Soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) in neonatal sepsis.

BACKGROUND: Efficient and accurate diagnosis of neonatal sepsis is challenging. The potential impact for a reduction in morbidity and mortality as well as antibiotic usage has stimulated the ongoing search for biomarkers of early sepsis. The objective of this pilot study was to quantify the levels of sTREM-1 and correlate with blood cultures and inflammatory markers in neonates evaluated for sepsis. METHODS: Neonates with suspected sepsis were enrolled (n = 83; Preterm n = 35; Term n = 48). Routine bloods for sepsis evaluation were included and plasma sTREM-1 levels were quantified by ELISA. RESULTS: Term and preterm neonates (n = 83; Preterm n = 35; Term n = 48) were enrolled and 16 neonates had positive blood cultures (preterm n = 15; term n = 1). sTREM-1 levels were not significantly different in infants with culture-positive or culture-negative sepsis (356 ± 218 pg/mL and 385 ± 254 pg/mL respectively). The immature-to-total granulocyte (I/T) ratio showed a significant positive correlation with sTREM-1 in the preterm group with positive blood cultures. Additionally, sTREM-1 showed a positive correlation with CRP in the preterm group with negative blood cultures. CONCLUSIONS: sTREM-1 was associated with traditional markers of inflammation (I/T ratio and CRP). However, in this cohort sTREM-1 did not improve the early detection of neonatal culture-positive sepsis.

Feature fusion of Raman chemical imaging and digital histopathology using machine learning for prostate cancer detection.

The diagnosis of prostate cancer is challenging due to the heterogeneity of its presentations, leading to the over diagnosis and treatment of non-clinically important disease. Accurate diagnosis can directly benefit a patient's quality of life and prognosis. Towards addressing this issue, we present a learning model for the automatic identification of prostate cancer. While many prostate cancer studies have adopted Raman spectroscopy approaches, none have utilised the combination of Raman Chemical Imaging (RCI) and other imaging modalities. This study uses multimodal images formed from stained Digital Histopathology (DP) and unstained RCI. The approach was developed and tested on a set of 178 clinical samples from 32 patients, containing a range of non-cancerous, Gleason grade 3 (G3) and grade 4 (G4) tissue microarray samples. For each histological sample, there is a pathologist labelled DP-RCI image pair. The hypothesis tested was whether multimodal image models can outperform single modality baseline models in terms of diagnostic accuracy. Binary non-cancer/cancer models and the more challenging G3/G4 differentiation were investigated. Regarding G3/G4 classification, the multimodal approach achieved a sensitivity of 73.8% and specificity of 88.1% while the baseline DP model showed a sensitivity and specificity of 54.1% and 84.7% respectively. The multimodal approach demonstrated a statistically significant 12.7% AUC advantage over the baseline with a value of 85.8% compared to 73.1%, also outperforming models based solely on RCI and mean and median Raman spectra. Feature fusion of DP and RCI does not improve the more trivial task of tumour identification but does deliver an observed advantage in G3/G4 discrimination. Building on these promising findings, future work could include the acquisition of larger datasets for enhanced model generalization.

Investigating the role of the IGF axis as a predictor of biochemical recurrence in prostate cancer patients post-surgery.

