OBJECTIVE: To determine the polymorphism in +252 site of tumor necrosis factor-beta(TNF-beta) gene in patients with or without endometriosis, to evaluate the levels of TNF-alpha and TNF-beta in the serum with or without endometriosis, to explore the relation between polymorphism of TNF-beta gene and the genetic susceptibility of endometriosis, and to explore the pathogenic mechanism of endometriosis at gene level. METHODS: By polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method, polymorphism on +252 site of TNF-beta gene was measured in 82 patients with endometriosis (the endometriosis group) and 80 patients without endometriosis (the control group). With the sandwich-enzyme-linked immunosorbent assay (ELISA), the levels of TNF-alpha and TNF-beta in the serum of the two groups were determined. RESULTS: The TNF-beta level in the serum in the endometriosis group with TNF-beta gene +252 site AA genotype significantly increased, compared with GG genotype (t=2.029, P<0.05); while TNF-alpha and TNF-beta level in the serum had no statistical significance in patients with other genotypes in TNF-beta gene +252 site in the endometriosis group and the control group. CONCLUSION TNF-beta gene +252 site AA genotype might be enhance TNF-beta level in the serum of patients with endometriosis.
Adolescent ; Adult ; Endometriosis ; blood ; genetics ; Female ; Humans ; Lymphotoxin-alpha ; blood ; genetics ; Polymorphism, Genetic ; Tumor Necrosis Factor-alpha ; blood ; Young Adult

OBJECTIVETo investigate the relationship between the presence of the TNF2 allele and plasma concentrations of tumor necrosis factor-alpha (TNF alpha) and soluble TNF receptor (sTNF-R) with the development of acute severe pancreatitis (ASP) and severe sepsis.

METHODSGenomic DNA was prepared from peripheral blood leukocytes. The TNF1 and TNF2 biallelic polymorphisms were identified by analyzing NcoI-digested DNA fragments obtained from PCR products. Plasma levels of TNF alpha and sTNF-R were measured by EASIA.

RESULTSThe overall TNF2 allele frequency in ASP patients was comparable to that found in healthy volunteers (29.2% vs. 29.3%, P > 0.05). Severe sepsis occurred in 26 of 72 patients. Patients with severe sepsis showed a significantly higher prevalence of TNF2 than those without (46.2% vs. 19.6%, P < 0.05). Plasma TNF alpha, sTNF-R I, and sTNF-R II levels were (36 +/- 31) pg/ml, (5.4 +/- 3.5) ng/ml, and (11.2 +/- 7.8) ng/ml, respectively, in patients with severe sepsis, and (31 +/- 25) pg/ml, (4.6 +/- 3.8) ng/ml, and (8.8 +/- 6.6) ng/ml in non-severe sepsis subjects. Differences in TNF levels were not statistically significant between patients with ASP and control group (P > 0.05). Moreover, there was no correlation between TNF2 allele frequency and TNFalpha levels [(37 +/- 31) pg/ml vs. (31 +/- 25) pg/ml in TNF2 group and TNF1 group, respectively, P > 0.05].

CONCLUSIONSOur results suggest that there is no relationship between ASP and the TNF2 allele, but that the TNF2 allele is associated with a susceptibility to severe sepsis as a result of ASP.

Acute Disease ; Adult ; Female ; Humans ; Male ; Middle Aged ; Pancreatitis ; genetics ; Polymorphism, Genetic ; Receptors, Tumor Necrosis Factor ; blood ; Sepsis ; genetics ; Tumor Necrosis Factor-alpha ; analysis ; genetics

OBJECTIVETo approach the effect of tumor necrosis factor-alpha (TNF-alpha) and tumor necrosis factor receptor II (TNFRII) gene polymorphisms on genetic susceptibility of coal worker's pneumoconiosis and their relationship with pulmonary fibrosis.

METHODSTwo hundred and thirty-four cases of coal worker's pneumoconiosis (CWP) and four hundred and forty coal mine workers (controls) were selected, and the cases of CWP were divided into three subgroups based on the various stages of I, II and III. 3 ml peripheral vein blood was drawn from every subject. Using polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLP) techniques, TNF-alpha and TNFRII gene polymorphisms were analyzed.

RESULTSIn both group matching and 1:1 paired matching, there was no significant difference between CWP workers and controls in distribution frequencies of G/A + A/A (TNF-alpha -308) and T/G + G/G (TNFRII 196) genotypes. The distribution frequency of G/A + A/A genotype in CWP with stage III (20.00%) was higher than those in control (10.91%), and CWP cases with stage I (10.34%) and II (7.50%) respectively. The risk of CWP with stage III in those with G/A + A/A genotype was 2-fold higher than with G/G genotype (OR = 3.00, 95% CI: 0.35 approximately 25.84) for 1:1 paired matching.

