Biomedical subjects

P Bufler

Publications and source records attributed to P Bufler.

9 recordsLinked to original sources

PubMed2004

Effect of advanced glycation end products on endotoxin-induced TNF-alpha, IL-1beta and IL-8 in human peripheral blood mononuclear cells.

BACKGROUND/AIMS: Advanced glycated end products (AGE) are endogenous proteins that have formed covalent complexes with sugars by a nonenzymatic process. Being proinflammatory molecules, AGE are thought to contribute to chronic systemic and local inflammatory processes associated with pathological changes in various diseases. In patients with end-stage renal disease, AGE are believed to play a role in the progression of atherosclerosis and worsening of renal failure. In patients receiving hemodialysis, AGE are thought to contribute to the inflammatory components of the therapy, particularly in diabetic patients. METHODS: In the present study, AGE were produced using 5% human serum albumin (HSA) and 50% glucose, both used for intravenous infusion into humans and both released after strict control for endotoxin content. The presence of AGE formed by HSA and glucose was confirmed using 2 independent assays. The inflammatory properties of these AGE were assessed using synthesis and release of the proinflammatory cytokines interleukin-1 (IL-1), tumor necrosis factor (TNF) and IL-8, a chemokine. RESULTS: Alone, AGE did not induce these cytokines from peripheral blood mononuclear cells (PBMC) obtained from 14 healthy human donors. However, in the presence of 1 or 10 ng/ml of endotoxin, AGE augmented the production of IL-1 and TNF above that induced by endotoxin alone. Although the amount of augmentation of LPS-induced cytokines by AGE varied between the blood donors, the response was consistently observed and reached statistical significance. The augmentation of cytokine production was confirmed using AGE prepared with different lots of HSA and glucose. CONCLUSION: These results demonstrate that in the strict absence ofendotoxins, AGE are formed that do not stimulate cytokine production from PBMC of healthy donors, however, AGE significantly augment the synthesis and release of proinflammatory cytokine in the presence of low concentrations of endotoxins. The data suggest that renal replacement therapies should consider the role of microbial products in potentiating the biological consequences of naturally formed AGE and their potential to contribute to systemic and local inflammation in renal replacement therapies. Therefore, although the formation of AGE is unavoidable, excluding microbial products during renal replacement therapy should reduce the pathological consequences of AGE.

Adult↗

PubMed2002

Changes of exhaled nitric oxide during steroid treatment of childhood asthma.

Exhaled nitric oxide (eNO) is elevated in several inflammatory airway diseases and is significantly reduced by anti-inflammatory treatment with inhaled steroids. The aim of this randomized, open clinical trial was to evaluate eNO in relation to conventional lung function parameters at rest and after exercise during sequential changes of inhaled steroids in children with persistent asthma. The study consisted of a 4 week run-in period, a 4 week washout phase and a randomized treatment period during which only one group was treated again with inhaled budesonide. After run-in, eNO was reduced to normal values, and rose again during washout. In the patients randomized to steroid treatment, eNO was again decreased, whereas it remained unchanged in the untreated patients. Forced expiratory volume in one second and forced vital capacity at rest and after exercise improved significantly after run-in, but showed no difference after randomization. However there was a strong correlation of eNO with patient compliance. Exhaled nitric oxide was able to differentiate between children briefly treated with or without steroids, the conventional lung-function variables however could not. In practice exhaled nitric oxide may thus be a valuable parameter to monitor adherence to steroids, but less suitable to describe physiologically relevant impairments of lung function.

Adolescent↗

PubMed2001

Surfactant proteins D and A in sputum.

The surfactant proteins SP-D and SP-A are collectins and central components of the innate immune system of the peripheral lungs. They bind to carbohydrates on microorganisms and promote their removal. In the larger airways their role and concentrations are not yet known. Sputum may thus potentially be useful to characterize the collectins in this compartment. Induced sputum was obtained from healthy adults and spontaneous sputum from patients with cystic fibrosis. ELISA and carbohydrate binding assays were used to quantify the amount and functional capacity of the collectins in plugs or sol-fractions, prepared by centrifugation of the plugs. SP-D and SP-A were detectable in only about 50% of the sputum plugs. Varying the salt, detergent and calcium conditions of the assay did not improve the results. In samples with reproducibly detectable SP-D or SP-A, their carbohydrate binding capacity was zero. Sputum sol fraction and LPS inhibited the binding of the collectins to carbohydrate in the presence of calcium. Whereas SP-D and SP-A are abundant in the peripheral lung, their presence in sputum derived from the larger airways is variable and their carbohydrate binding capacity is lost.

