Plasma TNF-alpha Research Articles

A human model of inflammatory cardio-metabolic dysfunction; a double blind placebo-controlled crossover trial

BackgroundChronic inflammation may contribute to insulin resistance (IR), metabolic syndrome and atherosclerosis although evidence of causality is lacking in humans. We hypothesized that very low-dose experimental endotoxemia would induce adipose tissue inflammation and systemic IR during a low-grade but asymptomatic inflammatory response and thus provide an experimental model for future tests of pharmacologic and genomic modulation of cardio-metabolic traits in humans.MethodsTen healthy, human volunteers (50% male, 90% Caucasian, mean age 22.7 ± 3.8) were randomized in a double-masked, placebo-controlled, crossover study to separate 36-hour inpatient visits (placebo versus intravenous-LPS 0.6 ng/kg). We measured clinical symptoms via the McGill pain questionnaire and serial vital signs. Plasma and serum were collected for measurement of cytokines, C-reactive protein, insulin and glucose, serial whole blood & subcutaneous adipose tissue mRNA expression were measured by real-time PCR. HOMA-IR, a well-validated measure of IR was calculated to estimate insulin resistance, and frequently sampled intravenous glucose tolerance testing (FSIGTT) was performed to confirm an insulin resistant state. We performed ANOVA and within subject ANOVA to understand the differences in cytokines, adipose tissue inflammation and IR before and after LPS or placebo.ResultsThere was no significant difference between placebo and LPS in clinical responses of symptom scores, body temperature or heart rate. However, low-dose endotoxemia induced a rapid and transient 25-fold induction of plasma TNF-alpha and 100-fold increase in plasma IL-6 (Figure 1B) (p < 0.001 for both) both peaking at two hours, followed by modest inflammation in adipose tissue with increases in mRNA levels of several inflammatory genes known to modulate adipose and systemic insulin resistance. Adipose tissue mRNA levels of IL-6 (peak 6-fold, ANOVA F = 27.5, p < 0.001) and TNF-alpha (peak 1.8-fold, F = 2.9, p = 0.01) increased with MCP-1 (peak 10-fold, F = 5.6, p < 0.01) and fractalkine (CX3CL1) (peak 15-fold, F = 13.3, p < 0.001). Finally, HOMA-IR was 32% higher following LPS compared to placebo (p < 0.01) and insulin sensitivity declined by 21% following LPS compared to placebo (p < 0.05).ConclusionsWe present a low dose human endotoxemia model of inflammation which induces adipose tissue inflammation and systemic insulin resistance in the absence of overt clinical response. Such a model has the potential for broad and safe application in the study of novel therapeutics and genomic influences in cardio-metabolic disease.

Abstract 308: Quercetin Intake During Exercise Reduces MCP1, Body Weight and Atherosclerotic Plaque in Mice

Introduction: Atherosclerosis is the major causes of cardiovascular diseases (CVD) and a chronic disease of the large arteries; the disease is mostly attributed to inflammation and oxidative stress. Chemokines are involved in the pathogenesis of atherosclerosis by promoting directed migration of inflammatory cells. Monocyte chemoattractant protein-1(MCP1) has been detected in atherosclerotic lesions by anti-MCP-1 antibody detection. MCP-1 is critical for the initiation and development of atherosclerotic lesions. Animal studies using a variety of antioxidants suggest that antioxidants may protect against CVD. Paradoxically, exercise which induces a severe oxidative stress resulting in the depletion of plasma and tissue antioxidants is an important deterrent of CVD. Aims and Objectives: In the current study we proposed that quercetin a flavonoid will have profound effects on the pathophysiology of atherosclerosis if combined with exercise, we hypothesized its action to be attributed to the inhibition of lipids oxidation, promoting clearance and improving the acute exercise induced oxidative stress resulting in lowering cholesterol, artery lipids deposition and decreasing the development of the plaque. Study Design: This study tested the possible beneficial effects of the combined exercise and quercetin on modulators of oxidation and inflammation in C57BL6 LDL -/- mice fed atherogenic diet. 40 mice were divided into four groups (10 each). These groups are as follows: Control mice, left untreated; control quercetin group, orally supplied with 100 μg/day of quercetin without exercising; exercise group without quercetin, and exercise group with quercetin supplements. The exercise groups were run on a treadmill for 30 minutes, 15m/m/ 5 days/week for 30 days. All animals were on atherogenic diet containing 1.5% cholesterol with total 42% Fat Kcal Diet. At the end of the month of treatment, mice were sacrificed, and atherosclerotic lesions in aorta were quantified. Liver, aorta and adipose tissue gene expressions for genes associated with oxidative stress, inflammation and lipoproteins were analyzed. Results and Conclusion: Mice on quercetin and exercise have significant (P&lt;0.001) drop in body weigh compared to control, these have been accompanied with a 78% drop in the atherosclerotic plaque development in exercising group compared to non-exercising mice without quercetin and 40% drop in atherosclerotic plaque size in exercising mice supplemented with quercetin compared to control group on quercetin. The changes were also accompanied with changes in plasma lipids and inflammatory markers. Although there is no significant differences in the HDL levels between exercising and non-exercising quercetin taking mice, HDL increased significantly (P&lt;0.001) in control on quercetin compared to controls off the treatment. Animals on quercetin and exercise regimen have significantly (P&lt;0.05) reduced plasma MCP-1 compared to controls on quercetin, interestingly sedentary mice on quercetin has 21% reduction in MCP-1 levels compared to the group without the treatment indicating that intake of quercetin may be responsible for the reduction in MCP-1 levels, body weight and atherosclerotic plaque size. However changes in plasmaTNF alpha levels between the groups with or without the treatments were not significant.

