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Combined MR, fluorescence and histology imaging strategy in a human breast tumor xenograft model
Abstract
Applications of molecular imaging in cancer and other diseases frequently require the combination of in vivo imaging modalities, such as MR and optical imaging, with ex vivo optical, fluorescence, histology and immunohistochemical imaging to investigate and relate molecular and biological processes to imaging parameters within the same region of interest. We have developed a multimodal image reconstruction and fusion framework that accurately combines in vivo MRI and MRSI, ex vivo brightfield and fluorescence microscopic imaging and ex vivo histology imaging. Ex vivo brightfield microscopic imaging was used as an intermediate modality to facilitate the ultimate link between ex vivo histology and in vivo MRI/MRSI. Tissue sectioning necessary for optical and histology imaging required the generation of a three-dimensional reconstruction module for two-dimensional ex vivo optical and histology imaging data. We developed an external fiducial marker-based three-dimensional reconstruction method, which was able to fuse optical brightfield and fluorescence with histology imaging data. The registration of the three-dimensional tumor shape was pursued to combine in vivo MRI/MRSI and ex vivo optical brightfield and fluorescence imaging data. This registration strategy was applied to in vivo MRI/MRSI, ex vivo optical brightfield/fluorescence and histology imaging datasets obtained from human breast tumor models. Three-dimensional human breast tumor datasets were successfully reconstructed and fused with this platform. Copyright © 2012 John Wiley & Sons, Ltd.
... A large number of modalities exist to image the breast, but until recently there have been few modalities or techniques to make quantitative comparisons between scales. 17,19,[38][39][40][41][42][43] The field's relative youth means that the quantitative links between the three scales are not well defined, leaving large Table 1 Three imaging scales for multiscale breast imaging: the cellular microscale, the tissue mesoscale, and the organ macroscale. These definitions are based on usage in prior literature. ...
... They developed workflows to register to magnetic resonance imaging (MRI) and widefield fluorescence. 40,103 There is much room to build on these and other similar methods to make 3D QHP an accessible and valuable tool for research. 18 One final limitation that should be considered is that QHP is inherently destructive to tissue and can only be done ex vivo. ...
... 103,[210][211][212] The penetration depth limit mostly restricts them to 2D imaging, but 3D imaging is possible ex vivo with serial sectioning of biopsies to obtain adjacent tissue slices and reconstructing those images. 40,103 They are being investigated for clinical use with IMA, where QMIB studies use histology for validation. 87,213 In addition, recent studies have shown improved resolution in some applications. ...
Breast cancer is the most common cancer among women worldwide and ranks second in terms of overall cancer deaths. One of the difficulties associated with treating breast cancer is that it is a heterogeneous disease with variations in benign and pathologic tissue composition, which contributes to disease development, progression, and treatment response. Many of these phenotypes are uncharacterized and their presence is difficult to detect, in part due to the sparsity of methods to correlate information between the cellular microscale and the whole-breast macroscale. Quantitative multiscale imaging of the breast is an emerging field concerned with the development of imaging technology that can characterize anatomic, functional, and molecular information across different resolutions and fields of view. It involves a diverse collection of imaging modalities, which touch large sections of the breast imaging research community. Prospective studies have shown promising results, but there are several challenges, ranging from basic physics and engineering to data processing and quantification, that must be met to bring the field to maturity. This paper presents some of the challenges that investigators face, reviews currently used multiscale imaging methods for preclinical imaging, and discusses the potential of these methods for clinical breast imaging.
... MALDI was capable of measuring ions of a wide mass range and to identify some of them using an MS/MS approach, whereas SIMS was used to indicate the localization of fragment ions. MALDI and SIMS were also applied by Chughtai et al. (2012) to show the distribution of electrolytes, lipids, and peptides in a mouse model of rheumatoid arthritis. SIMS measurements indicated the localization of potassium, calcium, cholesterol and several other lipids in various areas of the knee joint of a mouse, whereas MALDI was used to reveal the localization of different lipids and tryptic peptides from five different proteins. ...
... As an example of this approach, we can consider the study of Chughtai et al. (2012). The group proposed a 1:1 mixture of cresyl violet and Ponceau S, since both substances are clearly visible under a bright field microscope and can be detected in both the negative and positive ion modes by IMS ionization techniques. ...
... During the study, they revealed several proteins with altered abundance in response to infection in an organ-specific manner. Jiang et al. ( 2012) developed multimodal molecular image reconstruction, registration, and fusion platform to combine MRI, fluorescence, IMS, and histology imaging of a human breast tumor model, using fiducial markers. Oetjen et al. ( 2013) described 3D MALDI-imaging an experimental pipeline with PAXgene* fixation compatible with MRI (see Fig. 9). ...
Imaging Mass Spectrometry (IMS) is strengthening its position as a valuable analytical tool. It has unique ability to identify structures and to unravel molecular changes that occur in the precisely defined part of the sample. These unique features open new possibilities in the field of various aspects of biological research. In this review we briefly discuss the main imaging mass spectrometry techniques, as well as the nature of biological samples and molecules, which might be analyzed by such methodology. Moreover, a novel approach, where different analytical techniques might be combined with the results of IMS study, is emphasized and discussed. With such a fast development of IMS and related methods, we can foresee the promising future of this technique. © 2015 Wiley Periodicals, Inc. Mass Spec Rev.
© 2015 Wiley Periodicals, Inc.
... When tumors reached a volume of approximately 500-mm 3 , mice were sacrificed and tumors were removed. Each tumor was embedded into a gelatin block prepared using 15-mm×15-mm×5-mm cryomolds (Sakura Finetek, Torrance, CA, USA), 10 % gelatin, cooled to 30 °C in order to prevent tissue degradation, and three cresyl violet fiducial markers, which were injected inside the gelatin block next to the tumor as previously described121314 . This block was sectioned into serial 2- mm thick fresh tumor sections using an acrylic adjustable tissue slicer (12-mm depth up to 25-mm width; Braintree Scientific, Inc., Braintree, MA, USA) and tissue slicer blades (Braintree Scientific, Inc.). ...
... Since the spatial resolution of an MSI image is also much lower than that of an optical bright field/fluorescence image and the noise level of an MSI image is much higher than that of an optical bright field/fluorescence image, a nonlinear twodimensional median filtering method was applied to reduce the noise and preserve the tumor edge in the summed-up MSI image. A combined fiducial marker and tumor boundary-based image registration method [13, 14] was applied to both the summed-up MSI image and the optical bright field/fluorescence image. The tumor region was masked out by the active shape model in all images [13, 14]. ...
... A combined fiducial marker and tumor boundary-based image registration method [13, 14] was applied to both the summed-up MSI image and the optical bright field/fluorescence image. The tumor region was masked out by the active shape model in all images [13, 14]. The scattering distribution of the registered MSI ion intensity and optical fluorescence signal were plotted against each other, and the resulting Pearson's correlation coefficient was calculated to investigate the linearity between registered MSI ion image and optical fluorescence image. ...
Mass spectrometric imaging (MSI) in combination with electrospray mass spectrometry (ESI-MS) is a powerful technique for visualization and identification of a variety of different biomolecules directly from thin tissue sections. As commonly used tools for molecular reporting, fluorescent proteins are molecular reporter tools that have enabled the elucidation of a multitude of biological pathways and processes. To combine these two approaches, we have performed targeted MS analysis and MALDI-MSI visualization of a tandem dimer (td)Tomato red fluorescent protein, which was expressed exclusively in the hypoxic regions of a breast tumor xenograft model. For the first time, a fluorescent protein has been visualized by both optical microscopy and MALDI-MSI. Visualization of tdTomato by MALDI-MSI directly from breast tumor tissue sections will allow us to simultaneously detect and subsequently identify novel molecules present in hypoxic regions of the tumor. MS and MALDI-MSI of fluorescent proteins, as exemplified in our study, is useful for studies in which the advantages of MS and MSI will benefit from the combination with molecular approaches that use fluorescent proteins as reporters.
Figure
... Cresyl violet acetate and Ponceau S were prepared in 10% warm (37°C) gelatin at a concentration of 10 mg/ml, mixed in a 1:1 ratio, and injected into 10% gelatin blocks next to the tumor tissue. Each block was sectioned into serial 2 mm-thick fresh tumor sections using an acrylic adjustable tissue slicer (12 mm depth up to 25 mm width, Braintree Scientifi c, Inc.; Braintree, MA), and tissue slicer blades (Braintree Scientifi c, Inc.) as previously described ( 43,44 ). These serial fresh tumor xenograft sections were each placed on individual microscope slides (Fisherbrand catalog number 12-550-34, Fisher Scientifi c; Pittsburgh, PA), and stored in an ice box containing ice on the bottom, with the slides located on a perforated plate at approximately 1-cm above the ice to minimize tissue degradation ( 43,44 ). ...
... Each block was sectioned into serial 2 mm-thick fresh tumor sections using an acrylic adjustable tissue slicer (12 mm depth up to 25 mm width, Braintree Scientifi c, Inc.; Braintree, MA), and tissue slicer blades (Braintree Scientifi c, Inc.) as previously described ( 43,44 ). These serial fresh tumor xenograft sections were each placed on individual microscope slides (Fisherbrand catalog number 12-550-34, Fisher Scientifi c; Pittsburgh, PA), and stored in an ice box containing ice on the bottom, with the slides located on a perforated plate at approximately 1-cm above the ice to minimize tissue degradation ( 43,44 ). These fresh sections were imaged by of such heterogeneity is tissue hypoxia, a state of low oxygen tension (pO 2 values р 2.5 mmHg) observed in many solid tumors ( 2 ). ...
