis in cell and developing biology. Part Ⅰ:Formal methodology. ASME J. Biomech. Engng., 1987:109:10
20 Cheng LY, Deformation analysis in cell and developing biology. Part Ⅱ :Mechanical experiments on cells. ASME J. Biomech. Engng., 1987:109:18
21 Dong CR, Skalak R., Sung K-L P, Cytoplasmic rheology of passive neutrophis. Biorheology, 1991:28:557-567
22 Schmid-Schonbein GW, Skalak R, Continuum mechanical model leukocytes during protoped formation. ASME J. Biomech. Engng., 1984;106:10-18
23 Oster GF, Perelson AS, Cell Spreading and molitity: a model lamellipod. J.
Math. Biol. 1985;21:383-388
24 Skalak R, Zhu C, Thermodynamics and mechanics of cell activation. Bioraechanics of active raovement and deformation of cells. Akkas, N(ed.)NATO AST series H42,Springer-verlag, 1990:155-183
25 Zhu C, Skalak R, A continuum of modei of protrusion of pseudopod. Biophys. J. 1988:54:1115-1137
26 Zhu C, Skalak R, Schmid-Schonbein GW, One-dimensional steady continuum model of retraction of pseudopod in leukocytes. ASME J. Biomech. Engng. , 1989:112:69-77
27 Simon SI, Schmid-Schonbein GW, Kinematics of cytoplasmic deformation in neutrophils during active motions. ASME J. Biomech. Engng., 11990;12:283
28 Zhu C, Bao G, Wang N. Cell Mechanics: Mechanical response, cell adhesion, and molecular deformation. Annu. Rev. Biomed. Eng. 2000:2:189-226
29 Kaplanski G, Farnarier C, Tissot O, et al. Granulocyte-endothelium initial adhesion. Analysis of transient binding events mediated by E-selectin in a laminar shear flow. Biophys. J. 1993:64:1922-33
30 Alon R, Hammer DA, Springer TA, et al. Lifetime of the P-selectin-carbohydrate bond and its response to tensile force in hydrodynamic flow. Nature. 1995:374:539-42
31 Cozens-Roberts C, Lauffenburger DA, Quinn JA. Receptor-mediated cell attachment and deattchment kinetics.I. Probalistic model and analysis. Biophys. J. 1990:58:841-56
32 Tomczok J, Sliwa-Tomczok W, Klein CL, van Kooten TG, Kirkpatrick CJ. Biomaterial-induced alterations of human neutrophils under fluid shear stress: scanning electron microscopical study in vitro. Biomaterials. 1996;17(14) :1359-67.
33 Sirois E, Charara J, Ruel J, Dussault JC, Gagnon P, Doillon CJ. Endothelial cells exposed to erythrocytes under shear stress: an in vitro study. Biomaterials. 1998;19(21) :1925-34.
34 Davies PF. Flow-mediated endothelial mechanotransduction. Physiol. Rev. 1995:75:519-60
35 Davies PF, Barbee KA, Volin MV, et al. Spatial Relationships in early signaling events of flow-mediated endothelial mechamotransduction. Annu. Rev. Physiol. 1997:59:527-49
36 Stamenovic D, Wang N. Cellular response to mechanical stress. Invited Review: Engineering approaches to cytoskeletal mechanics. J. Appl. Physiol. 2000;89:2085-90
37 Fisher AB, Chien S, Barakat AI, et al. Endothelial celluar response to altered shear stress. Am J. Physiol Lung Cell Mol. Physiol. 2001;281:L529-33
38 Liu MY, Post M. Cellular Responses to Mechanical Stress. Invited Review: Mechanochemical signal transduction in the fetal lung. J Appl Physiol. 2000: 89: 2078-2084
39 Jen CJ, Jhiang SJ, Chen HI. Cellular rsponse to mechanical stress. Invited review: Efects of flow on vascular endothelial intracellular calcium signaling of rat aortas ex vivo. J. Appl. Physiol. 2000:89:1657-62
40 Gluchsmann A. . Studies on bone mechanics in vitro: Ⅱ, The role of tension and pressure in chondrogenesis. Anatomical Record. 1939:73:39-56.
