[ICS preserves interdisciplinary character. 32nd Annual Meeting of the International Continence Society (ICS) in Heidelberg, 28-30 August, 2002].
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Germany↗
Biomedical subjects
G Kramer
Publications and source records attributed to G Kramer.
Urolithiasis in allograft kidneys.
OBJECTIVES: Allograft stones are an uncommon clinical problem and management is mainly based on anecdotal experience, rather than analysis of larger series. METHODS: In an 8-year period, 19 patients were treated for 19 renal and 3 ureteral stones. In 9 patients, stones were transplanted and 10 formed de novo stones within a mean of 28 months (range 13 to 48) after transplantation. In 4 patients, stones were removed during transplantation. Seven patients were treated with extracorporeal shock wave lithotripsy (ESWL), 3 patients had stones removed percutaneously, 1 by antegrade ureteroscopy, and 1 at the time of ureteral reimplantation. Three patients passed stones spontaneously. RESULTS: In 3 of 4 patients with stones detected before transplantation, the procedure was completed successfully after endoscopic stone removal. Three of 5 patients with transplanted stones required emergency nephrostomy; 1 patient had permanent renal impairment. Three (42.9%) of 7 patients treated with ESWL needed transient nephrostomy; ultimately, all became stone free within a mean 15 days (range 10 to 40). Endoscopic stone removal always resulted in complete clearance without renal impairment. All patients were stone free during a follow-up of 29 months (range 13 to 48). CONCLUSIONS: Nine (47%) of 19 stones were actually transplanted. Therefore, intraoperative screening by ultrasonography with subsequent endoscopic removal is advisable. Small stones (4 mm or less) may be closely followed up, because they can pass spontaneously. ESWL is the treatment of choice for caliceal stones sized 5 to 15 mm. However, for stones greater than 15 mm or for ureteral stones, antegrade endoscopic procedures seem to be more favorable.
Adult↗
Differential expression of interleukin-15, a pro-inflammatory cytokine and T-cell growth factor, and its receptor in human prostate.
BACKGROUND: Pro-inflammatory interleukin (IL)-15 plays a major role in host defense and chronic inflammation by stimulating T-lymphocyte recruitment and growth. Expression of IL-15 and IL-15 receptor (IL-15R) in human prostate was examined. METHODS: Normal and benign hyperplastic (BPH) prostate specimens (n = 23) were analyzed for IL-15 and IL-15Ralpha-chain expression by immunohistochemistry and Real-Time-PCR/RT-PCR. Regulation of prostatic stromal cell (PSC) IL-15 mRNA and effect of IL-15 on prostatic cell growth were analysed in vitro. RESULTS: In normal prostate, anti-IL-15 and anti-IL-15Ralpha-chain reactivity were restricted to smooth muscle and stromal cells. However, in BPH, in addition epithelial cells frequently exhibited discrete anti-IL-15R and often intense, membranous anti-IL-15 reactivity. IL-15/IL-15R mRNA were detected in all prostatic cells types. In BPH tissues, IL-15 mRNA content was variable (15-fold). IL-15 mRNA synthesis of PSC was significantly up-regulated by IFN-gamma. Furthermore IL-15 strongly stimulated the growth of BPH-T-lymphocytes and weakly that of carcinoma cell lines, but not of stromal cells. CONCLUSIONS: Overexpression of IL-15 and IL-15Ralpha-chain in BPH and massive proliferation of BPH-T-lymphocytes induced by IL-15 suggest a role for IL-15 in prostatic inflammation. Since IFN-gamma, a T-lymphocyte product, stimulates prostatic IL-15 production; chronic inflammation might be triggered by this paracrine loop.
Adolescent↗
Binding specificity of Escherichia coli trigger factor.
