Induction of endothelial cell proliferation by recombinant and microparticle-tissue factor involves β1-integrin and extracellular signal regulated kinase activation

Objective: Increased levels of circulating tissue factor (TF) in the form of microparticles increase the risk of thrombosis. However, any direct influence of microparticle-associated TF on vascular endothelial cell proliferation is not known. In this study, the influence of recombinant and microparticle- associated TF on endothelial cell proliferation and mitogen-activated protein kinase signaling mechanisms was examined. Methods and Results: Incubation of human coronary artery endothelial cells with lipidated recombinant full-length TF, or TF-containing microparticles (50 to 200 pmol/L TF), increased the rate of cell proliferation and induced phosphorylation of extracellular signal regulated kinase 1 in a TF-dependent manner. Inhibition of extracellular signal regulated kinase 1/2 using PD98059 or extracellular signal regulated kinase 1/2 antisense oligonucleotides or inhibition of c-Jun N-terminal kinase reduced recombinant TF-mediated cell proliferation. PD98059 also reduced cell proliferation in response to TF-containing microparticles. Inclusion of FVIIa (5 nmol/L) and FXa (10 nmol/L) or preincubation of cells with an inhibitory anti-FVIIa antibody had no additional influence on TF-mediated cell proliferation. However, preincubation of exogenous TF with a β1-integrin peptide (amino acids 579 to 799) reduced TF-mediated proliferation. Conclusion: High concentrations of recombinant or microparticle-associated TF stimulate endothelial cell proliferation through activation of the extracellular signal regulated kinase 1/2 pathway, mediated through a novel mechanism requiring the interaction of exogenous TF with cell surface β1-integrin and independent of FVIIa. © 2010 American Heart Association, Inc

Filamin-A is required for the incorporation of tissue factor into cell-derived microvesicles

We previously reported that the incorporation of tissue factor (TF) into cell-derived microvesicles (MVs) is regulated by the phosphorylation of the cytoplasmic domain of TF. Since the cytoskeletal protein filamin-A is known to bind to the cytoplasmic domain of TF in a phosphorylation-dependent manner, the involvement of filamin-A in the incorporation of TF into MVs was examined. Endothelial cells were transfected to express TF, whereas MDA-MB-231 cells were used to examine endogenously expressed TF. MV release was induced by activating protease-activated receptor-2 (PAR2). Partial suppression of filamin-A expression using two different filamin-A siRNA sequences resulted in significant reductions in the incorporation of TF antigen into MVs as determined by TF-ELISA and western blot analysis, and was reflected in reduced thrombin-generation and FXa-generation capacities of these MVs. Deletion of the cytoplasmic domain of TF also resulted in reduced incorporation of TF into MVs, whereas the suppression of filamin-A expression had no additional effect on the incorporation of truncated TF into MVs. Partial suppression of filamin-A expression had no effect on the number and size distribution of the released MVs. However, >90 % suppression of filamin-A expression resulted in increased MV release, possibly as a result of increased instability of the plasma membrane and underlying cytoskeleton. In conclusion, the presence of filamin-A appears to be essential for the incorporation of TF into MVs following PAR2 activation, but is not required for the process of MV formation and release following PAR2 activation

Youth Employment

Youth employment is the norm in American society. Approximately 80% of youth report holding jobs during their high school years (National Research Council, 1998). Entry into the labor market often begins early, with about half of youth ages 12 and 13 reporting that they work (Rothstein & Herz, 2000). Although statistics are gathered regularly about youth employment in the general population, comparatively little was known about employment patterns of youth with disabilities until the National Longitudinal Transition Study (NLTS) collected data from 1987 to 1990 (see footnote 1). The National Longitudinal Transition Study-2 (NLTS2) (see footnote 2) began updating and expanding data on youth with disabilities in 2001, including information on employment. Information reported here comes from telephone interviews and a mail survey conducted in 2001 with parents and guardians of youth with disabilities, and from comparisons made with 1987 NLTS employment data. Findings from NLTS2 are generalizable to youth with disabilities nationally who were 13 to 16 years old in December of 2000, and to each of 12 federal disability categories and to each age group (e.g., all 13-year-old students with disabilities, all 14-year-old students with disabilities, etc.). According to parents' reports, almost 60% of youth with disabilities are employed during a 1-year period -- some at work-study jobs, but the vast majority at non-school-related jobs

