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PubMed1998

Photoprotection: sunscreens and the immunomodulatory effects of UV irradiation.

UV-B irradiation (UVR) of the host, in both humans and animal models, induces dose-related acute and chronic changes in skin which include erythema and photoageing, and induction of cancer. It can also induce modulation of immune responses of the host to antigens presented following irradiation. Commercially-available, broad-spectrum, high SPF (15, 15 + ) sunscreens protect against most effects of UV irradiation. An exception is the effects of UVR on immune responsiveness, with varying degrees of protection having been reported. We examined a system of UV-induced systemic suppression of contact hypersensitivity (CHS) responses in BALB/c mice. A range of commercially-available, broad spectrum, high SPF (15 + ) sunscreens demonstrated at best partial protection against systemic immunosuppression, yet were able to protect against two hallmarks of acute UVR-induced damage: skin oedema and keratinocyte proliferation. Two major models have been identified for the induction of immunosuppression following UVR, one identifying trans-urocanic acid (trans-UCA; deaminated histidine, located in the stratum corneum) as the critical photoreceptor, the other featuring DNA. UVR of trans-UCA produces cis-UCA, which itself is immunomodulatory. There was some abrogation of trans to cis isomerisation of urocanic acid in UV-irradiated, sunscreen-protected mice. However, the majority of the immunomodulation seen in these mice was abrogated by pretreatment with a monoclonal antibody to cis-urocanic acid. It is possible to induce formation of cis-urocanic acid in BALB/c skin in the absence of immunosuppression, using lower doses of UV radiation, indicating that formation of cis-urocanic acid in the stratum corneum is not necessarily sufficient to induce immunosuppression in the UV-irradiated host. The mechanisms of induction of the immunomodulated state in the UV-irradiated host are potentially diverse and the subject of ongoing debate. Our studies maintain a role for cis-UCA, and form the basis for further studies on its involvement in immunomodulation by UVR in sunscreen-protected hosts.

Animals↗
PubMed2001

Probing catalysis by Escherichia coli dTDP-glucose-4,6-dehydratase: identification and preliminary characterization of functional amino acid residues at the active site.

A model of the Escherichia coli dTDP-glucose-4,6-dehydratase (4,6-dehydratase) active site has been generated by combining amino acid sequence alignment information with the 3-dimensional structure of UDP-galactose-4-epimerase. The active site configuration is consistent with the partially refined 3-dimensional structure of 4,6-dehydratase, which lacks substrate-nucleotide but contains NAD(+) (PDB file ). From the model, two groups of active site residues were identified. The first group consists of Asp135(DEH), Glu136(DEH), Glu198(DEH), Lys199(DEH), and Tyr301(DEH). These residues are near the substrate-pyranose binding pocket in the model, they are completely conserved in 4,6-dehydratase, and they differ from the corresponding equally well-conserved residues in 4-epimerase. The second group of residues is Cys187(DEH), Asn190(DEH), and His232(DEH), which form a motif on the re face of the cofactor nicotinamide binding pocket that resembles the catalytic triad of cysteine-proteases. The importance of both groups of residues was tested by mutagenesis and steady-state kinetic analysis. In all but one case, a decrease in catalytic efficiency of approximately 2 orders of magnitude below wild-type activity was observed. Mutagenesis of each of these residues, with the exception of Cys187(DEH), which showed near-wild-type activity, clearly has important negative consequences for catalysis. The allocation of specific functions to these residues and the absolute magnitude of these effects are obscured by the complex chemistry in this multistep mechanism. Tools will be needed to characterize each chemical step individually in order to assign loss of catalytic efficiency to specific residue functions. To this end, the effects of each of these variants on the initial dehydrogenation step were evaluated using a the substrate analogue dTDP-xylose. Additional steady-state techniques were employed in an attempt to further limit the assignment of rate limitation. The results are discussed within the context of the 4,6-dehydratase active site model and chemical mechanism.

Amino Acid Substitution↗
PubMed2006

Parent and family factors associated with child adjustment to pediatric cancer.

