Antidepressants: general practitioners' opinions and clinical practice.

OBJECTIVE: To describe and compare general practitioners' (GPs) opinions on antidepressant drugs and their prescriptions to depressed patients. METHOD: Between November 2000 and July 2001 a representative sample of French GPs was asked their opinion of the 15 most prescribed antidepressants, and then to describe the treatments of the current depressive episode of four depressive patients each, their changes and the reasons thereof. RESULTS: One hundred and eighty-one GPs and 778 patients participated. The best-ranked antidepressants by the GPs were paroxetine, fluoxetine, sertraline and clomipramine for efficacy, and paroxetine, tianeptine, sertraline and fluoxetine for tolerability. In patients, the drugs most often prescribed were fluoxetine, paroxetine, and sertraline. Those least often stopped for intolerance were moclobemide (0%), dosulepine (0%), venlafaxine (4.5%) and citalopram (5.0%), and maprotiline (0%), citalopram (1.7%) and venlafaxine (2.3%) for lack of efficacy. The best predictor for prescription of antidepressants was the GPs' overall ranking, itself depending on opinions of the tolerability and efficacy of the drug. However, opinions of tolerability and efficacy were not related to the rates of treatment discontinuation for intolerability or inefficacy. CONCLUSION: Prescriber opinion does not seem related to actual product performance.

Diagnostic depressive symptoms of the mixed bipolar episode.

BACKGROUND: There is not yet consensus on the best diagnostic definition of mixed bipolar episodes. Many have suggested the DSM-III-R/-IV definition is too rigid. We propose alternative criteria using data from a large patient cohort. METHODS: We evaluated 237 manic in-patients using DSM-III-R criteria and the Scale for Manic States (SMS). A bimodally distributed factor of dysphoric mood has been reported from the SMS data. We used both the factor and the DSM-III-R classifications to identify candidate depressive symptoms and then developed three candidate depressive symptom sets. Using ROC analysis we determined the optimal threshold number of symptoms in each set and compared the three ROC solutions. The optimal solution was tested against the DSM-III-R classification for crossvalidation. RESULTS: The optimal ROC solution was a set, derived from both the DSM-III-R and the SMS, and the optimal threshold for diagnosis was two or more symptoms. Applying this set iteratively to the DSM-III-R classification produced the identical ROC solution. The prevalence of mixed episodes in the cohort was 13.9% by DSM-III-R, 20.2% by the dysphoria factor and 27.4% by the new ROC solution. CONCLUSIONS: A diagnostic set of six dysphoric symptoms (depressed mood, anhedonia, guilt, suicide, fatigue and anxiety), with a threshold of two symptoms, is proposed for a mixed episode. This new definition has a foundation in clinical data, in the proved diagnostic performance of the qualifying symptoms, and in ROC validation against two previous definitions that each have face validity.

Cu,Zn superoxide dismutase structure from a microbial pathogen establishes a class with a conserved dimer interface.

Macrophages and neutrophils protect animals from microbial infection in part by issuing a burst of toxic superoxide radicals when challenged. To counteract this onslaught, many Gram-negative bacterial pathogens possess periplasmic Cu,Zn superoxide dismutases (SODs), which act on superoxide to yield molecular oxygen and hydrogen peroxide. We have solved the X-ray crystal structure of the Cu,Zn SOD from Actinobacillus pleuropneumoniae, a major porcine pathogen, by molecular replacement at 1.9 A resolution. The structure reveals that the dimeric bacterial enzymes form a structurally homologous class defined by a water-mediated dimer interface, and share with all Cu,Zn SODs the Greek-key beta-barrel subunit fold with copper and zinc ions located at the base of a deep loop-enclosed active-site channel. Our structure-based sequence alignment of the bacterial enzymes explains the monomeric nature of at least two of these, and suggests that there may be at least one additional structural class for the bacterial SODs. Two metal-mediated crystal contacts yielded our C222(1) crystals, and the geometry of these sites could be engineered into proteins recalcitrant to crystallization in their native form. This work highlights structural differences between eukaryotic and prokaryotic Cu,Zn SODs, as well as similarities and differences among prokaryotic SODs, and lays the groundwork for development of antimicrobial drugs that specifically target periplasmic Cu,Zn SODs of bacterial pathogens.

Crystallographic structure reveals phosphorylated pilin from Neisseria: phosphoserine sites modify type IV pilus surface chemistry and fibre morphology.