The Effectiveness and Impressions Between Responsive and Non-Responsive Websites Displayed on a Computer, Tablet and Cell Phone: Displaying Websites of Eichler Homes

The researcher designed two types of websites for Eichler Homes in Thousand Oaks. The researcher created both responsive and non-responsive websites. The researcher performed a study and observed the data. Afterwards, the data was analyzed to see people’s preferences and which one they liked best. Most of the general public is not knowledgeable about responsive design. In business, people need to decide for themselves if responsive design is worth the investment since most people aren’t aware of it. As responsive design continues to grow with the use of mobile devices most people aren’t educated about responsive vs. non- responsive design. Results concluded that the responsive website received a more popular vote

'Shell is Proud to Present… The Spirit Sings’: Museum Sponsorship and Public Relations in Oil Country

This article re-examines the renowned Canadian exhibition, The Spirit Sings: Artistic Traditions of Canada’s First Peoples (1988) through a lens of corporate, national, and institutional interests. The author positions The Spirit Sings as a productive historical case study for contemporary questions of decolonization and divestment in museums. Using archival and interview findings from her doctoral research, the author highlights the sponsorship and public relations elements of the exhibition, which she argues have been missing from past analyses. Ultimately, the author uses this case study to question the relevance of current debates over oil sponsorship for museums that operate within extractive economies. The article concludes by calling for further critical research around the organizational processes of museums and their participation in corporate legitimation

Accumulation of tissue factor in endothelial cells induces cell apoptosis, mediated through p38 and p53 activation

We previously reported that high levels of tissue factor (TF) can induce cellular apoptosis in endothelial. In this study, TF-mediated mechanisms of induction of apoptosis were explored. Endothelial cells were transfected to express wild-type TF. Additionally, cells were transfected to express Asp253-substituted, or Ala253-substitued TF to enhance or prevent TF release respectively. Alternatively, cells were pre-incubated with TF-rich and TF-poor microvesicles. Cell proliferation, apoptosis and the expression of cyclin D1, p53, bax and p21 were measured following activation of cells with PAR2-agonist peptide. Greatest levels of cell proliferation and cyclin D1 expression were observed in cells expressing wild-type or Asp253-substituted TF. In contrast, increased cellular apoptosis was observed in cells expressing Ala253-substituted TF, or cells pre-incubated with TF-rich microvesicles. The level of p53 protein, p53-phosphorylation at ser33, p53 nuclear localisation and transcriptional activity, but not p53 mRNA, were increased in cells expressing wild-type and Ala253-substituted TF, or in cells pre-incubated with TF-rich microvesicles. However, the expression of bax and p21 mRNA, and Bax protein were only increased in cells pre-incubated with TF-rich microvesicle and in cells expressing Ala253-substituted TF. Inhibition of the transcriptional activity of p53 using pifithrin-α suppressed the expression of Bax. Finally, siRNA–mediated suppression of p38α, or inhibition using SB202190 significantly reduced the p53 protein levels, p53 nuclear localisation and transcriptional activity, suppressed Bax expression and prevented cellular apoptosis. In conclusion, accumulation of TF within endothelial cell, or sequestered from the surrounding can induce cellular apoptosis through mechanisms mediated by p38, and involves the stabilisation of p53

