A role for Rho kinase in vascular contraction evoked by sodium fluoride

Agonist and depolarization-induced vascular smooth muscle contractions involve the activation of Rho-kinase pathway. However, there are no reports addressing the question whether this pathway is involved in NaF-induced vascular contractions. We hypothesized that Rho-kinase plays a role in vascular contraction evoked by sodium fluoride in rat aortae. In both physiological salt solution and calcium-free solution with 2 mM EGTA, cumulative addition of NaF increased vascular tension in concentration-dependent manners. Effects of Rho-kinase inhibitor (Y27632) on phosphorylation of myosin light chain (MLC20) and myosin targeting subunit (MYPT1(Thr696)) of myosin light chain phosphatase as well as NaF-induced contractions were determined using isolated tissue and the Western blot experiments. Y27632 inhibited NaF-induced contractions in a concentration-dependent manner. NaF increased phosphorylation of MLC20 and MYPT1(Thr696), which were also inhibited by Y27632. However, MLCK inhibitor (ML-7) or PKC inhibitor (Ro31-8220) did not inhibit the NaF-induced contraction. These results indicate that activation of Rho-kinase and the subsequent phosphorylation of MYPT1(Thr696) play important roles in NaF-induced contraction of rat aortae.     

Role of a reaction coordinate in free energy calculations

Abstract

The free energy calculation method emerges as a viable technique for ‘in-silico’ calorimetry. Efficient sampling techniques and the good choice of a reaction path connecting the reactant and the product state enable accurate computations of the free energy differences. We argue that in many cases the thermodynamic integration technique has the lowest variance when the transformation between the reactant and the product state proceeds along the natural path of the studied chemical reaction. We provide examples of free energy calculations for the fragmentation of the charged clusters and the swapping reaction of oligomer formation in proteins that follow a tentative reaction mechanism.     

COX-2 Inhibition Combined with Radiation Reduces Orthotopic Glioma Outgrowth by Targeting the Tumor Vasculature

Cyclooxygenase 2 (COX-2) inhibitors have been shown to enhance tumor''s response to radiation in several animal models. The strong association of COX-2 and angiogenesis suggests that the tumor vasculature may be involved in this process. The current study investigated whether treatment with the COX-2 inhibitor E-6087 could influence response to local radiation in orthotopically growing murine gliomas and aimed to analyze the involvement of the tumor vasculature. GL261 glioma cells were injected into the cerebrum of C57bl/6 mice. From day 7 after tumor cell injection, mice were treated with COX-2 inhibitor at 50 mg/kg i.p. every third day. Radiation consisted of three fractions of 2 Gy given daily from day 9 to day 11. Mice were killed at day 21. The COX-2 inhibitor significantly enhanced the response to radiation, reducing mean volume to 32% of tumors treated with radiation only. The combination treatment neither increased apoptosis of tumor cells or stromal cells nor affected tumor microvascular density. In vitro, E-6087 and its active metabolite did not affect clonogenic survival of GL261 cells or human umbilical vein endothelial cell after radiation. In vivo, however, there was a nonsignificant increase in Angiopoietin (Ang)-1 and Tie-2 mRNA levels and a decrease of Ang-2 mRNA levels after combination treatment. These changes coincided with a significant increase in α-smooth muscle actin-positive pericyte coverage of tumor vessels. In conclusion, the antitumor effect of radiation on murine intracranial glioma growth is augmented by combining with COX-2 inhibition. Our findings suggest an involvement of the tumor vasculature in the observed effects.     

Long-Term Clinical Outcome of Internal Globus Pallidus Deep Brain Stimulation for Dystonia

Background

GPi (Internal globus pallidus) DBS (deep brain stimulation) is recognized as a safe, reliable, reversible and adjustable treatment in patients with medically refractory dystonia.

Objectives

This report describes the long-term clinical outcome of 36 patients implanted with GPi DBS at the Neurosurgery Department of Seoul National University Hospital.

Methods

Nine patients with a known genetic cause, 12 patients with acquired dystonia, and 15 patients with isolated dystonia without a known genetic cause were included. When categorized by phenomenology, 29 patients had generalized, 5 patients had segmental, and 2 patients had multifocal dystonia. Patients were assessed preoperatively and at defined follow-up examinations postoperatively, using the Burke-Fahn-Marsden dystonia rating scale (BFMDRS) for movement and functional disability assessment. The mean follow-up duration was 47 months (range, 12–84)

Results

The mean movement scores significantly decreased from 44.88 points preoperatively to 26.45 points at 60-month follow up (N = 19, P = 0.006). The mean disability score was also decreased over time, from 11.54 points preoperatively to 8.26 points at 60-month follow up, despite no statistical significance (N = 19, P = 0.073). When analyzed the movement and disability improvement rates at 12-month follow up point, no significant difference was noted according to etiology, disease duration, age at surgery, age of onset, and phenomenology. However, the patients with DYT-1 dystonia and isolated dystonia without a known genetic cause showed marked improvement.

Conclusions

GPi DBS is a safe and efficient therapeutic method for treatment of dystonia patients to improve both movement and disability. However, this study has some limitations caused by the retrospective design with small sample size in a single-center.     