Clinical Significance and Expression of PAF and TNF-alpha in Seminal Plasma of Leukocytospermic Patients

Objective. Discuss the changes and roles of PAF in the reproductive tract infection by observing the expression of platelet activating factor (PAF) and tumor necrosis factor α (TNF-α) in seminal plasma of patients with leukocytospermia. Methods. The seminal plasma was obtained from 22 cases of leukocytospermia and 15 cases of normal males; the peroxidase dyeing method was adopted for seminal plasma white blood count; the ELISA was adopted to test PAF and TNF-α concentration in seminal plasma. Result. PAF concentration (2.14 ± 0.43 ng/mL) of leukocytospermia group was significantly lower than the normal group (6.21 ± 1.38 ng/mL, P < 0.01) while TNF-α (5.51 ± 1.46 ng/mL) was significantly higher than that of normal group (3.48 ± 1.08 ng/mL). There was negative correlation between PAF and TNF-α , (r = −0.68, P < 0.01); the same situation existed in PAF and WBC (r = −0.62, P < 0.01); but TNF-α was positively correlated to WBC (r = 0.77, P < 0.01). Conclusion. (1) Low expression of PAF and high expression of TNF-α in leukocytospermia affect the sperm motility, which is one of the reasons that leads to infertility. (2) Lower expression of PAF has its particularity during the reproductive tract infection.

Tumor necrosis factor-alfa and interleukin-4 in cerbrospinal fluid and plasma in different clinical forms of multiple sclerosis

Multiple sclerosis (MS) is an immune-mediated central nervous system disease characterized by inflammation, demyelination and axonal degeneration. Cytokines are proven mediators of immunological process in MS. The aim of this study was to investigate whether there is a difference in the production of the tumor necrosis factor alpha (TNF-alpha) and interleukin-4 (IL-4) in cerebrospinal fluid (CSF) and plasma in the MS patients and the controls (other neurological non-inflammatory diseases) and to determine a possible difference in these cytokines in plasma and CSF in different clinical forms of MS. This study involved 60 consecutive MS patients--48 patients with relapsing-remitting MS (RRMS) and 12 patients with secondary progressive MS (SPMS). The control group consisted of 20, age and sex matched, non-immunological, neurological patients. According to the clinical presentation of MS at the time of this investigation, 34 (56.7%) patients had relapse (RRMS), 14 (23.3%) were in remission (RRMS), while the rest of the patients, 12 (20.0%), were SPMS. TNF-alpha and IL-4 concentrations were measured in the same time in CSF and plasma in the MS patients and the controls. Extended disability status score (EDSS), albumin ratio and IgG index were determined in all MS patients. The MS patients had significantly higher CSF and plasma levels of TNF-alpha than the controls (p < 0.001 for both samples). IL-4 CSF levels were significantly lower in the MS patients than in the controls (p < 0.001), however plasma levels were similar. The patients in relapse (RRMS) and with progressive disease (SPMS) had higher concentrations of CSF TNF-alpha levels than the patients in remission (p < 0.001). IL-4 CSF levels in relapse (RRMS) and SPMS groups were lower than in the patients in remission. The patients in remission had an unmeasurable plasma TNF-alpha level and the patients with SPMS had significantly lower IL-4 levels in plasma than the patients in relapse and remission (p < 0.001). The only significant correlation between cytokine level with either EDSS, or albumin ratio, or IgG index, was found between CSF TNF-alpha levels and albumin ratio in the patients with relapse (R square = 0.431, p < 0.001). According to the obtained data MS relapse was characterized by high concentrations of TNF-alpha in CSF and plasma and low concentrations of IL-4 in CSF. Remission was characterized by high concentrations of IL-4 and low concentrations of TNF-alpha both in CSF and plasma. SPMS was characterized with lower concentrations of TNF-alpha and IL-4 compared to relapse, both in CSF and plasma.

