Insulin degludec does not increase antibody formation versus insulin glargine: an evaluation of phase IIIa trials.

We examined insulin antibody formation in patients with type 1 (T1D) or type 2 diabetes (T2D) treated with once-daily insulin degludec (IDeg) or insulin glargine (IGlar) to evaluate the impact of antibody formation on efficacy and safety. Insulin antibodies were measured using subtraction radioimmunoassays in six phase IIIa clinical trials using IDeg (n = 2250) and IGlar (n = 1184). Spearman's correlation coefficient was used to evaluate associations between cross-reacting antibodies and change from baseline glycated haemoglobin (HbA1c) and insulin dose. IDeg- and IGlar-specific antibodies remained low [<1% bound/total radioactivity (B/T)] and with low levels of antibodies cross-reacting with human insulin in patients with T1D (<20% B/T) and T2D (<6% B/T). Spearman's correlation coefficients between insulin antibody levels and change in HbA1c or insulin dose were low in both treatment groups. No clinically meaningful differences in adverse event (AE) rates were observed in patients with >10% B/T or without an absolute increase in antibodies cross-reacting with human insulin. IDeg treatment resulted in few immunogenic responses in patients with T1D and T2D; antibody formation was not associated with change in HbA1c, insulin dose or rates of AEs.

Effects and barriers to deployment of telehealth wellness programs for chronic patients across 3 European countries.

BACKGROUND: Benefits of cardiopulmonary rehabilitation (CPR) in patients with chronic obstructive pulmonary disease (COPD) are well established, but long-term sustainability of training-induced effects and its translation into healthy lifestyles are unsolved issues. It is hypothesized that Integrated Care Services supported by Information and Communication Technologies (ICS-ICT) can overcome such limitations. In the current study, we explored 3 ICS-ICT deployment experiences conducted in Barcelona, Trondheim and Athens. METHODS: In the 3 sites, a total of 154 patients completed an 8-week supervised CPR program. Thereafter, they were allocated either to an ICS-ICT group or to usual care (CPR + UC) during a follow-up period of at least 12 months with assessment of 6-min walking test (6MWT) as main outcome variable at all time points in the 3 sites. Because real deployment was prioritized, the interventions were adapted to site heterogeneities. RESULTS: In the ICS-ICT group from Barcelona (n = 77), the use of the personal health folder (PHF) was the cornerstone technological tool to empower COPD patients for self-management showing high applicability and user-acceptance. Long-term sustainability of training-induced increase in exercise capacity was observed in ICS-ICT compared to the control group (p = 0.01). Likewise, ICS-ICT enhanced the activities domain of the SGRQ (p < 0.01) and daily physical activity (p = 0.03), not seen in controls. No effects of ICS-ICT were observed in Trondheim (n = 37), nor in Athens (n = 40), due to technological and/or organizational limitations. CONCLUSIONS: The study results suggest the potential of the ICS-ICT Barcelona's approach to enhance COPD management. Moreover, it allowed identification of the factors limiting transferability to the other sites. The research prompts the need for large multicenter trials specifically designed to assess effectiveness, efficiencies and transferability of this type of intervention.

The urokinase plasminogen activator receptor-associated protein/endo180 is coexpressed with its interaction partners urokinase plasminogen activator receptor and matrix metalloprotease-13 during osteogenesis.

The urokinase plasminogen activator receptor-associated protein/Endo180 (uPARAP/Endo180) is a newly discovered member of the macrophage mannose receptor family that was reported to interact with ligand-bound urokinase plasminogen activator receptor (uPAR), matrix metalloprotease-13 (MMP-13), and collagen V on the cell surface. We have determined the sites of expression of this novel receptor during murine postimplantation development. uPARAP/Endo180 was expressed in all tissues undergoing primary ossification, including the developing bones of the viscerocranium and calvarium that ossify intramembranously, and developing long bones undergoing endochondral ossification. uPARAP/Endo180 mRNA was expressed by both immature osteoblasts and by mature osteocalcin-producing osteoblasts-osteocytes, and was coexpressed with MMP-13. Interestingly, osteoblasts also expressed uPAR. Besides bone-forming tissues, uPARAP/Endo180 expression was detected only in a mesenchymal condensation of the midbrain and in the developing lungs. The data suggest a function of this novel protease receptor in bone development, possibly mediated through its interactions with uPAR, MMP-13, or collagen V.

