Academic literature on the topic 'R-30ib'

Hemophilia A (HA) is the most common X-linked bleeding disorder caused by a deficiency in coagulation factor VIII (FVIII). Inhibitor development in up to 30% of HA patients is the most challenging complication in HA treatment and has been associated with several patient-related risk factors such as gene mutation and residual plasma factor VIII concentrations. Treatment related risk factors are under debate. However, HCV infections were recently shown to correlate with low level inhibitor abundance, potentially as a result of chemokine and cytokine regulation. Therefore, the aim of this study was to evaluate a potential connection between proinflammatory cytokine levels and FVIII-specific B cell responses in HA patients without infections, but with other comorbidities that might modulate cytokine levels. Thus, several clinical parameters and concentrations of plasmatic proinflammatory cytokines (IL-1ß, IL-6, IL-8, IL-10, IL-12p70, TNF, IFNa, IFNg, TNFa, MCP-,1 IL-17A, IL-18, IL-23 and IL-33) were quantified from different patient samples. At first, the influence of proinflammatory cytokines on inhibitor development was assessed in a HA patient, who had an intron 22 inversion and was also diagnosed with type 1 diabetes. Prophylactic treatment began at the age of 9 months with 30IU/kg Octanate, and after four days of exposure, the formation of inhibitors was initially detected. Longitudinal cytokine analysis in plasma samples of that patient revealed differential regulation patterns of cytokines. IL-1b, IFNg, TNFa, IL-6, IL-10 and IL-12p70 tended to increase upon inhibitor occurrence, whereas other cytokines remained unchanged (MCP-1, IL-8, IL-17A, IL-18 and IL-23) or decreased (IFNa2 and IL-33). Next, a correlation analysis was performed, assuming that proinflammatory cytokines might stimulate B cells to produce antibodies. This analysis involved all cytokines that were found to be elevated upon inhibitor formation. The comparison of clinical and laboratory data from that patient revealed a strong association of inhibitor levels and IL-10 expression (Pearson r = 0,9391; R squared = 0,8818) as well as with IL-6 (Pearson r = 0,9290; R squared = 0,8630). Furthermore, a modest correlation was observed for IL-1b (Pearson r = 0,5277; R squared = 0,2785), IFNg (Pearson r = 0,3413; R squared = 0,1165), IL-12p70 (Pearson r = 0,3547; R squared = 0,1258), and TNFa (Pearson r = 0,3070; R squared = 0,09422). Other proinflammatory cytokines with a rather stable profile upon inhibitor initiation like MCP-1 (Pearson r = -0,06810; R squared = 0,004638) and IL-18 (Pearson r = 0,2245; R squared = 0,05042) seemed not to be directly correlated with inhibitor occurrence. Finally, we aim to investigate the potential impact of IL-10 and IL-6 on inhibitor production and/or FVIII immune tolerization across various patient groups and conditions. Therefore, we will compare patients with detectable inhibitor titers, those who have successfully eradicated inhibitors, and individuals without inhibitor titers (with normal recovery). Preliminary data suggest a positive correlation between both cytokines and active inhibitor presence. However, to reach a definitive conclusion, the sample size will be/should be increased.

Abstract Wire and arc additive manufacturing (WAAM) is a popular direct energy deposition (DED) method for producing large-scale metallic components. The main advantages of the technique are a high deposition rate and low cost. Furthermore, the utilization of the WAAM is prevalent in the aerospace industry. The AA5087 aluminium alloy with 4.5 wt.% of magnesium has been investigated because of its excellent properties. The present research deals with the study of thermal cycles and fields developed in the alloy during additive manufacturing with two different Cold metal transfer (CMT) modes, namely conventional (CMT) and cycle-step (CMT-CS). The welding system was equipped with a Fronius TransPulse Synergic 3200 CMT power source, a Fanuc Arc Mate 1000iC 6-axes robot with an R 30iA control unit, a welding torch, and a 1-axis positioner. The AA5087 aluminium alloy welding wire with a diameter of 1.2 mm was deposited onto the AA5083 aluminium alloy plate with dimensions of 70 mm x 200 mm x 3 mm during the experiment. The thermal cycles were documented using an Ahlborn Almemo 5690-2 measuring station equipped with K-type thermocouples. The thermal fields were monitored with a FLIR E95 thermography camera. The results showed the evident influence of arc mode on the temperatures developed in manufactured aluminium alloy parts during the process of WAAM.

