β2 integrin-mediated cell-cell contact transfers active myeloperoxidase from neutrophils to endothelial cells

Atherosclerosis and vasculitis both feature inflammation mediated by neutrophil-endothelial-cell (EC) contact. Neutrophil myeloperoxidase (MPO) can disrupt normal EC function, although the mechanism(s) by which MPO is transferred to EC are unknown. We tested the hypothesis that close, beta2-integrin-dependent neutrophil-EC contact mediates MPO transfer from neutrophils to EC. We used sensitive MPO assays and flow cytometry to detect MPO in EC and demonstrate that EC acquired MPO when contacted by neutrophils directly but not when EC and neutrophils were separated in transwells. The transfer was dependent on neutrophil number, exposure time, and incubation temperature. Transfer occurred in several EC types, increased with endotoxin, was not accompanied by MPO release into the medium and was not abrogated by inhibiting degranulation to secretagogues. Confocal microscopy showed MPO internalization by EC with cytoplasmic and nuclear staining. Neutrophils and EC formed intimate contact sites demonstrated by electron microscopy. Blocking CD11b or CD18 beta2-integrin chains, or using neutrophils from CD11b gene-deleted mice, reduced MPO transfer. EC-acquired MPO was enzymatically active, as demonstrated by its ability to oxidize the fluorescent probe aminophenyl fluorescein in the presence of a hydrogen peroxide source. The data suggest an alternative to EC uptake of soluble MPO, namely the cell contact-dependent, {beta}2-integrin-mediated transfer from neutrophils. The findings could be of therapeutic relevance in atherosclerosis and vasculitis

The repertory of bone marrow progenitor cells associated with lymphogenic metastasis in patients with invasive carcinoma of no special type

The high mortality of patients with breast cancer is determined by metastatic disease. It is thought that the metastatic disease development associated with the repertory of bone marrow progenitor cells in breast cancer patients. In our study the correlation between the bone marrow progenitor cells presences in the tumor and blood of patients and the lymphogenic metastasis development was studied. The main clinical and pathological parameters of 24 patients with invasive breast carcinoma of non-specific type were analyzed. Endothelial progenitor cells, mesenchymal stem cells, macrophage precursors, hematopoietic progenitor cells were detected with specific antibodies against CD34, CD133, CD90, VEGFR1, CD11b, CD45, CD202 in the cell-rich fluid from frozen tumor. The amount of MCP-1 in the patients blood serum was assessed by enzymelinked immunosorbent assay (ELISA), at a wavelength of 450 nm. The cytokines concentration was calculated from the calibration plot. The program package Statistica 10.0. was used for statistical data processing. The high risk of lymphogenic metastasis in patients who didn't complete a neoadjuvant chemotherapy course was associated with the number of HPC, EPC and MSC in tumor and MCP-1 in blood

Competitively disrupting the neutrophil-specific receptor-autoantigen CD177:proteinase 3 membrane complex reduces anti-PR3 antibody-induced neutrophil activation

CD177 is a neutrophil-specific receptor presenting the proteinase 3 (PR3) autoantigen on the neutrophil surface. CD177 expression is restricted to a neutrophil subset, resulting in CD177(pos)/mPR3(high) and CD177(neg)/mPR3(low) populations. The CD177(pos)/mPR3(high) subset has implications for anti-neutrophil cytoplasmic autoantibody (ANCA)-associated autoimmune vasculitis (AAV), wherein patients harbor PR3-specific ANCAs that activate neutrophils for degranulation. Here we generated high-affinity anti-CD177 monoclonal antibodies, some of which interfered with PR3 binding to CD177 (PR3 "blockers") as determined by surface plasmon resonance spectroscopy, and used them to test the effect of competing PR3 from the surface of CD177(pos) neutrophils. Because intact anti-CD177 antibodies also caused neutrophil activation, we prepared non-activating Fab fragments of a PR3 blocker and non-blocker that bound specifically to CD177(pos) neutrophils. We observed that Fab blocker clone 40, but not non-blocker clone 80, dose-dependently reduced anti-PR3 antibody binding to CD177(pos) neutrophils. Importantly, preincubation with clone 40 significantly reduced respiratory burst in primed neutrophils challenged with either monoclonal antibodies to PR3 or PR3-ANCA IgG from AAV patients. After separating the two CD177/mPR3 neutrophil subsets from individual donors by magnetic sorting, we found that PR3-ANCAs provoked significantly more superoxide production in CD177(pos)/mPR3(high) than in CD177(neg)/mPR3(low) neutrophils, and that anti-CD177 Fab clone 40 reduced the superoxide production of CD177(pos) cells to the level of the CD177(neg) cells. Our data demonstrate the importance of the CD177:PR3 membrane complex in maintaining a high ANCA epitope density and thereby underscore the contribution of CD177 to the severity of PR3-ANCA diseases