The murine receptor for urokinase-type plasminogen activator is primarily expressed in tissues actively undergoing remodeling.

uPAR is a cellular receptor for urokinase plasminogen activator, an enzyme involved in extracellular matrix degradation during processes involving tissue remodeling. We have expressed a recombinant soluble form of murine uPAR and raised rabbit polyclonal antibodies to study the expression of uPAR by immunohistochemistry. The immunohistochemical localization of uPAR was determined in normal mouse organs and in tumors formed by the highly metastatic Lewis lung carcinoma. uPAR immunoreactivity was found in the lungs, kidneys, and spleen, and in endothelial cells in the uterus, urinary bladder, thymus, heart, liver, and testis. No uPAR immunoreactivity was detected in muscle. In general, strong uPAR immunoreactivity was observed in organs undergoing extensive tissue remodeling, as exemplified by trophoblast cells in placenta, and in migrating, but not resting, keratinocytes at the edge of incisional wounds. Staining was not detected in any tissue sections derived from uPAR-deficient mice, thus confirming the specificity of the immunohistochemical staining of uPAR in normal mouse tissues. In Lewis lung carcinoma, uPAR immunoreactivity was found in the tumor cells of the primary tumor and in lung metastases. (J Histochem Cytochem 49:237-246, 2001)

Lactational competence and involution of the mouse mammary gland require plasminogen.

Urokinase-type plasminogen activator expression is induced in the mouse mammary gland during development and post-lactational involution. We now show that primiparous plasminogen-deficient (Plg(-/-)) mice have seriously compromised mammary gland development and involution. All mammary glands were underdeveloped and one-quarter of the mice failed to lactate. Although the glands from lactating Plg(-/-) mice were initially smaller, they failed to involute after weaning, and in most cases they failed to support a second litter. Alveolar regression was markedly reduced and a fibrotic stroma accumulated in Plg(-/-) mice. Nevertheless, urokinase and matrix metalloproteinases (MMPs) were upregulated normally in involuting glands of Plg(-/-) mice, and fibrin did not accumulate in the glands. Heterozygous Plg(+/-) mice exhibited haploinsufficiency, with a definite, but less severe mammary phenotype. These data demonstrate a critical, dose-dependent requirement for Plg in lactational differentiation and mammary gland remodeling during involution.

Generation of high-affinity rabbit polyclonal antibodies to the murine urokinase receptor using DNA immunization.

The urokinase receptor (uPAR) is a glycolipid anchored cell surface glycoprotein that plays a central role in extracellular proteolysis during tissue remodeling processes and cancer invasion. By intramuscular (i.m.) injection of rabbits with plasmid DNA coding for a carboxy-terminally truncated secreted form of the murine uPAR (muPAR), specific anti-sera with a titer of 64,000, as measured by ELISA, have been obtained. Rabbits received a total of 10 monthly injections of 1 mg DNA in phosphate-buffered saline. The antibody titer peaked between the 5th and 7th injection and slowly declined after the 8th injection. After the final immunization the immune response persisted for at least 6 months without further injections. The antibodies generated by DNA immunization were useful for immunohistochemistry and immunoblotting, recognizing the antigen both in its native and in its reduced and alkylated form. Using the antibodies in immunoblotting muPAR was identified in lysates of peritoneal macrophages, spleen and lung tissue. Both the intact and cleaved form of muPAR were identified in lysates of a murine monocyte cell line P388D.1. No cross-reaction with human uPAR was observed. In immunohistochemical analysis of normal mouse lung tissue uPAR immunoreactivity was located in the alveoli and pulmonary vessels, whereas the bronchial epithelium was negative. These results demonstrate that DNA immunization of rabbits using i.m. injection is a very effective and easy method to raise polyclonal antibodies which can be used for characterization and localization of muPAR in mouse tissue.

Open laboratory information systems: a case study.