Adolescent↗

PubMed2001

Dumping syndrome: a common problem following Nissen fundoplication in young children.

Within a 3-year period we observed six children, aged 3 months to 4 years, with dumping syndrome (DS) following Nissen fundoplication. Five were neurologically normal and one was slightly mentally retarded. Symptoms included postprandial tachycardia, diaphoresis, lethargy, severe retching, meteorism or gas-bloat syndrome, watery diarrhea, refusal to eat with failure to thrive, and developmental delay. Results of an oral glucose tolerance test did not correlate with the severity of symptoms. In one child severe retching led to recurrent paraesophageal gastric herniation, necessitating two reoperations and a gastrostomy. A lack of meal stimulated pancreatic polypeptide release in this patient indicated vagal damage. Initially five of the six children needed continuous intragastric feeding until bolus application of a carbohydrate-modified diet was tolerated. We conclude that the diagnosis of DS is often delayed in spite of characteristic clinical signs. Intensive postoperative follow-up after antireflux surgery should be mandatory in children.

Child, Preschool↗

PubMed1997

Nebulization of a bovine surfactant in cystic fibrosis: a pilot study.

Cystic fibrosis (CF) is a lethal disorder which results in excessive airway secretions and in chronic inflammation of the airways. In vitro and in vivo studies have shown that a lack of surfactant results in the closure of the small airways. In this pilot study, we aimed to determine whether surfactant administered by aerosol might improve lung function on a short-term basis in patients with CF. In a randomized, crossover double-blind pilot study, 120 mg of a lipid-extracted bovine surfactant (Alveofact) or placebo was aerosolized to five young adult patients with CF over a period of 30 min for five consecutive days. The sample size had the power of 90% to detect an increase in forced expiratory volume in one second (FEV1) of 15% (p < 0.05). Jet nebulization of surfactant produced particles of which more than 75% were the respirable range (< 5 microns). The inhalations were well tolerated. No changes in serum antibody titres against the surfactant proteins-B and -C (SP-B/SP-C) were observed. No differences in FEV1 and forced vital capacity were found before, and 30 or 90 min after, the inhalation. This pilot study shows no acute or short-term benefits of surfactant inhalation in young adults with cystic fibrosis. However, a beneficial effect of exogenous surfactant cannot be excluded before other reasons for a lack of effect, such as insufficient quantity delivered, inhomogeneous distribution or inhibition of the surfactant in the lungs, have been completely ruled out.

Administration, Inhalation↗

PubMed1996

The myeloid differentiation antigen CD14 is N- and O-glycosylated. Contribution of N-linked glycosylation to different soluble CD14 isoforms.

The myeloid differentiation antigen CD14 acts as the major receptor for bacterial lipopolysaccharide (LPS). A soluble form of the protein (sCD14) is present in human serum which functions as a soluble LPS receptor. We have compared the isoform patterns of soluble CD14 derived from human serum and of the recombinant proteins produced by CHO cells transfected with either the wild-type CD14 gene or with a cDNA coding for a truncated protein which lacks the C-terminal 21 amino acids [sCD14-(1-335)-peptide]. Using SDS/PAGE, two dominant isoforms (53 and 50 kDa) and two minor forms (46 and 43 kDa) can be detected in serum as well as in the supernatants of both transfectants. sCD14 is a glycoprotein which carries N- and O-linked carbohydrates. The different isoforms of sCD14-(1-335)-peptide are due to differences in the content of N-linked sugars. However after the removal of N- and O-linked carbohydrates from serum- and CHO-derived wild-type proteins, two isoforms are still present. These results indicate that N-linked glycosylation contributes to but does not fully explain the different forms of soluble CD14. We further examined whether the mutation of individual N-linked glycosylation sites influences the expression of membrane-bound and soluble CD14 forms and the ability of the membrane-bound molecule to bind LPS. As with the wild-type proteins, the different isoforms of the soluble mutants are partially due to differences in N-linked glycosylation. A truncated mutant which lacks the two N-terminal glycosylation sites {[Asp18, Asp132]CD14-(1-335)peptide} does not give rise to multiple forms on SDS gels. Like CD14-(1-335)-peptide, this mutant is not expressed on the cell surface suggesting that a smaller isoform present in the wild-type preparations results from proteolytic cleavage of the membrane-bound molecule. N-linked carbohydrates do not seem to be important for the binding of LPS to membrane-bound CD14.