The clinical study about the effect of rosuvastatin on blood lipemia and inflammatory factors

Objective: To investigate the association of genetic polymorphisms of the mitogen-activated protein kinase activated protein kinase 2 gene (MK2)with lowdensity lipoprotein cholesterol (LDL-C) inXinjiangUrgur population. Methods: 350 Uygur males and 595 females were recruited randomly from HETIAN area. The 2 single nucleotide polymorphisms (rs44890 and rs45514798) inMK2 genewere selected and genotyped by Taqman-PCR in the recruited 945 Uygur subjects. All subjects underwent questionnaire, physical examination, measurement of lipid levels and plasma TNF-alpha. Results: (1) In males, the concentration of total cholesterol (TC) [TT:CT:CC:(4.35±1.20) vs (4.69±1.34) vs (4.83±1.44), P=0.033] and TNF-alpha [TT:CT:CC:(106.63±62.39) vs (128.44 ±86.15) vs (153.06±82.99), P=0.001] was different in 3 genotypes of rs44890. But the LDL-C levels inTT, CT, CC genotypes of rs44890were not different neither in males nor in females [male: (2.64±1.16) vs (2.81± 1.28) vs (3.04±1.32), PN0.05; female: (2.42±1.11) vs (2.36±0.99) vs (2.43±1.05), PN0.05]. None of the alleles and genotypes frequencies of rs44890 and rs45514798 were different between high LDL-C group and control group. Linear regression analysis indicated that BMI (beta=0.089) and TNF-alpha (beta=0.002) were significant factors for LDL-C in males, while the age, BMI, and waist/hip ratio for LDL-C in females. However, rs44890 presented as effective factor on plasma TNF-alpha. (3) Two SNPs (rs44890 and rs45514798) were not involved in the logistic regression model, adjusting for gender, age, smoking, alcohol intake,waist/hip ratio, BMI andTNF-alpha, suggestingwhichmay not be effective factors of higher LDL-C. Conclusion: Our data showed negative association of polymorphisms inMK2 genewith LDL-C in Uygur population from HETIAN area of Xinjiang.

Negative association of MK2 gene polymorphisms with low density lipoprotein cholesterol in Uygur population from HETIAN area of Xinjiang

Objective: To investigate the association of genetic polymorphisms of the mitogen-activated protein kinase activated protein kinase 2 gene (MK2) with low density lipoprotein cholesterol (LDL-C) in Xinjiang Urgur population. Methods: 350 Uygur males and 595 females were recruited randomly from HETIAN area. The 2 single nucleotide polymorphisms (rs44890 and rs45514798) in MK2 gene were selected and genotyped by Taqman-PCR in the recruited 945 Uygur subjects. All subjects underwent questionnaire, physical examination, measurement of lipid levels and plasma TNF-alpha. Results: (1) In males, the concentration of total cholesterol (TC) [TT:CT:CC:(4.35±1.20) vs (4.69±1.34) vs (4.83±1.44), P=0.033] and TNF-alpha [TT:CT:CC:(106.63±62.39) vs (128.44±86.15) vs (153.06±82.99), P=0.001] was different in 3 genotypes of rs44890. But the LDL-C levels in TT, CT, CC genotypes of rs44890 were not different neither in males nor in females [male: (2.64±1.16) vs (2.81±1.28) vs (3.04±1.32), P>0.05; female: (2.42±1.11) vs (2.36±0.99) vs (2.43±1.05), P>0.05]. None of the alleles and genotypes frequencies of rs44890 and rs45514798 were different between high LDL-C group and control group. Linear regression analysis indicated that BMI (beta=0.089) and TNF-alpha (beta=0.002) were significant factors for LDL-C in males, while the age, BMI, and waist/hip ratio for LDL-C in females. However, rs44890 presented as effective factor on plasma TNF-alpha. (3) Two SNPs (rs44890 and rs45514798) were not involved in the logistic regression model, adjusting for gender, age, smoking, alcohol intake, waist/hip ratio, BMI and TNF-alpha, suggesting which may not be effective factors of higher LDL-C. Conclusion: Our data showed negative association of polymorphisms in MK2 gene with LDL-C in Uygur population from HETIAN area of Xinjiang.

Plasma L-Cystine/L-Glutamate Imbalance Increases Tumor Necrosis Factor-Alpha from CD14+ Circulating Monocytes in Patients with Advanced Cirrhosis

