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Biomedical subjects

M Müller

Publications and source records attributed to M Müller.

At least 199 records · Page 11Linked to original sources
PubMed2002

In vitro radiosensitivity measured in lymphocytes and fibroblasts by colony formation and comet assay: comparison with clinical acute reactions to radiotherapy in breast cancer patients.

PURPOSE: To compare colony-forming and comet assays on fibroblasts and lymphocytes of 32 breast cancer patients irradiated after breast-conserving operations and to correlate the results with acute clinical radiation reactions in the skin. MATERIAL AND METHODS: Skin fibroblasts were isolated and cultivated before radiotherapy and lymphocytes were drawn prior to the first and directly after the final external irradiation. The colony-forming assay was performed with fibroblasts and the comet assay with lymphocytes and fibroblasts of breast cancer patients according to standard protocols. The clinical radiation reactions of the patients were graded according to the RTOG system. RESULTS: No significant correlation (p =0.09) was detected between clinical acute skin reactions and the in vitro clonogenic data in fibroblasts. Results of the comet assay in lymphocytes, however, showed a significant correlation (p <0.05) with the clinical data when patients were divided into two groups with average and elevated acute reactions. Apart from initial damage, fibroblasts did not show significant differences between the two patient groups. Repeated comet assays in lymphocytes of the same patient drawn before treatment and before and after external radiotherapy demonstrated good reproducibility of the test and no significant impact of preceding radiation treatment. There was a good correlation (r =0.65) between the comet assay results in fibroblasts and lymphocytes of the same individual. CONCLUSIONS: In this cohort of patients, a significant correlation between the in vitro results of the comet assay in lymphocytes and clinical acute reactions was detected. The results of the comet assay and of fibroblast colony formation did not correlate with in vitro radiosensitivity.

Breast Neoplasms↗
PubMed2002

Incidence and risk calculation of inotropic support in patients undergoing cardiac surgery with cardiopulmonary bypass using an automated anaesthesia record-keeping system.

BACKGROUND: This retrospective study analysed the effects of preoperative and intraoperative factors on the occurrence of inotropic support after cardiopulmonary bypass (CPB). METHODS: The data sets of 1471 adult patients having received elective cardiac surgery with CPB were recorded using an online anaesthesia record-keeping system. Patients were judged to have required inotropic drug support if they had received one or a combination of the positive inotropic drugs, epinephrine, dobutamine and enoximone. The effects of age, height, weight, body mass index, gender, chronic heart failure, documented preoperative myocardial infarction, left main coronary artery disease, preoperative history of hypertension, chronic renal failure, diabetes mellitus, chronic obstructive pulmonary disease (COPD), preoperative medical treatment, type of surgical procedure, duration of CPB, duration of aortic clamping and reperfusion time were analysed by logistic regression for predictive power of the need for positive inotropic drugs. RESULTS: Of the patients, 32.4% received positive inotropic drugs in the operating theatre after weaning from CPB. The overall 30-day mortality was 2.2%. Of non-survivors, 81.8% received inotropes compared with 18.2% of survivors (P < 0.01). The numbers of previous myocardial infarctions (odds ratio (OR), 2.01), congestive heart failures New York Heart Association class > 2 (OR, 1.85), COPD (OR, 1.85) and age > 65 yr (OR, 1.62), aortic cross clamping time of > 90 min (OR, 2.32) and coronary artery bypass surgery (OR, 0.43) all represented influential factors within the logistic regression model. CONCLUSION: The knowledge of these risk factors should be useful in increasing the anaesthetist's vigilance in those patients most at risk for inotropic support and in providing for more timely therapeutic intervention and optimizing anaesthesia management.

Aged↗
PubMed2002

Quantification of the expression level of the gene encoding the catalytic subunit of telomerase in testicular tissue specimens predicts successful sperm recovery.

