Combined fused in sarcoma-positive (FUS+) basophilic inclusion body disease and atypical tauopathy presenting with an amyotrophic lateral sclerosis/motor neurone disease (ALS/MND)-plus phenotype.

AIMS: Amyotrophic lateral sclerosis/motor neurone disease (ALS/MND) is characterized by the presence of inclusions containing TDP-43 within motor neurones. In rare cases, ALS/MND may be associated with inclusions containing other proteins, such as fused in sarcoma (FUS), while motor system pathology may rarely be a feature of other neurodegenerative disorders. We here have investigated the association of FUS and tau pathology. METHODS: We report a case with an ALS/MND-plus clinical syndrome which pathologically demonstrated both FUS pathology and an atypical tauopathy. RESULTS: Clinical motor involvement was predominantly present in the upper motor neurone, and was accompanied by extrapyramidal features and sensory involvement, but with only minimal cognitive impairment. The presentation was sporadic and gene mutation screening was negative. Post mortem study demonstrated inclusions positive for FUS, including basophilic inclusion bodies. This was associated with 4R-tauopathy, largely as non-fibrillary diffuse phospho-tau in neurones, with granulovacuolar degeneration in a more restricted distribution. Double-staining revealed that neurones contained both types of protein pathology. CONCLUSION: FUS-positive basophilic inclusion body disease is a rare cause of ALS/MND, but in this case was associated with an unusual atypical tauopathy. The coexistence of two such rare neuropathologies raises the question of a pathogenic interaction.

Association of placental weight with cerebral palsy: population-based cohort study in Norway.

OBJECTIVE: To study the risk of cerebral palsy (CP) associated with placental weight, and also with placental weight/birthweight ratio and placental weight/birth length ratio. DESIGN: Population-based cohort study. SETTING: Perinatal data in the Medical Birth Registry of Norway were linked with clinical data in the CP Register of Norway. POPULATION: A total of 533 743 singleton liveborn children in Norway during 1999-2008. Of these, 779 children were diagnosed with CP. METHODS: Placental weight, placental weight/birthweight ratio, and placental weight/birth length ratio were grouped into gestational age-specific quartiles. Odds ratios (OR) with 95% confidence intervals (95% CI) for CP were calculated for children with exposure variables in the lowest or in the highest quartile, using the second to third quartile as the reference. MAIN OUTCOME MEASURES: CP and CP subtypes. RESULTS: Overall, children with low placental weight had increased risk for CP (OR 1.5, 95% CI 1.2-1.7). Low placental weight/birthweight ratio (OR 1.2, 95% CI 1.0-1.4) and low placental weight/birth length ratio (OR 1.5, 95% CI 1.2-1.8) were also associated with increased risk for CP. In children born at term, low placental weight was associated with a twofold increase in risk for spastic bilateral CP (including both quadriplegia and diplegia) (OR 2.1, 95% CI 1.5-2.9). In children born preterm, high placental ratios were associated with increased risk for spastic quadriplegia. CONCLUSIONS: Our results suggest that placental dysfunction may be involved in causal pathways leading to the more severe subtypes of CP. TWEETABLE ABSTRACT: Low placental weight increases the risk for cerebral palsy, especially for the spastic bilateral subtype.

Activation of adult rat CNS endothelial cells by opioid-induced toll-like receptor 4 (TLR4) signaling induces proinflammatory, biochemical, morphological, and behavioral sequelae.