41 白灵,樊瑜波,张明等,离体培养细胞的力学实验方法,生物医学工程学杂志, 19(2) :324-328,2002。
42 Mitchison JM, Swann MM. The mechanical properties of the red cell surface: Ⅰ. The cell elastimeter. J Exp Biol 1954:31:443-460.
43 Usami S, Wung S-L, et al. . Locomotion forces generated by a polymorphonnclear leukocyte. Biophys J. 1992:63:1663-1666,.
44 Miyazaki H, Hayashi K. Tensile Property Of Fibroblasts From The Rabbit Patellar Tendon. BIO'99: Cell and Tissue Engineering Ⅰ, Ⅱ, Ⅲ, Ⅳ, VBIO'99.
45 Lakshni S, Eidsath CA, Saxinger WC. A simple method for measurement of cell-substrate attachment force: application to HIV-1 Tat. Journal of Cell
Science. 1997:110:249-256.
46 Tokunaga O, Watanable T. Properties of endothelial cells and smooth muscle cell cultured in ambient pressure. In Vitro Cell Dev Biol. 1987;23:52-534.
47 Hishikawa k, Nakaki T, Marumo T, et al. Pressure promotes DNA synthesis in rat cultured vascular smooth muscle cells. J Clin Invest, 1994:93: 1975-80.
48 Imamura K, Ozawa H, Hiriade T, et al. Continuously applied compressive pressure induces bone resorption by a mechanism involving proslaglandin E2 synthesis. Journal of cell Physiology. 1990:144:222-228.
49 BrightonCT, FisherJRS, Levine SE, et al. The biochemical pathway mediating the proliferative response of bone cells to a mechanical stimulus. J of Bone and Joint Surg. 1996; 78(9) : 1337-47
50 Acevedo AD, Bowser SS, Gerritsen ME, et al. Morphological and prol iferative responses of endothelial cells to hydrostatic pressure:role of fibroblast growth factor. J Cell Physiol. 1993; 157: 603-614.
51 Parkkinen JJ, Lammi MJ, Inkinen R, et al. Influence of short-term hydrostatic pressure on organization of stress fibers in cultured chondrocytes. J of Orthop Res. 1995:13:495-502.
52 San RLY, Kim Young-Jo, Doong YH, et al. Biosynthetic response of cartilage explants to dynamic compression. J of Orthop Res. 1989; 7(5) : 619-636.
53 Buschmann MD, Gluzband YA, Grodzinskt AJ, et al. Mechanical compression modulates matrix biosynthesis in chondrocyte/agarose cultur. J cell Sci. 1995; 108: 1497-1508.
54 Buschmann MD, Gluzband YA., Grodzinskt A. J. , et ai. . Chondrocytes in agarose culture synthesize a mechanically functional extracellular matrix. Journal of Orthopaedic research 10, 745-758,1992.
55 Sah RL, Frippel SB, Grodzinsky AJ. Differential effects of serum, Insulin-like growth factor-I, and fibroblast growth factor-2 on the maintenance of cartilage physical properties during long-term culture. J of Orthop Res. 1996:14:44-52.
56 Burton-Wurster N, Vernier-Singer M, Farquhar T, et al. Effect of compressive loading and unloading on the synthesis of total protein, proteoglycan and fibronectin by canine cartilage explants. J. Orthop Res. 1993:11:717-729.
57 Torzilli PA, Grigiene R, Huang C, et al. Characterization of cartilage metabolic response to static and dynamic stress using a mechanical explant test system. J Biomech. 1997:30:1-9.
58 Tanaka S.M.. A new mechanical stimulator for cultured bone cells using piezoelectric actuator. J. Biomech. 32: 427-430,1999.
59 Mauck RL, Solta MA, Wang CC, et al. Functional tissue engineering of articular cartilage through dynamic loading of chondrocyte-seeded agarose gels. Transactions of the ASME, 2000;22:252-60.
60 Leung DYM, Glagov S, Mathews MBA. New in vitro system for studying cell response to mechanical stimulation-different effects of cyclic stretching and agitation on smooth muscle cell biosynthesis. Experimental cell research 1977; 109: 285-298.
