Prostate cancer tissue classification by multiphoton imaging, automated image analysis and machine learning.

Prostate carcinoma, a slow-growing and often indolent tumour, is the second most commonly diagnosed cancer among men worldwide. The prognosis is mainly based on the Gleason system through prostate biopsy analysis. However, new treatment and monitoring strategies depend on a more precise diagnosis. Here, we present results by multiphoton imaging for prostate tumour samples from 120 patients that allow to obtain quantitative parameters leading to specific tumour aggressiveness signatures. An automated image analysis was developed to recognise and quantify stromal fibre and neoplastic cell regions in each image. The set of metrics was able to distinguish between non-neoplastic tissue and carcinoma areas by linear discriminant analysis and random forest with accuracy of 89% ± 3%, but between Gleason groups of only 46% ± 6%. The reactive stroma analysis improved the accuracy to 65% ± 5%, clearly demonstrating that stromal parameters should be considered as additional criteria for a more accurate diagnosis.

Canine mammary cancer tumour behaviour and patient survival time are associated with collagen fibre characteristics.

Precise diagnosis and prognosis are key in prevention and reduction of morbidity and mortality in all types of cancers. Here we show that changes in the collagen fibres in the main histological subtypes of canine mammary gland carcinomas are directly associated with the tumour behaviour and the animal survival time and could become a useful tool in helping with diagnosis. Imaging by second harmonic generation and multiphoton excited fluorescence microscopy were performed to evaluate the collagen and cellular segment parameters in cancer biopsies. We present a retrospective study of 45 cases of canine mammary cancer analysing 836 biopsies regions including normal mammary gland tissue, benign mixed tumours, carcinoma in mixed tumour, carcinosarcoma, micropapillary carcinoma and solid carcinoma. The image analyses and the comparison between the tumour types allowed to assess the collagen fibre changes during tumour progression. We demonstrate that the collagen parameters correlate with the clinical and pathological data, the results show that in neoplastic tissues, the collagen fibres are more aligned and shorter as compared to the normal tissues. There is a clear association of the mean fibre length with the dogs survival times, the carcinomas presenting shorter collagen fibres indicate a worse survival rate.

Canine mammary cancer diagnosis from quantitative properties of nonlinear optical images.

We present nonlinear microscopy imaging results and analysis from canine mammary cancer biopsies. Second harmonic generation imaging allows information of the collagen structure in the extracellular matrix that together with the fluorescence of the cell regions of the biopsies form a base for comprehensive image analysis. We demonstrate an automated image analysis method to classify the histological type of canine mammary cancer using a range of parameters extracted from the images. The software developed for image processing and analysis allows for the extraction of the collagen fibre network and the cell regions of the images. Thus, the tissue properties are obtained after the segmentation of the image and the metrics are measured specifically for the collagen and the cell regions. A linear discriminant analysis including all the extracted metrics allowed to clearly separate between the healthy and cancerous tissue with a 91%-accuracy. Also, a 61%-accuracy was achieved for a comparison of healthy and three histological cancer subtypes studied.

What are the attitudes and beliefs of oncologists regarding potential cancer rehabilitation in a tertiary cancer center?

Cancer Rehabilitation (CR) is an emerging field in Physical Medicine & Rehabilitation. Current literature highlights the effectiveness of cancer rehabilitation in improving functional outcomes, shorter length of hospital stay, and improved quality of life. Despite this, there are very few formalized CR programs across all of North America. We conducted a survey at a tertiary cancer center without a formalized CR program to assess the perceived need of such a program and its potential development. This survey of medical, surgical, radiation and pediatric oncologists demonstrated that 92.3% of 39 respondents felt CR was somewhat to very important, particularly for their patients' issues of fatigue, deconditioning, pain management and disposition planning. These findings highlight the value seen by oncologists in the need for further cancer rehabilitation access and formalized program development in order to meet patient needs for improving functional deficits, activities of daily living and quality of life.

Surface reconstruction amendment to the intrinsic sampling method.