Abstract Background Total calcium is a less accurate test in predicting ionized calcium (Ca2+) in patients suspected of calcium metabolic disease. Nevertheless, total calcium continues to be used as routine measurement instead of adjusted Ca2+ (at pH 7.4). In the current study we evaluate a new multichannel instrument, the ISE Module E1200 for adjusted Ca2+ (at pH 7.4), containing three different ion-selective electrode (ISE) units. Methods Serum from 1350 patients was compared to the ABL835 flex and KoneLab. Total calcium was also evaluated on the Dimension Vista 1500 system. Correlations between instruments were assessed by Deming regression and degree of agreement by Cohen’s kappa (κ). Results Analytical imprecisions for the three ISE units for adjusted Ca2+ (at pH 7.4) was between 0.36% and 2.52%, and for pH between 0.32% and 3.24%. Results were comparable for each ISE unit (r = 0.797–0.917; all P < 0.0001) and in high-throughput settings (r = 0.871; P < 0.0001). The degree of agreement between instruments was moderate to good (κ = 0.52–0.77). In contrast, there was a very poor agreement (κ = −0.14) for total calcium with discrepancy in 53.4% of the samples. Conclusions The new ISE Module E1200 is comparable with the ABL835 flex and KoneLab 30i and therefore may be used for routine analysis of serum adjusted Ca2+ (at pH 7.4). The measured adjusted Ca2+ (at pH 7.4) was less comparable with very poor agreement to total calcium measured on the Dimension Vista 1500 system.

Pilots of high-performance aircraft are subject to transient loss of consciousness due to cerebral ischemia resulting from sudden high gravitational stress. To assess the effects of gravitational stress-induced blackout on cerebral metabolism and electrical function, we developed an animal model in which global cerebral ischemia is produced repeatedly at short intervals. Rats were prepared by ligation of subclavian and external carotid arteries and the right carotid artery was cannulated bidirectionally to measure circle of Willis and systemic pressures. Ischemia was induced by inflation of an occluder about the left carotid artery. Interleaved 31P and 1H NMR spectra were acquired on a 4.7-T Biospec system simultaneously with EEG recordings. We report results from 20 experiments of 30-min duration in which rats were subject to 30 1-min ischemia:reflow cycles of 10I:50R, 20I:40R, 30I:30R, and 40I:20R [numbers are seconds of ischemia (I) and reflow (R) during each 1-min cycle]. During ischemia the graded delivery of the ischemic insult permitted direct correlations between 2- to 5- and 7- to 20-Hz EEG activity and progressive changes in pH, lactate, ATP, phosphocreatine (PCr) and Pi. The best correlations were found between EEG activity and pH and PCr; correlation coefficients ranged from 0.93 to 0.95. A loss of EEG activity was observed without significant sustained energy loss in all but the most severe cycle.

Abstract Introduction: Maternal obesity is an independent risk factor for reduced reproductive fitness. Decreased secretion of FSH in women with obesity is well documented but poorly understood. Furthermore, obese women secrete less protein and steroid hormones from their ovaries. In mice, prior studies have demonstrated that pulsatile release of FSH enhances ovarian function and fertility. Hypothesis: We hypothesize that insufficient FSH pulsatility, as seen in women with obesity, results in inadequate folliculogenesis and reduced ovarian steroid production. We attempt to correct pulsatile FSH secretion in obese women by administering exogenous FSH to compensate for the suppressed circulating ovarian hormones. Our primary outcome is the change in peak inhibin B between pre- and post-treatment. We present results from our interim analysis. Methods: Reproductive aged, regularly menstruating, normal weight (NW) (BMI 18.5-24.9) and obese (OB) (BMI >30) women were recruited for a 26hr study during the early follicular phase. Frequent blood sampling (q10min) for 10h was performed to obtain baseline hormone levels. At 10h, 3 mg of cetrorelix, a gonadotropin hormone antagonist, was given followed by a secondary dose (0.25mg) 6h later. At this time, hourly IV recombinant (r)FSH (30IU) was initiated and frequent blood sampling continued for 10h. LH, FSH, estradiol (E2) were measured by immunoassay (Advia Centaur XP, Siemens). Inhibin B was measured using an ELISA kit (Ansh labs). Differences between groups were modeled by linear regression, adjusted for age and cycle day (continuous). The relationship between change in peak inhibin B and change in peak E2 was estimated in a linear regression. Results: A total of 36 participants (19 NW and 17 OB) were included in our interim analysis. There were no differences in age, cycle day of study, race, and waist/hip ratio. Inhibin B and E2 rises following the intervention were statistically significant within each group. Peak Inhibin B and E2 levels following intervention were lower in obese women compared to normal weight (133.4 vs 202.5 pg/mL and 85.8 vs 126.4 pg/mL, respectively). The difference in pre and post peak inhibin B levels trended lower in the obese group (-40.1 (95%CI: -86.2, 6.1, p=0.087). No difference was seen in maximal E2 response. There was no relationship between inhibin B and E2 response [0.08 (95%CI -0.26, 0.42), p=0.634]. Conclusions: These early results suggest obese women may have a lower response to pulsatile rFSH as compared to normal weight counterparts even with intravenous administration. We speculate this may be due to decreased uptake of rFSH in obese patients or a sign of ovarian dysfunction in obese women. Additional subjects are recruited to detect these differences.

