This article introduces fully enclosed microfluidic paper-based analytical devices (microPADs) fabricated by printing toner on the top and bottom of the devices using a laser printer. Enclosing ...paper-based microfluidic channels protects the channels from contamination, contains and protects reagents stored on the device, contains fluids within the channels so that microPADs can be handled and operated more easily, and reduces evaporation of solutions from the channels. These benefits extend the capabilities of microPADs for applications as low-cost point-of-care diagnostic devices.
Parkinson's disease (PD) is the most common movement neurodegenerative movement disorder. An incomplete understanding of the molecular pathways involved in its pathogenesis impedes the development of ...effective disease-modifying treatments. To address this gap, we have previously generated a Drosophila model of PD that overexpresses PD pathogenic mutant form of the second most common causative gene of PD, Leucine-Rich Repeat Kinase 2 (LRRK2).
We employed this model in a genetic modifier screen and identified a gene that encodes for a core subunit of retromer - a complex essential for the sorting and recycling of specific cargo proteins from endosomes to the trans-Golgi network and cell surface. We present evidence that overexpression of the Vps35 or Vps26 component of the cargo-recognition subunit of the retromer complex ameliorates the pathogenic mutant LRRK2 eye phenotype. Furthermore, overexpression of Vps35 or Vps26 significantly protects from the locomotor deficits observed in mutant LRRK2 flies, as assessed by the negative geotaxis assay, and rescues their shortened lifespan. Strikingly, overexpressing Vps35 alone protects from toxicity of rotenone, a neurotoxin commonly used to model parkinsonism, both in terms of lifespan and locomotor activity of the flies, and this protection is sustained and even augmented in the presence of mutant LRRK2. Finally, we demonstrate that knocking down expression of Vps35 in dopaminergic neurons causes a significant locomotor impairment.
From these results we conclude that LRRK2 plays a role in the retromer pathway and that this pathway is involved in PD pathogenesis.
Calcium pyrophosphate dihydrate (CPPD) and basic calcium phosphate (BCP) crystals are important in the pathogenesis of osteoarthritis (OA) but are under recognized even in end stage disease. We ...determined the prevalence of these calcium crystals in synovial fluid (SF) of persons undergoing total knee arthroplasty for degenerative arthritis.
SF samples were obtained from 53 knee joints undergoing total arthroplasty for a pre-operative diagnosis of OA. SF were analyzed via compensated light microscopy for CPPD crystals and a semiquantitative radiometric assay for BCP crystals. Fifty pre-operative radiographs were analyzed and graded according to the scale of Kellgren and Lawrence.
Patients had an average age of 70 years at the time of surgery. CPPD and/or BCP crystals were identified in 60% of SF. Overall radiographic scores correlated with mean concentrations of BCP crystals. Higher mean radiographic scores correlated with the presence of calcium-containing crystals of either type in SF Radiographic chondrocalcinosis was identified in only 31% of those with SF CPPD.
Pathologic calcium crystals were present in a majority of SF at the time of total knee arthroplasty. Intraoperative SF analysis could conveniently identify pathologic calcium crystals providing information that may be relevant to the future care of the patient's replaced joint and that of other joints. This information could also potentially aid in predicting the likelihood of the need for contralateral total knee arthroplasty.
Surgeon visual estimation of shoulder range of motion (ROM) is commonplace in the outpatient office setting and routinely reported in clinical research, but the reliability and accuracy of this ...practice remain unclear. The purpose of this study is to establish the reliability and accuracy of remote visual estimation of shoulder ROM in healthy volunteers and symptomatic patients among a large group of shoulder surgeons. Our hypothesis is that remote visual estimation would be reliable and accurate compared with the digital goniometer method.
Fifty shoulder surgeon members of the PacWest Shoulder and Elbow Society independently determined the active shoulder forward flexion (FF), internal rotation at 90° abduction (IR90), external rotation at 90° abduction, external rotation at the side , and maximal spinal level reached with internal rotation (IRspine) through visual estimation of video recordings taken from 10 healthy volunteers and 10 symptomatic patients. Variations in measurements were quantified using the interobserver reliability through calculation of the intraclass correlation coefficient. Accuracy was determined through comparison with digital goniometer measurements obtained with an on-screen protractor application using Bland–Altman mean differences and 95% limits of agreement.
The interobserver reliability among examiners showed moderate to excellent correlation, with intraclass correlation coefficient ranging from 0.768 to 0.928 for the healthy volunteers and 0.739 to 0.878 for the symptomatic patients. Accuracy was limited, with upper limits of agreement exceeding the established minimal clinically important differences (MCIDs) for FF (20° vs. MCID of 14°) and IR90 (25° vs. 18°) in the healthy volunteers and for FF (33° vs. 16°), external rotation at 90° abduction (21° vs. 18°), and IR90 (31° vs. 20°) in the symptomatic patients.