BACKGROUND: Between 20% and 35% of prostate cancer (PCa) patients who undergo treatment with curative intent (ie, surgery or radiation therapy) for localized disease will experience biochemical recurrence (BCR). Alterations in the insulin-like growth factor (IGF) axis and PTEN expression have been implicated in the development and progression of several human tumors including PCa. We examined the expression of the insulin receptor (INSR), IGF-1 receptor (IGF-1R), PTEN, and AKT in radical prostatectomy tissue of patients who developed BCR post-surgery. METHODS: Tissue microarrays (TMA) of 130 patients post-radical prostatectomy (65 = BCR, 65 = non-BCR) were stained by immunohistochemistry for INSR, IGF-1R, PTEN, and AKT using optimized antibody protocols. INSR, IGF1-R, PTEN, and AKT expression between benign and cancerous tissue, and different Gleason grades was assessed. Kaplan-Meier survival curves were used to examine the relationship between proteins expression and BCR. RESULTS: INSR (P < 0.001), IGF-1R (P < 0.001), and AKT (P < 0.05) expression was significantly increased and PTEN (P < 0.001) was significantly decreased in cancerous versus benign tissue. There was no significant difference in INSR, IGF-1R, or AKT expression in the cancerous tissue of non-BCR versus BCR patients (P = 0.149, P = 0.990, P = 0.399, respectively). There was a significant decrease in PTEN expression in the malignant tissue of BCR versus non-BCR patients (P = 0.011). Combinational analysis of the tissue proteins identified a combination of decreased PTEN and increased AKT or increased INSR was associated with worst outcome. We found that in each case, our hypothesized worst group was most likely to experience BCR and this was significant for combinations of PTEN+INSR and PTEN+AKT but not PTEN+IGF-1R (P = 0.023, P = 0.028, P = 0.078, respectively). CONCLUSIONS: Low PTEN is associated with BCR and this association is strongly modified by high INSR and high AKT expression. Measurement of these proteins could help inform appropriate patient selection for postoperative adjuvant therapy and prevent BCR.

High dose statin prophylaxis in cardiopulmonary bypass related surgery: clinical utility.

BACKGROUND: Previous studies from our group demonstrated the anti-inflammatory properties of statins on cardiopulmonary bypass (CPB), through inhibition of neutrophil transendothelial migration. We sought to determine the utility of preoperative statin on patients undergoing cardiac surgery, to investigate any moderating effects on the systemic inflammatory response (SIRS) with CPB, and to evaluate any clinical impact on our patients. METHODS: This is a prospective, randomised controlled trial with national regulatory body approval. Eligible patients were already on oral statin therapy. They were then randomly assigned to either investigation arm (n = 15, atorvastatin 80 mg for 2 weeks before surgery) or control arm (n = 15, no change to current statin therapy). Blood and urine samples were collected at 3 timepoints. Postoperative clinical measures were documented. RESULTS: Patients in the investigation arm have significantly lower troponin level (p = 0.016), and lower level of urine neutrophil gelatinase-associated lipocalin (NGAL; p = 0.002); thus demonstrating a lesser degree of cardiac and renal injury in these patients. Higher level of Interleukin-8 (IL-8) at baseline (p = 0.036) and 4 h post cross-clamp removal (p = 0.035) in the investiation arm. A similar trend is also observed in Matrix Metalloproteinase-9 (MMP-9; p > 0.05). There were however no differences in clinical outcomes. CONCLUSIONS: Maximizing the dose of statin in patients waiting for cardiac surgery has measurable biological effects. There is evidence of less cardiac and renal damage. The use of preoperative statins and in particular, high dose preoperative statin therapy, may prove a useful new tool for optimal preparation of patients for cardiac surgery. TRIAL REGISTRATION: EudraCT no. 2012-003396-20 . Registered 05 November 2012.

Serial cytokine alterations and abnormal neuroimaging in newborn infants with encephalopathy.

AIM: Inflammatory cytokines may play a role in the final common pathway in the pathogenesis of hypoxic-ischaemic injury in experimental models. We aimed to profile the systemic pro-and anti-inflammatory response over the first week of life in term infants at risk of neonatal encephalopathy. METHOD: In a tertiary referral university neonatal intensive care unit, serial blood samples were analysed from 41 term infants (requiring resuscitation at birth) in this prospective observational pilot study. Serum levels of 10 pro-and anti-inflammatory cytokines were evaluated including interleukin(IL)-1α, IL-1ß, IL-6, IL-8, IL-10, tumour necrosis factor(TNF)-α, interferon (IFN)-γ, vascular endothelial growth factor (VEGF), granulocyte/colony-stimulating factor (G-CSF) and granulocyte macrophage/colony-stimulating factor (GM-CSF). RESULTS: Infants with neonatal encephalopathy and abnormal neuroimaging (n = 15) had significantly elevated granulocyte macrophage/colony-stimulating factor at 0-24 h and interleukin-8, interleukin-6 and interleukin-10 at 24-48 hour. Tumour necrosis factor-α and vascular endothelial growth factor levels were lower at 72-96 hour (p < 0.05). Significantly elevated levels of interleukin-10 were associated with mortality. CONCLUSION: Serum cytokine changes and innate immune dysregulation in the first week of life may be indicators of outcome in neonatal encephalopathy but require validation in larger studies.