BACKGROUND: The objective of the present study was to evaluate the quantitative detection of human telomerase reverse transcriptase (hTERT) mRNA as a new molecular diagnostic parameter in the work-up of testicular tissue specimens from patients presenting with non-obstructive azoospermia. M ETHODS: hTERT mRNA expression was quantified in 49 cryopreserved testicular tissue specimens by fluorescence real-time RT-PCR in a LightCycler. This was paralleled by conventional histological work-up in all tissue specimens and additional semithin sectioning preparation in cases with maturation arrest (n = 20) and Sertoli cell-only syndrome (SCOS; n = 12). RESULTS: The average normalized hTERT expression (N(hTERT)) was 136.1 +/- 41.7 copies (mean +/- standard deviation) in tissue specimens with presence of haploid germs cells, N(hTERT) = 48.2 +/- 21.0 copies in those with maturation arrest and N(hTERT) = 2.7 +/- 2.8 copies in those with SCOS. The discriminant analysis showed that detection of N(hTERT) was able correctly to classify 89.0% of the investigated tissue specimens. CONCLUSIONS: Our results demonstrate that quantitative detection of hTERT mRNA expression in testicular tissue enables a molecular-diagnostic subclassification of spermatogenesis disorders. Quantitative detection of hTERT in testicular biopsies is thus well suited for predicting successful sperm recovery in patients with azoospermia and is a useful molecular diagnostic parameter for supplementing the histopathological evaluation.

Adult↗
PubMed2002

Multidrug resistance related molecules in human and murine lung.

AIMS: Transporter proteins known to mediate multidrug resistance (MDR) in tumour cells--MDR1 P-glycoprotein (P-gp) and multidrug resistance related protein 1 (MRP1)--are thought to be involved in protecting the lungs against inhaled toxic pollutants. Recently, several new transporter family members have been identified--for example, MRP2, MRP3, and breast cancer resistance protein (BCRP). To study the possible contribution of these proteins and the earlier defined MDR1 and MDR3 P-gp molecules, MRP1, and the major vault protein (MVP) to lung functioning, their expression was analysed in normal lung tissue of humans and several animal species. METHODS: Frozen sections of normal lung tissues were examined for the expression of the multidrug resistance associated proteins, using an extended panel of monoclonal antibodies that specifically detect these proteins in immunohistochemical techniques. RESULTS: In line with earlier reports, the expression of MDR1 P-gp and MRP1 was readily detected in the apical and basolateral membranes, respectively, of the epithelial cell layers of the lungs. In addition, prominent cytoplasmic MVP staining was detected in these layers. In contrast, the recently discovered transporters were either undetectable or they were present at very low values in lung tissue. Immunohistochemical staining in tissues from mice, rats, and guinea pigs points to a strong evolutionary conservation for these transporter proteins. CONCLUSIONS: These results show that the "classic" MDR related molecules, MDR1 P-gp, MRP1, and MVP, should be considered the most important transporters in normal lung physiology. It will be of great interest to investigate differences in expression of both classic and newly defined transporters between normal individuals and-for example, patients with various bronchopulmonary pathological conditions.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2002

The secondary structure of RNA under tension.

We study the force-induced unfolding of random disordered RNA or single-stranded DNA polymers. The system undergoes a second-order phase transition from a collapsed globular phase at low forces to an extensive necklace phase with a macroscopic end-to-end distance at high forces. At low temperatures, the sequence inhomogeneities modify the critical behaviour. We provide numerical evidence for the universality of the critical exponents which, by extrapolation of the scaling laws to zero force, contain useful information on the ground-state (f = 0) properties. This provides a good method for quantitative studies of scaling exponents characterizing the collapsed globule. In order to get rid of the blurring effect of thermal fluctuations, we restrict ourselves to the ground state at fixed external force. We analyze the statistics of rearrangements, in particular below the critical force, and point out its implications for force-extension experiments on single molecules.

Journal Article↗
PubMed2002

Transvaginal bone anchors in female stress urinary incontinence: poor results.

