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Processing of medical images using real-time optical Fourier processing.

Optical image processing techniques are inherently fast in view of parallel processing. A self-adaptive optical Fourier processing system using photoinduced dichroism in a bacteriorhodopsin film was experimentally demonstrated for medical image processing. Application of this powerful analog all-optical interactive technique for cancer diagnostics is illustrated with two mammograms and a Pap smear. Microcalcification clusters buried in surrounding tissue showed up clearly in the processed image. By playing with one knob, which rotates the analyzer in the optical system, either the microcalcification clusters or the surrounding dense tissue can be selectively displayed. Bacteriorhodopsin films are stable up to 140 degrees C and environmentally friendly. As no interference is involved in the experiments, vibration isolation and even a coherent light source are not required. It may be possible to develop a low-cost rugged battery operated portable signal-enhancing magnifier.

Bacteriorhodopsins↗

Three-dimensional fluorescence lifetime tomography.

Near-infrared fluorescence tomography using molecularly targeted lifetime-sensitive, fluorescent contrast agents have applications for early-stage cancer diagnostics. Yet, although the measurement of fluorescent lifetime imaging microscopy (FLIM) is extensively used in microscopy and spectroscopy applications, demonstration of fluorescence lifetime tomography for medical imaging is limited to two-dimensional studies. Herein, the feasibility of three-dimensional fluorescence-lifetime tomography on clinically relevant phantom volumes is established, using (i) a gainmodulated intensified charge coupled device (CCD) and modulated laser diode imaging system, (ii) two fluorescent contrast agents, e.g., Indocyanine green and 3-3'-Diethylthiatricarbocyanine iodide differing in their fluorescence lifetime by 0.62 ns, and (iii) a two stage approximate extended Kalman filter reconstruction algorithm. Fluorescence measurements of phase and amplitude were acquired on the phantom surface under different target to background fluorescence absorption (70:1, 100:1) and fluorescence lifetime (1:1, 2.1:1) contrasts at target depths of 1.4-2 cm. The Bayesian tomography algorithm was employed to obtain three-dimensional images of lifetime and absorption owing to the fluorophores.

Algorithms↗

Focused array transducer for two-dimensional optoacoustic tomography.

Optoacoustic (OA) imaging utilizes short laser pulses to create acoustic sources in tissue and time resolved detection of generated pressure profiles for image reconstruction. The ultrasonic transients provide information on the distribution of optical absorption coefficient that can be useful for early cancer diagnostics. In this work a new design of wide-band array transducer is developed and tested. The array consists of 32 focused piezo-elements made of PVDF slabs imposed on a cylindrical surface. A single array element response to an OA signal coming from arbitrarily located point source is investigated theoretically and experimentally. The measured signals correspond well to numerically calculated ones. Focal zone maps of the elements with aperture angles 30 degrees and 60 degrees are presented and discussed; the resolution in direction perpendicular to the imaging plane is determined. Point spread function of the whole array is calculated using experimentally obtained signals from the sources located at different distances from the array. Backprojection algorithm is employed for reconstruction of the optoacoustic images. It is shown that the spatial resolution of the images yielded by the proposed array increases significantly compared to previous transducer designs.

Equipment Design↗

Mechanical spectral signatures of malignant disease? A small-sample, comparative study of continuum vs. nano-biomechanical data analyses.

Thin sections from human breast biopsies were employed to perform a differential analysis of the ultrasound spectral responses from invasive ductal carcinoma and normal tissue. A non-destructive testing methodology was employed, yielding the reflection coefficients as function of frequency in the clinical ultrasound range. The spectral responses were simulated both in the context of continuum and nano-biomechanics, with the objective of quantifying the physical properties that determine the differences in the spectral signature of normal vs. malignant tissue. The properties that were employed for the theoretical reconstruction of the spectra were: the density, the continuum and the nanomechanical elastic constants, and the nanomechanical theory internodal distance. The latter is a measure of the depth-of-penetration of mechanical actions between contiguous tissue elements. Together with vectorial descriptors of the tissue spatial arrangement, the internodal distance variable affords the quantitative incorporation of tissue architectural data in the theoretical model. In this paper, the validity of the nanomechanical approach to tissue characterization is discussed, and its potential extensions to biomolecular marker-based cancer diagnostics and therapeutics are considered.