Improving multivariable prostate cancer risk assessment using the Prostate Health Index.

OBJECTIVES: To analyse the clinical utility of a prediction model incorporating both clinical information and a novel biomarker, p2PSA, in order to inform the decision for prostate biopsy in an Irish cohort of men referred for prostate cancer assessment. PATIENTS AND METHODS: Serum isolated from 250 men from three tertiary referral centres with pre-biopsy blood draws was analysed for total prostate-specific antigen (PSA), free PSA (fPSA) and p2PSA. From this, the Prostate Health Index (PHI) score was calculated (PHI = (p2PSA/fPSA)*√tPSA). The men's clinical information was used to derive their risk according to the Prostate Cancer Prevention Trial (PCPT) risk model. Two clinical prediction models were created via multivariable regression consisting of age, family history, abnormality on digital rectal examination, previous negative biopsy and either PSA or PHI score, respectively. Calibration plots, receiver-operating characteristic (ROC) curves and decision curves were generated to assess the performance of the three models. RESULTS: The PSA model and PHI model were both well calibrated in this cohort, with the PHI model showing the best correlation between predicted probabilities and actual outcome. The areas under the ROC curve for the PHI model, PSA model and PCPT model were 0.77, 0.71 and 0.69, respectively, for the prediction of prostate cancer (PCa) and 0.79, 0.72 and 0.72, respectively, for the prediction of high grade PCa. Decision-curve analysis showed a superior net benefit of the PHI model over both the PSA model and the PCPT risk model in the diagnosis of PCa and high grade PCa over the entire range of risk probabilities. CONCLUSION: A logical and standardized approach to the use of clinical risk factors can allow more accurate risk stratification of men under investigation for PCa. The measurement of p2PSA and the integration of this biomarker into a clinical prediction model can further increase the accuracy of risk stratification, helping to better inform the decision for prostate biopsy in a referral population.

The analysis of serum response factor expression in bone and soft tissue prostate cancer metastases.

BACKGROUND: Castration-resistant prostate cancer (CRPC) represents a challenge to treat with no effective treatment options available. We recently identified serum response factor (SRF) as a key transcription factor in an in vitro model of castration resistance where we showed that SRF inhibition resulted in reduced cellular proliferation. We also demonstrated an association between SRF protein expression and CRPC in a cohort of castrate-resistant transurethral resections of the prostate (TURPS). The mechanisms regulating the growth of CRPC bone and visceral metastases have not been explored in depth due to the paucity of patient-related material available for analysis. In this study, we aim to evaluate SRF protein expression in prostate cancer (PCa) metastases, which has not previously been reported. METHODS AND RESULTS: We evaluated the nuclear tissue expression profile of SRF by immunohistochemistry in 151 metastatic sites from 42 patients who died of advanced PCa. No relationship between SRF nuclear expression and the site of metastasis was observed (P = 0.824). However, a negative association between SRF nuclear expression in bone metastases and survival from (a) diagnosis with PCa (P = 0.005) and (b) diagnosis with CRPC (P = 0.029) was seen. These results demonstrate that SRF nuclear expression in bone metastases is associated with survival, with patients with the shortest survival showing high SRF nuclear expression and patients with the longest survival having low SRF nuclear expression. CONCLUSION: Our study indicates that SRF is a key factor determining patients' survival in metastatic CRPC and therefore may represent a promising target for future therapies.

