Does treatment strategy influence the ability to achieve and sustain DMARD-free remission in patients with RA? Results of an observational study comparing an intensified DAS-steered treatment strategy with treat to target in routine care.

OBJECTIVES: To study the impact of treatment strategy on achieving and sustaining disease-modifying antirheumatic drug (DMARD)-free remission in patients with rheumatoid arthritis (RA). METHODS: Two hundred seventy-nine RA patients (median follow-up 7.8 years) were studied. Of these, 155 patients participated in a disease activity score (DAS) < 1.6 steered trial aimed at DMARD-free remission. Initial treatment comprised methotrexate with high-dose prednisone (60 mg/day) and a possibility to start biologicals after 4 months. In the same period and hospital, 124 patients were treated according to routine care, comprising DAS < 2.4 steered treatment. Percentages of DMARD-free remission (absence of synovitis for ≥ 1 year after DMARD cessation), late flares (recurrence of clinical synovitis ≥ 1 year after DMARD cessation), and DMARD-free sustained remission (DMARD-free remission sustained during complete follow-up) were compared between both treatment strategies. RESULTS: Patients receiving intensive treatment were younger and more often ACPA-positive. On a group level, there was no significant association between intensive treatment and DMARD-free remission (35% vs 29%, corrected hazard ratio (HR) 1.4, 95%CI 0.9-2.2), nor in ACPA-negative RA (49% versus 44%). In ACPA-positive RA intensive treatment resulted in more DMARD-free remission (25% vs 6%, corrected HR 4.9, 95%CI 1.4-17). Intensive treatment was associated with more late flares (20% versus 8%, HR 2.3, 95%CI 0.6-8.3). Subsequently, there was no difference in DMARD-free sustained remission on a group level (28% versus 27%), nor in the ACPA-negative (43% versus 42%) or ACPA-positive stratum (17% versus 6%, corrected HR 3.1, 95%CI 0.9-11). CONCLUSIONS: Intensive treatment did not result in more DMARD-free sustained remission, compared to routine up-to-date care. The data showed a tendency towards an effect of intensive treatment in ACPA-positive RA; this needs further investigation.

Does the presence of magnetic resonance imaging-detected osteitis at diagnosis with rheumatoid arthritis lower the risk for achieving disease-modifying antirheumatic drug-free sustained remission: results of a longitudinal study.

BACKGROUND: Although infrequent, some rheumatoid arthritis (RA) patients achieve disease-modifying antirheumatic drug (DMARD)-free sustained remission. The absence of RA-specific autoantibodies, such as anticitrullinated protein antibodies (ACPA), is known to be associated with this outcome but further mechanisms underlying the chronic nature of RA are largely unknown. Magnetic resonance imaging (MRI)-detected bone marrow edema (BME), or osteitis, strongly predicts erosive progression and is associated with ACPA positivity. Therefore, we hypothesized that the presence of MRI-detected osteitis is also predictive of not achieving DMARD-free sustained remission and that the presence of osteitis mediates the association between ACPA and DMARD-free sustained remission. METHODS: A 1.5 T unilateral hand and foot MRI was performed at disease presentation in 238 RA patients, evaluating BME, synovitis, and tenosynovitis (summed as MRI inflammation score). DMARD-free sustained remission, defined as the absence of clinical synovitis after DMARD cessation that persisted during the total follow-up, was assessed (median follow-up 3.8 years). Associations between the different MRI-detected inflammatory features and this outcome were studied. A mediation analysis was performed to study whether the presence of BME mediated the association between ACPA and DMARD-free sustained remission. Finally, patterns of MRI-detected inflammation with regard to DMARD-free sustained remission were studied using partial least squares (PLS) regression. RESULTS: Forty-six (19.3%) patients achieved DMARD-free sustained remission. ACPA positivity associated independently with remission (hazard ratio (HR) 0.16, 95% confidence interval (CI) 0.06-0.39). In contrast, no associations were observed between MRI-detected BME (HR 0.99, 95% CI 0.94-1.03), or other MRI inflammatory features, and achieving DMARD-free sustained remission. Thus, the presence of BME did not mediate the association between ACPA and DMARD-free sustained remission. Furthermore, a PLS analysis revealed that patients who did or did not achieve remission could not be distinguished by patterns of MRI-detected inflammation. CONCLUSIONS: At disease presentation, osteitis, as well as other MRI-detected inflammatory features, was not associated with achieving DMARD-free sustained remission over time. Thus, imaging predictors for joint damage and disease persistence differ. The processes mediating RA chronicity remain largely unsolved.