61 Ives CL, Eskin SG, McIntire LV. Mechanical effects on endothelial cell morphology in vitro assessment. In Vitro Cellular and Development Biology. 1986; 22: 500-507.
62 Carosi JA, Eskin SG, Mclntire IV. Cyclic strain effects on the production of vasoactive materials in endothelial cells. J cell Physiol. 1992; 15: 29-36,
63 Neidlinger-Wilke C, Wilke Hans-Joachim and Clues Lutz. Cyclic stretching of human osteoblasts affects proliferation and metabolism:a new experimental method and its application. J. of Orthop. Res. 12: 70-78,1994.
64 Owan I, Burr DB, Turner CH, et al. Mechanotransduction in bone: osteoblasts are more responsive to fluid forces than mechanical strain. American Journal of Physiology. 1997;273;C810-815.
65 Bottlang M., Simnacher M., Schmidt H., et al. . A cell strain system for small homogeneous strain applications. Biomedical Technik. 1997; 42: 305-309.
66 Winston FK, Macarak EJ, Gorfien F, et al. A system to reproduce and quantify the biomechanival environment of the cell. J Appl Physiol. 1989; 67: 397-405.
67 Gorfien SF, Winston FK, Thibault LE, et al. Effects of biaxial deformation on pulmonary artery endothelial celis. J cell Physiol. 1989; 139:492-500.
68 Belloli DM, Williams JL, Park JY. Calculated strain fields on cell stressing devices employing circular diaphragms as substrates. Trans Orthop Res Soc. 1991; 166:399.
69 Watson PA. Function follows form:generation of intraceller signals by cell deformation. FASEB J. 1991; 5:2013-9.
70 Williams JL, Chen JH, Belloli DM. Strain field on cell stressing devices employing damped circular elastic diaphragms as substrates. J Biomech Engng. 1992; 114: 377-384.
71 Brighton CT, Strafford B, Gross SB, et al. The proliferative and synthetic response of isolated calvarial bone cells of rats to cyclic biaxial mechanical strain. J Bone Joint Surg (AM). 1991; 73: 320-331.
72 Hasegawa S, Sato S, Saito S, et al. Mechanical stretchingd increases the number of cultured bone celis synthesizing DNA and alters their pattern of protein synthesis. Calcif Tissue Int. 1985; 37: 431-436.
73 Andersen KL, Norton LA. A device for the application of known simulated of thodontic forces to human celis in vitro. J Biomech. 1991; 24(7) : 649-654,
74 Hung CT, Williams JL. A method for inducing equi-biaxial and uniform strains in elastomeric membranes used as cell substrates. J Biomech. 1994; 27: 227-232.
75 Schaffer JL, Rizen M, L' Italien GJ, et al. Device for the application of a dynamically biaxially uniform and isotropic strain to a flexible cell culture membrane. J Orthop Res. 1994; 12: 709-719,.
76 Soma S, Matsumoto S, Takano-Yamaoto T. Enhancement by conditioned medium of stretched calvarial bone cells of the ostoclast-like cell formation
induced by parathyroid hormone in mouse bone marrow cultures. Arch Oral Biol. 1997; 42(3) : 205-211.
77 Vandenburgh HH. A computerized mechanical cell stimulator for tissue culture: effects on skeletal muscle organogenesis. In Vitro Cell Dev Biol. 1988; 24: 609-619.
78 Banes AJ, Gilbert J, Taylor D, et al. A new vacuum-operated stress-providing instrument that applies static or variable duration cyclic tension or compression to cells in vitro. J cell Sci. 1985; 75: 35-42.
79 Buckley MJ, Banes AJ, Jordan RD, et al. The effects of mechanical strain on osteoblasts in vitro. J Oral Maxillofac Surg. 1990; 48: 276-282.
80 Gillert JA, Weinhold PS, Banes AJ, et al. Strain Profiles for circular cell culture plates containing flexible surfaces employed to mechanically deform cells in vitro. J Biomechanics, 1994; 7: 169-77.
