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Frank Stuber

Publications and source records attributed to Frank Stuber.

13 recordsLinked to original sources
PubMed2006

Interleukin 10.G microsatellite in the promoter region of the interleukin-10 gene in severe sepsis.

BACKGROUND: The highly polymorphic interleukin 10.G (IL10.G) microsatellite located in the promoter region of the interleukin-10 (IL-10) gene exerts a positive transcriptional regulatory effect on IL-10 gene expression and correlates with the in vitro IL-10 secretion. This study was conducted to investigate whether IL10.G microsatellite is associated with the incidence and/or the outcome of severe sepsis. METHODS: One hundred and fifteen patients with severe sepsis who had been treated at the intensive care unit of the university hospital were studied. One hundred and forty-one healthy individuals served as controls. IL10.G microsatellite genotyping was performed with the following two methods: fluorescent based polymerase chain reaction (PCR) techniques and silver staining of the amplified DNA fragment in polyacrylamide gel. Alleles were defined according to the size of the amplified DNA product. RESULTS: Ten alleles and 36 genotypes were detected both in the patients with severe sepsis and in the healthy controls. Allele IL10.G9 and allele IL10.G13 were the commonest alleles with the frequencies of 32.6% and 21.3% respectively in the patients with severe sepsis, and 34% and 27% respectively in the healthy controls. The allele frequencies of IL10.G microsatellite were neither different between the patients with severe sepsis and the healthy controls (P > 0.05), nor between survivors and non-survivors (P > 0.05). However, the frequency of one common allele IL10.G13 was slightly lower in the patients with severe sepsis than in the healthy controls (21.3% vs 27%, P > 0.05), and the frequency of allele IL10.G9 was slightly higher in the non-survivors than in the survivors (37.1% vs 28.1%, P > 0.05). CONCLUSION: IL10.G microsatellite may neither contribute to the susceptibility to severe sepsis nor to the fatal outcome of severe sepsis.

Female↗
PubMed2006

Enantiomeric determination of tramadol and O-desmethyltramadol by liquid chromatography- mass spectrometry and application to postoperative patients receiving tramadol.

A liquid chromatographic-mass spectrometric assay with atmospheric pressure chemical ionization is presented for quantification (selected-ion mode) of tramadol (T) and O-desmethyltramadol (ODT) in blood plasma after liquid-liquid extraction. The enantiomeric separation was achieved on a Chiralpak AD column containing amylose tris-(3,5-dimethylphenylcarbamate) as chiral selector. The validation data were within acceptable limits. The assay was successfully applied to authentic plasma samples allowing confirmation of diagnosis of overdose or intoxication as well as monitoring of patients' compliance. In postoperative patients receiving T, mean therapeutic plasma concentrations were determined as follows: 422.8 and 440.8 ng/mL for (+)-T and (-)-T, respectively, and 23.7 and 40.7 ng/mL for (+)-ODT and (-)-ODT, respectively. Quantitative results demonstrated a great interindividual variability in postoperative plasma analgesic drug concentrations and, therefore, a wide therapeutic concentration range. The 7.7% of the samples that revealed negative results for ODT are likely explained by genetic polymorphisms resulting in absent enzyme activity of CYP2D6.

Analgesics, Opioid↗
PubMed2006

Protection against Pseudomonas aeruginosa pneumonia and sepsis-induced lung injury by overexpression of beta-defensin-2 in rats.

Beta-defensin-2 (BD-2), a small cationic antimicrobial peptide, was first described to be an inducible defensin at the epithelial surfaces. In vitro studies have demonstrated that it may play a pivotal role in the anti-inflammatory immune response in addition to its antimicrobial activity. The purpose of this study was to evaluate the effect of overexpression of BD-2 on lung injury to crudely investigate whether the function of BD-2 in the lung attributed to both antimicrobial action and modulation of the immune response. Recombinant adenovirus carrying an expression cassette of rat BD-2 or control adenovirus carrying empty vector was administered intratracheally to Sprague-Dawley rats 48 h before performing acute lung injury, which was induced either by Pseudomonas aeruginosa infection or by cecal ligation and double puncture (2CLP). In vivo antimicrobial activity of BD-2, histological changes of the lungs in both infectious and 2CLP models, pulmonary intracellular adhesion molecule-1 protein level, as well as the 7-day survival rate in the latter model were determined. Amounts of the P. aeruginosa in the lung with BD-2 overexpression were significantly lower compared with that in controls (2.87+/-0.76x10(4) colony-forming units [CFU]/mL vs. 2.49+/-0.74x10(6) CFU/mL, P<0.05). Overexpression of BD-2 reduced alveolar damage, interstitial edema, and infiltration of neutrophils in both models. Furthermore, in the 2CLP model, recombinant BD-2 not only significantly decreased protein levels of intracellular adhesion molecule-1 in lung tissue at 24, 36, and 72 h after 2CLP (P<0.05), but also significantly improved the survival of rats (P<0.05). The CFU of abdominal bacteria was comparable to that in the control rats (P>0.05). Therefore, overexpression of BD-2 protects against P. aeruginosa pneumonia and 2CLP-induced lung injury based on its antimicrobial and anti-inflammatory activities, respectively. Modulating the expression level of BD-2 may serve as an approach to attenuate lung injury.

