Effect of Age and Cytoskeletal Elements on the Indentation-Dependent Mechanical Properties of Chondrocytes

Chahine, Nadeen O.; Blanchette, Craig; Thomas, Cynthia B.; Lu, Jeffrey; Haudenschild, Dominik; Loots, Gabriela G.

doi:10.1371/journal.pone.0061651

Title: Effect of Age and Cytoskeletal Elements on the Indentation-Dependent Mechanical Properties of Chondrocytes

Journal Article · Tue Apr 16 00:00:00 EDT 2013 · PLoS ONE

DOI:https://doi.org/10.1371/journal.pone.0061651· OSTI ID:1627600

Chahine, Nadeen O. ^[1]; Blanchette, Craig ^[2]; Thomas, Cynthia B. ^[2]; Lu, Jeffrey ^[3]; Haudenschild, Dominik ^[3]; Loots, Gabriela G. ^[4]

Hofstra Univ. Manhasset, NY (United States). Hofstra North Shore LIJ. School of Medicine. The Feinstein Inst. for Medical Research
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States). Physical and Life Sciences Directorate. Biosciences and Biotechnology Division
Univ. of California, Davis, Sacramento, CA (United States). Medical Center. Dept. of Orthopaedic Surgery. Lawrence J. Ellison Musculoskeletal Research Center
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States). Physical and Life Sciences Directorate. Biosciences and Biotechnology Division; Univ. of California, Merced, CA (United States). School of Natural Sciences

Articular cartilage chondrocytes are responsible for the synthesis, maintenance, and turnover of the extracellular matrix, metabolic processes that contribute to the mechanical properties of these cells. Here, we systematically evaluated the effect of age and cytoskeletal disruptors on the mechanical properties of chondrocytes as a function of deformation. We quantified the indentation-dependent mechanical properties of chondrocytes isolated from neonatal (1-day), adult (5-year) and geriatric (12-year) bovine knees using atomic force microscopy (AFM). We also measured the contribution of the actin and intermediate filaments to the indentation-dependent mechanical properties of chondrocytes. By integrating AFM with confocal fluorescent microscopy, we monitored cytoskeletal and biomechanical deformation in transgenic cells (GFPvimentin and mCherry-actin) under compression. We found that the elastic modulus of chondrocytes in all age groups decreased with increased indentation (15–2000 nm). The elastic modulus of adult chondrocytes was significantly greater than neonatal cells at indentations greater than 500 nm. Viscoelastic moduli (instantaneous and equilibrium) were comparable in all age groups examined; however, the intrinsic viscosity was lower in geriatric chondrocytes than neonatal. Disrupting the actin or the intermediate filament structures altered the mechanical properties of chondrocytes by decreasing the elastic modulus and viscoelastic properties, resulting in a dramatic loss of indentation-dependent response with treatment. Actin and vimentin cytoskeletal structures were monitored using confocal fluorescent microscopy in transgenic cells treated with disruptors, and both treatments had a profound disruptive effect on the actin filaments. Here we show that disrupting the structure of intermediate filaments indirectly altered the configuration of the actin cytoskeleton. These findings underscore the importance of the cytoskeletal elements in the overall mechanical response of chondrocytes, indicating that intermediate filament integrity is key to the non-linear elastic properties of chondrocytes. This study improves our understanding of the mechanical properties of articular cartilage at the single cell level.

View Accepted Manuscript (DOE)

Cite

Export

Save

Research Organization:: Lawrence Livermore National Laboratory (LLNL), Livermore, CA (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division

Grant/Contract Number:: AC52-07NA27344

OSTI ID:: 1627600

Journal Information:: PLoS ONE, Vol. 8, Issue 4; ISSN 1932-6203

Publisher:: Public Library of ScienceCopyright Statement

Country of Publication:: United States

Language:: English

References (60)

