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390	JENNIFER A. McCANN, THOMAS J. WEBSTER, AND KAREN M. HABERSTROH

Fluid Shearing

BLOOD

GDP-p21 Ras GTP-p21ras

Membrane

MEKK Raf-1

JNKK MEK

JNK ERK

c-Jun p62 TCF

/c-Fos

Cytoplasm

AP-1
TRE	Nucleus
e.g., MCP-1	Nucleus

FIGURE 18.8. Schematic diagram of the Ras-MAPK signal transduction pathways in response to ﬂuid shear stress. (Reprinted from Molecular and Cellular Biology, Vol 16, Li, Shyy, Li, Lee, Su, Karin, and Chien, The Ras-JNK pathway is involved in shear-induced gene expression, 5947-5954, Copyright 1996, with permission from the American Society for Microbiology)

pathway, which allows for p21ras activation. Another potential upstream mediator involves increased phosphorylation and activity of focal adhesion kinase (FAK). This activation leads to recruitment of Src and continuation of FAK phosphorylation, which allows FAK to associate with the complex growth factor receptor-binding protein 2 (Grb2) and guanine nucleotide exchange factor, son of sevenless (Sos) [17]. It is this complex which further activates Ras (Figure 18.9).

Shear stress

Membrance

RTKs (e.g., Fik-1)

Nucleus

Shc

ERK

AP-1

Grb2-Sos

Ras

e.g., MCP-1

JNK

Cytoplasm

Grb2-Sos

TRE

Focal adhesion site

Shc integrins (e.g.,

αvβ3)

Extracellular matrix

FIGURE 18.9. A proposed transduction pathway in endothelial cells in response to shear stress. (Reprinted from the Journal of Biological Chemistry, Vol 274, Chen, Li, Kim, Li, Yuan, Chien, and Shyy, Mechanotransduction in response to shear stress, 18393-18400, Copyright 1999, with permission from the American Society for Biochemistry and Molecular Biology)

VASCULAR CELL RESPONSES TO FLUID SHEAR STRESS

391

Once activated, p21ras triggers Raf-1 and MEKK (MAPK kinase kinase); in turn these stimulate ERK (extracellular signal regulated kinases) and JNK (c-Jun NH2-terminal kinases), respectively (Figure 18.8). ERK and JNK are MAPKs; ERK mediates cell growth while JNK mediates programmed cell death. When shear stress alone is the extracellular stimulus, JNK is activated to a much greater extent than ERK. In the case of JNK activation, c-Jun is expressed and crosses into the nucleus where it interacts with the transcription factor AP-1. AP-1 then interacts with the 12-O-tetradecanoyl-phorbol-13-acetate-responsive element (TRE) which mediates gene expression [42]; for example, MCP-1 expression is mediated through TRE [72]. Activation of ERK1 and ERK2 result in up regulation of Elk-1, [28] which crosses into the nucleus to interact with SRE. In turn, SRE affects gene expression of such molecules as c-fos [85].

18.10.2. IKK-NF · κ B Pathway

The IKK-NK- κB pathway is likely initiated by the αvβ3 integrin, which is stimulated by shear stress as was previously discussed. NF·κB is a transcription factor which mediates gene expression [12]; laminar shear stress up regulates its transcriptional activity [73]. NF·κB activation requires phosphorylation and then degradation of the I κB proteins, which inhibit NF·κ B activity. This process is mediated by the IKKs (IkB kinases), which are in part mediated by αvβ3 integrin expression in response to ﬂow (Bhullar, 1998). The IKKs phosphorylate I κB, which subsequently degrades, allowing NF κB to translocate into the nucleus where it affects gene expression. It has also been suggested that MEKK is capable of stimulating this pathway, demonstrating the intricate and complex network involved in mechanotransduction [42].

18.11. APPLICATIONS TO CLINICAL TREATMENT

Understanding these critical signaling mechanisms involved in disease progression may allow for the design of pharmacological treatments with increased efﬁcacy. In addition, knowledge of the relationship between gene expression and cardiovascular disease development may lead to gene manipulation strategies to restore or prevent speciﬁc gene expression (Heiko, 2001). As an example, it may be possible to restore the effects of NO via ecNOS gene transfer. Whatever the strategy, ultimately, results obtained from the ﬂuid shear stress studies outlined in this chapter will aid physicians in diagnosing and treating the clinical presentation of atherosclerosis.