OBJECTIVE: This study evaluates the results of a minimally invasive technique for correcting female stress urinary incontinence by transvaginal implantation of pubic bone anchors. PATIENTS AND METHODS: Female stress urinary incontinence was treated by fixing a gelatin-coated Dacron sling between two miniature titanium anchors with Prolene sutures. RESULTS: A total of 26 patients (median age 57.2 years) underwent the sling procedure. The follow-up examination was performed after 11.4 months on average. Stress incontinence showed a median improvement from grade 2 to grade 0.5 (p = 0.01), although only 16 of the 26 patients were completely continent. Urethral pressure and functional length were not significantly influenced. Impaired vaginal wound healing was seen in 14 of the 26 patients (53.8%), and 13 of them underwent revision. All patients affected (15/26, 57.7%) as well as 1 with uneventful healing showed sensory urge symptoms or detrusor instability (7/26, 26.9%). The correlation between impaired wound healing and detrusor instability was highly significant (p < 0.003). 17 of the 26 patients (65.3%) were dissatisfied or very dissatisfied with the intervention. The unfavorable results did not significantly correlate with the patients' age, the number of previous operations, or the surgeon's skill. CONCLUSION: In view of the poor vaginal wound healing and the resultant irritative symptoms, transvaginal bone anchoring with fixation of a Dacron sling must be regarded as an unsuitable technique.

Bone Screws↗
PubMed2002

Short communication: Identification and characterization of multiple splicing forms of bovine prochymosin mRNA.

Bovine prochymosin (bPC) is an inactive precursor of the milk clotting enzyme chymosin (EC 3.4.23.4), that is present in the abomasum of suckling calves. We investigated the pattern of bPC mRNA expression in the calf stomach tissues by RT-PCR assay and sequence analysis of cloned RT-PCR products. We identified multiple isoforms appearing due to alternative splicing of bPC mRNA. Alternative mRNA forms were generated by skipping one to four full exons within the bPC gene. Various splicing events resulted in seven bPC transcripts, which are 99, 114, 213, 237, 336, 351 and 450 nucleotides shorter compared to full-length mRNA. Analysis of amino acid sequences deduced from alternatively spliced mRNA sequences showed no amino acid transversions and no protein reading frame shift for any splice forms.

Alternative Splicing↗
PubMed2001

Unique phylogenetic relationships of glucokinase and glucosephosphate isomerase of the amitochondriate eukaryotes Giardia intestinalis, Spironucleus barkhanus and Trichomonas vaginalis.

Glucokinase (GK) and glucosephosphate isomerase (GPI), the first two enzymes of the glycolytic pathway of the diplomonads Giardia intestinalis and Spironucleus barkhanus, Type I amitochondriate eukaryotes, were sequenced. GPI of the parabasalid Trichomonas vaginalis was also sequenced. The diplomonad GKs belong to a family of specific GKs present in cyanobacteria, in some proteobacteria and also in T. vaginalis, a Type II amitochondriate protist. These enzymes are not part of the hexokinase family, which is broadly distributed among eukaryotes, including the Type I amitochondriate parasite Entamoeba histolytica. G. intestinalis GK expressed in Escherichia coli was specific for glucose and glucosamine, as are its eubacterial homologs. The sequence of diplomonad and trichomonad GPIs formed a monophyletic group more closely related to cyanobacterial and chloroplast sequences than to cytosolic GPIs of other eukaryotes and prokaryotes. The findings show that certain enzymes of the energy metabolism of these amitochondriate protists originated from sources different than those of other eukaryotes. The observation that the two diplomonads and T. vaginalis share the same unusual GK and GPI is consistent with gene trees that suggest a close relationship between diplomonads and parabasalids. The intriguing relationships of these enzymes to cyanobacterial (and chloroplast) enzymes might reflect horizontal gene transfer between the common ancestor of the diplomonad and parabasalid lineages and the ancestor of cyanobacteria.

Animals↗
PubMed2001

Biocatalytic reduction of beta,delta-diketo esters: a highly stereoselective approach to all four stereoisomers of a chlorinated beta,delta-dihydroxy hexanoate.