Breast Neoplasms↗

Wnt/beta-catenin/tcf signaling: a critical pathway in gastrointestinal tumorigenesis.

Cancers of the gastrointestinal tract, including the liver, bile ducts, and pancreas, constitute the largest group of malignant tumors. Colorectal cancer is one of the most common neoplastic diseases in Western countries and one of the leading causes of cancer-related deaths. Inactivation of the adenomatous polyposis coli (APC) tumor-suppressor gene during early adenoma formation is thought to be the first genetic event in the process of colorectal carcinogenesis followed by mutations in oncogenes like K-Ras and tumor-suppressor genes like p53. Identification of the interaction of APC with the proto-oncogene beta-catenin has linked colorectal carcinogenesis to the Wnt-signal transduction pathway. The main function of APC is thought to be the regulation of free beta-catenin in concert with the glycogen synthase kinase 3beta (GSK-3beta) and Axin proteins. Loss of APC function, inactivation of Axin or activating beta-catenin mutations result in the cellular accumulation of beta-catenin. Upon translocation to the nucleus beta-catenin serves as an activator of T-cell factor (Tcf)-dependent transcription leading to an increased expression of several specific target genes including c-Myc, cyclin D1, MMP-7, and ITF-2. While APC mutations are almost exclusively found in colorectal cancers, deregulation of Wnt/beta-catenin/Tcf signaling is also common in other gastrointestinal and extra-gastrointestinal human cancers. In a fraction of hepatocellular carcinomas the Wnt pathway is deregulated by inactivation of Axin or stabilizing mutations of beta-catenin. The majority of hepatoblastomas and a group of gastric cancers also carry beta-catenin mutations. Clearly, this pathway harbors great potential for future applications in cancer diagnostics, staging, and therapy.

Colorectal Neoplasms↗

Density estimation for positron emission tomography.

PET (positron emission tomography) scans are still in the experimental phase, as one of the newest breast cancer diagnostic techniques. There are two traditional approaches to the computation of images from data collected in PET. In the first, standard CT (computed tomography) algorithms are used on rays designated by pairs of detectors receiving coincidence events. The problem generated by this approach is that generally only the mean can be used by such algorithms. With the relatively small numbers of events in PET, and with Poisson statistics for which variance equals the mean, the noise sensitivity of standard CT algorithms becomes limiting. This is exasperated further by 3D imaging with cylindrical arrays of detectors. Statistical CT algorithms take the variance into account. As in the list-mode approach, we consider each coincidence event individually. However, we estimate the location of the annihilation event that caused each coincidence event, one by one, based on the previously assigned location of events processed earlier. The estimated annihilation locations form the image. To accomplish this, we construct a probability distribution along each coincidence line. This is generated from previous annihilation points by density estimation. In this paper we present our density estimation approach to positron emission tomography. Nonparametric methods of density estimation are overviewed followed by numerical examples. Our goal here is to determine which density estimation approach is most suitable for PET.

Algorithms↗

Extracellular DNA in breast cancer: Cell-surface-bound, tumor-derived extracellular DNA in blood of patients with breast cancer and nonmalignant tumors.

A methylation-specific polymerase chain reaction technique was used to investigate aberrant promoter methylation of RASSF1A and HIC-1 genes in circulating extracellular DNA (exDNA) from the blood of breast cancer and fibroadenoma patients. Methylated DNA could be detected in the exDNA eluted from the surface of erythrocytes and leukocytes, even in the samples where no methylated DNA could be detected in plasma. The data obtained demonstrate that cell surface bound exDNA provides a valuable source of material for early noninvasive cancer diagnostics and monitoring.

Breast Neoplasms↗

Isolation and comparative study of cell-free nucleic acids from human urine.