The ribosome-associated chaperone trigger factor (TF) assists the folding of newly synthesized cytosolic proteins in Escherichia coli. Here, we determined the substrate specificity of TF by examining its binding to 2842 membrane-coupled 13meric peptides. The binding motif of TF was identified as a stretch of eight amino acids, enriched in basic and aromatic residues and with a positive net charge. Fluorescence spectroscopy verified that TF exhibited a comparable substrate specificity for peptides in solution. The affinity to peptides in solution was low, indicating that TF requires ribosome association to create high local concentrations of nascent polypeptide substrates for productive interaction in vivo. Binding to membrane-coupled peptides occurred through the central peptidyl-prolyl-cis/trans isomerase (PPIase) domain of TF, however, independently of prolyl residues. Crosslinking experiments showed that a TF fragment containing the PPIase domain linked to the ribosome via the N-terminal domain is sufficient for interaction with nascent polypeptide substrates. Homology modeling of the PPIase domain revealed a conserved FKBP(FK506-binding protein)-like binding pocket composed of exposed aromatic residues embedded in a groove with negative surface charge. The features of this groove complement well the determined substrate specificity of TF. Moreover, a mutation (E178V) in this putative substrate binding groove known to enhance PPIase activity also enhanced TF's association with a prolyl-free model peptide in solution and with nascent polypeptides. This result suggests that both prolyl-independent binding of peptide substrates and peptidyl-prolyl isomerization involve the same binding site.
Amino Acid Sequence↗
An additional serine residue at the C terminus of rhodanese destabilizes the enzyme.
The rhodanese coding sequence was extended at its 3' end by three base pairs to generate mutants coding for a serine or arginine residue at the carboxyl terminus of the protein. Wild-type and mutant coding sequences were expressed in a cell-free Escherichia coli system by coupled transcription/translation. Predominantly full-length protein was formed in all cases. The amount of protein synthesized was quantified by incorporation of radioactive leucine into polypeptides. Enzymatic activity of in vitro synthesized rhodanese was determined at different temperatures. Specific enzymatic activity was calculated and is assumed to reflect the portion of the protein that is in its native three-dimensional conformation. It was observed that rhodanese extended by one serine at the C terminus lost enzymatic activity when incubated above 30 degrees C, in contrast to wild-type protein or variant rhodanese extended by an arginine residue. Similarly, variant rhodanese with an additional serine residue was more susceptible to urea denaturation than the other two rhodanese species. These results are surprising in light of the crystal structure of the protein.
Animals↗
Folate status among patients with non-small cell lung cancer: a case-control study.
BACKGROUND AND OBJECTIVES: This case control study explored the purported inverse relationship between folate status and lung cancer development. METHODS: Folate status of 46 postoperative non-small cell lung cancer (NSCLC) patients was compared to that of 44 non-cancer patients. Cancer patients had completed treatment > 3 months prior and had no evidence of cancer. Ineligibility criteria for all patients included (1) > 2 alcoholic drinks/day (2) ongoing tobacco use, or (3) folate supplementation > 400 microg/day. RESULTS: No differences were found between groups in serum and RBC folate after adjustment for age and use of folate-interfering medications: geometric means (GM) x /geometric standard error (GSE): 7.9 ng/ml x /1.1 vs. 7.8 ng/ml x /1.1, respectively (P = 0.91) for serum folate; 264 ng/ml x /1.1 vs. 263 ng/ml x /1.1, respectively (P = 0.97) for RBC folate. Age- and creatinine-adjusted homocysteine was no different between groups: GM x /GSE: 9.4 micromol/L x /1.0 vs. 8.6 micromol/L x /1.0, respectively (P = 0.17). No difference were seen in folate intake. Frequencies of the homozygous genotype for the MTHFR polymorphism, an enzyme important in folate metabolism and associated with a reduced risk of other cancers, were no different. CONCLUSIONS: This case control study does not support the hypothesis that low folate is an independent risk factor for NSCLC.
Aged↗
High-grade prostate cancer is associated with low serum testosterone levels.
BACKGROUND: The aim of this study was to assess whether low serum testosterone levels in men with newly diagnosed prostate cancer have an association to the endocrine status, prostate-specific antigen (PSA) levels, Gleason score, and androgen receptor expression. METHODS: Besides a full clinical work-up, the following hormones were quantified in men with newly diagnosed prostate cancer by serum analysis: total testosterone, human luteinising hormone (hLH), human follicle stimulating hormone (hFSH), estradiol, and dehydroepiandrostendione (DHEA). In a subgroup of men, androgen receptor expression was determined immunohistochemically. RESULTS: One hundred and fifty six patients (65.7 +/- 8.5 yrs) with a mean PSA of 29.8 ng/ml (median: 7.4 ng/ml) were analysed. Fifty-two patients (33%) had a partial androgen deficiency (serum testosterone < 3.0 ng/ml). These men had lower hLH (3.3 vs. 5.9 mIU/ml), hFSH (6.2 vs. 8.4 mIU/ml), and estradiol (18.8 vs. 29.1 pg/ml) serum levels. Mean Gleason score was higher (7.4 vs. 6.2) in men with a low serum testosterone, PSA-levels were lower (25.3 vs. 31.9 ng/ml). Mean testosterone levels decreased from 4.1 +/- 1.7 ng/ml in patients with Gleason scores < or = 5 to 2.8 +/- 2.7 ng/ml with Gleason scores > or = 8. Androgen receptor expression was higher in patients with low serum testosterone. CONCLUSIONS: Patients with high Gleason score prostate cancer have lower testosterone and estradiol serum levels. The fact that gonadotropins were lower in parallel suggests a tumor-mediated suppression of the hypothalamic-pituitary-gonadal hormone axis particularly in men with high Gleason score tumours.
