[Diabetes and hearing impairment due to mitochondrial mutation].

Mutations in the mitochondrial genome can cause mitochondrial diabetes. We present two cases in which the same mutation, mtDNA3243A>G, caused two different phenotypes: maternally inherited diabetes and deafness and mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes. Mitochondrial disease can imitate a variety of common con-ditions and should be considered in the case of multisystem disease, complex neurological symptoms or neurological symptoms combined with symptoms of other organ systems.

Pharmacist advice is accepted more for medical than for surgical patients in an emergency department.

INTRODUCTION: Pharmacists' advice may reduce medication errors in the emergency department (ED). However, pharmacists' recommendations are of little value if not acknowledged by physicians. The aim of the present study was to analyze how often and which categories of pharmacist recommendations were taken into account by the physicians in a Danish ED. Special attention is paid to problems of significant or vital importance. MATERIAL AND METHODS: Clinical pharmacists reviewed patient files within 24 hours of admission, described medication issues and made recommendations for solutions. It was subsequently noted whether the recommendations were taken into account by the physicians. Independent specialists in internal medicine and geriatrics reviewed the recommendations and assessed whether they were of minimal, moderate, significant or vital importance. RESULTS: Among the 301 recommendations made, 59% were followed by the physicians. The physician followed the recommendation made for patients admitted with medical problems significantly more often than the recommendations made for surgical patients (69% versus 51%, p = 0.002). In 47%, the recommendations were of significant or vital importance. Even these recommendations were acknowledged more by the physicians caring for medical patients than by physicians caring for surgical patients (78% versus 57%, p = 0.009). The difference remained significant in the multivariate analysis. CONCLUSION: Even though the pharmacists' recommendations were followed in many cases in an ED, there is still room for improvement, especially for the surgical patients. FUNDING: The Amgros Research Foundation covered the costs for the independent specialists who reviewed the patient files. TRIAL REGISTRATION: NCT01723462 (clinicaltrial.gov).

Medication problems are frequent and often serious in a Danish emergency department and may be discovered by clinical pharmacists.

INTRODUCTION: Transferring a patient from one health-care sector to another implies a risk of medication errors. It is of interest to evaluate whether a specialist in clinical pharmacy is beneficial for the patients in the emergency departments (ED). The aim of the present study was to report the incidence, categories and seriousness of medication problems discovered by clinical pharmacists in an ED and to evaluate if it is possible for pharmacists to identify those groups of patients who are most at risk of medication problems. MATERIAL AND METHODS: A pharmacist reviewed the patient files in the ED. If the pharmacists provided any kind of recommendations, a note was made describing the problem and a suggestion for a solution. After the study period, two medical specialists reviewed the files and rated the suggestions according to four levels of importance. RESULTS: A total of 1,696 patient files were reviewed after excluding patients who had received no medication. A total of 420 pharmacist notes were written, corresponding to 25% of all the included admissions. 47% of the pharmaceutical suggestions were considered serious. Increasing age and one drug as opposed to 2-9 drugs were associated with serious recommendations. In the multivariate analysis, only age above 70 years remained of significance for the identification of patients with a risk of a serious medication problem. CONCLUSION: A considerable amount of serious pharmaceutical problems were found in the ED. These problems had not been observed by the physicians and they were especially prevalent among the elderly and patients who were only prescribed a single drug. FUNDING: The Amgros research foundation financed salaries for the independent specialists who reviewed the patient files. TRIAL REGISTRATION: not relevant.

Activity as a tool in language training for immigrants and refugees.

OBJECTIVE: The focus of this study is how skills acquired from everyday life in one's native country can represent a resource in language training and work for immigrants and refugees. The specific aim is to explore what significance activity and participation in activity have on language training. METHODS: This qualitative study is based on fieldwork carried out in relation to a group of illiterate immigrants at a centre for adult education. The sample consists of 11 adult immigrants and refugees, male and female, between the ages of 20 and 65. The interviews with all the participants were carried out with the help of an interpreter. RESULTS: The main findings were that the individual immigrant's history of activities received little attention during the language training. There was hardly any mention of previous experience from everyday life and work. By relying on different activities in the language training, the resources and background of the individual immigrant would have become more visible. Familiar activities from one's own culture enable communication when language skills are limited.

Support for a bipolar affective disorder susceptibility locus on chromosome 12q24.3.