... The remaining tumor regions were normoxic. Ion images of each individual m/z were overlaid with the corresponding optical images and coregistered by using the positions of fi ducial markers and tumor boundaries as previously described ( 43,44 ). The tumor region was masked by applying an active-shape model to the registered optical bright-fi eld images ( 48 ). ...
The lipid compositions of different breast tumor microenvironments are largely unknown due to limitations in lipid imaging techniques. Imaging lipid distributions would enhance our understanding of processes occurring inside growing tumors such as cancer proliferation and metastasis. Recent developments in MALDI mass spectrometry imaging (MSI) enable rapid and specific detection of lipids directly from thin tissue sections. In this study, we performed multimodal imaging of acylcarnitines, phosphatidylcholines (PC), a lysophosphatidylcholine (LPC) and a sphingomyelin (SM) from different microenvironments of breast tumor xenograft models, which carried tdTomato red fluorescent protein as a hypoxia-response element driven reporter gene. The MSI molecular lipid images revealed spatially heterogeneous lipid distributions within tumor tissue. Five of the most abundant lipid species, namely PC(16:0/16:0), PC(16:0/18:1), PC(18:1/18:1) PC(18:0/18:1) and SM(d18:1/16:0) sodium adduct were localized in viable tumor regions, while PC(18:0/22:1) and LPC(16:0/0:0) were detected in necrotic tumor regions. We identified a heterogeneous distribution of palmitoyl- and stearoylcarnitine, which mostly colocalized with hypoxic tumor regions. For the first time, we have applied a multimodal imaging approach that has combined optical imaging and MALDI-MSI with ion mobility separation to spatially localize and structurally identify acylcarnitines and a variety of lipid species present in breast tumor xenograft models.
... Thin sections (1-20µm) were cut in Samavati et al. (2011);Zhan et al. (2007); Burton et al. (2006). Thick sections (>20µm) were cut in Jiang et al. (2013); Osechinskiy and Kruggel (2010); Mazaheri et al. (2010);Singh et al. (2008). ...
... Edwards et al. (2005) used histology to identify the tumour boundaries in oral cancer patients with better accuracy in order to enable precise PET-guided resection. Jiang et al. (2013) combined in vivo MRI/MRSI, ex vivo brightfield/fluorescence microscopic imaging, and histology to study human breast cancer. Seeley et al. (2014) studied secondary breast cancer in the bone using diffusion weighted MRI, Matrix-Assisted Laser Desorption/Ionisation Imaging Mass Spectrometry and histology in order to observe changes caused by tumour cells in the bone at the protein level. ...
Histology permits the observation of otherwise invisible structures of the internal topography of a specimen. Although it enables the investigation of tissues at a cellular level, it is invasive and breaks topology due to cutting. Three-dimensional (3D) reconstruction was thus introduced to overcome the limitations of single-section studies in a dimensional scope. 3D reconstruction finds its roots in embryology, where it enabled the visualisation of spatial relationships of developing systems and organs, and extended to biomedicine, where the observation of individual, stained sections provided only partial understanding of normal and abnormal tissues. However, despite bringing visual awareness, recovering realistic reconstructions is elusive without prior knowledge about the tissue shape.
3D medical imaging made such structural ground truths available. In addition, combining non-invasive imaging with histology unveiled invaluable opportunities to relate macroscopic information to the underlying microscopic properties of tissues through the establishment of spatial correspondences; image registration is one technique that permits the automation of such a process and we describe reconstruction methods that rely on it. It is thereby possible to recover the original topology of histology and lost relationships, gain insight into what affects the signals used to construct medical images (and characterise them), or build high resolution anatomical atlases.
This paper reviews almost three decades of methods for 3D histology reconstruction from serial sections, used in the study of many different types of tissue. We first summarise the process that produces digitised sections from a tissue specimen in order to understand the peculiarity of the data, the associated artefacts and some possible ways to minimise them. We then describe methods for 3D histology reconstruction with and without the help of 3D medical imaging, along with methods of validation and some applications. We finally attempt to identify the trends and challenges that the field is facing, many of which are derived from the cross-disciplinary nature of the problem as it involves the collaboration between physicists, histopathologists, computer scientists and physicians.
... We have previously performed a comprehensive analysis of the combined in vivo 1 H MRSI and ex vivo optical imaging data obtained from the MDA-MB-231-HRE-tdTomato breast tumor model and showed a higher concentration of noninvasively detected tCho and mobile lipid droplets that colocalized with the tdTomato-fluorescing hypoxic regions, which indicated that hypoxia can up-regulate tCho and lipid CH3 levels in this breast tumor model. [16][17][18] Mass spectrometric imaging (MSI) enables the characterization and profiling of a plethora of molecules and, at the same time, reveals their individual spatial localizations in cancer tissue sections, without the need for labeling of these molecules. 19 Hence, MSI provides a unique way to make snapshots of molecular distributions in a high-throughput manner with high spectral and spatial resolution. ...
... shows an overview of our experimental and data analysis workflow including sample preparation of breast tumor tissue, 3D MALDI-MSI, and data processing in human MDA-MB-231-HRE-tdTomato breast tumor xenografts.18,34,35 MALDI-MSI was performed using the 10 μm sections on ITO slides. ...
Hypoxic areas are a common feature of rapidly growing malignant tumors and their metastases, and are typically spatially heterogeneous. Hypoxia has a strong impact on tumor cell biology and contributes to tumor progression in multiple ways. To date, only a few molecular key players in tumor hypoxia, such as for example hypoxia-inducible factor-1 (HIF-1), have been discovered. The distribution of biomolecules is frequently heterogeneous in the tumor volume, and may be driven by hypoxia and HIF-1α. Understanding the spatially heterogeneous hypoxic response of tumors is critical. Mass spectrometric imaging (MSI) provides a unique way of imaging biomolecular distributions in tissue sections with high spectral and spatial resolution. In this paper, breast tumor xenografts grown from MDA-MB-231-HRE-tdTomato cells, with a red fluorescent tdTomato protein construct under the control of a hypoxia response element (HRE)-containing promoter driven by HIF-1α, were used to detect the spatial distribution of hypoxic regions. We elucidated the 3D spatial relationship between hypoxic regions and the localization of small molecules, metabolites, lipids, and proteins by using principal component analysis - linear discriminant analysis (PCA-LDA) on 3D rendered MSI volume data from MDA-MB-231-HRE-tdTomato breast tumor xenografts. In this study we identified hypoxia-regulated proteins active in several distinct pathways such as glucose metabolism, regulation of actin cytoskeleton, protein folding, translation/ribosome, splicesome, the PI3K-Akt signaling pathway, hemoglobin chaperone, protein processing in endoplasmic reticulum, detoxification of reactive oxygen species, aurora B signaling/apoptotic execution phase, the RAS signaling pathway, the FAS signaling pathway/caspase cascade in apoptosis and telomere stress induced senescence. In parallel we also identified co-localization of hypoxic regions and various lipid species such as PC(16:0/18:1), PC(16:0/18:2), PC(18:0/18:1), PC(18:1/18:1), PC(18:1/18:2), PC(16:1/18:4), PC(18:0/20:3), PC(16:0/22:1), among others. Our findings shed light on the biomolecular composition of hypoxic tumor regions, which may be responsible for a given tumor's resistance to radiation or chemotherapy.
... 3D-MSI also opens the possibility of correlating the volumetric molecular information with data from other (3D) imaging modalities [76]. A common example is the combination of 3D MALDI-MSI data with standard histology techniques. ...
Mass spectrometry imaging (MSI) enables the visualization of molecular distributions on complex surfaces. It has been extensively used in the field of biomedical research to investigate healthy and diseased tissues. Most of the MSI studies are conducted in a 2D fashion where only a single slice of the full sample volume is investigated. However, biological processes occur within a tissue volume and would ideally be investigated as a whole to gain a more comprehensive understanding of the spatial and molecular complexity of biological samples such as tissues and cells. Mass spectrometry imaging has therefore been expanded to the 3D realm whereby molecular distributions within a 3D sample can be visualized. The benefit of investigating volumetric data has led to a quick rise in the application of single-sample 3D-MSI investigations. Several experimental and data analysis aspects need to be considered to perform successful 3D-MSI studies. In this review, we discuss these aspects as well as ongoing developments that enable 3D-MSI to be routinely applied to multi-sample studies.
... In developed countries, such as the USA, the imaging investigations used, as Michael K. Pinkert [19][20][21] highlighted in the paper on quantified imaging, are microcomputed tomography (microCT), fluoroscopic US, very high frequency US, fluoroscopic molecular tomography, with a very high diagnosis accuracy, sensitivity and specificity. This suggests investing both in the preclinical, clinical and operational-medical field, which should also be the case for the country to invest in order to decrease mortality. ...