OBJECTIVE: To identify factors that influence the association between parent and child distress among families of children with cancer and comparison peers. METHODS: Parent and child distress, social support, and family environment were assessed among families of 95 children with cancer (94 mothers, 67 fathers) and 98 comparison peers (97 mothers, 77 fathers). RESULTS: Significant associations were found between parent and child distress. For models examining the impact of fathers' distress on children, several moderators were identified (i.e., family environment, child age and gender, a cancer diagnosis, and treatment severity). Family environment also partially mediated father and child distress. CONCLUSIONS: Children whose parents were distressed were more likely to be distressed themselves. Subgroups of children were particularly vulnerable, indicating a need to identify further mechanisms of risk and resilience and to develop family-based interventions. Support was found for including fathers as independent sources of information in pediatric psychology research and clinical practice.

Adaptation, Psychological↗
PubMed2003

Characterization of 17beta-hydroxysteroid dehydrogenase type 7 in reproductive tissues of the marmoset monkey.

In contrast to the known rodent enzymes, the physiological significance of 17beta-hydroxysteroid dehydrogenase type 7 (17HSD7) and its presumed function in reproductive biology is not well understood in primates. As a first step, we recently cloned the complete coding regions of human and marmoset monkey (Callithrix jacchus) 17HSD7 (cj17HSD7). In the present work the complete cDNA of marmoset 17HSD1 (cj17HSD1), including the proximal promoter region, and a partial sequence of marmoset aromatase (cjARO) were sequenced in order to compare the expression of these estradiol synthesizing enzymes with that of 17HSD7 in a primate model and to identify tissues where 17HSD7 might participate in the pathway of estradiol synthesis. The gene structures of cj17HSD1 and cj17HSD7 were determined and proved to be very similar to the human orthologues. Northern hybridization showed that cjARO mRNA seems to be coexpressed preferably with cj17HSD1 in placenta, whereas in other tissues it is expressed in parallel only with cj17HSD7. Especially in corpora lutea, the cj17HSD7 transcript is detectable throughout the luteal phase of the ovarian cycle and increases during pregnancy, in parallel with the transcript of aromatase. Results were confirmed by immunoblots and immunohistochemistry using new polyclonal antisera directed against cj17HSD7 and cjARO protein. The enzymatic conversion of estrone to estradiol was assessed in marmoset corpora lutea. The pattern of coexpression with aromatase supports the hypothesis that luteal 17HSD7 complements placental 17HSD1, ensuring continued estradiol synthesis throughout pregnancy in primates.

17-Hydroxysteroid Dehydrogenases↗
PubMed2002

Predictive models and the effectiveness of strategies for improving outpatient follow-up under managed care.

OBJECTIVES: This study tested the accuracy of models for predicting rehospitalization in a managed behavioral health organization and tested the effectiveness of different care management strategies for enhancing outpatient treatment follow-up. METHODS: In a controlled study, patients with an inpatient mental health or substance use admission received one of three types of care management, distinguished by the level of care managers' involvement in discharge planning and postdischarge outreach: usual (N=31), enhanced (N=94), and intensive (N=74). The groups were compared with each other and with a cohort admitted in the year before the study that received usual care management (N=192) to determine whether differences existed in time to outpatient follow-up, amount of postdischarge care, and rehospitalization at 30, 60, and 180 days. RESULTS: No differences between groups were found. The majority of patients (69 percent) received outpatient care within 30 days of discharge. Prediction models using logistic regression suggested that the number of clinical and sociodemographic risk factors identified by care managers was related to the rate of rehospitalization at 60 and 180 days. Patients authorized to receive intermediate care (partial hospitalization or residential care) and those who failed to attend intermediate care if it was authorized were more likely than other patients to be rehospitalized at 30, 60, and 180 days. CONCLUSIONS: Outpatient follow-up after psychiatric hospitalization did not improve with increasingly intensive discharge planning and outreach. Improvement in prediction of risk of rehospitalization may increase opportunities to provide intensive interventions for difficult-to-engage patients.

Adult↗
PubMed1997

Three-year coronal caries incidence in older Canadian adults.