Cost-effectiveness analysis of new generation coronary CT scanners for difficult-to-image patients.

AIMS: New generation dual-source coronary CT (NGCCT) scanners with more than 64 slices were evaluated for patients with (known) or suspected of coronary artery disease (CAD) who are difficult to image: obese, coronary calcium score > 400, arrhythmias, previous revascularization, heart rate > 65 beats per minute, and intolerance of betablocker. A cost-effectiveness analysis of NGCCT compared with invasive coronary angiography (ICA) was performed for these difficult-to-image patients for England and Wales. METHODS AND RESULTS: Five models (diagnostic decision model, four Markov models for CAD progression, stroke, radiation and general population) were integrated to estimate the cost-effectiveness of NGCCT for both suspected and known CAD populations. The lifetime costs and effects from the National Health Service perspective were estimated for three strategies: (1) patients diagnosed using ICA, (2) using NGCCT, and (3) patients diagnosed using a combination of NGCCT and, if positive, followed by ICA. In the suspected population, the strategy where patients only undergo a NGCCT is a cost-effective option at accepted cost-effectiveness thresholds. The strategy of using NGCCT in combination with ICA is the most favourable strategy for patients with known CAD. The most influential factors behind these results are the percentage of patients being misclassified (a function of both diagnostic accuracy and the prior likelihood), the complication rates of the procedures, and the cost price of a NGCCT scan. CONCLUSION: The use of NGCCT might be considered cost-effective in both populations since it is cost-saving compared to ICA and generates similar effects.

How to prepare a systematic review of economic evaluations for clinical practice guidelines: database selection and search strategy development (part 2/3).

INTRODUCTION: This article is part of the series "How to prepare a systematic review of economic evaluations (EES) for informing evidence-based healthcare decisions", in which a five-step approach is proposed. Areas covered: This paper focuses on the selection of relevant databases and developing a search strategy for detecting EEs, as well as on how to perform the search and how to extract relevant data from retrieved records. Expert commentary: Thus far, little has been published on how to conduct systematic review EEs. Moreover, reliable sources of information, such as the Health Economic Evaluation Database, have ceased to publish updates. Researchers are thus left without authoritative guidance on how to conduct SR-EEs. Together with van Mastrigt et al. we seek to fill this gap.

How we do: optimizing bone marrow biopsy logistics for sign-out within 2 days.

Since the introduction of fast diagnostic tracks in many areas of oncology, the traditional processing of bone marrow biopsies (BMB), requiring either resin embedding or lengthy fixation and decalcification, is due to an upgrade. Thanks to a growing number of new commercially available tissue processors, microwave-enhanced processing is becoming a standard tool in the pathology laboratory, allowing rapid fixation and decalcification of BMB with preserved morphology and antigens. In this short report, we describe the use of a commercially available EDTA-based decalcification fluid (USEDECALC, Medite, Orlando, USA) in combination with the LOGOS J (Milestone, Bergamo, Italy), a closed microwave-enhanced tissue processor, for overnight fixation, decalcification, and paraffin impregnation of the BMB. This allows next-day reporting without impaired morphology or immunohistochemistry, and even improved DNA quality of the BMB.

Cost-effectiveness of Elective Endovascular Aneurysm Repair Versus Open Surgical Repair of Abdominal Aortic Aneurysms.

