Transcriptional profiling of luteinizing hormone receptor-deficient mice before and after testosterone treatment provides insight into the hormonal control of postnatal testicular development and Leydig cell differentiation.

Luteinizing hormone (LH) is a key regulator of male fertility through its effects on testosterone secretion by Leydig cells. Transcriptional control of this is, however, currently poorly understood. Mice in which the LH receptor is knocked out (LuRKO) show reduced testicular size, reduced testosterone, elevated serum LH, and a spermatogenic arrest that can be rescued by the administration of testosterone. Using genome-wide transcription profiling of LuRKO and control testes during postnatal development and following testosterone treatment, we show that the transcriptional effects of LH insensitivity are biphasic, with an early testosterone-independent phase and a subsequent testosterone-dependent phase. Testosterone rescue re-enables the second, testosterone-dependent phase of the normal prepubertal transcription program and permits the continuation of spermatogenesis. Examination of the earliest responses to testosterone highlights six genes that respond rapidly in a dose-dependent fashion to the androgen and that are therefore candidate regulatory genes associated with the testosterone-driven progression of spermatogenesis. In addition, our transcriptional data suggest a model for the replacement of fetal-type Leydig cells by adult-type cells during testicular development in which a testosterone feedback switch is necessary for adult Leydig cell production. LH signaling affects the timing of the switch but is not a strict requirement for Leydig cell differentiation.

Modulation of the mouse testis transcriptome during postnatal development and in selected models of male infertility.

The aim of this study is to develop an overview of genetic events during spermatogenesis using a novel, specifically targeted gonadal gene set. Two subtracted cDNA libraries enriched for testis specific and germ cell specific genes were constructed, characterized and sequenced. The combined libraries contain >1905 different genes, the vast majority previously uncharacterized in testis. cDNA microarray analysis of the first wave of murine spermatogenesis and of selected germ cell-deficient models was used to correlate the expression of groups of genes with the appearance of defined germ cell types, suggesting their cellular expression patterns within the testis. Real-time RT-PCR and comparison to previously known expression patterns confirmed the array-derived transcription profiles of 65 different genes, thus establishing high confidence in the profiles of the uncharacterized genes investigated in this study. A total of 1748 out of 1905 genes showed significant change during the first spermatogenic wave, demonstrating the successful targeting of the libraries to this process. These findings highlight unknown genes likely to be important in germ cell production, and demonstrate the utility of these libraries in further studies. Transcriptional analysis of well-characterized mouse models of infertility will allow us to address the causes and progression of the pathology in related human infertility phenotypes.

Self-directed learning: faculty and student perceptions.

This article reports the results of a qualitative study that explored faculty and student perceptions of self-directed learning (SDL) and investigated factors that facilitate or impede it. This study was conducted at McMaster University with faculty and students in a 4-year undergraduate nursing program. Data were collected from 47 faculty and 17 students by means of focus groups that were audiotaped and transcribed. Content analysis was conducted to identify common themes in faculty and student transcripts. The themes that emerged provide insight into the educational strategy of self-directed learning and can be summarized by the following major points: (1) commitment to SDL requires students and faculty to understand the value of empowering learners to take increased responsibility for decisions related to learning; (2) students engaged in self-directed learning undergo a transformation that begins with negative feelings (i.e., confusion, frustration, and dissatisfaction) and ends with confidence and skills for lifelong learning; and (3) faculty development is important to ensure high levels of competency in facilitating self-directed learning.

Crystal structure of the 100 kDa arsenite oxidase from Alcaligenes faecalis in two crystal forms at 1.64 A and 2.03 A.

BACKGROUND: Arsenite oxidase from Alcaligenes faecalis NCIB 8687 is a molybdenum/iron protein involved in the detoxification of arsenic. It is induced by the presence of AsO(2-) (arsenite) and functions to oxidize As(III)O(2-), which binds to essential sulfhydryl groups of proteins and dithiols, to the relatively less toxic As(V)O(4)(3-) (arsenate) prior to methylation. RESULTS: Using a combination of multiple isomorphous replacement with anomalous scattering (MIRAS) and multiple-wavelength anomalous dispersion (MAD) methods, the crystal structure of arsenite oxidase was determined to 2.03 A in a P2(1) crystal form with two molecules in the asymmetric unit and to 1.64 A in a P1 crystal form with four molecules in the asymmetric unit. Arsenite oxidase consists of a large subunit of 825 residues and a small subunit of approximately 134 residues. The large subunit contains a Mo site, consisting of a Mo atom bound to two pterin cofactors, and a [3Fe-4S] cluster. The small subunit contains a Rieske-type [2Fe-2S] site. CONCLUSIONS: The large subunit of arsenite oxidase is similar to other members of the dimethylsulfoxide (DMSO) reductase family of molybdenum enzymes, particularly the dissimilatory periplasmic nitrate reductase from Desulfovibrio desulfuricans, but is unique in having no covalent bond between the polypeptide and the Mo atom. The small subunit has no counterpart among known Mo protein structures but is homologous to the Rieske [2Fe-2S] protein domain of the cytochrome bc(1) and cytochrome b(6)f complexes and to the Rieske domain of naphthalene 1,2-dioxygenase.

