skip to main content


Title: Quantitative phase imaging of stromal prognostic markers in pancreatic ductal adenocarcinoma

New quantitative prognostic markers are needed for improved pancreatic ductal adenocarcinoma (PDAC) prognosis. Second harmonic generation microscopy has been used to show that collagen fiber alignment in PDAC is a negative prognostic factor. In this work, a series of PDAC and normal adjacent tissue (NAT) biopsies were imaged with spatial light interference microscopy (SLIM). Quantitative analysis performed on the biopsy SLIM images show that PDAC fiber structures have lower alignment per unit length, narrower width, and are longer than NAT controls. Importantly, fibrillar collagen in PDAC shows an inverse relationship between survival data and fiber width and length (p < 0.05).

 
more » « less
NSF-PAR ID:
10134346
Author(s) / Creator(s):
; ; ; ;
Publisher / Repository:
Optical Society of America
Date Published:
Journal Name:
Biomedical Optics Express
Volume:
11
Issue:
3
ISSN:
2156-7085
Page Range / eLocation ID:
Article No. 1354
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract

    Tissue biopsy evaluation in the clinic is in need of quantitative disease markers for diagnosis and, most importantly, prognosis. Among the new technologies, quantitative phase imaging (QPI) has demonstrated promise for histopathology because it reveals intrinsic tissue nanoarchitecture through the refractive index. However, a vast majority of past QPI investigations have relied on imaging unstained tissues, which disrupts the established specimen processing. Here we present color spatial light interference microscopy (cSLIM) as a new whole-slide imaging modality that performs interferometric imaging on stained tissue, with a color detector array. As a result, cSLIM yields in a single scan both the intrinsic tissue phase map and the standard color bright-field image, familiar to the pathologist. Our results on 196 breast cancer patients indicate that cSLIM can provide stain-independent prognostic information from the alignment of collagen fibers in the tumor microenvironment. The effects of staining on the tissue phase maps were corrected by a mathematical normalization. These characteristics are likely to reduce barriers to clinical translation for the new cSLIM technology.

     
    more » « less
  2. Contact guidance is a major physical cue that modulates cancer cell morphology and motility, and is directly linked to the prognosis of cancer patients. Under physiological conditions, particularly in the three-dimensional (3D) extracellular matrix (ECM), the disordered assembly of fibers presents a complex directional bias to the cells. It is unclear how cancer cells respond to these noncoherent contact guidance cues. Here we combine quantitative experiments, theoretical analysis, and computational modeling to study the morphological and migrational responses of breast cancer cells to 3D collagen ECM with varying degrees of fiber alignment. We quantify the strength of contact guidance using directional coherence of ECM fibers, and find that stronger contact guidance causes cells to polarize more strongly along the principal direction of the fibers. Interestingly, sensitivity to contact guidance is positively correlated with cell aspect ratio, with elongated cells responding more strongly to ECM alignment than rounded cells. Both experiments and simulations show that cell–ECM adhesions and actomyosin contractility modulate cell responses to contact guidance by inducing a population shift between rounded and elongated cells. We also find that cells rapidly change their morphology when navigating the ECM, and that ECM fiber coherence modulates cell transition rates between different morphological phenotypes. Taken together, we find that subcellular processes that integrate conflicting mechanical cues determine cell morphology, which predicts the polarization and migration dynamics of cancer cells in 3D ECM.

     
    more » « less
  3. Vitreous collagen structure plays an important role in ocular mechanics. However, capturing this structure with existing vitreous imaging methods is hindered by the loss of sample position and orientation, low resolution, or a small field of view. The objective of this study was to evaluate confocal reflectance microscopy as a solution to these limitations. Intrinsic reflectance avoids staining, and optical sectioning eliminates the requirement for thin sectioning, minimizing processing for optimal preservation of the natural structure. We developed a sample preparation and imaging strategy usingex vivogrossly sectioned porcine eyes. Imaging revealed a network of uniform diameter crossing fibers (1.1 ± 0.3 µm for a typical image) with generally poor alignment (alignment coefficient = 0.40 ± 0.21 for a typical image). To test the utility of our approach for detecting differences in fiber spatial distribution, we imaged eyes every 1 mm along an anterior-posterior axis originating at the limbus and quantified the number of fibers in each image. Fiber density was higher anteriorly near the vitreous base, regardless of the imaging plane. These data demonstrate that confocal reflectance microscopy addresses the previously unmet need for a robust, micron-scale technique to map features of collagen networksin situacross the vitreous.

     
    more » « less
  4. Abstract

    Tendon‐bone insertion is a functionally graded tissue, transitioning from 200 MPa tensile modulus at the tendon end to 20 GPa tensile modulus at the bone, across just a few hundred micrometers. In this study, we examine the porcine digital flexor tendon insertion tissue to provide a quantitative description of its collagen orientation and mineral concentration by using Fast Fourier Transform (FFT) based image analysis and mass spectrometry, respectively. Histological results revealed uniformity in global collagen orientation at all depths, indicative of mechanical anisotropy, although at mid‐depth, the highest fiber density, least amount of dispersion, and least cellular circularity were evident. Collagen orientation distribution obtained through 2D FFT of histological imaging data from fluorescent microscopy agreed with past measurements based on polarized light microscopy. Results revealed global fiber orientation across the tendon‐bone insertion to be preserved along direction of physiologic tension. Gradation in the fiber distribution orientation index across the insertion was reflective of a decrease in anisotropy from the tendon to the bone. We provided elemental maps across the fibrocartilage for its organic and inorganic constituents through time‐of‐flight secondary ion mass spectrometry (TOF‐SIMS). The apatite intensity distribution from the tendon to bone was shown to follow a linear trend, supporting past results based on Raman microprobe analysis. The merit of this study lies in the image‐based simplified approach to fiber distribution quantification and in the high spatial resolution of the compositional analysis. In conjunction with the mechanical properties of the insertion tissue, fiber, and mineral distribution results for the insertion from this may potentially be incorporated into the development of a structural constitutive approach toward computational modeling. Characterizing the properties of the native insertion tissue would provide the microstructural basis for developing biomimetic scaffolds to recreate the graded morphology of a fibrocartilaginous insertion. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3050–3058, 2017.

     
    more » « less
  5. Abstract

    Pancreatic ductal adenocarcinoma (PDAC), one of the most aggressive types of cancer, is characterized by aberrant activity of oncogenic KRAS. A nuclease-hypersensitive GC-rich region in KRAS promoter can fold into a four-stranded DNA secondary structure called G-quadruplex (G4), known to regulate KRAS expression. However, the factors that regulate stable G4 formation in the genome and KRAS expression in PDAC are largely unknown. Here, we show that APE1 (apurinic/apyrimidinic endonuclease 1), a multifunctional DNA repair enzyme, is a G4-binding protein, and loss of APE1 abrogates the formation of stable G4 structures in cells. Recombinant APE1 binds to KRAS promoter G4 structure with high affinity and promotes G4 folding in vitro. Knockdown of APE1 reduces MAZ transcription factor loading onto the KRAS promoter, thus reducing KRAS expression in PDAC cells. Moreover, downregulation of APE1 sensitizes PDAC cells to chemotherapeutic drugs in vitro and in vivo. We also demonstrate that PDAC patients’ tissue samples have elevated levels of both APE1 and G4 DNA. Our findings unravel a critical role of APE1 in regulating stable G4 formation and KRAS expression in PDAC and highlight G4 structures as genomic features with potential application as a novel prognostic marker and therapeutic target in PDAC.

     
    more » « less