Understanding the structural biology of type IV pili, fibres responsible for the virulent attachment and motility of numerous bacterial pathogens, requires a detailed understanding of the three-dimensional structure and chemistry of the constituent pilin subunit. X-ray crystallographic refinement of Neisseria gonorrhoeae pilin against diffraction data to 2.6 A resolution, coupled with mass spectrometry of peptide fragments, reveals phosphoserine at residue 68. Phosphoserine is exposed on the surface of the modelled type IV pilus at the interface of neighbouring pilin molecules. The site-specific mutation of serine 68 to alanine showed that the loss of the phosphorylation alters the morphology of fibres examined by electron microscopy without a notable effect on adhesion, transformation, piliation or twitching motility. The structural and chemical characterization of protein phosphoserine in type IV pilin subunits is an important indication that this modification, key to numerous regulatory aspects of eukaryotic cell biology, exists in the virulence factor proteins of bacterial pathogens. These O-linked phosphate modifications, unusual in prokaryotes, thus merit study for possible roles in pilus biogenesis and modulation of pilin chemistry for optimal in vivo function.

Signs and symptoms of mania in pure and mixed episodes.

BACKGROUND: Debate continues about the diagnosis of mixed mania and the restrictiveness of the DSM-III-R and DSM-IV criteria for Bipolar Disorder, mixed. Although awareness of dysphoric features during mania continues to grow, standard mania rating instruments do not adequately assess mixed states and there is a striking disparity between the dysphoric signs and symptoms emphasized in research studies and the commonly employed DSM criteria. METHODS: Three hundred sixteen inpatients meeting DSM-III-R criteria for Bipolar Disorder, manic or mixed, were evaluated by rating 20 signs and symptoms. The frequencies of these signs and symptoms were computed for both diagnostic subtypes and compared using chi2 statistics and conditional probability parameters. RESULTS: The most frequently noted signs and symptoms in mania are motor activation, accelerated thought process, pressured speech and decreased sleep. Although euphoric mood was present in a large portion of the cohort, irritability, dysphoric mood and mood lability were also prominent in the entire cohort. Dysphoric mood, mood lability, anxiety, guilt, suicidality, and irritability were the only symptoms significantly more common in the mixed group. In contrast, grandiosity, euphoric mood, and pressured speech were significantly more often observed in the pure manic group. Contrary to popular belief, paranoia did not differ significantly between the two groups. Suicidality was present in a non-trivial 7% of the entire cohort, including some subjects who did not meet the criteria for mixed mania. LIMITATIONS: The comparison of mixed and manic episodes requires the appropriate definition of mixed states. In the current report we use the DSM-III-R definition of Bipolar Disorder, mixed, which may be too rigid. CONCLUSIONS: The data underscore that mania is not a purely euphoric state. Substantial rates of dysphoria, lability, anxiety and irritability were noted in the "pure" manic patients, as well as in those who meet the full DSM criteria for Bipolar Disorder, mixed, suggesting, that perhaps a less restrictive definition of mixed states would be more appropriate.

Assemblies of replication initiator protein on symmetric and asymmetric DNA sequences depend on multiple protein oligomerization surfaces.

The pi35.0 protein of plasmid R6K regulates transcription and replication by binding a DNA sequence motif (TGAGR) arranged either asymmetrically into 22 bp direct repeats (DRs) in the gamma origin, or symmetrically into inverted half-repeats (IRs) in the operator of its own gene, pir. The binding patterns of the two natural forms of the pi protein and their heterodimers revealed that the predominant species, pi35.0 (35.0 kDa), can bind to a single copy of the DR as either a monomer or a dimer while pi30.5 (30.5 kDa) binds only as a dimer. We demonstrate that only one subunit of a pi35.0 dimer makes specific contact with DNA. Electron microscopic (EM) analysis of the nucleoprotein complexes formed by pi35.0 and DNA fragments containing all seven DRs revealed coupled ("hand-cuffed") DNA molecules that are aligned in a parallel orientation. Antiparallel orientations of the DNA were not observed. Thus, hand-cuffing depends on a highly ordered oligomerization of pi35.0 in such structures. The pi protein (pi35.0, pi30.5) binds to an IR as a dimer or heterodimer but not as a monomer. Moreover, a single amino acid residue substitution, F200S (pir200), introduced into pi30.5 severely destabilizes dimers of this protein in solution and concomitantly prevents binding of this protein to the IR. This mutation also changes the stability of pi35.0 dimers but it does not change the ability of pi35.0 to bind IRs. To explain these observations we propose that the diverse interactions of pi variants with DNA are controlled by multiple surfaces for protein oligomerization.

