Bovine scavenger receptor class A (SR-A) exhibit specific patterns of regulation in the endometrium during the oestrous cycle and early pregnancy.

In mammals, tight regulation of maternal endometrial function is critical for pregnancy success. In bovine species, endometrial expression of members of the scavenger receptor class A (SR-A) has been listed in high-throughput analyses, but very little is known about the involvement of these immune factors during implantation in mammals. To provide first insights into the contribution of SR-A to endometrial physiology, we analysed the expression and regulation of all members of SR-A (SR-A1, SR-A3-SR-A6) during the oestrous cycle and early pregnancy in cattle. Levels of SR-A1 were increased on Day 20 of pregnancy, whereas SR-A3 levels were increased on Day 13 of the oestrous cycle and of the pregnancy. Although SR-A4 levels were reduced on Day 20 of the oestrous cycle, they remained high in pregnant animals. SR-A5 levels increased by Day 13 of the oestrous cycle and decreased on Day 20, but remained high in pregnant animals. Interferon-τ does not affect SR-A gene expression, whereas progesterone regulates the expression of the SR-A3 and SR-A5 transcripts. Endometrial SR-A3 appeared significantly higher in cows carrying invitro-produced embryos than in AI cows. Our data suggest that members of the SR-A family are involved in endometrial remodelling and regulation of endometrial gland physiology, both processes being critical for implantation in mammals.

Analysis of STAT1 expression and biological activity reveals interferon-tau-dependent STAT1-regulated SOCS genes in the bovine endometrium.

Signal transducer and activator of transcription (STAT) proteins are critical for the regulation of numerous biological processes. In cattle, microarray analyses identified STAT1 as a differentially expressed gene in the endometrium during the peri-implantation period. To gain new insights about STAT1 during the oestrous cycle and early pregnancy, we investigated STAT1 transcript and protein expression, as well as its biological activity in bovine tissue and cells of endometrial origin. Pregnancy increased STAT1 expression on Day 16, and protein and phosphorylation levels on Day 20. In cyclic and pregnant females, STAT1 was located in endometrial cells but not in the luminal epithelium at Day 20 of pregnancy. The expression of STAT1 during the oestrous cycle was not affected by progesterone supplementation. In vivo and in vitro, interferon-tau (IFNT) stimulated STAT1 mRNA expression, protein tyrosine phosphorylation and nuclear translocation. Using chromatin immunoprecipitation in IFNT-stimulated endometrial cells, we demonstrated an increase of STAT1 binding on interferon regulatory factor 1 (IRF1), cytokine-inducible SH2-containing protein (CISH), suppressor of cytokine signaling 1 and 3 (SOCS1, SOCS3) gene promoters consistent with the induction of their transcripts. Our data provide novel molecular insights into the biological functions of STAT1 in the various cells composing the endometrium during maternal pregnancy recognition and implantation.

SOCS genes expression during physiological and perturbed implantation in bovine endometrium.

In mammals, suppressor of cytokine signalling (CISH, SOCS1 to SOCS7) factors control signalling pathways involved in the regulation of numerous physiological processes including pregnancy. In order to gain new insights into the biological functions of SOCS in the endometrium, a comprehensive analysis of SOCS gene expression was carried out in bovine caruncular (CAR) and intercaruncular (ICAR) tissues collected i) during the oestrous cycle, ii) at the time of maternal recognition of pregnancy and at implantation in inseminated females, iii) following uterine interferon-tau (IFNT) infusion at day 14 post-oestrus, iv) following a period of controlled intravaginal progesterone release and v) following transfer of embryos by somatic-cell nuclear transfer (SCNT). The regulatory effects of IFNT on in vitro cultured epithelial and stromal cells were also examined. Altogether, our data showed that CISH, SOCS4, SOCS5 and SOCS7 mRNA levels were poorly affected during luteolysis and pregnancy. In contrast, SOCS1, SOCS2, SOCS3 and SOCS6 mRNA levels were strongly up-regulated at implantation (day 20 of pregnancy). Experimental in vitro and in vivo models demonstrated that only CISH, SOCS1, SOCS2 and SOCS3 were IFNT-induced genes. Immunohistochemistry showed an intense SOCS3 and SOCS6 staining in the nucleus of luminal and glandular epithelium and of stromal cells of pregnant endometrium. Finally, SOCS3 expression was significantly increased in SCNT pregnancies in keeping with the altered immune function previously reported in this model of compromised implantation. Collectively, our data suggest that spatio-temporal changes in endometrial SOCS gene expression reflect the acquisition of receptivity, maternal recognition of pregnancy and implantation.

