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Biomedical Optics Express

Biomedical Optics Express

  • Editor: Joseph A. Izatt
  • Vol. 2, Iss. 8 — Aug. 1, 2011
  • pp: 2307–2316

Structural changes in mixed Col I/Col V collagen gels probed by SHG microscopy: implications for probing stromal alterations in human breast cancer

Visar Ajeti, Oleg Nadiarnykh, Suzanne M. Ponik, Patricia J. Keely, Kevin W. Eliceiri, and Paul J. Campagnola  »View Author Affiliations

Biomedical Optics Express, Vol. 2, Issue 8, pp. 2307-2316 (2011)

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Second Harmonic Generation (SHG) microscopy has been previously used to describe the morphology of collagen in the extracellular matrix (ECM) in different stages of invasion in breast cancer. Here this concept is extended by using SHG to provide quantitative discrimination of self-assembled collagen gels, consisting of mixtures of type I (Col I) and type V (Col V) isoforms which serve as models of changes in the ECM during invasion in vivo. To investigate if SHG is sensitive to changes due to Col V incorporation into Col I fibrils, gels were prepared with 0-20% Col V with the balance consisting of Col I. Using the metrics of SHG intensity, fiber length, emission directionality, and depth-dependent intensities, we found similar responses for gels comprised of 100% Col I, and 95% Col I/5% Col V, where these metrics were all significantly different from those of the 80% Col I/20% Col V gels. Specifically, the gels of lower Col V content produce brighter SHG, are characterized by longer fibers, and have a higher forward/backward emission ratio. These attributes are all consistent with more highly organized collagen fibrils/fibers and are in agreement with previous TEM characterization as well as predictions based on phase matching considerations. These results suggest that SHG can be developed to discriminate Col I/Col V composition in tissues to characterize and follow breast cancer invasion.

© 2011 OSA

OCIS Codes
(180.6900) Microscopy : Three-dimensional microscopy
(190.2620) Nonlinear optics : Harmonic generation and mixing
(190.4180) Nonlinear optics : Multiphoton processes
(290.7050) Scattering : Turbid media
(170.6935) Medical optics and biotechnology : Tissue characterization

ToC Category:

Original Manuscript: May 31, 2011
Revised Manuscript: July 12, 2011
Manuscript Accepted: July 14, 2011
Published: July 20, 2011

Visar Ajeti, Oleg Nadiarnykh, Suzanne M. Ponik, Patricia J. Keely, Kevin W. Eliceiri, and Paul J. Campagnola, "Structural changes in mixed Col I/Col V collagen gels probed by SHG microscopy: implications for probing stromal alterations in human breast cancer," Biomed. Opt. Express 2, 2307-2316 (2011)