This paper describes the incidence of coronal caries in a sample of older adults. A 3-year follow-up study was conducted of 493 community-dwelling adults aged 50 years and over in Ontario, Canada. The incidence of coronal caries was 57.0%, and the mean net DFS increment was 1.9 surfaces. In bivariate analysis, several variables were significantly associated with incidence and/or mean DFS increment. These included: age, marital status, baseline coronal DFS, number of teeth at baseline, mean periodontal attachment loss of 4 mm or more, and wearing partial dentures. In logistic regression analysis only four factors had significant independent effects. These were level of education, marital status, mean periodontal attachment loss and number of teeth at baseline. The predictive ability of this model was fair: accuracy 65.7%, sensitivity 80.2%, and specificity 46.2%. When logistic analysis was repeated separately for two age groups, different predictors had significant independent effects, and sensitivity and specificity values differed substantially. These findings indicate predictive models for caries incidence should include both clinical and non-clinical variables because both types of variables may help to explain different aspects of coronal caries experience. Further research is required to identify other factors associated with coronal caries in older adults.

Age Factors↗
PubMed2005

Predictors of primary breast cancers responsiveness to preoperative epirubicin/cyclophosphamide-based chemotherapy: translation of microarray data into clinically useful predictive signatures.

BACKGROUND: Our goal was to identify gene signatures predictive of response to preoperative systemic chemotherapy (PST) with epirubicin/cyclophosphamide (EC) in patients with primary breast cancer. METHODS: Needle biopsies were obtained pre-treatment from 83 patients with breast cancer and mRNA was profiled on Affymetrix HG-U133A arrays. Response ranged from pathologically confirmed complete remission (pCR), to partial remission (PR), to stable or progressive disease, "No Change" (NC). A primary analysis was performed in breast tissue samples from 56 patients and 5 normal healthy individuals as a training cohort for predictive marker identification. Gene signatures identifying individuals most likely to respond completely to PST-EC were extracted by combining several statistical methods and filtering criteria. In order to optimize prediction of non responding tumors Student's t-test and Wilcoxon test were also applied. An independent cohort of 27 patients was used to challenge the predictive signatures. A k-Nearest neighbor algorithm as well as two independent linear partial least squares determinant analysis (PLS-DA) models based on the training cohort were selected for classification of the test samples. The average specificity of these predictions was greater than 74% for pCR, 100% for PR and greater than 62% for NC. All three classification models could identify all pCR cases. RESULTS: The differential expression of 59 genes in the training and the test cohort demonstrated capability to predict response to PST-EC treatment. Based on the training cohort a classifier was constructed following a decision tree. First, a transcriptional profile capable to distinguish cancerous from normal tissue was identified. Then, a "favorable outcome signature" (31 genes) and a "poor outcome signature" (26 genes) were extracted from the cancer specific signatures. This stepwise implementation could predict pCR and distinguish between NC and PR in a subsequent set of patients. Both PLS-DA models were implemented to discriminate all three response classes in one step. CONCLUSION: In this study signatures were identified capable to predict clinical outcome in an independent set of primary breast cancer patients undergoing PST-EC.

Journal Article↗
PubMed2006

[Problems of implementing integration management at company level in small and medium-sized enterprises].

UNLABELLED: At company level responsibility increases for the employment of workers with health-related problems or disabilities, but realisation in small and medium-sized enterprises (SME) is lacking. Therefore a model is developed based on theory and a survey. SITUATION: Minimum requirements for "betriebliches Eingliederungsmanagement" (integration management at company level) according to section 84 (2) SGB IX Book 9 of the German Social Code, the main products of the international movement "disability management", a description of roles for realisation and the main sources of employers' support are described. Although external supporting of SMEs is unquestioned, it is expensive and retards own initiative and own activity counting solely on this. HYPOTHESIS: Only by developing a minimum of SME's awareness, acceptance and competence, this will open up to (currently suboptimal) external support. SURVEY: Goal is identifying SME managers' attitudes, activities, proposals and expectations referring integration management at company level to derive concepts of SME's support. 13 interviews are analysed by qualitative content analysis identifying the following barriers: information deficit, absence of priority, limited possibilities for transitional work, cost, partially limited workers' self-responsibility, illness as a "tabes" subject. Possibilities overcoming these barriers are delineated. On that basis a model is presented: pragmatically for realisation, a company contact person with minimum competence, uniform external support, institutional partners' integration and quality assurance according to disability management. OUTLOOK: Interlocking SME world and social insurance world means first to support SME's awareness, acceptance and competence, second to create for SME a central contact in the "rehabilitation jungle" and third to develop SME-suitable premiums according to section 84 (3) SGB IX, Book 9 of the German social code.