EuroSCORE and neutrophil adhesion molecules predict outcome post-cardiac surgery.

BACKGROUND: This study tested the hypothesis that surgical stress and the host response to this trauma trigger an inflammatory cascade in which the neutrophil plays a central role. We hypothesised that pre-operative neutrophil migratory responses will correlate with post-operative clinical outcome in our shock model of open-heart surgery patients. We also tested the hypothesis that surface expression of adhesion molecules involved in the migratory process - CD11b, CD47 and CD99 - could be used to predict outcome. We believe that combining neutrophil migratory response, CD11b, CD47 and CD99 with the logistic European System for Cardiac Operative Risk Evaluation (EuroSCORE) will strengthen the power of the EuroSCORE not only in predicting post-operative mortality but also other clinical endpoints. MATERIALS AND METHODS: Neutrophils were isolated pre-operatively from n = 31 patients undergoing open-heart surgery and allowed to migrate across endothelial monolayers in response to N-formyl-methionine-leucine-phenylalanine (fMLP). Isolated neutrophils were also assessed for surface expression of CD11b, CD47 and CD99 in response to fMLP by flow cytometry. Post-operative clinical parameters collected included days 1-5 white cell count and creatinine levels as well as intensive care unit (ICU) and post-operative hospital stay. RESULTS: Pre-operative surface expression of CD99 and CD47 correlates with post-operative creatinine levels (P < 0·05), a measurement of renal injury. We also show that while the logistic EuroSCORE alone can be used as a predictor of ICU stay, when combined with pre-operative CD99 surface expression, it improves its AUC value (0·794). CONCLUSION: Immunological markers, specifically the ability of the neutrophil to migrate, combined with the logistic EuroSCORE lead to improved sensitivity and specificity to predict patient outcome.

Characterisation and manipulation of docetaxel resistant prostate cancer cell lines.

BACKGROUND: There is no effective treatment strategy for advanced castration-resistant prostate cancer. Although Docetaxel (Taxotere®) represents the most active chemotherapeutic agent it only gives a modest survival advantage with most patients eventually progressing because of inherent or acquired drug resistance. The aims of this study were to further investigate the mechanisms of resistance to Docetaxel. Three Docetaxel resistant sub-lines were generated and confirmed to be resistant to the apoptotic and anti-proliferative effects of increasing concentrations of Docetaxel. RESULTS: The resistant DU-145 R and 22RV1 R had expression of P-glycoprotein and its inhibition with Elacridar partially and totally reversed the resistant phenotype in the two cell lines respectively, which was not seen in the PC-3 resistant sublines. Resistance was also not mediated in the PC-3 cells by cellular senescence or autophagy but multiple changes in pro- and anti-apoptotic genes and proteins were demonstrated. Even though there were lower basal levels of NF-κB activity in the PC-3 D12 cells compared to the Parental PC-3, docetaxel induced higher NF-κB activity and IκB phosphorylation at 3 and 6 hours with only minor changes in the DU-145 cells. Inhibition of NF-κB with the BAY 11-7082 inhibitor reversed the resistance to Docetaxel. CONCLUSION: This study confirms that multiple mechanisms contribute to Docetaxel resistance and the central transcription factor NF-κB plays an immensely important role in determining docetaxel-resistance which may represent an appropriate therapeutic target.

Benefits of heat treatment to the protease packed neutrophil for proteome analysis: halting protein degradation.