A stereoselective chemoenzymatic synthesis of all four stereoisomers of tert-butyl 6-chloro-3,5-dihydroxy-hexanoate (6a) is presented. The key step of the sequence is a highly regio- and enantioselective single-site reduction of tert-butyl 6-chloro-3,5-dioxohexanoate (1a) by two enantiocomplementary biocatalysts. Alcohol dehydrogenase from Lactobacillus brevis (recLBADH) afforded a 72% yield of enantiopure tert-butyl (S)-6-chloro-5-hydroxy-3-oxohexanoate [(S)-2a]. The enantiomer (R)-2a was prepared with 90-94% ee by Baker's yeast reduction in a biphasic system (50% yield). Both biotransformations were performed on a gram scale. The beta-keto group of the enantiomeric delta-hydroxy-beta-keto esters 2a thus obtained was reduced by syn- and anti-selective borohydride reductions. Permutation of the reduction methods yielded all four stereoisomers of the crystalline target compound 6a (> or = 99.3% ee, dr > or = 205:1), which is a versatile 1,3-diol building block. recLBADH accepts a variety of beta,delta-diketo esters as was determined in a photometric assay. tert-Butyl 3,5-dioxo-hexanoate (1b) and tert-butyl 3,5-dioxo-heptanoate (1c) were reduced on a preparative scale as well to afford the corresponding delta-hydroxy-beta-keto esters (R)-2b and (R)-2c with 99.4% ee and 98.1% ee, respectively.

Bacteria↗
PubMed2001

Effects of flavonols on the generation of superoxide anion radicals by xanthine oxidase and stimulated neutrophils.

The effects of three aglycon flavonols (myricetin, quercetin, and kaempferol) and the natural glycoside rutin on superoxide anion radical generating systems were investigated. Quercetin, myricetin, and kaempferol inhibited the formation of uric acid from xanthine by xanthine oxidase, while rutin was ineffective. The generation of superoxide anion radicals by this system was determined by either reduction of cytochrome c or Pholasin luminescence. A scavenging of superoxide was only observed for myricetin and to a small extent for rutin. All flavonols tested inhibited the Pholasin luminescence of fMet-Leu-Phe-stimulated neutrophils. Rutin influenced the oxidative burst of neutrophils in the same way as wortmannin and LY294002, two inhibitors of the phosphoinositide 3-kinase gamma. Indeed, rutin inhibited the activity of this enzyme, whereas the three other flavonols showed no effect. Thus, an inhibition of enzymes involved in signaling rather than a scavenging of superoxide anion radicals dominates in fMet-Leu-Phe-stimulated neutrophils exposed to flavonols.

Cytochrome c Group↗
PubMed2001

Penetration of dacarbazine and its active metabolite 5-aminoimidazole-4-carboxamide into cutaneous metastases of human malignant melanoma.

BACKGROUND: Dacarbazine has been on the market for approximately 3 decades but remains the most effective single agent available for the therapy of metastatic malignant melanoma (MMM). Most MMMs, however, respond poorly to dacarbazine therapy. Apart from tumor resistance at a molecular level, several studies support the notion that therapeutic failure in tumor therapy also might be attributed to an impaired transcapillary drug transfer. METHODS: On the basis of this hypothesis, the authors measured intratumor transcapillary transfer rates of dacarbazine and its active metabolite 5-aminoimidazole-4-carboxamide (AIC) by in vivo microdialysis after intravenous administration of dacarbazine at doses of 200 mg/m(2) to 1000 mg/m(2) (n = 7) in patients suffering from MMM. RESULTS: For all doses, area under the concentration curve (AUC) values for dacarbazine and AIC were not significantly different between plasma and tumor interstitium with AUC(tumor)/AUC(plasma) ratios of 0.97 +/- 0.08 (mean +/- standard error of the mean) for dacarbazine and 0.76 +/- 0.22 for AIC. AUC(0-240) values for dacarbazine and AIC measured in plasma correlated closely with corresponding AUC(0-240)values measured in the interstitium of MMMs with values of r(s) = 0.82 (P = 0.042) and r(s) = 0.90 (P = 0.037), respectively. CONCLUSIONS: The results of this study indicate favorable tumor penetration characteristics of dacarbazine and its active metabolite AIC. The relative lack of response to antineoplastic therapy with dacarbazine, thus might be explained by resistance of melanoma cells at a molecular level rather than by an inability of dacarbazine and AIC to penetrate into the interstitium of MMM.

Aged↗
PubMed2001

Molecular staging of surgical margins after radical prostatectomy by detection of telomerase activity.