Maturation-related compressive properties of rabbit knee articular cartilage and volume fraction of subchondral tissue Wei, Xiaochun; Räsänen, Tuomas; Messner, Karola Osteoarthritis and Cartilage, Vol. 6, Issue 6 https://doi.org/10.1053/joca.1998.0143	journal	November 1998
Cytochalasin D and platelet gelsolin accelerate actin polymer formation. A model for regulation of the extent of actin polymer formation in vivo Tellam, Ross; Frieden, Carl Biochemistry, Vol. 21, Issue 13 https://doi.org/10.1021/bi00256a027	journal	June 1982
The Chondrocyte: a cell Under Pressure Urban, J. P. G. Rheumatology, Vol. 33, Issue 10 https://doi.org/10.1093/rheumatology/33.10.901	journal	January 1994
Variations in the intrinsic mechanical properties of human articular cartilage with age, degeneration, and water content. Armstrong, C. G.; Mow, V. C. The Journal of Bone & Joint Surgery, Vol. 64, Issue 1 https://doi.org/10.2106/00004623-198264010-00013	journal	January 1982
Surface ultrastructure and mechanical property of human chondrocyte revealed by atomic force microscopy Hsieh, C. -H.; Lin, Y. -H.; Lin, S. Osteoarthritis and Cartilage, Vol. 16, Issue 4 https://doi.org/10.1016/j.joca.2007.08.004	journal	April 2008
Actin filament capping and cleaving activity of cytochalasins B, D, E, and H Urbanik, Else; Ware, Bennie R. Archives of Biochemistry and Biophysics, Vol. 269, Issue 1 https://doi.org/10.1016/0003-9861(89)90098-2	journal	February 1989
Vimentin contributes to changes in chondrocyte stiffness in osteoarthritis: VIMENTIN AFFECTS CHONDROCYTE STIFFNESS Haudenschild, Dominik R.; Chen, Jianfen; Pang, Nina Journal of Orthopaedic Research, Vol. 29, Issue 1 https://doi.org/10.1002/jor.21198	journal	November 2010
Intermediate-size Filaments: Changes in Synthesis and Distribution in Cells of the Myogenic and Neurogenic Lineages Holtzer, H.; Bennett, G. S.; Tapscott, S. J. Cold Spring Harbor Symposia on Quantitative Biology, Vol. 46, Issue 0 https://doi.org/10.1101/sqb.1982.046.01.033	journal	January 1982
An Automated Approach for Direct Measurement of Two-Dimensional Strain Distributions Within Articular Cartilage Under Unconfined Compression Wang, Christopher C-B.; Deng, Jian-Ming; Ateshian, Gerard A. Journal of Biomechanical Engineering, Vol. 124, Issue 5 https://doi.org/10.1115/1.1503795	journal	September 2002
Stress distributions and material properties determined in articular cartilage from MRI-based finite strains Butz, Kent D.; Chan, Deva D.; Nauman, Eric A. Journal of Biomechanics, Vol. 44, Issue 15 https://doi.org/10.1016/j.jbiomech.2011.08.005	journal	October 2011
Mechanical Properties and Gene Expression of Chondrocytes on Micropatterned Substrates Following Dedifferentiation in Monolayer Darling, Eric M.; Pritchett, Poston E.; Evans, Benjamin A. Cellular and Molecular Bioengineering, Vol. 2, Issue 3 https://doi.org/10.1007/s12195-009-0077-3	journal	August 2009
Mechanical compression modulates matrix biosynthesis in chondrocyte/agarose culture Buschmann, Michael D.; Gluzband, Yehezkiel A.; Grodzinsky, Alan J. Journal of Cell Science, Vol. 108, Issue 4 https://doi.org/10.1242/jcs.108.4.1497	journal	April 1995
Contact pressures in the human hip joint measured in vivo. Hodge, W. A.; Fijan, R. S.; Carlson, K. L. Proceedings of the National Academy of Sciences, Vol. 83, Issue 9 https://doi.org/10.1073/pnas.83.9.2879	journal	May 1986
Events in Articular Chondrocytes with Aging Leong, Daniel J.; Sun, Hui B. Current Osteoporosis Reports, Vol. 9, Issue 4 https://doi.org/10.1007/s11914-011-0070-3	journal	September 2011
Biosynthetic response of cartilage explants to dynamic compression Sah, Robert L. -Y.; Kim, Young-Jo; Doong, Joe-Yuan H. Journal of Orthopaedic Research, Vol. 7, Issue 5 https://doi.org/10.1002/jor.1100070502	journal	September 1989