81 Levesque MJ, Nerem RM. The elongation and orientation of cultured endothelial cells in response to shear stress. J Biomech Engng. 1985; 107: 341-347.
82 Frangos JA, Mclntire LV, Eskin SG. Shear stress induced stimulation of mammalian cell metabolism. Biotechnol. Bioeng. 32:1053-1060,1988.
83 Tseng H, Peterson TE, Berk BC. Fluid shear stress stimulates mitogen-activated protein kinase in endothelial celis. Circulation Research. 1995; 77: 869-878.
84 Li S, Piotrowicz RS, Levin EG, et al. Fluid shear stress induces the phosporylation of small heat shock proteins in vascular endothelial cells. Am J Physiol. (Cell Physiol) 1996; 271: C994-C1000.
85 Chun TH, Ogawa Y, Tamara N, et al. Shear stress augments expression of C-type natriuretic peptide and adrenomedullin. Hypertension. 1997; 29: 1296-1302.
86 Jacobs DR, Yellowley CE, Davis BR, et al. Differential effect of ststeady versus osciliating flow on bone cells. J Biomech. 1998; 31: 969-976.
87 Heimeke BP, Thakker DB, Goldman RD, et al. Spatiotemporal analysis of
flow-induced intermediate f ilament displacement in living endothelia cells. Biophysical Journal, 2001; 80: 184-194.
88 Cao J, Usami S, Dong C. Development of a side-view chamber for studying cell-surface adhesion under flow conditions. Annals of Biomedical Engineering. 1997; 25: 573-580.
89 Dewey CF Jr, Bussolari SR, Gimbrone MA, et al. The dynamic response of vascular endothelial cells to fluid shear stress. J Biomech Engng. 1981; 103: 177-185.
90 Tran-Son-Tay R, Sutera SP. Determination of RBC membrane viscosity form rheoscopic observations of tank-treading motion. Biophysical Journal. 1984; 46: 65-72.
91 Sutera SP, Mueller ER, Zahalak GI, et al. Extensional recovery of an intact erythrocyte form a tank-treading motion. Journal of Engineering. 1990; 112: 250-256.
92 Dewey CF Jr. Efforts of fluid flow on living vascular cells. J Biomech. Engng. 1984;106(11) : 31-35.
93 Grondelle V, Albertus G, Scott Worthen, et al. Altering hydrodynamic variables influences PGI2 production by isolated lungs and endothelial cells. J Appl Physiol: Respirat Environ. Exercise Physiol. 1984; 57(2) : 388-395.
94 Schnittler Hans-Joachim, Ralf P F, et al. Improved in vitro rheological system for studying the effect of fluid shear stress on cultured celis. Am J Physiol. 1993; 265(Cell Physiol, 34) : C289-C298.
95 Langille BL. Integrity of arterial endothelium following acute exposure to high shear stress. Biorheology. 1984; 21: 333-346.
96 Nomura H, Ishikawa C, Komatsuda T, et al. A disk-type apparatus for applying fluid shear stress on cultured endothelial cell. Biorheology. 1988; 25: 461-470.
97 Waters CM, Chang JY, Glucksberg MR, et al. Mechanical forces alter growth factor relearse by pleural mesothelial cells. Am J Physiol, 1997; 272:
L552-557.
98 Vaishnav RN, Patel DJ, Atabek HB, et al. Determination of local erosion stress of the canine endothelium using a jet impingement method. ASME J Biomech Engng. 1983; 105: 77-83.
99 Ohata H, Aizawa H, Momose K. Lysophosphatidic acid sensitizes mechanical stress-induced Ca2 response via activation of phospholipase C and tyrosine kinase in cultured smooth muscle cells. Life Sciences. 1997; 60: 1287-1295.
100 Barbee KA, Mundel T, Lal R, Davies PF. Subcellular distribution of shear stress at the surface of flow-aligned and nonaligned endothelial monolayers. The American Journal of physiology. 1995; 268(4 Pt 2) : H1765-72.
101 Barbee KA, Davies PF, Lal R. Shear stress-induced reorganization of the surface topography of living endothelial celis imaged by atomic force microscopy. Circulation Research. 1994; 74(1) : 163-71.