CNS immune signaling contributes to deleterious opioid effects including hyperalgesia, tolerance, reward, and dependence/withdrawal. Such effects are mediated by opioid signaling at toll-like receptor 4 (TLR4), presumptively of glial origin. Whether CNS endothelial cells express TLR4 is controversial. If so, they would be well positioned for activation by blood-borne opioids, contributing to opioid-induced pro-inflammatory responses. These studies examined adult primary rat CNS endothelial cell responses to (-)-morphine or its mu opioid receptor (MOR)-inactive metabolite morphine-3-glucuronide (M3G), both known TLR4 agonists. We demonstrate that adult rat CNS endothelial cells express functional TLR4. M3G activated nuclear factor kappaB (NF-κB), increased tumor necrosis factor-α (TNFα) and cyclooxygenase-2 (COX2) mRNAs, and released prostaglandin E2 (PGE2) from these cells. (-)-Morphine-induced upregulation of TNFα mRNA and PGE2 release were unmasked by pre-treatment with nalmefene, a MOR antagonist without TLR4 activity (unlike CTAP, shown to have both MOR- and TLR4-activity), suggestive of an interplay between MOR and TLR4 co-activation by (-)-morphine. In support, MOR-dependent Protein Kinase A (PKA) opposed TLR4 signaling, as PKA inhibition (H-89) also unmasked (-)-morphine-induced TNFα and COX2 mRNA upregulation. Intrathecal injection of CNS endothelial cells, stimulated in vitro with M3G, produced TLR4-dependent tactile allodynia. Further, cortical suffusion with M3G in vivo induced TLR4-dependent vasodilation. Finally, endothelial cell TLR4 activation by lipopolysaccharide and/or M3G was blocked by the glial inhibitors AV1013 and propentofylline, demonstrating endothelial cells as a new target of such drugs. These data indicate that (-)-morphine and M3G can activate CNS endothelial cells via TLR4, inducing proinflammatory, biochemical, morphological, and behavioral sequelae. CNS endothelial cells may have previously unanticipated roles in opioid-induced effects, in phenomena blocked by presumptive glial inhibitors, as well as TLR4-mediated phenomena more broadly.

Opioid activation of toll-like receptor 4 contributes to drug reinforcement.

Opioid action was thought to exert reinforcing effects solely via the initial agonism of opioid receptors. Here, we present evidence for an additional novel contributor to opioid reward: the innate immune pattern-recognition receptor, toll-like receptor 4 (TLR4), and its MyD88-dependent signaling. Blockade of TLR4/MD2 by administration of the nonopioid, unnatural isomer of naloxone, (+)-naloxone (rats), or two independent genetic knock-outs of MyD88-TLR4-dependent signaling (mice), suppressed opioid-induced conditioned place preference. (+)-Naloxone also reduced opioid (remifentanil) self-administration (rats), another commonly used behavioral measure of drug reward. Moreover, pharmacological blockade of morphine-TLR4/MD2 activity potently reduced morphine-induced elevations of extracellular dopamine in rat nucleus accumbens, a region critical for opioid reinforcement. Importantly, opioid-TLR4 actions are not a unidirectional influence on opioid pharmacodynamics, since TLR4(-/-) mice had reduced oxycodone-induced p38 and JNK phosphorylation, while displaying potentiated analgesia. Similar to our recent reports of morphine-TLR4/MD2 binding, here we provide a combination of in silico and biophysical data to support (+)-naloxone and remifentanil binding to TLR4/MD2. Collectively, these data indicate that the actions of opioids at classical opioid receptors, together with their newly identified TLR4/MD2 actions, affect the mesolimbic dopamine system that amplifies opioid-induced elevations in extracellular dopamine levels, therefore possibly explaining altered opioid reward behaviors. Thus, the discovery of TLR4/MD2 recognition of opioids as foreign xenobiotic substances adds to the existing hypothesized neuronal reinforcement mechanisms, identifies a new drug target in TLR4/MD2 for the treatment of addictions, and provides further evidence supporting a role for central proinflammatory immune signaling in drug reward.

Cytomegalovirus antibody status at 17-18 weeks of gestation and pre-eclampsia: a case-control study of pregnant women in Norway.