OBJECTIVE/BACKGROUND: The aim of this study was to estimate the lifetime cost-effectiveness of endovascular aneurysm repair (EVAR) versus open surgical repair (OSR) in the Netherlands, based on recently published literature. METHODS: A model was developed to simulate a cohort of individuals (age 72 years, 87% men) with an abdominal aortic aneurysm (AAA) diameter of at least 5.5 cm and considered fit for both repairs. The model consisted of two sub-models that estimated the lifetime cost-effectiveness of EVAR versus OSR: (1) a decision tree for the first 30 post-operative days; and (2) a Markov model for the period thereafter (31 days-30 years). RESULTS: In the base case analysis, EVAR was slightly more effective (4.704 vs. 4.669 quality adjusted life years) and less expensive (€24,483 vs. €25,595) than OSR. Improved effectiveness occurs because EVAR can reduce 30 day mortality risk, as well as the risk of events following the procedure, while lower costs are primarily due to a reduction in length of hospital stay. The cost-effectiveness of EVAR is highly dependent on the price of the EVAR device and the reduction in hospital stay, complications, and 30 day mortality. CONCLUSION: EVAR and OSR can be considered equally effective, while EVAR can be cost saving compared with OSR. EVAR can therefore be considered as a cost-effective solution for patients with AAAs.

Treatment variation in stent choice in patients with stable or unstable coronary artery disease.

AIM: Variations in treatment are the result of differences in demographic and clinical factors (e.g. anatomy), but physician and hospital factors may also contribute to treatment variation. The choice of treatment is considered important since it could lead to differences in long-term outcomes. This study explores the associations with stent choice: i.e. drug-eluting stent (DES) versus bare-metal stents (BMS) for Dutch patients diagnosed with stable or unstable coronary artery disease (CAD). METHODS & RESULTS: Associations with treatment decisions were based on a prospective cohort of 692 patients with stable or unstable CAD. Of those patients, 442 patients were treated with BMS or DES. Multiple logistic regression analyses were performed to identify variables associated with stent choice. Bivariate analyses showed that NYHA class, number of diseased vessels, previous percutaneous coronary intervention, smoking, diabetes, and the treating hospital were associated with stent type. After correcting for other associations the treating hospital remained significantly associated with stent type in the stable CAD population. CONCLUSIONS: This study showed that several factors were associated with stent choice. While patients generally appear to receive the most optimal stent given their clinical characteristics, stent choice seems partially determined by the treating hospital, which may lead to differences in long-term outcomes.

Disease-modifying antirheumatic drug-free sustained remission in rheumatoid arthritis: an increasingly achievable outcome with subsidence of disease symptoms.

OBJECTIVE: Disease-modifying antirheumatic drug (DMARD)-free sustained remission, the sustained absence of synovitis after cessation of DMARD therapy, is a relevant long-term outcome of rheumatoid arthritis (RA) if (1) its occurrence is promoted by treatment and (2) this status reflects resolution of symptoms and disability. This study investigated both items. METHODS: 1007 patients with RA diagnosed between 1993 and 2011, included in the Leiden Early Arthritis Clinic, were studied on achieving DMARD-free sustained remission. Patients included in 1993-1995 were initially treated with non-steroidal anti-inflammatory drugs, in 1996-1998 mild DMARDs were started early, from 1999 onwards methotrexate was initiated promptly and from 2005 onwards disease activity score (DAS)-steered treatment was common. Remission rates were compared using Kaplan-Meier curves and Cox proportional regression. RESULTS: In total, 155 patients achieved DMARD-free sustained remission. Specific treatment strategies were significantly associated with achieving remission (p<0.001). Cox regression adjusted for anticitrullinated protein antibody/rheumatoid factor, swollen joint count, erythrocyte sedimentation rate, C-reactive protein revealed HRs for DMARD-free sustained remission of 1.13 (95% CI 0.48 to 2.64) in patients diagnosed in 1996-1998, 2.39 (1.07 to 5.32) in patients treated with early methotrexate (inclusion 1999-2004) and 3.72 (1.60 to 8.62) in those treated early with methotrexate and DAS-steered therapy (inclusion 2005-2011). At the time of remission, the Health Assessment Questionnaire was at the level of the general population (median 0.13, IQR 0-0.63). Also, patient-rated visual analogue scale (VAS) morning stiffness, fatigue, pain and disease activity were low (median (IQR) mm, 14 (2-27), 10 (0-47), 6 (0-20), 7 (0-20), respectively). CONCLUSIONS: More intensive treatment strategies increased the chance for DMARD-free sustained remission, indicating that RA chronicity can be influenced. Patients with RA achieving DMARD-free sustained remission have a normalised functional status.