Recovery of elasticity of aged human epithelial cells in vitro Sokolov, Igor; Iyer, Swaminathan; Woodworth, Craig D. Nanomedicine: Nanotechnology, Biology and Medicine, Vol. 2, Issue 1 https://doi.org/10.1016/j.nano.2005.12.002	journal	March 2006
Optical determination of anisotropic material properties of bovine articular cartilage in compression Wang, Christopher C. -B.; Chahine, Nadeen O.; Hung, Clark T. Journal of Biomechanics, Vol. 36, Issue 3 https://doi.org/10.1016/s0021-9290(02)00417-7	journal	March 2003
Synthesis of cartilage matrix by mammalian chondrocytes in vitro. I. Isolation, culture characteristics, and morphology. Kuettner, K. E.; Pauli, B. U.; Gall, G. Journal of Cell Biology, Vol. 93, Issue 3 https://doi.org/10.1083/jcb.93.3.743	journal	June 1982
Organisation of the chondrocyte cytoskeleton and its response to changing mechanical conditions in organ culture Durrant, L. A.; Archer, C. W.; Benjamin, M. Journal of Anatomy, Vol. 194, Issue 3 https://doi.org/10.1046/j.1469-7580.1999.19430343.x	journal	April 1999
Cross-bridge cycling gives rise to spatiotemporal heterogeneity of dynamic subcellular mechanics in cardiac myocytes probed with atomic force microscopy Azeloglu, Evren U.; Costa, Kevin D. American Journal of Physiology-Heart and Circulatory Physiology, Vol. 298, Issue 3 https://doi.org/10.1152/ajpheart.00427.2009	journal	March 2010
Basic amino acid residue cluster within nuclear targeting sequence motif is essential for cytoplasmic plectin-vimentin network junctions. Nikolic, B.; Mac Nulty, E.; Mir, B. Journal of Cell Biology, Vol. 134, Issue 6 https://doi.org/10.1083/jcb.134.6.1455	journal	September 1996
Age-Dependent Changes in Microscale Stiffness and Mechanoresponses of Cells Zahn, Jasmin T.; Louban, Ilia; Jungbauer, Simon Small, Vol. 7, Issue 10 https://doi.org/10.1002/smll.201100146	journal	May 2011
Dynamic mechanical properties of the tissue-engineered matrix associated with individual chondrocytes Lee, BoBae; Han, Lin; Frank, Eliot H. Journal of Biomechanics, Vol. 43, Issue 3 https://doi.org/10.1016/j.jbiomech.2009.09.053	journal	February 2010
Compressive strains at physiological frequencies influence the metabolism of chondrocytes seeded in agarose Lee, David A.; Bader, Dan L. Journal of Orthopaedic Research, Vol. 15, Issue 2 https://doi.org/10.1002/jor.1100150205	journal	March 1997
Alteration of the distribution of intermediate filaments in PtK1 cells by acrylamide II: Effect on the organization of cytoplasmic organelles Eckert, Barry S. Cell Motility and the Cytoskeleton, Vol. 6, Issue 1 https://doi.org/10.1002/cm.970060104	journal	January 1986
Depth-dependent confined compression modulus of full-thickness bovine articular cartilage Schinagl, Robert M.; Gurskis, Donnell; Chen, Albert C. Journal of Orthopaedic Research, Vol. 15, Issue 4 https://doi.org/10.1002/jor.1100150404	journal	July 1997
Chondrocyte biosynthesis correlates with local tissue strain in statically compressed adult articular cartilage Wong, M.; Wuethrich, P.; Buschmann, M. D. Journal of Orthopaedic Research, Vol. 15, Issue 2 https://doi.org/10.1002/jor.1100150206	journal	March 1997
Determination of the Poisson's ratio of the cell: recovery properties of chondrocytes after release from complete micropipette aspiration Trickey, Wendy R.; Baaijens, Frank P. T.; Laursen, Tod A. Journal of Biomechanics, Vol. 39, Issue 1 https://doi.org/10.1016/j.jbiomech.2004.11.006	journal	January 2006
Loss of plectin causes epidermolysis bullosa with muscular dystrophy: cDNA cloning and genomic organization. McLean, W. H.; Pulkkinen, L.; Smith, F. J. Genes & Development, Vol. 10, Issue 14 https://doi.org/10.1101/gad.10.14.1724	journal	July 1996
Alterations in the Young’s modulus and volumetric properties of chondrocytes isolated from normal and osteoarthritic human cartilage Jones, Wendy R.; Ping Ting-Beall, H.; Lee, Greta M. Journal of Biomechanics, Vol. 32, Issue 2 https://doi.org/10.1016/S0021-9290(98)00166-3	journal	February 1999
