24-hour Time Point Research Articles

Alteration of the Structure and Dynamics of Venous Clot Formation in Human and Murine Sickle Cell Disease

Sickle cell disease (SCD) is associated with chronic activation of coagulation and an increased risk of venous thromboembolism. Traditionally, it is believed that during venous thrombosis, red blood cells (RBC) are simply trapped within fibrin-rich thrombi and do not actively affect thrombosis. However, a study from our group showed that factor XIII (FXIII) activity is critical for the retention of RBC within clots and directly affects thrombus size. Others reported that during clot contraction polyhedral shaped RBC formed a densely packed core and that SCD alters the formation of polyhedrocytes which may affect clot stability (Strauss et al, 2015, ASH abstract).We further investigated if SCD affects the structure and the dynamics of clot formation. Ex vivo clot retraction was performed using blood from sickle cell patients and Townes mice (a model of SCD). Citrated blood was added to siliconized wells of 96-well plates containing CaCl2 (10mM) and tissue factor (1pM) and incubated for 2 hours at 37°C. The number of RBC extruded from the clot was counted in serum by Hemavet™ and expressed as a percentage of initial RBC number in the anticoagulated blood. Morphology of the clots was evaluated using scanning and transmission electron microscopy (EM). Thrombosis in Townes SS (sickle) and AA (control) mice was studied using femoral vein thrombosis induced by electrolytic injury and inferior vena cava stenosis models.The number of mouse SS RBC in the serum extruded during clot contraction was dramatically reduced compared to the number of AA RBC (0.8±0.8% vs. 19.4±0.8%, n=3, p<0.0001, Fig. A). A similar result was observed for human RBC. Since SS mice and sickle patients have a lower hematocrit (HCT), we investigated if the number of RBC affects the extrusion of these cells during clot retraction. Indeed, lowering HCT in AA mouse blood reduced RBC extrusion from the clots. However, increasing HCT in SS mouse blood to that of AA blood did not increase the number of SS RBC extruded from the clot. Furthermore, inhibition of FXIIIa activity with T101 (10µM) increased the release of AA (by 64%, n=6, p<0.05) but not SS RBC from mouse clots. These data indicate that the entrapment of SS RBCs within the clot is not simply caused by lower RBC number, and is FXIII-independent. Mixing the platelet poor plasma (PPP) and cellular fraction of AA and SS mouse blood revealed that the entrapment of SS RBC is not mediated by PPP (AA blood recombined = 6.9±3.6%; SS blood recombined = 0.4±0.4%; AA cells/SS PPP = 6.7±6.4%; SS cells/AA PPP = 0.1±0.2%, n=3 per group).Clots formed ex vivo from AA blood had a gel-like, soft structure, whereas SS clots were more firm and stiff. EM demonstrated that RBC within AA clots had polyhedral shapes and were tightly packed in the central part of the clot. In contrast, most of the mouse SS RBC did not have polyhedral shapes, underwent sickling and were not compacted within the clot. They also formed long "spicule-like" processes that intertwined with fibrin fibers (Fig. C). Similar results were observed in blood of sickle patients; however, the sickled RBC phenotype was less prominent. Importantly, sickling of RBC was observed in clots formed in the inferior vena cava of SS mice, 2 hours after vessel stenosis (Fig. D). We also performed a tPA challenge assay on clots formed ex vivo from human blood and showed that SS clots challenged with low tPA concentration (0.6nM) were more resistant to fibrinolysis compared to AA clots (clot lysis time, 714±6 vs. 388.3±120.7 minutes, n=6, p=0.08).The electrolytic injury model of venous thrombosis was used to investigate the dynamics of clot formation in SS mice in vivo. Mice were infused with fluorescently labeled antibodies for fibrin (green) and platelets (red). Electrolytic injury was applied to the femoral vein; a relative intensity of fibrin and platelet accumulation was assessed by fluorescence microscopy for one hour at 10-minute intervals. SS mice have increased platelet and fibrin accumulation compared to AA mice (~ 2 fold, n=5-7, p<0.05 for 40, 50 and 60 minute time points). Interestingly, in the AA clots, platelets were mostly localized on the surface, in contrast to their widespread distribution throughout the clot in SS mice (Fig. E, yellow color).Our data demonstrated that SCD alters the structure and dynamics of venous clot formation. Experiments investigating the consequence of these observations in mouse models of stroke and pulmonary embolism are currently ongoing. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

The role of P2X3 receptors in bilateral masseter muscle allodynia in rats.