The intrinsic sampling method (ISM) is a powerful tool that allows the exploration of interfacial properties from molecular simulations by fitting a function that represents the local boundary between two phases. However, owing to the non-physical nature of an "intrinsic" surface, there remains an ambiguity surrounding the comparison of theoretical properties with the physical world. It is therefore important that the ISM remains internally consistent when reproducing simulated properties which match experiments, such as the surface tension or interfacial density distribution. We show that the current ISM procedure causes an over-fitting of the surface to molecules in the interface region, leading to a biased distribution of curvature at these molecular coordinates. We assert that this biased distribution is a cause of the disparity between predicted interfacial densities upon convolution to a laboratory frame, an artefact which has been known to exist since the development of the ISM. We present an improvement to the fitting procedure of the ISM in an attempt to alleviate the ambiguity surrounding the true nature of an intrinsic surface. Our "surface reconstruction" method is able to amend the shape of the interface so as to reproduce the global curvature distribution at all sampled molecular coordinates. We present the effects that this method has on the ISM predicted structure of a simulated Lennard-Jones fluid air-liquid interface. Additionally, we report an unexpected relationship between surface thermodynamic predictions of our reconstructed ISM surfaces and those of extended capillary wave theory, which is of current interest.

Unexpected finite size effects in interfacial systems: Why bigger is not always better-Increase in uncertainty of surface tension with bulk phase width.

We present an unexpected finite size effect affecting interfacial molecular simulations that is proportional to the width-to-surface-area ratio of the bulk phase Ll/A. This finite size effect has a significant impact on the variance of surface tension values calculated using the virial summation method. A theoretical derivation of the origin of the effect is proposed, giving a new insight into the importance of optimising system dimensions in interfacial simulations. We demonstrate the consequences of this finite size effect via a new way to estimate the surface energetic and entropic properties of simulated air-liquid interfaces. Our method is based on macroscopic thermodynamic theory and involves comparing the internal energies of systems with varying dimensions. We present the testing of these methods using simulations of the TIP4P/2005 water forcefield and a Lennard-Jones fluid model of argon. Finally, we provide suggestions of additional situations, in which this finite size effect is expected to be significant, as well as possible ways to avoid its impact.

Intranasal delivery of insulin and a nitric oxide synthase inhibitor in an experimental model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal neuromuscular disorder in which motor neurons may be targeted by oxidative and nitrergic stress without sufficient compensation by intrinsic support mechanisms. In this work, we addressed two key tenets of this hypothesis for the pathogenesis of ALS. Using superoxide dismutase (SOD) 1G93A mice, we studied the impact of reduction of nitrergic stress within the CNS with the use of a broad spectrum nitric oxide synthase (NOS) inhibitor, NG-nitro-l-arginine methyl ester. A separate cohort of SOD1G93A mice received direct insulin neurotrophic support, ligating receptors expressed upon motor neurons, to attempt protection against neuronal and functional motor dropout. For direct access, we used a novel form of intranasal delivery that provides peak concentration levels in the CNS within 1 h of delivery without systemic side effects at doses which previously rescued retrograde loss of motor axons after axotomy. To identify even minor impacts of these interventions on the outcome, we utilized an intensive program of serial behavioral and electrophysiological testing weekly, combined with endpoint quantitative morphometry and molecular analysis. This intensive evaluation enhanced our knowledge of the time course in SOD1G93A mice and impact of the SOD1G93A mutation upon motor neurons and their function. Neither intervention had even minimal impact upon slowing progression of disease in SOD1G93A mice. Our data argue against significant roles for nitrergic stress in promoting motor neuron loss and the importance of alternative neurotrophic support mechanisms that might support motor neurons and prevent disease progression in SOD1G93A mice.

The catalytic subunit of the DNA polymerase of herpes simplex virus type 1 interacts specifically with the C terminus of the UL8 component of the viral helicase-primase complex.

The herpes simplex virus type 1 (HSV-1) UL8 DNA replication protein is a component of a trimeric helicase-primase complex. Sixteen UL8-specific monoclonal antibodies (MAbs) were isolated and characterized. In initial immunoprecipitation experiments, one of these, MAb 804, was shown to coprecipitate POL, the catalytic subunit of the HSV-1 DNA polymerase, from extracts of insect cells infected with recombinant baculoviruses expressing the POL and UL8 proteins. Coprecipitation of POL was dependent on the presence of UL8 protein. Rapid enzyme-linked immunosorbent assays (ELISAs), in which one protein was bound to microtiter wells and binding of the other protein was detected with a UL8- or POL-specific MAb, were developed to investigate further the interaction between the two proteins. When tested in the ELISAs, five of the UL8-specific MAbs consistently inhibited the interaction, raising the possibility that these antibodies act by binding to epitopes at or near a site(s) on UL8 involved in its interaction with POL. The epitopes recognized by four of the inhibitory MAbs were approximately located by using a series of truncated UL8 proteins expressed in mammalian cells. Three of these MAbs recognized an epitope near the C terminus of UL8, which was subjected to fine mapping with a series of overlapping peptides. The C-terminal peptides were then tested in the ELISA for their ability to inhibit the POL-UL8 interaction: the most potent exhibited a 50% inhibitory concentration of approximately 5 microM. Our findings suggest that the UL8 protein may be involved in recruiting HSV-1 DNA polymerase into the viral DNA replication complex and also identify a potential new target for antiviral therapy.