Neutrophils, cells of the innate immune system, contain an array of proteases and reactive oxygen species-generating enzymes that assist in controlling the invasion of bacteria and pathogens. The high content of intracellular proteolytic enzymes makes them difficult cells to work with as they can degrade proteins of potential interest. Here, we describe the benefits of heat treatment of neutrophils in reducing protein degradation for subsequent proteome analysis. Neutrophils isolated from four healthy volunteers were each divided into three aliquots and subjected to different preparation methods for 2-DE: (i) Heat treatment, (ii) resuspension in NP40 lysis buffer and (iii) resuspension in standard 2-DE lysis buffer. Representative spots found to be statistically significant between groups (p<0.01) were excised and identified by LC-MS/MS, three of which were validated by immunoblotting. Heat-treated samples contained proteins in the high-molecular-weight range that were absent from NP40-treated samples. Moreover, NP40-treated samples showed an increase in spot number and volume at lower molecular weights suggestive of protein degradation. Incorporating heat treatment into sample preparation resulted in the identification of proteins that may not have previously been detected due to sample degradation, thus leading to a more comprehensive 2-DE map of the human neutrophil proteome.

Biomarker research in prostate cancer--towards utility, not futility.

The identification of an appropriate clinical question is critical for any biomarker project. Despite rapid advances in technology, few biomarkers have been forthcoming for prostate cancer. This could be because the clinical questions under investigation have not actually originated from clinical practice. These clinical questions are difficult to identify in the complex and heterogeneous pathogenesis of prostate cancer. In this Review, we have developed a prostate cancer 'roadmap' to identify the aspects of prostate cancer that may be amenable to biomarker discovery and serve as a guide for future projects in prostate cancer biomarker research.

Ensemble based system for whole-slide prostate cancer probability mapping using color texture features.

We present a tile-based approach for producing clinically relevant probability maps of prostatic carcinoma in histological sections from radical prostatectomy. Our methodology incorporates ensemble learning for feature selection and classification on expert-annotated images. Random forest feature selection performed over varying training sets provides a subset of generalized CIEL*a*b* co-occurrence texture features, while sample selection strategies with minimal constraints reduce training data requirements to achieve reliable results. Ensembles of classifiers are built using expert-annotated tiles from training images, and scores for the probability of cancer presence are calculated from the responses of each classifier in the ensemble. Spatial filtering of tile-based texture features prior to classification results in increased heat-map coherence as well as AUC values of 95% using ensembles of either random forests or support vector machines. Our approach is designed for adaptation to different imaging modalities, image features, and histological decision domains.

Applying random forests to identify biomarker panels in serum 2D-DIGE data for the detection and staging of prostate cancer.

In recent years, Prostate Specific Antigen (PSA) testing is widespread and has been associated with deceased mortality rates; however, this testing has raised concerns of overdiagnosis and overtreatment. It is clear that additional biomarkers are required. To identify these biomarkers, we have undertaken proteomics and metabolomics expression profiles of serum samples from BPH, Gleason score 5 and 7 using two-dimensional difference in gel electrophoresis (2D-DIGE) and nuclear magnetic resonance spectroscopy (NMR). Panels of serum protein biomarkers were identified by applying Random Forests to the 2D-DIGE data. The evaluation of selected biomarker panels has shown that they can provide higher prediction accuracy than the current diagnostic standard. With careful validation of these serum biomarker panels, these panels may potentially help to reduce unnecessary invasive diagnostic procedures and more accurately direct the urologist to curative surgery.

Effects of titanocene dichloride derivatives on prostate cancer cells, specifically DNA damage-induced apoptosis.

BACKGROUND: While locally advanced prostate cancer is initially treatable with androgen ablation, eventually cells develop a castrate-resistant phenotype. Currently, there are no effective treatments for this form of the disease with Docetaxel only providing a small survival advantage. In this study, the effects of novel derivatives of titanocene dichloride on prostate cancer cell lines has been investigated. METHODS: Cellular effects were assessed using the crystal violet assay and the clonogenic survival assay. Cell cycle and apoptosis were assessed by propidium iodide staining. DNA damage was analyzed by comet assay and Western analysis. DNA damage response inhibition was achieved by pre-incubation with an ATM/ATR inhibitor; CGK733 and DNA-PK inhibitor; DMNB. RESULTS: These analogs caused a reduction in cell number. In particular titanocene Y and C had significant effects in all cell lines. A reduction in clonogenic survival was found in response to titanocene Y in three cell lines while the PC-3 cells exhibited increased resistance.Further analysis showed no effect on cell cycle however, the analogs were found to induce apoptosis in a dose-dependent manner in all cell lines. These analogs associate with DNA, induce DNA damage and a differential damage response. Inhibition of key regulators of this DNA damage response sensitized the PC-3 cell line to titanocene-induced apoptosis and significantly reduced the clonogenic capacity of the cells. CONCLUSION: These results demonstrate the mechanism of action of these novel titanocene dichloride analogs and their potential use in castrate-independent advanced prostate cancer.