102 Davies PF, Mundel T, Barbee KA. A mechanism for heterogeneous endothelial responses to flow in vivo and in vitro. Journal of Biomechanics; 1995; 28 (12) : 1553-60
103 Barbee KA. Changes in surface topography in endothelial monolayers with time at confluence: influence on subcellular shear stress distribution due to flow. Biochemistry and cell biology. 1995; 73(7-8) : 501
104 SatcherRLJr, Bussolari SR, Gimbrone MA Jr, et al. The distribution of fluid forces on model arterial endothelium using computational fluid dynamics. Journal of Biomechanical Engineering. 1992; 114(3) : 309-16
105 Sato M, Nagayama K, kataoka N, et al. Local mechanical properties measured by atomic force microscopy for cultured bovine endothelial cells exposed to shear stress. Journal of Biomechanics. 2000; 33: 127-35
106 Charras GT, Horton MA. Single cell mechanotransduction and its modulation analyzed by atomic force microscope indentation. Biophys J. 2002; 82(6) : 2970-81
107 Radmacher M. Measuring the elastic properties of living cells by the atomic
force microscope. Methods Cell Biol. 2002; 68:67-90
108 Sugawara M, Ishida Y, Wada H. Local mechanical properties of guinea pig outer hair cells measured by atomic force microscopy (1). Hear Res. 2002; 174(1-2): 222-9
109 Dregelid E, Svendsen E. Endothelial cell injury in human saphenous veins after manipulation and tweezer grasping. J Cardiovasc Surg (Torino). 1988; 29(4): 464-9
110 Holm A, Sundqvist T, Oberg A, et al. Mechanical manipulation of polymorphonuclear leukocyte plasma membranes with optical tweezers causes influx of extracellular calcium through membrane channels. Med Biol Eng Comput. 1999; 37(3): 410-2
111 Fujii T, Sun YL, An KN, et al. Mechanical properties of single hyaluronan molecules. J Biomech 2002; 35(4): 527-31
112 Alenghat FJ, Fabry B, Tsai KY, et al. Analysis of cell mechanics in single vinculin-deficient cells using a magnetic tweezer. Biochem Biophys Res Commun. 2000; 277(1): 93-9
113 邹远文,细胞力学脉动流实验方法与测控系统研究。博士学位论文,2002
114 白灵,改进的Flow Chamber系统的研制与应用。硕士学位论文,2002
114 郭欣,樊瑜波,宋锦磷,陈君楷。张应力作用下细胞培养膜上应力分布的三维有限元分析。生物医学工程学杂志。2002,19(1):60-63
115 膜式张应力施加装置的研制及细胞培养膜的三维有限元应力分析。硕士学位论文,2002。
117 Arnoczky SP, Lavagnino M, Whallon JH, et al. In situ cell nucleus deformation in tendons under tensile load; a morphological analysis using confocal laser microscopy. J Orthop Res. 2002; 20(1):29-35
118 Vanni S, Lagerholm BC, Otey C, et al. Internet-Based Image Analysis quantifies Contractile Behavior of Individual Fibroblasts inside Model Tissue. Biophys J. 2003; 84(4): 2715-27
119 Yoshikawa N, Ariyoshi H, Aono Y. Gradients in cytoplasmic calcium concentration ([Ca~2]) in migrating human umbilical vein endothelial cells
(HUVECs) stimulated by shear stress.
119 Aherty JT, Pierce JE, Ferrand VJ, et al. Endothelial nuclear patterns in the canine arterial tree with particular reference to hemodynamic events. Circulation Research. 1972; 30(1) : 23
120 Baker JW, Thubrikar MJ, Parekh JS, et al. Change in endothelial cell morphology at arterial branch sites caused by a reduction of intramural stress. Atherosclerosis. 1991; 89(2-3) : 209
121 Cornhill JF, levesque MJ, Hederick EE, et al. Quantitative Study of the Rabbit aortic endothelium using vascular casts. Atherosclerosis. 1980; 35: 321
122 Nerem RM, Levesque MJ, Cornhill JF. Vascular endothelial morphology as an indicator of the pattern of blood flow. Journal of Biomechanical Engineering. 1981; 103(3) : 172
123 Kataoka N, Ujita S, Sato M. Effect of flow direction on the morphological responses of cultured bovine aortic endothelial celis. Medical & Biological Engineering & Computing. 1998; 36(1) : 122.
