The importance of antioxidant status in gastric intestinal metaplasia

BackgroundOxidative stress status in different cancer types was investigated before, but not studied in gastric intestinal metaplasia to the best of our knowledge. Purpose of this study is to examine whether there is a difference between oxidative stress status in patients with intestinal metaplasia (IM) compared to individuals without IM, we compared the serum levels of disulfide (SS), total thiol (TT) and native thiol (NT).MethodsThis was a prospective, non-randomized casecontrol study including 67 patients with histopathologically confirmed IM and 60 individuals demographically matched in terms of age, gender, BMI, smoking status, and chronic diseases as control group.ResultsThe mean NT, TT and NT to TT (NT/TT) ratios were statistically significantly higher in IM group compared to controls ((351.71 ± 81.9 mol/L vs. 271.82 ± 54.13 mol/L, p=0.000), (391.5 ± 92.69 mol/L vs. 308.59 ± 55.53 mol/L, p=0.000) and (0.89 ± 0.6 vs. 0.87 ± 0.29, p=0.022), respectively). The mean SS to TT (SS/TT) ratio was significantly lower in IM group than control group (0.050 ± 0.31 vs. 0.060 ± 0.014, P=0.022). Median SS and mean SS/NT ratio was similar in both groups (16.3 (3.3-78) vs. 18.3 (10-32.7), p=0.271 and 0.055 ± 0.041 vs. 0.070 ± 0.019, p=0.068, respectively). In ROC analysis, cut off value of SS/NT for IM was found 0.062, in regression analysis, SS/NT <0.062 was found as an independently prognostic marker for IM (OR, 2.38; 95%CI: 1.168-4.865, P=0.017).ConclusionsSS/NT ratio lower than 0.062 was found as an independently prognostic marker for IM. This ratio could help to distinguish which patients should be followed closely for gastric cancer.     

Variation in haematological parameters in children less than five years of age with asymptomatic Plasmodium infection: implication for malaria field studies

During the season of high malaria transmission, most children are infected by Plasmodium, which targets red blood cells (RBCs), affecting haematological parameters. To describe these variations, we examined the haematological profiles of two groups of children living in a malaria-endemic area. A cross-sectional survey was conducted at the peak of the malaria transmission season in a rural area of Burkina Faso. After informed consent and clinical examination, blood samples were obtained from the participants for malaria diagnosis and a full blood count. Of the 414 children included in the analysis, 192 were not infected with Plasmodium, whereas 222 were asymptomatic carriers of Plasmodium infection. The mean age of the infected children was 41.8 months (range of 26.4-57.2) compared to 38.8 months (range of 22.4-55.2) for the control group (p = 0.06). The asymptomatic infected children tended to have a significantly lower mean haemoglobin level (10.8 g/dL vs. 10.4 g/dL; p < 0.001), mean lymphocyte count (4592/µL vs. 5141/µL; p = 0.004), mean platelet count (266 x 103/µL vs. 385 x 103/µL; p < 0.001) and mean RBC count (4.388 x 106/µL vs. 4.158 x 106/µL; p < 0.001) and a higher mean monocyte count (1403/µL vs. 1192/µL; p < 0.001) compared to the control group. Special attention should be applied when interpreting haematological parameters and evaluating immune responses in asymptomatic infected children living in malaria-endemic areas and enrolled in vaccine trials.     

Sensitivity of Plasmodium falciparum to Antimalarial Drugs in Hainan Island,China

Pyronaridine and artesunate have been shown to be effective in falciparum malaria treatment. However, pyronaridine is rarely used in Hainan Island clinically, and artesunate is not widely used as a therapeutic agent. Instead, conventional antimalarial drugs, chloroquine and piperaquine, are used, explaining the emergence of chloroquine-resistant Plasmodium falciparum. In this article, we investigated the sensitivity of P. falciparum to antimalarial drugs used in Hainan Island for rational drug therapy. We performed in vivo (28 days) and in vitro tests to determine the sensitivity of P. falciparum to antimalarial drugs. Total 46 patients with falciparum malaria were treated with dihydroartemisinin/piperaquine phosphate (DUO-COTECXIN) and followed up for 28 day. The cure rate was 97.8%. The mean fever clearance time (22.5±10.6 hr) and the mean parasite clearance time (27.3±12.2 hr) showed no statistical significance with different genders, ages, temperatures, or parasite density (P>0.05). The resistance rates of chloroquine, piperaquine, pyronarididine, and artesunate detected in vitro were 71.9%, 40.6%, 12.5%, and 0%, respectively (P<0.0001). The resistance intensities decreased as follows: chloroquine>piperaquine>pyronarididine>artesunate. The inhibitory dose 50 (IC₅₀) was 3.77×10^-6 mol/L, 2.09×10^-6 mol/L, 0.09×10^-6 mol/L, and 0.05×10^-6 mol/L, and the mean concentrations for complete inhibition (CIMC) of schizont formation were 5.60×10^-6 mol/L, 9.26×10^-6 mol/L, 0.55×10^-6 mol/L, and 0.07×10^-6 mol/L, respectively. Dihydroartemisinin showed a strong therapeutic effect against falciparum malaria with a low toxicity.     