OBJECTIVE: Linkage and association studies of bipolar affective disorder (BAD) point out chromosome 12q24 as a region of interest. METHODS: To investigate this region further, we conducted an association study of 22 DNA markers within a 1.14 Mb region in a Danish sample of 166 patients with BAD and 311 control individuals. Two-hundred and four Danish patients with schizophrenia were also included in the study. RESULTS: We observed highly significant allelic and genotypic association between BAD and two highly correlated markers. The risk allele of both markers considered separately conferred an odds ratio of 2 to an individual carrying one risk allele and an odds ratio of 4 for individuals carrying both risk alleles assuming an additive genetic model. These findings were supported by the haplotype analysis. In addition, we obtained a replication of four markers associated with BAD in an earlier UK study. The most significantly associated marker was also analyzed in a Scottish case-control sample and was earlier associated with BAD in the UK cohort. The association of that particular marker was strongly associated with BAD in a meta-analysis of the Danish, Scottish and UK sample (P=0.0003). The chromosome region confined by our most distant markers is gene-poor and harbours only a few predicted genes. This study implicates the Slynar locus. We confirmed one annotated Slynar transcript and identified a novel transcript in human brain cDNA. CONCLUSION: This study confirms 12q24.3 as a region of functional importance in the pathogenesis of BAD and highlights the importance of focused genotyping.

Further immunohistochemical characterization of BRD1 a new susceptibility gene for schizophrenia and bipolar affective disorder.

We have recently shown that the gene BRD1 is associated with schizophrenia and bipolar affective disorder and that the BRD1 protein (BRD1) which is expressed in neurons may occur in a short and a long variant. The aim of the study was to generate polyclonal antibodies against new BRD1 epitopes enabling discrimination between the long and short BRD1 variants, and elucidate the BRD1 distribution in several human tissues, including the CNS. Polyclonal rabbit antibodies were raised against three different BRD1 epitopes. One (67) was specific for the long BRD1 variant, whereas the two others (63/64 and 65/66) like the original monoclonal mouse antibody (K22) were predicted to stain both variants. Immunohistochemical staining procedures were subsequently performed on paraffin-embedded human cerebral cortex and microarray slides containing 30 different human tissues. Western blotting confirmed the predicted specificity of the developed antibodies. K22, 63/64 and 65/66 displayed a similar neuronal staining pattern characterized by a distinct but weak nuclear staining, while the surrounding cytoplasm and proximal dendrites were more intensely stained. Interestingly, staining with 67 generated in contrast primarily an intense nuclear staining. The new antibodies resulted, furthermore, in a prominent neuroglial reaction characterized by staining of cell bodies, nuclei and glial processes. The tissue microarray analysis revealed that BRD1 was widely distributed in human tissues. The particular expression profile, e.g., the degree of nuclear and/or cytoplasmatic staining, seemed, however, to be highly tissue dependent. These results suggest a general role of BRD1 in the cell and stress that the two BRD1 variants may play different roles in the etiology of psychiatric disease.

Dynamic regulation of integrin activation by intracellular and extracellular signals controls oligodendrocyte morphology.

BACKGROUND: Myelination requires precise control of oligodendrocyte morphology and myelin generation at each of the axons contacted by an individual cell. This control must involve the integration of extracellular cues, such as those on the axon surface, with intrinsic developmental programmes. We asked whether integrins represent one class of oligodendrocyte cell-surface receptors able to provide this integration. RESULTS: Integrins signal via a process of activation, a conformational change that can be induced either by "outside-in" signals comprising physiological extracellular matrix ligands (mimicked by the pharmacological use of the divalent cation manganese) or "inside-out" signalling molecules such as R-Ras. Increasing levels of outside-in signalling via the laminin receptor alpha6beta1 integrin were found to promote oligodendrocyte processing and myelin sheet formation in culture. Similar results were obtained when inside-out signalling was increased by the expression of a constitutively-active R-Ras. Inhibiting inside-out signalling by using dominant-negative R-Ras reduces processes and myelin sheets; importantly, this can be partially rescued by the co-stimulation of outside-in signalling using manganese. CONCLUSION: The balance of the equilibrium between active and inactive integrins regulates oligodendrocyte morphology, which is itself regulated by extrinsic and intrinsic cues so providing a mechanism of signal integration. As laminins capable of providing outside-in signals are present on axons at the time of myelination, a mechanism exists by which morphology and myelin generation might be regulated independently in each oligodendrocyte process.

Integrins direct Src family kinases to regulate distinct phases of oligodendrocyte development.