CONCLUSIONSTNF-alpha and TNFRII gene polymorphisms does not play an important role in susceptibility to CWP of Han race. TNF-alpha gene promoter polymorphisms might be related with the degree of severe pulmonary fibrosis in CWP.

Aged ; Coal Mining ; Genetic Predisposition to Disease ; Genotype ; Humans ; Male ; Middle Aged ; Pneumoconiosis ; blood ; genetics ; Polymorphism, Restriction Fragment Length ; Pulmonary Fibrosis ; blood ; genetics ; Receptors, Tumor Necrosis Factor, Type II ; blood ; genetics ; Tumor Necrosis Factor-alpha ; analysis ; genetics

A main symptom of equine metabolic syndrome (EMS) in ponies is pathological obesity characterized by abnormal accumulation of fat deposits and inflammation. In this study, we analyzed the expression of two pro-inflammatory cytokines, interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha), in subcutaneous adipose tissue and the correlation with serum concentrations in peripheral blood of Welsh ponies. Based on clinical examination findings, the animals were divided into two groups: ponies affected with EMS (n = 8) and obese ponies (n = 8). The adipose tissue was examined using immunohistochemical analysis while concentrations IL-6 and TNF-alpha were measured using enzyme-linked immunosorbent assays (ELISAs). Additionally, histological characterization of the adipose tissue was performed. The results obtained showed that IL-6 expression in adipose tissue biopsies derived from animals with EMS was enhanced while TNF-alpha levels of both groups were comparable. Compared to the obese ponies, EMS animals also had significantly elevated levels of serum IL-6 and TNF-alpha. Histological analysis revealed macrophage infiltration and fibrosis in adipose tissue preparations from the EMS group. These data suggest that IL-6 may play a key role in the course of EMS in Welsh ponies. Our findings also demonstrated that analysis of pro-inflammatory cytokines levels in serum may serve as an additional tool for diagnosing EMS.
Adipose Tissue/*metabolism ; Animals ; Female ; Horse Diseases/blood/*metabolism ; Horses ; Interleukin-6/blood/genetics/*metabolism ; Male ; Metabolic Syndrome X/metabolism/*veterinary ; Tumor Necrosis Factor-alpha/blood/genetics/*metabolism

Animals ; Interleukin-1 ; biosynthesis ; genetics ; Liver ; blood supply ; Lung ; metabolism ; Male ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Wistar ; Reperfusion Injury ; metabolism ; Tumor Necrosis Factor-alpha ; biosynthesis ; genetics

OBJECTIVETo investigate the antagonistic effect and mechanism of the effect of cyproheptadine (Cyp) on endotoxic shock in rats.

METHODSEndotoxic shock was produced in rats by i.v. injection of lipopolysaccharides (LPS) (5 mg/kg). Tumor necrosis factor (TNF(alpha)) mRNA expression was assessed by Northern blot. Plasma TNF(alpha) content was measured by radioimmunoassay. Plasma superoxide dismutase (SOD) activity and malondialdehyde (MDA) content were measured. The intracellular free calcium concentration ([Ca(2+)](i)) in single endothelial cells was determined by laser scanning confocal microscopy (LSCM).

RESULTSCyp 5 mg/kg injected immediately after i.v. LPS raised the mean arterial blood pressure (MABP) of shocked rats and improved their 24 h survival rate. Meanwhile, Cyp markedly decreased TNF(alpha) mRNA levels in rat liver (18 +/- 10 vs. LPS + saline 38 +/- 10, P < 0.01) as well as plasma TNF(alpha) content [(7.8 +/- 2.4) microg/L vs. LPS + saline (21.5 +/- 3.2) microg/L, P < 0.01)]. It enhanced plasma SOD activity [(1037.2 +/- 112.8) NU/L vs LPS + saline (615.4 +/- 92.6) NU/L, P < 0.01], reduced the MDA content [(5.2 +/- 1.1) micromol/L vs. LPS + saline (9.8 +/- 1.5) micromol/L, P < 0.01], and inhibited TNF(alpha)-induced [Ca(2+)](i) elevation.

CONCLUSIONCyp exerts an anti-endotoxic shock effect by inhibiting TNF(alpha) gene expression, enhancing SOD activity, reducing lipid peroxidation, and preventing [Ca(2+)](i) overload.

