Double-Stranded RNA Attenuates the Barrier Function of Human Pulmonary Artery Endothelial Cells

Circulating RNA may result from excessive cell damage or acute viral infection and can interact with vascular endothelial cells. Despite the obvious clinical implications associated with the presence of circulating RNA, its pathological effects on endothelial cells and the governing molecular mechanisms are still not fully elucidated. We analyzed the effects of double stranded RNA on primary human pulmonary artery endothelial cells (hPAECs). The effect of natural and synthetic double-stranded RNA (dsRNA) on hPAECs was investigated using trans-endothelial electric resistance, molecule trafficking, calcium (Ca2+) homeostasis, gene expression and proliferation studies. Furthermore, the morphology and mechanical changes of the cells caused by synthetic dsRNA was followed by in-situ atomic force microscopy, by vascular-endothelial cadherin and F-actin staining. Our results indicated that exposure of hPAECs to synthetic dsRNA led to functional deficits. This was reflected by morphological and mechanical changes and an increase in the permeability of the endothelial monolayer. hPAECs treated with synthetic dsRNA accumulated in the G1 phase of the cell cycle. Additionally, the proliferation rate of the cells in the presence of synthetic dsRNA was significantly decreased. Furthermore, we found that natural and synthetic dsRNA modulated Ca2+ signaling in hPAECs by inhibiting the sarco-endoplasmic Ca2+-ATPase (SERCA) which is involved in the regulation of the intracellular Ca2+ homeostasis and thus cell growth. Even upon synthetic dsRNA stimulation silencing of SERCA3 preserved the endothelial monolayer integrity. Our data identify novel mechanisms by which dsRNA can disrupt endothelial barrier function and these may be relevant in inflammatory processes

Hepatitis B virus infected physicians and disclosure of transmission risks to patients: A critical analysis

BACKGROUND: The potential for transmission of blood-borne pathogens such as hepatitis B virus from infected healthcare workers to patients is an important and difficult issue facing healthcare policymakers internationally. Law and policy on the subject is still in its infancy, and subject to a great degree of uncertainty and controversy. Policymakers have made few recommendations regarding the specifics of practice restriction for health care workers who are hepatitis B seropositive. Generally, they have deferred this work to vaguely defined "expert panels" which will have the power to dictate the conditions under which infected health care workers may continue to practice. DISCUSSION: In this paper we use recent Canadian policy statements as a critical departure point to propose more specific recommendations regarding disclosure of transmission risks in a way that minimizes practice restriction of hepatitis B seropositive health care workers without compromising patient safety. The range of arguments proposed in the literature are critically examined from the perspective of ethical analysis. SUMMARY: A process for considering the ethical implications of the disclosure of the sero-status of health care workers is advanced that considers the varied perspectives of different stakeholders

Alterations in Vitamin D signalling and metabolic pathways in breast cancer progression: a study of VDR, CYP27B1 and CYP24A1 expression in benign and malignant breast lesions Vitamin D pathways unbalanced in breast lesions

<p>Abstract</p> <p>Background</p> <p>Breast cancer is a heterogeneous disease associated with different patient prognosis and responses to therapy. Vitamin D has been emerging as a potential treatment for cancer, as it has been demonstrated that it modulates proliferation, apoptosis, invasion and metastasis, among others. It acts mostly through the Vitamin D receptor (VDR) and the synthesis and degradation of this hormone are regulated by the enzymes CYP27B1 and CYP24A1, respectively. We aimed to study the expression of these three proteins by immunohistochemistry in a series of breast lesions.</p> <p>Methods</p> <p>We have used a cohort comprising normal breast, benign mammary lesions, carcinomas <it>in situ </it>and invasive carcinomas and assessed the expression of the VDR, CYP27B1 and CYP24A1 by immunohistochemistry.</p> <p>Results</p> <p>The results that we have obtained show that all proteins are expressed in the various breast tissues, although at different amounts. The VDR was frequently expressed in benign lesions (93.5%) and its levels of expression were diminished in invasive tumours (56.2%). Additionally, the VDR was strongly associated with the oestrogen receptor positivity in breast carcinomas. CYP27B1 expression is slightly lower in invasive carcinomas (44.6%) than in benign lesions (55.8%). In contrast, CYP24A1 expression was augmented in carcinomas (56.0% in <it>in situ </it>and 53.7% in invasive carcinomas) when compared with that in benign lesions (19.0%).</p> <p>Conclusions</p> <p>From this study, we conclude that there is a deregulation of the Vitamin D signalling and metabolic pathways in breast cancer, favouring tumour progression. Thus, during mammary malignant transformation, tumour cells lose their ability to synthesize the active form of Vitamin D and respond to VDR-mediated Vitamin D effects, while increasing their ability to degrade this hormone.</p