Consequences of the loss of O-linked glycosylation of meningococcal type IV pilin on piliation and pilus-mediated adhesion.

Pili, which are assembled from protein subunits called pilin, are indispensable for the adhesion of capsulated Neisseria meningitidis (MC) to eukaryotic cells. Both MC and Neisseria gonorrhoeae (GC) pilins are glycosylated, but the effect of this modification is unknown. In GC, a galactose alpha-1,3-N-acetyl glucosamine is O-linked to Ser-63, whereas in MC, an O-linked trisaccharide is present between residues 45 and 73 of pilin. As Ser-63 was found to be conserved in pilin variants from different strains, it was replaced by Ala in two MC variants to test the possible role of this residue in pilin glycosylation and modulation of pili function. The mutated alleles were stably expressed in MC, and the proteins they encoded migrated more quickly than the normal protein during SDS-PAGE. As controls, neighbouring Asn-61 and Ser-62 were replaced by an Ala with no effect on electrophoretic mobility. Silver staining of purified pilin obtained from MC after oxidation with periodic acid confirmed the loss of glycosylation in the Ser-63-->Ala pilin variants. Mass spectrometry of HPLC-purified trypsin-digested peptides of pilin and Ser-63-->Ala pilin confirmed that peptide 45-73 has the molecular size of a glycopeptide in the wild type. In strains producing non-glycosylated pilin variants, we observed that (i) no truncated S pilin monomer was produced; (ii) piliation was slightly increased; and (iii) presumably as a consequence, adhesiveness for epithelial cells was increased 1.6- to twofold in these derivatives. In addition, pilin monomers and/or individual pilus fibres, obtained after solubilization of a crude pili preparation in a high pH buffer, were reassociated into insoluble aggregates of pili more completely with non-glycosylated variants than with the normal pilin. Taken together, these data eliminate a major role for pilin glycosylation in piliation and subsequent pilus-mediated adhesion, but they demonstrate that glycosylation facilitates solubilization of pilin monomers and/or individual pilus fibres.

A factor analysis of the signs and symptoms of mania.

BACKGROUND: No adequate factor analyses of signs and symptoms of mania have been reported. From limited past reports, the view has arisen that 2 main symptom clusters (euphoric-grandiose and paranoid-destructive) occur in patients with mania, along with so-called core symptoms of psychomotor pressure. In this view, dysphoric mania is associated with paranoid-destructive symptoms and with psychosis. METHODS: We rated 237 patients with DSM-III-R-defined bipolar disorder, manic (n = 204) or mixed (n = 33), on 15 classic features of mania and 5 features related to dysphoric mood. Principal components factor analysis was applied to the ratings. RESULTS: Five clearly interpretable and clinically relevant factors were identified. The first and strongest factor represented dysphoria in mania, with strong positive loadings for depressed mood, lability, guilt, anxiety, and suicidal thoughts and behaviors and a strong negative loading for euphoric mood. Factors 2 through 5 represented psychomotor acceleration, psychosis, increased hedonic function, and irritable aggression, respectively. The distribution of weighted scores on factor 1 was bimodal, whereas the corresponding distributions of factors 2 through 5 were unimodal. Contrary to all past reports, no general factor denoting overall severity of mania was found. Factors previously proposed by Beigel and Murphy were not confirmed. CONCLUSIONS: Five independent factors representing dysphoric mood, psychomotor pressure, psychosis, increased hedonic function, and irritable aggression were identified. The conventional view of symptom factors in mania was not confirmed. Dysphoric features are statistically salient in patients with mania, and the bimodal distribution of the dysphoria factor is consistent with the possibility that mixed bipolar disorder is a distinct state.

The performance of DSM-III-R major depression criteria in the diagnosis of bipolar mixed states.

Two hundred thirty-seven (237) manic patients diagnosed by DSM-III-R criteria as either purely manic (204) or mixed bipolar (33) were reviewed for analysis of the diagnostic performance of the DSM-III-R criteria required to diagnose the mixed bipolar state. We calculated the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and diagnostic efficiency of each of the 9 DSM-III-R criteria for major depression in this cohort. As predicted, four of the major depression criteria had low diagnostic utility, with PPV's less than 0.3. Those items were: weight change; sleep disturbance; psychomotor change; and diminished ability to think or concentrate or indecisiveness. Four symptoms: anhedonia, fatigue, feelings of worthlessness or guilt, and recurrent thoughts of death or suicidal ideation had acceptable utility for the diagnosis of mixed states.

Type-4 pilus-structure: outside to inside and top to bottom--a minireview.

