Patient-reported outcome measures for physical function in cancer patients: content comparison of the EORTC CAT Core, EORTC QLQ-C30, SF-36, FACT-G, and PROMIS measures using the International Classification of Functioning, Disability and Health.

BACKGROUND: Patient-reported physical function (PF) is a key endpoint in cancer clinical trials. Using complex statistical methods, common metrics have been developed to compare scores from different patient-reported outcome (PRO) measures, but such methods do not account for possible differences in questionnaire content. Therefore, the aim of our study was a content comparison of frequently used PRO measures for PF in cancer patients. METHODS: Relying on the framework of the International Classification of Functioning, Disability and Health (ICF) we categorized the item content of the physical domains of the following measures: EORTC CAT Core, EORTC QLQ-C30, SF-36, PROMIS Cancer Item Bank for Physical Function, PROMIS Short Form for Physical Function 20a, and the FACT-G. Item content was linked to ICF categories by two independent reviewers. RESULTS: The 118 items investigated were assigned to 3 components ('d - Activities and Participation', 'b - Body Functions', and 'e - Environmental Factors') and 11 first-level ICF categories. All PF items of the EORTC measures but one were assigned to the first-level ICF categories 'd4 - Mobility' and 'd5 - Self-care', all within the component 'd - Activities and Participation'. The SF-36 additionally included item content related to 'd9 - Community, social and civic life' and the PROMIS Short Form for Physical Function 20a also included content related to 'd6 - domestic life'. The PROMIS Cancer Item Bank (v1.1) covered, in addition, two first-level categories within the component 'b - Body Functions'. The FACT-G Physical Well-being scale was found to be the most diverse scale with item content partly not covered by the ICF framework. DISCUSSION: Our results provide information about conceptual differences between common PRO measures for the assessment of PF in cancer patients. Our results complement quantitative information on psychometric characteristics of these measures and provide a better understanding of the possibilities of establishing common metrics.

Diagnosed with a common cancer at an unusual age: causal attributions of survivors of adolescent and young adult colorectal cancer.

BACKGROUND: Insight into the causes of colorectal cancer (CRC) in adolescent and young adult (AYA) patients is limited. Without definitive information, patients often shape their own beliefs on the cause of their illness, developing causal attributions. This study aims to examine the causal attributions of CRC in AYA patients, compare these with middle-aged and older adults CRC patient groups and explore the association between causal attributions and psychological distress. METHODS: Two cross-sectional questionnaire studies were conducted among CRC survivors diagnosed between 1998 and 2007 using the population-based PROFILES registry. Three groups were defined by age at diagnosis: AYA (18-39 years; n = 67), middle-aged (40-70 years; n = 1993) and older adult survivors (70+ years; n = 1922). Causal attributions were assessed in a single free-text item from the Brief Illness Perception Questionnaire and psychological distress measured by the Hospital Anxiety and Depression Scale. RESULTS: AYA survivors most often attributed their CRC to heredity (36.4%), lifestyle (14.5%) and chance (10.9%). AYA survivors attributed their CRC to these causes more frequently than middle-aged and older adult CRC survivors. AYA survivors who attributed their CRC to causes out of their control (chance, heredity) showed significantly lower mean scores on anxiety (4.3 vs. 5.6; p < 0.01) compared to AYAs who reported causes within their control (lifestyle, stress). No significant difference was found for depression. CONCLUSION: AYA patients with CRC may benefit from in-depth discussion about the lack of known (biological) causes and how this does not directly imply a lifestyle or stress cause.

Beyond Teenage and Young Adult Cancer Care: Care Experiences of Patients Aged 25-39 Years Old in the UK National Health Service.