Cell-free nucleic acids (NA) from human urine were investigated. Concentrations of DNA and RNA in the urine of healthy people were independent of gender and were in the range of 6 ng/mL to 50 ng/mL and 24 ng/mL to 140 ng/mL, respectively. DNA fragments of 150-400 bp represent the main part of cell-free DNA, along with DNA fragments up to 1,300 bp, which were found in male urine, and DNA fragments up to 19 kbp, which were found in female urine. Analysis of circulating DNA, isolated from blood of breast cancer patients and cell-free DNA isolated from their urine by methylation-specific PCR, demonstrates that the presence of methylated promoters of RASSF1A and RARbeta2 genes in plasma was accompanied by the detection of the same methylated markers in urine. The data obtained demonstrate applicability of cell-free urine DNA in cancer diagnostics.

Biomarkers, Tumor↗

Nipple aspirate fluid expression of urokinase-type plasminogen activator, plasminogen activator inhibitor-1, and urokinase-type plasminogen activator receptor predicts breast cancer diagnosis and advanced disease.

BACKGROUND: Tumor expression of urokinase-type plasminogen activator (uPA), plasminogen activator inhibitor-1 (PAI-1), and uPA receptor (uPAR) are breast cancer prognostic factors. Less is known about their usefulness in breast cancer diagnosis. Nipple aspirate fluid (NAF) is secreted into the breast duct and collected noninvasively, making it potentially useful both in breast cancer diagnosis and prognosis. We determined the association of uPA, PAI-1, and uPAR levels in NAF with breast cancer (1) detection and (2) advanced disease. METHODS: A total of 88 NAF specimens were collected from women with or without breast cancer, and uPA, PAI-1, and uPAR expression were measured by enzyme-linked immunosorbent assay. RESULTS: uPA and uPAR were independent predictors of cancer presence; uPAR was also an independent predictor of advanced disease stage. Higher PAI-1 expression in breast cancer that was found with univariate analysis was not observed after logistic regression was applied. CONCLUSIONS: NAF evaluation of uPA, uPAR, and, perhaps, PAI-1 (significant only in univariate analysis) may provide useful breast cancer diagnostic and prognostic information.

Adult↗

The role of adult stem cells in carcinogenesis.

The role of adult stem cells in tissue regeneration has attracted much interest because of its enormous therapeutic potential. Adult stem cells reside in every tissue of the body and have the ability to self-renew and to give rise to a high number of differentiated progeny. These are indeed the essential steps in tissue regeneration, which in some cases occurs constitutively, whereas in other cases happens in response to an injury. The identification of the key factors involved in self-renewal and differentiation pathways is at the top of the agenda of stem cell biologists. Interestingly, a number of factors that govern the fate of adult stem cells also play a role in malignant cell transformation, such as the Wnt cascade, Oct-4, Bmi-1 and Evi1. In addition, rare subpopulations of cancer stem cells were identified for leukemia and other solid tumors. These rare fractions of cancer stem cells are in large part responsible for maintaining the tumor mass, as they have the capacity to self-renew and to generate a high number of progeny via differentiation program. These findings can be considered a major breakthrough in cancer research. Currently, oncologists and stem cell biologists have two main goals: i) the assessment of the consistency of the line that divides adult stem cells and carcinogenesis; ii) to better characterize the biology of cancer stem cells. This new line of investigation may allow for the development of novel therapeutics and cancer diagnostics.

AC133 Antigen↗

Time-resolved optical tomography using short-pulse laser for tumor detection.

Our objective is to perform a comprehensive experimental and numerical analysis of the short-pulse laser interaction with a tissue medium with the goal of tumor-cancer diagnostics. For a short-pulse laser source, the shape of the output signal is a function of the optical properties of the medium, and hence the scattered temporal optical signal helps in understanding the medium characteristics. Initially experiments are performed on tissue phantoms embedded with inhomogeneities to optimize the time-resolved optical detection scheme. Both the temporal and the spatial profiles of the scattered reflected and transmitted optical signals are compared with the numerical modeling results obtained by solving the transient radiative transport equation using the discrete ordinates technique. Next experiments are performed on in vitro rat tissue samples to characterize the interaction of light with skin layers and to validate the time-varying optical signatures with the numerical model. The numerical modeling results and the experimental measurements are in excellent agreement for the different parameters studied. The final step is to perform in vivo imaging of anesthetized rats with tumor-promoting agents injected inside skin tissues and of an anesthetized mouse with mammary tumors to demonstrate the feasibility of the technique for detecting tumors in an animal model.