Disrupting the CD177:proteinase 3 membrane complex reduces anti-PR3 antibody-induced neutrophil activation

CD177 is a neutrophil-specific receptor presenting proteinase 3 (PR3) autoantigen on the neutrophil surface. CD177 expression is restricted to a neutrophil subset resulting in CD177(pos)/mPR3(high) and CD177(neg)/mPR3(low) populations. The size of the CD177(pos)/mPR3(high) subset has implications for anti-neutrophil cytoplasmic autoantibody (ANCA)-associated autoimmune vasculitis (AAV) where patients harbor PR3-specific ANCA that activate neutrophils for degranulation. We generated high affinity anti-CD177 monoclonal antibodies, some of which interfered with PR3 binding to CD177 (PR3 "blockers") as determined by surface plasmon resonance spectroscopy, and used them to test the effect of competing PR3 from the surface of CD177(pos) neutrophils. Because intact anti-CD177 antibodies also caused neutrophil activation, we prepared non-activating Fab fragments of a PR3 blocker and non-blocker that bound specifically to CD177(pos) neutrophils by flow cytometry. We observed that Fab blocker clone 40, but not non-blocker clone 80, dosedependently reduced anti-PR3 antibody binding to CD177(pos) neutrophils. Importantly, preincubation with clone 40 significantly reduced respiratory burst in primed neutrophils challenged either with monoclonal antibodies to PR3 or PR3- ANCA IgG from AAV patients. After separating the two CD177/mPR3 neutrophil subsets from individual donors by magnetic sorting, we found that PR3-ANCA provoked significantly more superoxide production in CD177(pos)/mPR3(high) than in CD177(neg)/mPR3(low) neutrophils, and that anti- CD177 Fab clone 40 reduced the superoxide production of CD177(pos) cells to the level of the CD177(neg) cells. Our data demonstrate the importance of the CD177:PR3 membrane complex in maintaining a high ANCA epitope density and thereby underscore the contribution of CD177 to the severity of PR3-ANCA diseases

Correlation of the ratio of metastatic to non-metastatic cancer cases with the degree of socioeconomic deprivation among Texas counties

<p>Abstract</p> <p>Background</p> <p>Previous studies have demonstrated that cancer registrations and hospital discharge rate are closely correlated with census data-based socioeconomic deprivation indices. We hypothesized that communities with higher degrees of socioeconomic deprivation tend to have a higher ratio of metastatic to non-metastatic cancer cases (lung, breast, prostate, female genital system, colorectal cancers or all types of cancers combined). In this study, we investigate the potential link between this ratio and the Wellbeing Index (WI) among Texas counties.</p> <p>Results</p> <p>Cancer data in 2000 were provided by the Texas Cancer Registry, while data on the ten socioeconomic variables among the 254 Texas counties in 2000 for building the WI were obtained from U.S. Census Bureau. The ten socioeconomic status variables were subjected to the principal component analysis, and the first principal component scores were grouped into deciles for the WI (1 to 10) and the 254 Texas counties were classified into 10 corresponding groups. Weighted linear regression analyses and a Cochran-Armitage trend test were performed to determine the relationship between the ratio of age-adjusted metastatic to non-metastatic cancer incidence cases and WI. The ratios of metastatic to non-metastatic cases of female genital system cancer (<it>r</it>²= 0.84, <it>p </it>= 0.0002), all-type cancers (<it>r</it>²= 0.73, <it>p </it>= 0.0017) and lung cancer (<it>r</it>²= 0.54, <it>p </it>= 0.0156) at diagnosis were positively correlated with WI.</p> <p>Conclusions</p> <p>The ratios of metastatic to non-metastatic cases of all-type, female genital system and lung cancers at diagnosis were statistically correlated with socioeconomic deprivation. Potential mediators for the correlation warrant further investigation in order to reduce health disparities associated with socioeconomic inequality.</p

CSF2-dependent monocyte education in the pathogenesis of ANCA-induced glomerulonephritis