AIMS: Adolescents and young adults aged 15-39 years with cancer face unique medical, practical and psychosocial issues. In the UK, principal treatment centres and programmes have been designed to care for teenage and young adult patients aged 13-24 years in an age-appropriate manner. However, for young adults (YAs) aged 25-39 years with cancer, little access to age-specific support is available. The aim of this study was to examine this possible gap by qualitatively exploring YA care experiences, involving patients as research partners in the analysis to ensure robust results. MATERIALS AND METHODS: We conducted a phenomenological qualitative study with YAs diagnosed with any cancer type between ages 25 and 39 years old in the last 5 years. Participants took part in interviews or focus groups and data were analysed using inductive thematic analysis. Results were shaped in an iterative process with the initial coders and four YA patients who did not participate in the study to improve the rigor of the results. RESULTS: Sixty-five YAs with a range of tumour types participated. We identified seven themes and 13 subthemes. YAs found navigating the healthcare system difficult and commonly experienced prolonged diagnostic pathways. Participants felt under-informed about clinical details and the long-term implications of side-effects on daily life. YAs found online resources overwhelming but also a source of information and treatment support. Some patients regretted not discussing fertility before cancer treatment or felt uninformed or rushed when making fertility preservation decisions. A lack of age-tailored content or age-specific groups deterred YAs from accessing psychological support and rehabilitation services. CONCLUSIONS: YAs with cancer may miss some benefits provided to teenagers and young adults in age-tailored cancer services. Improving services for YAs in adult settings should focus on provision of age-specific information and access to existing relevant support.

Statin-mediated cytoprotection of human vascular endothelial cells: a role for Kruppel-like factor 2-dependent induction of heme oxygenase-1.

BACKGROUND: Heme oxygenase-1 (HO-1), by exerting anti-inflammatory, antiproliferative, antiapoptotic and antioxidant effects in the vasculature, protects against atherosclerosis and post-transplant vasculopathy. We noted the overlap between the effects of HO-1 and those attributed to 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins). This led to an investigation of the role of HO-1 in statin-mediated cytoprotection in primary human endothelial cells (ECs), and the ability of Kruppel-like factor 2 (KLF2) to regulate HO-1 function. METHODS/RESULTS: Treatment of human umbilical vein and aortic ECs with atorvastatin significantly upregulated HO-1 promoter activity, mRNA expression and protein expression, increasing HO-1 enzymatic activity as shown by raised intracellular bilirubin IXalpha. This effect was indirect, dependent upon inhibition of HMG-CoA reductase and geranylgeranylation, and independent of nitric oxide or changes in mRNA stability. Atorvastatin protected ECs against the generation of reactive oxygen species and H(2)O(2)-induced injury. HO-1 inhibition, with small interfering RNA (siRNA) or zinc protoporphyrin IX, abrogated atorvastatin-mediated cytoprotection. Atorvastatin upregulated KLF2 expression, whereas KLF2 siRNA attenuated statin-induced HO-1 and its associated antioxidant cytoprotective effects. Iron chelation, adenoviral-mediated overexpression of ferritin or supplementation of culture media with biliverdin reversed the inhibitory effects of HO-1 and KLF2 siRNA, suggesting that bile pigments and ferritin mediate the antioxidant actions of statin-induced HO-1. CONCLUSIONS: We have identified a novel link between KLF2 and HO-1 in human vascular ECs, demonstrating that atorvastatin-mediated HO-1 upregulation, and its associated antioxidant effect, is KLF2-dependent. The relationship between KLF2 and HO-1 is likely to represent an important component of the vasculoprotective profile of statins.

Isolation of endothelial cells from murine tissue.

The isolation and long-term culture of murine endothelial cells (ECs) has often proven a difficult task. In this paper we describe a quick, efficient protocol for the isolation of microvascular endothelial cells from murine tissues. Murine lung or heart are mechanically minced and enzymatically digested with collagenase and trypsin. The single cell suspension obtained is then incubated with an anti-CD31 antibody, anti-CD105 antibody and with biotinylated isolectin B-4. Pure EC populations are finally obtained by magnetic bead separation using rat anti-mouse Ig- and streptavidin-conjugated microbeads. EC cultures are subsequently expanded and characterised. The surface molecule expression by the primary cultures of murine EC obtained from lung and heart tissue is analysed and compared to that of a murine endothelioma and of primary cultures of murine renal tubular epithelial cells. The phenotype and morphology of these cultures remain stable over 10-15 passages in culture, and no overgrowth of contaminating cells of non-endothelial origin is observed at any stage.

Effect of aprotinin on endothelial cell activation.