Elevated serum squalene and cholesterol synthesis markers in pregnant obese women with gestational diabetes mellitus

We examined serum cholesterol synthesis and absorption markers and their association with neonatal birth weight in obese pregnancies affected by gestational diabetes mellitus (GDM). Pregnant women at risk for GDM (BMI >30 kg/m²) were enrolled from maternity clinics in Finland. GDM was determined from the results of an oral glucose tolerance test. Serum samples were collected at six time-points, one in each trimester of pregnancy, and at 6 weeks, 6 months, and 12 months postpartum. Analysis of serum squalene and noncholesterol sterols by gas-liquid chromatography revealed that in subjects with GDM (n = 22), the serum Δ8-cholestenol concentration and lathosterol/sitosterol ratio were higher (P < 0.05) than in the controls (n = 30) in the first trimester, reflecting increased cholesterol synthesis. Also, subjects with GDM had an increased ratio of squalene to cholesterol (100 × μmol/mmol of cholesterol) in the second (11.5 ± 0.5 vs. 9.1 ± 0.5, P < 0.01) and third (12.1 ± 0.8 vs. 10.0 ± 0.7, P < 0.05) trimester. In GDM, the second trimester maternal serum squalene concentration correlated with neonatal birth weight (r = 0.70, P < 0.001). In conclusion, in obesity, GDM associated with elevated serum markers of cholesterol synthesis. Correlation of maternal serum squalene with neonatal birth weight suggests a potential contribution of maternal cholesterol synthesis to newborn weight in GDM.     

Electronegative Low-Density Lipoprotein Increases C-Reactive Protein Expression in Vascular Endothelial Cells through the LOX-1 Receptor

Objectives

Increased plasma C-reactive protein (CRP) levels are associated with the occurrence and severity of acute coronary syndrome. We investigated whether CRP can be generated in vascular endothelial cells (ECs) after exposure to the most electronegative subfraction of low-density lipoprotein (LDL), L5, which is atherogenic to ECs. Because L5 and CRP are both ligands for the lectin-like oxidized LDL receptor-1 (LOX-1), we also examined the role of LOX-1.

Methods and Results

Plasma LDL samples isolated from asymptomatic hypercholesterolemic (LDL cholesterol [LDL-C] levels, 154.6±20 mg/dL; n = 7) patients and normocholesterolemic (LDL-C levels, 86.1±21 mg/dL; P<0.001; n = 7) control individuals were chromatographically resolved into 5 subfractions, L1-L5. The L5 percentage (L5%) and the plasma L5 concentration ([L5]  =  L5% × LDL-C) in the patient and control groups were 8.1±2% vs. 2.3±1% (P<0.001) and 12.6±4 mg/dL vs. 1.9±1 mg/dL (P<0.001), respectively. In hypercholesterolemic patients treated with atorvastatin for 6 months (10 mg/day), [L5] decreased from 12.6±4 mg/dL to 4.5±1.1 mg/dL (P = 0.011; n = 5), whereas both [L5] and L5% returned to baseline levels in 2 noncompliant patients 3 months after discontinuation. In cultured human aortic ECs (HAECs), L5 upregulated CRP expression in a dose- and time-dependent manner up to 2.5-fold (P<0.01), whereas the least electronegative subfraction, L1, had no effect. DiI-labeled L1, internalized through the LDL receptor, became visible inside HAECs within 30 seconds. In contrast, DiI-labeled L5, internalized through LOX-1, became apparent after 5 minutes. L5-induced CRP expression manifested at 30 minutes and was attenuated by neutralizing LOX-1. After 30 minutes, L5 but not L1 induced reactive oxygen species (ROS) production. Both L5-induced ROS and CRP production were attenuated by ROS inhibitor N-acetyl cysteine.

Conclusions

Our results suggest that CRP, L5, and LOX-1 form a cyclic mechanism in atherogenesis and that reducing plasma L5 levels with atorvastatin disrupts the vascular toxicity of L5.     