The EU-AIM project "OpenLabs" specified an open systems architecture for a clinical laboratory information system (LIS) environment and coined the term "open LIS". Since this conception has not yet been realized into systems and modules commercially available, two university hospitals in Norway had to specify an alternative model for openness when they, in a joint project, screened the market for a new LIS. To be open, a LIS must be (1) based on main-trend hardware and system software running under a standardized operating system, (2) designed for network-based distributed computing, (3) have sufficient documentation available to local staff, (4) be configurable through parameters, tables and scripts, and (5) have general and configurable access mechanisms both to real-time processes and to the database. The present study describes the selection process and examines the compliance of the selected LIS (NetLab) with this model of openness. It is shown that local staff have been able to adapt this LIS to various and changing requirements in the laboratory.

Methods for chemical analysis of contaminated soil samples--tests of their reproducibility between Nordic laboratories.

In an effort to develop common analytical methods for contaminated soil samples the Environmental Authorities of the Nordic countries have, together with Nordtest, published the report Nordic Guidelines for Chemical Analysis of Contaminated Soil Samples. The aim of these guidelines has been to describe analytical methods which could be accepted in all the Nordic countries and in that way contribute to reducing the variation in the analytical results between laboratories. The methods covered, reflects environmental concerns and priorities in the Nordic countries for now, i.e. heavy metals, chlorophenols, creosote, volatile organic compounds, PCB, THC and PAH. The repeatability and reproducibility of the guideline methods were determined in a Nordic inter-laboratory test in 1996, and the results showed some variations. The analytical methods and the results from the inter-laboratory tests are given for heavy metals, chlorophenols, creosote, volatile organic compounds and PCB.

[Sedimentation rate and renal cancer]. / Senkningsreaksjon og nyrekreft.

This paper is based on two articles published in 1996 in Journal of Internal Medicine. One is an evaluation of the erythrocyte sedimentation rate (ESR) measured in 3910 subjectively healthy Norwegians of both sexes, with an age range of 20 to 90 years. The authors underline the difference between a population-based and a subject-based upper ESR reference level, and the benefit of knowing each single healthy person's subject-based ESR level. The other article shows that, in 70% of a sample of 236 randomly selected patients with renal cell carcinoma, a diagnosis could have been indicated several years before clinical symptoms or signs became apparent. Therefore, the ESR might be a predictive test for many such cancer cases. This type of cancer has a good prognosis when treated surgically at an early stage. In Norway, 432 new cases occurred in 1993, 264 in men and 168 in women. If our observations are generally applicable, and proper attention had been paid to a slowly increasing ESR value over time, about 300 of these cases could have been operated on much earlier. It is suggested that physicians and public might both benefit from this knowledge.

Population-based erythrocyte sedimentation rates in 3910 subjectively healthy Norwegian adults. A statistical study based on men and women from the Oslo area.

OBJECTIVES: To establish age- and sex-specific reference limits for the erythrocyte sedimentation rate (ESR) in asymptomatic Norwegian adults. DESIGN: Single ESR recordings were obtained by the classical or a modified Westergren method from 2145 men and 1765 women (93% being blood donors) with age range 20-90 years, and analysed statistically. RESULTS: There was a significant positive association between ESR level and age, consistent with a parabolic pattern in men but a linear one in women. The mean values for men were about 3 mm h-1 at 20 years, 6 mm h-1 at 55 years, and 10 mm h-1 at 90 years, and 6, 9, and 11 mm h-1 respectively for women. These averages (predicted by regression lines) were significantly higher in women up to the age of 75 years, after which the estimated sex-specific 95% confidence limits for mean values were found to overlap. CONCLUSIONS: The upper reference levels expected to be exceeded only by chance in 5% of single individual recordings at the ages of 20, 55 or 90 years, respectively, were estimated to be 12, 14 and 19 mm h-1 for men, and 18, 21 and 23 mm h-1 for women. Higher values should be controlled and, if confirmed, lead to a clinical check-up. However, about 76% of our overall material had ESR values lower than 9 mm h-1. Knowledge of each person's baseline ESR value might increase the disease-predictive ability of the test. If several measurements over years reveal a steeper rise with age than depicted in our population-based curves, it should be taken seriously, even when each reading is below the population-based reference limits.

Rising erythrocyte sedimentation rate during several years before diagnosis can be a predictive factor in 70% of renal cell carcinoma patients. The benefit of knowing subject-based reference values.

