The nanomechanical signature of breast cancer

Journal name:
Nature Nanotechnology
Volume:
7,
Pages:
757–765
Year published:
DOI:
doi:10.1038/nnano.2012.167
Received
Accepted
Published online

Abstract

Cancer initiation and progression follow complex molecular and structural changes in the extracellular matrix and cellular architecture of living tissue. However, it remains poorly understood how the transformation from health to malignancy alters the mechanical properties of cells within the tumour microenvironment. Here, we show using an indentation-type atomic force microscope (IT-AFM) that unadulterated human breast biopsies display distinct stiffness profiles. Correlative stiffness maps obtained on normal and benign tissues show uniform stiffness profiles that are characterized by a single distinct peak. In contrast, malignant tissues have a broad distribution resulting from tissue heterogeneity, with a prominent low-stiffness peak representative of cancer cells. Similar findings are seen in specific stages of breast cancer in MMTV-PyMT transgenic mice. Further evidence obtained from the lungs of mice with late-stage tumours shows that migration and metastatic spreading is correlated to the low stiffness of hypoxia-associated cancer cells. Overall, nanomechanical profiling by IT-AFM provides quantitative indicators in the clinical diagnostics of breast cancer with translational significance.

At a glance

Figures

  1. Nanomechanical signatures of human breast tissue.
    Figure 1: Nanomechanical signatures of human breast tissue.

    a, Top: schematic of an ultrasound-guided biopsy from a patient with a suspicious lesion. Middle: multiple stiffness maps (20 × 20 µm each) are recorded in a defined geometrical pattern across the entire specimen. Bottom: top view of an oriented, immobilized biopsy in Ringer's solution with the cantilever positioned for IT-AFM. Scale bar, 500 µm. b, Top left: stiffness distribution for normal mammary gland tissue is unimodal. Top right: post-AFM H&E-stained section reveals the terminal ductal lobular unit of a normal mammary gland fenced by interstitial connective tissue. Middle left: biopsy-wide histogram for a benign lesion reveals a unimodal but broader stiffness distribution with an increase in stiffness compared with the healthy biopsy. Middle right: H&E-stained section reveals extensive fibrotic stroma interspersed with fibroblasts typical for fibroadenoma. Bottom left: heterogeneous stiffness distribution with a characteristic soft peak for malignant tumour tissue is consistent with histopathology (bottom right), revealing an invasive breast carcinoma with infiltrating nests of cancer cells that have evoked a dense fibrous tissue response. Scale bar applies to all images, 50 µm.

  2. Stiffness varies from core to periphery in human cancer biopsies.
    Figure 2: Stiffness varies from core to periphery in human cancer biopsies.

    a, Post-AFM histological overview of the entire cancer biopsy with reference to the areas mapped in detail. Scale bar, 500 µm. b, Top: representative AFM stiffness map (24 × 24 pixels) of the core region visualizes pronounced softness within a narrow peak of specific stiffness values (middle). Bottom: the local histology shows that the core region is densely populated with cancer cells. c, Top: typical stiffness map (24 × 24 pixels) of the tumour periphery demonstrates stiff features. Middle: the corresponding stiffness distribution is broader and shifted towards stiffer values. Bottom: post-AFM histopathology reveals that the tumour periphery predominantly comprises fibrotic tissue. Scale bar, 50 µm (also applies to image in b).

  3. Correlating the nanomechanical response with tumour progression in MMTV-PyMT mice.
    Figure 3: Correlating the nanomechanical response with tumour progression in MMTV-PyMT mice.

    a, Left: stiffness values of normal ductal epithelium follow a uniform Gaussian distribution. Right: post-AFM H&E-stained section shows a non-lactating mammary gland with a duct surrounded by adipose tissue. b, Left: in the premalignant lesion, the contribution of stiffer features increases, as seen by a broader stiffness distribution with an indication of bimodality. Right: H&E-stained sections of the same tissue show extensive proliferation of epithelial cells and an increase of the surrounding stromal components. c, Left: in early cancer, the significant softening produces a characteristic peak that dominates the prominent bimodality of the stiffness distribution. Right: in the H&E section, cancer cells are delineated by stromal tissue. Scale bar in a applies to images in b and c, 50 µm.