Skillful Coaching: New Directions In Teaching Health Assessment

The purpose of this study was to examine differences in nursing student’s assessment skills before and after the implementation of a cognitive apprenticeship didactic approach including think-aloud and critical dialogue.  In the nursing care of clients, a through and accurate holistic health assessment is essential.  The content of the assessment task is taught in all nursing programs.  Students must be competent not only in performing the examination but also in evaluating the information obtained and integrating it with the health history and lab findings.  This extensive criteria makes assessment a challenging skill to master.  Problems arise when students in nursing attempt to apply the theoretical assessment approaches to the clinical setting with real clients.  Consequently, student’s performance outcomes are often unsatisfactory, as the teaching format usually focuses on the students’ acquisition of domain knowledge and psychomotor skills rather than on shaping the thinking processes involved.  Students’ common problems with physical assessment shows that they struggle to manipulate the abstract positions involved

Back To The Future A Joint Adventure For Addressing The Nursing Shortage

One of the greatest challenges facing managers in the health care industry is the recurrent nursing shortage.  This paper presents a ‘Back to the Future’ innovative approach that addresses this issue.  The ‘Berger Project’, a joint venture between Berger Health Care System (BHCS) and a nursing education program at Ohio University-Chillicothe, began in spring 2003.  The venture incorporates a strategy that goes ‘BACK’in time by shifting the educational setting for nursing students from the traditional college campus to the health care organization campus which was a commonplace setting  ‘BACK’ in the 1940’s and 1950’s to resolve the nursing shortages--present and ‘FUTURE’.  The project will culminate with an evaluation process guided by comparative research studies of the nursing students in the traditional setting and those in the ‘Berger Project’ that investigate the differences in the level of critical thinking, empowerment and retention of the graduates of the two nursing programs

Exploring justice tensions in the Barnahus model

Barnahus is to support children and their families during the justice and recovery process from experiencing abuse or violence. Yet, many perspectives of justice exist in the multi-disciplinary systems involved in Barnahus. The intersection of these systems can cause tension, which affects service delivery and, ultimately, the experience of the justice and recovery journey. The purpose of this chapter is to outline the needs, rights, and responsibilities of those involved in Barnahus as key stakeholders, including children, their families, criminal justice professionals, social and health care professionals, and professionals working in non-governmental organisations (NGOs) that support children and families. We also discuss the justice-related tensions that arise in the Barnahus model and how Barnahus can situate and advance a child-friendly justice model

Peptidyl-prolyl isomerase 1 (Pin1) preserves the phosphorylation state of tissue factor and prolongs its release within microvesicles

© 2017 Elsevier B.V. The exposure and release of TF is regulated by post-translational modifications of its cytoplasmic domain. Here, the potential of Pin1 to interact with the cytoplasmic domain of TF, and the outcome on TF function was examined. MDA-MB-231 and transfected-primary endothelial cells were incubated with either Pin1 deactivator Juglone, or its control Plumbagin, as well as transfected with Pin1-specific or control siRNA. TF release into microvesicles following activation, and also phosphorylation and ubiquitination states of cellular-TF were then assessed. Furthermore, the ability of Pin1 to bind wild-type and mutant forms of overexpressed TF-tGFP was investigated by co-immunoprecipitation. Additionally, the ability of recombinant or cellular Pin1 to bind to peptides of the C-terminus of TF, synthesised in different phosphorylation states was examined by binding assays and spectroscopically. Finally, the influence of recombinant Pin1 on the ubiquitination and dephosphorylation of the TF-peptides was examined. Pre-incubation of Pin1 with Juglone but not Plumbagin, reduced TF release as microvesicles and was also achievable following transfection with Pin1-siRNA. This was concurrent with early ubiquitination and dephosphorylation of cellular TF at Ser253. Pin1 co-immunoprecipitated with overexpressed wild-type TF-tGFP but not Ser258 → Ala or Pro259 → Ala substituted mutants. Pin1 did interact with Ser258-phosphorylated and double-phosphorylated TF-peptides, with the former having higher affinity. Finally, recombinant Pin1 was capable of interfering with the ubiquitination and dephosphorylation of TF-derived peptides. In conclusion, Pin1 is a fast-acting enzyme which may be utili sed by cells to protect the phosphorylation state of TF in activated cells prolonging TF activity and release, and therefore ensuring adequate haemostasis