Longitudinal Study of Mammary Epithelial and Fibroblast Co-Cultures Using Optical Coherence Tomography Reveals Morphological Hallmarks of Pre-Malignancy

Chhetri, Raghav K.; Phillips, Zachary F.; Troester, Melissa A.; Oldenburg, Amy L.

doi:10.1371/journal.pone.0049148

Cited by 36 publications

(44 citation statements)

References 44 publications

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“…GFP expression was obtained through a lentiviral infection with puromycin as the selection antibiotic. MCF10A is a mammary epithelial cell line (MEC), which was maintained as previously published (47) in DMEM/F12 media. For the collagen study, collagen I (BD Biosciences) at 1−3 mg/mL final concentration was gelled in DMEM/F12 media.…”

Section: Methodsmentioning

confidence: 99%

“…For the collagen study, collagen I (BD Biosciences) at 1−3 mg/mL final concentration was gelled in DMEM/F12 media. For studies with cells, 1:1 collagen I: Matrigel (BD Biosciences) was prepared and seeded with RMF-GFP or MEC as described previously (47), with the final collagen I concentration at 1 mg/mL. RMF-GFP 3D cultures were seeded at three different concentrations (100, 300, and 500 cells per μL) and were incubated for 48, 72, and 96 h. MEC 3D cultures were seeded at 30 cells per μL and were incubated for 2 wk.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Probing biological nanotopology via diffusion of weakly constrained plasmonic nanorods with optical coherence tomography

Chhetri

Blackmon

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Biological materials exhibit complex nanotopology, i.e., a composite liquid and solid phase structure that is heterogeneous on the nanoscale. The diffusion of nanoparticles in nanotopological environments can elucidate biophysical changes associated with pathogenesis and disease progression. However, there is a lack of methods that characterize nanoprobe diffusion and translate easily to in vivo studies. Here, we demonstrate a method based on optical coherence tomography (OCT) to depth-resolve diffusion of plasmon-resonant gold nanorods (GNRs) that are weakly constrained by the biological tissue. By using GNRs that are on the size scale of the polymeric mesh, their Brownian motion is minimally hindered by intermittent collisions with local macromolecules. OCT depth-resolves the particleaveraged translational diffusion coefficient (D T ) of GNRs within each coherence volume, which is separable from the nonequilibrium motile activities of cells based on the unique polarized light-scattering properties of GNRs. We show how this enables minimally invasive imaging and monitoring of nanotopological changes in a variety of biological models, including extracellular matrix (ECM) remodeling as relevant to carcinogenesis, and dehydration of pulmonary mucus as relevant to cystic fibrosis. In 3D ECM models, D T of GNRs decreases with both increasing collagen concentration and cell density. Similarly, D T of GNRs is sensitive to human bronchial-epithelial mucus concentration over a physiologically relevant range. This novel method comprises a broad-based platform for studying heterogeneous nanotopology, as distinct from bulk viscoelasticity, in biological milieu.dynamic light scattering | plasmon resonance | nanoparticle diffusion | diffusion in mucus | diffusion in extracellular matrix B iological fluids (e.g., blood, mucus, saliva, synovial fluids) and soft solids [e.g., collagen, extracellular matrix (ECM), cytoskeleton] consist of a milieu of small molecules (making up the solvent) and large molecules (proteins and polymers making up the mesh or matrix) that are collectively responsible for their viscoelastic nature. Traditional rheological methods characterize the bulk viscoelastic properties of such biological media. However, nanoscopic objects, such as viruses and drugs that are smaller than the polymeric correlation length of the biological tissue network, encounter mechanical environments that are entirely different from that described by bulk viscoelasticity. For instance, at the nanoscale, mucus is a heterogeneous network of mucin fibers, nonmucin proteins, cell debris, lipids, DNA, actin filaments, and salts in a low-viscosity interstitial fluid (1). Similarly, tissue ECM, although soft solid-like in bulk, is an interconnected mesh of elongated protein fibers riddled with pores that are filled with low-viscosity interstitial fluids.Microrheological techniques based on the generalized StokesEinstein relation (2-5) are capable of converting the observed diffusion of probes of controlled hydrodynamic size into a mea...