Background and AimsThe innate immune cells can not normally respond to the pathogen in patients with decompensated cirrhosis. Previous studies reported that antigen-presenting cells take up L-Cystine (L-Cys) and secrete substantial amounts of L-Glutamate (L-Glu) via the transport system Xc- (4F2hc+xCT), and that this exchange influences the immune responses. The aim of this study is to investigate the influence of the plasma L-Cys/L-Glu imbalance observed in patients with advanced cirrhosis on the function of circulating monocytes.MethodsWe used a serum-free culture medium consistent with the average concentrations of plasma amino acids from patients with advanced cirrhosis (ACM), and examined the function of CD14+ monocytes or THP-1 under ACM that contained 0–300 nmol/mL L-Cys with LPS. In patients with advanced cirrhosis, we actually determined the TNF-alpha and xCT mRNA of monocytes, and evaluated the correlation between the plasma L-Cys/L-Glu ratio and TNF-alpha.ResultsThe addition of L-Cys significantly increased the production of TNF alpha from monocytes under ACM. Monocytes with LPS and THP-1 expressed xCT and a high level of extracellular L-Cys enhanced L-Cys/L-Glu antiport, and the intracellular GSH/GSSG ratio was decreased. The L-Cys transport was inhibited by excess L-Glu. In patients with advanced cirrhosis (n = 19), the TNF-alpha and xCT mRNA of monocytes were increased according to the Child-Pugh grade. The TNF-alpha mRNA of monocytes was significantly higher in the high L-Cys/L-Glu ratio group than in the low ratio group, and the plasma TNF-alpha was significantly correlated with the L-Cys/L-Glu ratio.ConclusionsA plasma L-Cys/L-Glu imbalance, which appears in patients with advanced cirrhosis, increased the TNF-alpha from circulating monocytes via increasing the intracellular oxidative stress. These results may reflect the immune abnormality that appears in patients with decompensated cirrhosis.

Early systemic inflammatory response in mice after a single oral gavage with live Escherichia coli is evidenced by increased TNF-alpha and nitric oxide production

Twenty-four female BALB/c mice were orally inoculated with 108 CFU Escherichia coli ATCC 25922 and euthanized 2.5, 7, 13 and 25h post-inoculation. The levels of organ nitric oxide (NO) and plasma endotoxin, TNF-alpha and nitrite/nitrate (NOx) were compared to those found in sham-inoculated mice, to evaluate systemic host-response to a low-level oral exposure to Gram-negative bacteria. Organ bacterial culture and immunohistochemistry for iNOS were performed on lungs, liver, kidneys and brain from all mice. Organ NO and plasma TNF-alpha levels were higher in E. coli-inoculated animals, but no differences were detected in plasma endotoxin levels, NOx or iNOS immunostaining for any of the animal groups. Single oral gavage with live E. coli stimulates an early systemic immune response in clinically healthy mice as evidenced by increased plasma TNF-alpha and organ NO levels, but bacteremia and endotoxemia are not related to this inflammatory response.

血液透析および持続携行式腹膜透析患者の末梢血単核球におけるTNF-α, IL-1βおよびIL-6遺伝子発現

Inflammatory cytokines, including TNF-alpha, IL-1 beta and IL-6, have been reported to be responsible for acute or chronic complications and biocompatibility issues in dialysis patients have focused on the role of these cytokines. Expression of mRNA for TNF-alpha, interleukin (IL)-1 beta and IL-6 in peripheral blood mononuclear cells (PBMC) and their plasma concentrations was examined in 10 hemodialysis and 10 continuous ambulatory peritoneal dialysis (CAPD) patients. Plasma TNF-alpha and IL-1 beta levels in hemodialysis, CAPD and in every healthy subject were below the detection limits. Plasma IL-6 concentration was increased significantly in hemodialysis and CAPD patients when compared to normal controls (p < 0.01). Expression of mRNA for TNF-alpha, IL-1 beta and IL-6 in PBMCs was increased significantly (p < 0.01) in hemodialysis and CAPD patients compared to normal subjects. These mRNA expressions were lower in CAPD patients than in hemodialysis patients (p < 0.05). Significant correlation was observed between the plasma IL-6 concentration and PBMC IL-6 mRNA expression in patients on hemodialysis or CAPD (p < 0.01). IL-6 mRNA expression in PBMCs did not correlate with the duration of hemodialysis or CAPD, blood urea nitrogen (BUN) or serum creatinine levels. These findings suggest that mRNA expression indicates stimulation by dialysis procedures in patients, and CAPD is tolerated better than hemodialysis by patients with end-stage renal failure.

Elevated plasma level of HMGB1 is associated with disease activity and combined alterations with IFN-alpha and TNF-alpha in systemic lupus erythematosus

Recent studies indicate that high-mobility group box protein 1 (HMGB1) contributes to the pathogenesis of diverse autoimmune disorders. It induces the production of interferon-alpha (IFN-alpha) and tumor necrosis factor alpha (TNF-alpha) in vitro. In the present study, plasma HMGB1, TNF-alpha, and IFN-alpha were determined with ELISA in 37 patients with systemic lupus erythematosus (SLE) and 39 age- and sex-matched healthy controls (HC). The possible associations of these cytokines with disease activities, autoantibodies, and certain laboratory parameters were also explored. The plasma levels of HMGB1, TNF-alpha, and IFN-alpha were increased in SLE patients compared with those of HC (P<0.05). Moreover, the levels of HMGB1 and TNF-alpha in the active SLE patients were elevated compared with those in inactive patients and HC. Additionally, plasma HMGB1 was positively correlated with peripheral neutrophils, and plasma TNF-alpha was positively correlated with anti-Sm, ESR and CRP, while plasma IFN-alpha was inversely correlated with the age and platelet level in SLE patients. Our data indicated that increased plasma HMGB1 was associated with disease activity in SLE, which was similar to TNF-alpha. High level of plasma IFN-alpha may be related to nephritis and thrombocytopenia in SLE.