NS1643 Interacts around L529 of hERG to Alter Voltage Sensor Movement on the Path to Activation

Activators of hERG1 such as NS1643 are being developed for congenital/acquired long QT syndrome. Previous studies identify the neighborhood of L529 around the voltage-sensor as a putative interacting site for NS1643. With NS1643, the V_1/2 of activation of L529I (−34 ± 4 mV) is similar to wild-type (WT) (−37 ± 3 mV; P > 0.05). WT and L529I showed no difference in the slope factor in the absence of NS1643 (8 ± 0 vs. 9 ± 0) but showed a difference in the presence of NS1643 (9 ± 0.3 vs. 22 ± 1; P < 0.01). Voltage-clamp-fluorimetry studies also indicated that in L529I, NS1643 reduces the voltage-sensitivity of S4 movement. To further assess mechanism of NS1643 action, mutations were made in this neighborhood. NS1643 shifts the V_1/2 of activation of both K525C and K525C/L529I to hyperpolarized potentials (−131 ± 4 mV for K525C and −120 ± 21 mV for K525C/L529I). Both K525C and K525C/K529I had similar slope factors in the absence of NS1643 (18 ± 2 vs. 34 ± 5, respectively) but with NS1643, the slope factor of K525C/L529I increased from 34 ± 5 to 71 ± 10 (P < 0.01) whereas for K525C the slope factor did not change (18 ± 2 at baseline and 16 ± 2 for NS1643). At baseline, K525R had a slope factor similar to WT (9 vs. 8) but in the presence of NS1643, the slope factor of K525R was increased to 24 ± 4 vs. 9 ± 0 mV for WT (P < 0.01). Molecular modeling indicates that L529I induces a kink in the S4 voltage-sensor helix, altering a salt-bridge involving K525. Moreover, docking studies indicate that NS1643 binds to the kinked structure induced by the mutation with a higher affinity. Combining biophysical, computational, and electrophysiological evidence, a mechanistic principle governing the action of some activators of hERG1 channels is proposed.     

Thiol/disulphide homeostasis in pregnant women with Familial Mediterranean fever

Objective: To investigate the presence of oxidative stress (OS) in pregnant women with Familial Mediterranean fever (FMF) in the first trimester by evaluating thiol/disulphide homeostasis.

Study design: A total of 31 pregnant women with a diagnosis of FMF, between 11⁰ and 13⁶ weeks of gestation, were compared with 51 healthy pregnant controls at the same gestational weeks. A recently defined method was used to measure plasma native thiol, total thiol and disulphide levels.

Results: There were no differences between groups in terms of maternal age, body mass index and numbers of gravida and parity. Antenatal complications (45.2% vs. 9.8%, P?=?0.001) and primary caesarean section (22.6% vs. 5.9%, P?=?0.037) were higher in the FMF group. Pregnant women with FMF had significantly lower first trimester serum levels of native thiol (297.5?μmol/l (153.2–441.8) vs. 366.1?μmol/l (288.7–432.4), P?=?0.000), total thiol (327.2?μmol/l (171.0–471.0) vs. 389.9?μmol/l (317.1–449.8), P?=?0.000) and higher levels of disulphide (14.2?±?4.5?μmol/l vs. 12.4?±?3.4?μmol/l, P?=?0.045). No differences were found in these parameters among FMF patients with and without antenatal complications.

Conclusions: The main outcome demonstrates a relation between OS and pregnant women with FMF in the first trimester of gestation. OS in the first trimester may be a major aetiological factor of unfavourable pregancy outcomes in this group of patients.     

Dynamic thiol/disulphide homeostasis as indicator of oxidative stress in automotive workers

Abstract

Purpose: To examine thiol-disulphide homeostasis auto painters.

Materials and methods: A total of 115 male workers, including 60 auto painters workers and 55 reference group, of the painting and assembly line units respectively, were included in the study. Thiol-disulphide parameters and ischaemia-modified albumin (IMA) of groups were determined. Urinary hippuric acid, (HA) phenol, hexanedione, trichloroacetic acid, arsenic and blood lead and manganese were analysed.

Results: The median urinary HA level was significantly higher in auto painters when compared to the reference group [(2461 (1212) vs. 520 (513) µgr/L), (p?<?0.001)] . The mean disulphide level [19.7 (4.3) vs 0.15.1(4.1) μmol/L, (p?<?0.001)], the disulphide/native thiol ratio [4.72 (1.47) vs. 3.13 (1.21, (p?<?0.001)] and the disulphide/total thiol ratio [4.31 (1.23) vs. 2.94 (1.06), (p?<?0.001)] were higher in auto painters when compared to the reference group. There was a statistically significant positive correlation between urinary HA and disulphide concentrations (r?=?0.536 and p?<?0.001), disulphide/native thiol ratio (r?=?0.564 and p?<?0.001) and the disulphide/total thiol ratio (r?=?0.564 and p?<?0.001) and IMA (r?=?0.396 and p?<?0.001).

Conclusion: The results presented in this study showed that oxidative stress can be associated with occupational exposure to toluene denoted by alteration of thiol disulphide homeostasis and ischaemia-modified albumin levels.     