124 Azuma N, Akasaka N, Kito H, et al. Role of p38 MAP kinase in endothelial cell alignment induced by fluid shearstress. American Journal of Physiology: Heart and Circulatory Physiology, 2001; 280(1) : H189-97
125 Palmer BM, Bizios R. Quantitative characterization of vascular endothelial cell morphology and orientation using fourier Transform analysis. Journal of Biomechanical Engineering. 1997; 119(4) : 159
126 Dewey, CF. Effects of fluid flow on living vascular cells. Journal of Biomechanical Engineering. 1984; 106(1) : 31-35
127 Dieterich P, Odenthal-Schnittler M, Mrowietz C, et al. Quantitative morphodynamics of endothelial cells within confluent cultures in response to fluid shear stress. Biophysical Journal. 2000; 79(3) : 1285-97
128 Barbee KA, Mundel T, Lal R, Davies PF. Subcellular distribution of shear stress at the surface of flow-aligned and nonaligned endothelial monolayers.
The American Journal of Physiology, 1995; 268(4 Pt 2) : H1765-72.
129 Davies PF, Mundel T, Barbee KA. A mechanism for heterogeneous endothelial responses to f low in vivo and in vi tro. Journal of Biomechanics, 1995; 28(12) : 1553
130 Satcher RL Jr, Bussolari SR, Gimbrone MA Jr, et al. The distribution of fluid forces on model arterial endothelium using computational fluid dynamics. Journal of Biomechanical Engineering, 1992; 14(3) : 309
131 Sato M, Nagayama K, kataoka N, et al. Local mechanical properties measured hy atomic force microscopy for cultured bovine endothelial cells exposed to shear stress. Journal of Biomechanics, 2000; 33:127
132 Stamatas GN, Mc Intire LV. Rapid flow-induced responses in endothelial cells. Biotechnology Progress. 2001; 17(3) : 383
133 Hazel AL, Pedley TJ. Vascular endothelial cells minimize the total force on their nuclei. Biophysical Journal, 2000; 78(1) : 47
134 GalbraithCG, SkalakR, Chien S. Shear stress induces spatial reorganization of the endothelial cell cytoskeleton. Cell Motility and the Cytoskeleton, 1998; 40(4) : 317
135 Satcher R, Dewey CF Jr, Hartwig JH. Mechanical remodeling of the endothelial surface and actin cytoskeleton induced by fluid flow. Microcirculation, 1997; 4(4) : 439
136 Helmke BP, Thakker DB, Goldman RD. et al. Spatiotemporal analysis of flow-induced intermediate filament displacement in living endothelial cells. Biophysical Journal, 2001; 80(1) : 184
137 Cornhill JF, levesque MJ, Hederick EE, et al. Quantitative Study of the Rabbit aortic endothelium using vascular casts. Atherosclerosis 1980, 35: 321-337
138 程范军,杨爱德,费洪宝等。急性粒细胞白血病细胞形态定量初步研究。同济医科 大学学报。1994;2391) :65~6
139 Cammer M, Wyckoff J, Segall JE. Computer-assisted analysis of single-cell
behavior. Methods in Molecular. 1997; 75: 459-70
140 Albertini MC, Teodori L, Piatti E, et al. Automated analysis of morphometric parameters for accurate definition of erythrocyte cell shape. Cytometry 2003; 52A(1) : 12-8
141 Segovia C, Hutchinson I, Laing DG, et al.A quantitative study of fungiform papillae and taste pore density in adults and children. Brain Res Dev Brain Res 2002; 138(2) : 135-46
142 Yamauchi Y, Ikeda R, Michitaka K, et al. Morphometric analysis of lymphatic vessels in primary biliary cirrhosis. Hepatol Res. 2002; 24(2) : 107