Animals↗
PubMed2005

Screening of copy number polymorphisms in human beta-defensin genes using modified real-time quantitative PCR.

Defensins are cationic antimicrobial peptides, which play an important role in host immune defense to some infectious diseases as well as immune disease and skin disease. Recent studies identified that the genes coding for human beta-defensin 2 (DEFB4), human beta-defensin 3 (DEFB103) and human beta-defensin 4 (DEFB104) showed variation in copy numbers. This variation may have an impact on gene expression levels. Here, we have demonstrated a real-time PCR-based method to measure beta-defensin gene copy number. Using this relative real-time quantitative PCR, we developed a new rapid and reliable approach, which involves amplification of the target locus (DEFB4 or DEFB103 or DEFB104) and the single-copy reference locus (human serum albumin, ALB) in a single PCR reaction. A calibrator was prepared by recombining one copy of the target gene and one copy of the reference gene into a plasmid. After correcting the PCR amplification efficiency, which differed between the defensin gene and ALB gene, and normalization by the calibrator, the ratio of the copy number of the target gene to that of the reference gene in an unknown sample was determined. This normalized ratio directly related to the gene copy number. The assay was validated using previously genotyped samples, which demonstrated high accuracy and reliability of the method. Furthermore, this method was used to screen the copy number variations of these three beta-defensin genes in healthy blood donors. This method proved to be a reliable and fast tool to genotype gene copy number variations in projects associating genomic variations with gene expression or with population phenotypes in epidemiologic studies.

Base Sequence↗
PubMed2005

Cytokine promoter polymorphisms in severe sepsis.

The need to develop individualized risk profiles and drug therapy regimens motivates interest in genetic studies of critically ill patients. Gene promoter variants may predict interindividual variability in response to inflammatory stimuli, such as infection and trauma. Genomic variations also may affect gene expression profiles, as well as the structure and production of proteins. The genes involved in inflammation are numerous, as are genomic variations within most of those genes. Cytokine genes involved in inflammatory cascades are important candidate genes that may determine the extent of a person's response to injury. Understanding the genetic determination of the inflammatory process includes the possibility of developing valuable diagnostic tools and new therapeutic approaches in severe sepsis. To date, specific patterns of markers of genomic variation reliably indicating at-risk populations do not exist. Evaluation of possible genomic markers for risk stratification of patients with sepsis and persons at high risk of developing organ failure has begun at a level of well-powered genetic epidemiological research. Cytokine promoter variants may contribute substantially to studies of genetic predisposition of sepsis because they operate in a gene region of high regulatory activity.

Animals↗
PubMed2005

Postoperative analgesia in children--current practice in Germany.

German departments of anaesthesia were surveyed to determine current practice of postoperative pain management in children. The response rate of the survey was 58.6%: Questionnaires of 383 departments in which paediatric surgery was performed could be analyzed. 37.3% operated an acute pain service (APS). In 58.8% of the hospitals, postoperative pain management in children was mainly performed by surgeons or pediatricians. Anaesthesiologists or an APS were in charge for pain management in children in 38.6% of the institutions. Non-opioid analgesics were the drugs most widely used (93.4%), whereas i.v. opioids were never used in 20.9% of the hospitals and used less than once a week in 28.7%. The intramuscular route was chosen at least occasionally by 27.7% of the respondents. Peripheral and central regional techniques were performed in most of the departments, however, frequency of use varied considerably between hospitals running or not running an APS. The majority performed the techniques of regional anaesthesia less than once a week. The basic primary quality criterion of pain therapy, a regular measurement and documentation of pain scores, was performed in only 4% of the institutions. Paediatric pain management does not meet quality criteria and standards of care already established in adults. In the future, additional education of the medical staff considering analgesic techniques and measurement of pain scores has to be emphasized.

Analgesics↗
PubMed2005

Human genotyping.

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Genetic Techniques↗
PubMed2003

Impact of CYP2D6 genotype on postoperative tramadol analgesia.