Healthy birth weight results in higher vitamin A storage in neonate piglets administered high-dose supplements

A proposed intervention for newborn infants in countries with suspected vitamin A (VA) deficiency is to administer 50,000 IU retinyl palmitate at birth to reduce mortality risk. However, no studies have investigated birth weight effects. In this study, low birth weight (LBW; <1 kg, n = 18) and healthy birth weight (HBW) piglets (>1.5 kg, n = 18) from VA-depleted sows were dosed with 25,000 or 50,000 IU retinyl palmitate (26.2 or 52.4 µmol retinol equivalents) at birth to compare VA reserves. Blood was collected at varying times (n = 3–5/time/dose), and piglets were killed at 12 or 24 h for blood, liver, kidneys, spleen, lungs, adrenal gland, and intestinal contents. HBW piglets had significantly higher birth, death, and organ weights than LBW (P < 0.0001 for all). HBW and LBW piglets, which received VA, had higher liver and kidney VA concentrations (0.18 ± 0.09, 0.24 ± 0.10 µmol/g liver and 13.4 ± 4.1, 14.2 ± 4.5 nmol/g kidney, respectively) than controls (n = 10) (0.051 ± 0.01 µmol/g liver and 1.01 ± 0.43 nmol/g kidney) (P = 0.0061 and < 0.0001, respectively). Total liver (9.75 ± 5.16 µmol) and kidney retinol (204 ± 79.1 nmol) were higher in HBW than LBW piglets (P < 0.0001). Extrahepatic tissues, except lung, had higher VA concentration than controls (P < 0.0001). Serum retinol and ester concentrations were higher in treated than control piglets (P = 0.0028, P < 0.0001, respectively), and significantly changed during the times sampled (P = 0.022, P = 0.011, respectively). Peak serum retinyl ester concentrations, which occurred at 3 h, were higher in piglets that received 50,000 IU (4.2 ± 4.4 µmol/L) than 25,000 IU (2.7 ± 2.3 µmol/L) (P = 0.031). Regardless of dose amount, HBW piglets stored more supplemental VA than LBW piglets when administered at birth.     

A Pro-Inflammatory Role of C5L2 in C5a-Primed Neutrophils for ANCA-Induced Activation

Background

The complement system is crucial for the development of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). In particular, C5a and its receptor on neutrophils, CD88, play a central role. The functional role of the second receptor of C5a, C5L2, remains unclear. In the current study, we investigated the role of C5L2 in C5a-primed neutrophils for ANCA-induced activation.

Methods

The effect of blocking C5L2 by anti-human C5L2 blocking antibody were tested on respiratory burst and degranulation of C5a-primed neutrophils activated with ANCA, as well as on membrane-bound proteinase 3 (mPR3) and concentration of myeloperoxidase (MPO) in supernatant of C5a-primed neutrophils. An antagonist for CD88 was also employed.

Results

Blocking C5L2 resulted in a significantly decreased MPO concentration in the supernatant of C5a-primed neutrophils. mPR3 expression increased from 209.0±43.0 in untreated cells to 444.3±60.8 after C5a treatment (P<0.001), and decreased to 375.8±65.44, 342.2±54.3 and 313.7±43.6 by pre-incubating blocking C5L2 antibody at 2.5 µg/ml, 5 µg/ml or 10 µg/ml (compared with C5a-priming group, P<0.001, P<0.001, and P<0.001), respectively. In C5a-primed neutrophils, subsequently activating with MPO-ANCA-positive IgG, the MFI value was 425.8±160.6, which decreased to 292.8±141.2, 289.7±130.0 and 280.3±136.4 upon pre-incubation with mouse anti-human C5L2 blocking antibody at 2.5 µg/ml, 5 µg/ml or 10 µg/ml (compared with C5a-primed neutrophils, for MPO-ANCA-positive IgG-induced activation, P<0.05, P<0.05, and P<0.05), respectively. Blocking C5L2 also resulted in significantly decreased C5a-primed neutrophils for PR3-ANCA-positive IgG-induced activation. Moreover, the lactoferrin concentration in the supernant significantly decreased in pre-incubation with anti-human C5L2 blocking antibody, compared with C5a-primed neutrophils induced by PR3- or MPO-ANCA-positive IgG.

Conclusions

C5L2 may be implicated in the pro-inflammatory role in C5a-primed neutrophils for ANCA-induced activation.     