143 Ghauharali RI, Driel RV, Brakenhoff GJ. Structure-oriented fluorescence photobleaching analysis: a method for double fluorescent labeling studies. Journal of microscopy. 1997; 185(3) :375-84
144 Wong K, Wessels D, Krob SL, et al. Forced expression of a dominant-negative chimeric tropomyosin causes abnormal motile behavior during cell division. Cell Motil Cytoskeleton. 2000; Feb; 45(2) :121-32
145 Arribas SM, Gonzalez C, Graham D, et al. Cellular changes induced by chronic nitric oxide inhibition in intact rat basilar arteries revealed by confocal microscopy. J Hypertens. 1997; 15(12 Pt 2) : 1685-93
146 Arribas SM, Hillier C, Gonzalez C, et al. Cellular aspects of vascular remodeling in hypertension revealed by confocal microscopy. Hypertension 1997; 30(6) : 1455-64
147 Klimaschewski L, Nindl W, Pimpl M, et al. Biolistic transfection and morphological analysis of cultured sympathetic neurons. J Neurosci Methods. 200; 113(1) : 63-71
148 Cheng W, Tam PK. Apoptosis in murine duodenum during embryonic development. Pediatr Surg Int. 2000; 16(7) : 485-7
149 Carai A, Diaz G, Santa Cruz R, et al. Computerized quantitative color analysis for histological study of pulmonary f ibrosis. Anticancer Res. 2002; 22(6C): 3889-94
150 Bagyanszki M, Kovacs EG, Resch BA, et al. Computer-aided morphometric analysis of the developing concentric structure of the human fetal intestinal tube. Histol Histopathol. 2002; 17(3): 731-7
151 韩林,张宝仁,柳兆荣等,稳态层流下的内皮细胞形态特征变化。第二军医大学学报,1999;20(10):801-3
152 Cheung SW, Cho P. Endothelial ceils analysis with the TOPCON specular microscope SP-2000P and IMAGEnet system. Current Eye Research. 2000; 21(4): 788-98
153 王速,赵捷,吉田洋二等,流体力学模型个区域内皮细胞的形态学变化。中国医科大学学报,1996;25(6):551-5
154 Berns GS, Berns MW. Computer-based tracking of living cells. Experimental cell research, 1982;142:103-9
155 Siegert F, Weijer CJ, Nomura A, et al. A gradient method for the quantitative analysis of cell movement and tissue flow and its application to the analysis of multicellular Dictyostelium development. Journal of Cell Science. 1994; 107(1): 97-104.
156 Helmke BP, Thakker DB, Goldman RD, et al. Spatiotemporal analysis of flow-induced intermediate filament displacement in living endothelial cells. Biophysical Journal, 2001;80(1):184-94.
157 舍英,伊力奇,呼和巴特尔。现代光学显微镜。科学出版社,1997,北京
158 JVC—C680使用说明书
159 Piarea 150ES产品说明书,2001
160 张松梅编著,C++语言教程,电子科技大学出版社,1998
161 周长发著,精通Visual c++图象编程,电子工业出版社,2000
162 Kruglinski DJ, Wingo S,Shepherd G.著,希望图书创作室译,Visual C++6.0技术内幕。北京希望电子出版社,1999
163 阮秋琦编著,数字图象处理学。电子工业出版社,北京,2001
164 章毓晋编著,图象工程上册—图象处理和分析。清华大学出版社,1999
165 Canny J. A Computational approach to edge dextion. IEEE-PAMI, 1986, 8:
679-98
166 章毓晋著,图象分割。科学出版社,北京,2001
167 Ziou D, Tabbone S. Edge detection techniques-An overview. IEEE Pattern Recognition and Image Analysis. 1998; 8(4): 1-41
168 Buxton B. Early Image Processing: Structural Techniques Motivated by Human Visual Response. Unsiversity of Surrey, 1984
169 Canny J. A computational approach to edge detection. IEEE PAMI, 1986; 8: 679-98
170 崔屹编著,图象处理与分析—数学形态学方法及应用。科学出版社,北京2002
171 Vincent L, Morphological grayscale reconstruction in image analysis: applications and efficient algorithms. IEEE Transactions on image processing 1993; 2(2):176-201
