Signalling danger: endoplasmic reticulum stress and the unfolded protein response in pancreatic islet inflammation.

Protein synthesis is increased by several-fold in stimulated pancreatic beta cells. Synthesis and folding of (pro)insulin takes place in the endoplasmic reticulum (ER), and beta cells trigger the unfolded protein response (UPR) to upgrade the functional capacity of the ER. Prolonged or excessive UPR activation contributes to beta cell dysfunction and death in type 2 diabetes, but there is another side of the UPR that may be of particular relevance for autoimmune type 1 diabetes, namely, the cross-talk between the UPR and innate immunity/inflammation. Recent evidence, discussed in this review, indicates that both saturated fats and inflammatory mediators such as cytokines trigger the UPR in pancreatic beta cells. The UPR potentiates activation of nuclear factor κB, a key regulator of inflammation. Two branches of the UPR, namely IRE1/XBP1s and PERK/ATF4/CHOP, mediate the UPR-induced sensitisation of pancreatic beta cells to the proinflammatory effects of cytokines. This can contribute to the upregulation of local inflammatory mechanisms and the aggravation of insulitis. The dialogue between the UPR and inflammation may provide an explanation for the parallel increase in the prevalence of childhood obesity and type 1 diabetes.

Adherence to Enhanced Recovery After Surgery and length of stay after colonic resection.

AIM: The Enhanced Recovery After Surgery (ERAS) programme is a multimodal approach to improve peri-operative care in colon surgery. The aim of this study was to report on the adherence to and outcomes of ERAS in the first years after implementation. METHOD: Data of patients undergoing elective colon resections for malignancy in 2006 until 2010 were compared with patients receiving conventional care in 2005. Retrospective analysis was performed including length of stay (LOS), protocol adherence and complications. The predictive values of ERAS items and baseline characteristics on LOS and complications were analysed using univariate and multivariate analysis. RESULTS: Length of stay (LOS) was significantly shorter in 2006 and 2007 (P ≤ 0.009 and P ≤ 0.004) but not in 2008 and 2009. The mean adherence rate to the ERAS items was 84.1% in 2006 and 2007 and 72.4% in 2008 and 2009 (P < 0.001). In 2005, 2008 and 2009 LOS was significantly shorter for laparoscopically operated patients than for patients with open resections (P < 0.002, P < 0.001 and P < 0.004 respectively). Multivariate analysis showed that age, laparoscopic surgery, removal of nasogastric tube before extubation, mobilization within 24 h after surgery, starting nonsteroidal anti-inflammatory drugs at day 1 and removal of thoracic epidural analgesia at day 2 were independent predictors of LOS. CONCLUSION: Strict adherence to the ERAS protocol was associated with reduced LOS and improved outcome in elective colon surgery for malignancy. These benefits were lost when protocol adherence was lower. Embedding the ERAS protocol into an organization and repetitive education are vital to sustain its beneficial effects on LOS and outcome.

Is ARE/poly(U)-binding factor 1 (AUF1) a new player in cytokine-mediated beta cell apoptosis?

Type 1 diabetes is a chronic autoimmune disease involving the progressive loss of beta cell mass. Cytokines released by immune cells are early contributors to beta cell apoptosis. Thus, an understanding of the signal transduction mechanisms induced by cytokines in beta cells is necessary for the rational design of novel therapies to prevent or to cure this disease. Cytokine-mediated beta cell apoptosis is a complex phenomenon that includes activation of the transcription factors signal transducer and activator of transcription 1 and nuclear factor κB (NFκB), c-Jun N-terminal kinase, endoplasmic reticulum (ER) stress and the intrinsic mitochondrial apoptotic pathway. NFκB has both a pro-inflammatory and a pro-apoptotic role in beta cells. One of the mechanisms by which NFκB contributes to beta cell apoptosis is via activation of ER stress. The role for ER stress in beta cell apoptosis is not completely clarified but involves production of C/EBP homologous protein and activation of the intrinsic mitochondrial apoptotic pathway. In this issue of Diabetologia, Roggli et al (DOI 10.1007/s00125-011-2399-7) report on a new player in this elaborate response, the RNA-binding protein ARE/poly(U)-binding factor 1. This commentary discusses these findings and their relevance to the field.

Deletion of C/EBP homologous protein (Chop) in C57Bl/6 mice dissociates obesity from insulin resistance.