Residents' medical information needs in clinic: are they being met?

PURPOSE: Little is known about how often residents encounter unanswered clinical questions in their training. This knowledge would facilitate the development of curricula to help residents practice evidence-based medicine. This study was conducted to determine the frequency, characteristics, and pursuit of residents' clinical questions. SUBJECTS AND METHODS: Residents in a university-based primary care internal medicine program were observed in two hospital-based teaching clinics. Residents were interviewed after each patient encounter to determine whether they had any remaining clinical questions. At the end of each clinic session, they recorded their level of agreement with a series of statements about factors that were expected to motivate residents to seek the answers to each question. One week later, residents were contacted to determine if they had pursued these questions. RESULTS: Sixty-four residents were interviewed after 401 (99%) of 404 patient encounters. They identified 280 new questions, approximately 2 questions for every 3 patients. The most common types of questions were related to therapy (38%) or diagnosis (27%). The residents were subsequently contacted about 277 (99%) of their questions. Of these, only 80 (29%) were pursued, most commonly by consulting textbooks (31%), original articles (21%), or attending physicians (17%). In a multivariable analysis, belief that the patient expected the answer (odds ratio [OR] = 2.3, 95% confidence interval [CI]: 1.3 to 4.0, P = 0.004) and fear of malpractice exposure (OR = 2.1, 95% CI: 1.0 to 4.3, P = 0.05) were associated with information pursuit. Lack of time (60%) and forgetting the question (29%) were the most frequent reasons for failing to pursue a question. CONCLUSION: Residents frequently encountered new clinical questions in the outpatient clinic, but infrequently answered them. Efforts to demonstrate the feasibility of timely searches, remind them of their questions, and reinforce the exigency (educational if not clinical) of all questions may reclaim missed opportunities for self-directed learning.

Crystal structure of the complex of brugia malayi cyclophilin and cyclosporin A.

The resistance of the human parasite Brugia malayi to the antiparasitic activity of cyclosporin A (CsA) may arise from the presence of cyclophilins with relatively low affinity for the drug. The structure of the complex of B. malayi cyclophilin (BmCYP-1) and CsA, with eight independent copies in the asymmetric unit, has been determined at a resolution of 2.7 A. The low affinity of BmCYP-1 for CsA arises from incomplete preorganization of the binding site so that the formation of a hydrogen bond between His132 of BmCYP-1 and N-methylleucine 9 of CsA is associated with a shift in the backbone of approximately 1 A in this region.

Determination of Fe-ligand bond lengths and the Fe-N-O bond angles in soybean ferrous and ferric nitrosylleghemoglobin a using multiple-scattering XAFS analyses.

The NO adducts of leghemoglobin (Lb) are implicated in biological processes, but only the adduct with ferrous Lb (Lb(II)NO) has been characterized previously. We report the first characterization of ferric nitrosylleghemoglobin (Lb(III)NO) and XAS experiments performed on frozen aqueous solutions of Lb(II)NO and Lb(III)NO at 10 K. The XANES and electronic spectra of the NO adducts are similar in shape and energies to the myoglobin (Mb) analogues. The environment of the Fe atom has been refined using multiple-scattering (MS) analyses of the XAFS data. For Lb(II)NO, the MS analysis resulted in an averaged Fe-N(p)(pyrrole) distance of 2.02 A, an Fe-N(epsilon)(imidazole) distance of 1.98 A, an Fe-N(NO) distance of 1.77 A, and an Fe-N-O angle of 147 degrees. The Fe-N(NO) distance and Fe-N-O angle obtained from the analysis of Lb(II)NO are in good agreement with those determined crystallographically for [Fe(TPP)(NO)] (TPP, tetraphenylporphyrinato), with and without 1-methylimidazole (1-MeIm) as the sixth ligand, and the MS XAFS structures reported previously for the myoglobin (Mb(II)NO) analogue and [Fe(TPP)(NO)]. The MS analysis of Lb(III)NO yielded an average Fe-N(p) distance of 2.00 A, an Fe-N(epsilon) distance of 1.89 A, an Fe-N(NO) distance of 1.68 A, and an Fe-N-O angle of 173 degrees. These bond lengths and angles are consistent with those determined previously for the myoglobin analogue (Mb(III)NO) and the crystal structures of the model complexes, [Fe(III)(TPP)(NO)(OH(2))](+) and [Fe(OEP)(NO)](+) (OEP, octaethylporphyrinato). The final XAFS R values were 16.1 and 18.2% for Lb(II)NO and Lb(III)NO, respectively.