Longer storage of red blood cells is associated with increased in vitro erythrophagocytosis.

BACKGROUND AND OBJECTIVES: Refrigerated storage of red blood cells (RBCs) induces numerous changes that may target the cells for erythrophagocytosis following transfusion. The influence of storage upon the phagocytosis of unseparated and fractionated young and old stored RBCs was investigated using two in vitro quantitative phagocytosis assays. MATERIALS AND METHODS: Leucocyte-depleted RBC units were sampled at day 1 or 42 of storage. Young and old RBCs were fractionated at day 1 by density centrifugation and stored in paediatric packs for up to 42 days. RBCs were labelled with the fluorescent dye PKH26 and incubated with the human monocytic cell line THP-1. Erythrophagocytosis was quantified by flow cytometry and plate fluorometric assays. RESULTS: A higher proportion of THP-1 cells phagocytosed RBCs stored for 42 days compared with 1 day (41% and 24% respectively; P<0·0001). This was associated with an increased mean number of RBCs phagocytosed per THP-1 cell (5·2±0·6 and 3·3±0·2 respectively; P<0·002). Erythrophagocytosis of fractionated young and old RBCs increased with longer storage duration up to 28 days (P<0·05). However, no significant differences were observed between erythrophagocytosis of young and old RBCs. CONCLUSION: The susceptibility of stored RBCs to erythrophagocytosis significantly increased with longer storage time of the RBC units. Storage duration of RBCs had a greater influence on in vitro erythrophagocytosis than the chronological age of the RBCs at donation.

Immunity and inflammation in the uterus.

Microbes often infect the uterus and particularly the endometrium of animals. Infections are most commonly associated with natural service, pregnancy and the post-partum period, leading to inflammation with the elaboration of cytokines, chemokines and prostaglandins. Clinical diseases such as metritis, endometritis and abortion are important causes of infertility. The adaptive immune response to infection has been characterized previously, so the present review aims to highlight the emerging role for innate immunity in the endometrium. The detection of microbes and the innate immune response depends on the detection of pathogen-associated molecular patterns by pattern recognition receptors. The main families of pattern recognition receptors are Toll-like receptors (TLRs), nucleotide oligomerization domain-like receptors, retinoic acid-inducible gene I-like receptors and C-type lectin receptors. These receptors are most often expressed by hematopoietic cells, but the epithelial and stromal cells of the endometrium also possess functional receptors. For example, endometrial cells express TLR4 for recognition of the lipopolysaccharide endotoxin of Gram-negative bacteria, leading to secretion of IL-6, IL-8 and prostaglandin E(2) . It is likely that the epithelial and stromal cells provide a first line of defence in the endometrium to alert hematopoietic cells to the presence of microbes within the uterus.

Clinical validation of an autoantibody test for lung cancer.

BACKGROUND: Autoantibodies may be present in a variety of underlying cancers several years before tumours can be detected and testing for their presence may allow earlier diagnosis. We report the clinical validation of an autoantibody panel in newly diagnosed patients with lung cancer (LC). PATIENTS AND METHODS: Three cohorts of patients with newly diagnosed LC were identified: group 1 (n = 145), group 2 (n = 241) and group 3 (n = 269). Patients were individually matched by gender, age and smoking history to a control individual with no history of malignant disease. Serum samples were obtained after diagnosis but before any anticancer treatment. Autoantibody levels were measured against a panel of six tumour-related antigens (p53, NY-ESO-1, CAGE, GBU4-5, Annexin 1 and SOX2). Assay sensitivity was tested in relation to demographic variables and cancer type/stage. RESULTS: The autoantibody panel demonstrated a sensitivity/specificity of 36%/91%, 39%/89% and 37%/90% in groups 1, 2 and 3, respectively, with good reproducibility. There was no significant difference between different LC stages, indicating that the antigens included covered the different types of LC well. CONCLUSION: This assay confirms the value of an autoantibody panel as a diagnostic tool and offers a potential system for monitoring patients at high risk of LC.