AimTo determine the relationship between bilateral allodynia induced by masseter muscle inflammation and P2X3 receptor expression changes in trigeminal ganglia (TRG) and the influence of intramasseteric P2X3 antagonist administration on bilateral masseter allodynia.MethodsTo induce bilateral allodynia, rats received a unilateral injection of complete Freund’s adjuvant (CFA) into the masseter muscle. Bilateral head withdrawal threshold (HWT) was measured 4 days later. Behavioral measurements were followed by bilateral masseter muscle and TRG dissection. Masseter tissue was evaluated histopathologically and TRG tissue was analyzed for P2X3 receptor mRNA expression by using quantitative real-time polymerase chain reaction (PCR) analysis. To assess the P2X3 receptor involvement in nocifensive behavior, two doses (6 and 60 μg/50 μL) of selective P2X3 antagonist A-317491 were administrated into the inflamed masseter muscle 4 days after the CFA injection. Bilateral HWT was measured at 15-, 30-, 60-, and 120-minute time points after A-317491 administration.ResultsHWT was bilaterally reduced after the CFA injection (P < 0.001). Intramasseteric inflammation was confirmed ipsilaterally to the CFA injection. Quantitative real-time PCR analysis demonstrated enhanced P2X3 expression in TRG ipsilaterally to CFA administration (P < 0.01). In comparison with controls, the dose of 6 μg of A-317491 significantly increased bilateral HWT at 15-, 30-, and 60-minute time points after the A-317491 administration (P < 0.001), whereas the dose of 60 μg of A-317491 was efficient at all time points ipsilaterally (P = 0.004) and at 15-, 30-, and 60-minute time points contralaterally (P < 0.001).ConclusionUnilateral masseter inflammation can induce bilateral allodynia in rats. The study provided evidence that P2X3 receptors can functionally influence masseter muscle allodynia and suggested that P2X3 receptors expressed in TRG neurons are involved in masseter inflammatory pain conditions.

In vitro comparison of three earwax removal formulations for the disintegration of earwax

Introduction: Impacted cerumen is a widespread reason that patients visit their health care providers. It effects approximately 2-6% of the general population and disproportionately impacts up to 65% of patients over 65. This study compared a new cerumen (earwax) removal product (Solution 1; EOS-002; a glycolic acid/bicarbonate formulation) versus two commercially available products (Solution 2 and Solution 3; both containing carbamide peroxide 6.5%) for their cerumenolytic activity in vitro. Methods: Samples of human cerumen were placed in 10 x 75 mm polypropylene test tubes. Approximately 1 mL of each test solution was added and incubated at room temperature for 30 minutes. The vials were shaken at the 15- and 30-minute time points to simulate rinsing in a clinical setting. Breakdown of the cerumen was graded at 5-, 10-, 15-, and 30-minute time points in a masked manner on a 5-point scale (Grade 0 = no change; Grade 4 = complete disintegration). Results: Significantly greater disintegration of the cerumen was observed in the samples exposed to EOS-002 at every time point (P &lt; 0.0001). At 5 minutes, disintegration was observed in 39 out of 43 samples exposed to EOS-002, 0 out of 24 samples exposed to Solution 2, and 1 out of 19 samples exposed to Solution 3. Mean disintegration scores at 5, 10, 15, and 30 minutes were 1.65, 2.38, 2.95, and 3.24 for EOS-002; 0, 0, 0, and 0.2 for Solution 2; and 0.05, 0.13, 0.16, and 0.21 for Solution 3, respectively. Discussion: EOS-002 exhibited a significantly greater ability to breakdown cerumen than the two other products. Disintegration of cerumen occurred with EOS-002 within 5 minutes in 91% (39/43) of the samples. Therefore, EOS-002 provides rapid disintegration of human cerumen in vitro.

Intravenous nutrients for preventing inadvertent perioperative hypothermia in adults.