OBJECTIVE: To assess the association between maternal cytomegalovirus (CMV) antibodies in mid-pregnancy and pre-eclampsia. DESIGN: Nested case-control study. SETTING: Pregnancies registered in the Norwegian Mother and Child Cohort Study (MoBa): a large population-based pregnancy cohort (1999-2006). SAMPLE: A cohort of 1500 women with pre-eclampsia and 1000 healthy pregnant women. METHODS: Plasma samples and pregnancy-related information were provided by the MoBa. Antibody status (CMV IgG and CMV IgM) and levels (CMV IgG) at 17-18 weeks of gestation were determined by enzyme-linked immunosorbent assay (ELISA). MAIN OUTCOME MEASURE: A diagnosis of pre-eclampsia, as defined in the Medical Birth Registry of Norway. RESULTS: There was no evidence of an effect of CMV IgG seropositivity on the likelihood of developing pre-eclampsia, and CMV IgG antibody levels among women who were seropositive did not differ between groups. Adjusted for maternal age, parity and smoking, the odds ratio for pre-eclampsia in women seropositive for CMV IgG was 0.89 (95% CI 0.74-1.05; P = 0.17). The proportions of women who were seropositive for IgM did not differ between women with pre-eclampsia and women who were healthy (P = 0.98). Among nulliparous women, the proportion of women who were seropositive for CMV IgG was slightly lower among women with pre-eclampsia (53.5%) than among healthy women (59.8%) (P = 0.03). Subgroup analyses were performed for women with early or late onset pre-eclampsia, with preterm delivery and/or with neonates that were small for gestational age, but antibody status did not differ between pre-eclampsia subtypes and controls. CONCLUSIONS: The presence of maternal antibodies to CMV was not associated with pre-eclampsia in our study. The results suggest that CMV infection is unlikely to be a major cause of pre-eclampsia.

Clinical and cellular effects of hypothermia, acidosis and coagulopathy in major injury.

BACKGROUND: Hypothermia, acidosis and coagulopathy have long been considered critical combinations after severe injury. The aim of this review was to give a clinical update on this triad in severely injured patients. METHODS: A non-systematic literature search on hypothermia, acidosis and coagulopathy after major injury was undertaken, with a focus on clinical data from the past 5 years. RESULTS: Hypothermia (less than 35 °C) is reported in 1·6-13·3 per cent of injured patients. The occurrence of acidosis is difficult to estimate, but usually follows other physiological disturbances. Trauma-induced coagulopathy (TIC) has both endogenous and exogenous components. Endogenous acute traumatic coagulopathy is associated with shock and hypoperfusion. Exogenous effects of dilution from fluid resuscitation and consumption through bleeding and loss of coagulation factors further add to TIC. TIC is present in 10-34 per cent of injured patients, depending on injury severity, acidosis, hypothermia and hypoperfusion. More expedient detection of coagulopathy is needed. Thromboelastography may be a useful point-of-care measurement. Management of TIC is controversial, with conflicting reports on blood component therapy in terms of both outcome and ratios of blood products to other fluids, particularly in the context of civilian trauma. CONCLUSION: The triad of hypothermia, acidosis and coagulopathy after severe trauma appears to be fairly rare but does carry a poor prognosis. Future research should define modes of early detection and targeted therapy.

AV-65, a novel Wnt/ß-catenin signal inhibitor, successfully suppresses progression of multiple myeloma in a mouse model.

Multiple myeloma (MM) is a malignant neoplasm of plasma cells. Although new molecular targeting agents against MM have been developed based on the better understanding of the underlying pathogenesis, MM still remains an incurable disease. We previously demonstrated that ß-catenin, a downstream effector in the Wnt pathway, is a potential target in MM using RNA interference in an in vivo experimental mouse model. In this study, we have screened a library of more than 100 000 small-molecule chemical compounds for novel Wnt/ß-catenin signaling inhibitors using a high-throughput transcriptional screening technology. We identified AV-65, which diminished ß-catenin protein levels and T-cell factor transcriptional activity. AV-65 then decreased c-myc, cyclin D1 and survivin expression, resulting in the inhibition of MM cell proliferation through the apoptotic pathway. AV-65 treatment prolonged the survival of MM-bearing mice. These findings indicate that this compound represents a novel and attractive therapeutic agent against MM. This study also illustrates the potential of high-throughput transcriptional screening to identify candidates for anticancer drug discovery.

Vitamin D homeostasis is compromised due to increased urinary excretion of the 25-hydroxycholecalciferol-vitamin D-binding protein complex in the Zucker diabetic fatty rat.