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Probing biological nanotopology via diffusion of weakly constrained plasmonic nanorods with optical coherence tomography

Chhetri

Blackmon

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…We have previously shown how OCT of 3D mammary epithelial cell (MEC) organoids provides the ability to non-invasively and longitudinally monitor the evolution of organoids over weeks [12]. We have also shown how coupling temporal fluctuation analysis with polarization-sensitive OCT in these models allows for preferential image contrast of MEC organoids against a background of diffusive gold nanorod probes [13].…”

Section: Introductionmentioning

confidence: 99%

“…It is noted that the basal-like subtype is a type of breast cancer associated with a particularly poor prognosis [16]. We have recently shown that the morphology of basal-like MEC organoids changes with progression (normal versus pre-malignant) and depends upon the presence of fibroblasts in 3D co-culture [12]. Here we study the fluctuation spectra of MEC in the same co-culture model to understand how the apparent motility is affected by stromal cell signaling.…”

Section: Introductionmentioning

confidence: 99%

Inverse-power-law behavior of cellular motility reveals stromal–epithelial cell interactions in 3D co-culture by OCT fluctuation spectroscopy

Oldenburg¹,

Yu²,

Gilliss³

et al. 2015

Optica

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The progression of breast cancer is known to be affected by stromal cells within the local microenvironment. Here we study the effect of stromal fibroblasts on the in-place motions (motility) of mammary epithelial cells within organoids in 3D co-culture, inferred from the speckle fluctuation spectrum using optical coherence tomography (OCT). In contrast to Brownian motion, mammary cell motions exhibit an inverse power-law fluctuation spectrum. We introduce two complementary metrics for quantifying fluctuation spectra: the power-law exponent and a novel definition of the motility amplitude, both of which are signal- and position-independent. We find that the power-law exponent and motility amplitude are positively (p<0.001) and negatively (p<0.01) correlated with the density of stromal cells in 3D co-culture, respectively. We also show how the hyperspectral data can be visualized using these metrics to observe heterogeneity within organoids. This constitutes a simple and powerful tool for detecting and imaging cellular functional changes with OCT.

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“…The source of contrast for OCT is the scattering properties of the sample. OCT has been used in organoids to monitor cell movement and extracellular matrix invasion (31,32). Further, functional OCT techniques such as photothermal OCT, a method that detects heat generated by laser-light absorption by gold nanoparticles or other strong absorbers, can assess protein expression or the presence of functionalized nanoparticles (33).…”

Section: Optical Imaging Of Organoidsmentioning

confidence: 99%

Functional Optical Imaging of Primary Human Tumor Organoids: Development of a Personalized Drug Screen

2017

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Primary tumor organoids are a robust model of individual human cancers and present a unique platform for patient-specific drug testing. Optical imaging is uniquely suited to assess organoid function and behavior because of its subcellular resolution, penetration depth through the entire organoid, and functional endpoints. Specifically, optical metabolic imaging (OMI) is highly sensitive to drug response in organoids, and OMI in tumor organoids correlates with primary tumor drug response. Therefore, an OMI organoid drug screen could enable accurate testing of drug response for individualized cancer treatment. The objective of this perspective is to introduce OMI and tumor organoids to a general audience in order to foster the adoption of these techniques in diverse clinical and laboratory settings.

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Longitudinal Study of Mammary Epithelial and Fibroblast Co-Cultures Using Optical Coherence Tomography Reveals Morphological Hallmarks of Pre-Malignancy

Cited by 36 publications

References 44 publications

Probing biological nanotopology via diffusion of weakly constrained plasmonic nanorods with optical coherence tomography

Probing biological nanotopology via diffusion of weakly constrained plasmonic nanorods with optical coherence tomography

Inverse-power-law behavior of cellular motility reveals stromal–epithelial cell interactions in 3D co-culture by OCT fluctuation spectroscopy

Functional Optical Imaging of Primary Human Tumor Organoids: Development of a Personalized Drug Screen

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