Inadvertent perioperative hypothermia (a drop in core temperature to below 36°C) occurs because normal temperature regulation is disrupted during surgery, mainly because of the effects of anaesthetic drugs and exposure of the skin for prolonged periods. Many different ways of maintaining body temperature have been proposed, one of which involves administration of intravenous nutrients during the perioperative period that may reduce heat loss by increasing metabolism, thereby increasing heat production. To assess the effectiveness of preoperative or intraoperative intravenous nutrients in preventing perioperative hypothermia and its complications during surgery in adults. We searched the Cochrane Central Register of Controlled Trials (CENTRAL; November 2015) in the Cochrane Library; MEDLINE, Ovid SP (1956 to November 2015); Embase, Ovid SP (1982 to November 2015); the Institute for Scientific Information (ISI) Web of Science (1950 to November 2015); and the Cumulative Index to Nursing and Allied Health Literature (CINAHL, EBSCO host; 1980 to November 2015), as well as the reference lists of identified articles. We also searched the Current Controlled Trials website and ClincalTrials.gov. Randomized controlled trials (RCTs) of intravenous nutrients compared with control or other interventions given to maintain normothermia in adults undergoing surgery. Two review authors extracted data and assessed risk of bias for each included trial, and a third review author checked details if necessary. We contacted some study authors to request additional information. We included 14 trials (n = 565), 13 (n = 525) of which compared intravenous administration of amino acids to a control (usually saline solution or Ringer's lactate). The remaining trial (n = 40) compared intravenous administration of fructose versus a control. We noted much variation in these trials, which used different types of surgery, variable durations of surgery, and different types of participants. Most trials were at high or unclear risk of bias owing to inappropriate or unclear randomization methods, and to unclear participant and assessor blinding. This may have influenced results, but it is unclear how results might have been influenced.No trials reported any of our prespecified primary outcomes, which were risk of hypothermia and major cardiovascular events. Therefore, we decided to analyse data related to core body temperature instead as a primary outcome. It was not possible to conduct meta-analysis of data related to amino acid infusion for the 60-minute and 120-minute time points, as we observed significant statistical heterogeneity in the results. Some trials showed that higher temperatures were associated with amino acids, but not all trials reported statistically significant results, and some trials reported the opposite result, where the amino acid group had a lower core temperature than the control group. It was possible to conduct meta-analysis for six studies (n = 249) that provided data relating to the end of surgery. Amino acids led to a statistically significant increase in core temperature in comparison to those receiving control (MD = 0.46°C 95% CI 0.33 to 0.59; I2 0.0%; random-effects; moderate quality evidence).Three trials (n = 155) reported shivering as an outcome. Meta-analysis did not show a clear effect, and so it is uncertain whether amino acids reduce the risk of shivering (RR 0.36, 95% CI 0.13 to 1.00; I2 = 93%; random-effects model; very low-quality evidence). Intravenous amino acids may keep participants up to a half-degree C warmer than the control. This difference was statistically significant at the end of surgery, but not at other time points. However, the clinical importance of this finding remains unclear. It is also unclear whether amino acids have any effect on the risk of shivering and if intravenous nutrients confer any other benefits or harms, as high-quality data about these outcomes are lacking.

Effects of PAR2 antagonist on inflammatory signals and tight junction expression in protease‐activated canine primary epithelial keratinocytes

Effects of <scp>PAR</scp>2 antagonist on inflammatory signals and tight junction expression in protease‐activated canine primary epithelial keratinocytes

Time-Dependent Effects of Arthroscopic Conditions on Human Articular Cartilage: An In Vivo Study

To assess time-dependent effects of arthroscopic conditions on human articular cartilage invivo. From each of 10 patients undergoing multiligament reconstruction of the knee under our routine arthroscopic conditions (gravity irrigation of 0.9% normal saline solution at room temperature with 150cmH2O [110mm Hg] pressure and pneumatic tourniquet under 270mm Hg pressure), cartilage specimens were harvested from the lateral edge of the femoral notch at the beginning of the operation (baseline) and at 15-, 30-, 45-, and 60-minute time points during the operation. H&E staining and safranin O staining were used to evaluate the tissue structure, chondrocytes, and extracellular matrix (ECM) of the articular cartilage. Chondrocyte viability was evaluated, and a biochemical examination of the ECM was performed to detect changes in glycosaminoglycan and collagen content. The expression levels of genes associated with proinflammatory cytokines, ECM metabolism, and chondrocyte apoptosis of the articular cartilage were evaluated. At the 45- and 60-minute time points, an obvious impairment of tissue structure, a significant decrease in glycosaminoglycan content, and a significantly lower percentage chondrocyte viability were observed, as compared with baseline (P < .05). Regarding the tissue collagen content, no significant change was detected at any time point (P > .05). The gene expression examination at the 45- and 60-minute time points detected significant upregulation of interleukin 1β and tumor necrosis factor α (P < .05), indicating an inflammatory response by the chondrocytes, and significant upregulation of aggrecanase 1 (P < .05), which indicates catabolism or the disturbance of aggrecan metabolism. Under current arthroscopic conditions, an operative duration of 45minutes or longer can cause detrimental structural, biochemical, and metabolic effects on human articular cartilage. Arthroscopic skills should always be improved to reduce operation time and thus minimize the potential detrimental effects of arthroscopic conditions on articular cartilage.

Optimal Time Points for Scintigraphic Imaging of Pleuroperitoneal Shunts.

Nuclear imaging can confirm pleuroperitoneal shunt as the cause of pleural effusion. No society guidelines exist for scintigraphic pleuroperitoneal shunt detection. Our institutional protocol was evaluated to determine optimal imaging time points for shunt detection. Pleuroperitoneal shunt studies over 4 years were blindly reviewed by 2 nuclear radiologists. Data from blinded review included presence or absence of pleuroperitoneal shunt, laterality of shunt and time points for shunt detection. Chart review yielded 30 studies. Three cases were excluded because of improper injection. Imaging was positive for pleuroperitoneal shunt in 81% (22/27) of cases. In positive cases, activity was identified in the right hemithorax in 82% (18/22), left hemithorax in 9% (2/22), and bilaterally in 9% (2/22). One-hour imaging demonstrated 91% (20/22) of positive cases. The remaining 2 positive cases were negative at 1 hour but positive after 4 hours. No study was negative at 1 and 4 hours and positive at 24 hours. All negative cases (5/27) were confirmed on 24-hour imaging. The majority of positive pleuroperitoneal shunt examinations will demonstrate activity in the right hemithorax on 1-hour imaging. Although no case was negative at 1 and 4 hours and positive at 24 hours, imaging at 24 hours may still be necessary to confirm absence of shunt. Therefore, optimal imaging time points consist of early 1-hour and delayed 24-hour images if the 1-hour time point was negative. The 4-hour time point may be considered optional, thereby potentially optimizing patient safety and resource utilization.