The use of Photovoice to document and characterize the food security of users of community food programs in Iqaluit, Nunavut.

INTRODUCTION: Food insecurity is a chronic problem affecting Inuit communities. The most comprehensive assessment of Inuit food security to-date, the Inuit Health Survey, reported that 70% of Inuit pre-school children lived in 'food insecure' households. Food banks and soup kitchens are relatively new in the Arctic but the number of users is increasing. Little is known about the experience and determinants of food insecurity among food program users who are often among the most marginalized (socially and economically) in communities. The use of participatory research methods when working in the north of Canada can promote meaningful knowledge exchange with community members and this approach was used in the present 'Photovoice' research. Photovoice uses photography to develop a baseline understanding of an issue, in this case the experience and determinants of food insecurity among users of community food programs in Iqaluit, Nunavut. The target population includes those who face significant social and economic marginalization, an often neglected group in Arctic food systems research. METHODS: Eight regular users of food programs were recruited and engaged in a Photovoice research project to document factors determining their daily food consumption. The research method was introduced in workshops and discussion included the ethical concerns related to photography and how to take pictures. Participants were supplied with digital cameras, and asked to answer the following question using photography: 'What aspects of your everyday life affect what you eat and how much you have to eat?'. In the final workshop, photographs were discussed among the group and participants identified key themes in the photographs, offering an understanding of food insecurity from their perspectives. The group then discussed what should be done with the knowledge gained. RESULTS: Factors improving food security were the customary systems for sharing 'country food', and the presence of social support networks in the community, such as the Food Bank, the Soup Kitchen and the Tukisigiarvik Center. Factors identified as negatively affecting food security were the high cost of food in the Arctic, and substance abuse. The participants decided by consensus whether and how the knowledge from this project would be disseminated. They decided that a museum exhibit of the photographs in the summer of 2010 and promotion of the results among policy-makers in Nunavut were of high priority. CONCLUSION: The use of participatory research approaches such as Photovoice offers promise for exploring food security issues among similarly disadvantaged and vulnerable populations elsewhere in the Arctic. This approach was found to be a useful method for gathering and sharing research data because the data was generated and analysed by the participants. The clear and concise messages developed by the participants can be used to inform policy. This research method can assist in making a valuable contribution to health research, both in the Arctic and worldwide, because it promotes an understanding of the experiences of individuals from their own perspective.

Technical validation of an autoantibody test for lung cancer.

BACKGROUND: Publications on autoantibodies to tumour-associated antigens (TAAs) have failed to show either calibration or reproducibility data. The validation of a panel of six TAAs to which autoantibodies have been described is reported here. MATERIALS AND METHODS: Three separate groups of patients with newly diagnosed lung cancer were identified, along with control individuals, and their samples used to validate an enzyme-linked immunosorbant assay. Precision, linearity, assay reproducibility and antigen batch reproducibility were all assessed. RESULTS: For between-replicate error, samples with higher signals gave coefficients of variation (CVs) in the range 7%-15%. CVs for between-plate variation were only 1%-2% higher. For between-run error, CVs were in the range 15%-28%. In linearity studies, the slope was close to 1.0 and correlation coefficient values were generally >0.8. The sensitivity and specificity of individual batches of antigen varied slightly between groups of patients; however, the sensitivity and specificity of the panel of antigens as a whole remained constant. The validity of the calibration system was demonstrated. CONCLUSIONS: A calibrated six-panel assay of TAAs has been validated for identifying nearly 40% of primary lung cancers via a peripheral blood test. Levels of reproducibility, precision and linearity would be acceptable for an assay used in a regulated clinical setting.

Quantitation of proteins and internal antigen pools by a monoclonal sandwich radioimmune assay.