Core fucosylation and alpha2-3 sialylation in serum N-glycome is significantly increased in prostate cancer comparing to benign prostate hyperplasia.

One of the most urgent requirements in prostate cancer diagnosis is the development of a blood-based test which would be able to distinguish prostate cancer from benign prostate hyperplasia (BPH). Previously published results found a significant difference between specific glycan levels in patients with advanced prostate cancer and healthy controls. N-Glycans from the whole serum glycoproteins were measured using our fully quantitative high-throughput N-glycan analysis in combination with exoglycosidase digestions in sera from 13 BPH and 34 prostate cancer samples (17 Gleason score 5 and 17 Gleason score 7). The levels of core-fucosylated biantennary glycans and α2-3-linked sialic acids were significantly increased in prostate cancer patients compared with patients with BPH. Triantennary trigalactosylated glycans and tetraantennary tetrasialylated glycans with outer arm fucose were significantly decreased, and tetraantennary tetrasialylated glycans increased in Gleason 7 compared with Gleason 5. All these glycans can distinguish prostate cancer patients from BPH or Gleason 7 from Gleason 5 prostate cancer patients better than the current clinical test, prostate-specific antigen; therefore, their measurement may provide a new noninvasive approach to diagnose prostate cancer. However, additional validation studies would need to be carried out to further support this finding. Decreases in triantennary trigalactosylated glycans and/or bisected core-fucosylated biantennary monosialylated glycans and increases in tetraantennary tetrasialylated glycans correlate with perineural invasion, which could further help to diagnose tumor spread and predict patients' survival.

The epigenome as a therapeutic target in prostate cancer.

During cancer development and progression, tumor cells undergo abnormal epigenetic modifications, including DNA methylation, histone deacetylation and nucleosome remodeling. Collectively, these aberrations promote genomic instability and lead to silencing of tumor-suppressor genes and reactivation of oncogenic retroviruses. Epigenetic modifications, therefore, provide exciting new avenues for prostate cancer research. Promoter hypermethylation is widespread during neoplastic transformation of prostate cells, which suggests that restoration of a 'normal' epigenome through treatment with inhibitors of the enzymes involved could be clinically beneficial. Global patterns of histone modifications are also being defined and have been associated with clinical and pathologic predictors of prostate cancer outcome. Although treatment for localized prostate cancer can be curative, the development of successful therapies for the management of castration-resistant metastatic disease is urgently needed. Reactivation of tumor-suppressor genes by demethylating agents and histone deacetylase inhibitors could be a potential treatment option for patients with advanced disease.

Integrating biobanks: addressing the practical and ethical issues to deliver a valuable tool for cancer research.

Cancer is caused by complex interactions between genes, environment and lifestyles. Biobanks of well-annotated human tissues are an important resource for studying the underlying mechanisms of cancer. Although such biobanks exist, their integration to form larger biobanks is now required to provide the diversity of samples that are needed to study the complexity and heterogeneity of cancer. Clear guidelines and policies are also required to address the challenges of integrating individual institutional or national biobanks and build public trust. This Science and Society article highlights some of the main practical and ethical issues that are undergoing discussion in the integration of tissue biobanks for cancer.

Prostate cancer grading: the effect of stratification of needle biopsy Gleason Score 4 + 3 as high or intermediate grade.