Alcohol Withdrawal Increases Protein Kinase A Activity in the Rat Inferior Colliculus.

Cyclic AMP-dependent protein kinase A (PKA) signaling is a key target for the action of alcohol and may therefore play a role in the pathophysiology of alcohol withdrawal seizures (AWSs). Here, we investigated the role of PKA activity with respect to increased seizure susceptibility in rats that were subjected to alcohol withdrawal. Adult male Sprague Dawley rats received 3 daily doses of ethanol (EtOH) (or vehicle) for 4 consecutive days. Rats were then tested for susceptibility to acoustically evoked AWSs 3, 24, and 48hours after the last alcohol dose. In separate experiments, the inferior colliculus (IC) was collected at these same time points from rats subjected to alcohol withdrawal and control rats following alcohol withdrawal. PKA activity, catalytic Cα (PKACα ) protein, regulatory RIIα (PKARIIα ) protein, and RIIβ (PKARIIβ ) protein were measured in the IC. Lastly, insitu pharmacological studies were performed to evaluate whether inhibiting PKA activity in the IC suppressed AWSs. In the EtOH-treated group, AWSs were observed at the 24-hour time point, but not at the 3-hour or 48-hour time points. In the IC, PKA activity was significantly higher both 3hours (i.e., before AWS susceptibility) and 24hours after the last alcohol dose (when AWS susceptibility peaked) than in control rats. Consistent with these findings, protein levels of the PKACα subunit were significantly increased in the IC both 3 and 24hours after the last alcohol dose. Lastly, insitu inhibition of PKA activity within the IC suppressed AWSs. The increase in PKA activity and PKACα protein expression in the IC preceded the occurrence of AWSs, and inhibiting PKA activity within the IC suppressed acoustically evoked AWSs. Together, these findings suggest that altered PKA activity plays a key role in the pathogenesis of AWSs.

Delayed Mitogen-Activated Protein Kinase/Extracellular Signal–Regulated Kinase Inhibition by Trametinib Attenuates Systemic Inflammatory Responses and Multiple Organ Injury in Murine Sepsis*

The mitogen-activated protein kinase/extracellular signal-regulated kinase signaling pathway is an essential component of innate immunity necessary for mediating proinflammatory responses in the setting of sepsis. We previously demonstrated that the mitogen-activated protein kinase 1/2 inhibitor trametinib prevents endotoxin-induced renal injury in mice. We therefore assessed efficacy of trametinib in a more clinically relevant experimental model of sepsis. Controlled in vivo laboratory study. University animal research laboratory. Male C57BL/6 mice. Mice were subjected to cecal ligation and puncture to induce sepsis or underwent sham operation as controls. Six hours after cecal ligation and puncture, mice were randomized to four experimental groups as follows: 1) sham control; 2) sham control + trametinib (1 mg/kg, IP); 3) cecal ligation and puncture; and 4) cecal ligation and puncture + trametinib. All animals received buprenorphine (0.05 mg/kg, SC) and imipenem/cilastatin (14 mg/kg, SC) in 1.5 mL of warm saline (40 mL/kg) at the 6-hour time point. Mice were euthanized at 18 hours after induction of cecal ligation and puncture. Trametinib inhibition of mitogen-activated protein kinase/extracellular signal-regulated kinase signaling 6 hours after cecal ligation and puncture attenuated increases in circulating proinflammatory cytokines (tumor necrosis factor-α, interleukin-1β, interleukin-6, and granulocyte macrophage colony-stimulating factor) and hypothermia at 18 hours. Trametinib also attenuated multiple organ injury as determined by serum creatinine, alanine aminotransferase, lactate dehydrogenase, and creatine kinase. At the organ level, trametinib completely restored peritubular capillary perfusion in the kidney. Restoration of microvascular perfusion was associated with reduced messenger RNA expression of well-characterized markers of proximal tubule injury. mitogen-activated protein kinase/extracellular signal-regulated kinase blockade attenuated cecal ligation and puncture-mediated up-regulation of cytokines (tumor necrosis factor-α, interleukin-1β) and restored interleukin-6 to control levels in the renal cortex, indicating the protective effects on the proximal tubule occur primarily through modulation of the proinflammatory response in sepsis. These data reveal that the mitogen-activated protein kinase/extracellular signal-regulated kinase inhibitor trametinib attenuates systemic inflammation and multiple organ damage in a clinically relevant model of sepsis. Because trametinib has been safely used in humans, we propose that this drug might represent a translatable approach to limit organ injury in septic patients.

