Prevalence of urinary tract cancer in the Spanish cohort of the IDENTIFY study.

INTRODUCTION: Malignant tumors of the urinary tract are associated with high morbidity and mortality, and their prevalence can vary worldwide. Recently, the IDENTIFY study has published results on the prevalence of urinary tract cancer at a global level. This study evaluates the prevalence of cancer within the Spanish cohort of the IDENTIFY study to determine whether the published results can be extrapolated to our population. PATIENTS AND METHODS: An analysis of the data from the Spanish cohort of patients in the IDENTIFY study was performed. This is a prospective cohort of patients referred to secondary care with suspected cancer, predominantly due to hematuria. Patients were recruited between December 2017 and December 2018. RESULTS: A total of 706 patients from 9 Spanish centers were analyzed. Of these, 277 (39.2%) were diagnosed with cancer: 259 (36.7%) bladder cancer, 10 (1.4%) upper tract urothelial carcinoma, 9 (1.2%) renal cancer and 5 (0.7%) prostate cancer. Increasing age (OR 1.05 (95% CI 1.03-1.06; P < 0.001)), visible hematuria (VH) OR 2.19 (95% CI 1.13-4.24; P = 0.02)) and smoking (ex-smokers: OR 2.11(95% CI 1.30-3.40; P = 0.002); smokers: OR 2.36 (95% CI 1.40-3.95; P = 0.001)) were associated with higher probability of bladder cancer. CONCLUSION: This study highlights the risk of bladder cancer in patients with VH and smoking habits. Bladder cancer presented the highest prevalence; higher than the prevalence reported in previous series and presented in the IDENTIFY study. Future work should evaluate other associated factors that allow us to create cancer prediction models to improve the detection of cancer in our patients.

Interpersonal psychotherapy group (IPT-G) for depression.

A case study of a time-limited interpersonal psychotherapy group (IPT-G) is presented to illustrate the use of interpersonal therapy (IPT) to treat patients with major depression in a group psychotherapy format. The use of individual outcome measures as a helpful adjunct to clinical psychotherapeutic practice is demonstrated. Because IPT-G has only a few exclusion criteria (active suicidality and significant borderline personality features), it can be used in a broad range of clinical settings. This clinical example demonstrates IPT-G to be a useful modality for addressing a common and difficult patient population.

Modulation of glycophorin A transmembrane helix interactions by lipid bilayers: molecular dynamics calculations.

Starting from the glycophorin A dimer structure determined by NMR, we performed simulations of both dimer and monomer forms in explicit lipid bilayers with constant normal pressure, lateral area, and temperature using the CHARMM potential. Analysis of the trajectories in four different lipids reveals how lipid chain length and saturation modulate the structural and energetic properties of transmembrane helices. Helix tilt, helix-helix crossing angle, and helix accessible volume depend on lipid type in a manner consistent with hydrophobic matching concepts: the most relevant lipid property appears to be the bilayer thickness. Although the net helix-helix interaction enthalpy is strongly attractive, analysis of residue-residue interactions reveals significant unfavorable electrostatic repulsion between interfacial glycine residues previously shown to be critical for dimerization. Peptide volume is nearly conserved upon dimerization in all lipid types, indicating that the monomeric helices pack equally well with lipid as dimer helices do with one another. Enthalpy calculations indicate that the helix-environment interaction energy is lower in the dimer than in the monomer form, when solvated by unsaturated lipids. In all lipid environments there is a marked preference for lipids to interact with peptide predominantly through one rather than both acyl chains. Although our trajectories are not long enough to allow a full thermodynamic treatment, these results demonstrate that molecular dynamics simulations are a powerful method for investigating the protein-protein, protein-lipid, and lipid-lipid interactions that determine the structure, stability and dynamics of transmembrane alpha-helices in membranes.

Structure-based prediction of the stability of transmembrane helix-helix interactions: the sequence dependence of glycophorin A dimerization.

The ability to predict the effects of point mutations on the interaction of alpha-helices within membranes would represent a significant step toward understanding the folding and stability of membrane proteins. We use structure-based empirical parameters representing steric clashes, favorable van der Waals interactions, and restrictions of side-chain rotamer freedom to explain the relative dimerization propensities of 105 hydrophobic single-point mutants of the glycophorin A (GpA) transmembrane domain. Although the structure at the dimer interface is critical to our model, changes in side-chain hydrophobicity are uncorrelated with dimer stability, indicating that the hydrophobic effect does not influence transmembrane helix-helix association. Our model provides insights into the compensatory effects of multiple mutations and shows that helix-helix interactions dominate the formation of specific structures.

A transmembrane helix dimer: structure and implications.