Superiority of zinc complex of acetylsalicylic acid to acetylsalicylic acid in preventing postischemic myocardial dysfunction

The pathophysiology of ischemic myocardial injury involves cellular events, reactive oxygen species, and an inflammatory reaction cascade. The zinc complex of acetylsalicylic acid (Zn(ASA)₂) has been found to possess higher anti-inflammatory and lower ulcerogenic activities than acetylsalicylic acid (ASA). Herein, we studied the effects of both ASA and Zn(ASA)₂ against acute myocardial ischemia. Rats were pretreated with ASA (75 mg/kg) or Zn(ASA)₂ (100 mg/kg) orally for five consecutive days. Isoproterenol (85 mg/kg, subcutaneously [s.c.]) was applied to produce myocardial infarction. After 17–22 h, animals were anesthetized with sodium pentobarbital (60 mg/kg, intraperitoneally [i.p.]) and both electrical and mechanical parameters of cardiac function were evaluated in vivo. Myocardial histological and gene expression analyses were performed. In isoproterenol-treated rats, Zn(ASA)₂ treatment normalized significantly impaired left-ventricular contractility index (E_max 2.6 ± 0.7 mmHg/µL vs. 4.6 ± 0.5 mmHg/µL, P < 0.05), increased stroke volume (30 ± 3 µL vs. 50 ± 6 µL, P < 0.05), decreased systemic vascular resistance (7.2 ± 0.7 mmHg/min/mL vs. 4.2 ± 0.5 mmHg/min/mL, P < 0.05) and reduced inflammatory infiltrate into the myocardial tissues. ECG revealed a restoration of elevated ST-segment (0.21 ± 0.03 mV vs. 0.09 ± 0.02 mV, P < 0.05) and prolonged QT-interval (79.2 ± 3.2 ms vs. 69.5 ± 2.5 ms, P < 0.05) by Zn(ASA)₂. ASA treatment did not result in an improvement of these parameters. Additionally, Zn(ASA)₂ significantly increased the mRNA-expression of superoxide dismutase 1 (+73 ± 15%), glutathione peroxidase 4 (+44 ± 12%), and transforming growth factor (TGF)-β₁ (+102 ± 22%). In conclusion, our data demonstrate that oral administration of zinc and ASA in the form of bis(aspirinato)zinc(II) complex is superior to ASA in preventing electrical, mechanical, and histological changes after acute myocardial ischemia. The induction of antioxidant enzymes and the anti-inflammatory cytokine TGF-β₁ may play a pivotal role in the mechanism of action of Zn(ASA)₂.     

The Intestinal Barrier in Irritable Bowel Syndrome: Subtype-Specific Effects of the Systemic Compartment in an In Vitro Model

BackgroundIrritable bowel syndrome (IBS) is a disorder with multifactorial pathophysiology. Intestinal barrier may be altered, especially in diarrhea-predominant IBS (IBS-D). Several mediators may contribute to increased intestinal permeability in IBS.AimWe aimed to assess effects of tryptase and LPS on in vitro permeability using a 3-dimensional cell model after basolateral cell exposure. Furthermore, we assessed the extent to which these mediators in IBS plasma play a role in intestinal barrier function.ResultsTryptase (20 and 50 mU) and LPS (6.25 – 50 ng/mL) significantly increased Caco-2 permeability versus control (all P< 0.05). Plasma of IBS-D only showed significantly elevated median tryptase concentrations (7.1 [3.9 – 11.0] vs. 4.2 [2.2 – 7.0] vs. 4.2 [2.5 – 5.9] μg/mL; P<0.05) and LPS concentrations (3.65 [3.00 – 6.10] vs. 3.10 [2.60-3.80] vs. 2.65 [2.40 – 3.40] EU/ml; P< 0.05) vs. IBS-C and HC. Also, plasma of IBS-D increased Caco-2 permeability versus HC (0.14450 ± 0.00472 vs. 0.00021 ± 0.00003; P < 0.001), which was attenuated by selective inhibition of tryptase and LPS (P< 0.05).ConclusionBasolateral exposure of spheroids to plasma of IBS-D patients resulted in a significantly increased FD4 permeation, which was partially abolished by selective inhibition of tryptase and LPS. These findings point to a role of systemic tryptase and LPS in the epithelial barrier alterations observed in patients with IBS-D.     

Increased levels of circulating microparticles in primary Sj?gren's syndrome,systemic lupus erythematosus and rheumatoid arthritis and relation with disease activity

Introduction

Cell stimulation leads to the shedding of phosphatidylserine (PS)-rich microparticles (MPs). Because autoimmune diseases (AIDs) are characterized by cell activation, we investigated level of circulating MPs as a possible biomarker in primary Sjögren''s syndrome (pSS), systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA).

Methods

We measured plasma levels of total, platelet and leukocyte MPs by prothrombinase capture assay and flow cytometry in 43 patients with pSS, 20 with SLE and 24 with RA and in 44 healthy controls (HCs). Secretory phospholipase A2 (sPLA2) activity was assessed by fluorometry. Soluble CD40 ligand (sCD40L) and soluble P-selectin (sCD62P), reflecting platelet activation, were measured by ELISA.