Adult↗
[Kidney Transplantation Study Circle of the German Urology Society Graduate and Continuing Education Committee. Report of the annual meeting, 12-14 October 2000, Rostock].
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Education, Medical, Continuing↗
Folding of a nascent peptide on the ribosome.
Even though very significant progress has been made recently in elucidating the structure of the bacterial ribosome and topological assignments of its functional parts, the molecular mechanism of how a peptide is formed and how the nascent peptides is folded on the ribosomes remains uncertain. Here, the current progress and remaining problems are considered from the standpoint of the authors. Topics considered include formation of peptide bonds and models that represent this process, the vicinity of RNA to the nascent peptide, the cotranslational folding hypothesis, evidence that some but not all nascent peptides pass through a region within the 50S ribosomal subunit, presumably the tunnel, in which they are folded and sheltered, pause-site peptides, and the involvement of chaperones in folding of nascent proteins on ribosomes. The chaperone-like activity of the large ribosomal subunit in renaturation of denatured proteins is reviewed. It is concluded that cotranslational folding of some but not all nascent peptides occurs in the large ribosomal subunit. It is suggested that this folding is facilitated by changes in the conformation of the ribosome that are related to the reaction cycle of peptide elongation.
Bacterial Proteins↗
Cotranslational folding--omnia mea mecum porto?
Evidence for cotranslational folding on both prokaryotic and eukaryotic ribosomes is reviewed. Molecular chaperones appear to assist only a small fraction of newly synthesized proteins in folding into their native conformation. The recently published crystal structure of the large ribosomal subunit at 2.5 A resolution has provided the basis for understanding where and how peptide synthesis takes place on the ribosome. The nascent peptide is concluded to pass through a tunnel that extends about 100 A between the peptidyl transferase center and its exit site. The minimum diameter of the tunnel and the apparent physical and chemical properties of its walls appear to preclude complex folding of the nascent peptide within most of the length of the tunnel. However, results indicate that nascent peptides that are protected within the ribosomes vary in length from about 30 to 72 amino acid residues. This suggests that nascent peptides have different conformations. It is hypothesized that folding of the nascent polypeptide into its native conformation starts in the distal portion of the tunnel, and proceeds at the surface of the ribosomal subunit in a depression or bay near the exit opening of the tunnel.
Animals↗
ILAE commission report: evaluations and awards at the 4th European Congress of Epileptology, Florence, 7-12 October 2000.
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Awards and Prizes↗
Local intratumoral tumor necrosis factor-alpha and systemic IFN-alpha 2b in patients with locally advanced prostate cancer.