18.12. SUMMARY

Vascular cell responses to hemodynamic forces and biochemical stimulation dictate vessel behavior. For this reason, endothelial cell responses to ﬂuid ﬂow have been widely studied using ﬂow models (e.g., the parallel plate ﬂow chamber) which allow scientists and engineers to investigate cell functions under well deﬁned ﬂows. In such models, cells sense the ﬂow via surface receptors, which activate one to many intracellular signaling cascades, thereby affecting the cell nucleus. In turn, cell functions including cell growth,

392	JENNIFER A. McCANN, THOMAS J. WEBSTER, AND KAREN M. HABERSTROH

mRNA expression, and protein production are altered. These responses are directly related to the ﬂow environment; laminar ﬂuid ﬂow yields healthy/physiological cell functions, while disturbed ﬂow patterns favor cardiovascular disease development. Understanding the relationships between such intracellular signaling pathways and cell functions in response to these ﬂow types (through studies such as those outlined in this chapter) will aid in understanding disease initiation and progression, as well as in the development of clinical treatment strategies.

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About the Editors

Professor Mauro Ferrari is a pioneer in the ﬁelds of bioMEMS and biomedical nanotechnology. As a leading academic, a dedicated entrepreneur, and a vision setter for the Nation’s premier Federal programs in nanomedicine, he brings a three-fold vantage perspective to his roles as Editor-in-Chief for this work. Dr. Ferrari has authored or co-authored over 150 scientiﬁc publications, 6 books, and over 20 US and International patents. Dr. Ferrari is also Editor-in-Chief of Biomedical Microdevices and series editor of the new Springer series on Emerging Biomedical Technologies.

Several private sector companies originated from his laboratories at the Ohio State University and the University of California at Berkeley over the years. On a Federal assignment as Special Expert in Nanotechnology and Eminent Scholar, he has provided the scientiﬁc leadership for the development of the Alliance for Cancer Nanotechnology of the National Cancer Institute, the world-largest medical nanotechnology operation to date. Dr. Ferrari trained in mathematical physics in Italy, obtained his Master’s and Ph.D. in Mechanical Engineering at Berkeley, attended medical school at The Ohio State University, and served in faculty positions in Materials Science and Engineering, and Civil and Environmental Engineering in Berkeley, where he was ﬁrst tenured. At Ohio State he currently serves as Professor of Internal Medicine, Division of Hematology and Oncology, as Edgar Hendrickson Professor of Biomedical Engineering, and as Professor of Mechanical Engineering. He is Associate Director of the Dorothy M. Davis Heart and Lung Research Institute, and the University’s Associate Vice President for Health Science, Technology and Commercialization.

Rashid Bashir completed his Ph.D. in 1992. From Oct 1992 to Oct 1998, he worked at National Semiconductor in the Process Technology Development Group as Sr. Engineering Manager. He is currently a Professor of Electrical and Computer Engineering and Courtesy Professor of Biomedical Engineering at Purdue University. He has authored or coauthored over 100 journal and conference papers, has over 25 patents, and has given over 30 invited talks. His research interests include biomedical microelectromechanical systems, applications of semiconductor fabrication to biomedical engineering, advanced semiconductor fabrication techniques, and nano-biotechnology. In 2000, he received the NSF Career Award for his work in Biosensors and BioMEMS. He also received the Joel and Spira Outstanding Teaching award from School of ECE at Purdue University, and the Technology Translation Award from the 2001 BioMEMS and Nanobiotechnology World Congress Meeting in

396	ABOUT THE EDITORS

Columbus, OH. He was also selected by National Academy of Engineering to attend the Frontiers in Engineering Workshop in Fall 2003. https://engineering.purdue.edu/LIBNA

Professor Steve Wereley completed his masters and doctoral research at Northwestern University and joined the Purdue University faculty in August of 1999 after a two-year postdoctoral appointment at the University of California Santa Barbara in the Department of Mechanical and Environmental Engineering. At UCSB he focused exclusively on developing diagnostic techniques for microscale systems, work which ultimately led to developing, patenting, and licensing to TSI, Inc., the micro-Particle Image Velocimetry technique. His current research interests include designing and testing microﬂuidic MEMS devices, investigating biological ﬂows at the cellular level, improving micro-scale laminar mixing, and developing new micro/nano ﬂow diagnostic techniques. Professor Wereley has co-authored Fundamentals and Applications of Microﬂuidics, Artech House, 2002.

Index

Page numbers with t and f indicate table and ﬁgure respectively.