Results

Patients with pSS showed increased plasma level of total MPs (mean ± SEM 8.49 ± 1.14 nM PS equivalent (Eq), P < 0.0001), as did patients with RA (7.23 ± 1.05 n PS Eq, P = 0.004) and SLE (7.3 ± 1.25 nM PS Eq, P = 0.0004), as compared with HCs (4.13 ± 0.2 nM PS Eq). Patients with AIDs all showed increased level of platelet MPs (P < 0.0001), but only those with pSS showed increased level of leukocyte MPs (P < 0.0001). Results by capture assay and flow cytometry were correlated. In patients with high disease activity according to extra-glandular complications (pSS), DAS28 (RA) or SLEDAI (SLE) compared with low-activity patients, the MP level was only slightly increased in comparison with those having a low disease activity. Platelet MP level was inversely correlated with anti-DNA antibody level in SLE (r = -0.65; P = 0.003) and serum β2 microglobulin level in pSS (r = -0.37; P < 0.03). The levels of total and platelet MPs were inversely correlated with sPLA2 activity (r = -0.37, P = 0.0007; r = -0.36, P = 0.002, respectively). sCD40L and sCD62P concentrations were significantly higher in pSS than in HC (P ≤ 0.006).

Conclusions

Plasma MP level is elevated in pSS, as well as in SLE and RA, and could be used as a biomarker reflecting systemic cell activation. Level of leukocyte-derived MPs is increased in pSS only. The MP level is low in case of more severe AID, probably because of high secretory phospholipase A2 (sPLA2) activity, which leads to consumption of MPs. Increase of platelet-derived MPs, sCD40L and sCD62P, highlights platelet activation in pSS.     

Probiotics Prevent Intestinal Barrier Dysfunction in Acute Pancreatitis in Rats via Induction of Ileal Mucosal Glutathione Biosynthesis

Background

During acute pancreatitis (AP), oxidative stress contributes to intestinal barrier failure. We studied actions of multispecies probiotics on barrier dysfunction and oxidative stress in experimental AP.

Methodology/Principal Findings

Fifty-three male Spraque-Dawley rats were randomly allocated into five groups: 1) controls, non-operated, 2) sham-operated, 3) AP, 4) AP and probiotics and 5) AP and placebo. AP was induced by intraductal glycodeoxycholate infusion and intravenous cerulein (6 h). Daily probiotics or placebo were administered intragastrically, starting five days prior to AP. After cerulein infusion, ileal mucosa was collected for measurements of E. coli K12 and ⁵¹Cr-EDTA passage in Ussing chambers. Tight junction proteins were investigated by confocal immunofluorescence imaging. Ileal mucosal apoptosis, lipid peroxidation, and glutathione levels were determined and glutamate-cysteine-ligase activity and expression were quantified. AP-induced barrier dysfunction was characterized by epithelial cell apoptosis and alterations of tight junction proteins (i.e. disruption of occludin and claudin-1 and up-regulation of claudin-2) and correlated with lipid peroxidation (r>0.8). Probiotic pre-treatment diminished the AP-induced increase in E. coli passage (probiotics 57.4±33.5 vs. placebo 223.7±93.7 a.u.; P<0.001), ⁵¹Cr-EDTA flux (16.7±10.1 vs. 32.1±10.0 cm/s10⁻⁶; P<0.005), apoptosis, lipid peroxidation (0.42±0.13 vs. 1.62±0.53 pmol MDA/mg protein; P<0.001), and prevented tight junction protein disruption. AP-induced decline in glutathione was not only prevented (14.33±1.47 vs. 8.82±1.30 nmol/mg protein, P<0.001), but probiotics even increased mucosal glutathione compared with sham rats (14.33±1.47 vs. 10.70±1.74 nmol/mg protein, P<0.001). Glutamate-cysteine-ligase activity, which is rate-limiting in glutathione biosynthesis, was enhanced in probiotic pre-treated animals (probiotics 2.88±1.21 vs. placebo 1.94±0.55 nmol/min/mg protein; P<0.05) coinciding with an increase in mRNA expression of glutamate-cysteine-ligase catalytic (GCLc) and modifier (GCLm) subunits.

Conclusions

Probiotic pre-treatment diminished AP-induced intestinal barrier dysfunction and prevented oxidative stress via mechanisms mainly involving mucosal glutathione biosynthesis.     