Is it cost-effective to use a test to decide which individuals with an intermediate cardiovascular disease risk would benefit from statin treatment?

BACKGROUND: The 2012 European guidelines recommend statins for intermediate-risk individuals with elevated cholesterol levels. Improved discrimination of intermediate-risk individuals is needed to prevent both cardiovascular disease (CVD) and statin side-effects (e.g. myopathy) efficiently since only 3-15 in every 100 individuals actually experience a cardiovascular event in the next 10 years. We estimated the potential cost-effectiveness of a hypothetical test which helps to determine which individuals will benefit from statins. METHODS AND RESULTS: Prognosis of different age- and gender-specific cohorts with an intermediate risk was simulated with a Markov model to estimate the potential costs and quality-adjusted life-years for four strategies: treat all with statins, treat none with statins, treat according to the European guidelines, or use a test to select individuals for statin treatment. The test-first strategy dominated the other strategies if the hypothetical test was 100% accurate and cost no more than €237. This strategy and the treat-all strategy were equally effective but the test generated lower costs by reducing statin usage and side-effects. The treat-none strategy was the least effective strategy. Threshold analyses show that the test must be highly accurate (especially sensitive) and inexpensive to be the most cost-effective strategy, since myopathy has a negligible impact on cost-effectiveness and statin costs are low. CONCLUSION: Use of a highly accurate prognostic test could reduce overall CVD risk, frequency of drug side-effects and lifetime costs. However, no additional test would add usefully to risk prediction over SCORE when it does not satisfy the costs and accuracy requirements.

Long-term outcome of rheumatoid arthritis defined according to the 2010-classification criteria.

OBJECTIVE: The 2010 American College of Rheumatology/European League Against Rheumatism (ACR/EULAR) criteria for rheumatoid arthritis (RA) have been thoroughly studied for the test characteristics but it is unclear whether '2010 RA' has a different phenotype than '1987 RA' when assessing the severity of the disease course. Therefore this study compared two long-term disease outcomes. METHODS: 1502 early arthritis patients that had no other diagnoses than RA or undifferentiated arthritis (UA) were studied on fulfilling the 1987 ACR criteria, 2010 criteria or both. The severity of joint damage was studied with yearly radiographs over 7 years. Achieving disease-modifying anti-rheumatic drug (DMARD)-free sustained remission was assessed over 10-years follow-up. Multivariate normal regression and Cox-proportional hazard regression were used, adjusting for age, gender and treatment. RESULTS: 550 patients fulfilled the 1987 criteria, 788 patients the 2010 criteria and 489 both criteria sets. Patients fulfilling the 2010 criteria developed less severe radiological joint damage (p=0.023) and achieved DMARD-free sustained remission more often (HR=1.18 (0.93-1.50)) than patients fulfilling the 1987 criteria, though the latter was not statistically significant. All 1987+2010- patients were anti-citrullinated peptide antibody (ACPA)-negative. When also applying the radiologic criterion of the 2010-criteria, half of the 1987+2010- patients became 2010 criteria positive, but results on the long-term outcome remained similar. CONCLUSIONS: '2010 RA' has a milder disease course than '1987 RA'. This may have important implications for basic scientific studies and clinical trials in RA.

Treatment costs of acute myocardial infarction in the Netherlands.

BACKGROUND: This study aimed to calculate the treatment costs of acute myocardial infarction (AMI) in the Netherlands for 2012. Also, the degree of association between treatment costs of AMI and some patient and hospital characteristics was examined. METHODS: For this retrospective cost analysis, patients were drawn from the database of the Diagnosis Treatment Combination (Diagnose Behandeling Combinatie, DBC) casemix system, which contains data on the resource use of all hospitalisations in the Netherlands. All costs were based on Euro 2012 cost data. RESULTS: The analysis was based on data of 25,657 patients. Mean treatment costs were estimated at 5021, with significant cost increases for patients with percutaneous coronary intervention (PCI) treatment. ST-segment elevation myocardial infarction (STEMI) patients receiving thrombolysis incurred the lowest ( 4286), while non-STEMI patients receiving PCI the highest costs ( 6060). Length of stay and hospital type were strong predictors of treatment costs. CONCLUSIONS: This study is the most extensive cost assessment of the treatment costs of AMI in the Netherlands thus far. Our results may be used as input for health-economic models and economic evaluations to support the decision making of registration, reimbursement and pricing of interventions in healthcare.