Altered serum concentrations of the major circulating form of vitamin D [25-hydroxycholecalciferol (25D(3))] and its active hormone derivative [1,25-dihydroxycholecalciferol (1,25D(3))] have been linked to non-insulin-dependent diabetes mellitus (NIDDM). However, a mechanistic basis for this occurrence has not been fully elucidated. Normally, renal reabsorption of vitamin D-binding protein-bound 25D(3) absolutely requires receptor-mediated endocytosis via a receptor complex containing megalin, cubilin, and disabled-2 (Dab2), whereas an absence of megalin or its endocytic partners can lead to a marked urinary loss of 25D and severe vitamin D deficiency. Therefore, we hypothesized that reduced serum vitamin D status in NIDDM may be due to reduced expression of megalin and/or its endocytic partners and increased urinary excretion of protein-complexed 25D(3). In the present study, we utilized Zucker diabetic fatty Rats (ZDF) to demonstrate that renal reuptake of the 25D(3)-DBP complex was compromised in ZDF animals, which was reflected by a reduction in expression of megalin and Dab2. Moreover, serum levels of both 25D(3) and 1,25D(3) were reduced, and urinary 25D(3), 1,25D(3), and DBP excretion were elevated in the ZDF animals compared with their lean controls regardless of vitamin D levels in the diet. Taken together, these are the first reports to our knowledge that associate compromised renal reabsorption of the 25D(3)-DBP complex with expression of megalin and its endocytic partners in NIDDM, which in turn can lead to compromised vitamin D status.

Cognitive dysfunction and health-related quality of life after a cardiac arrest and therapeutic hypothermia.

BACKGROUND: Evidence-based treatment protocols including therapeutic hypothermia have increased hospital survival to over 50% in unconscious out-of-hospital cardiac arrest survivors. In this study we estimated the incidence of cognitive dysfunctions in a group of cardiac arrest survivors with a high functional outcome treated with therapeutic hypothermia. Secondarily, we assessed the cardiac arrest group's level of cognitive performance in each tested cognitive domain and investigated the relationship between cognitive function and age, time since cardiac arrest and health-related quality of life (HRQOL). METHODS: We included 26 patients 13-28 months after a cardiac arrest. All patients were scored using the Cerebral Performance Category scale (CPC) and Mini-Mental State Examination (MMSE). Twenty-five of the patients were tested for cognitive function using the Cambridge Neuropsychological Test Automated Battery (CANTAB). These patients were tested using four cognitive tests: Motor Screening Test, Delayed Matching to Sample, Stockings of Cambridge and Paired Associate Learning from CANTAB. All patients filled in the Short Form-36 for the assessment of HRQOL. RESULTS: Thirteen of 25 (52%) patients were classified as having a cognitive dysfunction. Compared with the reference population, there was no difference in the performance in motor function and delayed memory but there were significant differences in executive function and episodic memory. We found no associations between cognitive function and age, time since cardiac arrest or HRQOL. CONCLUSION: Half of the patients had a cognitive dysfunction with reduced performance on executive function and episodic memory, indicating frontal and temporal lobe affection, respectively. Reduced performance did not affect HRQOL.

The interrater reliability of SAPS II and SAPS 3.

PURPOSE: In this study we evaluated and compared the interrater reliability of SAPS II and SAPS 3 in order to measure the consistency of performance among different raters. METHOD: Ten junior doctors working at two general ICUs were trained in the use of SAPS II and SAPS 3 using a 2.5-h training program. After training they scored 24 cases in both systems. Scores were analyzed using intraclass correlation coefficient (ICC) statistics. In order to identify variables with low reliability, subscores were analyzed using the ICC, and single-variables were compared to a template score using weighted kappa statistics. RESULTS: The ICC (95% CI) of the scores was 0.84 (0.74, 0.91) in SAPS II and 0.80 (0.68, 0.89) in SAPS 3, which is considered adequate for both systems. Mean mortality predictions among the raters had a range of 0.12 in SAPS II and 0.19 in SAPS 3. Administrative data including age had high reliability, whereas variables based on diagnostic information had only moderate reliability. Laboratory data had consistently higher reliability than variables based on the interpretation of charts. CONCLUSION: Both SAPS II and SAPS 3 have adequate interrater reliability, but the standardized mortality ratios are still likely to be influenced by the rater's scoring practice.

Impact of the post-World War II generation on intensive care needs in Norway.