We describe a monoclonal sandwich radioimmune assay (MSRA) that is capable of measuring 0.5 fmoles of protein antigens. The basis of the assay is the ability of most rabbit heteroantisera directed against protein antigens to precipitate labeled monoclonal Fab-antigen complexes. The assay offers the advantages of specificity, sensitivity, linearity and rapidity. The MSRA circumvents the need for preparation of either highly purified protein antigens for use in radioimmune inhibition assays or affinity purified antisera for sandwich immunoassays. Utilizing variations of MSRA protocols, we determined that 48% of FcR gamma 2b/gamma 1, 34% of an 90,000 Mr glycoprotein identified by monoclonal antibody 2D2C, and 24% of an 82,000 Mr glycoprotein identified by monoclonal antibody 2E2A were accessible to labeled Fab probes at the plasma membrane. The total amount of antigen per cell was determined by dividing the amount of antigen at the cell surface (determined from binding saturation assays) by the percentage of surface/total antigen. This value of total antigen agrees well with the total cellular antigen pool determined by MSRA after appropriate corrections for probe saturation and recovery of pre-formed Fab-antigen complexes were made.

Optimum field size and choice of isodose lines in electron beam treatment.

PURPOSE: A method is provided for the optimum field size and the choice of isodose line for the dose prescription in electron beam therapy. METHODS AND MATERIALS: Electron beam dose uniformity was defined in terms of target coverage factor (TCF) which is an index of dose coverage of a given treatment volume. The TCF was studied with respect to the field size, the beam energy, and the isodose level for prescription from the measured data for various accelerators. The effect of the TCF on air gap between electron applicator/cone and the surface was investigated. Electron beams from scattering foil and scanned beam units were analyzed for the target coverage. RESULTS: A mathematical method is provided to optimize a field size for target coverage by a given isodose line in terms of TCF which is strongly dependent on the type of accelerator and the design of the collimator. For a given type of collimating system, the TCF does not depend on the type of electron beam production (scattering foil or swept scanned beam). Selection of isodose line for dose prescription is very critical for the value of the TCF and the dose coverage. The TCF is inversely proportional to the isodose value selected for the treatment and nearly linear with field size and beam energy. Air gap between applicator and the surface reduces the dose uniformity. Tertiary collimator moderately improves the lateral coverage for high energy beams. CONCLUSIONS: To adequately cover the target volume in electron beam treatment, lateral and depth coverage should be considered. The coverage at depth is strongly dependent on the choice of isodose line or beam normalization. If the dose prescription is at dmax (i.e., the 100% isodose line is selected), the choice of beam energy is not critical for depth coverage since dmax is nearly independent of energy for smaller fields. The 100% isodose line should not be chosen for treatment because of the significant constriction of this isodose line and inadequate coverage at depth. For a higher TCF, a minimum air gap between the cone to the surface of the patient is desired. If such is not possible, then a tertiary collimator at the skin is required. Whenever, a tertiary collimator is used, it is advised to increase the collimator field size by a factor of 1.4.

Cell damage resulting from the labeling of rat lymphocytes and HeLa S3 cells with In-111 oxine.

Rat thoracic-duct lymphocytes and HeLa S3 cells were labeled in vitro with different amounts of indium-111 oxine. The labeled rat lymphocytes were tested for their ability to recirculate normally in syngeneic rats; the labeled HeLa S3 cells for their ability to divide to form colonies in tissue culture. Both cell types behaved normally by these criteria when labeled with small amounts of indium-111 oxine but at higher doses were obviously damaged. Evidence was obtained for the HeLa S3 cells that this damage was primarily radiation-induced. These findings may impose limitations on the use of In-111 oxine as a cell label for clinical purposes.

Low dose rate versus high dose rate intraluminal brachytherapy for malignant endobronchial tumors.

Although the evolution from low dose rate to high dose rate brachytherapy for malignant endobronchial malignancies was primarily based on economy, patient convenience, and radiation protection, the difference in therapeutic index, if any, between these two modalities must be kept in mind. Our experience with both methods permits assessment of the feasibility of replacing low dose rate brachytherapy with high dose rate brachytherapy. Results with our first 110 patients (group 1) treated with low dose rate brachytherapy (133 procedures) were compared with results with our initial 59 consecutive patients (group 2) treated with high dose rate brachytherapy (161 procedures). In group 1, patients were treated with one or two sessions of 30-60 Gy each calculated at a 1 cm radius. In patients in group 2, we aimed at three weekly sessions of 7 Gy each calculated at a 1 cm radius. External beam irradiation therapy had previously been given to 88% of patients in group 1 and to 85% of patients in group 2. Laser bronchoscopy was performed in 36% of patients in group 1 and in 24% of patients in group 2 before brachytherapy. Clinical or bronchoscopic improvement was noted in 72% of patients in group 1 and in 85% of patients in group 2 (p > 0.05). Complication rates were low and comparable. Survival was similar in both groups (median < 6 months). Although both low dose rate and high dose rate brachytherapy appear equally effective in palliation for malignant endobronchial obstruction, we are now practicing the latter exclusively.