Regulation of α-Transducin and α-Gustducin Expression by a High Protein Diet in the Pig Gastrointestinal Tract

BackgroundThe expression of taste receptors (TASRs) and their signalling molecules in the gastrointestinal (GI) epithelial cells, including enteroendocrine cells (EECs), suggests they participate in chemosensing mechanisms influencing GI physiology via the release of endocrine messengers. TASRs mediate gustatory signalling by interacting with different transducers, including α-gustducin (G_αgust) and α-transducin (G_αtran) G protein subunits. This study tested whether G_αtran and G_αgust immunoreactive (-IR) cells are affected by a short-term (3 days) and long-term (30 days) high protein (Hp) diet in the pig GI tract.ResultIn the stomach, G_αgust and G_αtran-IR cells contained serotonin (5-HT) and ghrelin (GHR), while in the small and large intestine, G_αgust and G_αtran-IR colocalized with 5-HT-, cholecystokinin (CCK)- and peptide YY (PYY)-IR. There was a significant increase in the density of G_αtran-IR cells in the pyloric mucosa in both short- and long-term Hp diet groups (Hp3 and Hp30) vs. the control group (Ctr) (P<0.05), while the increase of G_αgust-IR cells in the pyloric mucosa was significant in Hp30 group vs. Ctr and vs. Hp3 (P<0.05); these cells included G_αtran / 5HT-IR and G_αtran / GHR-IR cells (P<0.05 and P<0.001 vs. Ctr, respectively) as well as G_αgust /5-HT-IR or G_αgust / GHR-IR cells (P<0.05 and P<0.01 vs. Ctr, respectively). In the small intestine, we recorded a significant increase in G_αtran-IR cells in the duodenal crypts and a significant increase of G_αgust-IR cells in the jejunal crypts in Hp3 group compared to HP30 (P<0.05). With regard to the number of G_αtran-G_αgust IR cells colocalized with CCK or 5-HT, there was only a significant increase of G_αtran / CCK-IR cells in Hp3 group compared to Ctr (P = 0.01).ConclusionThis study showed an upregulation of selected subpopulations of G_αgust / G_αtran-IR cells in distinct regions of the pig GI tract by short- and long-term Hp diet lending support to TASR-mediated effects in metabolic homeostasis and satiety mechanisms.     

Low tidal volume protects pulmonary vasomotor function from “second-hit” injury in acute lung injury rats

Background

Sepsis could induce indirect acute lung injury(ALI), and pulmonary vasomotor dysfunction. While low tidal volume is advocated for treatment of ALI patients. However, there is no evidence for low tidal volume that it could mitigate pulmonary vasomotor dysfunction in indirect ALI. Our study is to evaluate whether low tidal volume ventilation could protect the pulmonary vascular function in indirect lipopolysaccharide (LPS) induced acute lung injury rats.

Methods

An indirect ALI rat model was induced by intravenous infusion of LPS. Thirty rats (n = 6 in each group) were randomly divided into (1)Control group; (2) ALI group; (3) LV group (tidal volume of 6mL/kg); (4) MV group (tidal volume of 12mL/kg); (5)VLV group (tidal volume of 3mL/kg). Mean arterial pressure and blood gas analysis were monitored every 2 hours throughout the experiment. Lung tissues and pulmonary artery rings were immediately harvested after the rats were bled to be killed to detect the contents of endothelin-1 (ET-1), endothelial nitric oxide synthase (eNOS) and TNF-α. Acetylcholine (Ache)-induced endothelium-dependent and sodium nitroprusside (SNP)-induced endothelium-independent relaxation of isolated pulmonary artery rings were measured by tensiometry.

Results

There was no difference within groups concerning blood pressure, PaCO₂ and SNP-induced endothelium-independent relaxation of pulmonary artery rings. Compared with MV group, LV group significantly reduced LPS-induced expression of ET-1 level (113.79 ± 7.33pg/mL vs. 152.52 ± 12.75pg/mL, P < 0.05) and TNF-α (3305.09 ± 334.29pg/mL vs.4144.07 ± 608.21pg/mL, P < 0.05), increased the expression of eNOS (IOD: 15032.05 ± 5925.07 vs. 11454.32 ± 6035.47, P < 0.05). While Ache (10^-7mol/L-10^-4mol/L)-induced vasodilatation was ameliorated 30% more in LV group than in MV group.

Conclusions

Low tidal volume could protect the pulmonary vasodilative function during indirect ALI by decreasing vasoconstrictor factors, increasing expressions of vasodilator factors in pulmonary endothelial cells, and inhibiting inflammation injuries.     

H-Index Is Important for Postural Control for People with Impaired Foot Sole Sensation

Introduction

People with Peripheral Neuropathy (PN), especially those with impaired sensory inputs through the small-afferent fiber (type II afferent fibers) reflex loop (SAF), might depend more on the large-afferent fiber (type I afferent fibers) reflex loop (LAF) for postural control.

Purpose

To examine whether the function of the LAF reflex loop, reflected by the H-reflex and ankle joint proprioception, influences postural control when the SAF reflex loop is impaired, as indicated by reduced foot sole cutaneous sensation.