Introduction. Leptin resistance (LR) is characterized by a weakening of the positive metabolic effect of leptin, despite its elevated level, while maintaining some of the pleiotropic effects, including those on the cardiovascular system. However, since most of the data have been obtained in cellular and animal models, the role of LR in cardiovascular diseases remains unclear. The lack of precise diagnostic criteria for assessing LR limits the study of this phenomenon. As a result, data on the incidence and contribution of LR in MI are critically scarce today, in addition, they are extremely contradictory. Purpose: to evaluate the prevalence of LR and its clinical and prognostic significance in conjunction with metabolic disorders in the hospital period of myocardial infarction (MI). Materials and methods. The study included 114 men with an established diagnosis of ST elevation MI. Patients on the 1st and 12th day of MI measured the concentration of leptin, the leptin receptor. The free leptin index (FLI) was calculated as the ratio of leptin concentration (ng/mL) to the concentration of soluble leptin receptor (ng/mL) multiplied by 100. LR was recorded at leptin > 6.45 ng/mL and LL >25. Assessment of glucose, lipid spectrum (total cholesterol, TAG, LDL-C, VLDL-C, HDL-C, FFA) in blood serum was performed using standard test systems from Thermo Fisher Sientific on an automatic biochemical analyzer Konelab 30i (Finland) , the content of C-peptide and insulin were determined using enzyme immunoassay (Monobind, USA) according to the protocol established by the manufacturer. To determine insulin resistance (IR), the QUICKI index was calculated, the severity of IR was assessed according to A. Katz et al. A comparative analysis of clinical and anamnestic characteristics and cardiovascular prognosis between patients with and without PR was carried out. Data analyzed using Statistica 10.0. Results. In patients with MI in the whole group in the hospital period, there was an increased content of leptin relative to the reference interval of 2.0-5.6 ng/ml. On the 1st and 12th days of the disease, the leptin concentration in patients with MI was 11.6 [6.6;20.5] ng/ml and 11.5 [5.4;13.9] ng/ml, respectively. . The content of the leptin receptor did not go beyond the established reference interval. LSI on the 1st day of the disease was 32.7 [14.3; 70.5], on the 12th day - 31.9 [16.2; 64.5]. The prevalence of LR in the hospital period for MI was 64%. LR was associated with CVD risk factors - hereditary burden for cardiovascular pathology (p=0.02), arterial hypertension (p=0.01), dyslipidemia (p=0.001), obesity (p=0.001). When assessing the metabolic profile, a statistically significant increase in the content of glucose (p=0.02), insulin (p=0.02) and C-peptide (p=0.03) on the 1st day of MI, insulin (p=0 .01) and C-peptide (p=0.03) on the 12th day of the disease, a decrease in the QUICKI index (p=0.03) throughout the hospital period in patients with PR compared with patients without PR. In the group of patients with PR, 45 people (61.8%) had a moderate and severe degree of IR, in patients without PR - in 12 patients (29.2%). When conducting a correlation analysis, a significantly significant direct relationship was found between the level of insulin on the 12th day of MI and LSI (r=0.509, p=0.02), as well as an inverse correlation between the QUICKI index on the 12th day and LSI (r= -0.367, p=0.01). Among the studied parameters of lipid metabolism, only the content of FFA on the 1st day of the disease in the group of patients with LR was higher than in the group without LR (p=0.03). Patients with LR were more often prone to early postinfarction angina (p=0.03), recurrent MI (p=0.001), rhythm and conduction disturbances (p=0.03) during the hospital period of MI. Prognostic significance in relation to the risk of adverse cardiovascular events in the hospital period of MI, according to logistic regression analysis, had LSI both on the 1st and 12th days of the disease, as well as FFA levels on the 1st day of the disease. Conclusion. Patients with MI are characterized by a high prevalence of LR in the hospital period. LR is associated with CVD risk factors, metabolic disorders, and IR formation. The identified features in the presence of LR can probably contribute to the development of adverse cardiovascular events in the hospital period of MI.