Gray matter alterations and correlation of nutritional intake with the gray matter volume in prediabetes.

The neurophysiology of prediabetes plays an important role in preventive medicine. The dysregulation of glucose metabolism is likely linked to changes in neuron-related gray matter. Therefore, we designed this study to investigate gray matter alterations in medication-naive prediabetic patients. We expected to find alterations in the gray matter of prediabetic patients.A total of 64 prediabetic patients and 54 controls were enrolled. All subjects received T1 scans using a 3-T magnetic resonance imaging machine. Subjects also completed nutritional intake records at the 24-hour and 3-day time points to determine their carbohydrate, protein, fat, and total calorie intake. We utilized optimized voxel-based morphometry to estimate the gray matter differences between the patients and controls. In addition, the preprandial serum glucose level and the carbohydrate, protein, fat, and total calorie intake levels were tested to determine whether these parameters were correlated with the gray matter volume.Prediabetic patients had lower gray matter volumes than controls in the right anterior cingulate gyrus, right posterior cingulate gyrus, left insula, left super temporal gyrus, and left middle temporal gyrus (corrected P < 0.05; voxel threshold: 33). Gray matter volume in the right anterior cingulate was also negatively correlated with the preprandial serum glucose level gyrus in a voxel-dependent manner (r = –0.501; 2-tailed P = 0.001).The cingulo-temporal and insula gray matter alterations may be associated with the glucose dysregulation in prediabetic patients.

The Effect of Acute Hyperglycemia on Muscular Strength, Power and Endurance.

Chronic hyperglycemia has been shown to cause skeletal muscle dysfunction, however, there is a paucity of understanding on the effect of acute hyperglycemia on skeletal muscle function. PURPOSE: The purpose of this study is to better understand the impact of acute hyperglycemia on skeletal muscle strength, power, and endurance. METHODS: Ten male collegiate athletes (age 21.5 ± 1.5 years, height 186 ± 2.03 cm, body mass 108.8 ± 7.6 kg, % body fat (7-site skinfold) 22.5 ± 3.09%) participated in 2 testing sessions, separated by 7 days and randomized for either high glucose (HG) or control (C) treatment groups. The HG group consumed a high glucose drink (2 g glucose/kg body weight) while controls consumed an isocaloric nutrition bar with an energy provision of 40% protein, 30% fat, and 30% carbohydrate. Blood glucose (BC) levels for HG and C were tested at 0 (basal) and 30, 60, 90, and 120 minutes (mins) post consumption. At 30 mins post consumption, muscular strength in the HG and C groups was assessed by a 1RM bench press (BP) test followed by lower body power at 60 mins via vertical jump test. Muscular endurance was examined with a 3-set-to-failure BP test at 90 mins. RESULTS: The HG group exhibited significantly greater BC values (p<0.05) at the 30, 60, 90, and 120 minute time points. Additionally, HG glucose area under the curve was significantly greater (p<0.05) than C and was positively correlated with %body fat, a finding that trended towards significance, r = 0.587, n= 10, p = 0.074. There were no between group differences in maximal strength, power or muscular endurance. CONCLUSION: Although performance measures were unaffected by acute hyperglycemia, we provide evidence that acute hyperglycemia can be induced and maintained in healthy, active and young subjects which may provide a valuable platform for future research on the etiological time course of metabolic diseases.

The Relationship Between Choroidal Expansion and Intraocular Pressure Rise During the Water Drinking Test in Healthy Subjects and Patients With Exfoliation Syndrome.

To evaluate the changes in choroidal thickness (ChT) and intraocular pressure (IOP) during the water drinking test (WDT) in patients with exfoliation syndrome (XFS). In this prospective study, 20 eyes of 20 patients with XFS and 20 eyes of 20 healthy subjects underwent measurements of subfoveal ChT, IOP, systolic and diastolic blood pressure before and 30 and 60 minutes after the WDT. WDT involved ingestion of 1 L of water within 5 minutes. χ test and analysis of variance were used for the statistical analysis. There were no significant differences between the 2 groups with respect to age (P = 0.238), sex (P = 1.000), central corneal thickness (P = 0.714), axial length (P = 1.000), diastolic blood pressure (P = 0.350), systolic blood pressure (P = 0.064), and IOP (P = 0.234) before water ingestion. Baseline ChT was lower in the XFS group (285.1 ± 69.1 μm) as compared with that of controls (339.4 ± 81.2 μm) (P = 0.028). ChT was found to be significantly increased at 30 and 60 minutes after water ingestion in the XFS group (P < 0.001), whereas in the control group an increase was found only at the 60-minute time-point (P = 0.028). Independent of baseline ChT, at 60-minute endpoint, the magnitude of choroidal expansion was higher in XFS eyes (13.2 ± 14.3%) compared with control eyes (6.2 ± 3.5%) (P = 0.028). The mean IOP was higher at all time-points with respect to their respective baseline values in both XFS (P = 0.002, P < 0.001) and control (P = 0.038, P = 0.002) eyes. Choroidal expansion during WDT does not seem to be an important mechanism for IOP elevation in healthy eyes or those with XFS. Furthermore, the similarity of IOP increase to controls suggests that trabecular outflow facility is not impaired in eyes with XFS.