A systematic review and economic evaluation of new-generation computed tomography scanners for imaging in coronary artery disease and congenital heart disease: Somatom Definition Flash, Aquilion ONE, Brilliance iCT and Discovery CT750 HD.

BACKGROUND: Computed tomography (CT) is important in diagnosing and managing many conditions, including coronary artery disease (CAD) and congenital heart disease. Current CT scanners can very accurately diagnose CAD requiring revascularisation in most patients. However, imaging technologies have developed rapidly and new-generation computed tomography (NGCCT) scanners may benefit patients who are difficult to image (e.g. obese patients, patients with high or irregular heart beats and patients who have high levels of coronary calcium or a previous stent or bypass graft). OBJECTIVE: To assess the clinical effectiveness and cost-effectiveness of NGCCT for diagnosing clinically significant CAD in patients who are difficult to image using 64-slice computed tomography and treatment planning in complex congenital heart disease. DATA SOURCES: Bibliographic databases were searched from 2000 to February/March 2011, including MEDLINE, MEDLINE In-Process and Other Non-Indexed Citations, EMBASE, Cochrane Database of Systematic Reviews (CDSR), Cochrane Central Register of Controlled Trials (CENTRAL), Database of Abstracts of Reviews of Effects (DARE), NHS Economic Evaluation Database (NHS EED), Health Technology Assessment (HTA) database and Science Citation Index (SCI). Trial registers and conference proceedings were searched. REVIEW METHODS: Systematic review methods followed published guidance. Risk of bias was assessed using QUADAS-2. Results were stratified by patient group. Summary sensitivity and specificity were calculated using a bivariate summary receiver operating characteristic, or random effects model. Heterogeneity was assessed using the chi-squared statistic and I(2)-statistic. Cost-effectiveness of NGCCT was modelled separately for suspected and known CAD, evaluating invasive coronary angiography (ICA) only, ICA after positive NGCCT (NGCCT-ICA), and NGCCT only. The cost-effectiveness of NGCCT, compared with 64-slice CT, in reducing imaging-associated radiation in congenital heart disease was assessed. RESULTS: Twenty-four studies reported accuracy of NGCCT for diagnosing CAD in difficult-to-image patients. No clinical effectiveness studies of NGCCT in congenital heart disease were identified. The pooled per-patient estimates of sensitivity were 97.7% [95% confidence interval (CI) 88.0% to 99.9%], 97.7% (95% CI 93.2% to 99.3%) and 96.0% (95% CI 88.8% to 99.2%) for patients with arrhythmias, high heart rates and previous stent, respectively. The corresponding estimates of specificity were 81.7% (95% CI 71.6% to 89.4%), 86.3% (95% CI 80.2% to 90.7%) and 81.6% (95% CI 74.7% to 87.3%), respectively. In patients with high coronary calcium scores, previous bypass grafts or obesity, only per-segment or per-artery data were available. Sensitivity estimates remained high (> 90% in all but one study). In patients with suspected CAD, the NGCCT-only strategy appeared most cost-effective; the incremental cost-effectiveness ratio (ICER) of NGCCT-ICA compared with NGCCT only was £71,000. In patients with known CAD, the most cost-effective strategy was NGCCT-ICA (highest cost saving, dominates ICA only). The ICER of NGCCT only compared with NGCCT-ICA was £726,230. For radiation exposure only, the ICER for NGCCT compared with 64-slice CT in congenital heart disease ranged from £521,000 for the youngest patients to £90,000 for adults. LIMITATIONS: Available data were limited, particularly for obese patients and patients with previous bypass grafts. All studies of the accuracy of NGCCT assume that the reference standard (ICA) is 100% sensitive and specific; however, there is some evidence that ICA may sometimes underestimate the extent and severity of stenosis. Patients with more than one criterion that could contribute to difficulty in imaging were often excluded from studies; the effect on test accuracy of multiple difficult to image criteria remains uncertain. CONCLUSIONS: NGCCT may be sufficiently accurate to diagnose clinically significant CAD in some or all difficult-to-image patient groups. Economic analyses suggest that NGCCT is likely to be considered cost-effective for difficult-to-image patients with CAD, at current levels of willingness to pay in the NHS. For patients with suspected CAD, NGCCT only would be most favourable; for patients with known CAD, NGCCT-ICA would be most favourable. No studies assessing the effects of NGCCT on therapeutic decision making, or subsequent patient outcomes, were identified. The ideal study to address these questions would be a large multi-centre RCT. However, one possible alternative might be to establish a multicentre tracker study. High-quality test accuracy studies, particularly in obese patients, patients with high coronary calcium, and those with previous bypass grafts are needed to confirm the findings of our systematic review. These studies should include patients with multiple difficult to image criteria. FUNDING: The National Institute for Health Research Health Technology Assessment programme. This project was funded by the HTA programme, on behalf of NICE, as project number 10/107/01.