Hamsters were exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days. Equal amounts of proteins from urinary bladder or liver extracts of control and arsenic-treated hamsters were labeled with Cy3 and Cy5 dyes, respectively. After differential in gel electrophoresis and analysis by the DeCyder software, several protein spots were found to be down-regulated and several were up regulated. Our experiments indicated that in the bladder tissues of hamsters exposed to arsenite, transgelin was down-regulated and GST-pi was up-regulated. The loss of transgelin expression has been reported to be an important early event in tumor progression and a diagnostic marker for cancer development [29-32]. Down-regulation of transgelin expression may be associated with the carcinogenicity of inorganic arsenic in the urinary bladder. In the liver of arsenite-treated hamsters, ornithine aminotransferase was up-regulated, and senescence marker protein 30 and fatty acid binding protein were down-regulated. The volume ratio changes of these proteins in the bladder and liver of hamsters exposed to arsenite were significantly different than that of control hamsters. Introduction Chronic exposure to inorganic arsenic can cause cancer of the skin, lungs, urinary bladder, kidneys, and liver [1-6]. The molecular mechanisms of the carcinogenicity and toxicity of inorganic arsenic are not well understood [7-9). Humans chronically exposed to inorganic arsenic excrete MMA(V), DMA(V) and the more toxic +3 oxidation state arsenic biotransformants MMA(III) and DMA (III) in their urine [10, 11], which are carcinogen [12]· After injection of mice with sodium arsenate, the highest concentrations of the very toxic MMA(III) and DMA(III) were in the kidneys and urinary bladder tissue, respectively, as shown by experiments of Chowdhury et al [13]. Many mechanisms of arsenic toxicity and carcinogenicity have been suggested [1, 7, 14] including chromosome abnormalities [15], oxidative stress [16, 17], altered growth factors [18], cell proliferation [19], altered DNA repair [20], altered DNA methylation patterns [21], inhibition of several key enzymes [22], gene amplification [23] etc. Some of these mechanisms result in alterations in protein expression. Methods for analyzing multiple proteins have advanced greatly in the last several years. In particularly, mass spectrometry (MS) and tandem MS (MS/MS) are used to analyze peptides following protein isolation using two-dimensional (2-D) gel electrophoresis and proteolytic digestion [24]. In the present study, Differential In Gel Electrophoresis (DIGE) coupled with Mass Spectrometry (MS) has been used to study some of the proteomic changes in the urinary bladder and liver of hamsters exposed to sodium arsenite in their drinking water. Our results indicated that transgelin was down-regulated and GST-pi was up-regulated in the bladder tissues. In the liver tissues ornithine aminotransferase was up-regulated, and senescence marker protein 30, and fatty acid binding protein were down-regulated. Materials and Methods Chemicals Tris, Urea, IPG strips, IPG buffer, CHAPS, Dry Strip Cover Fluid, Bind Silane, lodoacetamide, Cy3 and Cy5 were from GE Healthcare (formally known as Amersham Biosciences, Uppsala, Sweden). Thiourea, glycerol, SDS, DTT, and APS were from Sigma-Aldrich (St. Louis, MO, USA). Glycine was from USB (Cleveland, OH, USA). Acrylamide Bis 40% was from Bio-Rad (Hercules, CA, USA). All other chemicals and biochemicals used were of analytical grade. All solutions were made with Milli-Q water. Animals Male hamsters (Golden Syrian), 4 weeks of age, were purchased from Harlan Sprague Dawley, USA. Upon arrival, hamsters were acclimated in the University of Arizona animal care facility for at least 1 week and maintained in an environmentally controlled animal facility operating on a 12-h dark/12-h light cycle and at 22-24°C. They were provided with Teklad (Indianapolis, IN) 4% Mouse/Rat Diet # 7001 and water, ad libitum, throughout the acclimation and experimentation periods. Sample preparation and labelling Hamsters were exposed to sodium arsenite (173 mg) in drinking water for 6 days and the control hamsters were given tap water. On the 6th day hamsters were decapitated rapidly by guillotine. Urinary bladder tissues and liver were removed, blotted on tissue papers (Kimtech Science, Precision Wipes), and weighed. Hamster urinary bladder or liver tissues were homogenized in lysis buffer (30mMTris, 2M thiourea, 7M urea, and 4% w/w CHAPS adjusted to pH 8.5 with dilute HCI), at 4°C using