The three-dimensional structure of the dimeric transmembrane domain of glycophorin A (GpA) was determined by solution nuclear magnetic resonance spectroscopy of a 40-residue peptide solubilized in aqueous detergent micelles. The GpA membrane-spanning alpha helices cross at an angle of -40 degrees and form a small but well-packed interface that lacks intermonomer hydrogen bonds. The structure provides an explanation for the previously characterized sequence dependence of GpA dimerization and demonstrates that van der Waals interactions alone can mediate stable and specific associations between transmembrane helices.

Leucine side-chain rotamers in a glycophorin A transmembrane peptide as revealed by three-bond carbon-carbon couplings and 13C chemical shifts.

We have used a spin-echo difference NMR pulse sequence to measure three-bond J couplings between lambda- and alpha-carbons of the leucine residues in a micelle-associated helical peptide dimer that corresponds to residues 62-101 of the transmembrane erythrocyte protein glycophorin A. The observed 3J couplings correlate strongly with the 13C chemical shift of the lambda-methyl groups, and within experimental error both the shift distribution of the methyl carbons and the variations in 3J can be accounted for by variations in side-chain rotamer populations. We infer that all leucine side chains in this peptide dimer are in fast exchange among chi 2 rotamers and sample two of the three possible rotameric states, even when the side chain forms part of the dimer interface. The observed correlation of chemical shift with couplings can be traced to a gamma-gauche interaction of methyl and alpha-carbons. This correlation may provide an alternate route to rotamer analysis in some protein systems.

Mapping the lipid-exposed surfaces of membrane proteins.

Phospholamban forms a stable complex of five long transmembrane helices. We show that the relative rotational orientation of the helices in the pentameric complex can be distinguished by S-H to S-D exchange of cysteine sulphydryl groups located in the transmembrane segment of the protein and exposed to the lipid environment. Of the three cysteine residues in phospholamban, two residues (Cys 36 and Cys 46) are oriented towards the helix interface and protected from exchange, while the third cysteine (Cys 41) is oriented towards the lipid interface and undergoes exchange with water diffused into the bilayer. Distinguishing the external and internal faces of a membrane protein by sulphydryl exchange provides a general approach for determining the three-dimensional fold of membrane proteins and enhances model building efforts to generate high-resolution structures.

Time-limited group psychotherapy.

The presence of a time limit shapes the entire therapeutic process and culminates in the management of termination. This paper outlines a protocol for dealing with termination in time-limited psychotherapy groups: (1) Select suitable members with similar capacity to use a time-limited format; (2) use a closed group format; (3) clarify the time boundaries at the beginning; (4) reinforce the time frame during the last half of the group; (5) provide focused interventions; (6) forestall premature termination; (7) reinforce termination themes of deprivation, resentment and anger, rejection, grief and loss, responsibility for self; (8) structure the final session; and (9) plan a 4-month follow-up visit to encourage ongoing application. These guidelines provide the therapist with a general structure to ensure that major termination themes are systematically addressed. They deal with powerful concepts of maturation and self-responsibility and incorporate the principal strategies that distinguish time-limited from time-unlimited group psychotherapy.

Self-disclosure, feedback, and outcome in long-term inpatient psychotherapy groups.

Process and outcome measures were compared in two long-term psychotherapy groups. Measures of Self-Disclosure and Feedback were based on analysis of videotaped sessions by use of Bales's SYMLOG three-axial system. These measures were then combined to form a measure of Interpersonal Work. High Interpersonal Work scores, both in early sessions and throughout treatment, predicted better outcome for individual members at 18-month follow-up. Interpersonal Work scores in later group sessions were not related to outcome. Findings suggest that early sessions are a critical time for members to develop a working style in the group. Members who fail to do so have a less successful outcome even if their working styles improve in later sessions.

Glycoconjugates of the human trabecular meshwork: a lectin histochemical study.