The helper T lymphocyte precursor (HTLp) frequency does not predict outcome after HLA-identical sibling donor G-CSF-mobilised peripheral blood stem cell transplantation.

Here, we report the first study assessing the helper T lymphocyte precursor (HTLp) frequency as a predictor of outcome in patients undergoing allogeneic PBSC transplantation. The HTLp assay uses limiting dilution analysis to measure the frequency, in PBMCs from the donor, of T lymphocytes capable of producing IL-2 in response to histocompatibility antigenic differences on PBMCs from the recipient. This assay has shown promise as a functional histocompatibility assessment used to predict the risk of recipients of HLA-matched donor bone marrow developing severe acute GVHD: the higher the HTLp frequency, the greater the significance of any histoincompatibility, and the greater the risk of severe acute GVHD. In the current report, the HTLp frequency was measured in 28 HLA-identical sibling pairs who subsequently underwent allogeneic PBSC transplantation for haematological malignancies. The HTLp frequency did not predict for acute GVHD (P = 0.38), chronic GVHD (P = 0.95), transplant-related mortality (P = 0.79), relapse (P = 0.39) or overall survival (P = 0.84). Converting the HTLp frequency to HTLp infused per kilogram of recipient body weight also did not predict for any of the outcome measures. We conclude that, although the HTLp assay may be useful for patients undergoing BMT, it does not predict for outcome after HLA-identical sibling donor G-CSF-mobilised PBSC transplantation.

Improved detection of clinically significant host-reactive antigens prior to HLA-identical sibling peripheral blood stem cell transplantation using a dendritic cell-based helper T-lymphocyte precursor assay.

SUMMARY: Graft-versus-host disease (GVHD) due to host-reactive antigenic differences between HLA-identical pairs remains an important cause of morbidity and mortality after allogeneic transplantation. The helper T-lymphocyte precursor (HTLp) assay, using peripheral blood mononuclear cells (PBMCs), has been variably shown to detect such host-reactive differences. We assessed whether using dendritic cells (DCs) as the stimulator cells would improve the ability of the HTLp assay to detect these differences. We used PBMCs (standard HTLp assay) or monocyte-derived DCs (DC-HTLp assay) as the stimulator cells for 12 HLA-identical sibling pairs undergoing allogeneic peripheral blood stem cell transplantation. HTLp frequencies were greater by the DC-HTLp assay (median 1:77 712 vs 1:727 514; P=0.008). The standard HTLp assay did not predict for acute GVHD (P=0.42), whereas a trend was noted for the DC-HTLp assay (P=0.095). Of note, of seven patients developing moderately severe to severe acute GVHD, four had a significantly greater DC-HTLp frequency compared to the standard HTLp frequency, whereas all four patients who developed no to moderate acute GVHD had similar HTLp frequencies whether PBMCs or DCs were used as the stimulator cells. Although the small number of donor/recipient pairs assessed limits the strength of any conclusions, our study suggests that the DC-HTLp assay is better able to detect clinically significant host-reactive antigenic differences between HLA-identical siblings.

The inadequacy of rectal temperature measurements for assessing the effects of antiviral drugs on influenza virus infection of ferrets.

From 6 experiments in which 99 ferrets were infected with influenza virus A/Finland/74 and treated with various agents which suppress virus shedding and other parameters of infection, we assessed whether rectal temperature correlated with nasal virus shedding. A number of temperature and virus-shedding related parameters were determined for each experiment but statistical analysis showed little correlation between them, although an elevated temperature occurred at some time after infection. The pooled data also suggested that temperature and virus shedding parameters are not clearly related. The analysis indicates that intermittent rectal temperature measurements are unsatisfactory for determining the efficacy of anti-influenza agents in ferrets.