a glass homogenizer and a Teflon coated steel pestle; transferred to a 5 ml acid-washed polypropylene tube, placed on ice and sonicated 3 times for 15 seconds. The sonicate was centrifuged at 12,000 rpm for 10 minutes at 4°C. Small aliquots of the supernatants were stored at -80°C until use (generally within one week). Protein concentration was determined by the method of Bradford [25] using bovine serum albumin as a standard. Fifty micrograms of lysate protein was labeled with 400 pmol of Cy3 Dye (for control homogenate sample) and Cy5 Dye (for arsenic-treated urinary bladder or liver homogenate sample). The samples containing proteins and dyes were incubated for 30 min on ice in the dark. To stop the labeling reaction, 1uL of 10 mM lysine was added followed by incubation for 10 min on ice in the dark. To each of the appropriate dye-labeled protein samples, an additional 200 ug of urinary bladderor liver unlabeled protein from control hamster sample or arsenic-treated hamster sample was added to the appropriate sample. Differentially labeled samples were combined into a single Microfuge tube (total protein 500 ug); protein was mixed with an equal volume of 2x sample buffer [2M thiourea, 7M urea, pH 3-10 pharmalyte for isoelectric focusing 2% (v/v), DTT 2% (w/v), CHAPS 4% (w/v)]; and was incubated on ice in the dark for 10 min. The combined samples containing 500 ug of total protein were mixed with rehydration buffer [CHAPS 4% (w/v), 8M urea, 13mM DTT, IPG buffer (3-10) 1% (v/v) and trace amount of bromophenol blue]. The 450 ul sample containing rehydration buffer was slowly pipetted into the slot of the ImmobilinedryStripReswelling Tray and any large bubbles were removed. The IPG strip (linear pH 3-10, 24 cm) was placed (gel side down) into the slot, covered with drystrip cover fluid (Fig. 1), and the lid of the Reswelling Tray was closed. The ImmobillineDryStrip was allowed to rehydrate at room temperature for 24 hours. First dimension Isoelectric focusing (IEF) The labeled sample was loaded using the cup loading method on universal strip holder. IEF was then carried out on EttanIPGphor II using multistep protocol (6 hr @ 500 V, 6 hr @ 1000 V, 8 hr @ 8000 V). The focused IPG strip was equilibrated in two steps (reduction and alkylation) by equilibrating the strip for 10 min first in 10 ml of 50mM Tris (pH 8.8), 6M urea, 30% (v/v) glycerol, 2% (w/v) SDS, and 0.5% (w/v) DTT, followed by another 10 min in 10 ml of 50mM Tris (pH 8.8), 6M urea, 30% (v/v) glycerol, 2% (w/v) SDS, and 4.5% (w/v) iodoacetamide to prepare it for the second dimension electrophoresis. Second dimension SDS-PAGE The equilibrated IPG strip was used for protein separation by 2D-gel electrophoresis (DIGE). The strip was sealed at the top of the acrylamide gel for the second dimension (vertical) (12.5% polyacrylamide gel, 20x25 cm x 1.5 mm) with 0.5% (w/v) agarose in SDS running buffer [25 mMTris, 192 mM Glycine, and 0.1% (w/v) SDS]. Electrophoresis was performed in an Ettan DALT six electrophoresis unit (Amersham Biosciences) at 1.5 watts per gel, until the tracking dye reached the anodic end of the gel. Image analysis and post-staining The gel then was imaged directly between glass plates on the Typhoon 9410 variable mode imager (Sunnyvale, CA, USA) using optimal excitation/emission wavelength for each DIGE fluor: Cy3 (532/580 nm) and Cy5 (633/670 nm). The DIGE images were previewed and checked with Image Quant software (GE Healthcare) where all the two separate gel images could be viewed as a single gel image. DeCyde v.5.02 was used to analyze the DIGE images as described in the Ettan DIGE User Manual (GE Healthcare). The appropriate up-/down regulated spots were filtered based on an average volume ratio of ± over 1.2 fold. After image acquisition, the gel was fixed overnight in a solution containing 40% ethanol and 10% acetic acid. The fixed gel was stained with SyproRuby (BioRad) according to the manufacturer protocol (Bio-Rad Labs., 2000 Alfred Nobel Drive, Hercules, CA 94547). Identification of proteins by MS Protein spot picking and digestion Sypro Ruby stained gels were imaged using an Investigator ProPic and HT Analyzer software, both from Genomic Solutions (Ann Arbor, MI). Protein spots of interest that matched those imaged using the DIGE Cy3/Cy5 labels were picked robotically, digested using trypsin as described previously [24] and saved for mass spectrometry identification. Liquid chromatography (LC)- MS/MS analysis LC-MS/MS analyses were carried out using a 3D quadrupole ion trap massspectrometer (ThermoFinnigan