Jak2 V617F Induced TNF Resistance as a Mechanism of Clonal Expansion In Myeloproliferative Neoplasm

AbstractAbstract 795Elevated levels of pro-inflammatory cytokines such as TNF-alpha (TNF) have been implicated in the constitutional symptoms commonly seen in JAK2 V617F MPN. However, it is unclear to what extent they contribute to the disease process itself. We hypothesized that JAK2 V617F may confer a selective advantage on hematopoietic progenitors residing in the high cytokine milieu uniquely present MPN patients, focusing on the pro-inflammatory cytokine TNF.We used ELISA and qPCR to measure TNF in plasma and neutrophils of MPN patients and normal controls. All MPN subtypes had increased plasma concentrations of TNF, with primary myelofibrosis, polycythemia vera and essential thrombocythemia displaying 6, 4 and 2 fold higher mean concentrations of plasma TNF as compared to normal controls, respectively. Median neutrophil TNF mRNA was 2 fold higher in PV patients as compared to normal controls (p<0.001). We then tested the effect of TNF on CFU-GM and BFU-E formation from MPN vs normal mononuclear cells. At low concentrations (1ng/ml) of TNF we observed a decrease in BFU-E from normal controls to 82% of that without TNF but an increase in MPN BFU-E to 120% (p<0.05) of that without TNF. We also observed a blunting of TNF induced CFU-GM suppression in MPN versus normal. We genotyped colonies and found that in the presence of TNF colony formation was almost exclusively JAK2 V617F, demonstrating that TNF selects for JAK2 V617F colony growth. To distinguish between direct and indirect TNF effects (i.e. effects mediated through bystander cells) as well as to potentially augment the differential effects of TNF on normal and MPN by the use of a more homogenous cell population we performed equivalent colony formation experiments on CD34+ cells. The most notable difference observed was the effect 10ng/ml TNF on CFU-GM. TNF reduced CFU-GM colony formation by normal CD34+ cells to 50%, but increased CFU-GM colony formation by MPN cells to 120% (p<0.001). The suppressive effect of 10ng/ml on BFU-E was also blunted in MPN versus normal CD34+ cells (49% vs 30% of that without TNF) (p=0.054). Genotyping of MPN colonies allowed us to compare the effect of TNF on colony formation on three distinct populations: (1) JAK2WT MPN CD34+ cells, (2) JAK2V617F MPN CD34+ cells and (3) normal CD34+ cells. As compared to normal controls (59% decrease), JAK2 non-mutated MPN CD34+ cells are hypersensitive to myeloid colony suppression (78% decrease) whereas TNF paradoxically enhances colony formation by JAK2 V617F CD34+ cells (180% increase). This suggests that “pre-JAK2” CD34+ cells are functionally disadvantaged when compared to those from normal controls and implicates TNF in the selection and clonal expansion of the JAK2 V617F clone. To confirm the latter we tested whether ectopic expression of JAK2 V617F in murine hematopoietic progenitors blunts TNF-induced inhibition of myeloid colony formation. In the presence of TNF, colony formation of hematopoietic progenitors expressing empty vector or BCR-ABL (included as an additional control) was decreased to 60%, while TNF had no effect on progenitors with ectopic expression of JAK2 V617F. This suggests that JAK2 V617F directly imparts TNF resistance. To interrogate the role of TNF for the development of MPN in vivo, we infected TNF+/+ and TNF−/− bone marrow with JAK2 V617F-GFP retrovirus and transplanted the cells into genotypically matched recipients. Two strains of mice were used in independent experiments (C57BL6 and B6129S6). Equal numbers of TNF+/+ and TNF−/− GFP+ cells per mouse were injected. Although peripheral blood counts were comparable, JAK2 V617F burden (assessed by GFP%) was significantly lower in recipients of TNF−/− knockout bone marrow compared to recipients of TNF+/+ marrow. This difference in JAK2 V617F burden between the two groups was apparent in peripheral blood (47% vs 3.7%, p <0.05) and bone marrow (52% vs 2%, p<0.05) and the spleens of the TNF−/− were smaller (0.46g vs 0.11g, p=0.078).Our data suggest that JAK2 V617F-induced TNF resistance may drive the expansion of the mutant clone by conferring a growth advantage to JAK2 V617F cells over their JAK2 wild-type counterparts in the high cytokine microenvironment of MPN patients. Further, JAK2WT MPN cells exhibit intrinsic TNF hypersensitivity compared to normal cells, which greatly enhances this selective advantage. This JAK2V617F-negative state may play a role in the predisposition to acquire MPN. Disclosures:No relevant conflicts of interest to declare.