172 Digabel H, Lantuéjoul C. Iterative algorithms. In Actes du Second Symposium Européen d' Analyse Quantitative des Microstructures en Sciences des Matériaux, Biologie et Médecine, Caen, 4-7 October 1977(1978),J.-L. Chermant, Ed., Riederer Verlag, Stuttgart, pp. 85-99
173 Lantu éjoul C. La squelettisation et son application aux mesures topologiques des mosa ques polycristallines. PhD thesis, Ecole des Mines, Paris, 1978
174 Beucher S, Lantuéjoul C. Use of watersheds in contour detection. In Proc. International Workshop on Image Processing, Real-Time Edge and Motion Detection/Estimation, Rennes, setember, 1979
175 Nanjman L, Schmitt M. Geodesic saliency of watershed contours and hierarchical segmentation. IEEE. Trans. PAMI, 1996; 18(12): 1163-73
176 Vincent L, Soille P. Watersheds in digital spaces: an efficient algorithm based on immersion simulations. IEEE. Trans. PAMI, 1991; 13(6): 583-98
177 Beucher S, Watersheds of functions and picture segmentation. In Proc. IEEE Int. Conf. Acoustics, Speech, and Signal Processing, Paris, France, May 1982,1928-31
178 Vicent L, Beucher S. The morphological approach to segmentation: an introduction. School of Mines, Paris, France, Internal Rep. CMM, July 1989
179 Haralick R, Shapiro L. Survey: Image segmentation techniques. Comput. Vision, Graphics, Image Processing. 1985; 29: 100-32
180 Monga O, An optimal region growing algorithm for image segmentaion. Int. J. pattern Recog. Artificial Intell. 1987; 3(4)
181 Cornhill JF, levesque MJ, Hederick EE, et al. Quantitative Study of the Rabbit aortic endothelium using vascular casts. Atherosclerosis, 1980; 35: 321
182 Nerem RM, Levesque MJ, Cornhill JF. Vascular endothelial morphology as an indicator of the pattern of blood flow. Journal of Biomechanical Engineering. 1981; 103(3) :172
183 Stevens AI,Walnum C.著,林丽闽,别红霞等译,标准C++宝典。电子工业出版 社,北京,2001
184 张桂林,陈益新,曹伟烜等,基于跑长码的连通区域标记算法。华中理工大学学报, 1994:22(5) :11-14
185 Levesque MJ, Nerem RM. The Study of Rheological effects on vascular endothelial cells in culture. Biorheology. 1989; 26: 345-57
186 Dewey CF Jr, Effects of fluid flow on living vascular cells. ASMEJ Bio. Mech. Engr. 1984; 106: 31
187 White GE, Gimbrone MA Jr, Fujiwara K. Factors influencing the expression of stress fibers in vascular endothelial cells in situ. J Cell Boil. 1983; 97: 416
188 Sholley MM, Gimbrone MA Jr, Controm RS. Cell migration and replication in endothelial regeneration: study using irradiated endothelial culture. Lab Invest. 1977; 36: 18
189 Grabowski EF, Jaffe EA, Wekaler BB. Prostacyclin production by cultured endothelial cell monolayers exposed to step increases in shear stress. J Lab Clin Med. 1985; 105: 36
190 Lamontague D, Pohl U, Busse R. Mechanical deformation of vessel wall and shear stress determine the basal release of endothelium-derived relaxing factor in the intact rabbit coronary vascular bed. Circ Res. 1992; 70:123
191 Sprague EA, Steinbach BL, Nerem RM, et al. Influence of laminar steady-state fluid imposed wall shear stress on the binding, internalization and degradation of flow-density lipoproteins by cultured arterial endothelium. Lab. Invest, 1987; 76:648
192 艾珣,王克强,赵子琴等。切应力对培养人脐静脉内皮细胞形态的影响。解剖学杂志。1997:20(1),81-87
193 韩林,张宝仁,柳兆荣等。稳态层流下的内皮细胞形态特征变化。第二军医大学学报。1999;20(10),801-805