LCQ DECA XP PLUS; ThermoFinnigan, San Jose, CA) equipped with a Michrom Paradigm MS4 HPLC (MichromBiosources, Auburn, CA) and a nanospray source, or with a linear quadrupole ion trap mass spectrometer (ThermoFinnigan LTQ), also equipped with a Michrom MS4 HPLC and a nanospray source. Peptides were eluted from a 15 cm pulled tip capillary column (100 um I.D. x 360 um O.D.; 3-5 um tip opening) packed with 7 cm Vydac C18 (Vydac, Hesperia, CA) material (5 µm, 300 Å pore size), using a gradient of 0-65% solvent B (98% methanol/2% water/0.5% formic acid/0.01% triflouroacetic acid) over a 60 min period at a flow rate of 350 nL/min. The ESI positive mode spray voltage was set at 1.6 kV, and the capillary temperature was set at 200°C. Dependent data scanning was performed by the Xcalibur v 1.3 software on the LCQ DECA XP+ or v 1.4 on the LTQ [27], with a default charge of 2, an isolation width of 1.5 amu, an activation amplitude of 35%, activation time of 50 msec, and a minimal signal of 10,000 ion counts (100 ion counts on the LTQ). Global dependent data settings were as follows: reject mass width of 1.5 amu, dynamic exclusion enabled, exclusion mass width of 1.5 amu, repeat count of 1, repeat duration of a min, and exclusion duration of 5 min. Scan event series were included one full scan with mass range of 350-2000 Da, followed by 3 dependent MS/MS scans of the most intense ion. Database searching Tandem MS spectra of peptides were analyzed with Turbo SEQUEST, version 3.1 (ThermoFinnigan), a program that allows the correlation of experimental tandem MS data with theoretical spectra generated from known protein sequences. All spectra were searched against the latest version of the non redundant protein database from the National Center for Biotechnology Information (NCBI 2006; at that time, the database contained 3,783,042 entries). Statistical analysis The means and standard error were calculated. The Student's t-test was used to analyze the significance of the difference between the control and arsenite exposed hamsters. P values less than 0.05 were considered significant. The reproducibility was confirmed in separate experiments. Results Analysis of proteins expression After DIGE (Fig. 1), the gel was scanned by a Typhoon Scanner and the relative amount of protein from sample 1 (treated hamster) as compared to sample 2 (control hamster) was determined (Figs. 2, 3). A green spot indicates that the amount of protein from sodium arsenite-treated hamster sample was less than that of the control sample. A red spot indicates that the amount of protein from the sodium arsenite-treated hamster sample was greater than that of the control sample. A yellow spot indicates sodium arsenite-treated hamster and control hamster each had the same amount of that protein. Several protein spots were up-regulated (red) or down-regulated (green) in the urinary bladder samples of hamsters exposed to sodium arsenite (173 mg As/L) for 6 days as compared with the urinary bladder of controls (Fig. 2). In the case of liver, several protein spots were also over-expressed (red) or under-expressed (green) for hamsters exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days (Fig. 3). The urinary bladder samples were collected from the first and second experiments in which hamsters were exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days and the controls were given tap water. The urinary bladder samples from the 1st and 2nd experiments were run 5 times in DIGE gels on different days. The protein expression is shown in Figure 2 and Table 1. The liver samples from the 1st and 2nd experiments were also run 3 times in DIGE gels on different days. The proteins expression were shown in Figure 3 and Table 2. The volume ratio changed of the protein spots in the urinary bladder and liver of hamsters exposed to arsenite were significantly differences than that of the control hamsters (Table 1 and 2). Protein spots identified by LC-MS/MS Bladder The spots of interest were removed from the gel, digested, and their identities were determined by LC-MS/MS (Fig. 2 and Table 1). The spots 1, 2, & 3 from the gel were analyzed and were repeated for the confirmation of the results (experiments; 173 mg As/L). The proteins for the spots 1, 2, and 3 were identified as transgelin, transgelin, and glutathione S-transferase Pi, respectively (Fig. 2). Liver We also identified some of the proteins in the liver samples of hamsters exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days (Fig. 3). The spots 4, 5, & 6 from the gels were