Noninvasive Transcutaneous Vagus Nerve Stimulation Decreases Whole Blood Culture-Derived Cytokines and Chemokines: A Randomized, Blinded, Healthy Control Pilot Trial.

The purpose of this study was to test the transcutaneous noninvasive vagus nerve stimulator (nVNS) (gammaCore©) device to determine if it modulates the peripheral immune system, as has been previously published for implanted vagus nerve stimulators. A total of 20 healthy males and females were randomized to receive either nVNS or sham stimulation (SST). All subjects underwent an initial blood draw at 8:00 am, followed by stimulation with nVNS or SST at 8:30 am. Stimulation was repeated at 12:00 pm and 6:00 pm. Additional blood samples were withdrawn 90 min and 24 hour after the first stimulation session. After samples were cultured using the Myriad RBM TruCulture (Austin, TX) system (WBCx), levels of cytokines and chemokines were measured by the Luminex assay and statistical analyses within and between groups were performed using the Wilcoxon Signed Ranks Test and Mann-Whitney U with the statistical program R. A significant percent decrease in the levels of the cytokine interleukin [IL]-1β, tumor necrosis factor [TNF] levels, and chemokine, interleukin [IL]-8 IL-8, macrophage inflammatory protein [MIP]-1α, and monocyte chemoattractant protein [MCP]-1 levels was observed in the nVNS group non-lipopolysaccharide (LPS)-stimulated whole blood culture (n-WBCx) at the 24-hour time point (p < 0.05). In SST group, there was a significant percent increase in IL-8 at 90 min post-stimulation (p < 0.05). At 90 min, the nVNS group had a greater percent decrease in IL-8 concentration (p < 0.05) compared to SST group. The nVNS group had a greater percent decrease in cytokines (TNF, IL-1β) and chemokines (MCP-1 and IL-8) at 24 hour (p < 0.05) in comparison to SST. LPS-stimulated whole blood cultures (L-WBCx) did not show a significant decrease in cytokine levels in either the nVNS or SST group across any time points. The nVNS group showed a significant percent increase in LPS-stimulated IL-10 levels at the 24-hour time point in comparison to SST. nVNS downregulates inflammatory cytokine release suggesting that nVNS may be an effective anti-inflammatory treatment.

Physiological Responses and Hedonics During Prolonged Physically Interactive Videogame Play.

This study was designed to assess physiologic responses and hedonics (i.e., liking) during prolonged physically interactive videogame play. Participants (n = 24) completed three 30-minute videogame conditions on separate days in a random order. During two of the conditions participants played physically interactive videogames (Nintendo of America, Inc. [Redmond, WA] "Wii™ Fit" "Basic Run" and "Basic Step"). During the third condition participants played a traditional/sedentary game ("Tanks!"), which required minimal physical movement for gameplay. Oxygen consumption (VO2) was assessed using indirect calorimetry throughout each condition and averaged every 5 minutes. Liking was assessed via visual analog scale at the 15- and 30-minute time points during each condition. Mean VO2 was significantly (P < 0.001) greater during "Basic Run" (16.14 ± 5.8 mL/kg/minute, 4.6 ± 1.7 metabolic equivalents [METs]) than either "Basic Step" (11.4 ± 1.7 mL/kg/minute, 3.3 ± 0.5 METs) or the traditional/sedentary videogame (5.39 ± 1.0 mL/kg/minute, 1.5 ± 0.1 METs). "Basic Step" was also greater (P < 0.001) than the traditional/sedentary game. VO2 did not significantly (P = 0.25) fluctuate across the 30-minute session for any game. In other words, participants maintained a consistent physiologic intensity throughout each 30-minute condition. There were no differences (P ≥ 0.20) across gaming conditions or time for liking. Participants achieved and maintained moderate-intensity physical activity (≥3.0 METs) during both 30-minute physically interactive videogame conditions. Furthermore, because liking was similar across all gaming conditions, participants may be willing to substitute the physically interactive videogames in place of the traditional/sedentary game.

(+)-Pentazocine Reduces NMDA-Induced Murine Retinal Ganglion Cell Death Through a σR1-Dependent Mechanism.