The Response to Stimulation in Neurons and Astrocytes

Neurochemical Research - The brain uses mainly glucose as fuel with an index of glucose to oxygen utilization close to 6, the maximal index if all glucose was completely oxidized. However, this...     

Protective role of cytosolic NADP(+)-dependent isocitrate dehydrogenase, IDH1, in ischemic pre-conditioned kidney in mice

Ischemic pre-conditioning protects the kidney against subsequent ischemia/reperfusion (I/R). This study investigated the role of cytosolic NADP(+)-dependent isocitrate dehydrogenase (IDH1), a producer of NADPH, in the ischemic pre-conditioning. Mice were pre-conditioned by 30 min of renal ischemia and 8 days of reperfusion. In non-pre-conditioned mice 30 min of ischemia had significantly increased the levels of plasma creatinine, BUN, lipid peroxidation and hydrogen peroxide in kidneys, whereas in pre-conditioned mice, the ischemia did not increase them. The reductions of reduced glutathione and NADPH after I/R were greater in non-pre-conditioned mice than in pre-conditioned mice. Ischemic pre-conditioning prevented the I/R-induced decreases in IDH1 activity and expression, but not in glucose-6-phosphate dehydrogenase activity. In conclusion, protection of the kidney afforded by ischemic pre-conditioning may be associated with increased activity of IDH1 which relates to increased levels of NADPH, increased ratios of GSH/total glutathione, less oxidative stress and less kidney injury induced by subsequent I/R insult.     

Crystal structures of aprotinin and its complex with sucrose octasulfate reveal multiple modes of interactions with implications for heparin binding

The crystal structures of aprotinin and its complex with sucrose octasulfate (SOS), a polysulfated heparin analog, were determined at 1.7-2.6 Å resolutions. Aprotinin is monomeric in solution, which associates into a decamer at high salt concentrations. Sulfate ions serve to neutralize the basic amino acid residues of aprotinin to stabilize the decameric aprotinin. Whereas SOS interacts with heparin binding proteins at 1:1 molar ratio, SOS was surprisingly found to induce strong agglutination of aprotinins. Five molecules of aprotinin interact with one molecule of the sulfated sugar, which is stabilized by electrostatic interactions between the positively charged residues of aprotinin and sulfate groups of SOS. The multiple binding modes of SOS with five individual aprotinin molecules may represent the diverse patterns of potential heparin binding to aprotinin, reflecting the interactions of densely packed protein molecules along the heparin polymer.     

The effects of probiotics on PPI-triple therapy for Helicobacter pylori eradication

Background: This study was performed to evaluate whether the addition of probiotics to proton pump inhibitor (PPI)‐based triple therapy increases the likelihood of successful Helicobacter pylori eradication. Materials and Methods: Three hundred and forty‐seven H. pylori‐infected patients were randomized into a triple‐plus‐yogurt group (yogurt group, n = 168) or a triple‐only group (control group, n = 179). Triple therapy consisted of PPI b.i.d., clarithromycin 500 mg b.i.d., and amoxicillin 1 g b.i.d. for 7 days. Yogurt group received triple therapy for 1 week and one bottle of Will yogurt per day for at 3 weeks, starting on the first day of triple therapy. Will yogurt (a Korean brand) contains Lactobacillus acidophilus HY2177, Lactobacillus casei HY2743, Bifidobacterium longum HY8001, and Streptococcus thermophilus B‐1. ¹³C‐urea breath test was performed at least 4 weeks after completion of triple therapy. Eradication rates, compliances, and adverse events were compared. Results: By intention‐to treat analysis the H. pylori eradication rates in the yogurt group 79.2% (133 of 168) was similar to that in the control group 72.1% (129 of 179) (p = .124). However, by per‐protocol (PP) analysis, the eradication rate in the yogurt group, 87.5% (133 of 152) was higher than that in the control group, 78.7% (129 of 164) (p = .037). Common adverse events were metallic taste (11.8%) and diarrhea (8.6%). The frequency of adverse effects in the yogurt group 41.1% (69/168) were higher than in the control group, 26.3% (47 of 179) (p = .003). However, most adverse events were mild to moderate in intensity, and the severities of adverse effects were similar in both groups (p = .401). Conclusions: The addition of Will yogurt to triple therapy did not reduce the side‐effects of triple therapy. But it increased the H. pylori eradication rate by PP analysis, encouraging more research in this field.     

Ubiquitination of mammalian Pex5p, the peroxisomal import receptor

Protein translocation across the peroxisomal membrane requires the concerted action of numerous peroxins. One central component of this machinery is Pex5p, the cycling receptor for matrix proteins. Pex5p recognizes newly synthesized proteins in the cytosol and promotes their translocation across the peroxisomal membrane. After this translocation step, Pex5p is recycled back into the cytosol to start a new protein transport cycle. Here, we show that mammalian Pex5p is ubiquitinated at the peroxisomal membrane. Two different types of ubiquitination were detected, one of which is thiol-sensitive, involves Cys(11) of Pex5p, and is necessary for the export of the receptor back into the cytosol. Together with mechanistic data recently described for yeast Pex5p, these findings provide strong evidence for the existence of Pex4p- and Pex22p-like proteins in mammals.