analyzed and were repeated for the confirmation of the results. The proteins for the spots 4, 5, and 6 were identified as ornithine aminotransferase, senescence marker protein 30, and fatty acid binding protein, respectively (Fig. 3) Discussion The identification and functional assignment of proteins is helpful for understanding the molecular events involved in disease. Weexposed hamsters to sodium arsenite in drinking water. Controls were given tap water. DIGE coupled with LC-MS/MS was then used to study the proteomic change in arsenite-exposed hamsters. After electrophoresis DeCyder software indicated that several protein spots were down-regulated (green) and several were up-regulated (red). Our overall results as to changes and functions of the proteins we have studied are summarized in Table 3. Bladder In the case of the urinary bladder tissue of hamsters exposed to sodium arsenite (173 mg As/L) in drinking water for 6 days, transgelin was down-regulated and GST-pi was up-regulated. This is the first evidence that transgelin is down-regulated in the bladders of animals exposed to sodium arsenite. Transgelin, which is identical to SM22 or WS3-10, is an actin cross linking/gelling protein found in fibroblasts and smooth muscle [28, 29]. It has been suggested that the loss of transgelin expression may be an important early event in tumor progression and a diagnostic marker for cancer development [30-33]. It may function as a tumor suppressor via inhibition of ARA54 (co-regulator of androgen receptor)-enhanced AR (androgen receptor) function. Loss of transgelin and its suppressor function in prostate cancer might contribute to the progression of prostate cancer [30]. Down-regulation of transgelin occurs in the urinary bladders of rats having bladder outlet obstruction [32]. Ras-dependent and Ras-independent mechanisms can cause the down regulation of transgelin in human breast and colon carcinoma cell lines and patient-derived tumorsamples [33]. Transgelin plays a role in contractility, possibly by affecting the actin content of filaments [34]. In our experiments loss of transgelin expression may be associated or preliminary to bladder cancer due to arsenic exposure. Arsenite is a carcinogen [1]. In our experiments, LC-MS/MS analysis showed that two spots (1 and 2) represent transgelin (Fig. 2 and Table 1). In human colonic neoplasms there is a loss of transgelin expression and the appearance of transgelin isoforms (31). GST-pi protein was up-regulated in the bladders of the hamsters exposed to sodium arsenite. GSTs are a large family of multifunctional enzymes involved in the phase II detoxification of foreign compounds [35]. The most abundant GSTS are the classes alpha, mu, and pi classes [36]. They participate in protection against oxidative stress [37]. GST-omega has arsenic reductase activity [38]. Over-expression of GST-pi has been found in colon cancer tissues [39]. Strong expression of GST-pi also has been found in gastric cancer [40], malignant melanoma [41], lung cancer [42], breast cancer [43] and a range of other human tumors [44]. GST-pi has been up-regulated in transitional cell carcinoma of human urinary bladder [45]. Up-regulation of glutathione – related genes and enzyme activities has been found in cultured human cells by sub lethal concentration of inorganic arsenic [46]. There is evidence that arsenic induces DNA damage via the production of ROS (reactive oxygen species) [47]. GST-pi may be over-expressed in the urinary bladder to protect cells against arsenic-induced oxidative stress. Liver In the livers of hamsters exposed to sodium arsenite, ornithine amino transferase was over-expressed, senescence marker protein 30 was under-expressed, and fatty acid binding protein was under-expressed. Ornithine amino transferase has been found in the mitochondria of many different mammalian tissues, especially liver, kidney, and small intestine [48]. Ornithine amino transferase knockdown inhuman cervical carcinoma and osteosarcoma cells by RNA interference blocks cell division and causes cell death [49]. It has been suggested that ornithine amino transferase has a role in regulating mitotic cell division and it is required for proper spindle assembly in human cancer cells [49]. Senescence marker protein-30 (SMP30) is a unique enzyme that hydrolyzes diisopropylphosphorofluoridate. SMP30, which is expressed mostly in the liver, protects cells against various injuries by stimulating membrane calcium-pump activity [50]. SMP30 acts to protect cells from apoptosis [51]. In addition it protects