PurposeTo evaluate, in vivo, the effects of the sigma-1 receptor (σR1) agonist, (+)-pentazocine, on N-methyl-D-aspartate (NMDA)-mediated retinal excitotoxicity.MethodsIntravitreal NMDA injections were performed in C57BL/6J mice (wild type [WT]) and σR1−/− (σR1 knockout [KO]) mice. Fellow eyes were injected with phosphate-buffered saline (PBS). An experimental cohort of WT and σR1 KO mice was administered (+)-pentazocine by intraperitoneal injection, and untreated animals served as controls. Retinas derived from mice were flat-mounted and labeled for retinal ganglion cells (RGCs). The number of RGCs was compared between NMDA and PBS-injected eyes for all groups. Apoptosis was assessed using TUNEL assay. Levels of extracellular-signal–regulated kinases (ERK1/2) were analyzed by Western blot.ResultsN-methyl-D-aspartate induced a significant increase in TUNEL-positive nuclei and a dose-dependent loss of RGCs. Mice deficient in σR1 showed greater RGC loss (≈80%) than WT animals (≈50%). (+)-Pentazocine treatment promoted neuronal survival, and this effect was prevented by deletion of σR1. (+)-Pentazocine treatment resulted in enhanced activation of ERK at the 6-hour time point following NMDA injection. The (+)-pentazocine–induced ERK activation was diminished in σR1 KO mice.ConclusionsTargeting σR1 activation prevented RGC death while enhancing activation of the mitogen-activated protein kinase (MAPK), ERK1/2. Sigma-1 receptor is a promising therapeutic target for retinal neurodegenerative diseases.

Rapamycin Promotes the Autophagic Degradation of Oxidized Low-Density Lipoprotein in Human Umbilical Vein Endothelial Cells

Background/Aims: Oxidized low-density lipoprotein (ox-LDL) has been extensively implicated in the initiation of atherosclerosis. Our previous studies reported that ox-LDL could activate autophagy in human umbilical vein endothelial cells (HUVECs). Because of this, subsequent studies were designed to elucidate the possible role of the autophagic inducer, rapamycin, on ox-LDL degradation in endothelial cells. Methods: Intracellular cholesterol content was measured using a tissue total cholesterol assay kit. ox-LDL trafficking within endothelial cells was analyzed by flow cytometry. Levels of proteins involved in the autophagic process, microtubule-associated protein 1 light chain 3 (MAP1-LC3), lysosome-associated membrane protein 1 (LAMP1), Beclin 1 and p62, were assessed by Western blot analysis. Results: We discovered that rapamycin could decrease the ox-LDL content in HUVECs at the 3-hour time point. Rapamycin also mediated an obvious increase in Dil-labeled ox-LDL (Dil-ox-LDL)/LC3 and Dil-ox-LDL/LAMP1 co-localization, which was inhibited by 3-methyladenine (3-MA), an autophagic inhibitor. In addition, significant co-localization of LC3 and LAMP1 occurred in cells pretreated with rapamycin. In the presence of rapamycin, p62 levels were reduced, and autophagic flux was enhanced. Conclusion: These data demonstrate that the activation of the autophagy-lysosome pathway by rapamycin may accelerate ox-LDL degradation.

Validation of a Simple Thyroid Cancer Dosimetry Model Based on the Fractional Whole-Body Retention at 48 Hours Post-Administration of (131)I.

Standard dosimetric methods to determine the maximum tolerated activity (MTA) of (131)I for the treatment of metastatic, well-differentiated thyroid cancer (DTC) are time-consuming and require complex analysis. As a result, reliable, accurate, and simplified methods are desirable. The objective of this study was to evaluate the validity of a simple regression dosimetry model. Previously, the authors reported a bi-exponential model for estimating the MTA of (131)I for the treatment of metastatic DTC based on a limit of 2 Gy to the blood. This model uses the patient's body surface area (BSA) along with the fractional whole-body retention (WBR) at 48 hours following oral administration of a diagnostic dosage of (131)I. A bi-exponential regression model was developed between the MTA normalized to the patient's BSA and the percent retention value at the 48-hour time point (R): MTA (GBq)/BSA (m(2)) = (13.91 · e(-0.0387R) + 42.33 · e(-0.8522R)). In this study, the same model was applied to a different set of adult patients referred for dosimetry and possible (131)I treatment of DTC under conditions of thyroid hormone withdrawal or recombinant human thyrotropin (rhTSH) stimulation. All patients (n = 170; 96 female) referred to the authors' clinic for dosimetry and possible (131)I treatment for metastatic DTC during the collection period were included in this study, apart from those undergoing renal dialysis. The MTA predicted (MTAp) using the model described above was compared to the measured MTA (MTAm), with statistical analysis performed using ProStat v4.5. In this group, the MTAm ranged from 2.3 to 41.1 GBq. The linear correlation between the MTAp and MTAm was excellent (r = 0.96), with an average deviation of only ± 1.2%. However, to avoid overdosing a patient on the basis of the MTAp, a weighting factor (<1.0) should be applied (e.g., using a value of 0.7 would result in only one patient receiving a prescribed activity of (131)I that exceeded the MTAm [<3%]). The % 48-hour WBR as determined by the bi-exponential function noted herein with reasonable restrictions has been validated as a reliable simplified dosimetry model.

Clinical efficacy of percutaneous vertebroplasty combined with intensity-modulated radiotherapy for spinal metastases in patients with NSCLC.