Is forced dextrality an explanation for the fall in the prevalence of sinistrality with age? A study in northern England.

OBJECTIVES: The fall in the prevalence of left handedness with age has been attributed to either premature mortality or a cohort effect of forced dextrality. Evidence for forced dextrality was sought to differentiate between these competing theories. DESIGN: 6097 Edinburgh handedness inventories were used to calculate laterality quotients (LQ) with and without the questions relating to writing and drawing. These questions might be expected to be most influenced by forced dextrality. SETTING: The study was performed in a small industrial town in Lancashire, UK. PARTICIPANTS: Using the British family practitioner service where over 95% of the population are registered with a general practitioner a response rate of 82.17% was obtained with respect to the Edinburgh Inventory. RESULTS: Questions about writing and drawing on the Edinburgh Inventory contributed to the positivity (right handedness) of the mean LQ, but equally across the ages. When a negative LQ was used to define left handedness the prevalence of left handedness fell from 11.2% at age 15 to 4.4% at age 70. Removal of the questions about writing and drawing caused the prevalence of left handedness to fall from 10.5% at age 15 to 4.95% at age 70. CONCLUSIONS: Less than 20% of the fall in the prevalence of left handedness was accounted for by questions relating to writing and drawing. The fall in the prevalence of sinistrals in older age groups is not adequately explained by cohort effects of forced dextrality on the writing hand.

Structure of ferric soybean leghemoglobin a nicotinate at 2.3 A resolution.

Soybean leghemoglobin a is a small (16 kDa) protein facilitating the transport of O(2) to respiring N(2)-fixing bacteria at low free-O(2) tension. The crystal structure of soybean ferric leghemoglobin a nicotinate has been refined at 2.3 A resolution. The final R factor is 15.8% for 6877 reflections between 6.0 and 2.3 A. The structure of soybean leghemoglobin a (143 residues) is closely similar to that of lupin leghemoglobin II (153 residues), the proteins having 82 identical residues when the sequences are aligned. The new structure provides support for the conclusion that the unique properties of leghemoglobin arise principally from a heme pocket considerably larger and more flexible than that of myoglobin, a strongly ruffled heme group, and a proximal histidine orientation more favourable to ligand binding.

Impact of an evidence-based medicine curriculum based on adult learning theory.

OBJECTIVE: To develop and implement an evidence-based medicine (EBM) curriculum and determine its effectiveness in improving residents' EBM behaviors and skills. DESIGN: Description of the curriculum and a multifaceted evaluation, including a pretest-posttest controlled trial. SETTING: University-based primary care internal medicine residency program. PARTICIPANTS: Second- and third-year internal medicine residents (N = 34). INTERVENTIONS: A 7-week EBM curriculum in which residents work through the steps of evidence-based decisions for their own patients. Based on adult learning theory, the educational strategy included a resident-directed tutorial format, use of real clinical encounters, and specific EBM facilitating techniques for faculty. MEASUREMENTS AND MAIN RESULTS: Behaviors and self-assessed competencies in EBM were measured with questionnaires. Evidence-based medicine skills were assessed with a 17-point test, which required free text responses to questions based on a clinical vignette and a test article. After the intervention, residents participating in the curriculum (case subjects) increased their use of original studies to answer clinical questions, their examination of methods and results sections of articles, and their self-assessed EBM competence in three of five domains of EBM, while the control subjects did not. The case subjects significantly improved their scores on the EBM skills test (8.5 to 11.0, p = .001), while the control subjects did not (8.5 to 7.1, p = .09). The difference in the posttest scores of the two groups was 3.9 points (p = .001, 95% confidence interval 1.9, 5.9). CONCLUSIONS: An EBM curriculum based on adult learning theory improves residents' EBM skills and certain EBM behaviors. The description and multifaceted evaluation can guide medical educators involved in EBM training.

XFIT - an Interactive EXAFSAnalysis Program.

XFIT is an interactive and user-friendly program for the analysis of X-ray absorption fine structure (XAFS, EXAFS) curves. XFIT incorporates in a single package a number of features available in other existing programs: ab initio EXAFS calculation (using FEFF4.06/6.01), empirical EXAFS calculation (as in XFPAKG), allowance for polarization, use of Fourier filtering, and the application of constraints and restraints. Additional features not previously available are: simultaneous refinement with respect to several data sets, simultaneous refinement of several absorber sites, and Monte-Carlo error analysis. Applications including the analysis of EXAFS data from mixtures and the analysis of DAFS (diffraction anomalous fine structure) data are indicated.

Resistance to tomato spotted wilt virus infection in transgenic tobacco expressing the viral nucleocapsid gene.