Animals↗

Kinetworks protein kinase multiblot analysis.

The proteomics analysis of protein kinases and other cell-signaling proteins in tumor samples by traditional two-dimensional (2-D) gel electrophoresis is complicated by the low abundance of these regulatory proteins relative to metabolic enzymes and structural proteins. We present an antibody-based method called Kinetworks that relies on sodium dodecyl sulfate (SDS)-poly-acrylamide minigel electrophoresis and multilane immunoblotters to permit the specific and quantitative detection of 45 or more protein kinases or other signal transduction proteins at once. The technique can also permit the resolution of these proteins based on differences in their phosphorylation state and other forms of covalent modification. Kinetworks profiling of protein kinases in solid human tumors and cell lines can reveal profound differences in their expression and phosphorylation states, which can serve for the identification of cancer diagnostic markers and therapeutic targets for drug discovery.

Biomarkers, Tumor↗

[Neoplasms associated with esophageal epidermoid carcinoma].

INTRODUCTION: The esophageal epidermoid cancer has an important association with aerodigestive tract neoplasms and possibly share the same risk factors. Furthermore, other neoplasms can be associated with esophagus cancer. OBJECTIVE: To analyze retrospectively the patients with esophageal epidermoid cancer (EEC) and associated neoplasms, treated by the Esophagus Stomach and Small Intestine Group of Surgery at Hospital de Clinicas de Porto Alegre from January 1988 to June 1995. PATIENTS AND METHODS: Nineteen (7.28%) of the 261 studied patients had associated neoplasms to the EEC. Ten patients presented synchronic tumours and 9 metachronic ones. The predominant sex was the masculine with 17 cases. The mean age was 62.52 years in the moment of the esophageal cancer diagnostic. RESULTS: The aerodigestive tumours, squamous carcinomas in totality, represented the predominant associated neoplasm histological type in 68.42% of the cases. The most frequent associated aerodigestive tumours site was the respiratory tract (53.8%), followed by the oral cavity and oropharynx (23%) and larynx (23%). In our sample, twelve patients were smokers and 9 were alcohol abusers. In relation to the EEC treatment, surgery was performed in 6 patients. The associated neoplasm was treated with radical surgery in 11 patients and radiotherapy in 5. Surprisingly 4 cases (21%) of gastric adenocarcinoma associated to the EEC were diagnosed, treated with radical surgery in 3 patients. CONCLUSION: The authors call attention to the importance of a criterial staging as well as the follow up in patients with EEC owing to the significant association with others neoplasms, principally with aerodigestive tumours, and discuss the common risk factors possibility: tobacco and alcohol use. Important association with gastric neoplasms were found in this casuistry.

Adult↗

High-throughput microarray-based enzyme-linked immunosorbent assay (ELISA).

A new generation biochip is described as capable of supporting high-throughput (HT), multiplexed enzyme-linked immunosorbent assays (ELISAs). These biochips consist of an optically flat, glass plate containing 96 wells formed by an enclosing hydrophobic Teflon mask. The footprint dimensions of each well and the plate precisely match those of a standard microplate. Each well contains four identical 36-element arrays (144 elements per well) comprising 8 different antigens and a marker protein. Arrays are formed by a custom, continuous flow, capillary-based print head attached to a precise, high-speed, X-Y-Z robot. The array printing capacity of a single robot exceeds 20,000 arrays per day. Arrays are quantitatively imaged using a custom, high-resolution, scanning charge-coupled device (CCD) detector with an imaging throughout of 96 arrays every 30 s. Using this new process, arrayed antigens were individually and collectively detected using standard ELISA techniques. Experiments demonstrate that specific multiplex detection of protein antigens arrayed on a glass substrate is feasible. Because of the open microarray architecture, the 96-well microarray format is compatible with automated robotic systems and supports a low-cost, highly parallel assay format. Future applications of this new high-throughput screening (HTS) format include direct cellular protein expression profiling, multiplexed assays for detection of infectious agents and cancer diagnostics.