Breast cancer is a condition with the highest incidence of all neoplasms and a frequent cause of death. Due to increased incidence and mortality, this disease motivates healthcare professionals to redirect efforts to develop effective strategies for secondary prophylaxis. Imagistic investigations play an important role both in detecting lesions and in post-therapeutic evolutionary follow-up. The objective of the paper is to study cases of premalignant and malignant tumors, with a view to their imagistic identification confirmed in terms of histopathology, to highlight the accuracy of the imagistic examination as an important factor in the diagnosis and adaptation of an appropriate therapeutic attitude. The study was performed on a batch of 768 patients admitted to the Department of Surgery III, "Prof. Dr. Alexandru Trestioreanu" Institute of Oncology, Bucharest, Romania. The classical examined hypothesis is local examination, mammography, ultrasound, with its variations, and histopathological (HP) confirmation, either by thick-needle biopsy puncture and∕or tumor excision. By correlating with HP examination of the imagistic representation of the lesion, we can show the importance or limitation of each imagistic investigation, but especially its usefulness in the choice of therapeutic behavior. Breast cancer screening using classical techniques currently requires implementation of modern techniques to diagnose this disease.
... 5, 6, 7, 8 The former approach requires complex computational models that are particularly susceptible to complications due to data formatting, imaging artifacts, and changes caused by tissue processing. 4 On the other hand, the latter approach introduces the possibility of contaminating or otherwise harming the tissue itself. 9 The approach described here improves the transition between modalities through the use of postmortem MRI to bridge the gap between in vivo MRI and histology. Postmortem MRI provides three-dimensional (3D) images of the brain at higher resolution than can be achieved in vivo and furthermore provides the data needed for producing a morphologically accurate model of the brain surface. ...
Magnetic resonance imaging (MRI) allows for the delineation between normal and abnormal tissue on a macroscopic scale, sampling an entire tissue volume three-dimensionally. While MRI is an extremely sensitive tool for detecting tissue abnormalities, association of signal changes with an underlying pathological process is usually not straightforward. In the central nervous system, for example, inflammation, demyelination, axonal damage, gliosis, and neuronal death may all induce similar findings on MRI. As such, interpretation of MRI scans depends on the context, and radiological-histopathological correlation is therefore of the utmost importance. Unfortunately, traditional pathological sectioning of brain tissue is often imprecise and inconsistent, thus complicating the comparison between histology sections and MRI. This article presents novel methodology for accurately sectioning primate brain tissues and thus allowing precise matching between histology and MRI. The detailed protocol described in this article will assist investigators in applying this method, which relies on the creation of 3D printed brain slicers. Slightly modified, it can be easily implemented for brains of other species, including humans.
In this paper, we present an easy-to-follow procedure for the analysis of tissue sections from 3D cell cultures (spheroids) by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) and laser scanning confocal microscopy (LSCM). MALDI MSI was chosen to detect the distribution of the drug of interest, while fluorescence immunohistochemistry (IHC) followed by LSCM was used to localize the cells featuring specific markers of viability, proliferation, apoptosis and metastasis. The overlay of the mass spectrometry (MS) and IHC spheroid images, typically without any morphological features, required fiducial-based coregistration. The MALDI MSI protocol was optimized in terms of fiducial composition and antigen epitope preservation to allow MALDI MSI to be performed and directly followed by IHC analysis on exactly the same spheroid section. Once MS and IHC images were coregistered, the quantification of the MS and IHC signals was performed by an algorithm evaluating signal intensities along equidistant layers from the spheroid boundary to its center. This accurate colocalization of MS and IHC signals showed limited penetration of the clinically tested drug perifosine into spheroids during a 24 h period, revealing the fraction of proliferating and promigratory/proinvasive cells present in the perifosine-free areas, decrease of their abundance in the perifosine-positive regions and distinguishing between apoptosis resulting from hypoxia/nutrient deprivation and drug exposure.
Magnetic resonance spectroscopy (MRS) or spectroscopic imaging (MRSI) enables the detection of metabolites, amino acids, and lipids, among other biomolecules, in tumors of live mouse models of cancer. Tumor-bearing mice are anesthetized by breathing isoflurane in a magnetic resonance (MR) scanner dedicated to small animal MR. Here we describe the overall setup and steps for measuring ¹H and ³¹P MRS and ¹H MRSI of orthotopic breast tumor models in mice with surface coils. This protocol can be adapted to the use of volume coils to measure ¹H and ³¹P MRS(I) of tumor models that grow inside the body. We address issues of animal handling, setting up the measurement, measurement options, and data analysis.
Although tumor hypoxia is associated with tumor aggressiveness and resistance to cancer treatment, many details of hypoxia-induced changes in tumors remain to be elucidated. Mass spectrometry imaging (MSI) is a technique that is well suited to study the biomolecular composition of specific tissue regions, such as hypoxic tumor regions. Here, we investigate the use of pimo-nidazole as exogenous hypoxia marker for matrix-assisted laser desorption/ionization (MALDI) MSI. In hypoxic cells, pimoni-dazole is reduced and forms reactive products that bind to thiol groups in proteins, peptides and amino acids. We show that a reductively activated pimonidazole metabolite can be imaged by MALDI-MSI in a breast tumor xenograft model. Immunohistochemical detection of pimonidazole adducts on adjacent tissue sections confirmed that this metabolite is localized to hypoxic tissue regions. We used this metabolite to image hypoxic tissue regions and their associated lipid and small molecule distributions with MALDI-MSI. We identified a heterogeneous distribution of 1-methylnicotinamide and acetylcarnitine, which mostly co-localized with hypoxic tumor regions. As pimonidazole is a widely used immunohistochemical marker of tissue hypoxia, it is likely that the presented direct MALDI-MSI approach is also applicable to other tissues from pimonidazole-injected animals or humans.
The objective of our study was to determine the relative accuracy of mammography, sonography, and MRI in predicting residual tumor after neoadjuvant chemotherapy for breast cancer as compared with the gold standards of physical examination and pathology.
Forty-one women with stage IIB-III palpable breast cancer were prospectively enrolled in a study investigating the effects of sequential single-agent chemotherapy (doxorubicin followed by paclitaxel or vice versa) on tumor imaging. The study cohort consisted of the first 31 patients (age range, 31-65 years; mean, 45 years) who completed the protocol. All underwent physical examination, mammography, sonography, and MRI before and after receiving each neoadjuvant chemotherapeutic drug. Imaging studies were reviewed by two radiologists using conventional lexicons for lesion analysis, and the findings were compared with clinical response and pathology results.
Complete, partial, and stable clinical response as defined by clinical examination was seen in 15, 14, and two of the 31 patients, respectively. Agreement rates about the degree of response were 32%, 48%, and 55%, respectively, for mammography, sonography, and MRI compared with clinical evaluation and did not differ statistically. Agreement about the rate of response as measured by clinical examination, mammography, sonography, and MRI compared with the gold standard (pathology) was 19%, 26%, 35%, and 71%, respectively. Of the four, MRI agreed with the gold standard significantly more often (p < 0.002 for all three paired comparisons with MRI). When there was disagreement with the gold standard, none of the four exhibited a significant tendency to either under- or overestimate.
MRI appears to provide the best correlation with pathology-better than physical examination, mammography, and sonography-in patients undergoing neoadjuvant chemotherapy. However, MRI may overestimate (6%) or underestimate (23%) residual disease in approximately 29% of the patients (95% confidence interval, 14-48%).
Active Appearance Models (AAMs) have been shown to be useful for interpreting images of deformable objects. Here we place the AAM matching algorithm in a statistical framework, allowing extra constraints to be applied. This enables the models to be combined with other methods of object location. We demonstrate how user interaction can be used to guide the search and give results of experiments showing the effect of constraints on the performance of model matching.
The CD44 transmembrane glycoproteins play multifaceted roles in tumor progression and metastasis. CD44 expression has also been associated with stem-like breast cancer cells. Hypoxia commonly occurs in tumors and is a major cause of radiation and chemo-resistance. Hypoxia is known to inhibit differentiation and facilitates invasion and metastasis. Here we have investigated the effect of hypoxia on CD44 and two of its isoforms in MDA-MB-231 and SUM-149 triple negative human breast cancer cells and MDA-MB-231 tumors using imaging and molecular characterization.
The roles of hypoxia and hypoxia inducible factor (HIF) in regulating the expression of CD44 and its variant isoforms (CD44v6, CD44v7/8) were investigated in human breast cancer cells, by quantitative real-time polymerase chain reaction (qRT-PCR) to determine mRNA levels, and fluorescence associated cell sorting (FACS) to determine cell surface expression of CD44, under normoxic and hypoxic conditions. In vivo imaging studies with tumor xenografts derived from MDA-MD-231 cells engineered to express tdTomato red fluorescence protein under regulation of hypoxia response elements identified co-localization between hypoxic fluorescent regions and increased concentration of (125)I-radiolabeled CD44 antibody.
Our data identified HIF-1α as a regulator of CD44 that increased the number of CD44 molecules and the percentage of CD44 positive cells expressing variant exons v6 and v7/8 in breast cancer cells under hypoxic conditions. Data from these cell studies were further supported by in vivo observations that hypoxic tumor regions contained cells with a higher concentration of CD44 expression.
