Rapid and label-free nanomechanical detection of biomarker transcripts in human RNA.

The availability of entire genome sequences has triggered the development of microarrays for clinical diagnostics that measure the expression levels of specific genes. Methods that involve labelling can achieve picomolar detection sensitivity, but they are costly, labour-intensive and time-consuming. Moreover, target amplification or biochemical labelling can influence the original signal. We have improved the biosensitivity of label-free cantilever-array sensors by orders of magnitude to detect mRNA biomarker candidates in total cellular RNA. Differential gene expression of the gene 1-8U, a potential marker for cancer progression or viral infections, has been observed in a complex background. The measurements provide results within minutes at the picomolar level without target amplification, and are sensitive to base mismatches. This qualifies the technology as a rapid method to validate biomarkers that reveal disease risk, disease progression or therapy response. We foresee cantilever arrays being used as a tool to evaluate treatment response efficacy for personalized medical diagnostics.

Evaluation of the quality of patient information to support informed shared decision-making.

OBJECTIVES: (a) To find out how much patient information material on display in family physicians' offices refers to management choices, and hence may be useful to support informed and shared decision-making (ISDM) by patients and (b) to evaluate the quality of print information materials exchanged during the consultation, i.e. brought in by patients or given out by family physicians. DESIGN: All print information available for patients and exchanged between physicians and patients was collected in a single complete day of the office practices of 21 family physicians. A published and validated instrument (DISCERN) was used to assess quality. SETTING AND PARTICIPANTS: Community office practices in the greater Vancouver area, British Columbia, Canada. The physicians were purposefully recruited by their association with the medical school Department of Family Practice, their interest in providing patients with print information and their representation of a range of practice types and location. MAIN VARIABLES STUDIED: The source of the pamphlets and these categories: available in the physicians' offices; exchanged between physician and patient; and produced with the explicit or apparent intent to support evidence-based patient choice. MAIN OUTCOME MEASURES: The quality of the print information to support ISDM, as measured by DISCERN and the ease of use and reliability of the DISCERN tool. RESULTS AND CONCLUSIONS: Fewer than 50% of pamphlets available in these offices fulfilled our minimum criteria for ISDM (mentioned more than one management option). Offices varied widely in the proportion of pamphlets on display that supported ISDM and how particular the physician was in selecting materials. The DISCERN tool is quick, valid and reliable for the evaluation of patient information. The quality of patient information materials used in the consultation and available in these offices was below midpoint on the DISCERN score. Major deficiencies were with respect to the mention of choices, risks, effect of no treatment or uncertainty and reliability (source, evidence-base). Good quality information can be produced; some is available locally.

The effect of low dose methotrexate on bone density.

OBJECTIVE: High dose methotrexate (MTX) has been linked with bone loss in oncology patients. However, it is unclear whether longterm low dose MTX used in the treatment of inflammatory arthritis is associated with bone loss. We compared the effect of low dose MTX on bone density in prospectively recruited patients with rheumatoid arthritis (RA) and psoriasis/psoriatic arthritis (Ps/PsA). METHODS: Thirty RA patients and 30 Ps/PsA patients taking MTX were compared to controls not taking MTX (30 with RA, 27 Ps/PsA). Bone mineral density (BMD) of the radius, lumbar spine, trochanter, and femoral neck was measured using Lunar dual energy x-ray absorptiometry. Student t tests were used to detect differences in bone density (using Z scores) of the MTX group versus controls for both the RA and Ps/PsA groups. Analysis of covariance was used to examine for confounders including disease duration, disease activity, age, and sex. RESULTS: BMD of the radius/femoral neck/trochanter did not differ significantly between the MTX treated groups and controls when analyzed by Z scores. The mean difference between the MTX group and controls of the femoral neck was 0.040 (95% CI -0.40, 0.12) and 0.060 (95% CI -0.30, 0.15) for the RA and Ps/PsA groups, respectively. The absolute BMD of the lumbar spine (L2-L4) was higher in the RA MTX group than in controls. Analysis of covariance did not reveal an effect of study group on bone density. CONCLUSION: This study suggests that low dose MTX does not have a negative effect on bone density, at either cortical or trabecular sites.