93 Cytokines Associate with Tissue Factor But Not with Thrombin Formation in Preterm Infants

Background: Tissue factor (TF), the in vivo initiator of coagulation cascade, is a central link between inflammatory and coagulation pathways.Objective: To evaluate associations between cytokine concentrations, TF and thrombin formation in preterm infants.Methods: We measured proinflammatory cytokines (IL-6, IL-8, TNF-alpha), anti-inflammatory cytokine IL-10, free TF and, as indicator of thrombin generation, prothrombin fragment 1+2 (F1+2) in 56 VLBW infants (GA 23+5-31+5 wks, BW 600-1500g). Cord blood and blood samples on day 1, 3, and 5-7 were collected. From plasma cytokines were analyzed with Luminex technique, free TF and F1+2 were analyzed with Elisa tests (American Diagnostica, Dade Behring).Results: IL-6 correlated with free TF in cord samples and on days 1 and 3 (R=0.53, p=0.013, n=21; R=0.380, p=0.019, n=38; R=0.373, p=0.042, n=30). IL-8 correlated with free TF on days 1 and 3 (R=0.597, p< 0.001, n=38; R=0.462, p=0.009, n=31). TNF-alpha and IL-10 both correlated with free TF in cord samples (R=0.536, p=0.012, n=31, and R=0.610, p=0.003, n=22). No correlations were found between cytokines and F1+2.Conclusions: Both pro- and anti-inflammatory cytokines correlated with high TF concentration. A difference seems to exist between in utero (cord) and postnatal TF regulation: in cord plasma TNF-alpha as well as IL-10 showed strong associations with TF but lost this role to IL-8 postnatally. Free TF failed to contribute to the rate of thrombin generation. This shift of TF from coagulation to inflammation may have effects on pathogenesis of lung injury, as well as on extrapulmonary morbidity in preterm infants.

Could nephrotoxicity due to colistin be ameliorated with the use of N-acetylcysteine?

The protective effect of N-acetylcysteine (NAC) on nephrotoxicity due to contrast nephropathy and reperfusion-induced ischemia has been reported in experimental models. However, its efficacy on colistin-induced nephrotoxicity has not been elucidated yet. The primary aim of this study was to evaluate the nephrotoxic effect of colistin and to investigate the possible protective effect of NAC on colistin-induced nephrotoxicity. The secondary aim was to research the systemic effects of nephrotoxicity-induced oxidative stress on the lung. Eighteen female Sprague-Dawley rats were randomly assigned and were given (a) 1 ml/kg sterile saline, (b) 300,000 IU/kg/day colistin, and (c) 300,000 IU/kg/day colistin and 150 mg/kg NAC for six consecutive days. Plasma blood urea nitrogen (BUN), creatinine, urinary creatinine, urinary protein, plasma TNF-alpha levels, renal tissue superoxide dismutase (SOD) and malondialdehyde (MDA) activity and immunocytochemical findings were evaluated. Colistin exerted nephrotoxicity and achieved a significant increase in plasma BUN and creatinine levels and renal tissue SOD levels. NAC exhibited no significant effect on biochemical parameters but reduced renal tissue SOD level and reversed immunocytochemical staining of inducible nitric oxide synthase (i-NOS) and neurotrophin-3. Increased oxidative stress in the lung tissue of the rats treated with colistin has also been documented. Additionally, NAC significantly reduced the immunostaining of endothelial NOS (e-NOS) and i-NOS in the lung tissue. Colistin-induced renal toxicity may be attributable to oxidative damage. The combined treatment of colistin plus NAC seems to have a beneficial role in restoration of the oxidant injury which may be related to its antioxidant effect.

Elevated expression of tumor necrosis factor‐α signaling molecules in colonic tumors of Zucker obese (fa/fa) rats

Zucker obese rats are highly sensitive to colon cancer and possess a plethora of metabolic abnormalities including elevated levels of cytokine tumor necrosis factor-alpha (TNF-alpha). The main objective of this study was to determine if physiologically elevated TNF-alpha affects colonic tumor phenotype with regard to an altered TNF-alpha signaling pathway. Zucker obese (fa/fa, homozygous recessive for dysfunctional leptin receptors), Zucker lean (Fa/fa, Fa/Fa) and Sprague-Dawley (SD) rats were injected twice with azoxymethane (10 mg/kg) over 2 weeks. After 30 weeks, the animals were terminated and physiological and tumor parameters were assessed. Obese rats had notably higher body and organ weights as well as plasma TNF-alpha, insulin and leptin levels than lean or SD animals. A 100% tumor incidence and significantly higher tumor size, multiplicity and burden were found in obese rats compared to the lean group that had 47.8% tumor incidence. The SD group had the lowest tumor incidence (20.0%). Tumors from obese animals had higher protein levels of TNF-alpha, TNF-alpha-receptor-2 (TNFR2), nuclear transcription factor-kappaB (NF-kappaB) and IkappaB-kinasebeta (IKKbeta) compared to lean animals. In both obese and lean groups, expression levels of these proteins were higher in tumors than in surrounding, normal-appearing colonic mucosae. These findings support an important role for TNF-alpha signaling in tumorigenesis and demonstrate that tumors growing in an obese state had significantly different expression levels of TNFR2 and NF-kappaB, proteins known to play a critical role in growth and survival, than those growing in the lean state. It is concluded that the physiological state of the host intricately affects tumor phenotype.