ObjectiveThis study aimed to evaluate the safety and efficacy of percutaneous vertebroplasty (PVP) combined with intensity-modulated radiotherapy (IMRT) for metastatic lesions of patients with non-small-cell lung cancer (NSCLC) at centrum vertebrae.MethodsA total of 39 patients with spinal metastatic NSCLC (stage IV) were treated with PVP followed by IMRT (30 Gy/10F/2 W) for metastatic lesion at centrum vertebrae under local anesthesia. Retrospective analysis was done with medical records and radiological data. The change of visual analog scale (VAS), activities of daily living, and kyphotic angle was measured preoperatively. The presence of complications was assessed preoperatively (baseline) at 24 hours, 1 week, and 1, 3, 6, 12, and 24 months postoperatively, or until the patient died or was lost to follow-up. Survival was assessed in the group.ResultsA total of 39 consecutive patients were successfully treated with PVP via a translateral approach and IMRT. Their mean VAS score decreased from 7.93±1.09 preoperatively to 4.14±1.15 by the 24-hour postoperative time point and was 3.92±1.23 at 1 week, 4.27±1.93 at 1 month, 3.24±1.35 at 3 months, 2.27±0.96 at 6 months, and 2.59±1.55 at 12 months after the procedure. The mean VAS score at all of the postoperative time points was decreased significantly from the preoperative baseline score (P<0.05). Activities of daily living evaluation showed that the patients had a significantly high life quality after the combined approach (50.9±11.7 vs 82.3±9.9, P<0.05). No severe complications were observed. Mild complications included two cases (5.13%) of asymptomatic cement leakage into the epidural space and one case (2.56%) of paravertebral leakage. Median survival time was extended to 13 months.ConclusionThe safety and efficacy of PVP combined with IMRT in patients with NSCLC with metastatic lesions at centrum vertebrae and the ability to prevent the diseased vertebrae from further deformation and tumor infiltration into the vertebral body were demonstrated. PVP combined with IMRT should be the optimal technique for treatment of vertebral compression pain resulting from spinal metastatic NSCLC.

Does intravenous atropine affect stroke volume variation in man?

ObjectivesCurrently there are no reports of the effect of increasing heart rate (HR) induced by intravenous atropine on stroke volume variation (SVV). We hypothesized that increasing HR alters the value of SVV. This prospective study aimed to investigate changes in SVV values by increasing HR induced by intravenous atropine in patients with good cardiac function. We also re-evaluated the effect of intravenous atropine alone on hemodynamics including new hemodynamic parameters such as SVV. MethodsPatients were chosen as participants of this study if, 30 minutes after anesthesia induction, HR was below 65 beats/min. Baseline hemodynamic values were recorded, and then the patients received intravenous atropine (0.01 mg/kg; max 0.5 mg). These values were recorded again after intravenous atropine every minute for 5 minutes. ResultsTen American Society of Anesthesiologists (ASA) physical status I–II patients aged 37–65 years who were scheduled for elective surgery were included. Intravenous atropine significantly increased HR at the 1–5 minute time points, mean arterial pressure at the 1–4 minute time points, and cardiac output at the 1–3 minute time points compared with baseline values but did not significantly change SVV, stroke volume index, pressure of end-tidal CO2, and systemic vascular resistance. ConclusionAdministration of intravenous atropine did not change SVV, and we present this as a novel finding.

Thrombin-activatable Fibrinolysis Inhibitor (TAFI) as a Novel Prognostic Factor After Orthotropic Liver Transplantation: A Pilot Study

BackgroundThrombin-activatable fibrinolysis inhibitor (TAFI), a liver-produced coagulation factor, has been associated with higher mortality in cirrhotic patients, but there has not been any description of its role in perioperative care in liver transplantation cases. MethodsA total of 21 patients were included. Serum TAFI levels were determined at 3 time points: preoperatively (TAFI pre), immediately postoperative (TAFI PO), and 24 hours postoperatively (TAFI 24 h). The main outcome was the physiological pattern of TAFI in the perioperative period of liver transplantation. The secondary outcomes were the association between TAFI and early allograft dysfunction (EAD) as well as that of TAFI and 6-month mortality. ResultsTAFI levels increased at the 24-hour time point, compared to the other 2 time points (TAFI pre, P = .007; TAFI PO, P = .0001). Early allograft dysfunction occurred in 2 of 21 patients, both demonstrating lower TAFI 24 h levels compared to those who did not develop this complication (3.0 ± 0.2 vs 1.5 ± 0.3; P = .0001). Three patients who died all demonstrated lower levels of TAFI pre (1.3 ± 0.1 vs 2.5 ± 0.5; P = .001) and TAFI PO (1.2 ± 0.1 vs 2.4 ± 0.4; P = .001) compared to the survivors. ConclusionsThese findings suggest that the determination of TAFI levels—both pre- and postoperatively—may be of clinical relevance in liver transplant recipients.