Legalized, regulated, but unfunded: midwifery's laborious professionalization in Alberta, Canada, 1975-99.

In 1992, Alberta became the second Canadian province to legalize midwifery. This happened even though there were only approximately 20 midwives in practice at the time, and despite strong opposition from the medical and nursing professions. Between 1992 and 1999. Alberta established a regulatory framework for midwifery as a profession but. unlike Ontario and British Columbia, failed to pay midwives out of the provincial health care budget. This sent midwifery in Alberta into a crisis as many midwives closed their practices. This article first considers why midwifery was legalized and then professionalized in Alberta. Our answer emphasizes the leading role of state health bureaucrats in promoting midwifery as part of the state's challenge to medical dominance. Second. the article addresses why midwifery received so little governmental support at the same time that it attained professional status. This analysis includes a comparison with how midwifery developed in Ontario and British Columbia. Our conclusion is that midwifery in Alberta became a victim in the post-1993 period when a new Right government set aside bureaucratic initiatives in health care and committed itself to major cuts in government spending.

Chemical force microscopy with active enzymes.

The adhesion forces have been measured between an atomic force microscope tip derivatized with an active enzyme, shikimate kinase, and an ATP mimic immobilized on a gold surface. Experiments with competitive binding of other ligands in solution show that the observed adhesion forces arise predominantly from specific interactions between the immobilized enzyme and surface-bound adenine derivative. These experiments represent a step in the development of a screening methodology based upon chemical force microscopy.

Does the site of postgraduate family medicine training predict performance on summative examinations? A comparison of urban and remote programs.

BACKGROUND: The location of postgraduate medical training is shifting from teaching hospitals in urban centres to community practice in rural and remote settings. We were interested in knowing whether learning, as measured by summative examinations, was comparable between graduates who trained in urban centres and those who trained in remote and rural settings. METHODS: Family medicine training programs in Ontario were selected as a model of postgraduate medical training. The results of the 2 summative examinations--the Medical Council of Canada Qualifying Examination (MCCQE) Part II and the College of Family Physicians of Canada (CFPC) certification examination--for graduates of the programs at Ontario's 5 medical schools were compared with the results for graduates of the programs in Sudbury and Thunder Bay from 1994 to 1997. The comparability of these 2 cohorts at entry into training was evaluated using the results of their MCCQE Part I, completed just before the family medicine training. RESULTS: Between 1994 and 1997, 1013 graduates of family medicine programs (922 at the medical schools and 91 at the remote sites) completed the CFPC certification examination; a subset of 663 completed both the MCCQE Part I and the MCCQE Part II. The MCCQE Part I results for graduates in the remote programs did not differ significantly from those for graduates entering the programs in the medical schools (mean score 531.3 [standard deviation (SD) 69.8] and 521.8 [SD 74.4] respectively, p = 0.33). The MCCQE Part II results did not differ significantly between the 2 groups either (mean score 555.1 [SD 71.7] and 545.0 [SD 76.4] respectively, p = 0.32). Similarly, there were no consistent, significant differences in the results of the CFPC certification examination between the 2 groups. INTERPRETATION: In this model of postgraduate medical training, learning was comparable between trainees in urban family medicine programs and those in rural, community-based programs. The reasons why this outcome might be unexpected and the limitations on the generalizability of these results are discussed.

Synovial cyst of the proximal tibiofibular joint.

We describe a case of an unusual synovial cyst originating from the proximal tibiofibular joint. This cyst presented as a painless, soft tissue swelling just distal to the fibular head. We review the literature on this topic regarding clinical presentation, imaging, and management.

Activin-induced factors maintain goosecoid transcription through a paired homeodomain binding site.

Previous studies in both Xenopus and zebrafish have shown that goosecoid is one of the first genes to be transcribed at the onset of gastrulation. Goosecoid transcription still initiates when embryos are treated with protein synthesis inhibitors, indicating that it is mediated by preexisting factors and suggesting that goosecoid transcription is immediately downstream of the maternal mesoderm-inducing signal. However, goosecoid transcription continues long after this maternal signal has ceased to be active, indicating that there are mechanisms to maintain activin-induced transcription. Our study has focused on understanding the factors required to maintain this transcription. We have defined an element within the zebrafish goosecoid promoter that is sufficient for activin inducibility in both Xenopus and zebrafish embryos. This element, the goosecoid activin element, interacts with two developmentally regulated proteins from Xenopus embryos. A maternal protein interacts through cleavage stages until the midblastula transition, and a second protein binds from the onset of gastrulation. The second protein is zygotically expressed, and its binding is required for activin inducibility in our assay system. We suggest that the zygotic protein we have identified is a good candidate to be involved in the maintenance of goosecoid transcription. Furthermore, this zygotic protein is likely to contain a paired class homeodomain since a consensus binding site for such proteins is present within the goosecoid activin element and is essential for its function.