To examine tolerability and activity of local, intratumoral tumor necrosis factor-alpha (TNF-alpha) and systemic interferon-alpha2b (IFN-alpha2b) in locally advanced, hormone-resistant prostate cancer (LA-HRPC), 10 patients with LA-HRPC (T4N x M0, n = 3, T4N x M1, n = 5; T4N1M1, n = 2) were treated with recombinant TNF-alpha injected locally into prostate tumor tissue at 4-week intervals (maximum of four cycles) combined with intermittent subcutaneous (s.c.) administration of 5 x 10(6) IU IFN-alpha2b. Twenty-nine TNF-alpha cycles were administered. Despite significant TNF-alpha leakage into the systemic circulation 2 h after intraprostatic application (from a mean of 9 to a mean of 416 pg/ml; p = 0.0034), TNF-alpha (and IFN-alpha2b) was well tolerated (WHO grade 1-2 toxicity), possibly because of its rapid neutralization by increasing soluble 55-kDa and 75-kDa TNF receptor levels in the serum (mean increase 268% and 91%, respectively) at the same time. TNF-alpha induced prostate tumor cell necrosis in all patients, leading to a significant reduction of prostate volume in 9 of 10 cases (mean 38%; p = 0.0025). The significant short-term increase of prostate-specific antigen (PSA) (mean 65%; p < 0.001), tissue polypeptide-specific antigen (TPS) (mean 85%; p = 0.001), and possibly interleukin-8 (IL-8) (mean 2687%; p < 0.009) serum levels within 4 h after TNF-alpha confirmed the cytotoxic effect in vivo. In the long term, serum PSA levels dropped by 18%-87%, reaching the nadir value 7 weeks after baseline. Objective responses of metastases were not seen. Intraprostatic administration of TNF-alpha is feasible at a tolerable toxicity in patients with LA-HRPC and, thus, may be a new treatment option for these patients.
Aged↗
Human accessory cells activate fresh, normal, tumor-distant T lymphocytes but not tumor-infiltrating T lymphocytes to lyse autologous tumor cells in a primary cytotoxic T lymphocyte assay in renal cell carcinoma.
OBJECTIVES: Search for an ideal responder T-lymphocyte source for adoptive T-lymphocyte therapy in renal cell carcinoma (RCC). METHODS: Cytotoxic T-lymphocyte (CTL) activity of (a) normal, tumor-distant, renal T lymphocytes, (b) tumor-infiltrating T lymphocytes and (c) peripheral blood T lymphocytes against autologous tumor epithelial cells (EC) of 10 patients with organ-confined, primary RCC was analyzed in a primary CTL assay. Freshly enriched T lymphocytes were cultured with or without autologous, mitomycin-C-treated normal or tumor EC in the presence or absence of antigen-presenting cells (APC) for 7 days. RESULTS: Both tissue T-lymphocyte populations displayed a similar CD4:CD8 ratio (1:1). Elevated CD62L coexpression of CD4+ T lymphocytes in normal, tumor-distant, renal tissue resulted in a significantly higher transient T-cell activation than that seen in renal tumor tissue (46 vs. 27%; p = 0.002). All trials to induce significant lysis of autologous, renal tumor EC in tumor-infiltrating and peripheral blood T lymphocytes failed. Only when normal, tumor-distant, renal T lymphocytes were stimulated by autologous APC and tumor EC was significant autologous tumor EC lysis obtained (mean 14%; p<0.05). Costimulation by anti-CD3 (mean 21%; p<0.05) or interleukin-2 (mean 31%; p<0.05) further increased tumor EC lysis significantly. CONCLUSIONS: Increased turnover of T lymphocytes in normal, tumor-distant, renal tissue was associated with a higher yield of pre-CTL which can be transformed into a functionally active effector T-cell pool by stimulation via antigen plus APC. Thus, tumor-distant renal tissue has to be included in the tissue-sampling procedure for adoptive immunotherapy.
Antigen-Presenting Cells↗
Treating epilepsy in the elderly: safety considerations.
The incidence of epilepsy increases with advancing age. Epilepsy in the elderly has different aetiologies from that in younger populations, cerebrovascular disease being the most common condition associated with seizures. Partial seizures are the predominant seizure type in older patients. A diagnosis of epilepsy in the elderly is based mainly on the history and is frequently delayed. In addition, seizure imitators are especially frequent. In many cases ancillary tests for diagnosis may show normal age-related variants, sometimes making results difficult to interpret. Treating epilepsy in the elderly is problematic due to a number of issues that relate to age and comorbidity. The physical changes associated with increasing age frequently lead to changes in the pharmacokinetics of many anticonvulsants. The treatment of epilepsy in the elderly is also complicated by the existence of other diseases that might affect the metabolism or excretion of anticonvulsants and the presence of concomitant medications that might interact with them. Moreover, specific trials of anticonvulsants in the aged population are scarce. General guidelines for treatment include starting at lower doses, slowing the titration schedule, individualising the choice of anticonvulsant to the characteristics of the patient, avoiding anticonvulsants with important cognitive or sedative adverse effects, and where possible, treating with monotherapy.
Aged↗
Strong coupling constant from scaling violations in fragmentation functions.