We have recently proposed a computational model of the N. gonorrhoeae pilus fiber based on the high resolution X-ray crystal structure of the component protein pilin, combined with available biophysical and genetic data [Parge et al. (1995) Nature 378, 32-38]. In parallel, we have used anti-peptide antibodies to distinguish buried and exposed regions of pilin within the assembled fiber [Forest et al. (1996) Infect. Immun. 64, 644-652]. This mini-review addresses the properties of the current pilus model and the locations of end-exposed epitopes. The fiber forms a three-layered structure of coiled conserved alpha helices surrounded by beta-sheet, with the hypervariable region as the most highly exposed portion. Overall the pilus model developed from diffraction and antibody mapping is expected to be representative of type-4 pili with general implications for type-4 assembly, function, and interactions with other proteins and cell membranes.

Assembly and antigenicity of the Neisseria gonorrhoeae pilus mapped with antibodies.

The relationship between the sequence of Neisseria gonorrhoeae pilin and its quaternary assembly into pilus fibers was studied with a set of site-directed antibody probes and by mapping the specificities of antipilus antisera with peptides. Buried and exposed peptides in assembled pili were identified by competitive immunoassays and immunoelectron microscopy with polyclonal antibodies raised against 11 peptides spanning the pilin sequence. Pili did not compete significantly with pilin subunits for binding to antibodies against residues 13 to 31 (13-31) and 18-36. Pilus fibers competed well with pilin protein subunits for binding to antibodies raised against peptides 37-56, 58-78, 110-120, 115-127, 122-139, and 140-159 and competed weakly for antibodies against residues 79-93 and 94-108. Antibodies to sequence-conserved residues 37-56 and to semiconserved residues 94-108 preferentially bound pilus ends as shown by immunoelectron microscopy. The exposure of pilus regions to the immune system was tested by peptide mapping of antiserum specificities against sets of overlapping peptides representing all possible hexameric or octameric peptides from the N. gonorrhoeae MS11 pilin sequence. The immunogenicity of exposed peptides incorporating semiconserved residues 49-56 and 121-126 was revealed by strong, consistent antigenic reactivity to these regions measured in antipilus sera from rabbits, mice, and human and in sera from human volunteers with gonorrhea. The conservation and variation of antigenic responses among these three species clarify the relevance of immunological studies of other species to the human immune response against pathogens. Overall, our results explain the extreme conservation of the entire N-terminal one-third of the pilin protein by its dominant role in pilus assembly: hydrophobic residues 1-36 are implicated in buried lateral contacts, and polar residues 37-56 are implicated in longitudinal contacts within the pilus fiber.

Family policies for an aging society: moving to the twenty-first century.

Most family policy implicitly or explicitly focuses on families with young children, but the revolution in longevity suggests the value of a life course focus, aimed at promoting the effectiveness of families and individuals at all ages and stages. Gerontologists can make a contribution by documenting and describing the gaps between needs and resources of families at all life stages, developing family indicators of social change, and sensitizing both decision makers and the public to the unintended consequences of existing or proposed policies.

Structure of the fibre-forming protein pilin at 2.6 A resolution.

The crystallographic structure of Neisseria gonorrhoeae pilin, which assembles into the multifunctional pilus adhesion and virulence factor, reveals an alpha-beta roll fold with a striking 85 A alpha-helical spine and an O-linked disaccharide. Key residues stabilize interactions that allow sequence hypervariability, responsible for pilin's celebrated antigenic variation, within disulphide region beta-strands and connections. Pilin surface shape, hydrophobicity and sequence variation constrain pilus assembly to the packing of flat subunit faces against alpha 1 helices. Helical fibre assembly is postulated to form a core of coiled alpha 1 helices banded by beta-sheet, leaving carbohydrate and hypervariable sequence regions exposed to solvent.

Isomorphous binding of mercury-substituted thiosaccharides to pertussis toxin crystals yields crystallographic phases.

An isomorphous derivative of pertussis toxin crystals was prepared using a 2-alpha-mercuric analog of N-acetyl neuraminic acid in a method analogous to the use of inhibitors labelled with heavy atoms to solve crystal structures of enzymes. This derivative exploits the specific binding between pertussis toxin and terminal sialic acid residues on receptor glycoproteins. Difference Patterson maps yielded heavy-atom sites which refined with good statistics, indicating that the protein probably does not undergo a conformational change on receptor binding. Mercuric analogs of other monosaccharides should be easily obtainable using the same synthetic strategy, suggesting a general method for derivatizing crystals of carbohydrate-binding proteins.