OBJECTIVES: A diagnosis of renal cell carcinoma (RCC) early enough for potentially curative surgery is difficult. We wanted to establish whether the erythrocyte sedimentation rate (ESR) in RCC patients had begun to rise before the appearance of any symptoms or signs and, if so, when. DESIGN: A retrospective study of the evolution of the ESR in 236 randomly selected RCC patients during several years before diagnosis, comparing the results with previously obtained population-based control values. RESULTS: It is generally held that RCC patients have a high ESR at diagnosis. In our material, however, 29.7% of the RCC cases had an ESR that at this time was at or below the population-based upper reference limit; it had not increased significantly, neither with time before diagnosis, nor with age. In 70.3% of RCC patients the ESR was increased and had been significantly rising for up to 6 years or more before diagnosis. This had not been adequately responded to, probably because the physicians lacked knowledge of the patients' baseline ESR, and because none of the prediagnostic readings had been above the population-based reference limit. CONCLUSIONS: Systematic ESR graphic recordings over time will enable a physician to determine each individual's baseline value, and hence note any continuously rising trend, which should lead to further investigations, e.g. an ultrasound kidney examination. This may provide an early clue to many otherwise non-symptomatic RCC cases. It is time for a reappraisal of the predictive value of the ESR to discover early RCC, and possibly other diseases as well.

The CCL conferences revisited.

The history of the ten conferences on Computing in Clinical Laboratories (CCL) since the first one in Birmingham, UK, 1975, mirrors the developments in medical laboratory computing during nearly twenty years.

Two distinct phases of apoptosis in mammary gland involution: proteinase-independent and -dependent pathways.

Postlactational involution of the mammary gland is characterized by two distinct physiological events: apoptosis of the secretory, epithelial cells undergoing programmed cell death, and proteolytic degradation of the mammary gland basement membrane. We examined the spatial and temporal patterns of apoptotic cells in relation to those of proteinases during involution of the BALB/c mouse mammary gland. Apoptosis was almost absent during lactation but became evident at day 2 of involution, when beta-casein gene expression was still high. Apoptotic cells were then seen at least up to day 8 of involution, when beta-casein gene expression was being extinguished. Expression of sulfated glycoprotein-2 (SGP-2), interleukin-1 beta converting enzyme (ICE) and tissue inhibitor of metalloproteinases-1 was upregulated at day 2, when apoptotic cells were seen initially. Expression of the matrix metalloproteinases gelatinase A and stromelysin-1 and the serine proteinase urokinase-type plasminogen activator, which was low during lactation, was strongly upregulated in parallel starting at day 4 after weaning, coinciding with start of the collapse of the lobulo-alveolar structures and the intensive tissue remodeling in involution. The major sites of mRNA synthesis for these proteinases were fibroblast-like cells in the periductal stroma and stromal cells surrounding the collapsed alveoli, suggesting that the degradative phase of involution is due to a specialized mesenchymal-epithelial interaction. To elucidate the functional role of these proteinases during involution, at the onset of weaning we treated mice systemically with the glucocorticoid hydrocortisone, which is known to inhibit mammary gland involution. Although the initial wave of apoptotic cells appeared in the lumina of the gland, the dramatic regression and tissue remodeling usually evident by day 5 was substantially inhibited by systemic treatment with hydrocortisone. mRNA and protein for gelatinase A, stromelysin-1 and uPA were weakly induced, if at all, in hydrocortisone-treated mice. Furthermore, mRNA for membrane-type matrix metalloproteinase decreased after hydrocortisone treatment and paralleled the almost complete inhibition of activation of latent gelatinase A. Concomitantly, the gland filled with an overabundance of milk. Our data support the hypothesis that there are at least two distinct phases of involution: an initial phase, characterized by induction of the apoptosis-associated genes SGP-2 and ICE and apoptosis of fully differentiated mammary epithelial cells without visible degradation of the extracellular matrix, and a second phase, characterized by extracellular matrix remodeling and altered mesenchymal-epithelial interactions, followed by apoptosis of cells that are losing differentiated functions.

RefVal: a program implementing the recommendations of the International Federation of Clinical Chemistry on the statistical treatment of reference values.