OBJECTIVES: Myeloid cell activation by antineutrophil cytoplasmic antibody (ANCA) is pivotal for necrotising vasculitis, including necrotising crescentic glomerulonephritis (NCGN). In contrast to neutrophils, the contribution of classical monocyte (CM) and non-classical monocyte (NCM) remains poorly defined. We tested the hypothesis that CMs contribute to antineutrophil cytoplasmic antibody-associated vasculitis (AAV) and that colony-stimulating factor-2 (CSF2, granulocyte-macrophage colony-stimulating factor (GM-CSF)) is an important monocyte-directed disease modifier.METHODS: Myeloperoxidase (MPO)-immunised MPO(-/-) mice were transplanted with haematopoietic cells from wild-type (WT) mice, C-C chemokine receptor 2 (CCR2)(-/-) mice to abrogate CM, or transcription factor CCAAT-enhancer-binding protein beta (C/EBPß)(-/-) mice to reduce NCM, respectively. Monocytes were stimulated with CSF2, and CSF2 receptor subunit beta (CSF2rb)-deficient mice were used. Urinary monocytes and CSF2 were quantified and kidney expression was analysed. CSF2-blocking antibody was used in the nephrotoxic nephritis (NTN) model. RESULTS: Compared with WT mice, CCR2(-/-) chimeric mice showed reduced circulating CM and were protected from NCGN. C/EBPß(-/-) chimeric mice lacked NCM but developed NCGN similar to WT chimeric mice. Kidney and urinary CSF2 were upregulated in AAV mice. CSF2 increased the ability of ANCA-stimulated monocytes to generate interleukin-1ß and to promote T17(H) effector cell polarisation. CSF2rb(-/-) chimeric mice harboured reduced numbers of kidney T17(H) cells and were protected from NCGN. CSF2 neutralisation reduced renal damage in the NTN model. Finally, patients with active AAV displayed increased urinary CM numbers, CSF2 levels and expression of GM-CSF in infiltrating renal cells. CONCLUSIONS: CMs but not NCMs are important for inducing kidney damage in AAV. CSF2 is a crucial pathological factor by modulating monocyte proinflammatory functions and thereby T17(H) cell polarisation

Stereotactically guided breast biopsy: a review

The aims of this review are to compare and contrast the available stereotactic equipment, and to describe the variety of needle types used and their affect on pathological results and subsequent patient management. Initial stereotactic devices were “added-on” to analogue mammography units and have been replaced by prone or ducubitus equipment using digital image acquisition. Biopsies use either 14-G core biopsy (CB) needles or vacuum-assisted biopsies (VAB). Vacuum-assisted biopsy systems consistently out-perform 14-G CB with reduced need for diagnostic or multi-treatment surgery. The false-negative rate is 8% for 14-G CB compared with 0.7% for VAB. There is a risk of underestimating the disease present for lesions of uncertain malignant potential (Cat B3) and suspicious of malignancy (Cat B4) results with 25% of patients with a B3 biopsy found to have cancer at subsequent surgery and 66% of those with a B4 biopsy. A CB diagnosis of in situ malignancy is upgraded to invasive disease at surgery in 15-36% of patients undergoing CB and of the order of 10% with VAB. A high degree of diagnostic accuracy and hence safe patient care can only be achieved by meticulous attention to technique and multi-disciplinary cooperation

Reliability and validity of needle biopsy evaluation of breast-abnormalities using the B-categorization – design and objectives of the Diagnosis Optimisation Study (DIOS)

<p>Abstract</p> <p>Background</p> <p>The planned nationwide implementation of mammography screening 2007 in Germany will increase the occurrence of mammographically detected breast abnormalities. These abnormalities are normally evaluated by minimal invasive core biopsy. To minimize false positive and false negative histological findings, quality assurance of the pathological evaluation of the biopsies is essential. Various guidelines for quality assurance in breast cancer diagnosis recommend applying the B-classification for histopathological categorization. However, to date there are only few studies that reported results about reliability and validity of B-classification. Therefore, objectives of our study are to determine the inter- and intraobserver variability (reliability study) and construct and predictive validity (validity study) of core biopsy evaluation of breast abnormalities. This paper describes the design and objectives of the DIOS Study.</p> <p>Methods/Design</p> <p>All consecutive asymptomatic and symptomatic women with breast imaging abnormalities who are referred to the University Hospital of Halle for core breast biopsy over a period of 24 months are eligible. According to the sample size calculation we need 800 women for the study. All patients in the study population underwent clinical and radiological examination. Core biopsy is performed by stereotactic-, ultrasound- or magnetic resonance (MR) guided automated gun method or vacuum assisted method. The histopathologic agreement (intra- and interobserver) of pathologists and the histopathologic validity will be evaluated. Two reference standards are implemented, a reference pathologist and in case of suspicious or malignant findings the histopathologic result of excision biopsy. Furthermore, a self administrated questionnaire which contains questions about potential risk factors of breast cancer, is sent to the participants approximately two weeks after core biopsy. This enables us to run a case-control-analysis (woman with breast cancer histological verified after excision are defined as cases, woman without malignant breast lesions are defined as controls) to investigate the predictive values of various risk factors on breast cancer risk.</p> <p>Conclusion</p> <p>The analysis of reliability and validity of the histopathological evaluation of core biopsy specimens of breast abnormalities is intended to provide important information needed for a high quality in breast cancer diagnostic and for planning of treatment strategies.</p