Zonal changes in the three-dimensional morphology of the chondron under compression: The relationship among cellular, pericellular, and extracellular deformation in articular cartilage Choi, Jae Bong; Youn, Inchan; Cao, Li Journal of Biomechanics, Vol. 40, Issue 12 https://doi.org/10.1016/j.jbiomech.2007.01.009	journal	January 2007
Non-Hertzian Approach to Analyzing Mechanical Properties of Endothelial Cells Probed by Atomic Force Microscopy Costa, Kevin D.; Sim, Alan J.; Yin, Frank C-P. Journal of Biomechanical Engineering, Vol. 128, Issue 2 https://doi.org/10.1115/1.2165690	journal	November 2005
Imaging and Probing Cell Mechanical Properties With the Atomic Force Microscope Costa, Kevin D. Cell Imaging Techniques https://doi.org/10.1007/978-1-59259-993-6_17	book	January 2006
Biphasic Creep and Stress Relaxation of Articular Cartilage in Compression: Theory and Experiments Mow, V. C.; Kuei, S. C.; Lai, W. M. Journal of Biomechanical Engineering, Vol. 102, Issue 1 https://doi.org/10.1115/1.3138202	journal	February 1980
Effect of tissue maturity on cell viability in load-injured articular cartilage explants Levin, Adam S.; Chen, Chih-Tung Christopher; Torzilli, Peter A. Osteoarthritis and Cartilage, Vol. 13, Issue 6 https://doi.org/10.1016/j.joca.2005.01.006	journal	June 2005
Incongruity versus Instability in the Etiology of Posttraumatic Arthritis McKinley, Todd O.; Rudert, M. James; Koos, Daniel C. Clinical Orthopaedics & Related Research, Vol. 423 https://doi.org/10.1097/01.blo.0000131639.89143.26	journal	January 2004
Local stimulation of proteoglycan synthesis in articular cartilage explants by dynamic compression in vitro Parkkinen, Jyrki J.; Lammi, Mikko J.; Helminen, Heikki J. Journal of Orthopaedic Research, Vol. 10, Issue 5 https://doi.org/10.1002/jor.1100100503	journal	September 1992
Passive mechanical behavior of human neutrophils: effect of cytochalasin B Tsai, M. A.; Frank, R. S.; Waugh, R. E. Biophysical Journal, Vol. 66, Issue 6 https://doi.org/10.1016/s0006-3495(94)81012-4	journal	June 1994
Age-related changes in the morphology and deformational behavior of knee joint cartilage Hudelmaier, M.; Glaser, C.; Hohe, J. Arthritis & Rheumatism, Vol. 44, Issue 11 https://doi.org/10.1002/1529-0131(200111)44:11<2556::AID-ART436>3.0.CO;2-U	journal	January 2001
Two-dimensional strain fields on the cross-section of the bovine humeral head under contact loading Canal, Clare E.; Hung, Clark T.; Ateshian, Gerard A. Journal of Biomechanics, Vol. 41, Issue 15 https://doi.org/10.1016/j.jbiomech.2008.08.031	journal	November 2008
Plectin Transcript Diversity: Identification and Tissue Distribution of Variants with Distinct First Coding Exons and Rodless Isoforms Elliott, Candace E.; Becker, Bruno; Oehler, Susanne Genomics, Vol. 42, Issue 1 https://doi.org/10.1006/geno.1997.4724	journal	May 1997
Viscoelastic Cell Mechanics and Actin Remodelling Are Dependent on the Rate of Applied Pressure Pravincumar, Priyanka; Bader, Dan L.; Knight, Martin M. PLoS ONE, Vol. 7, Issue 9 https://doi.org/10.1371/journal.pone.0043938	journal	September 2012
Modelling of location- and time-dependent deformation of chondrocytes during cartilage loading Wu, J. Z.; Herzog, W.; Epstein, M. Journal of Biomechanics, Vol. 32, Issue 6 https://doi.org/10.1016/s0021-9290(99)00034-2	journal	June 1999
Identification of a 3Kbp Mechanoresponsive Promoter Region in the Human Cartilage Oligomeric Matrix Protein Gene Amanatullah, Derek F.; Lu, Jeffrey; Hecht, Jacqueline Tissue Engineering Part A, Vol. 18, Issue 17-18 https://doi.org/10.1089/ten.tea.2011.0497	journal	September 2012