BACKGROUND: Cardiopulmonary bypass surgery is often accompanied by a systemic inflammatory response, which can lead to postoperative complications in high-risk patients. This is mediated in part through a systemic rise in inflammatory cytokine levels and the sequestration of leukocytes within organs. Aprotinin has previously been shown to exert an anti-inflammatory effect by preventing the capacity of leukocytes to transmigrate through vascular endothelium. Here we have focused on whether aprotinin has an effect on endothelial cell activation and adhesion molecule expression in response to tumor necrosis factor-alpha, particularly with reference to whether aprotinin inhibits tumor necrosis factor-stimulated neutrophil transendothelial migration. METHODS AND RESULTS: Intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and E-selectin expression was studied in tumor necrosis factor-alpha-activated human umbilical vein endothelial cells in the presence of aprotinin at 200, 800, and 1600 kIU/mL. Aprotinin inhibited tumor necrosis factor-alpha-stimulated expression of intercellular adhesion molecule-1 (P =.019 at 1600 kIU/mL) and vascular cell adhesion molecule-1 (P =.003 at 1600 kIU/mL) but not E-selectin. Similar results were obtained in the dermal microvascular endothelial cell line, HMEC-1, which exhibited diminished intercellular adhesion molecule-1 expression in the presence of aprotinin (P =.040 at 800 kIU/mL and P <.001 at 1600 kIU/mL). Aprotinin also significantly inhibited neutrophil transmigration across tumor necrosis factor-alpha-activated human umbilical vein endothelial cells (P =.046 at 1600 kIU/mL). CONCLUSIONS: We have demonstrated that aprotinin inhibits intercellular adhesion molecule-1 and vascular cell adhesion molecule-1, but not E-selectin, expression on tumor necrosis factor-alpha-activated endothelial cells and that transendothelial migration by neutrophils is also specifically suppressed under these conditions. Our results indicate that endothelial cells can be specifically targeted by aprotinin, therefore adding to our understanding of the anti-inflammatory mechanism of action of aprotinin during cardiopulmonary bypass.

An anti-inflammatory property of aprotinin detected at the level of leukocyte extravasation.

BACKGROUND: Aprotinin is a serine protease inhibitor used extensively in cardiac operations to reduce postoperative bleeding. It has also been used in trials aimed at reducing the systemic inflammatory response to cardiopulmonary bypass. It remains unclear whether the anti-inflammatory action of aprotinin is related to its general ability to suppress leukocyte activation or whether aprotinin can exercise effects during the leukocyte-endothelial cell adhesion cascade. METHODS: We used intravital microscopy to study the 3 main stages of the adhesion cascade (leukocyte rolling, firm adhesion, and extravasation) within the mesenteric microcirculation of rats. This in vivo technique allows leukocyte recruitment to be viewed directly through the transparent mesentery of anesthetized animals. RESULTS: Aprotinin, given by continuous infusion at a clinically relevant dose, exerted no effect on the rolling or firm adhesion responses toward local chemoattractant N -formyl-methyl-leucyl-phenylalanine but significantly inhibited extravasation of leukocytes (73% at 40 minutes, P =.04) into surrounding tissues. In parallel in vitro experiments, aprotinin (used at 200, 800, and 1600 kIU/mL) dose dependently inhibited neutrophil transmigration through cultured endothelial cells in response to 3 different chemoattractants: N -formyl-methyl-leucyl-phenylalanine (P <.001 at 800 and 1600 kIU/mL), interleukin 8 (P <.05 at 200 kIU/mL and P <.001 at 800 and 1600 kIU/mL), and platelet-activating factor (P <.05 at 1600 kIU/mL). CONCLUSIONS: Our studies have therefore revealed a novel anti-inflammatory mechanism of aprotinin operating at the level of leukocyte extravasation. These findings may be relevant in the prevention of systemic inflammation after cardiopulmonary bypass through the use of protease inhibitors.

A comparison of primary endothelial cells and endothelial cell lines for studies of immune interactions.

