The effects of paraoxon and obidoxime on coagulation.

<p><b>(A)</b> Prothrombin time, <b>(B)</b> Thrombin Time (TT) and <b>(C)</b> activated partial thromboplastin time (APTT) are significantly prolonged by obidoxime as well as by its combination with paraoxon.</p

The Organophosphate Paraoxon and Its Antidote Obidoxime Inhibit Thrombin Activity and Affect Coagulation <i>In Vitro</i>

<div><p>Organophosphates (OPs) are potentially able to affect serine proteases by reacting with their active site. The potential effects of OPs on coagulation factors such as thrombin and on coagulation tests have been only partially characterized and potential interactions with OPs antidotes such as oximes and muscarinic blockers have not been addressed. In the current study, we investigated the <i>in vitro</i> interactions between coagulation, thrombin, the OP paraoxon, and its antidotes obidoxime and atropine. The effects of these substances on thrombin activity were measured in a fluorescent substrate and on coagulation by standard tests. Both paraoxon and obidoxime but not atropine significantly inhibited thrombin activity, and prolonged prothrombin time, thrombin time, and partial thromboplastin time. When paraoxon and obidoxime were combined, a significant synergistic effect was found on both thrombin activity and coagulation tests. In conclusion, paraoxon and obidoxime affect thrombin activity and consequently alter the function of the coagulation system. Similar interactions may be clinically relevant for coagulation pathways in the blood and possibly in the brain.</p></div

Paraoxon and obidoxime inhibit thrombin activity <i>in vitro</i>.

<p><b>(A)</b> Varying concentrations of paraoxon (500 nM, 5 μM, 28 μM, 50 μM, 0.28 mM and 0.5 mM) inhibit thrombin activity. <b>(B)</b> Obidoxime inhibits thrombin activity at varying concentrations. In the presence of 3 nM obidoxime there is no significant change in thrombin activity. <b>(C)</b> Thrombin activity is significantly inhibited when obidoxime is applied together with paraoxon.</p

At two weeks upon injury, brain RNA levels of prothrombin, PAR1 and factor X were comparable between mTBI exposed animals and controls.

<p>At two weeks upon injury, brain RNA levels of prothrombin, PAR1 and factor X were comparable between mTBI exposed animals and controls.</p

Long Term Potentiation (LTP) is equally evoked in mTBI and control animals at two weeks following injury.

<p>(a) While twenty fours upon mTBI, animals exhibited a lower LTP compared to controls., (b) no difference in LTP could be detected in mTBI exposed animals vs. control at two weeks upon injury. Sample illustrations are at the indicated time points, the arrow indicates the time of high frequency stimulation delivery.</p

At two weeks upon injury, brain thrombin activity and concentration are similar between mTBI exposed animals and controls.

<p>(a) Thrombin activity as well as its protein levels (b) assessed as described in the methods section were comparable between mTBI and control brain slices. (c) PAR1 and (d) factor X were slightly elevated in mTBI animals compared to control. Refer to text for statistics.</p

Expression of Snord3A in scrapie-infected brain’s.

<p><u>Main picture</u>: Total RNA from brains of scrapie infected mice (6 and 24 months old mice, 4 in each group) and age matched wt controls were amplified for Snord3A by real time PCR as described. Relative RNA expression levels were normalized in reference to UBC. Relative Snord3A expression level in scrapie infected mice is indicated by fold change versus age-matched controls. <u>Insert</u>: Immunoblotting of PrP^Sc in brains of infected mice (2 young, two old). (** P value <0.001).</p

Elevation of Snord3aA expression in Tg MHu2M E199K mice brains is disease dependent.

<p>Total RNA extracted from brains of TgMHu2ME199K (tg), PrP ^0/0 (ko) and wt mice, od designated ages were reverse transcribed into cDNA and amplified by Real Time PCR as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054433#s4" target="_blank">Materials and methods</a>. Each group comprised 4–7 mice. Gene expression changes are depicted in relevance to those measured at 3 months old wt mice. Relative expression levels were normalized in reference to UBC. (** P value <0.001).</p

Elevation of Snord3A expression in Tg MHu2M E199K MEFs and in copper treated mice.

<p>Total RNA from brains of TgMHu2ME199K and wild-type mice as described, as well as from designated MEFs were amplified for Snord3A by Real time PCR. Relative RNA expression levels were normalized in reference to UBC and <i>β-actin</i> (Respectively). a: Scheme of accumulation of PrP and MEF survival during copper treatment (as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054433#pone.0054433-Canello2" target="_blank">[32]</a>). b: Snord3A levels in designated MEFs (wt: cells from wt mice; tg: cells from TgMHu2ME199K mice; KO: cells from PrP^0/0 mice. c: Snord3A levels in TgMHu2ME199K and wild-type mice brain after 75 days of copper treatment versus age-matched control (** P value <0.001).</p

Modeling of Cognitive Impairment by Disease Duration in Multiple Sclerosis: A Cross-Sectional Study

<div><p>Background/Aims</p><p>Large-scale population studies measuring rates and dynamics of cognitive decline in multiple sclerosis (MS) are lacking. In the current cross-sectional study we evaluated the patterns of cognitive impairment in MS patients with disease duration of up to 30 years.</p><p>Methods</p><p>1,500 patients with MS were assessed by a computerized cognitive battery measuring verbal and non-verbal memory, executive function, visual spatial perception, verbal function, attention, information processing speed and motor skills. Cognitive impairment was defined as below one standard deviation (SD) and severe cognitive impairment as below 2SD for age and education matched healthy population norms.</p><p>Results</p><p>Cognitive performance in our cohort was poorer than healthy population norms. The most frequently impaired domains were information processing speed and executive function. MS patients with secondary-progressive disease course performed poorly compared with clinically isolated syndrome, relapsing-remitting and primary progressive MS patients. By the fifth year from disease onset, 20.9% of patients performed below the 1SD cutoff for impairment, p = 0.005, and 6.0% performed below the 2SD cutoff for severe cognitive impairment, p = 0.002. By 10 years from onset 29.3% and 9.0% of patients performed below the 1SD and 2SD cutoffs, respectively, p = 0.0001. Regression modeling suggested that cognitive impairment may precede MS onset by 1.2 years.</p><p>Conclusions</p><p>The rates of cognitive impairment in this large sample of MS patients were lower than previously reported and severe cognitive impairment was evident only in a relatively small group of patients. Cognitive impairment differed significantly from expected normal distribution only at five years from onset, suggesting the existence of a therapeutic window during which patients may benefit from interventions to maintain cognitive health.</p></div