A proposed mechanism of action of cytochalasin D on muscle actin Selden, Lynn A.; Gershman, Lewis C.; Estes, James E. Biochemical and Biophysical Research Communications, Vol. 95, Issue 4 https://doi.org/10.1016/s0006-291x(80)80115-x	journal	August 1980
Mechanisms of neurotoxicity related to selective disruption of microtubules and intermediate filaments Sager, Polly R.; Matheson, Dale W. Toxicology, Vol. 49, Issue 2-3 https://doi.org/10.1016/0300-483X(88)90034-0	journal	May 1988
Viscoelastic properties of zonal articular chondrocytes measured by atomic force microscopy Darling, E. M.; Zauscher, S.; Guilak, F. Osteoarthritis and Cartilage, Vol. 14, Issue 6 https://doi.org/10.1016/j.joca.2005.12.003	journal	June 2006
Tensile properties of human knee joint cartilage: I. Influence of ionic conditions, weight bearing, and fibrillation on the tensile modulus: TENSILE MODULUS OF HUMAN KNEE CARTILAGE Akizuki, Shaw; Mow, Van C.; Müller, Francisco Journal of Orthopaedic Research, Vol. 4, Issue 4 https://doi.org/10.1002/jor.1100040401	journal	January 1986
The effects of matrix compression on proteoglycan metabolism in articular cartilage explants Guilak, F.; Meyer, B.; Ratcliffe, A. Osteoarthritis and Cartilage, Vol. 2, Issue 2 https://doi.org/10.1016/s1063-4584(05)80059-7	journal	June 1994
Flow-independent viscoelastic properties of articular cartilage matrix Hayes, W. C.; Bodine, A. J. Journal of Biomechanics, Vol. 11, Issue 8-9 https://doi.org/10.1016/0021-9290(78)90075-1	journal	January 1978
In-situ measurements of chondrocyte deformation under transient loading Chahine, No; Hung, Ct; Ateshian, Ga European Cells and Materials, Vol. 13 https://doi.org/10.22203/eCM.v013a11	journal	May 2007
The Role of F-Actin in Hypo-Osmotically Induced Cell Volume Change and Calcium Signaling in Anulus Fibrosus Cells Pritchard, Scott; Guilak, Farshid Annals of Biomedical Engineering, Vol. 32, Issue 1 https://doi.org/10.1023/b:abme.0000007795.69001.35	journal	January 2004
Integrin???Interleukin-4 Mechanotransduction Pathways in Human Chondrocytes Salter, D. M.; Millward-Sadler, S. J.; Nuki, G. Clinical Orthopaedics and Related Research, Vol. 391 https://doi.org/10.1097/00003086-200110001-00006	journal	January 2001
Cytochalasins inhibit nuclei-induced actin polymerization by blocking filament elongation. Lin, D. C.; Tobin, K. D.; Grumet, M. Journal of Cell Biology, Vol. 84, Issue 2 https://doi.org/10.1083/jcb.84.2.455	journal	February 1980
The mechanical environment of the chondrocyte: a biphasic finite element model of cell–matrix interactions in articular cartilage Guilak, Farshid; Mow, Van C. Journal of Biomechanics, Vol. 33, Issue 12 https://doi.org/10.1016/S0021-9290(00)00105-6	journal	December 2000
Distribution and Ultrastructure of Plectin Arrays in Subclones of Rat Glioma C6 Cells Differing in Intermediate Filament Protein (Vimentin) Expression Foisner, Roland; Bohn, Wolfgang; Mannweiler, Klaus Journal of Structural Biology, Vol. 115, Issue 3 https://doi.org/10.1006/jsbi.1995.1055	journal	November 1995
Tensile Properties of Articular Cartilage Kempson, G. E.; Freeman, M. A. R.; Swanson, S. A. V. Nature, Vol. 220, Issue 5172 https://doi.org/10.1038/2201127b0	journal	December 1968
The role of the cytoskeleton in the viscoelastic properties of human articular chondrocytes Trickey, Wendy R.; Vail, T. Parker; Guilak, Farshid Journal of Orthopaedic Research, Vol. 22, Issue 1 https://doi.org/10.1016/S0736-0266(03)0150-5	journal	January 2004
Plectin contributes to mechanical properties of living cells Na, Sungsoo; Chowdhury, Farhan; Tay, Bernard American Journal of Physiology-Cell Physiology, Vol. 296, Issue 4 https://doi.org/10.1152/ajpcell.00604.2008	journal	April 2009
Commentary: Role of plectin in cytoskeleton organization and dynamics Wiche, Gerhard Journal of Cell Science, Vol. 111, Issue 17 https://doi.org/10.1242/jcs.111.17.2477	journal	September 1998