BACKGROUND: The further course of prostate cancer (PC) after radical prostatectomy (RPX) is decisively influenced by the local tumor stage. Although it is thus far possible to assess the risk of local recurrence from the pathohistology, precise predictions cannot be made. A more precise evaluation would be desirable, mainly for early planning of adjuvant therapy. Other authors have shown that telomerase activity may be a marker for malignant potential. We assessed the detection of telomerase activity using the telomeric repeat amplification protocol (TRAP) in surgical margins compared to conventional histopathological examination. METHODS: Ninety-two patients with local PC who underwent RPX were examined. After RPX biopsies were obtained from four defined areas of the prostatic fossa and processed by TRAP assay for telomerase activity using a standard protocol. RESULTS: In 5 of 48 patients (10.4%) with organ-confined prostate carcinoma (pT2) telomerase activity could be detected. Seven of 47 patients (14.9%) with locally advanced PC (> pT2) had at least one positive specimen. CONCLUSIONS: The results obtained in our study indicate that detection of telomerase activity by TRAP assay may be a suitable parameter for molecular staging of surgical margins, because of the high tumor-specificity. Further follow-up must clarify whether patients with positive molecular detection have an increased risk of local recurrence.

Biomarkers, Tumor↗
PubMed2001

Ethanol-induced apoptosis in hepatoma cells proceeds via intracellular Ca(2+) elevation, activation of TLCK-sensitive proteases, and cytochrome c release.

Ethanol is known to induce apoptosis in hepatocytes. However, intracellular signaling events of ethanol-induced death are still only partially understood. We studied such processes in ethanol-induced apoptosis in HepG2 cells as a model system for human liver cells. We determined the incidence of apoptosis by DNA fragmentation and tested the effects of various known inhibitors. Ethanol induces apoptosis in HepG2 cells in a dose- and time-dependent manner as well as in rat primary hepatocytes. This effect was not mediated through the death receptor CD95 and the tumor necrosis factor receptors. It was efficiently inhibited by the caspase inhibitor N-benzyloxycarbonyl-Asp-Glu-Val-Asp-fluoromethylketone (zVAD-fmk), the Ca(2+) chelator EGTA, and the serine protease inhibitor N-p-tosyl-l-lysine chloromethyl ketone (TLCK). Upon ethanol treatment, the intracellular calcium ion concentration was increased and cytochrome c was released from the mitochondria, and caspases were activated. EGTA and TLCK could inhibit cytochrome c release from the mitochondria. Furthermore, overexpression of Bcl-x(L) saved cells from ethanol-induced apoptosis. These data suggest that ethanol-induced apoptosis in liver cells is initiated by the intracellular Ca(2+) elevation in the cytoplasm and activation of TLCK-sensitive serine proteases. Our data provide new insight into ethanol-induced apoptosis in liver cells and may lead to therapeutic strategies to prevent liver damage.

Amino Acid Chloromethyl Ketones↗
PubMed2001

Spinodal decomposition in a binary polymer mixture: dynamic self-consistent-field theory and Monte Carlo simulations.

We investigate how the dynamics of a single chain influences the kinetics of early stage phase separation in a symmetric binary polymer mixture. We consider quenches from the disordered phase into the region of spinodal instability. On a mean field level we approach this problem with two methods: a dynamical extension of the self-consistent-field theory for Gaussian chains, with the density variables evolving in time, and the method of the external potential dynamics where the effective external fields are propagated in time. Different wave vector dependencies of the kinetic coefficient are taken into account. These early stages of spinodal decomposition are also studied through Monte Carlo simulations employing the bond fluctuation model that maps the chains-in our case with 64 effective segments-on a coarse grained lattice. The results obtained through self-consistent-field calculations and Monte Carlo simulations can be compared because the time, length, and temperature scales are mapped onto each other through the diffusion constant, the chain extension, and the energy of mixing. The quantitative comparison of the relaxation rate of the global structure factor shows that a kinetic coefficient according to the Rouse model gives a much better agreement than a local, i.e., wave vector independent, kinetic factor. Including fluctuations in the self-consistent-field calculations leads to a shorter time span of spinodal behavior and a reduction of the relaxation rate for smaller wave vectors and prevents the relaxation rate from becoming negative for larger values of the wave vector. This is also in agreement with the simulation results.

Journal Article↗