BACKGROUND: A high birth rate during the first two decades following World War II has increased the proportion of elderly people in present-day society and, consequently, the demand for health-care services. The impact on intensive care services may become dramatic because the age distribution of critically ill patients is skewed towards the elderly. We have used registry data and population statistics to forecast the demand for intensive care services in Norway up until the year 2025. METHODS: Data collected by the Norwegian intensive care registry (NIR), showing the age distribution in Norwegian intensive care units (ICU) during the years 2006 and 2007, were used with three different Norwegian prognostic models of population growth for the years 2008-2025 to compute the expected increase in intensive care unit bed-days (ICU bed-days). RESULTS: The elderly were overrepresented in Norwegian ICUs in 2006-2007, with patients from 60 to 79 years of age occupying 44% of ICU bed-days. Population growth from 2008 to 2025 was estimated to be from 11.1 to 26.4%, depending on the model used. Growth will be much larger in the age group 60-79 years. Other factors kept unchanged, this will result in an increase in the need for intensive care (ICU bed-days) of between 26.1 and 36.9%. CONCLUSION: The demand for intensive care beds will increase markedly in Norwegian hospitals in the near future. This will have serious implications for the planning of infrastructure, education of health care personnel, as well as financing of our health care system.

A comparison of SAPS II and SAPS 3 in a Norwegian intensive care unit population.

BACKGROUND: Simplified Acute Physiology Score (SAPS II) is the most widely used general severity scoring system in European intensive care medicine. Because its performance has been questioned in several external validation studies, SAPS 3 was recently released. To our knowledge, there are no published validation studies of SAPS II or SAPS 3 in the Scandinavian countries. We aimed to evaluate and compare the performance of SAPS II and SAPS 3 in a Norwegian intensive care unit (ICU) population. METHOD: Prospectively collected data from adult patients admitted to two general ICUs at two different hospitals in Norway were used. Probability of mortality was calculated using the SAPS 3 global equation (SAPS 3 G), the SAPS 3 Northern European equation (SAPS 3 NE), and the original SAPS II equation. Performance was assessed by the standardized mortality ratio (SMR), area under receiving operating characteristic, and the Hosmer and Lemeshow goodness-of-fit C test. RESULTS: One thousand eight hundred and sixty-two patients were included after excluding readmissions, and patients who were admitted after coronary surgery or burns. The SMRs were SAPS 3 G 0.71 (0.65, 0.78), SAPS 3 NE 0.74 (0.68, 0.81), and SAPS II 0.82 (0.75, 0.91). Discrimination was good in all systems. Only the SAPS 3 equations displayed satisfactory calibration, as measured by the Hosmer-Lemeshow test. CONCLUSION: The performance of SAPS 3 was satisfactory, but not markedly better than SAPS II. Both systems considerably overestimated mortality and exhibited good discrimination, but only the SAPS 3 equations showed satisfactory calibration. Customization of these equations based on a larger cohort is recommended.

Percutanous dilatational tracheotomy in the ICU: a Norwegian survey focusing on perceived risk and safety attitudes.

BACKGROUND AND OBJECTIVES: Despite its popularity, serious complications do occur with percutaneous dilatational tracheotomy in the ICU. The associated risks in daily practice are probably underestimated and may reflect system flaws in training and team function. This study was performed to obtain an impression of risk perception and safety culture in connection with percutaneous dilatational tracheotomy in Norwegian ICUs. METHODS: The Medical Director or intensivist on-call in the 30 ICUs participating in the Norwegian Intensive Care Registry was telephone interviewed using a semi-structured questionnaire. Data on the practice of tracheotomy and a qualitative assessment of complications experienced during the last 2 years were collected. In the second part, percutaneous dilatational tracheotomy operators in two ICUs were questioned about their perception of risk with percutaneous dilatational tracheotomy and asked to assess their own abilities as percutaneous dilatational tracheotomy operators and the training they had undergone. RESULTS: Of the 30 ICUs, 23 used percutaneous dilatational tracheotomy. The majority reported knowledge of severe complications like bleeding, hypoxia and tube dislodgment. Percutaneous dilatational tracheotomy-related deaths were also reported. Operators rated themselves relatively low and indicated the absence of any organized training. They acknowledged the known hazards related to percutaneous dilatational tracheotomy and suggested measures like fibreoptic guidance during the percutaneous dilatational tracheotomy and fewer operators with more experience as well as better team training, to improve patient safety. CONCLUSION: Based on the frequent reporting of serious complications and the suggested safety precautions, we conclude that the percutaneous dilatational tracheotomy is considered a high-risk procedure and that there is still room for improving the safety of this much used ICU procedure.