Cited By (7)

Experimental study of the difference in deformation between normal and pathological, renal and bladder, cells induced by acoustic radiation force Wang, Haibin; Qiao, Yupei; Liu, Jiehui European Biophysics Journal, Vol. 49, Issue 2 https://doi.org/10.1007/s00249-020-01422-3	journal	January 2020
Deformability of Human Mesenchymal Stem Cells Is Dependent on Vimentin Intermediate Filaments Sharma, Poonam; Bolten, Zachary T.; Wagner, Diane R. Annals of Biomedical Engineering, Vol. 45, Issue 5 https://doi.org/10.1007/s10439-016-1787-z	journal	January 2017
Effects of Osmolarity on the Spontaneous Calcium Signaling of In Situ Juvenile and Adult Articular Chondrocytes Zhou, Yilu; David, Michael A.; Chen, Xingyu Annals of Biomedical Engineering, Vol. 44, Issue 4 https://doi.org/10.1007/s10439-015-1406-4	journal	July 2015
The intrinsic stiffness of human trabecular meshwork cells increases with senescence Morgan, Joshua T.; Raghunathan, Vijay Krishna; Chang, Yow-Ren Oncotarget, Vol. 6, Issue 17 https://doi.org/10.18632/oncotarget.3798	journal	April 2015
Proteome Alterations in Equine Osteochondrotic Chondrocytes Chiaradia, Elisabetta; Pepe, Marco; Orvietani, Pier Luigi International Journal of Molecular Sciences, Vol. 20, Issue 24 https://doi.org/10.3390/ijms20246179	journal	December 2019
Effects of Inflammation on Multiscale Biomechanical Properties of Cartilaginous Cells and Tissues Nguyen, Q. T.; Jacobsen, T. D.; Chahine, N. O. ACS Biomaterials Science & Engineering, Vol. 3, Issue 11 https://doi.org/10.1021/acsbiomaterials.6b00671	journal	March 2017
Effects of mechanical stress on chondrocyte phenotype and chondrocyte extracellular matrix expression Liu, Qiang; Hu, Xiaoqing; Zhang, Xin Scientific Reports, Vol. 6, Issue 1 https://doi.org/10.1038/srep37268	journal	November 2016

Similar Records

Estradiol influences the mechanical properties of human fetal osteoblasts through cytoskeletal changes

Journal Article · Fri Jul 06 00:00:00 EDT 2012 · Biochemical and Biophysical Research Communications · OSTI ID:1627600

Muthukumaran, Padmalosini; Lim, Chwee Teck; Lee, Taeyong

AFM indentation study of breast cancer cells

Journal Article · Fri Oct 03 00:00:00 EDT 2008 · Biochemical and Biophysical Research Communications · OSTI ID:1627600

Li, Q S; Lee, G Y.H.; Ong, C N; +1 more

Quantification of stromal vascular cell mechanics with a linear cell monolayer rheometer

Journal Article · Thu Jan 15 00:00:00 EST 2015 · Journal of Rheology · OSTI ID:1627600

Fuller, Gerald G.; Shen, Wen-Jun; Khor, Victor K.; +1 more

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
Science & Technology - Other Topics

Title: Effect of Age and Cytoskeletal Elements on the Indentation-Dependent Mechanical Properties of Chondrocytes

Citation Formats

References (60)

Cited By (7)

Similar Records

Related Subjects