Despite generally high intersurgeon reliability in the visual estimation of shoulder ROM, there was questionable accuracy when compared to digital goniometer measurements,with measurement errors often exceeding established MCID values. Given the potential implications for the clinical response to treatment and the significance of research findings, the adoption of validated instruments to measure ROM and the standardization of examination procedures should be considered.
Neutrophils respond to ischemic injury by infiltrating the myocardium via the vascular wall. During this process, neutrophils are activated and release inflammatory mediators. Some of these mediators ...are metabolites of arachidonic acid. We have reported that neutrophils metabolize arachidonic acid to 20-HETE, a cytochrome P450 metabolite. We investigated the effects of 20-HETE on coronary vascular tone by examining 20-HETE-induced changes in isometric tension in bovine coronary artery rings precontracted with the thromboxane-mimetic, U46619. 20-HETE relaxed precontracted coronary rings in a concentration-dependent manner (EC50 of 3 x 10(-7) mol/L). Pretreatment with indomethacin, a cyclooxygenase inhibitor, shifted the concentration-response curve to the right (EC50 of 1 x 10(-6) mol/L); maximal relaxations were not affected. This suggested that 20-HETE-induced relaxations were, in part, dependent on the cyclooxygenase pathway. Relaxations to 20-HETE were not significantly changed in endothelium-denuded rings. To determine whether metabolism of 20-HETE to a vasoactive compound might explain the relaxations caused by 20-HETE, rings of coronary artery were incubated with 3H 20-HETE. The incubation buffer was extracted and the 3H products resolved on reverse-phase HPLC. Both denuded and intact arteries failed to metabolize 3H 20-HETE. To investigate whether 20-HETE-induced relaxations were related to release of prostacyclin, we measured the release of 6-keto PGF1alpha, the stable metabolite of prostacyclin, from bovine coronary arteries. 20-HETE (1 x 10(-6) mol/L) stimulated an increase in 6-keto PGF1alpha in intact vessels (908 +/- 138 pg/mL versus 1402 +/- 157 pg/mL, basal versus stimulated). Thus, 20-HETE-induced relaxations are due, in part, to the stimulation of the release of the dilatory prostanoid, prostacyclin.
Vacuolar protein sorting 35 Linhart, Radek; Wong, Sarah Anne; Cao, Jieyun ...
Molecular neurodegeneration,
06/2014, Letnik:
9
Journal Article
Recenzirano
Background Parkinson's disease (PD) is the most common movement neurodegenerative movement disorder. An incomplete understanding of the molecular pathways involved in its pathogenesis impedes the ...development of effective disease-modifying treatments. To address this gap, we have previously generated a Drosophila model of PD that overexpresses PD pathogenic mutant form of the second most common causative gene of PD, Leucine-Rich Repeat Kinase 2 (LRRK2). Findings We employed this model in a genetic modifier screen and identified a gene that encodes for a core subunit of retromer - a complex essential for the sorting and recycling of specific cargo proteins from endosomes to the trans-Golgi network and cell surface. We present evidence that overexpression of the Vps35 or Vps26 component of the cargo-recognition subunit of the retromer complex ameliorates the pathogenic mutant LRRK2 eye phenotype. Furthermore, overexpression of Vps35 or Vps26 significantly protects from the locomotor deficits observed in mutant LRRK2 flies, as assessed by the negative geotaxis assay, and rescues their shortened lifespan. Strikingly, overexpressing Vps35 alone protects from toxicity of rotenone, a neurotoxin commonly used to model parkinsonism, both in terms of lifespan and locomotor activity of the flies, and this protection is sustained and even augmented in the presence of mutant LRRK2. Finally, we demonstrate that knocking down expression of Vps35 in dopaminergic neurons causes a significant locomotor impairment. Conclusions From these results we conclude that LRRK2 plays a role in the retromer pathway and that this pathway is involved in PD pathogenesis. Keywords: Parkinson's disease, LRRK2, VPS35, Retromer, Endolysosomal pathway, Drosophila, Genetics, Rotenone, Neurodegeneration, Endosomes, Lysosome, VPS26
BACKGROUND:Injuries to the hallucal sesamoid complex are uncommon, but they can cause significant pain. The medial sesamoid is the most common site of pain, but the fibular sesamoid can also become ...symptomatic. The most common clinical entities that lead to chronic fibular sesamoid pain are fracture nonunion and osteonecrosis. The purpose of this study was to describe the technique and determine the results of the dorsolateral approach for fibular sesamoid excision.