Severity scoring in the ICU: a review.

BACKGROUND: Patients in the intensive care unit (ICU) require huge resources because of the dysfunction of several of their vital organs. The heterogeneity and complexity of the ICU patient have generated interest in systems able to measure severity of illness as a method of predicting outcome, comparing quality-of-care and stratification for clinical trials. METHODS: By searching Medline and EMBASE for publications describing scoring systems in the ICU, the most frequently used systems, defined as resulting in more than 50 references, are included in this review. Scoring systems belong to one of four classes prognostic, single-organ failure, trauma scores and organ dysfunction (OD). The different systems are described and discussed. RESULTS: Three different prognostic scoring systems, including several versions, four single OD scores and three OD scores, were included in this review. CONCLUSION: Different forms of scoring systems are frequently used in the ICU. They have become a necessary tool to describe ICU populations and to explain differences in mortality. As there are several pitfalls related to the interpretation of the numbers supplied by the systems, they should not be used without knowledge on the science of severity scoring.

Frontotemporal lobar degeneration and ubiquitin immunohistochemistry.

We set out to determine the frequency of the different pathologies underlying frontotemporal degeneration (FTD) in our brain bank series, by reviewing all cases of pathologically diagnosed FTD over the last 12 years. We identified and reviewed 29 cases of FTD and classified them using the most recent consensus criteria with further histological analysis of 6 initially unclassifiable cases. Detailed histological analysis of these 6 cases revealed variable numbers of ubiquitin-positive (tau and alpha-synuclein-negative) inclusions in 5 cases, consistent with the diagnosis of frontotemporal lobar degeneration with ubiquitin-only-immunoreactive neuronal changes (FTLD-U). As a consequence of the current re-evaluation, 18 (62%) of the 29 cases with FTD have underlying pathology consistent with FTLD-U. Therefore in our brain bank series of frontotemporal degeneration, most cases were non-tauopathies with FTLD-U accounting for 62% of all the diagnoses.

Immunohistochemical localization of metallothionein in benign and malignant epithelial ovarian tumors.

Metallothioneins (MTs) are a group of low-molecular-weight proteins that are overexpressed in a variety of human neoplasms and are related to differentiation and prognosis in some tumor types. This study investigated immunohistochemically detectable metallothionein expression in benign and malignant ovarian surface epithelial tumors of serous, mucinous, and endometrioid types. MT expression was observed in 56% of carcinomas (n = 139) and in 2% of benign neoplasms (n = 81). Of the malignant tumors, MT expression was found in 68% of endometrioid, 56% of mucinous, and 52% of serous neoplasms. There was increased MT expression in grade 3 carcinomas (64%) as compared with grade 2 (60%) and grade 1 (23%). The overexpression of MT in malignant as opposed to benign ovarian surface epithelial tumors may suggest a role in tumorigenesis. Analogous to the situation in endometrial carcinomas, there is a tendency toward higher expression in poorly differentiated tumors. Whether high MT expression is an independent prognostic factor and increased expression indicates chemotherapy resistance in ovarian cancer, as has been previously suggested, should be determined by further studies.

Cooperative hemoglobins: conserved fold, diverse quaternary assemblies and allosteric mechanisms.

Assembly of hemoglobin subunits into cooperative complexes produces a remarkable variety of architectures, ranging in oligomeric state from dimers to complexes containing 144 hemoglobin subunits. Diverse stereochemical mechanisms for modulating ligand affinity through intersubunit interactions have been revealed from studies of three distinct hemoglobin assemblages. This mechanistic diversity, which occurs between assemblies of subunits that have the same fold, provides insight into the range of regulatory strategies that are available to protein molecules.

Oncostatin M promotes biphasic tissue factor expression in smooth muscle cells: evidence for Erk-1/2 activation.