Persons with Disabilities↗
PubMed1992

Oxidative fragmentation of collagen and prolyl peptide by Cu(II)/H2O2. Conversion of proline residue to 2-pyrrolidone.

Oxidative degradation of collagen and the model peptides by Cu(II)/H2O2 has been studied. The depolymerization of collagen was predominantly observed by use of gel filtration chromatography. Polyproline was used as a model for collagen, and the oxidative modification was examined by amino acid analysis. Glutamic acid and gamma-aminobutyric acid were identified in the hydrolysates of oxidized polyproline. The formation of glutamic acid was reduced by treatment with NaBH4. The model peptide, (Pro-Pro-Gly)10, was also degraded by Cu(II)/H2O2, and a new N-terminal glycine was generated in proportion to the reaction time. Hydroxyl radical scavengers show only partial inhibition of the degradation of (Pro-Pro-Gly)10. In order to estimate the fragmentation mechanism, we used N-tert-butoxycarbonyl (Boc)-L-prolylglycine as a model for collagen and (Pro-Pro-Gly)10. The degradation products were isolated and characterized. Then N-tert-Boc-2-pyrrolidone, which provides gamma-aminobutyric acid by acid hydrolysis, was identified. The formation of a 2-pyrrolidone compound from oxidized Boc-L-prolylglycine is direct evidence for the scission of the peptide bond. The time-dependent formation of N-tert-Boc-2-pyrrolidone and liberation of glycine from N-tert-Boc-L-prolylglycine exposed to Cu(II)/H2O2 was observed. These results suggest that the cleavage of the peptide bond (Pro-Gly) was caused by oxidation of the proline residue, which led to the formation of the 2-pyrrolidone compound. We confirmed that proline oxidation leads to the fragmentation of proteins, accompanied by the formation of a 2-pyrrolidone structure.

Amino Acid Sequence↗
PubMed1993

Mechanical properties of vesicles. II. A model for osmotic swelling and lysis.

Vesicle polydispersity and leakage of solutes from the vesicle lumen influence the measurement and analysis of osmotically induced vesicle swelling and lysis, but their effects have not been considered in previous studies of these processes. In this study, a model is developed which expressly includes polydispersity and leakage effects. The companion paper demonstrated the preparation and characterization of large unilamellar lipid vesicles. A dye release technique was employed to indicate the leakage of solutes from the vesicles during osmotic swelling. Changes in vesicle size were monitored by dynamic light scattering (DLS). In explaining the results, the model identifies three stages. The first phase involves differential increases in membrane tension with strain increasing in larger vesicles before smaller ones. In the second phase, the yield point for lysis (leakage) is reached sequentially from large sizes to small sizes. In the final phase, the lumen contents and the external medium partially equilibrate under conditions of constant membrane tension. When fit to the data, the model yields information on polydispersity-corrected values for membrane area compressibility, Young's modulus, and yield point for lysis.

Biomechanical Phenomena↗
PubMed2001

Circular structure of the MCMI-III personality disorder scales.

Millon's (1987) circular model of personality disorders was examined in a large sample of psychiatric patients (N = 2,366) who completed the Millon Clinical Multiaxial Inventory-III (MCMI-III; Millon, 1997) as part of routine assessment after presentation for treatment. Principal components analyses were conducted to identify the first two dimensions in MCMI-III base rate scores, weighted and unweighted raw scores, and nonoverlapping scale scores. Similar analyses were made on these scores when acquiescence was partialled out. Circular plots of the scales were examined against Millon's hypothesized arrangement and the model was tested using confirmatory factor analysis. Results replicated those of Strack, Lorr, and Campbell (1990) with the MCMI-II. Millon's horizontal Impassive-Expressive dimension was recovered in both regular and residual scores but the vertical axis appeared to represent an Impulsivity-Compulsivity dimension rather than the Autonomous-Enmeshed continuum envisioned by Millon. Although scale order followed Millon's predictions for the most part, a number of departures from theoretical expectations were noted and none of the score sets yielded a good fit to the hypothetical structure. Millon's model appears to have promise as a circumplex that can encompass all of the personality disorders but changes are needed to rectify discrepancies between the theory and empirical findings.

Adolescent↗
PubMed1997

The functional architecture of an acetylcholine receptor-mimicking antibody.

