SPARC oppositely regulates inflammation and fibrosis in bleomycin-induced lung damage

Fibrosis results from inflammatory tissue damage and impaired regeneration. In the context of bleomycin-induced pulmonary fibrosis, we demonstrated that the matricellular protein termed secreted protein acidic and rich in cysteine (SPARC) distinctly regulates inflammation and collagen deposition, depending on its cellular origin. Reciprocal Sparc(-/-) and wild-type (WT) bone marrow chimeras revealed that SPARC expression in host fibroblasts is required and sufficient to induce collagen fibrosis in a proper inflammatory environment. Accordingly, Sparc(-/-) >WT chimeras showed exacerbated inflammation and fibrosis due to the inability of Sparc(-/-) macrophages to down-regulate tumor necrosis factor production because of impaired responses to tumor growth factor-β. Hence, the use of bone marrow cells expressing a dominant-negative form of tumor growth factor-β receptor type II under the monocyte-specific CD68 promoter, as a decoy, phenocopied Sparc(-/-) donor chimeras. Our results point to an unexpected dual role of SPARC in oppositely influencing the outcome of fibrosis.     

Carbon Black Nanoparticle Intratracheal Instillation Does Not Alter Cardiac Gene Expression

Exposure to nanoparticles has been associated with inflammation-related progression of atherosclerosis. To examine nanoparticle-induced cardiac effects in more detail, we characterized heart gene expression profiles alongside plasma proteins associated with cardiovascular disease in C57BL/6 mice intratracheally instilled with vehicle or 0.162 mg Printex 90 carbon black nanoparticles (CBNPs). Mice were killed 1, 3, and 28 days after the exposure and expression profiles were derived using DNA microarrays. Cardiac gene expression was unperturbed by CBNP exposure in two independent experiments, despite substantive changes in pulmonary and hepatic gene expression. MicroRNAs were not affected. Plasma levels of cell adhesion molecules (sE-selectin, sICAM-1, sVCAM-1) and total PAI-1 were immediately increased up to day 3, whereas Apo-A1 and Apo-E were marginally decreased on day 1. These data suggest that though adverse cardiovascular effects are likely following CBNP exposure, these effects are unlikely to be mediated by major direct effects on cardiac gene expression.     

Genotyping analysis and 18FDG uptake in breast cancer patients: a preliminary research

Background

Diagnostic imaging plays a relevant role in the care of patients with breast cancer (BC). Positron Emission Tomography (PET) with 18F-fluoro-2-deoxy-D-glucose (FDG) has been widely proven to be a clinical tool suitable for BC detection and staging in which the glucose analog supplies metabolic information about the tumor. A limited number of studies, sometimes controversial, describe possible associations between FDG uptake and single nucleotide polymorphisms (SNPs). For this reason this field has to be explored and clarified. We investigated the association of SNPs in GLUT1, HIF-1a, EPAS1, APEX1, VEGFA and MTHFR genes with the FDG uptake in BC.

Methods

In 26 caucasian individuals with primary BC, whole-body PET-CT scans were obtained and quantitative analysis was performed by calculating the maximum Standardized Uptake Value normalized to body-weight (SUVmax) and the mean SUV normalized to body-weight corrected for partial volume effect (SUVpvc). Human Gene Mutation Database and dbSNP Short Genetic Variations database were used to analyze gene regions containing the selected SNPs. Patient genotypes were obtained using Sanger DNA sequencing analysis performed by Capillary Electrophoresis.

Results

BC patients were genotyped for the following nine SNPs: GLUT1: rs841853 and rs710218; HIF-1a: rs11549465 and rs11549467; EPAS1: rs137853037 and rs137853036; APEX1: rs1130409; VEGFA: rs3025039 and MTHFR: rs1801133. In this work correlations between the nine potentially useful polymorphisms selected and previously suggested with tracer uptake (using both SUVmax and SUVpvc) were not found.

Conclusions

The possible functional influence of specific SNPs on FDG uptake needs further studies in human cancer. In summary, this is the first pilot study, to our knowledge, which investigates the association between a large panel of SNPs and FDG uptake specifically in BC patients. This work represents a multidisciplinary and translational medicine approach to study BC where, the possible correlation between SNPs and tracer uptake, may be considered to improve personalized cancer treatment and care.     

Development and characterization of a novel mAb against bilitranslocase - a new biomarker of renal carcinoma

Background

Bilitranslocase (TC 2.A.65.1.1) is a bilirubin-specific membrane transporter, found on absorptive (stomach and intestine) and excretory (kidney and liver) epithelia and in vascular endothelium. Polyclonal antibodies have been raised in rabbits in the past, using a synthetic peptide corresponding to AA65-77 of rat liver bilitranslocase, as an antigen. Affinity-purified antibodies from immune sera have been found to inhibit various membrane transport functions, including the bilirubin uptake into human hepatocytes and the uptake of some flavonoids into human vascular endothelial cells. It was described by means of immunohistochemistry using polyclonal antibodies that bilitranslocase expression is severely down-regulated in clear cell renal carcinoma. The aim of our work was development and characterization of high-affinity, specific mAbs against bilitranslocase, which can be used as a potential diagnostic tool in renal cell carcinoma as well as in a wide variety of biological assays on different human tissues.

Materials and methods

Mice were immunized with a multi-antigen peptide corresponding to segment 65–75 of predicted primary structure of the bilitranslocase protein. By a sequence of cloning, immune- and functional tests, we aimed at obtaining a specific monoclonal antibody which recognizes a 37 kDa membrane protein, and influences the transport activity of bilitranslocase.