Specific integrins expressed on oligodendrocytes, the myelin-forming cells of the central nervous system, promote either differentiation and survival or proliferation by amplification of growth factor signaling. Here, we report that the Src family kinases (SFKs) Fyn and Lyn regulate each of these distinct integrin-driven behaviors. Fyn associates with alpha6beta1 and is required to amplify platelet-derived growth factor survival signaling, to promote myelin membrane formation, and to switch neuregulin signaling from a phosphatidylinositol 3-kinase to a mitogen-activated protein kinase pathway (thereby changing the response from proliferation to differentiation). However, earlier in the lineage Lyn, not Fyn, is required to drive alphaVbeta3-dependent progenitor proliferation. The two SFKs respond to integrin ligation by different mechanisms: Lyn, by increased autophosphorylation of a catalytic tyrosine; and Fyn, by reduced Csk phosphorylation of the inhibitory COOH-terminal tyrosine. These findings illustrate how different SFKs can act as effectors for specific cell responses during development within a single cell lineage, and, furthermore, provide a molecular mechanism to explain similar region-specific hypomyelination in laminin- and Fyn-deficient mice.

Inhibition of proteolysis by the proform of eosinophil major basic protein (proMBP) requires covalent binding to its target proteinase.

By proteolytic cleavage of insulin-like growth factor binding proteins, the metalloproteinase pregnancy-associated plasma protein-A (PAPP-A) is able to control the biological activity of insulin-like growth factors. PAPP-A circulates in pregnancy as a proteolytically inactive complex, disulfide bound to the proform of eosinophil major basic protein (proMBP). We here demonstrate that co-transfection of mammalian cells with PAPP-A and proMBP cDNA results in the formation of a covalent PAPP-A/proMBP complex in which PAPP-A is inhibited. Formation of the complex also occurs when PAPP-A and proMBP synthesized separately are incubated. Complex formation was monitored by Western blotting, and by using an immunoassay specific for the complex. Using mutagenesis, we further demonstrate that the complex forms in a specific manner and depends on the presence of two proMBP cysteine residues. Mutated proMBP, in which Cys-51 and -169 are replaced by serine, is unable to form the covalent complex with PAPP-A. Of particular interest, such mutated proMBP further lacks the ability to inhibit PAPP-A. For the first time, this conclusively demonstrates that proMBP is a proteinase inhibitor. We further conclude that proMBP inhibits PAPP-A in an unusual manner, not paralleled by other proteinase inhibitors of our knowledge, which requires proMBP to be covalently bound to PAPP-A by disulfide bonds. ProMBP binding to PAPP-A most likely either abrogates substrate access to the active site of PAPP-A or induces a conformational change in the structure of PAPP-A, as we, by further mutagenesis, were able to exclude that the inhibitory mechanism of proMBP is based on a cysteine switch-like mechanism.

Expression of recombinant murine pregnancy-associated plasma protein-A (PAPP-A) and a novel variant (PAPP-Ai) with differential proteolytic activity.

Murine pregnancy-associated plasma protein-A (PAPP-A) cDNA encoding a 1545 amino-acid protein has been cloned. We have also identified and cloned cDNA that encodes a novel variant of PAPP-A, PAPP-Ai, carrying a 29-residue highly basic insert. The point of insertion corresponds to a junction between two exons in the human PAPP-A gene. The human intron flanked by these exons does not encode a homologous corresponding insert, which is unique to the mouse. The overall sequence identity between murine and human PAPP-A is 91%, and murine PAPP-A contains sequence motifs previously described in the sequence of human PAPP-A. Through expression in mammalian cells, we show that murine PAPP-A and PAPP-Ai are active metalloproteinases, both capable of cleaving insulin-like growth factor binding protein (IGFBP)-4 and -5. Cleavage of IGFBP-4 is dramatically enhanced by the addition of IGF, whereas cleavage of IGFBP-5 is slightly inhibited by IGF, as previously established with human PAPP-A. Surprisingly, however, quantitative analyses demonstrate that the murine PAPP-Ai cleaves IGFBP-4 very slowly compared to PAPP-A, even though its ability to cleave IGFBP-5 is unaffected by the presence of the insert. By RT-PCR analysis, we find that both variants are expressed in several tissues. The level of mRNA in the murine placenta does not exceed the levels of other tissues analyzed. Furthermore, the IGFBP-4-proteolytic activity of murine pregnancy serum is not elevated. This is in striking contrast to the increase seen in human pregnancy serum, and the expression of PAPP-A in the human placenta, which exceeds other tissues at least 250-fold. Interestingly, the position of the insert of PAPP-Ai, within the proteolytic domain, lies in close proximity to the cysteine residue, which in human PAPP-A forms a disulfide bond with the proform of eosinophil major basic protein (proMBP). ProMBP functions as a proteinase inhibitor in the PAPP-A-proMBP complex, but whether any mechanistic parallel on regulation of proteolytic activity can be drawn between the insert of PAPP-Ai and the linkage to proMBP is not known. Importantly, these data support the development of the mouse as a model organism for the study of PAPP-A, which must take into account the differences between the mouse and the human.