Malpha2-3 is a monoclonal antibody that partially mimics the nicotinic acetylcholine receptor (AChR). Its three-dimensional structure has been previously predicted by molecular modeling, suggesting that 29 complementarity determining region (CDR) residues and 2 framework residues are exposed to solvent. To identify the antibody residues that bind to the antigen, i.e. snake toxin that binds specifically to AChR, we (i) produced the scFv form of Malpha2-3 fused to alkaline phosphatase, in the periplasmic space of Escherichia coli; (ii) submitted approximately 75% of exposed residues of the fused scFv to individual or combined mutations, and (iii) identified the residues whose mutations affect scFv binding to the toxin, using a sensitive enzyme-linked immunosorbent assay. 11 critical residues were identified, including 8 heavy chain residues, 2 framework residues, and 1 light chain residue. They cover a surface of approximately 800 A2, with a subset of most critical residues (VHD31, VHY32, and VHG101) and several aromatic residues. This functional architecture not only constitutes a plausible complementary binding surface for the snake toxin but also offers a structural basis to ultimately understand the capacity of the antibody to partially mimic AChR.

Alkaline Phosphatase↗
PubMed1999

Crystal structure of penicillin G acylase from the Bro1 mutant strain of Providencia rettgeri.

Penicillin G acylase is an important enzyme in the commercial production of semisynthetic penicillins used to combat bacterial infections. Mutant strains of Providencia rettgeri were generated from wild-type cultures subjected to nutritional selective pressure. One such mutant, Bro1, was able to use 6-bromohexanamide as its sole nitrogen source. Penicillin acylase from the Bro1 strain exhibited an altered substrate specificity consistent with the ability of the mutant to process 6-bromohexanamide. The X-ray structure determination of this enzyme was undertaken to understand its altered specificity and to help in the design of site-directed mutants with desired specificities. In this paper, the structure of the Bro1 penicillin G acylase has been solved at 2.5 A resolution by molecular replacement. The R-factor after refinement is 0.154 and R-free is 0.165. Of the 758 residues in the Bro1 penicillin acylase heterodimer (alpha-subunit, 205; beta-subunit, 553), all but the eight C-terminal residues of the alpha-subunit have been modeled based on a partial Bro1 sequence and the complete wild-type P. rettgeri sequence. A tightly bound calcium ion coordinated by one residue from the alpha-subunit and five residues from the beta-subunit has been identified. This enzyme belongs to the superfamily of Ntn hydrolases and uses Ogamma of Ser beta1 as the characteristic N-terminal nucleophile. A mutation of the wild-type Met alpha140 to Leu in the Bro1 acylase hydrophobic specificity pocket is evident from the electron density and is consistent with the observed specificity change for Bro1 acylase. The electron density for the N-terminal Gln of the alpha-subunit is best modeled by the cyclized pyroglutamate form. Examination of aligned penicillin acylase and cephalosporin acylase primary sequences, in conjunction with the P. rettgeri and Escherichia coli penicillin acylase crystal structures, suggests several mutations that could potentially allow penicillin acylase to accept charged beta-lactam R-groups and to function as a cephalosporin acylase and thus be used in the manufacture of semi-synthetic cephalosporins.

Amino Acid Sequence↗
PubMed1975

Fragments of bovine serum albumin produced by limited proteolysis. Isolation and characterization of tryptic fragments.

Several fragments of bovine serum albumin have been isolated following limited tryptic hydrolysis and their positions then determined in the bovine serum albumin sequence published by J. R. Brown ((1975), Fed. Proc., Fed. Am. Soc. Exp. Biol. 34, 591). When bovine serum albumin was coupled to palmityl-aminoethylamino-agarose and digested with trypsin, two fragments were obtained: (a) peptide 115-184, containing the highly aromatic disulfide loop 3 of Brown's model, and (b) a larger fragment, residues 377-581, containing disulfide loops 7-9. This fragment constitutes the third of the three domains of the albumin molecule. From bovine serum albumin digested in solution, peptide 115-184 was again obtained, as well as (c) a 39,000-dalton fragment identified as residues 198-581, loops 4-9 of the second and third domains, but with a long, tryptophan-containing segment 204-238 missing from loop 4. The ability to isolate these fragments without cleaving disulfide bridges is partial confirmation of the proposed model of bovine serum albumin as a series of nine independent loops.