METHODS:During an 11-year span, 8 patients underwent fibular sesamoidectomy using a dorsolateral approach after a minimum of 6 months of nonoperative treatment. The mean age was 33 years (range, 22 to 43 y). The average follow-up was 97 months (range, 24 to 167 mo). Patients were assessed using the AOFAS forefoot grading scale and a subjective rating for walking, pain, and overall satisfaction.
RESULTS:Fibular sesamoidectomy was performed for osteonecrosis in 3 patients and for nonunion in 5 patients. Four patients had work-related injuries. Two injuries were due to trauma and the rest were chronic, without a known cause. The average length of nonoperative care was 107 weeks and included rest, injections, physiotherapy, bracing, casting, NSAIDs, and orthotics. Overall, the patient subjective satisfaction was 5 excellent and 3 good. The mean AOFAS forefoot score was 91 and average time to return to activity was 15 weeks. The mean pain rating was 1.3/5, and the mean subjective walking score was 4.625/5.
DISCUSSION:Compared with previously published reports, our results for isolated fibular sesamoidectomy show similar satisfaction rates with equivalent time to return to activities and a low complication rate while avoiding a plantar incision.
LEVEL OF EVIDENCE:Diagnostic Level 3. See Instructions for Authors for a complete description of levels of evidence.
This thesis examines linkages between transit extensions and displacement through gentrification and explores Somerville's readiness to address displacement associated with a proposed transit ...extension. Somerville, Massachusetts, experienced economic growth and gentrification following the 1984 Red Line transit extension to Davis Square and now is anticipating a Green Line extension. This study includes a review of gentrification literature, a case study of planning for Davis Square in Somerville prior to the Red Line extension, and recent interviews with practitioners from the Somerville community in the areas of affordable housing, transportation, and planning to identify measures being taken to prevent displacement associated with the proposed Green Line extension. Displacement prevention measures are being implemented in Somerville, suggesting that lessons learned from the 1984 transit extension are being applied, within the limits of political feasibility and available funding.
Department of Pharmacology and Toxicology, Medical College of
Wisconsin, Milwaukee, Wisconsin 53226
Endothelium-derived hyperpolarizing
factor (EDHF) is released in response to agonists such as ACh ...and
bradykinin and regulates vascular smooth muscle tone. Several studies
have indicated that ouabain blocks agonist-induced,
endothelium-dependent hyperpolarization of smooth muscle. We have
demonstrated that epoxyeicosatrienoic acids (EETs), cytochrome
P -450 metabolites of arachidonic acid, function as
EDHFs. To further test the hypothesis that EETs represent EDHFs, we
have examined the effects of ouabain on the electrical and mechanical
effects of 14,15- and 11,12-EET in bovine coronary arteries. These
arteries are relaxed in a concentration-dependent manner to 14,15- and
11,12-EET (EC 50 = 6 × 10 7 M),
bradykinin (EC 50 = 1 × 10 9 M),
sodium nitroprusside (SNP; EC 50 = 2 × 10 7 M), and bimakalim (BMK; EC 50 = 1 × 10 7 M). 11,12-EET-induced relaxations were
identical in vessels with and without an endothelium. Potassium
chloride (1-15 × 10 3 M) inhibited
3 Houabain binding to smooth muscle cells but failed to
relax the arteries. Ouabain (10 5 to 10 4 M)
increased basal tone and inhibited the relaxations to bradykinin, 11,12-EET, and 14,15-EET, but not to SNP or BMK. Barium (3 × 10 5 M) did not alter EET-induced relaxations and ouabain
plus barium was similar to ouabain alone. Resting membrane potential
( E m ) of isolated smooth muscle cells was
50.2 ± 0.5 mV. Ouabain (3 × 10 5 and 1 × 10 4 M) decreased E m
( 48.4 ± 0.2 mV), whereas 11,12-EET (10 7 M)
increased E m ( 59.2 ± 2.2 mV). Ouabain
inhibited the 11,12-EET-induced increase in E m .
In cell-attached patch clamp studies, 11,12-EET significantly increased
the open-state probability ( NP o ) of a calcium-activated potassium channel compared with control cells (0.26 ± 0.06 vs. 0.02 ± 0.01). Ouabain did not change
NP o but blocked the 14,15-EET-induced increase
in NP o . These results indicate that:
1 ) EETs relax coronary arteries in an
endothelium-independent manner, 2 ) unlike EETs, potassium
chloride does not relax the coronary artery, and 3 ) ouabain
inhibits bradykinin- and EET-induced relaxations as has been reported
for EDHF. These findings provide further evidence that EETs are EDHFs.
potassium channels; endothelium-derived hyperpolarizing factor; membrane potential; bimakalim; sodium nitroprusside; potassium; bradykinin
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