The remarkable spectrum of methotrexate toxicities.

This article outlines a general scheme for categorizing medication-related adverse events. This is followed by a review of the less well-recognized adverse events attributed to low-dose methotrexate therapy. Known and suspected risk factors are described and causative mechanisms are suggested. Ultimately, this article aims at increasing the readers awareness of uncommon or underreported methotrexate-associated adverse events so that prescribing and monitoring practices can be tailored to enhance the safe use of this valuable antirheumatic agent.

Combination treatment of severe rheumatoid arthritis with cyclosporine and methotrexate for forty-eight weeks: an open-label extension study. The Methotrexate-Cyclosporine Combination Study Group.

OBJECTIVE: To determine whether the clinical benefit and favorable safety profile previously noted with the combination of cyclosporine (CSA) and methotrexate (MTX) given for 24 weeks in patients with rheumatoid arthritis (RA) would be maintained for a further 24 weeks, and whether the addition of CSA in patients who had previously been randomized to receive placebo + MTX would result in clinical benefit. METHODS: Eligible subjects from the initial study (weeks 0-24), in which the addition of placebo or CSA to MTX therapy was compared in patients with RA that was partially responsive to MTX, were enrolled. Patients who had received CSA + MTX continued this regimen for a further 24 weeks (weeks 24-48) (group 1; n = 48), and patients who had initially received placebo + MTX now received CSA + MTX for 24 weeks (weeks 24-48) (group 2; n = 44), in an open-label extension study. The primary outcome measures were the number of tender joints, number of swollen joints, physician and patient global assessments, pain, functional disability as measured by the modified Health Assessment Questionnaire, and erythrocyte sedimentation rate. RESULTS: Of the 92 patients enrolled, 80 (87%) completed the extension study. In patients in group 1, the clinically and statistically significant improvement in response outcomes previously noted at week 24, ranging from 25% to 50%, was maintained through week 48. In patients in group 2, the addition of CSA resulted in significant clinical improvement. By week 48, most outcome measures in group 2 patients were similar to those in group 1 patients. CSA treatment resulted in a small increase in serum creatinine levels, but only 1 patient was withdrawn from the study for this reason. CONCLUSION: The clinical improvement previously observed in patients treated with the CSA + MTX combination for 24 weeks was maintained for 24 subsequent weeks, without serious adverse effects, and was also observed in the patients whose treatment was switched from placebo + MTX to CSA + MTX.

LEF-1/TCF proteins mediate wnt-inducible transcription from the Xenopus nodal-related 3 promoter.

The Xenopus nodal-related 3 gene (Xnr3) is expressed in the Spemann organizer of the embryo and encodes a member of the transforming growth factor beta family that mediates some activities of the organizer. Xnr3 is transcriptionally activated by wnt signaling during gastrulation in the Xenopus embryo. Here we show that a small region of the Xnr3 promoter is sufficient to confer wnt-inducible transcription. By mutational analysis of the promoter, we have identified two distinct sequence elements required for the response to wnt signals. One regulatory sequence interacts with a factor which accumulates in Xenopus gastrulae independent of wnt signaling. The other functionally important site can bind mammalian LEF-1 protein, a member of the LEF-1/TCF family of transcription factors. In addition, misexpression of LEF-1 in embryo explants induces transcription of the endogenous Xnr3 gene. Taken together, these data provide further evidence for a role of LEF-1/TCF proteins in wnt signaling and identify the Spemann organizer-specific gene Xnr3 as a direct target of these transcription factors in vertebrates.

Interferon-alpha-dependent activation of Tyk2 requires phosphorylation of positive regulatory tyrosines by another kinase.