1-bromonaphthalene (1-BrNp), 333

2-[Methoxy(polyethylenoxy)propyl] trimethoxysilane (PEG-silane), 31

3,3-Dithiobis [sulfo-succinimidylpropionate] (DTSSP), 31

3T3 ﬁbroblast cells, 174 5-carboxymethoxy-2-nitrobenzyl (CMNB)-caged

ﬂuorescein, 333

Absorption measurements of a pH sensitive dye, 12 AC electrokinetic phenomena, 244

AC electroosmosis, 244 dielectrophoresis, 244 electrophoresis, 244

AC electroosmosis. See AC electrokinetic phenomena AC MHD micropump, 141–142

measurements in, 152,187 Acrylic. See PMMA, 98 Active mixers, 233

Active valves, 228 Adhesive bonding, 111

Adhesive stamping technique, 28 Airy function, 266 Amperometric detection, 57 Amplitude modulation, 76 Analytical instruments, 95 Anionic networks, 122 Antibodies, characteristics of, 59

Antibody binding sites, free streptavidin blocking of, 48

Antibody-antigen bindings, 28

Antibody-antigen interactions. See Protein-protein bindings, 30

Antigen antibody interaction, 10 Anti-lysozyme F-ab fragment, 7

Array of signaling systems characteristic of cell-based sensors, 87

Arthrobacter nicotiane microorganism, 10

Artiﬁcial membranes, use of, 11 Atherosclerosis initiation, 371 Atherosclerotic vessels,

pathology, 374 physiology, 372

Atomic force microscope, 196 Avidin oxidase, 7

AZ 4620, 95

Bacteria, 171 Big I/O, 28

Biochip capability, 89

Biochip technologies, application of, 18 BioChip, deﬁnition of, 190

Biochips

microarray systems, 14 integrated biochip systems, 16

Biocompatibility of materials & processing, biocompatibility tests, 113

material response, 113

tissue & cellular response, 113 Biocompatibility, 63–64

Biological ﬂurescence. See Bioluminescence Bioluminescence, 57, 195

Bioluminescent catabolic reporter bacterium, 57

Biomedical micro devices (BMMD), 95, 113 advantages of, 110

drawbacks of, 95 fabrication of, 94 size of, 96

Biomedical micro devices, materials for, polymers, 95–96

silicon & glass, 93–94

Biomedical micro devices, packaging of, adhesive bonding, 110

interconnects, 109

thermal direct bonding, 109

398

BioMEMS for cellular detection electrical detection, 197 mechanical detection, 196 optical detection, 194–195

BioMEMS for cellular manipulation and separation dielectrophoresis, 191–196

electrophoresis, 189 BioMEMS sensors, use of, 194 Biomimetic ligand, 10

Biomimetic receptors, construction methods of recombinant techniques, 11

Biomolecular reaction arrays deﬂection of DNA, 29–31 deﬂection of PSA, 28–30

Biomolecular separators, 304 Biomolecules, immobilization of, 25 Bioreceptors, bases of

cellular systems, 9 enzymes, 9 non-enzymatic proteins, 9

Bioreceptors, forms of antigen interactions, 3 cellular interactions, 5 enzymatic interactions, 5

interactions using biomimetic materials, 5 nucleic acid interactions, 5

Bioreceptors, schematic diagram of, 6f Bioreceptors, types of

antibody bioreceptors, 5 biomimetic bioreceptors, 10 cellular bioreceptors, 9 enzyme bioreceptors, 8 nucleic acid bioreceptors, 14

Bioreceptors, types of, 14 antibody, 5–8 biomimetic, 10 cellular, 9–10

enzyme, 8 nucleic acid, 14

Biosensing process, principle of, 4f Biosensing system

chip assembly, 67–68 environmental chamber, 68

Biosensor for protein and amino acid estimation, 13 Biosensor, deﬁnition of, 55

Biosensors, application of, 66,80,82 biological monitoring, 5 environmental sensing applications, 5

Biotin, 11

Biotinylated antibody, 7 Bisensors

types of bioreceptors. See Bioreceptors, types of types of transducers. See Transducers, forms of

Blood ﬂow, hemodynamics of, 375 cyclic stain forces on, 377

shear stress force on, 380

INDEX

Bnzo[a]pyrene tetrol (BPT), 7–8 Bovine Serum Albumen (BSA), 45

BRCA1 gene. See cancer gene, screening of Breast cancer gene BRCA1, 18

Bromine production, 13

Brownian diffusion effects in nPIV. See Nano-particle image velocimetry

Brownian motion of particles, 162, 248, 275, 338 Bulk micromachining, 94

Bulk silicon micromachining, 105

Calcium Green-1, 58 Calibration of the sensor, 14

Caliper life science microﬂuidic chip, 365 Cancer gene, screening of, 15

Cantilever bending, 21, 23 Cantilever deﬂection, 30

Cantilever response to DNA hybridization plotted as a function of target DNA concentration, 30f

Cantilever sensor, noise of, 24 Cantilever, fabrication of, 36

Cantilever, surface-stress change, measurement of, 27

Cantilever’s deﬂection, 23 Cantilevers curvature, use of, 26

Cantilevers, signal-to-noise ratio of, 27 Capacitance cytometry, 152

Cardiac pacemakers, 205 Cardiovascular diseases, 376 Cascaded sensing, 79 Catch-and-tell operation, 58 Causative agent, 10