Adsorption↗

Clinical experiences with direct magnification mammography with the DIMA Plus M11.

OBJECTIVE: The evaluation of mammographies carried out by conventional technique (standard imaging and focal imaging with 1.9x magnification) should be compared with a direct magnification image (standard imaging with 1.7x magnification, focal imaging with 4x magnification and preparation imaging with 7x magnification) provided by the mammographic device DIMA Plus M11 of the company feinfocus Medizintechnik. MATERIAL AND METHODS: Out of over 1,000 mammographies (DIMA technique) 50 histologically proved cases were selected for evaluation. Within a three months period these cases underwent conventional standard mammography as well as 1.9x magnification and DIMA-mammographies. The second X-ray was carried out when it was necessary for a pre-operative marking. RESULTS: When mammographies of mammaries, which were radiologically transparent and easily compressible, where taken by DIMA-technique, they showed a distinct advantage, especially in unclear micro-calcification cases, in comparison to the mammographies carried out by the conventional standard imaging. CONCLUSIONS: Direct magnification images carried out by DIMA Plus M11 provide a better breast cancer diagnostic. This refers in particular to focal images with 4x magnification and to digital mammography, which is yet being developed.

Breast Neoplasms↗

Detection of tumor messenger RNA in the serum of patients with malignant melanoma.

Serum RNases are known to be elevated in patients with cancer. Consequently, it is not clear whether human mRNA with sufficient integrity as to permit reverse transcription-PCR (RT-PCR) amplification is detectable in serum. We examined serum from six patients with malignant melanoma for human tyrosinase mRNA using RT-PCR. Serum from 20 normal volunteers served as controls. Tyrosinase mRNA could be demonstrated in serum from four of the six melanoma patients with detection by gel electrophoresis and confirmation by blotting amplified product to a tyrosinase-specific probe. The serum remained tyrosinase mRNA positive, even if passed through a 0.45 microm filter prior to RNA extraction, indicating that the mRNA was extracellular at the time of extraction. Tyrosinase mRNA could not be detected in any control serum (0 of 20 individuals). The presence and integrity of amplifiable RNA was confirmed in all serum specimens (patients and controls) by RT-PCR amplification of c-abl mRNA. Amplifiable RNA could be demonstrated regardless of whether serum was freshly drawn or stored frozen for several years. We conclude that human mRNA can be extracted and amplified from serum. The ability to amplify tumor mRNA from serum may have important utility in cancer diagnostics and monitoring.

Blotting, Southern↗

[Molecular pathology: intersection of morphology, biochemistry and genetics].

Molecular analysis of samples of cells and tissues plays an important part in clinical pathology, as a supplement to classical morphological examination. This holds true without reserve for immunohistochemistry, by now indispensable for clinical pathology. It was only recently that molecular genetic analysis was introduced into the pathological laboratory and it may be expected that this technique will play an important part, especially in cancer diagnostics. Where haemato-oncological malignancies and soft tissue sarcomas are concerned, this is already the case. Molecular tumour analysis will gain momentum due to advanced automated analysis using DNA arrays. Morphological aspects will long remain the foundation of tumour classification, but in the future not without major support from molecular analysis.

Cytogenetic Analysis↗

Diagnosis and treatment of breast cancer.

This article presents a thorough review of the diagnosis and treatment of breast cancer, the most common type of cancer found among American women. Topics covered in this article include risk factors, screening, types of breast cancer, diagnostic procedures, surgical and nonsurgical treatments, staging, genetic testing, and nutrition and complementary therapies.

Breast Neoplasms↗