OBJECTIVES: To assess the discrepancy between needle biopsy (NB) and radical prostatectomy (RP) Gleason score (GS) in Irish men, specifically the influence of the stratification of GS 4 + 3 on overall levels of agreement, levels of discrepancy and kappa coefficients, as the GS assigned to prostate cancer NBs affects clinical decision-making and influences future therapeutic strategies. PATIENTS AND METHODS: We reviewed retrospectively a database of the discrepancies between NB and RP Gleason grades (GG) from 2003 to 2008. All patients had clinically localized prostate cancer, and none had had neoadjuvant therapy. Grading of 206 NB specimens was compared with their corresponding RP specimens. The discrepancy rate between NB and RP GS was assessed for each combination of GG. Intermediate- (GS 7, defined as GS 3 + 4 alone vs GS 7) and high-grade (GS 4 + 3 and GS 8-10 vs GS 8-10) classifications were compared. The level of agreement and the kappa coefficient for each system was assessed. RESULTS: In NB, GS 6 was most frequently diagnosed (53%); after RP, GS 3 + 4 was most frequent (36%). In 42% of cases the exact GG remained unchanged after RP, increasing to 48% for GS 6 and GS 3 + 4. Overall 42% of cases showed an increase in their GG. In GS 6 NBs, the rate of increase in the primary GG or increase in the GS was 52%. Biopsy GS 6 and 3 + 4 showed the highest levels of agreement between NB and RP. Low-grade prostate cancer on NB was upgraded in 52% of cases; high-grade prostatic adenocarcinoma was downgraded in 27-77% of cases depending on the grading system used. CONCLUSIONS: Classification of high-grade prostate cancer as GS 4 + 3 and GS 8-10 results in higher levels of agreement between NB and RP GS. Reliable identification of well differentiated prostatic adenocarcinoma in NB specimens represents an ongoing diagnostic challenge, necessitating careful preoperative consideration of the definitive grade of a patient's disease.

Pre-operative determination of an individual's neutrophil response: a potential predictor of early cardiac transplant cellular rejection.

BACKGROUND: Success of heart transplantation is affected by allograft rejection, which is known as a lymphocyte-mediated process. Recent studies suggest that neutrophils contribute to this process. We hypothesized that the severity of cardiac rejection can be predicted by evaluating an individual's neutrophil transendothelial migration potential, which can be assessed through artificial stimulation of neutrophils. METHODS: Eleven patients were recruited from the active heart transplant list. Pre-operative blood samples were stained with neutrophil adhesion molecule (CD11b, CD62L, and PSGL-1) antibodies before and after in vitro stimulation with phorbol 12-myristate 13-acetate (PMA; 1 ng/ml) and lipopolysaccharide (LPS 1 microg/ml), and assessed using flow cytometry. Stimulated neutrophil responses were correlated with rejection grade of the first endomyocardial biopsy sampled 10 days post-operatively. RESULTS: Neutrophil adhesion molecules are upregulated after artificial stimulation. Pre-operative neutrophil surface CD11b expression after in vitro lipopolysaccharide stimulation correlated with rejection grade detected in the first endomyocardial biopsy sample (R = 0.677; p = 0.022). CONCLUSION: Pre-operative neutrophil response to in vitro stimuli predicted the rejection grade in the first post-transplant endomyocardial biopsy specimen, suggesting that neutrophils may contribute more to cardiac allograft rejection than previously thought.

digeR: a graphical user interface R package for analyzing 2D-DIGE data.

UNLABELLED: 2D Difference In-Gel Electrophoresis (2D-DIGE) or 2D gel technology is being used as a routine proteomics technique for biomarker discovery. Analyzing such high-dimensional data requires multivariate analysis techniques to be applied. In addition, protein post-translational modification (PTM) information from the 2D gel data is usually overlooked. We report on an R package, digeR, with an easy to use graphical user interface for analyzing 2D-DIGE (2D gel) data. It provides a tool for visually looking for potential PTM changes from different biological states and support biomarker discovery through multivariate analysis techniques. AVAILABILITY: digeR package is freely available from the CRAN: http://cran.r-project.org/web/packages/digeR/index.html.