A recombinant plasmid containing the entire tomato spotted with virus (TSWV) nucleocapsid gene, with the exception of nucleotide encoding three N-terminal amino acids, was isolated by screening a complementary DNA library, prepared against random primed viral RNA, using a specific monoclonal antibody. The insert contained in plasmid pTSW1 was repaired and amplified by polymerase chain reaction, and the complete nucleocapsid protein gene was introduced into Nicotiana tabacum 'Samsun' by leaf disk transformation using Agrobacterium tumefaciens. Transgenic plants expressing the viral nucleocapsid protein were resistant to subsequent infection following mechanical inoculation with TSWV as indicated by a lack of systemic symptoms and little or no systemic accumulation of virus as determined by double antibody sandwich enzyme-liked immunosorbent assay. These results further extend the applicability of coat protein-mediated resistance, as previously demonstrated for a number of simple plant viruses composed of a positive-sense RNA genome encapsidated with a single species of coat protein, to a membrane-encapsidated, multi-component, negative-sense RNA virus.

Alumni perspectives comparing a general internal medicine program and a traditional medicine program.

OBJECTIVE: To evaluate a primary care internal medicine curriculum, the authors surveyed four years (1983-1986) of graduates of the primary care and traditional internal medicine residency programs at their institution concerning the graduates' preparation. DESIGN: Mailed survey of alumni of a residency training program. SETTING: Teaching hospital alumni. SUBJECTS/METHODS: Of 91 alumni of an internal medicine training program for whom addresses had been found, 82 (90%) of the residents (20 primary care and 62 traditional) rated on a five-point Likert scale 82 items for both adequacy of preparation for practice and importance of training. These items were divided into five groups: traditional medical disciplines (e.g., cardiology), allied disciplines (e.g., orthopedics), areas related to medical practice (e.g., patient education), basic skills and knowledge (e.g., history and physical), and technical procedures. MAIN RESULTS: Primary care residents were more likely to see themselves as primary care physicians versus subspecialists (84% versus 45%). The primary care graduates felt significantly better prepared in the allied disciplines and in areas related to medical practice (p less than 0.01). There was no significant difference overall in perceptions of preparation in the traditional medical disciplines, basic skills and knowledge, and procedures. The same results were obtained when the authors looked only at graduates from the two programs who spent more than 50% of their time as primary care physicians versus subspecialists. There was no significant difference between the two groups in the perceived importance of these areas to current practice. CONCLUSIONS: These results suggest that the primary care curriculum has prepared residents in areas particularly relevant to primary care practice. Additionally, these individuals feel as well prepared as do their colleagues in the traditional medical disciplines, basic skills and knowledge, and procedural skills.

Hand preference in a normal population.

The Edinburgh Inventory was used to assess handedness in a normal population. The incidence of strong right-handedness increased with age and the incidence of left-handedness fell with age, both significantly. A non-significant increase in the incidence of left-handedness in twins was observed. The difference in the incidence of left-handedness for males and females failed to reach significance but more males were left-handed than females. Any difference in the incidence of left-handedness between the sexes must be small (less than 1.5%) and this has major theoretical implications.

Interaction of daily injections and subcutaneous reservoirs of melatonin on the reproductive physiology of female Syrian hamsters.

This study investigated the ability of continuously available melatonin (from sc melatonin-beeswax pellets that were implanted bi-weekly) in overcoming the antigonadotrophic action of single daily melatonin (25 microgram) injections given late in the light period to female Syrian hamsters kept in light:dark cycles of 14:10. The melatonin-beeswax pellets contained either 1, 50, 500 microgram, 1 or 10 mg melatonin and 25 mg beeswax. Daily melatonin injections into hamsters implanted with beeswax only caused vaginal acyclicity in 60% of the animals within 6 weeks. After 10 weeks of treatment, uterine weights and pituitary levels of immunoreactive prolactin were still greatly depressed. Beeswax pellets containing either 500 microgram, 1 or 10 mg melatonin overcame the inhibitory effects of daily melatonin injections. Thus, the vaginal cycles and pituitary hormone levels of these animals were indistinguishable from those of untreated control animals. Beeswax pellets containing either 1 or 50 microgram melatonin were incapable of counteracting the antigonadotrophic effects of daily injections of melatonin. This is the first report that continuously available melatonin can negate the effects of daily injections of melatonin in female hamsters. When measured after 10 weeks, none of the treatment procedures significantly influenced circulating levels of luteinizing hormone or prolactin.

First results from the selective chopper radiometer on Nimbus 4.

This article describes the temperature profiles through the atmosphere that are being obtained from the radiometer on Nimbus 4.

Group B streptococci in neonatal deaths.

Clinical microbiologists are reminded to look for group B streptococci in neonatal infants. We report four deaths, one with meningitis, occurring among 14 such infected newborn infants at our institution; of the nursing staff, 31% were also carriers of group B streptococci.