We apply the Delaunay Tessellation Field Estimator (DTFE) to reconstruct and analyse the matter distribution and cosmic velocity flows in the local Universe on the basis of the PSCz galaxy survey. The prime objective of this study is the production of optimal resolution 3D maps of the volume-weighted velocity and density fields throughout the nearby universe, the basis for a detailed study of the structure and dynamics of the cosmic web at each level probed by underlying galaxy sample. Fully volume-covering 3D maps of the density and (volume-weighted) velocity fields in the cosmic vicinity, out to a distance of 150 h−1 Mpc, are presented. Based on the Voronoi and Delaunay tessellation defined by the spatial galaxy sample, DTFE involves the estimate of density values on the basis of the volume of the related Delaunay tetrahedra and the subsequent use of the Delaunay tessellation as natural multidimensional (linear) interpolation grid for the corresponding density and velocity fields throughout the sample volume. The linearized model of the spatial galaxy distribution and the corresponding peculiar velocities of the PSCz galaxy sample, produced by Branchini et al., forms the input sample for the DTFE study. The DTFE maps reproduce the high-density supercluster regions in optimal detail, both their internal structure as well as their elongated or flattened shape. The corresponding velocity flows trace the bulk and shear flows marking the region extending from the Pisces–Perseus supercluster, via the Local Superclusters, towards the Hydra–Centaurus and the Shapley concentration. The most outstanding and unique feature of the DTFE maps is the sharply defined radial outflow regions in and around underdense voids, marking the dynamical importance of voids in the local Universe. The maximum expansion rate of voids defines a sharp cut-off in the DTFE velocity divergence probability distribution function. We found that on the basis of this cut-off DTFE manages to consistently reproduce the value of Ωm≈ 0.35 underlying the linearized velocity data set.
This paper describes building models which represent the appearance of an object (in particular, a face) as seen from two or more di erent viewpoints simultaneously. A small number of 2D linear statistical models are sucient to capture the shape and appearance of a face from a wide range of viewpoints. Given multiple images of the same face we can learn a coupled model describing the relationship between the frontal appearance and the pro le of a face. This relationship can be used to predict...
Phosphocholine (PC) and total choline (tCho) are increased in malignant breast tumors. In this study, we combined magnetic resonance spectroscopic imaging (MRSI), mass spectrometry (MS) imaging, and pathologic assessment of corresponding tumor sections to investigate the localization of choline metabolites and cations in viable versus necrotic tumor regions in the nonmetastatic MCF-7 and the highly metastatic MDA-MB-231 breast cancer xenograft models. In vivo three-dimensional MRSI showed that high tCho levels, consisting of free choline (Cho), PC, and glycerophosphocholine (GPC), displayed a heterogeneous spatial distribution in the tumor. MS imaging performed on tumor sections detected the spatial distributions of individual PC, Cho, and GPC, as well as sodium (Na+) and potassium (K+), among many others. PC and Cho intensity were increased in viable compared with necrotic regions of MDA-MB-231 tumors, but relatively homogeneously distributed in MCF-7 tumors. Such behavior may be related to the role of PC and PC-related enzymes, such as choline kinase, choline transporters, and others, in malignant tumor growth. Na+ and K+ colocalized in the necrotic tumor areas of MDA-MB-231 tumors, whereas in MCF-7 tumors, Na+ was detected in necrotic and K+ in viable tumor regions. This may be attributed to differential Na+/K+ pump functions and K+ channel expressions. Principal component analysis of the MS imaging data clearly identified different tumor microenvironmental regions by their distinct molecular signatures. This molecular information allowed us to differentiate between distinct tumor regions and tumor types, which may, in the future, prove clinically useful in the pathologic assessment of breast cancers.
Purpose
A NIR tomography system that combines frequency domain (FD) and continuous wave (CW) measurements was used to image normal and malignant breast tissues.
Methods
FD acquisitions were confined to wavelengths less than 850 nm because of detector limitations, whereas light from longer wavelengths (up to 948 nm) was measured in CW mode with CCD‐coupled spectrometer detection. The two data sets were combined and processed in a single spectrally constrained reconstruction to map concentrations of hemoglobin, water, and lipid, as well as scattering parameters in the breast.
Results
Chromophore concentrations were imaged in the breasts of nine asymptomatic volunteers to evaluate their intrasubject and intersubject variability. Normal subject data showed physiologically expected trends. Images from three cancer patients indicate that the added CW data is critical to recovering the expected increases in water and decreases in lipid content within malignancies. Contrasts of 1.5 to twofold in hemoglobin and water values were found in cancers.
Conclusions
In vivo breast imaging with instrumentation that combines FD and CW NIR data acquisition in a single spectral reconstruction produces more accurate hemoglobin, water, and lipid results relative to FD data alone.
Metastasis continues to be one of the major causes of mortality from prostate cancer. Because human malignant cell lines metastasize more readily from orthotopic sites than from heterotopic sites, to identify metastasis-permissive tumor microenvironments, we used noninvasive imaging to compare the in vivo vascular, metabolic, and physiologic characteristics of a human prostate cancer xenograft implanted orthotopically in the prostate or s.c. in the flank. Hypoxia was detected in these xenografts by placing an enhanced green fluorescence protein optical reporter under the control of a hypoxia response element. A multiparametric analysis of hypoxia, extracellular pH, vascularization, and metabolism provided a characterization of environments that are permissive for metastasis to occur. We found that orthotopic tumors, which metastasized more easily, were characterized by higher vascular volume, permeability, and total choline and a more acidic extracellular pH. Interestingly, metastatic deposits in the lymph nodes as well as cancer cells in ascites fluid were found to be hypoxic, explaining, in part, the refractory nature of metastatic disease. These results also provide the basis for clinically translatable noninvasive imaging markers for predicting metastatic risk in prostate cancer.
In vivo knowledge of the spatial distribution of viable, necrotic, and hypoxic areas can provide prognostic information about the risk of developing metastases and regional radiation sensitivity and may be used potentially for localized dose escalation in radiation treatment. In this study, multimodality in vivo magnetic resonance imaging (MRI) and positron emission tomography (PET) imaging using stereotactic fiduciary markers in the Dunning R3327-AT prostate tumor were performed, focusing on the relationship between dynamic contrast-enhanced (DCE) MRI using Magnevist (Gd-DTPA) and dynamic (18)F-fluoromisonidazole ((18)F-Fmiso) PET. The noninvasive measurements were verified using tumor tissue sections stained for hematoxylin/eosin and pimonidazole. To further validate the relationship between (18)F-Fmiso and pimonidazole uptake, (18)F digital autoradiography was performed on a selected tumor and compared with the corresponding pimonidazole-stained slices. The comparison of Akep values (kep = rate constant of movement of Gd-DTPA between the interstitial space and plasma and A = amplitude in the two-compartment model (Hoffmann U, Brix G, Knopp MV, Hess T and Lorenz WJ (1995). Magn Reson Med 33, 506-514) derived from DCE-MRI studies and from early (18)F-Fmiso uptake PET studies showed that tumor vasculature is a major determinant of early (18)F-Fmiso uptake. A negative correlation between the spatial map of Akep and the slope map of late (last 1 hour of the dynamic PET scan) (18)F-Fmiso uptake was observed. The relationships between DCE-MRI and hematoxylin/eosin slices and between (18)F-Fmiso PET and pimonidazole slices confirm the validity of MRI/PET measurements to image the tumor microenvironment and to identify regions of tumor necrosis, hypoxia, and well-perfused tissue.
The objective of this work was to develop and then validate a stereotactic fiduciary marker system for tumor xenografts in rodents which could be used to co-register magnetic resonance imaging (MRI), PET, tissue histology, autoradiography, and measurements from physiologic probes. A Teflon fiduciary template has been designed which allows the precise insertion of small hollow Teflon rods (0.71 mm diameter) into a tumor. These rods can be visualized by MRI and PET as well as by histology and autoradiography on tissue sections. The methodology has been applied and tested on a rigid phantom, on tissue phantom material, and finally on tumor bearing mice. Image registration has been performed between the MRI and PET images for the rigid Teflon phantom and among MRI, digitized microscopy images of tissue histology, and autoradiograms for both tissue phantom and tumor-bearing mice. A registration accuracy, expressed as the average Euclidean distance between the centers of three fiduciary markers among the registered image sets, of 0.2 +/- 0.06 mm was achieved between MRI and microPET image sets of a rigid Teflon phantom. The fiduciary template allows digitized tissue sections to be co-registered with three-dimensional MRI images with an average accuracy of 0.21 and 0.25 mm for the tissue phantoms and tumor xenografts, respectively. Between histology and autoradiograms, it was 0.19 and 0.21 mm for tissue phantoms and tumor xenografts, respectively. The fiduciary marker system provides a coordinate system with which to correlate information from multiple image types, on a voxel-by-voxel basis, with sub-millimeter accuracy--even among imaging modalities with widely disparate spatial resolution and in the absence of identifiable anatomic landmarks.
The development of therapeutic inhibitors of key signaling pathways has been hampered by the inability to assess the effect of a drug on its target in the patient. 17-allylaminogeldanamycin (17-AAG) is the first Hsp90 inhibitor to be tested in a clinical trial. It causes the degradation of HER2 and other Hsp90 targets, and has antitumor activity in preclinical models. We have developed a method for imaging the inhibition of Hsp90 by 17-AAG. We labeled an F(ab')2 fragment of the anti-HER2 antibody Herceptin with 68Ga, a positron emitter, which allows the sequential positron-emission tomographic imaging of HER2 expression. We have used this method to quantify as a function of time the loss and recovery of HER2 induced by 17-AAG in animal tumors. This approach allows noninvasive imaging of the pharmacodynamics of a targeted drug and will facilitate the rational design of combination therapy based on target inhibition.