Genetic polymorphisms result in absent enzyme activity of CYP2D6 (poor metabolizers, PM) in about 10% of the Caucasian population. This study investigates whether the PM genotype has an impact on the response to tramadol analgesia in postoperative patients. A prospective study design was used and 300 patients recovering from abdominal surgery were enrolled. After titration of an individual loading dose, patients could self-administer 1 ml bolus doses of the drug combination tramadol 20 mg/ml, dipyrone 200 mg/ml and metoclopramide 0.4 mg/ml via patient-controlled analgesia (PCA). Patients' genotype was analyzed considering the most prevalent PM associated CYP2D6 mutations using a real-time PCR and hybridization based genotyping method. Demographic data, surgery related variables, pain scores, analgesic consumption and need for rescue medication were compared between extensive metabolizers (EM) and PM. The primary outcome criterion 'response' was defined as responder or non-responder status by the need for rescue medication and patients' satisfaction at the final interview. Demographic and surgery related data were comparable between EM (n=241) and PM (n=30). The percentage of non-responders was significantly higher in the PM group (46.7%) compared with the EM group (21.6%; p=0.005). Tramadol loading dose amounted to 108.2+/-56.9 and 144.7+/-22.6 mg (p<0.001) in EM and PM, respectively. More patients displaying the PM genotype needed rescue medication in the recovery room and during PCA period than patients with at least one wild type allele (21.6 versus 43.3%, p=0.02). PM for CYP2D6 showed a lower response rate to postoperative tramadol analgesia than EM. Therefore, CYP2D6 genotype has an impact on analgesia with tramadol. Pharmacogenetics may explain some of the varying response to pain medication in postoperative patients.

Adult↗
PubMed2003

IL-10 polymorphism is associated with increased incidence of severe sepsis.

OBJECTIVE: To investigate whether three biallelic polymorphisms at positions -592, -819 and -1082 in the promoter region of the IL-10 gene are associated with increased incidence of severe sepsis. METHODS: The IL-10 -592, -819 and -1082 polymorphisms were typed using polymerase chain reaction followed by digestion with the restriction enzymes RsaI, MaeIII and MnlI, respectively. RESULTS: Patients with severe sepsis were more likely to have IL-10 -1082 allele 1, compared with controls (P < 0.05). Genotype distribution of the IL-10 -1082 polymorphism significantly differed between patients and controls (P < 0.05). However, the allele frequencies and genotype distribution of the IL-10 -1082 polymorphism did not differ between surviving and dead patients (P > 0.05). No significant differences in the genotype distribution and allele frequencies of the IL-10 -592 and IL-10 -819 polymorphisms were observed between patients with severe sepsis and healthy controls, nor between surviving and dead patients (P > 0.05). CONCLUSIONS: The polymorphism at position -1082 in the promoter region of the IL-10 gene may be associated with susceptibility to severe sepsis. In contrast, the other two highly linked IL-10 polymorphisms are not associated with incidence or the outcome of severe sepsis.

Alleles↗
PubMed2002

Real-time PCR for detection and differentiation of gram-positive and gram-negative bacteria.

We developed a consensus real-time PCR protocol that enables us to detect spiked bacterial 16S DNA from specimens such as water, urine, plasma, and sputum. The technique allows an exact Gram stain classification of 17 intensive care unit-relevant bacteria by means of fluorescence hybridization probes. All tested bacteria were identified correctly, and none gave a false-positive signal with the opposite Gram probe.

Bacterial Infections↗
PubMed2002

Comparison of different decontamination methods for reagents to detect low concentrations of bacterial 16S DNA by real-time-PCR.

Contamination of polymerase chain reaction (PCR) reagents continues to be a major problem when consensus primers are used for detection of low concentrations of bacterial DNA. We designed a real-time polymerase chain reaction (PCR) for quantification of bacterial DNA by using consensus primers that bind specifically to the 16S region of bacterial DNA. We have tested four different methods of decontamination of PCR reagents in a project aimed at detecting bacterial DNA at low concentrations: deoxyribonuclease (DNAse) treatment, restriction endonuclease digestion, UV irradiation, and 8-methoxypsoralen in combination with long-wave UV light to intercalate contaminating DNA into double-stranded DNA. All four methods result in inhibition of the PCR reaction, and most of the decontamination procedures failed to eliminate the contaminating bacterial DNA. Only the DNAse decontamination proved to be efficient in eliminating contaminating DNA while conserving PCR efficiency. All four decontamination methods are time consuming and have the possibility of carrying new contamination into the reaction mixture. However, decontamination with DNAse may help, together with the use of highly purified PCR reagents, in detecting small amounts of bacterial DNA in clinical specimens.

Artifacts↗