Effects of the HIV-1 Protein Tat on Myocardial Function and Response to Endotoxin

HIV-1 infection has been associated with cardiomyopathy in a subset of patients. In order to determine whether HIV-1 alters myocardial function or the myocardial response to stress, transgenic mice that express the HIV-1 protein Tat were used. Heart function was assessed using the isolated working heart preparation. Response to infection was assessed by measuring heart function at various times after endotoxin administration. Since cytokines are implicated in myocardial dysfunction, plasma tumor necrosis factor alpha (TNF-alpha) and interleukin-6 (IL-6) and myocardial mRNA and protein levels of TNF-alpha and IL-6 were determined. Tat by itself did not cause myocardial dysfunction; however, 4h after endotoxin, myocardial function was more severely compromised in the Tat mice than in control mice. Plasma TNF-alpha levels were elevated at 2h and higher in the control group but myocardial levels were similar in the two groups. Plasma IL-6 was increased but myocardial levels were different only at 24h at which time myocardial function was no longer depressed. Tat expression, by itself, did not impair intrinsic myocardial function but did increase myocardial injury induced by endotoxin. Although cytokines are associated with dysfunction, TNF-alpha and IL-6 were probably not responsible for the exaggerated dysfunction in Tat mice receiving endotoxin.

Lack of therapeutic effects of gabexate mesilate on the hepatic encephalopathy in rats with acute and chronic hepatic failure

Inflammation plays a pivotal role in liver injury. Gabexate mesilate (GM, a protease inhibitor) inhibits inflammation by blocking various serine proteases. This study examined the effects of GM on hepatic encephalopathy in rats with acute and chronic liver failure. Acute and chronic liver failure (cirrhosis) were induced by intraperitoneal TAA administration (350 mg/kg/day for 3 days) and common bile duct ligation, respectively, in male Sprague-Dawley rats. Rats were randomized to receive either GM (50 mg/10 mL/kg) or saline intraperitoneally for 5 days. Severity of encephalopathy was assessed by the Opto-Varimex animal activity meter and hemodynamic parameters, mean arterial pressure and portal pressure, were measured (only in chronic liver failure rats). Plasma levels of liver biochemistry, ammonia, nitrate/nitrite, interleukins (IL) and tumor necrosis factor (TNF)-alpha were determined. In rats with acute liver failure, GM treatment significantly decreased the plasma levels of alanine aminotransferase (P = 0.02), but no significant difference of motor activity, plasma levels of ammonia, IL-1beta, IL-6, IL-10 and TNF-alpha or survival was found. In chronic liver failure rats, GM significantly lowered the plasma TNF-alpha levels (P = 0.04). However, there was no significant difference of motor activity, other biochemical tests or survival found. GM-treated chronic liver failure rats had higher portal pressure (P = 0.04) but similar mean arterial pressure in comparison with saline-treated rats. Chronic GM treatment does not have a major effect on hepatic encephalopathy in rats with TAA-induced acute liver failure and rats with chronic liver failure induced by common bile duct ligation.

Virological and immunological features of active cytomegalovirus infection in nonimmunosuppressed patients in a surgical and trauma intensive care unit

Cytomegalovirus (CMV) reactivation occurs frequently in critically ill patients. The natural course of CMV infection and the interaction between CMV and the adaptive immune system in this setting remain poorly defined. Fifty-three CMV-seropositive patients in a surgical and trauma intensive care unit were included in this study. The CMV DNA load in tracheal aspirates (TA) and plasma (PL) was monitored by qPCR. CMV-specific T-cell immunity was assessed by intracellular cytokine staining. Plasma TNF-alpha levels were determined by ELISA. CMV reactivation occurred in 39.7% of patients (23% had CMV DNA detected only in TA). The analysis of TA allowed an earlier diagnosis in 28% of patients. Clearance of CMV DNAemia preceded that of CMV DNA in TA in some episodes. Peak CMV DNA levels were significantly higher in TA than in PL (P = 0.02). CMV reactivation developed in the presence of CMV-specific T cells. Termination of CMV reactivation was associated with an expansion of functional CMV-specific T cells. Plasma levels of TNF-alpha did not allow for the prediction of the occurrence of CMV reactivation. CMV-specific T-cell immunity is preserved in most critically ill patients experiencing CMV reactivation. Analysis of respiratory specimens is imperative for an optimal monitoring of CMV reactivation in this setting.