Numerous investigations have shown that both tissue and cell distribution profiles of anticancer drugs can be controlled by their entrapment in submicronic colloidal systems (nanoparticles). The rationale behind this approach is to increase antitumor efficacy, while reducing systemic side-effects. This review provides an update of tumor targeting with conventional or long-circulating nanoparticles. The in vivo fate of these systems, after intravascular or tumoral administration, is discussed, as well as the mechanism involved in tumor regression. Nanoparticles are also of benefit for the selective delivery of oligonucleotides to tumor cells. Moreover, certain types of nanoparticles showed some interesting capacity to reverse MDR resistance, which is a major problem in chemotherapy. The first experiments, aiming to decorate nanoparticles with molecular ligand for 'active' targeting of cancerous cells, are also discussed here. The last part of this review focus on the application of nanoparticles in imaging for cancer diagnosis.
Model-based vision is firmly established as a robust approach to recognizing and locating known rigid objects in the presence of noise, clutter, and occlusion. It is more problematic to apply model-based methods to images of objects whose appearance can vary, though a number of approaches based on the use of flexible templates have been proposed. The problem with existing methods is that they sacrifice model specificity in order to accommodate variability, thereby compromising robustness during image interpretation. We argue that a model should only be able to deform in ways characteristic of the class of objects it represents. We describe a method for building models by learning patterns of variability from a training set of correctly annotated images. These models can be used for image search in an iterative refinement algorithm analogous to that employed by Active Contour Models (Snakes). The key difference is that our Active Shape Models can only deform to fit the data in ways consistent with the training set. We show several practical examples where we have built such models and used them to locate partially occluded objects in noisy, cluttered images.
This paper is concerned with the development of entropy-based registration criteria for automated 3D multi-modality medical image alignment. In this application where misalignment can be large with respect to the imaged field of view, invariance to overlap statistics is an important consideration. Current entropy measures are reviewed and a normalised measure is proposed which is simply the ratio of the sum of the marginal entropies and the joint entropy. The effect of changing overlap on current entropy measures and this normalised measure are compared using a simple image model and experiments on clinical image data. Results indicate that the normalised entropy measure provides significantly improved behaviour over a range of imaged fields of view.
Volume rendering and surface rendering are the two main methodologies currently available for visualizing three-dimensional medical image information. The relative merits of these methodologies in portraying medically relevant information are not yet established scientifically and hence the claims of superiority are controversial at present. This article summarizes the position statement of the author on this subject at a panel moderated by him at this conference.
Born in Sidney, Ohio, on May 6, 1929, Paul Lauterbur (Fig. 1) received his B.S. in chemistry in 1951 from the Case Institute of Technology in Cleveland. After two years of U.S. military service where he began research on the phenomenon of nuclear magnetic resonance, he went on to receive a Ph.D. in chemistry from the University of Pittsburgh in 1962. Since 1963 he has been affiliated with the State University of New York at Stony Brook, most recently as Adjunct University Professor. Since 1985 he has been a full-time faculty member at the University of Illinois at Urbana-Champaign and now holds appointments there in the Department of Medical Information Science of the College of Medicine, the Department of Chemistry, the Biophysics Program of the Department of Physiology and Biophysics, the Bioengineering Program of the Department of Electrical and Computer Engineering, and the Center for Advanced Study. In addition to this, he is Director of the Biomedical Magnetic Resonance Laboratory there. Dr. Lauterbur also serves as a Professor in the University of Illinois at Chicago College of Medicine and, to date, has 124 publications to his credit. He is a Fellow of the American Physical Society and the American Association for the Advancement of Science and a member of the National Academy of Sciences. His many awards include the Gold Medal of the Society of Magnetic Resonance in Medicine (1982), the American Physical Society Prize in Biological Physics (1983), The Franklin Institute Howard N. Potts Medal (1984), The Michelson-Morley Award, Case Western Reserve University (1984). The Albert Lasker Clinical Research Award (1984), The Kosar Memorial Award, Society of Photographic Scientists and Engineers (1985), The Charles F. Kettering Prize, General Motors Cancer Research Foundation (1985); The Gairdner Foundation International Award (1985); The Harvey Prize in Science and Technology, Technion (1986); The Roentgen Medal (1987); The Institute of Electrical and Electronics Engineers Medal of Honor (1987); The National Medal of Science (1987); The Gold Medal of the Radiological Society of North America (1987); and the National Medal of Technology (1988).
MARSHALL R. URIST, M.D.
(C) Lippincott-Raven Publishers.
Multimodal imaging represents one of the current trends in the development of biophotonics imaging technologies. This paper briefly reviews current multimodal biophotonics imaging platforms in macroscopic, microscopic, and mesoscopic (or millimeter) scales. We also present a combined optical coherence tomography and line-scan fluorescence laminar optical tomography system for co-registered structural and molecular imaging with millimeter-scale imaging depth. Experimental results using a capillary phantom filled with the fluorescence dye Cy5.5 and a human breast cancer xenograft model are presented.
It is shown that transverse relaxation measurements obtained from CPMG echo trains are valid even in the presence of strong, static magnetic field gradients. In the context of in situ measurements for water or oil exploration, low magnetic fields and short echo spacings are utilized to minimize diffusional effects. Under these conditions, it is shown that, for T1 = T2, the inverse Laplace transformation of the echo train is essentially independent of field homogeneity. For T1 ≠ T2, the error in determining T2 does not exceed ∼12%, even for high T1/T2 ratios. In most porous media, T1/T2 is less than 3, in which case the error is below 8%. Analytical expressions for the echo amplitudes including relaxation are derived based on the density-matrix formalism. We define recursion relations that give the density matrix at echo i by a simple multiplication of the density matrix at echo i − 1 with a set of operators describing the evolution between consecutive echoes. The echo intensity is shown to be a function of the ratio between the radiofrequency strength and the receiver bandwidth. The optimal signal-to-noise ratio is obtained when this ratio is unity. The paper provides the theoretical framework for interpreting data obtained in situ by a modern NMR logging instrument. Furthermore, the results are directly applicable to magnetic resonance imaging.
We consider the problem of aligning histological sections for 3D reconstruction and analysis. The method we propose is based on a block-matching strategy that allows us to compute local displacements between the sections. We then collect these local measures to estimate a rigid transformation. Our emphasis is on the necessity to use a robust approach for this estimation step. The process is integrated within a multi-scale scheme to improve both accuracy and computation time. We prove experimentally that we can reach sub-pixel accuracy and we show some results of aligning histological sections from a rat's brain and a rhesus monkey's brain.
Volumetric rendering speaks volumes for data 20 orders of magnitude apart—from human anatomy to neuroanatomy, and from electrostatic charges of macromolecules to failure analysis of manufactured parts.
By performing registration of preoperative multiprotocol in vivo magnetic resonance (MR) images of the prostate with corresponding whole-mount histology (WMH) sections from postoperative radical prostatectomy specimens, an accurate estimate of the spatial extent of prostate cancer (CaP) on in vivo MR imaging (MRI) can be retrospectively established. This could allow for definition of quantitative image-based disease signatures and lead to development of classifiers for disease detection on multiprotocol in vivo MRI. Automated registration of MR and WMH images of the prostate is complicated by dissimilar image intensities, acquisition artifacts, and nonlinear shape differences.
The authors present a method for automated elastic registration of multiprotocol in vivo MRI and WMH sections of the prostate. The method, multiattribute combined mutual information (MACMI), leverages all available multiprotocol image data to drive image registration using a multivariate formulation of mutual information.
Elastic registration using the multivariate MI formulation is demonstrated for 150 corresponding sets of prostate images from 25 patient studies with T2-weighted and dynamic-contrast enhanced MRI and 85 image sets from 15 studies with an additional functional apparent diffusion coefficient MRI series. Qualitative results of MACMI evaluation via visual inspection suggest that an accurate delineation of CaP extent on MRI is obtained. Results of quantitative evaluation on 150 clinical and 20 synthetic image sets indicate improved registration accuracy using MACMI compared to conventional pairwise mutual information-based approaches.
The authors' approach to the registration of in vivo multiprotocol MRI and ex vivo WMH of the prostate using MACMI is unique, in that (1) information from all available image protocols is utilized to drive the registration with histology, (2) no additional, intermediate ex vivo radiology or gross histology images need be obtained in addition to the routinely acquired in vivo MRI series, and (3) no corresponding anatomical landmarks are required to be identified manually or automatically on the images.
Some of the most exciting advances in molecular-functional imaging of cancer are occurring at the interface between chemistry and imaging. Several of these advances have occurred through the development of novel imaging probes that report on molecular pathways, the tumor micro-environment and the response of tumors to treatment; as well as through novel image-guided platforms such as nanoparticles and nanovesicles that deliver therapeutic agents against specific targets and pathways. Cancer cells have a remarkable ability to evade destruction despite the armamentarium of drugs currently available. While these drugs can destroy cancer cells, normal tissue toxicity is a major limiting factor, a problem further compounded by poor drug delivery. One major challenge for chemistry continues to be to eliminate cancer cells without damaging normal tissues. Here we have selected examples of MRI and optical imaging, to demonstrate how integrating imaging with novel probes can facilitate the successful treatment of this multifaceted disease.