the liver from toxic agents [52]. The livers of SMP30 knockout mice accumulate phosphatidylethanolamine, cardiolipin, phosphatidyl-choline, phosphatidylserine, and sphingomyelin [53]. Liver fatty acid binding protein (L-FABP) also was down- regulated. Decreased liver fatty acid-binding capacity and altered liver lipid distribution hasbeen reported in mice lacking the L-FABP gene [54]. High levels of saturated, branched-chain fatty acids are deleterious to cells and animals, resulting in lipid accumulation and cytotoxicity. The expression of fatty acid binding proteins (including L-FABP) protected cells against branched-chain saturated fatty acid toxicity [55]. Limitations: we preferred to study the pronounced spots seen in DIGE gels. Other spots were visible but not as pronounced. Because of limited funds, we did not identify these others protein spots. In conclusion, urinary bladders of hamsters exposed to sodium arsenite had a decrease in the expression of transgelin and an increase in the expression of GST-pi protein. Under-expression of transgelin has been found in various cancer systems and may be associated with arsenic carcinogenicity [30-33). Inorganic arsenic exposure has resulted in bladder cancer as has been reported in the past [1]. Over-expression of GST-pi may protect cells against oxidative stress caused by arsenite. In the liver OAT was up regulated and SMP-30 and FABP were down regulated. These proteomic results may be of help to investigators studying arsenic carcinogenicity. The Superfund Basic Research Program NIEHS Grant Number ES 04940 from the National Institute of Environmental Health Sciences supported this work. Additional support for the mass spectrometry analyses was provided by grants from NIWHS ES06694, NCI CA023074 and the BIOS Institute of the University of Arizona. Acknowledgement The Author wants to dedicate this paper to the memory of his former supervisor Dr. H. VaskenAposhian who passed away in September 6, 2019. He was an emeritus professor of the Department of Molecular and Cellular Biology at the University of Arizona. 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Lipopolysaccharide (LPS) is a gram‐negative bacterial marker that might be released into circulation during exercise‐heat stress. LPS induces an innate immune response by binding LPS‐binding protein (LBP) and its cognate receptor, TLR4 (toll‐like receptor 4), expressed on CD16+CD14+ circulating immune cells. LPS‐induced TLR4 activation results in robust immune responses which drive endotoxemia, sepsis, and putatively, exertional heatstroke pathophysiology.PURPOSEWe aimed to test the hypothesis that competing in prolonged cycling (50–100 mile event) or high‐intensity running (7 mile race) events in hot and humid environments (heat index>80°F) result in LPS‐induced TLR4 activation and changes in cellular gene expression.METHODSWe recruited 64 subjects participating in the Hotter n' Hell Hundred (HHH) ultraendurance cycling event (Wichita Falls, TX) and 36 subjects racing in the Falmouth (FAL) Road Race (Falmouth, MA). We sampled venous blood pre (PRE) and immediately post (IP) each event, and also 30 minutes after IP (30IP) in FAL. We isolated circulating PBMCs (peripheral blood mononuclear cells) which were stained for surface markers CD14, CD16, and TLR4 using fluor‐conjugated antibodies. We used multi‐color digital flow cytometry on‐site to analyze cell populations. We also isolated anticoagulant‐treated plasma samples from whole blood for analysis of circulating LPS (Hycult Biotech, ELISA) and LBP (R&D ELISA) concentrations. At both events, we measured environmental and physiological variables. We statistically analyzed data using repeated measures ANOVA (LSD post hoc, α‐level:p≤0.05) for differences (PRE vs. IP, PRE vs. 30IP).RESULTSCirculating LPS increased in both HHH riders and FAL racers by 25% (p<0.05) and 39% (p<0.04), respectively. By 30IP, FAL racers' circulating LPS levels had returned to PRE values (p>0.19). In both races, circulating LBP remained unchanged (p>0.33). CD14+CD16+ cells expressing TLR4 in circulation were significantly increased at IP (p<0.05).CONCLUSIONWe conclude that circulating LPS is increased post‐exercise heat stress and analysis of downstream activation is required to determine role of LPS in exercise‐heat stress.Support or Funding InformationWe would like to thank Connecticut Institute for Clinical and Translational Science and McNair Scholars Program for funding the research and thank Hotter'N Hell Hundred and Falmouth Road Race for their support.