Papillary cystadenoma of epididymis: component of von Hippel-Lindau syndrome.

A case of epididymal cystadenoma, thought to be a component of the von Hippel-Lindau syndrome, is presented. The syndrome is reviewed, and the importance of long-term urologic follow-up for possible presentation of renal cell carcinoma is discussed.

Treatment of high-grade low-stage prostate cancer by high-dose-rate brachytherapy.

BACKGROUND AND PURPOSE: The best management of patients with low-stage, high-grade prostate cancer remains unclear. In an attempt to improve the outcomes of this high-risk group, we have offered those with Gleason > or =7 cancers removable-source high-dose-rate (HDR) brachytherapy in combination with external-beam radiation. PATIENTS AND METHODS: We reviewed the clinical histories of 61 consecutive patients with high-grade clinical stage T1-T2 lesions who received the combination radiation therapy between March 1997 and November 1998. The average Gleason score was 7.5. The HDR brachytherapy was given in three sessions with removable-source afterloaded (192)Ir to a minimum peripheral dose of 6 Gy. Conformal external-beam radiation in 25 fractions to a dose of 50 Gy was given beginning 1 week later. Patients with prostate volumes >40 cc received a luteinizing hormone-releasing hormone analog before brachytherapy. RESULTS: Among the 52 patients available for follow-up (average duration 11.8 months), there has been one death from prostate cancer. After treatment, only one patient had an initial rise in serum prostate specific antigen (PSA) concentration. In addition to the patient who died, there have been three confirmed treatment failures. Toxicity was mild, with only two patients having RTOG grade 3 or 4 effects. Neither of them required surgery. CONCLUSION: Although long-term results are not available, available data suggest that HDR brachytherapy plus external-beam radiation is at least as effective as any single therapy for high-risk, low-stage prostate cancer. The toxicity is acceptable.

Computing illumination-invariant descriptors of spatially filtered color image regions.

Spatial filters provide a useful and efficient means of analyzing an input color image into components that capture different spatial properties. Representations based on spatial filtering have restricted usefulness for recognition, however, because the output of a spatial filter across an image depends on the scene illumination conditions. We use a physically accurate linear model for spectral reflectance to derive invariants of distributions in spatially filtered color images that do not depend on the scene illumination. These invariants can be used for the illumination-invariant recognition of regions following an arbitrary linear filtering operation. We describe a method for illumination correction based on color distributions and introduce an illumination change consistency constraint that is useful for verifying matches obtained using the invariants. We show, using a set of classification experiments, that the filtered distribution invariants can significantly improve the capability of a recognition system in environments where illumination cannot be controlled.

An analytical and experimental study of the performance of Markov random fields applied to textured images using small samples.

We investigate to what extent textures can be distinguished using conditional Markov fields and small samples. We establish that the least square (LS) estimator is the only reasonable choice for this task, and we prove its asymptotic consistency and normality for a general class of random fields that includes Gaussian Markov fields as a special case. The performance of this estimator when applied to textured images of real surfaces is poor if small boxes are used (20x20 or less). We investigate the nature of this problem by comparing the behavior predicted by the rigorous theory to the one that has been experimentally observed. Our analysis reveals that 20x20 samples contain enough information to distinguish between the textures in our experiments and that the poor performance mentioned above should be attributed to the fact that conditional Markov fields do not provide accurate models for textured images of many real surfaces. A more general model that exploits more efficiently the information contained in small samples is also suggested.

Feature extraction for texture discrimination via random field models with random spatial interaction.

In this paper, we attack the problem of distinguishing textured images of real surfaces using small samples. We first analyze experimental data that results from applying ordinary conditional Markov fields. In the face of the disappointing performance of these models, we introduce a random field with spatial interaction that is itself a random variable (usually referred to as a random field in a random environment). For this class of models, we establish the power spectrum and the autocorrelation function as well-defined quantities, and we devise a scheme for the estimation of related parameters. The new set of features that resulted from this approach was applied to real images. Accurate discrimination was observed even for boxes of size 10x16.