There is increasing interest in the registration of 3-D histopathology with 3-D in vivo imaging, for example, to validate tumor boundary delineation for targeted radiation cancer therapy. However, accurate correlation is compromised by tissue distortion induced by histopathological processing. Reference landmarks that are visible in both data sets are required. In this study, two iridescent acrylic paints, "Bronze" (containing iron oxide coated mica particles) and "Stainless Steel" (containing iron, chromium, and nickel), were evaluated for creating MRI-visible and histology-visible fiducial markers at 7 T, where resolution is more similar to histology, but artifacts are accentuated. Furthermore, a straight-line paint-track fiducial method was developed to assist in registration and 3-D histopathology reconstruction.
First, the paints were injected into ex vivo porcine tissue samples, which were MR imaged prefixation and postfixation, and subsequently prepared for hematoxylin and eosin staining to verify stability through histopathological processing. Second, the severity of marker susceptibility artifacts produced was compared while using spin-echo and gradient-echo MRI pulse sequences. Finally, multiple paint tracks were injected prefixation through an ex vivo canine prostate sample to validate the potential for line-based registration between MR images of prefixation and postfixation tissue and whole mount histology slides.
The Stainless Steel paint produced excessive susceptibility artifacts and image distortion, while the Bronze paint created stable and appropriate markers in MRI and histology. The Bronze paint produced artifacts approximately three times larger in gradient-echo than in spin-echo MR images. Finally, the paint-track fiducials were visible in the prefixation and postfixation MRI and on whole mount histology.
The Bronze iridescent acrylic paint is appropriate for fiducial marker creation in MRI at 7 T. The straight-line paint-track fiducials may assist 3-D histopathology reconstruction and can provide important information on the deformation effects of fixation, and hence may improve registration accuracy.
Multinuclear magnetic resonance spectroscopy (MRS) has evolved into a highly versatile, noninvasive technique that is finding multiple applications in medical diagnosis, monitoring therapy, and research. The increasing interface of chemistry with the fields of MRSI and molecular biology, and the resulting advances in theranostic contrast agent design, are providing new advances in the applications of MRS for molecular characterization and molecular-targeted medicine, especially in cancer. MRS or MRSI detection of the 1H or 31P MRS signals of the chemical structures of glucose, lactate, N-acetyl aspartate, and citrate can assist in the diagnosis of cancer. MRS detection of 13C-labeled metabolites can be performed following administration of suitable 13C-labeled substrates in cancer cells and solid tumors to study glycolysis or other metabolic pathways such as choline metabolism. High-resolution magic angle spinning (HR MAS) 1H MRS is a relatively new technology for examining intact biological tissue, such as biopsy specimens, at high spectral resolution.
The purposes of this study were to present magnetic resonance imaging (MRI) findings in patients with breast cancer soon after the excisional biopsy and before definitive surgery and to assess the diagnostic accuracy of MRI for prediction of residual cancer.
Breast MRI was performed on 51 patients with breast cancer diagnosed by excisional biopsy. The morphologic type of postexcisional site was categorized into regular thin rim enhancement (P1), peripheral nodular enhancement (P2), satellite nodule within 2 mm from the cavity margin (P3), and irregularly thickened wall enhancement (P4). Enhancement pattern was categorized into "washout," "plateau," and "progressive" enhancement. All patients underwent definitive surgery after MRI. Magnetic resonance imaging findings were correlated with the pathologic findings.
The sensitivity, specificity, accuracy, positive predictive value and negative predictive value of MRI for prediction of residual cancer were 92.1%, 69.2%, 78.4%, 88.6%, 56.3%, respectively.
MRI is a sensitive method for prediction of residual cancer after excisional biopsy.
To apply an intensity-based nonrigid registration algorithm to MRI-guided prostate brachytherapy clinical data and to assess its accuracy.
A nonrigid registration of preoperative MRI to intraoperative MRI images was carried out in 16 cases using a Basis-Spline algorithm in a retrospective manner. The registration was assessed qualitatively by experts' visual inspection and quantitatively by measuring the Dice similarity coefficient (DSC) for total gland (TG), central gland (CG), and peripheral zone (PZ), the mutual information (MI) metric, and the fiducial registration error (FRE) between corresponding anatomical landmarks for both the nonrigid and a rigid registration method.
All 16 cases were successfully registered in less than 5 min. After the nonrigid registration, DSC values for TG, CG, PZ were 0.91, 0.89, 0.79, respectively, the MI metric was -0.19 +/- 0.07 and FRE presented a value of 2.3 +/- 1.8 mm. All the metrics were significantly better than in the case of rigid registration, as determined by one-sided t-tests.
The intensity-based nonrigid registration method using clinical data was demonstrated to be feasible and showed statistically improved metrics when compare to only rigid registration. The method is a valuable tool to integrate pre- and intraoperative images for brachytherapy.
Inflammation and neovascularization play critical roles in the stability of atherosclerotic plaques. Whole-body quantitative assessment of these plaque features may improve patient risk-stratification for life-threatening thromboembolic events and direct appropriate intervention. In this report, we determined the utility of the MR contrast agent gadofluorine-M (GdF) for staging plaque stability and compared this to the conventional agent Gd-DTPA.
Five control and 7 atherosclerotic rabbits were sequentially imaged after administration of Gd-DTPA (0.2 mmol/kg) and GdF (0.1 mmol/kg) using a T(1)-weighted pulse sequence on a 3-T MRI scanner. Diseased aortic wall could be distinguished from normal wall based on wall-to-muscle contrast-to-noise values after GdF administration. RAM-11 (macrophages) and CD-31 (endothelial cells) immunostaining of MR-matched histological sections revealed that GdF accumulation was related to the degree of inflammation at the surface of plaques and the extent of core neovascularization. Importantly, an MR measure of GdF accumulation at both 1 and 24 hours after injection but not Gd-DTPA at peak enhancement was shown to correlate with a quantitative histological morphology index related to these 2 plaque features.
GdF-enhanced MRI of atherosclerotic plaques allows noninvasive quantitative information about plaque composition to be acquired at multiple time points after injection (within 1 and up to 24 hours after injection). This dramatically widens the imaging window for assessing plaque stability that is currently attainable with clinically approved MR agents, therefore opening the possibility of whole-body (including coronary) detection of unstable plaques in the future and potentially improved mitigation of cataclysmic cardiovascular events.
Despite diversity in genetic events in oncogenesis, cancer cells exhibit a common set of functional characteristics. Otto Warburg discovered that cancer cells have consistently higher rates of glycolysis than normal cells. The underlying mechanisms leading to the Warburg phenomenon include mitochondrial changes, upregulation of rate-limiting enzymes/proteins in glycolysis and intracellular pH regulation, hypoxia-induced switch to anaerobic metabolism, and metabolic reprogramming after loss of p53 function. The regulation of energy metabolism can be traced to a "triad" of transcription factors: c-MYC, HIF-1 and p53. Oncogenetic changes involve a nonrandom set of gene deletions, amplifications and mutations, and many oncogenes and tumor suppressor genes cluster along the signaling pathways that regulate c-MYC, HIF-1 and p53. Glycolysis in cancer cells has clinical implications in cancer diagnosis, treatment and interaction with diabetes mellitus. Many drugs targeting energy metabolism are in development. Future advances in technology may bring about transcriptome and metabolome-guided chemotherapy.
Filner and Varner3 showed that GA induced isolated aleurone tissue to synthesize hydrolytic enzymes de novo. Experiments with actinomycin D showed that RNA synthesis appeared to be necessary for both enzyme synthesis and secretion*.Fig.
A method for obtaining the three-dimensional distribution of chemical shifts in a spatially inhomogeneous sample using Fourier transform NMR is presented. The method uses a sequence of pulsed field gradients to measure the Fourier transform of the desired distribution on a rectangular grid in (k,t) space. Simple Fourier inversion then recovers the original distribution. An estimated signal/noise ratio of 20 in 10 min is obtained for an "image" of the distribution of a 10 mM phosphorylated metabolite in the human head at a field of 20 kG with 2-cm resolution.
An automated multimodal warping based on mutual information metric (MI) as a mapping cost function is demonstrated. Mutual information (I) is calculated from a two-dimensional (2D) gray scale histogram of an image pair, and MI (= -I) provides a matching cost function which can be effective in registration of two- or three-dimensional data sets independent of modality. Most histological image data, though information rich and high resolution, present nonlinear deformations due to the specimen sectioning and need reconstitution into deformation-corrected volumes prior to geometric mapping to an anatomical volume for spatial analyses. Section alignment via automatic 2D registrations employing MI as a global cost function and thin-plate-spline (TPS) warping is applied to deoxy-D-[14C]glucose autoradiographic image slices of a rat brain with video reference images of the uncut block face to reconstitute a cerebral glucose metabolic volume data. Unlike the traditional feature-based TPS warping algorithms, initial control points are defined independently from feature landmarks. Registration quality using automated multimodal image warping is validated by comparing MIs of the image pair registered by automated affine registration and manual warping method. The MI proves to be a robust objective matching cost function effective for automatic multimodality warping for 2D data sets and can be readily applied to volume registrations.