The herpes simplex virus type 1 UL8 protein influences the intracellular localization of the UL52 but not the ICP8 or POL replication proteins in virus-infected cells.

We have developed a panel of 14 monoclonal antibodies (MAbs) to POL, the catalytic subunit of herpes simplex virus type 1 (HSV-1) DNA polymerase encoded by gene UL30, and one MAb to the UL52 protein, another of the seven proteins essential for replication of HSV DNA. The approximate locations of the epitopes of the polymerase-specific MAbs were identified using truncated polymerase molecules, and the antibodies were characterized in a number of immunological assays allowing eight different specificities to be recognized. These MAbs, together with a polyclonal antibody raised in rabbits against a third DNA replication protein, ICP8, were used to localize the respective proteins by immunofluorescence in cells infected with wild-type HSV-1 or the DNA replication-defective mutants ambUL8 or 2-2. In BHK cells infected with ambUL8, a mutant with an amber termination codon within the coding region of gene UL8, the UL52 protein did not enter the nucleus, although ICP8 and POL entered the nucleus in a normal fashion. The failure of the UL52 protein to be correctly transported to the nucleus was also observed in both HFL and Vero cells infected with ambUL8. In contrast, UL52 protein was transported to the nucleus in BHK cells infected with wild-type HSV-1 or with 2-2, a mutant lacking a functional UL9 protein.

Rapid urease test in the management of Campylobacter pyloridis-associated gastritis.

Campylobacter pyloridis colonization of the stomach may be an etiological factor in gastritis and peptic ulceration. Campylobacter pyloridis produces large amounts of urease, and the presence of this enzyme in gastric mucosa usually indicates infection with the organism. In this paper we describe the use of a rapid urease test (CLOtest) to detect C. pyloridis infection in gastric mucosal biopsies. In 141 consecutive endoscopy cases, antral biopsies were taken for culture and histology, and an extra biopsy was inserted into the CLOtest gel. There were 79 patients infected with C. pyloridis, 78 of whom were detected by CLOtest: 75% were positive at 20 min, 92% at 3 h, and 98% at 24 h. There were no false positive results. Eighteen infected patients were rebiopsied after a course of amoxycillin and bismuth subcitrate. Active chronic gastritis resolved in eight of nine who were cleared of the organism, but histological gastritis was unchanged in nine patients who were still infected. CLOtest is a simple, sensitive, and highly specific test that enables the endoscopist to diagnose C. pyloridis infection in the endoscopy room. A negative test after antibiotic therapy correlates with clearance of the bacteria and healing of active gastritis.

Antibacterial action of bismuth in relation to Campylobacter pyloridis colonization and gastritis.

Colloidal bismuth subcitrate (CBS, De-Nol) heals duodenal ulcers but with a lower relapse rate than cimetidine, perhaps due to inhibition of Campylobacter pyloridis (CP) organisms. To test this hypothesis we studied gastric mucosal histology in three groups of ulcer patients treated with either cimetidine, CBS, or CBS in combination with an antibiotic. Cimetidine had no effect on CP or gastric mucosal histology but with CBS therapy there was a significant reduction in the number of bacteria (p less than 0.0001). However, relapse of both CP infection and gastritis usually occurred once CBS was withdrawn. When CBS was combined with amoxycillin or tinidazole, long-term disappearance of both CP bacteria and gastritis was achieved (p less than 0.0001). In ultrastructural studies 30-90 min after single oral doses of CBS or bismuth subsalicylate, CP had detached from the gastric epithelial cells and exhibited structural degradation associated with the selective deposition of a particulate bismuth complex within and upon the surface of the organisms. In vitro, CP and other campylobacters were inhibited by bismuth compounds at 25 mg/l but they were resistant to cimetidine and ranitidine. CBS has a powerful antibacterial effect against CP but relapse of infection is common after CBS alone. In combination with antibiotics however, eradication of CP and long-term healing of gastritis occurs. In such cases the gastroduodenal mucosa is intact, and less likely to ulcerate.