RefVal is a computer program that implements the recommendations of the International Federation of Clinical Chemistry on the statistical treatment of reference values. The program performs the following main tasks: graphical display of the distribution of reference values, identification or elimination of outliers, testing of the fit of the distribution to Gaussian shape (coefficients of skewness and kurtosis, Anderson-Darling's test, Cramér-von Mises' test, Kolmogorov-Smirnov's test), non-parametric and parametric estimation of reference limits (fractiles, percentiles). The parametric estimation method is based on a two-stage mathematical transformation of data: (1) Manly's exponential transformation (to remove skewness) and (2) John and Draper's modulus transformation (to adjust for remaining kurtosis). The program exists in different versions. The paper describes two of these: (1) a library of FORTRAN functions and subroutines and (2) a Pascal PC program that runs under MS-DOS.

The receptor for urokinase-type plasminogen activator is not essential for mouse development or fertility.

The urokinase-type plasminogen activator receptor (uPAR) gene was disrupted in mice in order to explore the role of cell surface-associated plasminogen activation in development and hemostasis. Homozygous, uPAR-/- mice were born and survived to adulthood with no overt phenotypic abnormalities. There was no indication of loss of fetal animals based on the Mendelian pattern of transmission of the mutant uPAR gene. uPAR-/- mice carried no detectable uPAR in lung, spleen, and other tissues when measured both immunologically by Western blot analysis and functionally by ligand cross-linking analyses. In addition, activated peritoneal macrophages collected from uPAR-/- mice failed to promote plasminogen activation in vitro. The loss of the receptor also resulted in a redistribution of uPA in some tissues but had no impact on pro-uPA activation in the urogenital tract. Thus, in the absence of other challenging factors such as infection, injury, or other functional deficits, uPAR deficiency does not compromise fertility, development, or hemostasis. These mice provide a means to test the proposed function of uPA/uPAR in wound repair, atherogenesis, and tumor cell invasion in vivo.

Multivariate reference regions.

A high frequency of "false positive" conclusions is often the outcome when several laboratory test results are interpreted by comparing each single test result with the corresponding reference interval, i.e., by multiple univariate comparisons. The adequate solution to this problem is the application of a multivariate reference region for the test profile. This region is a straightforward extension of the univariate reference interval to the multidimensional situation.

Subject-based reference values.

Medical decisions based on comparison with group-based reference value have often less than desired sensitivity to significant changes in the biochemical or physiological state of an individual under investigation. Subgrouping of the reference values according to sex, age, or other criteria does frequently not solve this problem. The better solution is to compare new values with old results from the same individual, i.e., with subject-based reference values. Two types of criteria may be used: those based on critical differences between two results (the reference change limit) and those based on time-series models.

A cleaved form of the receptor for urokinase-type plasminogen activator in invasive transplanted human and murine tumors.

It was recently found that urokinase-type plasminogen activator (uPA) is involved in the cleavage of its receptor (uPAR) on cultured cells, thereby releasing one of the receptor's 3 domains (the ligand binding domain I) and leaving the 2 others [uPAR(2 + 3)] anchored to the cell surface. With monoclonal antibodies (MAbs) we have now identified human uPAR(2 + 3) in lysates of invasive human MDA-MB-231 mammary carcinomas xenografted into nude mice. The production of peptide antibodies recognizing different domains of murine uPAR made it possible to identify a similar cleaved form of uPAR, murine uPAR(2 + 3), in extracts of primary Lewis lung carcinomas. Cleavage of uPAR also occurs in cultured MDA-MB-231 cells and Lewis lung carcinoma cells. This cleavage is inhibited by anticatalytic antibodies to either human or murine uPA, respectively, indicating that it is catalyzed by either uPA or plasmin generated by uPA. The amount of uPAR(2 + 3) may therefore be directly related to the activity of the uPA system and it is possible that the level of uPAR(2 + 3) in cancer tissue may prove to be a stronger prognostic parameter than the levels of either full-length uPAR or UPA.

RefVal for windows--migrating to the Windows environment with the help of Visual Basic.

REFVAL is a program which implements the statistical treatment of reference values as recommended by the IFCC Expert Panel on Theory of Reference Values. The authors have used Microsoft Visual Basic to produce a new version for the Window environment.