Animals ; Cyproheptadine ; pharmacology ; Histamine H1 Antagonists ; pharmacology ; Malondialdehyde ; blood ; Rats ; Rats, Wistar ; Shock, Septic ; drug therapy ; metabolism ; Superoxide Dismutase ; blood ; Tumor Necrosis Factor-alpha ; biosynthesis ; genetics

BACKGROUNDTumor necrosis factor-alpha (TNF-&alpha;) is a pleiotropic proinflammatory cytokine and contributes to many kinds of cardiovascular diseases via its receptors (TNFR1/TNFR2). We hypothesize that TNF-&alpha; plays a role in the pathogenesis of chronic atrial fibrillation (AF).

METHODSSixty-seven consecutive patients who were scheduled to have cardiac surgery were enrolled into the study. Thirty-one patients with rheumatic heart disease (RHD) and AF were enrolled as study group (AF group). The sinus rhythm (SR) control groups consisted of 20 patients with RHD and 16 patients with coronary artery disease (CAD). Peripheral blood sample was collected before the operation. About 5 mm(3) left atrial tissue was disserted during the operation and was separated into three parts for Western blotting, real time polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC) analysis.

RESULTSCompared with the controls (RHD SR and CAD SR), the levels of TNF-&alpha; ((14.40 ± 5.45) pg/ml vs. (4.20 ± 3.19) pg/ml vs. (2.68 ± 2.20) pg/ml, P = 0.000) and its soluble receptor 1 (sTNFR1) ((1623.9 ± 558.6) pg/ml vs. (1222.3 ± 175.6) pg/ml vs. (1387.5 ± 362.2) pg/ml, P = 0.001) in plasma were higher in patients with AF. TNF-&alpha; level had positive correlation with the left atrial diameter (LAD) (r = 0.642, P = 0.000). Western blotting analysis showed that the protein levels of TNF-&alpha; (0.618 ± 0.236 vs. 0.234 ± 0.178 vs. 0.180 ± 0.103, P = 0.000) were higher in patients with AF. The RT-PCR analysis results demonstrated that the mRNA expression of TNF-&alpha; (0.103 ± 0.047 vs. 0.031 ± 0.027 vs. 0.023 ± 0.018, P = 0.000) increased in patients with AF. IHC analysis displayed that, comparing to the SR, the expression of TNF-&alpha; (0.125 ± 0.025 vs. 0.080 ± 0.027 vs. 0.070 ± 0.023, P = 0.000) increased in the AF group. The protein level and mRNA expression of TNF-&alpha; also had positive correlation with left atrium diameter (LAD) (r = 0.415, P = 0.000 and r = 0.499, P = 0.000).

CONCLUSIONSThe results revealed that TNF-&alpha; elevated in the plasma and left atrial tissue and had positive correlation with LAD in patients of chronic AF. TNF-&alpha; might involve in the pathogenesis of chronic AF.

Aged ; Atrial Fibrillation ; blood ; metabolism ; Blotting, Western ; Female ; Heart Atria ; metabolism ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Real-Time Polymerase Chain Reaction ; Tumor Necrosis Factor-alpha ; blood ; genetics ; metabolism

OBJECTIVETo investigate the association of single nucleus polymorphisms(SNP)of tumor necrosis factor alpha (TNF-&alpha;) gene (-308 G>A and -238 G>A genotypes) with susceptibility to primary myelodysplastic syndromes (MDS).

METHODSTwo SNPs (TNF-&alpha;-308 G>A,TNF-&alpha;-238 G>A) of TNF-&alpha; gene were detected by Taqman probes in 341 MDS patients and 365 unrelated-healthy controls.

RESULTSCompared to healthy controls, the frequency of TNF-&alpha;-308 AA+AG genotype and A allele increased (18% vs 10%, P=0.015, 9% vs 5%, P=0.021, respectively) in refractory cytopenia with multilineage dysplasia (RCMD) patients. There was no correlation of TNF-&alpha;-308 G>A genotype and allele frequency between MDS and controls. No difference in the genotype and allele frequency of TNF-&alpha;-238 G>A were found between controls and MDS or the subtypes of MDS (P>0.05). We did not find any linkage between plasma level of TNF-&alpha; and TNF-&alpha;-308 G>A or TNF-&alpha;-238 G>A genotype. Statistic differences were observed between platelet count[58(1-611)×10⁹/L vs 90(7-352)×10⁹/L]and bone marrow blasts in MDS patients carrying TNF-&alpha;-308 G>A GG and AA+AG genotype (P=0.024, 0.019, respectively).

CONCLUSIONTNF-&alpha;-308 G>A polymorphism was correlated with susceptibility to MDS-RCMD.