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  1. C. Ricciardelli and R. J. Rodgers, “Extracellular matrix of ovarian tumors,” Semin. Reprod. Med. 24(4), 270–282 (2006). [CrossRef] [PubMed]
  2. S. M. Pupa, S. Ménard, S. Forti, and E. Tagliabue, “New insights into the role of extracellular matrix during tumor onset and progression,” J. Cell. Physiol. 192(3), 259–267 (2002). [CrossRef] [PubMed]
  3. N. Théret, O. Musso, B. Turlin, D. Lotrian, P. Bioulac-Sage, J. P. Campion, K. Boudjéma, and B. Clément, “Increased extracellular matrix remodeling is associated with tumor progression in human hepatocellular carcinomas,” Hepatology 34(1), 82–88 (2001). [CrossRef] [PubMed]
  4. P. M. McGowan and M. J. Duffy, “Matrix metalloproteinase expression and outcome in patients with breast cancer: analysis of a published database,” Ann. Oncol. 19(9), 1566–1572 (2008). [CrossRef] [PubMed]
  5. J. M. Pellikainen, K. M. Ropponen, V. V. Kataja, J. K. Kellokoski, M. J. Eskelinen, and V. M. Kosma, “Expression of matrix metalloproteinase (MMP)-2 and MMP-9 in breast cancer with a special reference to activator protein-2, HER2, and prognosis,” Clin. Cancer Res. 10(22), 7621–7628 (2004). [CrossRef] [PubMed]
  6. S. Y. Huang, M. Van Arsdall, S. Tedjarati, M. McCarty, W. J. Wu, R. Langley, and I. J. Fidler, “Contributions of stromal metalloproteinase-9 to angiogenesis and growth of human ovarian carcinoma in mice,” J. Natl. Cancer Inst. 94(15), 1134–1142 (2002). [PubMed]
  7. H. Hugo, M. L. Ackland, T. Blick, M. G. Lawrence, J. A. Clements, E. D. Williams, and E. W. Thompson, “Epithelial--mesenchymal and mesenchymal--epithelial transitions in carcinoma progression,” J. Cell. Physiol. 213(2), 374–383 (2007). [CrossRef] [PubMed]
  8. T. Sørlie, C. M. Perou, R. Tibshirani, T. Aas, S. Geisler, H. Johnsen, T. Hastie, M. B. Eisen, M. van de Rijn, S. S. Jeffrey, T. Thorsen, H. Quist, J. C. Matese, P. O. Brown, D. Botstein, P. Eystein Lønning, and A. L. Børresen-Dale, “Gene expression patterns of breast carcinomas distinguish tumor subclasses with clinical implications,” Proc. Natl. Acad. Sci. U.S.A. 98(19), 10869–10874 (2001). [CrossRef] [PubMed]
  9. P. J. Campagnola and L. M. Loew, “Second-harmonic imaging microscopy for visualizing biomolecular arrays in cells, tissues and organisms,” Nat. Biotechnol. 21(11), 1356–1360 (2003). [CrossRef] [PubMed]
  10. Y. R. Shen, The Principles of Nonlinear Optics (John Wiley and Sons, 1984).
  11. S. V. Plotnikov, A. C. Millard, P. J. Campagnola, and W. A. Mohler, “Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres,” Biophys. J. 90(2), 693–703 (2006). [CrossRef] [PubMed]
  12. R. Lacomb, O. Nadiarnykh, S. S. Townsend, and P. J. Campagnola, “Phase Matching considerations in Second Harmonic Generation from tissues: Effects on emission directionality, conversion efficiency and observed morphology,” Opt. Commun. 281(7), 1823–1832 (2008). [CrossRef] [PubMed]
  13. R. M. Williams, W. R. Zipfel, and W. W. Webb, “Interpreting second-harmonic generation images of collagen I fibrils,” Biophys. J. 88(2), 1377–1386 (2005). [CrossRef] [PubMed]
  14. F. Légaré, C. Pfeffer, and B. R. Olsen, “The role of backscattering in SHG tissue imaging,” Biophys. J. 93(4), 1312–1320 (2007). [CrossRef] [PubMed]
  15. P. P. Provenzano, K. W. Eliceiri, J. M. Campbell, D. R. Inman, J. G. White, and P. J. Keely, “Collagen reorganization at the tumor-stromal interface facilitates local invasion,” BMC Med. 4(1), 38 (2006). [CrossRef] [PubMed]
  16. P. P. Provenzano, D. R. Inman, K. W. Eliceiri, J. G. Knittel, L. Yan, C. T. Rueden, J. G. White, and P. J. Keely, “Collagen density promotes mammary tumor initiation and progression,” BMC Med. 6(1), 11 (2008). [CrossRef] [PubMed]
  17. M. W. Conklin, J. C. Eickhoff, K. M. Riching, C. A. Pehlke, K. W. Eliceiri, P. P. Provenzano, A. Friedl, and P. J. Keely, “Aligned collagen is a prognostic signature for survival in human breast carcinoma,” Am. J. Pathol. 178(3), 1221–1232 (2011). [CrossRef] [PubMed]
  18. E. Sahai, J. Wyckoff, U. Philippar, J. E. Segall, F. Gertler, and J. Condeelis, “Simultaneous imaging of GFP, CFP and collagen in tumors in vivo using multiphoton microscopy,” BMC Biotechnol. 5(1), 14 (2005). [CrossRef] [PubMed]