Three-dimensional reconstruction of tumor microvasculature: simultaneous visualization of multiple components in paraffin-embedded tissue.

Three-dimensional (3D) visualization of microscopic structures may provide useful information about the exact 3D configuration, and offers a useful tool to examine the spatial relationship between different components in tissues. A promising field for 3D investigation is the microvascular architecture in normal and pathological tissue, especially because pathological angiogenesis plays a key role in tumor growth and metastasis formation. This paper describes an improved method for 3D reconstruction of microvessels and other microscopic structures in transmitted light microscopy. Serial tissue sections were stained for the endothelial marker CD34 to highlight microvessels and corresponding images were selected and aligned. Alignment of stored images was further improved by automated non-rigid image registration, and automated segmentation of microvessels was performed. Using this technique, 3D reconstructions were produced of the vasculature of the normal brain. Also, to illustrate the complexity of tumor vasculature, 3D reconstructions of two brain tumors were performed: a hemangioblastoma and a glioblastoma multiforme. The possibility of multiple component visualization was shown in a 3D reconstruction of endothelium and pericytes of normal cerebellar cortex and a hemangioblastoma using alternate staining for CD34 and alpha-smooth muscle actin in serial sections, and of a GBM using immunohistochemical double staining. In conclusion, the described 3D reconstruction procedure provides a promising tool for simultaneous visualization of microscopic structures.

Cutaneous reflexes from the foot during gait in hereditary spastic paraparesis.

OBJECTIVE: It is known that P2 cutaneous reflexes from the foot show phase-dependent modulation during gait. The role of the motor cortex and the cortico-spinal tract in these reflexes and their modulation is unknown. Patients with hereditary spastic paraparesis (HSP) have a lesion in the cortico-spinal tract and may show deficits in P2 reflexes and/or their modulation. METHODS: Reflex responses of tibialis anterior and biceps femoris after sural nerve stimulation in 10 HSP-patients were compared with those in 10 healthy subjects. The reflexes were studied at two different moments in the step cycle during walking on a treadmill. RESULTS: Both patients and controls showed a phase-dependent modulation of P2 responses. For the individual muscles, no significant difference in reflex activity was observed between HSP-patients and the controls. However, when all muscles were taken together, the reflex activity for the controls was significantly higher than for the patients. CONCLUSIONS: The results of this study suggest that the cortico-spinal tract is involved in the regulation of the amplitude of the P2 responses and their phase-dependent modulation.

Three dimensional imaging of mammary ductal carcinoma in situ: clinical implications.