Tyk2 and JAK1, members of the Janus kinase (JAK) family of protein tyrosine kinases, are required for interferon-alpha/beta binding and signaling. Both enzymes are associated with the interferon-alpha/beta receptor, and upon ligand binding, they undergo tyrosine phosphorylation and catalytic activation in an interdependent manner. To identify residues involved in Tyk2 regulation and to understand the basis of the interdependence of Tyk2 and JAK1, six mutated versions of Tyk2 bearing single or multiple point mutations in the tyrosine kinase domain were studied in a cell line lacking endogenous Tyk2. The Y1054F/Y1055F substitutions in the putative activation loop prevented ligand-dependent activation of Tyk2, without abolishing its catalytic potential. The K930R mutation in the ATP binding site generated a kinase-negative protein, which however, still became phosphorylated upon interferon-alpha treatment. The Y1054F/Y1055F substitutions in this kinase-negative Tyk2 abolished the induced phosphorylation. These results indicate that Tyk2 is activated by phosphorylation on Tyr-1054 and/or Tyr-1055 and that this phosphorylation requires another kinase, most likely JAK1. While the Tyk2 forms mutated on Tyr-1054 and Tyr-1055 or on Lys-930 allowed some inducible gene expression, the combination of the three point mutations totally abolished signaling.

Progression of rheumatoid arthritis following bone marrow transplantation. A case report with a 13-year followup.

The progression of rheumatoid arthritis (RA) is documented in a patient receiving a sex-mismatched, allogeneic bone marrow transplant (BMT) for gold-induced marrow aplasia. DNA typing confirmed a high probability of a full donor engraftment (complete chimerism). Although the RA was in complete remission 2 years post-BMT, clinical, laboratory, histologic, and radiologic evidence of the recurrence of synovitis from 3-13 years post-BMT is presented. Implications of these observations for theories of the pathogenesis of RA and the future of immunotherapies are discussed.

Factors influencing the emigration of physicians from Canada to the United States.

OBJECTIVE: To determine whether location of postgraduate medical training and other factors are associated with the emigration of physicians from Canada to the United States. DESIGN: Case-control study, physicians were surveyed with the use of a questionnaire mailed in May 1994 (with a reminder sent in September 1994), responses to which were accepted until Dec. 31, 1994. PARTICIPANTS: Physicians randomly selected from the CMA database, 4000 with addresses in Canada and 4000 with current addresses in the United States and previous addresses in Canada. OUTCOME MEASURES: Sex, age, location of undergraduate and postgraduate medical training, qualifications, practice location, opinions concerning residence decisions, current satisfaction and plans. RESULTS: The overall response rate was 49.6% (50.0% among physicians in the United States and 49.2% among those in Canada). Age and sex distributions were similar among the 8000 questionnaire recipients and the nearly 4000 respondents. Physicians living in the United States were more likely to be older (mean 53.2 v. 49.6 years of age), male (87% v. 75%) and specialists (79% v. 52%) than those practising in Canada. Postgraduate training in the United States was associated with subsequent emigration (odds ratio 9.2, 95% confidence interval 7.8 to 10.7). However, in rating the importance of nine factors in the decision to emigrate or remain in Canada, there was no significant difference between the two groups in the rating assigned to location of postgraduate training. Professional factors rated most important by most physicians in both groups were professional/clinical autonomy, availability of medical facilities and job availability. Remuneration was considered an equally important factor by those in Canada and in the United States. Six of seven personal/family factors were rated as more important to their choice of practice location by respondents in Canada than by those in the United States. Current satisfaction was significantly higher among respondents in the United States. Most physicians in each group planned to continue practising at their current location. Of Canadian respondents, 22% indicated that they were more likely to move to the United States than they were a year beforehand, whereas 4% of US respondents indicated that they were more likely to return to Canada. CONCLUSIONS: Factors affecting the decision to move to the United States or remain in Canada can be categorized as "push" factors (e.g., government involvement) and "pull" factors (e.g., better geographic climate in the US). Factors can also be categorized by whether they are amenable to change (e.g., availability of medical facilities) or cannot be managed (e.g., proximity of relatives). An understanding of the reasons why physicians immigrate to the United States or remain in Canada is essential to planning physician resources nationally.