Animals↗

PubMed1995

CD40 function in nonhematopoietic cells. Nuclear factor kappa B mobilization and induction of IL-6 production.

CD40 is a member of the TNF receptor family that was first characterized as an important T-B cell interaction molecule. This receptor is also expressed on many other cell types, including normal basal epithelium, carcinomas, and transformed cell lines. The functions of CD40 in non-B cells are largely unknown. Our studies demonstrate that CD40 mediates nuclear factor kappa B (NF-kappa B) mobilization and IL-6 production in nonhematopoietic cells. Stimulation of the transformed fibroblast cell line SV80 with CD40 ligand (CD40L) or anti-CD40 Ab resulted in the production of IL-6; this could be increased by IFN-gamma pretreatment, which is known to up-regulate CD40 expression. Studies with transfectants overexpressing CD40 demonstrated that activation of CD40 alone is sufficient to induce IL-6 production. The transcription factor NF-kappa B appears to play a central role in CD40-mediated activation of the IL-6 gene; NF-kappa B mobilization directly preceded CD40-mediated IL-6 production, and suppression of NF-kappa B mobilization with the metabolic inhibitor D609 also suppressed the IL-6 response. A striking similarity to the requirements for TNF-induced IL-6 production, which is mediated by the p55TNF receptor in SV80 cells, was observed. In view of the intracellular homologies between CD40 and the p55TNF receptor, it should be considered that the two receptors share common components in their signaling pathways that lead to IL-6 production.

Base Sequence↗

PubMed1995

Soluble lipopolysaccharide receptor (CD14) is released via two different mechanisms from human monocytes and CD14 transfectants.

The receptor for lipopolysaccharide LPS (CD14) exists in a membrane-associated (mCD14) and a soluble form (sCD14). Previous studies indicate that monocytes produce sCD14 by limited proteolysis of the membrane-bound receptor. In this study we demonstrate that human monocytes also produce sCD14 by a protease-independent mechanism. To investigate the molecular nature of this second pathway we studied sCD14 formation in the monocytic cell line Mono Mac 6 (MM6) and in CD14 transfectants. Both MM6 and the CD14 transfectants constitutively produce sCD14 by a protease-independent mechanism. Structural analysis of sCD14 produced by the CD14 transfectants reconfirmed the presence of the COOH terminus predicted from the cDNA. Since glycosylphosphatidylinositol anchor attachment is associated with the removal of a hydrophobic C-terminal signal peptide, our finding demonstrates that the transfectants secrete sCD14 which escaped this posttranslational modification. Identical results obtained for sCD14 derived from peritoneal dialysis fluid of a patient with kidney dysfunction show the in vivo relevance of this pathway for sCD14 production.

Amino Acid Sequence↗

PubMed1995

Functional discrepancies between tumor necrosis factor and lymphotoxin alpha explained by trimer stability and distinct receptor interactions.

Tumor necrosis factor (TNF) and lymphotoxin alpha (LT alpha) are closely related cytokines which bind with nearly identical affinities to the same pair of cell surface receptors, p55 and p75TNFR. Therefore it is assumed that TNF and LT alpha are redundant cytokines. This study, however, demonstrates that TNF and LT alpha differ significantly with regard to their mitogenic and cytotoxic potentials. LT alpha's superior mitogenic effect could be explained by its formation of a more stable trimer. In contrast to the TNF trimer, which disintegrated under physiological conditions into biologically inactive monomers, the LT alpha trimer remained stable for several days. Accordingly, LT alpha more effectively induced fibroblast growth which demands long-term presence of the cytokine. TNF's superior cytotoxicity, which requires only short-term impact of the cytokine, could be attributed to a distinct interaction with the human p55TNFR. This was demonstrated in NIH 3T3 cells transfected with the human p55TNFR, where cytotoxicity is mediated exclusively by the transfected receptor. Although the p55ATNFR had virtually identical affinities for TNF and LT alpha, as defined by Scatchard analysis, it nevertheless discriminated between binding of each cytokine and showed a 200-fold enhanced cytotoxicity mediated by TNF.

3T3 Cells↗