Tissue factor (TF), a transmembrane glycoprotein, initiates the extrinsic coagulation cascade. TF is known to play a major role in mediating thrombosis and thrombotic episodes associated with the progression of atherosclerosis. Macrophages at inflammatory sites, such as atherosclerotic lesions, release numerous cytokines that are capable of modulating TF expression. This study examined the role of oncostatin M (OSM), a macrophage/ T-lymphocyte-restricted cytokine, in the expression of TF in vascular smooth muscle cells (SMCs). It is reported here that OSM stimulated a biphasic and sustained pattern of TF messenger RNA (mRNA). The effect of OSM on TF mRNA expression was regulated at the transcriptional level as determined by nuclear run-offs and transient transfection of a TF promoter-reporter gene construct. OSM-induced TF expression was regulated primarily by the transcription factor NF-kappaB. Activation of NF-kappaB by OSM did not require IkappaB-alpha degradation. Inhibition of MEK activity by U0126 prevented OSM-induced TF expression by suppressing NF-kappaB DNA binding activity as determined by gel-shift analysis. Further, inhibition of Erk-1/2 protein by antisense treatment resulted in suppression of TF mRNA expression, indicating a role for Erk-1/2 in modulating NF-kappaB DNA binding activity. These studies suggest that the induced expression of TF by OSM is primarily through the activation of NF-kappaB and that activation of NF-kappaB is regulated in part by the MEK/Erk-1/2 signal transduction pathway. This study indicates that OSM may play a key role in promoting TF expression in SMCs within atherosclerotic lesions.

A strong intronic enhancer element of the EGFR gene is preferentially active in high EGFR expressing breast cancer cells.

Hormone-independent human breast cancer is characterized by estrogen receptor (ER) loss and the acquisition of high epidermal growth factor receptor (EGFR) levels. Despite the tendency for an inverse correlation between EGFR and ER, EGFR is a strong prognostic indicator for poor survival rate independent of ER status suggesting that EGFR overexpression is an important step in the progression to estrogen independence. We have previously shown that several DNase I hypersensitive sites which correspond to potential regulatory regions reside within the EGFR gene first intron exclusively in hormone-independent breast cancer cells. CAT assays investigating the transcriptional activity of the first intron of EGFR indicate that a 140 bp region has an enhancer ability specifically in these hormone-independent breast cancer cells. The DNA-protein interaction that occurs in this enhancer was localized to a 35 bp region and displayed enhancer-like activity in the same hormone-independent breast cancer cells. Furthermore, the protein that binds to this 35 bp region seems to be ubiquitous in the cell lines tested but in higher abundance in high EGFR expressing cells. Identifying the specific regulatory elements involved in EGFR up-regulation could lead to the development of therapies for preventing and treating estrogen-independent breast cancer.

Induction of the urokinase plasminogen activator system by oncostatin M promotes endothelial migration.

Oncostatin M (OSM) is an inflammatory cytokine produced by activated macrophages and T-lymphocytes. We have previously demonstrated that OSM-induced endothelial cell migration, unlike endothelial cell proliferation and spindle formation, is independent of basic fibroblast growth factor expression (Wijelath et al. [1997] J. Cell. Sci. 110:871-879). To better understand the mechanism of OSM-induced endothelial cell migration, this study examined the potential role of the plasminogen activator system in promoting OSM mediated endothelial cell migration. OSM stimulated increased mRNA levels of urokinase-plasminogen activator (uPA) and urokinase-plasminogen activator receptor (uPAR) in a time and dose-dependent manner. Transcriptional run-off and mRNA stability analysis demonstrated that the increase in uPA and uPAR mRNA levels was due to both increased gene transcription and mRNA stability. The increase in mRNA correlated with increased protein levels of both uPA and uPAR. This increase was reflected in elevated levels of membrane-bound plasmin activity. OSM-induced endothelial cell migration was only partially dependent on plasmin activity since incubating endothelial cells without plasminogen or, in the presence of aprotinin, resulted in suppression of endothelial cell migration, indicating that OSM promoted endothelial cell migration through both a plasmin-dependent and -independent mechanism. Our results imply a role for OSM in promoting endothelial cell migration via a plasmin-dependent pathway and a uPAR-mediated pathway. Together, these and other recent studies support a role for OSM in modulating the different phases of angiogenesis.