Methods

Thirteen participants (8 women, 5 men) diagnosed with PN and 12 age-matched controls (7 women, 5 men) completed the testing protocol. Measures of interest included the H-index, active (AAP) and passive (PAP) ankle proprioception, plantar pressure sensitivity (PPS), average sway velocity (V_AVG) and area (A₉₅) during 30 seconds eyes-closed standing, 6-minute walk distance (6MWD) and timed up-and-go duration (TUG).

Results

Statistically significant group-dependent regression was observed between V_AVG and H-index. Compared to the control group, the PN group demonstrated reduced PPS (2.0 ± 1.9 vs. 4.2 ± 1.2, P < .05) and H-index (63.6 ± 10.9 vs. 76.4 ± 16.0, P < .05), greater V_AVG (3.5 ± 2.1 vs. 1.6 ± 0.6cm/s, P < .05) and A₉₅ (10.0 ± 10.1 vs. 2.5 ± 1.5cm², P < .05), shorter 6MWD (442.2 ± 93.0 vs. 525.3 ± 68.2m, P < .05), and longer TUG (9.4 ± 1.6 vs. 6.5 ± 1.3s, P < .05). Within the PN group, but not the control group, the H-index was correlated with V_AVG (r = -.56, P < .05). Moreover, within the PN group only, PAP scores were correlated with 6MWD (r = -.68, P < .05) and TUG (r = -.59, P < .05) performance. No other statistically significant group difference, correlation or group-dependent regression was observed.

Conclusion

V_AVG, 6MWD, and TUG correlated with LAF reflex loop function observed among those with impaired functioning of the SAF reflex loop. This observation suggests that the LAF reflex loop may be critical to the control of balance in those individuals suffering from small-fiber PN.     

A comparison and optimization of methods and factors affecting the transformation of Escherichia coli

DNA manipulation routinely requires competent bacteria that can be made using one of numerous methods. To determine the best methods, we compared four commonly used chemical methods (DMSO, MgCl₂–CaCl₂, CaCl₂ and Hanahan''s methods) on frequently used Escherichia coli (E. coli) strains: DH5α, XL-1 Blue, SCS110, JM109, TOP10 and BL21-(DE3)-PLysS. Hanahan''s method was found to be most effective for DH5α, XL-1 Blue and JM109 strains (P<0.05), whilst the CaCl₂ method was best for SCS110, TOP10 and BL21 strains (P<0.05). The use of SOB (super optimal broth) over LB [Luria–Bertani (broth)] growth media was found to enhance the competency of XL-1 Blue (P<0.05), dampened JM109′s competency (P<0.05), and had no effect on the other strains (P>0.05). We found no significant differences between using 45 or 90 s heat shock across all the six strains (P>0.05). Through further optimization by means of concentrating the aliquots, we were able to get further increases in transformation efficiencies. Based on the optimized parameters and methods, these common laboratory E. coli strains attained high levels of TrE (transformation efficiency), thus facilitating the production of highly efficient and cost-effective competent bacteria.     

Promoter Polymorphism in the Serotonin Transporter (5-HTT) Gene Is Significantly Associated with Leukocyte Telomere Length in Han Chinese

The serotonin transporter gene (5-HTT)-linked polymorphic region (5-HTTLPR) plays an important role in modulating mood and behavior by regulating 5-HTT expression and thereby controlling the concentration of serotonin (5-HT) in brain synapses: The homozygous shorter allele (S/S) in 5-HTTLPR results in lower 5-HTT expression coupled with stronger psycho-pathological reactions to stressful experiences compared to the homozygous long (L/L) and heterozygous (S/L) alleles. Psychological insults and mood disorders have been shown to cause accelerated telomere shortening, a marker of biological aging, however, it is currently unclear whether the allelic variants of 5-HTTLPR affect telomere length (TL) in the healthy population without mood disorders. In the present study, we determined the relationship between TL and the 5-HTTLPR variants in healthy Han Chinese. The 5-HTTLPR genotyping and leukocyte TL analysis of 280 young female Han Chinese freshmen showed a significantly shorter TL in 149 of them carrying the 5-HTTLPR S/S version compared to those (131) with the L/S or L/S plus L/L genotypes (mean ± SD, 0.533±0.241 for S/S vs 0.607±0.312 for L/S, P  =  0.034; or vs 0.604±0.313 for L/S plus L/L, P  =  0.038). Similar results were achieved in the other cohort including 220 adult healthy individuals of different age, gender and profession (0.691±0.168 for S/S vs 0.729±0.211 for L/S, P  =  0.046, or vs 0.725±0.213 for L/S plus L/L, P  =  0.039). Taken together, shorter leukocyte TL is significantly associated with the 5-HTTLPR S/S allelic variant, which may be implicated in psychological stress-related health problems.