CCD screen, 26 CCD spot, 26

Cell based biosensors designs & methods of biosensing system, 67

cell culture, 68–69

cell manipulation techniques, 64

cell manipulation using dielectrophoresis (DEP), 66

cell types and parameters for dielectrophoretic patterning, 67–68

experimental measurement system, 67 principles of dielectrophoresis (DEP), 89 requirements for cell based sensors, 63

Cell based biosensors, measurements of

inﬂuence of geometry and environmental factors on the noise spectrum, 74

interpretation of bioelectric noise, 76 long-term signal recording in vivo, 69 selection of chemical agent, 76 signal processing, 76

Cell based biosensors, types of

cellular microorganism based biosensors, 57–58 extra cellular potential based biosensors, 61–63 ﬂuorescence based cellular biosensors, 58–59

INDEX

impedance based cellular biosensors, 59 intracellular potential based biosensors, 59–61

Cell based sensing, 56

Cell biosensor for environmental monitoring, 200

Cell biosensor, speciﬁc for, formaldehyde, 57 Cell culture

neuron culture, 68

primary osteoblast culture, 69–70 Cell monitoring-system, 198

Cell organelles, 10

Cell patterning methods

biocompatible silane elastomers, use of, 64 micro-contact printing (µCP), 64 topographical method, 64

Cell positioning over the electrodes, technique for isolating and positioning, 69

Cell signal transducting, approaches or, 57 Cell-based biosensors, applications of, 87 Cell-gel sensors. See Microﬂuidic tectonics,

sensors in

Cells, use of, in environmental applications, 57 Central difference interrogation (CDI), 271 Charged couple device (CCD) camera, 26–27 Charged couple device (CCD) detection, 26 Chemical agent sensing

cascaded sensing of chemical agents using single osteoblast, 87

comparison of detection limits and response times, 82

EDTA sensing using single osteoblast, 85 effect of verifying concentration of chemical

agents, 85

ethanol sensing using single osteoblast, 79 hydrogen peroxide servicing using single

osteoblast, 79–81

pyrethroid servicing using single osteoblast, 79–80

Chemiluminescence, 194 CHF3 RIE, 36

Chip assembly, integration with silicone chamber, 67

Chip-base capillary electrophoresis devices, 223 Chip-based fabrication, advantages of, 45 Cholera toxins, binding of, 11 Claussius-Mossotti factor 65, 164–168,190,261 CMOS biochip coupled to multiplex capillary

electrophoresis (CE) system, 17 CMOS technology, 17, 211 Cochlear prosthetics, 205

Codeine, 11

Coenzyme, 8

Colloid concentration, 46, 50 Colloids, binding capacity of, 46 Colloids, binding of, to antibodies, 46 Compartmentalization, 231–232

399

Complementary metal oxide silicon technology. See CMOS technology

Computer generated holograms (CGHS), 350–351

Cone and plate viscometer, 382 Conformal mapping, 176

Continuous window shifting (CWS), 269 Continuum model of EOF, 324

experimental data of, 320–324 Coulomb body forces in ﬂuid, 244 Coulombic interactions, 9

Coulter counter principle, 199 Coulter counter technique, 35 Coulter counter

application of, 36 sensitivity of, 37

Covalently binding antibodies, self assembled method for, 14

COX-2 mRNA expression, 384 Carcinogen benzo[a]pyrene, 7 Crossover frequency, 65 Cy5-labeled antibody, 17 Cyclooxygenase (COX) 2, 388 Cyclopentanone (CP), 97 Cytosensor microphysiometer, 198 Cytotoxicity, 114

Czarnik’s compound, 58

DC MHD micropump, 144 de Silva’s compound, 58

Debye-Huckel approximation, 306 Deconvolution method, 262–263

drawbacks of, 263 hypothesis of, 263

Deep reactive etching (DRIE), 95 Deionized (DI), 99

DEP. See Dielectrophoresis Detection of E. coli, 18

Detect-to-treat. See Cell based sensors Detect-to-warn. See Cell based sensors Diagnostic wireless microsystems, 210–213 Dielctrophoretic force, 245

Dielectrophorectic device, performance quantiﬁcation experiments of, 266–268

Dielectrophoresis (DEP), 64, 66, 79, 160, 162–165, 190–196. See also AC electrokinetic phenomena

applications of, 192–193 Claussius-Mossotti factor, 164–168 electrothermal forces, effect of, 171–172 interaction with gravitational force, 169–171 interaction with hydrodynamic drag forces,

170–171

multipolar effects, 167–169 negative, 190

positive, 190 scaling, 168–169

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