Results

On the basis of previous results, specific IgM monoclonal antibodies were produced in BALB/c mice, in order to further improve and extend the immunological approach to the study of bilitranslocase in renal cancer cells as well as to develop its potential diagnostics use.

Conclusions

In this article we show an immunological approach, based on newly developed monoclonal antibodies, to a detailed biochemical and functional characterization of a protein whose gene and protein structure is still unknown. We were able to demonstrate our novel mAb as a tumor marker candidate of renal cell carcinoma, which may prove useful in the diagnostic procedures.     

A pilot quasi-experimental study to determine the feasibility of implementing a partograph e-learning tool for student midwife training in Nairobi

Objective

the partograph is a tool used globally to record labour progress. Although it has the potential to improve maternal and neonatal outcomes, some midwives struggle with using it in practice. Training in partograph use is limited, and the theory is often divorced from practice. Innovative ways of improving training are urgently required. We therefore aimed to determine whether the use of an e-learning tool is beneficial for learning partograph skills.

Design

an uncontrolled before-and-after study was conducted, informed by Kirkpatrick's four-stage model of evaluation; we report on the first two stages. We included a cohort of third and fourth year midwifery students who were studying at one university in Nairobi. The same hypothetical case scenario was used, pre- and post-implementation of the World Health Organization partograph e-learning tool, to assess students' partograph completion ability. Views on the tool were also sought, using semi-structured questionnaires. Data were analysed using standard statistical techniques and framework analysis.

Findings

92 (88%) students participated. Students expressed positive views about the e-learning tool. However, the mean post-intervention score (27.21) was less than half of the maximum obtainable score. There was some improvement in test scores; year three mean score pre-intervention was 21.39 (SD 5.72), which increased to 25.10 (5.41) post-intervention (paired-t=3.47, p=0.001); year four mean score pre-intervention was 24.39 (5.98) which increased to 29.30 (6.77) post-intervention (paired t=3.85, df=91, p<0.001). In the post-test, year four students scored higher than year three students (unpaired t=3.28, df=90, p=0.001). Students were unable to plot cervical dilatation correctly, once established labour had been confirmed.

Key conclusion

e-Learning training is acceptable to student midwives and has the potential to be an effective means of teaching the practical application of the partograph. However, in this study, their inability to correctly plot transference from the latent to active phase of labour suggests that the partograph itself may be too complicated. Modifications and further evaluation of the e-learning tool would be required before any widespread implementation. Furthermore, students need the clinical support to operationalise their learning; educating qualified midwives and obstetricians to be positive role models when completing the partograph would be one potential solution. Further research is required, taking on board the recommendations from our pilot study, to investigate the impact of partograph e-learning on practice and clinical outcomes.     

Liver dysfunction in severe ovarian hyperstimulation syndrome

We report the case of a 32-year-old woman suffering from severe liver dysfunction in the course of ovarian hyperstimulation syndrome (OHSS). Complications occurred after successful fertilization subsequent to ovarian stimulation with human menopausal gonadotropin followed by ovulation induction with human chorionic gonadotropin. Because of nausea, vomiting, abdominal distention and enlarged ovaries on an ultrasound examination, she was admitted on the diagnosis of OHSS. During the course of hospitalization severe hepatic injury developed. An increase of more than 100-fold in blood aminotransferase activity was observed. Applied treatment resulted in gradual reduction of ovarian size and resolution of ascites, as well as pleural and pericardial effusions. The patient was discharged from hospital after 46 days. Follow-up examinations at the 13th and 32nd weeks of gestation did not reveal any abnormalities. Pregnancy developed without complications and the woman went into spontaneous labor, giving birth to a viable child at 38 weeks' gestation. Taking into account the above case and previously published reports, the issue of liver dysfunction may have a great impact on the understanding both the pathology and the treatment of OHSS.     

Therapeutic potential of MEK inhibition in acute myelogenous leukemia: rationale for “vertical” and “lateral” combination strategies

In hematological malignancies, constitutive activation of the RAF/MEK/ERK pathway is frequently observed, conveys a poor prognosis, and constitutes a promising target for therapeutic intervention. Here, we investigated the molecular and functional effects of pharmacological MEK inhibition in cell line models of acute myeloid leukemia (AML) and freshly isolated primary AML samples. The small-molecule, ATP-non-competitive, MEK inhibitor PD0325901 markedly inhibited ERK phosphorylation and growth of several AML cell lines and approximately 70?% of primary AML samples. Growth inhibition was due to G(1)-phase arrest and induction of apoptosis. Transformation by constitutively active upstream pathway elements (HRAS, RAF-1, and MEK) rendered FDC-P1 cells exquisitely prone to PD0325901-induced apoptosis. Gene and protein expression profiling revealed a selective effect of PD0325901 on ERK phosphorylation and compensatory upregulation of the RAF/MEK and AKT/p70( S6K ) kinase modules, potentially mediating resistance to drug-induced growth inhibition. Consequently, in appropriate cellular contexts, both "vertical" (i.e., inhibition of RAF and MEK along the MAPK pathway) and "lateral" (i.e., simultaneous inhibition of the MEK/ERK and mTOR pathways) combination strategies may result in synergistic anti-leukemic effects. Overall, MEK inhibition exerts potent growth inhibitory and proapoptotic activity in preclinical models of AML, particularly in combination with other pathway inhibitors. Deeper understanding of the molecular mechanisms of action of MEK inhibitors will likely translate into more effective targeted strategies for the treatment of AML.