Case-Control Studies ; Gene Frequency ; Genetic Predisposition to Disease ; Genotype ; Humans ; Myelodysplastic Syndromes ; genetics ; Polymorphism, Single Nucleotide ; Tumor Necrosis Factor-alpha ; blood ; genetics

Objective To explore the aptamer specific binding blood group antigen-binding adhesin (BabA) of Helicobacter pylori (H.pylori) for blocking of H.pylori adhering host cell. Methods H.pylori strain was cultured and its genome was extracted as templates to amplify the BabA gene by PCR with designed primers. The BabA gene obtained was cloned and constructed into prokaryotic expression plasmid, which was induced by isopropyl beta-D-galactoside (IPTG) and purified as target. The single stranded DNA (ssDNA) aptamers that specifically bind to BabA were screened by SELEX. Enzyme-linked oligonucleotide assay (ELONA) was used to detect and evaluate the characteristics of candidate aptamers. The blocking effect of ssDNA aptamers on H.pylori adhesion was subsequently verified by flow cytometry and colony counting at the cell level in vitro and in mouse model of infection, respectively. Meanwhile, the levels of cytokines, interleukin 6 (IL-6), IL-8, tumor necrosis factor α (TNF-α), IL-10 and IL-4 in the homogenate of mouse gastric mucosa cells were detected by ELISA. Results The genome of H.pylori ATCC 43504 strains was extracted and the recombinant plasmid pET32a-BabA was constructed. After induction and purification, the relative molecular mass (M_r) of the recombinant BabA protein was about 39 000. The amino acid sequence of recombinent protein was consistent with BabA protein by peptide mass fingerprint (PMF). Five candidate aptamers were selected to bind to the above recombinent BabA protein by SELEX. The aptamers A10, A30 and A42 identified the same site, while A3, A16 and the above three aptamers identified different sites respectively. The aptamer significantly blocked the adhesion of H.pylori in vitro. Animal model experiments showed that the aptamers can block the colonization of H.pylori in gastric mucosa by intragastric injection and reduce the inflammatory response. The levels of IL-4, IL-6, IL-8 and TNF-α in gastric mucosal homogenates in the model group with aptamer treatment were lower than that of model group without treatment. Conclusion Aptamers can reduce the colonization of H.pylori in gastric mucosa via binding BabA to block the adhesion between H.pylori and gastric mucosal epithelial cells.
Animals ; Mice ; Helicobacter pylori/genetics* ; Interleukin-4 ; Interleukin-6 ; Interleukin-8 ; Tumor Necrosis Factor-alpha ; Stomach ; Oligonucleotides ; Adhesins, Bacterial/genetics* ; Blood Group Antigens

The aim of this study was to detect the expressions of transforming growth factor (TGFβ(1)), tumor necrosis factor alpha (TNF&alpha;) and leukemia inhibitory factor (LIF) in newly diagnosed patients with acute myeloid leukemia (AML) and investigate the association between serum levels of various cytokines and clinical outcomes. The levels of TGFβ1, TNF&alpha; and LIF in patient's plasma were detected by enzyme-linked immunosorbent assays (ELISA) and were compared with healthy controls; bone marrow cell morphology, immunology, cytogenetics examinations (MIC) were performed meanwhile. The results showed that levels of TGFβ1, TNF&alpha; and LIF were elevated in AML patients as compared with the controls (13.08±9.77 ng/ml, 10.67±15.11 pg/ml, 4.23±4.73 pg/ml vs 8.23±3.12 ng/ml, 5.86±3.05 pg/ml, 2.78±1.22 pg/ml) (p all<0.05). The three cytokines and MIC examination analysis indicated that level of LIF was abnormally elevated in M5 patients (7.14±6.62 pg/ml); TNF&alpha; was abnormally elevated in M4 and M3 patients especially M4; TGFβ1 level in M6 and M2 patients was higher than others. TGFβ1 plasma concentration in low-risk group the lowest (10.45±4.73 ng/ml), and that in middle risk group was the highest (16.13±13.76 ng/ml) (p<0.05); the levels of other two kinds of factors in the chromosome karyotype groups showed no significant difference. It is concluded that TGFβ1, TNF&alpha; and LIF expressions showed increased level in the untreated patients with de novo AML, the TGFβ1 level among which is associated with the prognosis of patients.
Adolescent ; Adult ; Aged ; Case-Control Studies ; Female ; Hematopoietic System ; metabolism ; Humans ; Karyotyping ; Leukemia Inhibitory Factor ; blood ; Leukemia, Myeloid, Acute ; blood ; diagnosis ; genetics ; Male ; Middle Aged ; Neoplasm Staging ; Prognosis ; Transforming Growth Factor beta1 ; blood ; Tumor Necrosis Factor-alpha ; blood ; Young Adult