We present a new determination of the strong coupling constant alpha(s) through the scaling violations in the fragmentation functions for charged pions, charged kaons, and protons. In our fit we include the latest e+e- annihilation data from CERN LEP1 and SLAC SLC on the Z-boson resonance and older, yet very precise, data from SLAC PEP at center-of-mass energy sqaure root of s = 29 GeV. At next-to-leading order, we find alpha(s)(5)(M(Z)) = 0.1170+/-0.0073. A new world average of alpha(s) is given.
Journal Article↗
Expression of different coding sequences in cell-free bacterial and eukaryotic systems indicates translational pausing on Escherichia coli ribosomes.
Five different coding sequences of bacterial or eukaryotic origin in plasmids under the T7 promoter were expressed in a cell-free system derived from Escherichia coli. Translation on E. coli ribosomes resulted in a full-length product only in four of the five coding sequences tested. A unique pattern of less than full-length polypeptides was generated in each case. Many of these polypeptides on E. coli ribosomes reacted with a puromycin derivative, cytidylic acid-puromycin, which was radioactively labeled. Thus these incomplete polypeptides can be defined as nascent peptides bound to the ribosomal P site. Certain nascent peptides could be shifted into full-length protein indicating that they resulted from translational pausing. In contrast to these results, expression of the same coding sequences in a wheat germ or reticulocyte cell-free system resulted in a 80-90% full-length product with no evidence for nascent polypeptides and translational pausing.
Cell-Free System↗
Automated data acquisition by confocal laser scanning microscopy and image analysis of triple stained immunofluorescent leukocytes in tissue.
The aim of the study was to develop a novel system permitting automated analysis of multicolor immunofluorescence-staining of cells in solid tissues which would be comparable to the analytical capacity of flow cytometry. In the user friendly automated data acquisition and image processing system which was established, the software includes a set of pre-defined processing steps for improved object identification and can be either interactive or fully automatic. As with multi color flow cytometry, stained cells can be analyzed in a fully automated manner regardless of tissue type. The software organizes computerized sample movement, autofocus, laser readjustment, data capture and storage as well as calculation. Data are presented as histograms indicating the staining intensity and frequency of each antigen, and in dotplots with each channel plotted against the other. The calculated statistics give information about how many of the cells are single-, double- or triple-reactive and how intensely they react with the respective antibodies. Comparison of data generated by this automated fluorescence confocal laser scanning microscopy (AF-CLSM) with flow cytometry using triple stained peripheral blood lymphocytes revealed a highly significant correlation between the methods (P<0.001). A correlation was also observed when sections triple stained for anti-CD45, -CD3 and -CD8 were analyzed by AF-CLSM and the data were compared to visual/manual cell counting (P<0.01). The AF-CLSM system permits for the first time, fast (online) and reproducible analysis of immunofluorescence staining of an unlimited number of cells in tissue sections. The software, including a manual, is available for a small fee to cover the costs of printing and postage.
CD3 Complex↗
Fluorophores at the N terminus of nascent chloramphenicol acetyltransferase peptides affect translation and movement through the ribosome.
Structurally different fluorescent probes were covalently attached to methionyl-tRNA(f) and tested for their incorporation into nascent peptides and full-length protein using an Escherichia coli cell-free coupled transcription/translation system. Bovine rhodanese and bacterial chloramphenicol acetyltransferase (CAT) were synthesized using derivatives of cascade yellow, eosin, pyrene, or coumarin attached to [(35)S]Met-tRNA(f). All of the probes tested were incorporated into polypeptides, although less efficiently when compared with formyl-methionine. Eosin, the largest of the fluorophores used with estimated dimensions of 20 x 11 A, caused the largest reduction in product formed. The rate of initiation was reduced with the fluorophore-Met-tRNA(f) compared with fMet-tRNA(f) with pyrene having the least and eosin the biggest effect. Analysis of the nascent polypeptides showed that the modifications at the N terminus affected the rate at which nascent CAT peptides were elongated causing accumulation of peptides of about 4 kDa, possibly by steric hindrance inside the tunnel within the 50 S ribosomal subunit. Fluorescence measurements indicate that the probe at the N terminus of nascent pyrene-CAT peptides is in a relatively hydrophilic environment. This finding is in agreement with recent data showing cross-linking of the N terminus of nascent peptides to nucleotides of the 23 S ribosomal RNA.
Animals↗