The purpose of this study was to assess the suitability of using endothelial cell (EC) lines for studies of endothelial/immune interactions. The immortal human EC lines HMEC-1, ECV304 and EaHy926 were compared to human umbilical vein endothelial cells (HUVEC) for constitutive and induced expression of surface antigens known to be involved in interactions with T cells. These cell lines were also compared to HUVEC in transendothelial migration assays. Flow cytometry was used to measure cell surface expression of platelet/endothelial cell adhesion molecule-1 (PECAM-1), intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), E-selectin, major histocompatibility complex (MHC) class I and MHC class II, CD40, CD95 (fas) and lymphocyte function associated antigen-3 (LFA-3) before and after treatment with the cytokines tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). Polymerase chain reaction (PCR) was used to detect expression of the MHC class II transactivator. Significant differences were found in the ability to respond to cytokines between HUVEC and the cell lines, the greatest differences being induction of VCAM-1 and E-selectin in response to TNF-alpha and induction of MHC class II antigens in response to IFN-gamma. Thus unlike HUVEC, induction of VCAM-1 and E-selectin was not detectable on EaHy926 and ECV304 and barely detectable on HMEC-1. MHC class II antigens were not induced on ECV304 in response to IFN-gamma and nor was the class II transactivator (CIITA). Unlike HUVEC and the other cell lines, ECV304 were constitutively negative for PECAM-1. Constitutive and induced expression of MHC class I, ICAM-1, LFA/3, CD40 and fas were most conserved between the cell lines and showed little difference to HUVEC. The migration of peripheral blood mononuclear cells (PBMC) through all cell lines was significantly reduced compared to through HUVEC, suggesting that there is a functional difference between the cell lines with regard to interactions with lymphocytes. In conclusion this study has demonstrated significant differences in the ability of endothelial cell lines to respond to cytokines compared to primary HUVEC cultures. In particular ECV304 compares very poorly with HUVEC. Whether these differences are caused by immortalization procedures or reflect heterogeneity of EC arising from different vascular beds is discussed.

Induction of decay-accelerating factor by thrombin through a protease-activated receptor 1 and protein kinase C-dependent pathway protects vascular endothelial cells from complement-mediated injury.

There is increasing evidence for functional crosstalk between inflammatory and thrombotic pathways in inflammatory vascular diseases such as atherosclerosis and vasculitis. Thus, complement activation on the endothelial cell (EC) surface during inflammation may generate thrombin via the synthesis of tissue factor. We explored the hypothesis that thrombin induces EC expression of the complement-regulatory proteins decay-accelerating factor (DAF), membrane cofactor protein (MCP), and CD59 and that this maintains vascular integrity during coagulation associated with complement activation. Thrombin increased DAF expression on the surface of ECs by 4-fold in a dose- and time-dependent manner as measured by flow cytometry. DAF up-regulation was first detectable at 6 hours and maximal 24 hours poststimulation, whereas no up-regulation of CD59 or MCP was seen. Thrombin-induced expression required increased DAF messenger RNA and de novo protein synthesis. The response depended on activation of protease-activated receptor 1 (PAR1) and was inhibited by pharmacologic antagonists of protein kinase C (PKC), p38 and p42/44 mitogen-activated protein kinase, and nuclear factor-kappa B. The increased DAF expression was functionally relevant because it significantly reduced C3 deposition and complement-mediated EC lysis. Thus, thrombin-generated at inflammatory sites in response to complement activation-is a physiologic agonist for the PKC-dependent pathway of DAF regulation, thereby providing a negative feedback loop protecting against thrombosis in inflammation. (Blood. 2000;96:2784-2792)

Inhibition of the transendothelial migration of human lymphocytes but not monocytes by phosphodiesterase inhibitors.