Pharmacologically directed strategies in academic anticancer drug discovery based on the European NCI compounds initiative

Background: The European NCI compounds programme, a joint initiative of the EORTC Research Branch, Cancer Research Campaign and the US National Cancer Institute, was initiated in 1993. The objective was to help the NCI in reducing the backlog of in vivo testing of potential anticancer compounds, synthesised in Europe that emerged from the NCI in vitro 60-cell screen. Methods: Over a period of more than twenty years the EORTC—Cancer Research Campaign panel reviewed ~2000 compounds of which 95 were selected for further evaluation. Selected compounds were stepwise developed with clear go/no go decision points using a pharmacologically directed programme. Results: This approach eliminated quickly compounds with unsuitable pharmacological properties. A few compounds went into Phase I clinical evaluation. The lessons learned and many of the principles outlined in the paper can easily be applied to current and future drug discovery and development programmes. Conclusions: Changes in the review panel, restrictions regarding numbers and types of compounds tested in the NCI in vitro screen and the appearance of targeted agents led to the discontinuation of the European NCI programme in 2017 and its transformation into an academic platform of excellence for anticancer drug discovery and development within the EORTC-PAMM group. This group remains open for advice and collaboration with interested parties in the field of cancer pharmacology

Cerebrospinal fluid matrix metalloproteinase-9 increases during treatment of recurrent malignant gliomas

<p>Abstract</p> <p>Background</p> <p>Matrix metalloproteinases (MMPs) are enzymes that promote tumor invasion and angiogenesis by enzymatically remodeling the extracellular matrix. MMP-2 and MMP-9 are the most abundant forms of MMPs in malignant gliomas, while a 130 kDa MMP is thought to be MMP-9 complexed to other proteinases. This study determined whether doxycycline can block MMP activity <it>in vitro</it>. We also measured MMP-2 and MMP-9 levels in cerebrospinal fluid (CSF) from patients with recurrent malignant gliomas.</p> <p>Methods</p> <p>To determine whether doxycycline can block MMP activity, we measured the extent of doxycyline-mediated MMP-2 and MMP-9 inhibition <it>in vitro </it>using epidermal growth factor receptor (EGFR) transfected U251 glioma cell lines. MMP activity was measured using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) zymography. In addition, patients underwent lumbar puncture for CSF sampling at baseline, after 6 weeks (1 cycle), and after 12 weeks (2 cycles), while being treated with a novel chemotherapy regimen of irinotecan, thalidomide, and doxycycline designed to block growth/proliferation, angiogenesis, and invasion. Irinotecan was given at 125 mg/m²/week for 4 weeks in 6-week cycles, together with continuous doxycycline at 100 mg twice daily on Day 1 and 50 mg twice daily thereafter. Daily thalidomide dose in our cohort was 400 mg. Tumor progression was monitored by magnetic resonance imaging (MRI).</p> <p>Results</p> <p>Doxycyline <it>in vitro </it>completely abolished MMP-9 activity at 500 μg/ml while there was only 30 to 50% inhibition of MMP-2 activity. Four patients respectively completed 4, 3, 1, and 2 cycles of irinotecan, thalidomide, and doxycycline. Patient enrollment was terminated after one patient developed radiologically defined pulmonary embolism, and another had probable pulmonary embolism. Although CSF MMP-2 and 130 kDa MMP levels were stable, MMP-9 level progressively increased during treatment despite stable MRI.</p> <p>Conclusion</p> <p>Doxycycline can block MMP-2 and MMP-9 activities from glioma cells <it>in vitro</it>. Increased CSF MMP-9 activity could be a biomarker of disease activity in patients with malignant gliomas, before any changes are detectable on MRI.</p