Amino Acid Sequence↗
PubMed2004

Sequence of pattern onset responses in the human visual areas: an fMRI constrained VEP source analysis.

We measured the timing of activity in distinct functional areas of the human visual cortex after onset of a visual pattern. This is not possible with visual evoked potentials (VEPs) or magnetic fields alone, and direct combination of functional magnetic resonance imaging (fMRI) with electromagnetic data has turned out to be difficult. We tested a relatively new approach, where both position and orientation of the active cortex was given to the VEP source model. Subjects saw the same visual patterns flashed ON and OFF, both when recording VEPs and fMRI responses. We identified the positions and orientations of the activated cortex in four retinotopic areas in each individual, and the corresponding dipoles were seeded to model the individual evoked potential data. Unexplained variance, comprising signals from other areas, was inversely modeled. Despite the partially a priori fixed model and optimized signal-to-noise ratio of VEP data, full separation of retinotopic areas was only seldom possible due to crosstalk between the adjacent sources, but separation was usually possible between areas V1 and V3/V3a. Whereas the latencies generally followed the hierarchical organization of cortical areas (V1-V2-V3), with around 25 ms between the strongest responses, an early activation emerged 10-20 ms after V1, close to the temporo-occipital junction (LO/V5) and with an additional 20-ms latency in the corresponding region of the opposite hemisphere. Our approach shows that it is feasible to directly seed information from fMRI to electromagnetic source models and to identify the components and dynamics of VEPs in different retinotopic areas of a human individual.

Adult↗
PubMed2005

Recovery of impaired songs following unilateral but not bilateral lesions of nucleus uvaeformis of adult zebra finches.

Zebra finches utilize neural circuits in both cerebral hemispheres to produce their learned songs. Although direct reciprocal connections do not exist between song control nuclei across hemispheres, premotor activity in these nuclei during singing is precisely and continuously coordinated between the hemispheres. We hypothesized that this interhemispheric coordination is mediated by bilateral feedback projections from medullary and midbrain song control nuclei to the thalamic song control nucleus uvaeformis (Uva). Consistent with our hypothesis, bilateral lesions of Uva severely impaired singing. This impairment was long-lasting, as it persisted for at least 35 days after the lesions. Unilateral lesions of Uva on either side also resulted in an immediate singing impairment. However, song recovered substantially after less than 15 days, suggesting a possible compensation by the unlesioned side. Although the acoustic structure of individual syllables recovered fully after unilateral lesioning, subtle changes in the sequencing of syllables were observed after song recovery, suggesting that the lesion led to an alteration in the functioning of the remaining song control network. These results demonstrate that the adult songbird brain can adjust to damage to certain parts of the song control network and recover from their associated singing deficits. The well-identified and localized central neural pathways mediating birdsong production provide an advantageous model system to analyze systematically the sensorimotor contexts and the specific sites and mechanisms for behavioral recovery following partial damage to a behavior-producing neural circuit.

Animals↗
PubMed1985

Epidermal growth factor from the mouse. Physical evidence for a tiered beta-sheet domain: two-dimensional NMR correlated spectroscopy and nuclear Overhauser experiments on backbone amide protons.

When H2O-exchanged, lyophilized mouse epidermal growth factor (mEGF) is dissolved in deuterium oxide at low pH (i.e., below approximately 6.0), 13 well-resolved, amide proton resonances are observed in the downfield region of an NMR spectrum (500 MHz). Under the conditions of these experiments, the lifetimes of these amide protons in exchange for deuterons of the deuterium oxide solvent suggest that these amide protons are hydrogen-bonded, backbone amide protons. Several of these amide proton resonances show splittings (i.e., JNH alpha-CH) of approximately 8-10 Hz, indicating that their associated amide protons are in some type of beta-structure. Selective nuclear Overhauser effect (NOE) experiments performed on all amide proton resonances strongly suggest that all 13 of these backbone amide protons are part of a single-tiered beta-sheet structural domain in mEGF. Correlation of 2D NMR correlated spectroscopy data, identifying scaler coupled protons, with NOE data, identifying protons close to the irradiated amide protons, allows tentative assignment of some resonances in the NOE difference spectra to specific amino acid residues. These data allow a partial structural model of the tiered beta-sheet domain in mEGF to be postulated.

Amino Acid Sequence↗