  4. Correlating local nanomechanical properties and ECM structure in late cancer.
    Figure 4: Correlating local nanomechanical properties and ECM structure in late cancer.

    a, Consecutive stiffness maps (24 × 24 pixels) across the sample demonstrate a significant increase in stiffness and mechanical heterogeneity from core to periphery (left to right). b, Corresponding stiffness distributions change from the core to the periphery. c, IHC analysis (brown staining) reveals associated structural and morphological changes in collagen I (top) and laminin-1 (bottom) from core to periphery. Scale bar applies to all images, 50 µm.

  5. Stiffness profiles of primary tumour and lung metastasis reveal a common phenotype.
    Figure 5: Stiffness profiles of primary tumour and lung metastasis reveal a common phenotype.

    a, Left: representative AFM stiffness map (24 × 24 pixels) demonstrates nanomechanical heterogeneity among cancer cells in primary lesions. Right: the corresponding stiffness distribution from the map reveals two peaks representing distinct phenotypes of cancer cells, a softer one at 0.45 ± 0.15 kPa and another at 1.26 ± 0.43 kPa. b, Left: representative AFM stiffness map (24 × 24 pixels) of a metastatic lesion and the corresponding stiffness distribution (right) reveal a peak value that is almost identical to the softer peak detected at the primary tumour site (indicated by the red dashed line).

  6. Dissemination of hypoxic cancer cells increases with tumour progression.
    Figure 6: Dissemination of hypoxic cancer cells increases with tumour progression.

    Paraffin-embedded sections of mammary gland tissues of pimonidazole (hypoxyprobe)-treated MMTV-PyMT mice at different stages were immunolabelled. a, Hypoxia is absent in normal glandular tissue (left) and in premalignant lesions (middle). In contrast, pimonidazole-positive cells (brown staining) reveal central hypoxia in early cancer (right). b, In advanced cancer stages, hypoxic cells abundant in the core region (left) are streaming towards tumour blood vessels (middle) and have disseminated to the tumour periphery (right). Scale bar applies to all images, 50 µm.

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Author information

  1. These authors contributed equally to this work

    • Marko Loparic,
    • Christophe A. Monnier,
    • Ellen C. Obermann &
    • Rosanna Zanetti-Dallenbach

Affiliations

  1. Biozentrum and the Swiss Nanoscience Institute, University of Basel, 4056 Basel, Switzerland

    • Marija Plodinec,
    • Marko Loparic,
    • Christophe A. Monnier,
    • Philipp Oertle,
    • Janne T. Hyotyla &
    • Roderick Y. H. Lim
  2. Maurice E. Mueller Institute for Structural Biology, Biozentrum, University of Basel, 4056 Basel, Switzerland

    • Marija Plodinec,
    • Marko Loparic,
    • Ueli Aebi &
    • Cora-Ann Schoenenberger
  3. Institute of Pathology, University Hospital Basel, 4031 Basel, Switzerland

    • Ellen C. Obermann
  4. Department of Gynecology and Gynecological Oncology, University Hospital Basel, University of Basel, 4031 Basel, Switzerland

    • Rosanna Zanetti-Dallenbach
  5. Mechanisms of Cancer, Friedrich Miescher Institute for Biomedical Research, 4058 Basel, Switzerland

    • Mohamed Bentires-Alj

Contributions

M.P., R.Y.H.L. and C-A.S. conceived the study and designed experiments. M.P., M.L. and R.Y.H.L. developed all customized hardware and software solutions for AFM. M.P. and E.C.O. performed pathohistological and IHC analysis of human and murine tissues. R.Z.D. recruited patients and provided human biopsies. M.P., C.A.M. and P.O. performed AFM experiments. M.P., M.L, C.A.M., J.T.H., P.O. and R.Y.H.L. analysed AFM data. M.B-A. provided MMTV-PyMT mice and was involved in the analysis of murine tissues. M.P., U.A., R.Y.H.L. and C-A.S. wrote the paper. All authors discussed the results and commented on the manuscript.

Competing financial interests

The University of Basel has filed patents on the technology and intellectual property related to this work based on the inventions of M.P., M.L. and R.Y.H.L.

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