Twelve specimens of resin-embedded human trabecular meshwork were probed with a panel of 21 biotinylated lectins, using an avidin-biotin peroxidase revealing system, in order to determine the normal pattern of saccharide expression in this tissue. High-mannose, intermediate and hybrid N-linked glycans, and complex N-linked bisected and non-bisected bi/tri-antennate glycans, as shown by the binding of Canavalia ensiformis (ConA), Pisum sativum (PSA), Lens culinaris (LCA) agglutinins and Phaseolus vulgaris erythroagglutinin (ePHA), were strongly expressed by the canal of Schlemm endothelium and juxtacanalicular tissue, but less so by the corneoscleal meshwork. Highly branced complex glycans were not found, as there was no binding by Phaseolus vulgaris leukoagglutinin (IPHA). Sialyl residues, especially those alpha 2,6-linked as demonstrated by strong Sambucus nigra (SNA) lectin staining, were also abundant in this area. N-acetyllactosamine sequences and some O-linked glycans were present in the trabecular meshwork, as shown by Solanum tuberosum (STA), Datura stramonium (DSA), and Jacalin (Jac) lectin binding, while fucose residues were not detected by Tetragonolobus purpureas (LTA) or Ulex europaeus-1 (UEA-1) agglutinins. These results indicate similarities with renal glomerular and vascular endothelium, although the lack of binding with UEA-1 agglutinin suggests differences which may relate to the specialized function of the trabecular meshwork. This study provides a baseline for comparative analysis of the glycans of human trabecular meshwork in pathological conditions such as primary open-angle glaucoma.

Overexpression, purification, and characterization of Escherichia coli acyl carrier protein and two mutant proteins.

A synthetic gene of 237 bases encoding the 77-residue acyl carrier protein (ACP) from Escherichia coli, along with two mutant genes, ACP-I54V and ACP-A59V, were subcloned into the pET11a-pLysS E. coli overexpression system under the control of the bacteriophage T7 promoter. This efficient expression system and a simplified purification protocol yielded more than 120 mg/l of pure protein. The construct produced a mixture of holo-ACP and apo-ACP and two HPLC procedures were developed to separate the two species. This overexpression system allows cost-effective growths of 13C- and 15N-labeled protein for structural and other studies on ACP. In the course of the work on the mutants of ACP, an apparent homologous recombination event led, in one case, to reversion to a wild-type protein, suggesting that precautions to prevent such reversion should be taken.

Structural organization of the pentameric transmembrane alpha-helices of phospholamban, a cardiac ion channel.

Phospholamban is a 52 amino acid calcium regulatory protein found as pentamers in cardiac SR membranes. The pentamers form through interactions between its transmembrane domains, and are stable in SDS. We have employed a saturation mutagenesis approach to study the detailed interactions between the transmembrane segments, using a chimeric protein construct in which staphylococcal nuclease (a monomeric soluble protein) is fused to the N-terminus of phospholamban. The chimera forms pentamers observable in SDS-PAGE, allowing the effects of mutations upon the oligomeric association to be determined by electrophoresis. The disruptive effects of amino acid substitutions in the transmembrane domain were classified as sensitive, moderately sensitive or insensitive. Residues of the same class lined up on faces of a 3.5 amino acids/turn helical projection, allowing the construction of a model of the interacting surfaces in which the helices are associated in a left-handed pentameric coiled-coil configuration. Molecular modeling simulations (to be described elsewhere in detail) confirm that the helices readily form a left-handed coiled-coil helical bundle and have yielded molecular models for the interacting surfaces, the best of which is identical to that predicted by the mutagenesis. Residues lining the pore show considerable structural sensitivity to mutation, indicating that care must be taken in interpreting the results of mutagenesis studies of channels. The cylindrical ion pore (minimal diameter of 2 A) appears to be defined largely by hydrophobic residues (I40, L43 and I47) with only two mildly polar elements contributed by sulfurs in residues C36 and M50.

Where is here and when is now? The adaptational challenge of mental health reform for group psychotherapy.

The structure of health care delivery is in a period of rapid change. An understanding of psychotherapy service use patterns indicates the importance of time categories as well as the relevance of group techniques. Traditional interpersonal and psychodynamic group techniques may be modified for time-limited use without sacrificing basic values and with demonstrated effectiveness. Adapting to a more fiscally stringent practice environment requires the clinician to undertake a difficult process of transition. As professionals our task is to prepare ourselves for this challenge and to let the health care system know that proper practice guidelines are necessary to achieve effective treatment for our patients.

Localisation of alpha(2,3) and alpha(2,6) linked terminal sialic acid groups in human trabecular meshwork.

Sialic acid specific lectins were used to localise isomers of sialyl glycosides in human trabecular meshwork (TM) at the ultrastructural level. A lectin immunogold method demonstrated that sialic groups were concentrated on the endothelial surface of Schlemm's canal (SC) and in the adjacent juxta-canalicular tissue (JCT). One sialyl glycoside, alpha(2,6) linked N-acetyl neuraminic acid, was present mainly on the luminal aspect of the SC endothelium and in the cytoplasm of the JCT cells. Another, alpha(2,3) linked N-acetyl neuraminic acid, was localised predominantly to the extracellular fibrillar material of the JCT. The existence of a topographical segregation of these two sialyl glycosides within the TM supports the view that highly charged anionic molecules may be of significance in regulating aqueous outflow.