Anti-inflammatory action of lipid-bound apoA-I cysteine mutants

To determine the anti-inflammatory functions of different cysteine mutants of apolipoprotein A-I recombinant HDLs. The recombinant HDLs (named rHDL52, rHDL107, rHDL173, rHDLwt) were reconstituted by mixing wild types or their mutants with dipalmitoyl phosphatidylcholine. And the in vivo effects on lipopolysaccharide (LPS)-induced endotoxemia were examined in mice. The plasma levels of plasma tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta and IL-6 in by ELISA were tested. And we also set up two control groups: LPS and saline. The rHDL52 mice had a significant decrease of plasma TNF-alpha and IL-1beta and the protection of lung against acute injury. 24 h post-injection as compared with rHDLwt group [TNF-alpha: (135.28 +/- 12.84) pg/ml, IL-1beta: (82.00 +/- 8.19) pg/ml], the rHDL52 mice exhibited a higher capability of lowering the plasma levels of TNF-alpha and IL-1beta [(39.66 +/- 2.44) pg/ml, (66.83 +/- 6.24) pg/ml, both P < 0.05]. And, as indicated by histological sections of lung tissue, the rHDL52 mice could also lower the infiltration of inflammatory cells in lung. rHDL52 has a higher anti-inflammation capability than wild type rHDLwt.

Effects of One Year Treatment of Vildagliptin Added to Pioglitazone or Glimepiride in Poorly Controlled Type 2 Diabetic Patients

The aim of the study was to compare the effects of vildagliptin added to pioglitazone or glimepiride on metabolic and insulin resistance related-indices in poorly controlled type 2 diabetic patients (T2DM). 168 patients with T2DM were randomized to take either pioglitazone 30 mg once a day plus vildagliptin 50 mg twice a day or glimepiride 2 mg 3 times a day plus vildagliptin 50 mg twice a day. We evaluated body weight, body mass index (BMI), glycated hemoglobin (HbA1c), fasting plasma glucose (FPG), postprandial plasma glucose (PPG), fasting plasma insulin (FPI), homeostasis model assessment insulin resistance index (HOMA-IR), homeostasis model assessment beta-cell function index (HOMA-beta), fasting plasma proinsulin (FPPr), proinsulin/fasting plasma insulin ratio (Pr/FPI ratio), adiponectin (ADN), resistin (R), tumor necrosis factor-alpha (TNF-alpha), and high sensitivity C-reactive protein (Hs-CRP) at their baseline values, and after 3, 6, 9, and 12 months of treatment. We observed a similar improvement of HbA1c, FPG, PPG, and Hs-CRP compared to baseline in the 2 groups. Fasting plasma insulin, FPPr, Pr/FPI ratio, R, and TNF-alpha were significantly decreased and ADN was significantly increased with pioglitazone plus vildagliptin, but not with glimepiride plus vildagliptin. HOMA-IR, and HOMA-beta values obtained with pioglitazone plus vildagliptin were significantly better than the values obtained with glimepiride plus vildagliptin. Pioglitazone plus vildagliptin were found to be more effective in preserving beta-cell function, and in reducing insulin resistance, and inflammatory state parameters.

Involvement of Various Molecular Events in Cellular Injury Induced by Smokeless Tobacco

Smokeless tobacco (ST) consumption is implicated in the pathogenesis of oral diseases, including cancer. However, its pathological effect in other organs is not well understood. In the present study, the effect of aqueous extract of smokeless tobacco (AEST) prepared from "gutkha" (a form of ST) on the xenobiotic drug-metabolizing enzymes, histopathological changes, and damage to the genetic material in lung, liver, and kidney of rats was evaluated. Animals were orally administered AEST at a low dose (L-AEST, 96 mg/kg body wt/day) for 2 (L-AEST(2)) and 28 weeks (L-AEST(28)) and at a high dose (H-AEST, 960 mg/kg body wt/day) for 2 weeks (H-AEST(2)). Real-time PCR and immunohistological studies showed that administration of L-AEST(2) did not induce the expression of phase I cytochrome P450s (CYP1A1, 1A2, and 2E1) and phase II mu-glutathione-s-transferase (GST-mu) drug-metabolizing enzymes in lung, liver, and kidney. Although H-AEST(2) administration significantly induced both gene and protein expression of CYP1A1, 1A2, and 2E1 in all of the above organs, it mildly expressed the phase II detoxifying enzyme, GST-mu, in type I and type II epithelial cells of lung and in proximal tubular cells of kidney. L-AEST(28) enhanced the gene and protein expression of CYP1A1, 1A2, and 2E1 in lung, liver, and kidney in a differential manner and induced the expression of GST-mu in lung and kidney. L-AEST(28) induced the micronuclei formation in the peripheral blood mononuclear cells, TNF-alpha in plasma, and myeloperoxidase activity in the organs. L-AEST(28) significantly enhanced Bax, p53, and NF-kappaB and decreased Bcl-2 gene expressions differentially in an organ-specific manner. The differential changes in these organs due to AEST might be due to their different physiological functions and variable sensitivities toward the metabolites of AEST, which create a microenvironment favorable for AEST-induced pathogenesis. This study broadens the insight into the different molecular mechanisms in various organs, which appear to be deregulated due to AEST. Understanding these processes may help in clinical treatment planning strategies for tobacco-related diseases.