  19. S. J. Lin, S. H. Jee, C. J. Kuo, R. J. Wu, W. C. Lin, J. S. Chen, Y. H. Liao, C. J. Hsu, T. F. Tsai, Y. F. Chen, and C. Y. Dong, “Discrimination of basal cell carcinoma from normal dermal stroma by quantitative multiphoton imaging,” Opt. Lett. 31(18), 2756–2758 (2006). [CrossRef] [PubMed]
  20. R. Cicchi, S. Sestini, V. De Giorgi, P. Carli, D. Massi, and F. S. Pavone, “Basal cell carcinoma imaging and characterization by multiple nonlinear microscopy techniques,” Biophys. J. , 157a–157a (2007).
  21. E. Brown, T. McKee, E. diTomaso, A. Pluen, B. Seed, Y. Boucher, and R. K. Jain, “Dynamic imaging of collagen and its modulation in tumors in vivo using second-harmonic generation,” Nat. Med. 9(6), 796–801 (2003). [CrossRef] [PubMed]
  22. O. Nadiarnykh, R. B. LaComb, M. A. Brewer, and P. J. Campagnola, “Alterations of the extracellular matrix in ovarian cancer studied by Second Harmonic Generation imaging microscopy,” BMC Cancer 10(1), 94 (2010). [CrossRef] [PubMed]
  23. S. H. Barsky, C. N. Rao, G. R. Grotendorst, and L. A. Liotta, “Increased content of Type V Collagen in desmoplasia of human breast carcinoma,” Am. J. Pathol. 108(3), 276–283 (1982). [PubMed]
  24. D. E. Birk, J. M. Fitch, J. P. Babiarz, K. J. Doane, and T. F. Linsenmayer, “Collagen fibrillogenesis in vitro: interaction of types I and V collagen regulates fibril diameter,” J. Cell Sci. 95(Pt 4), 649–657 (1990). [PubMed]
  25. P. J. Keely, A. M. Fong, M. M. Zutter, and S. A. Santoro, “Alteration of collagen-dependent adhesion, motility, and morphogenesis by the expression of antisense alpha 2 integrin mRNA in mammary cells,” J. Cell Sci. 108(Pt 2), 595–607 (1995). [PubMed]
  26. O. Nadiarnykh, R. B. Lacomb, P. J. Campagnola, and W. A. Mohler, “Coherent and incoherent SHG in fibrillar cellulose matrices,” Opt. Express 15(6), 3348–3360 (2007). [CrossRef] [PubMed]
  27. D. J. Hulmes, “Building collagen molecules, fibrils, and suprafibrillar structures,” J. Struct. Biol. 137(1-2), 2–10 (2002). [CrossRef] [PubMed]
  28. B. Eyden and M. Tzaphlidou, “Structural variations of collagen in normal and pathological tissues: role of electron microscopy,” Micron 32(3), 287–300 (2001). [CrossRef] [PubMed]
  29. D. E. Birk, “Type V collagen: heterotypic type I/V collagen interactions in the regulation of fibril assembly,” Micron 32(3), 223–237 (2001). [CrossRef] [PubMed]
  30. R. Lacomb, O. Nadiarnykh, and P. Campagnola, “Quantitative second harmonic generation imaging of the diseased state osteogenesis imperfecta: experiment and simulation,” Biophys. J. 94(11), 4504–4514 (2008). [CrossRef]
  31. R. LaComb, O. Nadiarnykh, S. Carey, and P. J. Campagnola, “Quantitative second harmonic generation imaging and modeling of the optical clearing mechanism in striated muscle and tendon,” J. Biomed. Opt. 13(2), 021109 (2008). [CrossRef] [PubMed]
  32. C. B. Raub, V. Suresh, T. Krasieva, J. Lyubovitsky, J. D. Mih, A. J. Putnam, B. J. Tromberg, and S. C. George, “Noninvasive assessment of collagen gel microstructure and mechanics using multiphoton microscopy,” Biophys. J. 92(6), 2212–2222 (2007). [CrossRef] [PubMed]
  33. F. Tiaho, G. Recher, and D. Rouède, “Estimation of helical angles of myosin and collagen by second harmonic generation imaging microscopy,” Opt. Express 15(19), 12286–12295 (2007). [CrossRef] [PubMed]
  34. P. J. Su, W. L. Chen, Y. F. Chen, and C. Y. Dong, “Determination of collagen nanostructure from second-order susceptibility tensor analysis,” Biophys. J. 100(8), 2053–2062 (2011). [CrossRef] [PubMed]
  35. C. Luparello, F. David, G. Campisi, and R. Sirchia, “T47-D cells and type V collagen: a model for the study of apoptotic gene expression by breast cancer cells,” Biol. Chem. 384(6), 965–975 (2003). [CrossRef] [PubMed]
  36. X. X. Han and E. Brown, “Measurement of the ratio of forward-propagating to back-propagating second harmonic signal using a single objective,” Opt. Express 18(10), 10538–10550 (2010). [CrossRef] [PubMed]

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