The conservation treatment of ductal carcinoma in situ (DCIS) is based on the surgical excision of the tumour together with irradiation of the remaining breast. Because short-term recurrence is almost certainly caused by residual tumour, an attempt should be made to verify the adequacy of the excision by assessing the specimen margin. The reliability of histologic margin assessment is influenced by the growth pattern of DCIS within the ductal tree and by the distance between tumour foci. Using an original stereoscopic technique, the present study of 60 mastectomy specimens shows that continuous and multifocal growth patterns are usual. A multifocal distribution (defined as gap of 4 cm or more between tumour foci) was found in only a single case. The growth pattern is related to DCIS type. Poorly-differentiated DCIS shows continuous growth, in contrast to the well-differentiated DCIS, which has a multicentric distribution. Irrespective of histologic type, however, only 8% of DCIS have a multifocal distribution with gaps greater than 10 mm. Therefore, with careful assessment, the likelihood of a false free margin seems theoretically low and should encourage the use of conserving treatment for eradicable DCIS.

Purification and characterization of rabbit pancreas protein kinase C.

Protein kinase C was purified 6,900-fold from rabbit pancreas with a total yield of 15% by a procedure involving ammonium sulfate fractionation, diethyl aminoethyl ion exchange chromatography, hydroxylapatite chromatography, and finally protamine-agarose affinity chromatography. After these purification steps the protein kinase C preparation contained two major protein bands as judged by silver staining after SDS-polyacrylamide gel electrophoresis: 80 and 69-kDa bands. Monoclonal antibodies directed against bovine brain protein kinase C (alpha- and beta-subtype) recognized only the 80-kDa band. On the other hand, both the 80 and 69-kDa proteins were recognized by a polyclonal monospecific antibody directed against rat brain protein kinase C. Analysis of rabbit pancreas protein kinase C subtypes by means of hydroxylapatite chromatography showed the presence of the III (alpha) subtype as the major subtype. The enzyme depended absolutely on the presence of both phosphatidylserine and Ca2+ for its activity, with apparent Ka values of 3.1 micrograms/ml and 247 microM for phosphatidylserine and Ca2+, respectively. When dioctanoylglycerol or the phorbol ester 12-O-tetradecanoyl-phorbol 13 acetate (TPA) was present, the Ka value for Ca2+ decreased to 10 and 18 microM, respectively. In the presence of the phorbol ester, pancreatic protein kinase C could be activated without added Ca2+. The enzyme also required Mg2+ for its activity. The Ka value was 3.6 mM and maximal activity was reached at 10 mM Mg2+. Pancreatic protein kinase C activity showed a broad pH dependence, with optimal activity at pH 6.75. The Km value for ATP and for histone-H1 was 8.5 microM and 20.4 micrograms/ml, respectively. The present study shows that the kinetic properties of protein kinase C purified from rabbit pancreas closely resemble those found in other tissues.

An original stereomicroscopic analysis of the mammary glandular tree.

In the 1970s, Wellings developed and reported extensively on a technique for a three-dimensional (3D) analysis of breast lesions. Drawbacks of this subgross sampling technique were that it was laborious, rather time-consuming and only allowed prospective studies. Furthermore, the stereomicroscopic aspect of the lesions studied was not diagnostic and each sample had to be studied histologically after paraffin embedding to determine the diagnosis. The present study introduces an original method enabling the exploration of the 3D structure of the mammary glandular tree from a paraffin-embedded sample. This procedure is quicker than the Wellings' technique, permits retrospective study and enables a 3D analysis of previously identified histological structures. Stereomicroscopic aspects of non-malignant lesions such as single multiple or metaplastic cysts, adenosis, ductal-lobular hyperplasia and malignant in situ neoplasms are illustrated. Our results confirm Wellings' concept that most minimal lesions arise in the terminal ductulo-lobular units. We also show that ductal carcinoma in situ may grow continuously by extending through the glandular tree but may also have a multifocal or stepwise progression in some cases.

An antigen resembling HL-A7 on the leukocytes of vervet monkeys.

Human leukocyte typing sera of known specificities were used to test the leukocyte antigens of vervet monkeys. The results suggest that these leukocytes contained an antigen resembling the HL-A7 antigen of human leukocytes. This is similar to a previous observation with leukocytes from baboons. These findings are consistent with the suggestion that the 4a/4b complex is the precursor substance from which the other specificities have evolved.