Nonsurgical and surgical periodontal therapy in single-rooted teeth

The purpose of this study was to compare the effect of tooth related and patient related factors on the success of non-surgical and surgical periodontal therapy. In 41 patients (22 female) with untreated and/or recurrent periodontitis, no therapy, scaling and root planing (SRP), or access flap (AF) were assigned according to probing pocket depth (PPD). PPD and vertical relative attachment level (RAL-V) were obtained initially, 3 and 6 months after therapy. Baseline data were compared according to therapy, jaw, tooth type, and site. Factors influencing clinical parameters were identified using multilevel analyses. Baseline PPDs were deeper interproximally, in the maxilla and at premolars compared to buccal/oral sites, mandibular, and anterior teeth. At 6 months, PPD reduction and RAL-V gain were significantly greater at sites receiving SRP and AF as compared to untreated sites (p < 0.001). PPD reduction and RAL-V gain were significantly less (p < 0.005) in smokers as compared to nosmokers and at interproximal sites (p < 0.0001) as compared to buccal/oral sites. RAL-V gain was less in aggressive periodontitis, and PPD reduction was less in the maxilla (p < 0.001). In sites with greater bone loss and infrabony defects, a poorer response was observed regarding RAL-V gain or PPD reduction, respectively. The conclusions of the study are the following: (1) Nonsurgical and surgical periodontal therapies are effective in single-rooted teeth; (2) severe interproximal bone loss and infrabony defects deteriorate clinical results; and (3) there seem to be more defect-associated (tooth, site) factors influencing treatment outcome than patient-associated factors

Prevalence of contagious and environmental mastitis-causing bacteria in bulk tank milk and its relationships with milking practices of dairy cattle herds in São Miguel Island (Azores)

This study aimed to assess the degree of contamination of bulk tank milk (BTM) by Staphylococcus spp. and coliform bacteria and to identify major milking practices that help perpetuate them in dairy cattle herds in São Miguel Island. In July 2014, BTM was sampled and a survey concerning local milking practices was conducted on 100 herds. Semi quantitative multiplex polymerase chain reaction detected coagulase-negative staphylococci, Escherichia coli, Staphylococcus aureus, and other coliform bacteria (Klebsiella oxytoca, Klebsiella pneumoniae, andSerratia marcescens) in 100, 75, 59, and 35 % of BTM, respectively. According to multivariable univariate models, on herds not using hot water for cleaning the milking machine and teat liners, there was at least 3.4 more odds (P<0.01) to have S. aureus or coliform bacteria contamination in BTM. The likelihoodoffinding S.aureus inBTMwas higher(P<0.001)on herds without high hygiene during milking, when milking mastitic cows at the end, on abrupt cessation of milking at dry-off, and official milk control implementation. The glove use also favored (odds ratio (OR) 5.8; P<0.01)thedetection ofcoliformbacteriainBTM.Poormilkingpracticesidentified in this study should be avoided in order to decrease S. aureus and coliform bacteria contamination of BTM. Other factors associated with milk quality in São Miguel Island also should be further investigated

A novel approach to modelling water transport and drug diffusion through the stratum corneum

<p>Abstract</p> <p>Background</p> <p>The potential of using skin as an alternative path for systemically administering active drugs has attracted considerable interest, since the creation of novel drugs capable of diffusing through the skin would provide a great step towards easily applicable -and more humane- therapeutic solutions. However, for drugs to be able to diffuse, they necessarily have to cross a permeability barrier: the <it>stratum corneum </it>(SC), the uppermost set of skin layers. The precise mechanism by which drugs penetrate the skin is generally thought to be diffusion of molecules through this set of layers following a "tortuous pathway" around corneocytes, i.e. impermeable dead cells.</p> <p>Results</p> <p>In this work, we simulate water transport and drug diffusion using a three-dimensional porous media model. Our numerical simulations show that diffusion takes place through the SC regardless of the direction and magnitude of the fluid pressure gradient, while the magnitude of the concentrations calculated are consistent with experimental studies.</p> <p>Conclusions</p> <p>Our results support the possibility for designing arbitrary drugs capable of diffusing through the skin, the time-delivery of which is solely restricted by their diffusion and solubility properties.</p