We have generated a spatially accurate, high-resolution three-dimensional (3D) volume of brain anatomy from cryosectioned whole human head. The head of a female cadaver was cryosectioned on a heavy duty cryomacrotome (PMV, Stockholm Sweden) modified for quantitative digital image capture. Serial images (1024(2), 24-bit) were captured directly from the cryoplaned specimen blockface in 500-micron intervals and reconstructed to a 3D data volume. Data were placed into the Talairach coordinate system to create a volume of brain anatomy for atlas reference. We resampled the volume at 500 microns along the sagittal, coronal, and horizontal planes and enhanced the images by digitally editing the background. The spatial resolution of the original digitized images provided sufficient anatomic detail to clearly delineate gray and white matter and neural structures, including major fiber pathways, subthalamic nuclei, and laminae. We developed a compact disk and controlling software program to enable the viewer to select planes of orientation, display, and copy individual to sections at higher resolution. Animation proved useful in the conveyance of system anatomy as structures are shown traversing through the neuroaxis. Postmortem cryosectioning paired with this computerized presentation allowed the complete 3D volume data to be distributed and shared as an educational, clinical, and research resource.
The association between [18F]fluorodeoxyglucose positron emission tomography (FDG-PET) counts obtained 8 h before death and neurofibrillary tangle (NFT) staining density in a patient with Alzheimer's disease (AD) was evaluated. In our patient FDG-PET counts were globally decreased with a greater focal deficit in the left medial temporal region independent of volume loss. After death, whole-brain sections derived from cryomacrotome sectioning were stained for NFTs by the Gallyas method and elastically warped into their native space enabling registration with premortem FDG-PET data. Gallyas staining density was localized to the paralimbic cortex of the basal forebrain, medial temporal, and orbital frontal regions. The poor correlation between NFT staining density and hypometabolism on FDG-PET implicates alternate mechanisms underlying the metabolic defect in AD.
Transduction of mitogenic signals in cells can be mediated by molecules derived from the synthesis and breakdown of the major membrane phospholipid, phosphotidylcholine. Studies were performed on human mammary epithelial cells in culture to understand the impact of malignant transformation and progression on membrane phospholipid metabolism. In the model system used here, phosphocholine levels and total choline-containing phospholipid metabolite levels increased with progression from normal to immortalized to oncogene-transformed to tumor-derived cells. These changes occurred independently of cell doubling time. A "glycerophosphocholine to phosphocholine switch" was apparent with immortalization. This alteration in phenotype of increased phosphocholine relative to glycerophosphocholine was observed in oncogene-transformed and for all human breast tumor cell lines analyzed. The results demonstrate that progression of human mammary epithelial cells from normal to malignant phenotype is associated with altered membrane choline phospholipid metabolism.
Using optimized, asymmetric radiofrequency (RF) pulses for slice selection, the authors demonstrate that stimulated echo acquisition mode (STEAM) localization with ultra-short echo time (1 ms) is possible. Water suppression was designed to minimize sensitivity to B1 inhomogeneity using a combination of 7 variable power RF pulses with optimized relaxation delays (VAPOR). Residual water signal was well below the level of most observable metabolites. Contamination by the signals arising from outside the volume of interest was minimized by outer volume saturation using a series of hyperbolic secant RF pulses, resulting in a sharp volume definition. In conjunction with FASTMAP shimming (Gruetter Magn Reson Med 1993;29: 804-811), the short echo time of 1 msec resulted in highly resolved in vivo 1H nuclear magnetic resonance spectra. In rat brain the water linewidths of 11-13 Hz and metabolite singlet linewidths of 8-10 Hz were measured in 65 microl volumes. Very broad intense signals (delta v(1/2) > 1 kHz), as expected from membranes, for example, were not observed, suggesting that their proton T2 are well below 1 msec. The entire chemical shift range of 1H spectrum was observable, including resolved resonances from alanine, aspartate, choline group, creatine, GABA, glucose, glutamate, glutamine, myo-inositol, lactate, N-acetylaspartate, N-acetylaspartylglutamate, phosphocreatine, and taurine. At 9.4 T, peaks close to the water were observed, including the H-1 of alpha-D-glucose at 5.23 ppm and a tentative H-1 resonance of glycogen at 5.35 ppm.
We present a method for coregistration and warping of magnetic resonance images (MRI) to histological sections for comparison purposes. This methodology consists of a modified head and hat surface-based registration algorithm followed by a new automated warping approach using nonlinear thin plate splines to compensate for distortions between the data sets. To test the methodology, 15 male Wistar rats were subjected to focal cerebral ischemia via permanent occlusion of the middle cerebral artery. The MRI images were acquired in separate groups of animals at 16-24 h (n = 9) and 48-168 h (n = 6) postocclusion. After imaging, animals were immediately sacrificed and hematoxylin- and eosin-stained brain sections were obtained for histological analysis. The MRI was coregistered and warped to histological sections. The MRI lesion areas were defined using the Eigenimage (EI) filter technique. The EI is a linear filter that maximizes the projection of a desired tissue (ischemic tissue) while it minimizes the projection of undesired tissues (nonischemic tissue) onto a composite image called an EI. When using coregistration without warping the MRI lesion area demonstrated poor correlation (r = 0.55, p > 0.01) with a percent difference between the two lesion areas of 22.5% +/- 10.8%. After warping, the MRI and histology had significant correlation (r = 0.97, p < 0.01) and a decreased percent difference of 5.56% +/- 4.31%. This methodology is simple and robust for coregistration and warping of MRI to histological sections and can be utilized in many applications for comparison of MRI to histological data.
The spin dynamics for Carr-Purcell-Meiboom-Gill-like sequences is analyzed in grossly inhomogeneous B(0) and B(1) fields. This problem is important for many applications, especially when the bandwidth of the signal is excitation limited. Examples include stray-field NMR or inside-out NMR probes used in well logging. The amplitudes of the first few echoes exhibit a characteristic transient behavior but quickly approach a smooth asymptotic behavior. For simple Hamiltonians without scalar or dipolar couplings, the evolution of a refocusing subcycle for a given isochromat is described by a rotation. Simple expressions for the signal of the Nth echo are derived in terms of these effective rotations that have a simple geometrical interpretation. It is shown that the asymptotic behavior is controlled by the direction of the axis of these effective rotations and the signal is dominated by magnetization "spin-locked" to the rotation axis. The phase of the signal is independent of the details of the field inhomogeneities. Relaxation in inhomogeneous fields leads to a signal decay that is in general nonexponential with an initial decay rate that is a weighted sum of T(-1)(1) and T(-1)(2). At long times, the echo amplitudes decay to a finite value. Phase cycling eliminates this offset. The effect of diffusion is also studied. This analysis has been applied to an inside-out NMR well logging apparatus. Good quantitative agreement is found between measurements and calculations that are based on the measured B(0) and B(1) field maps.
The rate of conversion of D-[1-(13)C]glucose into [3-(13)C]lactate (apparent glycolytic rate) has been determined in C3H murine mammary carcinomas in vivo using tumor-selective (13)C nuclear magnetic resonance spectroscopy with (1)H-(13)C cross-polarization. Under conditions of acute hypoxia induced by breathing carbon monoxide at 660 ppm, the apparent glycolytic rate was 0.0239 +/- 0.0019 min(-1). The proportion of (13)C label incorporated into [4-(13)C]glutamate (measured in tumor extracts) was 25-fold lower than that incorporated into [3-(13)C]lactate, reflecting a very limited oxidative metabolism during this hypoxic episode. For animals breathing air or carbogen (95% O(2) + 5% CO(2)), the calculated glycolytic rates were correspondingly lower (0.0160 +/- 0.0021 min(-1) and 0.0050 +/- 0.0011 min(-1), respectively). Although (13)C labeling of glutamate at C4 was still an order of magnitude lower than that for lactate at C3 (11-fold for air and 9-fold for carbogen), these ratios did show a greater degree of oxidative metabolism than that seen in animals breathing carbon monoxide at 660 ppm. The marked difference in apparent glycolytic rate for this tumor model between well-oxygenated and hypoxic conditions demonstrates a substantial Pasteur effect (inhibition of glycolysis by oxygen). Dynamic (13)C nuclear magnetic resonance spectroscopy provides a noninvasive estimate of tumor glycolysis that can be used to evaluate the relationship between oxygenation and energy metabolism, and this has potential consequences for the sensitivity of hypoxic cells to treatment and their ability to promote angiogenesis.
To develop an optical imaging method to determine the expression level of tumoral matrix metalloproteinase-2 (MMP-2) in vivo.
An optical contrast agent was developed that was highly activatable by means of MMP-2-induced conversion. Signal characteristics of the probe were quantified ex vivo with a recombinant enzyme. Animal tumor models were established with MMP-2-positive (human fibrosarcoma cell line, n = 4) and MMP-2-negative (well-differentiated mammary adenocarcinoma, n = 4) tumor cell lines. Both tumors were implanted into nude mice and were optically imaged after intravenous administration of the MMP-2-sensitive probe.
The MMP-2-sensitive probe was activated by MMP-2 in vitro, producing up to an 850% increase in near-infrared fluorescent signal intensity. This activation could be blocked by MMP-2 inhibitors. MMP-2-positive tumors were easily identified as high-signal-intensity regions as early as 1 hour after intravenous injection of the MMP-2 probe, while contralateral MMP-2-negative tumors showed little to no signal intensity. A nonspecific control probe showed little to no activation in MMP-2-positive tumors.
It is feasible to image MMP-2 enzyme activity in vivo by using near-infrared optical imaging technology and "smart" matrix metalloproteinase-sensitive probes.