This study describes an in vitro model of peripheral blood mononuclear cell (PBMC) migration through human endothelial cells, held on polycarbonate inserts, which allows automatic differential counting of migrated cells as lymphocytes and monocytes. Using this system it was found that treatment of PBMC with the phosphodiesterase (PDE) inhibitors theophylline (at 1 and 10 micrograms/ml) and RO-20-1724 (at 1 microM) inhibited the migration of the lymphocyte component to 64.2+/-16.4%, 48.9+/-3.0% and 47.5+/-5.8% of the control values, respectively, while the migration of the monocytes component was largely unaffected. The PDE inhibitors needed to be present during the assay to inhibit migration, whereas pre-treatment of either the endothelium or the PBMC did not consistently effect lymphocyte migration. The drugs also inhibited the migration of lymphocytes through control inserts, either uncoated or coated with fibronectin, suggesting that some of the inhibition is an effect on lymphocyte motility rather than lymphocyte-endothelial interactions. Lymphocyte migration through fibronectin-coated filters was significantly enhanced compared with uncoated filters. Activation of the PBMC by anti-CD3 MoAb increased motility and migration by up to 300%. This migration appeared to be greatly inhibited by the PDE inhibitors, although the effect was complicated by problems of lymphocyte aggregation. This study provides a novel method of measuring mononuclear cell transendothelial migration, and suggests a possible role of PDE inhibitors in reducing this progress.

The effects of monocytes on the transendothelial migration of T lymphocytes.

In vivo cell-mediated immune reactions are characterized by mixtures of monocytes and T cells. The purpose of this study was to investigate the role of monocytes on T-cell migration and induction of endothelial adhesion molecules. The in vitro model consisted of adding peripheral blood mononuclear cells (PBMC), T cells or mixtures of monocytes and T cells, to endothelial cells on a porous membrane and using flow cytometry to distinguish between the monocyte and lymphocyte components. PBMC and PBMC supernatants were highly potent at upregulating intercellular adhesion molecule-1 (ICAM-1) and inducing expression of vascular cell adhesion molecule-1 (VCAM-1) and E-selectin. Induction by supernatants was inhibited by antibodies to tumour necrosis factor-alpha (TNF-alpha) and interleukin-1 (IL)-1 beta. Using monocyte-enriched populations, as few as one monocyte to 100 endothelial cells was sufficient to upregulate adhesion molecules. Fixed monocytes also induced adhesion molecules and expressed surface-bound cytokines. In contrast, highly purified unstimulated T cells were not found to induce adhesion molecules at 4, 6, 24 or 48 hr of coculture. Purified T cells showed low-level migration through resting (VCAM-1 negative) endothelium, which was approximately doubled by addition of small numbers of monocytes or TNF-alpha. In conclusion, monocytes, via cell surface or released cytokines play an essential role in allowing large-scale recruitment of T cells to inflammatory sites in vivo.

Induction of endothelial cell decay-accelerating factor by vascular endothelial growth factor: a mechanism for cytoprotection against complement-mediated injury during inflammatory angiogenesis.

OBJECTIVE: Decay-accelerating factor (DAF) is a widely expressed, multifunctional cell surface protein involved in complement regulation and cell signaling. Previous studies have demonstrated that endothelial cell (EC) DAF is up-regulated by tumor necrosis factor alpha and inhibits complement binding. Because vascular endothelial growth factor (VEGF) is cytoprotective to endothelium and is expressed at sites of chronic inflammation, we hypothesized that VEGF may induce DAF expression during inflammatory angiogenesis. METHODS: Human umbilical vein and dermal microvascular EC were isolated using routine procedures, and the regulation and function of DAF, as well as other complement-regulatory proteins (membrane cofactor protein and CD59), were analyzed following stimulation with VEGF. RESULTS: Incubation of large- or small-vessel EC with VEGF led to increased expression of DAF, with maximal expression after 48-72 hours of stimulation. This effect depended on the activation of protein kinase C (PKC) and required increased steady-state messenger RNA levels and de novo protein synthesis. Although VEGF-induced EC proliferation was inhibited by both p38 and p42/44 mitogen-activated protein kinase (MAPK) antagonists, DAF up-regulation in response to VEGF was only sensitive to inhibition of p38 MAPK. VEGF-stimulated EC showed a 60% reduction in C3 deposition following complement activation, and this resulted in a marked reduction in complement-mediated EC lysis. These protective effects were abolished by anti-DAF monoclonal antibody 1H4. CONCLUSION: This study confirms the importance of PKC for the regulation of DAF expression by EC and reveals VEGF to be a physiologic agonist for this pathway. The up-regulation of DAF expression by VEGF may represent an important mechanism for the protection of EC from complement-mediated injury during angiogenesis in inflammatory rheumatic diseases.