Numerous investigations have shown that both tissue and cell distribution profiles of anticancer drugs can be controlled by their entrapment in submicronic colloidal systems (nanoparticles). The rationale behind this approach is to increase antitumor efficacy, while reducing systemic side-effects. This review provides an update of tumor targeting with conventional or long-circulating nanoparticles. The in vivo fate of these systems, after intravascular or tumoral administration, is discussed, as well as the mechanism involved in tumor regression. Nanoparticles are also of benefit for the selective delivery of oligonucleotides to tumor cells. Moreover, certain types of nanoparticles showed some interesting capacity to reverse MDR resistance, which is a major problem in chemotherapy. The first experiments, aiming to decorate nanoparticles with molecular ligand for 'active' targeting of cancerous cells, are also discussed here. The last part of this review focus on the application of nanoparticles in imaging for cancer diagnosis.
Matrix Metalloproteinases (MMPs) are cell-secreted soluble and membrane-tethered enzymes that degrade extracellular matrix (ECM) proteins. These proteases play a key role in diverse physiological and pathological processes, including embryonic development, wound repair, inflammatory diseases and cancer. Yet, there is insufficient knowledge on the mode by which cell-produced MMPs conduct their action on the ECM. Specifically, the localization and the mode of the degradation within the pericellular space are of great interest. To provide new insights to these questions we utilized Fourier transform infrared (FTIR) micro-spectroscopy to follow proteolytic processes, induced by invasive cancer cells, on insoluble collagen-based matrices. Here we show that FTIR micro-spectroscopy have a great potential for monitoring degradation events near cells. Using this tool we demonstrate that the net proteolysis is unevenly distributed around the cell boundary. The degradation patterns show different levels of proteolytic activity by MMPs within the pericellular space. In addition, our spectral analysis suggests that the enzymatic proteolysis of the collagen-based matrices induces unwinding of the triple helical structures of the macromolecules within the collagen network.
The aims of this study were to assess the diagnostic accuracy of MRI in evaluating patients for residual cancer, identify the prevalence of multicentric or multifocal disease, and evaluate the impact of MRI on surgical treatment planning. SUBJECTS AND METHODS. Of 101 potentially eligible patients, 80 candidates for breast conservation therapy who had primary breast cancer in 82 breasts diagnosed by excisional biopsy with close or positive margins were included in the study group. All patients underwent contrast-enhanced MRI before further surgery and subsequently underwent either reexcision lumpectomy or mastectomy with histopathologic correlation.
Residual carcinoma, either invasive or in situ, was present in 59.8% of the breasts. The sensitivity and specificity of MRI for detecting residual disease were 61.2% and 69.7%, respectively. Twenty-three additional lesions distant from the biopsy site were identified in 19 breasts, and 18 suspicious lesions underwent biopsy. Histology results indicated that six lesions were malignant, so the overall prevalence was 7.3%. The positive predictive value of identifying an additional suspicious lesion was 33.3%. In 24 breasts, MRI changed which procedure would be performed next from reexcision lumpectomy to mastectomy (n = 9), biopsy of an additional lesion in the ipsilateral (n = 12) or contralateral (n = 2) breast, or neoadjuvant chemotherapy (n = 1). Approximately 25% of the breasts underwent mastectomy as definitive surgical treatment.
Overlap in the appearances of benign and malignant lesions limits MRI evaluation for residual disease. MRI can show additional suspicious lesions that are likely to be multicentric or multifocal disease. These findings changed the original treatment plan for approximately 30% of breasts.
In the past years, the development of 3-D medical imaging has enabled the 3-D imaging of in vivo tissues, from an anatomical (MR, CT) or even functional (fMRI, PET, SPECT) point of view. However, despite immense technological progress, the resolution of these images is still short of the level of anatomical or functional details that in vitro imaging (e.g., histology, autoradiography) permits. The motivation of this work is to compare fMRI activations to activations observed in autoradiographic images from the same animals. We aim to fuse post-mortem autoradiographic data with a pre-mortem anatomical MR image. We first reconstruct a 3-D volume from the 2-D autoradiographic sections, coherent both in geometry and intensity. Then, this volume is fused with the MR image. This way, we ensure that the reconstructed 3-D volume can be superimposed onto the MR image that represents the reference anatomy. We demonstrate that this fusion can be achieved by using only simple global transformations (rigid and/or affine, 2-D and 3-D), while yielding very satisfactory results.
Integrin alpha(v)beta(3) is a widely-recognized target for the development of targeted molecular probes for imaging pathological conditions. alpha(v)beta(3) is a cell-surface receptor protein that is upregulated in various pathological conditions including osteoporosis, rheumatoid arthritis, macular degeneration, and cancer. The synthesis of an alpha(v)beta(3)-targeted optical probe 7 from compound 1, and its in vitro and in vivo characterization is described. A series of aliphatic carbamate derivatives of the potent non-peptide integrin antagonist 1 was synthesized and the binding affinity to alpha(v)beta(3) was determined in both enzyme linked immunosorbent assay (ELISA) and cell adhesion inhibition assays. The hydrophobic carbamate-linked appendages improved the binding affinity of the parent compound for alpha(v)beta(3) by 2-20 times. A Boc-protected neopentyl derivative in the series is shown to have the best binding affinity to alpha(v)beta(3) (IC(50)=0.72 nM) when compared to compound 1 as well as to c-RGDfV. Optical probe 7 utilizes the neopentyl linker and demonstrates increased binding affinity and significant tumor cell uptake in vitro as well as specific tumor accumulation and retention in vivo. These results illustrate the potential of employing integrin-targeted molecular probes based on 1 to image a multitude of diseases associated with alpha(v)beta(3) overexpression.
The hypoxia-inducible factor 1 (HIF-1) plays a critical role in cellular responses to hypoxia. The aim of the present study was to evaluate which genes are induced by hypoxia, and whether this induction is mediated by HIF-1, by expression microarray analysis of wt and HIF-1alpha null mouse fibroblasts. Forty-five genes were up-regulated by hypoxia and 40 (89%) of these were regulated by HIF-1. Of the 114 genes down-regulated by hypoxia, 19 (17%) were HIF-1-dependent. All glycolytic enzymes were strongly up-regulated by hypoxia in a HIF-1-dependent manner. Genes already known to be related to hypoxia, such as glucose transporter 1, BNIP3, and hypoxia-induced gene 1, were induced. In addition, multiple new HIF-1-regulated genes were identified, including genes involved in metabolism (adenylate kinase 4, galactokinase), apoptosis (galectin-3 and gelsolin), and invasion (RhoA). Genes down-regulated by hypoxia were involved in cytoskeleton maintenance (Rho kinase), mRNA processing (heterogeneous nuclear ribonucleoprotein H1 and splicing factor), and DNA repair (REV3). Furthermore, seven cDNAs from genes with unknown function or expressed sequence tags (ESTs) were up-regulated and 27 such cDNAs were down-regulated. In conclusion, hypoxia causes down- rather than up-regulation of gene expression and HIF-1 seems to play a major role in the regulation of hypoxia-induced genes.
The purpose of this study was to assess the value of MRI measurements of breast tumor size for predicting recurrence-free survival (RFS) in patients undergoing neoadjuvant (preoperative) chemotherapy and to compare the predictive value of MRI with that of established prognostic indicators.
The study included 62 patients undergoing neoadjuvant chemotherapy. The longest diameter and volume of each tumor were measured on MRI before and after one and four cycles of treatment. Change in diameter on clinical examination, tumor size at pathology, and the number of positive nodes were determined. Each measure of tumor extent was assessed for the ability to predict RFS.
Univariate Cox analysis showed initial MRI volume was the strongest predictor of RFS (p = 0.002). Final change in MRI volume (p = 0.015) was more predictive than change in diameter on MRI (p = 0.077) or clinical examination (p = 0.27). Initial diameter on MRI (p = 0.003) and clinical examination (p = 0.033), tumor size at pathology (p = 0.016), and number of positive nodes (p = 0.045) were also significantly predictive of RFS. Early change in MRI volume (p = 0.071) and diameter (p = 0.081) after one chemotherapy cycle showed trends of association with RFS. Multivariate analysis showed initial MRI volume (p = 0.005) and final change in MRI volume (p = 0.003) were significant independent predictors.
MRI tumor volume was more predictive of RFS than tumor diameter, suggesting that volumetric changes measured using MRI may provide a more sensitive assessment of treatment efficacy.
We developed a three-dimensional (3D) registration method to align medical scanner data with histological sections. After acquiring 3D medical scanner images, we sliced and photographed the tissue using, a custom apparatus, to obtain a volume of tissue section images. Histological samples from the sections were digitized using a video microscopy system. We aligned the histology and medical images to the reference tissue images using our 3D registration method. We applied the method to correlate in vivo magnetic resonance (MR) and histological measurements for radio-frequency thermal ablation lesions in rabbit thighs. For registration evaluation, we used an ellipsoid model to describe the lesion surfaces. The model surface closely fit the inner (M1) and outer (M2) boundaries of the hyperintense region in MR lesion images, and the boundary of necrosis (H1) in registered histology images. We used the distance between the model surfaces to indicate the 3D registration error. For four experiments, we measured a registration accuracy of 0.96+/- 0.13 mm (mean+/-SD) from the absolute distance between the M2 and H1 model surfaces, which compares favorably to the 0.70 mm in-plane MR voxel dimension. This suggests that our registration method provides sufficient spatial correspondence to correlate 3D medical scanner and histology data.