AIMS/HYPOTHESIS: Endoplasmic reticulum (ER) stress has been implicated in the development of type 2 diabetes, via effects on obesity, insulin resistance and pancreatic beta cell health. C/EBP homologous protein (CHOP) is induced by ER stress and has a central role in apoptotic execution pathways triggered by ER stress. The aim of this study was to characterise the role of CHOP in obesity and insulin resistance. METHODS: Metabolic studies were performed in Chop ( -/- ) and wild-type C57Bl/6 mice, and included euglycaemic-hyperinsulinaemic clamps and indirect calorimetry. The inflammatory state of liver and adipose tissue was determined by quantitative RT-PCR, immunohistology and macrophage cultures. Viability and absence of ER stress in islets of Langerhans was determined by electron microscopy, islet culture and quantitative RT-PCR. RESULTS: Systemic deletion of Chop induced abdominal obesity and hepatic steatosis. Despite marked obesity, Chop ( -/- ) mice had preserved normal glucose tolerance and insulin sensitivity. This discrepancy was accompanied by lower levels of pro-inflammatory cytokines and less infiltration of immune cells into fat and liver. CONCLUSIONS/INTERPRETATION: These observations suggest that insulin resistance is not induced by fat accumulation per se, but rather by the inflammation induced by ectopic fat. CHOP may play a key role in the crosstalk between excessive fat deposition and induction of inflammation-mediated insulin resistance.

Sustained production of spliced X-box binding protein 1 (XBP1) induces pancreatic beta cell dysfunction and apoptosis.

AIMS/HYPOTHESIS: Pro-inflammatory cytokines involved in the pathogenesis of type 1 diabetes deplete endoplasmic reticulum (ER) Ca2+ stores, leading to ER-stress and beta cell apoptosis. However, the cytokine-induced ER-stress response in beta cells is atypical and characterised by induction of the pro-apoptotic PKR-like ER kinase (PERK)-C/EBP homologous protein (CHOP) branch of the unfolded protein response, but defective X-box binding protein 1 (XBP1) splicing and activating transcription factor 6 activation. The purpose of this study was to overexpress spliced/active Xbp1 (XBP1s) to increase beta cell resistance to cytokine-induced ER-stress and apoptosis. METHODS: Xbp1s was overexpressed using adenoviruses and knocked down using small interference RNA in rat islet cells. In selected experiments, Xbp1 was also knocked down in FACS-purified rat beta cells and rat fibroblasts. Expression and production of XBP1s and key downstream genes and proteins was measured and beta cell function and viability were evaluated. RESULTS: Adenoviral-mediated overproduction of Xbp1s resulted in increased XBP1 activity and induction of several XBP1s target genes. Surprisingly, XBP1s overexpression impaired glucose-stimulated insulin secretion and increased beta cell apoptosis, whereas it protected fibroblasts against cell death induced by ER-stress. mRNA expression of Pdx1 and Mafa was inhibited in cells overproducing XBP1s, leading to decreased insulin expression. XBP1s knockdown partially restored cytokine/ER-stress-driven insulin and Pdx1 inhibition but had no effect on cytokine-induced ER-stress and apoptosis. CONCLUSIONS/INTERPRETATION: XBP1 has a distinct inhibitory role in beta cell as compared with other cell types. Prolonged XBP1s production hampers beta cell function via inhibition of insulin, Pdx1 and Mafa expression, eventually leading to beta cell apoptosis.

Presentation, diagnosis, and management of multiple system atrophy in Europe: final analysis of the European multiple system atrophy registry.

Multiple system atrophy (MSA) is a Parkinson's Disease (PD)-like α-synucleinopathy clinically characterized by dysautonomia, parkinsonism, cerebellar ataxia, and pyramidal signs in any combination. We aimed to determine whether the clinical presentation of MSA as well as diagnostic and therapeutic strategies differ across Europe and Israel. In 19 European MSA Study Group centres all consecutive patients with a clinical diagnosis of MSA were recruited from 2001 to 2005. A standardized minimal data set was obtained from all patients. Four-hundred thirty-seven MSA patients from 19 centres in 10 countries were included. Mean age at onset was 57.8 years; mean disease duration at inclusion was 5.8 years. According to the consensus criteria 68% were classified as parkinsonian type (MSA-P) and 32% as cerebellar type (MSA-C) (probable MSA: 72%, possible MSA: 28%). Symptomatic dysautonomia was present in almost all patients, and urinary dysfunction (83%) more common than symptomatic orthostatic hypotension (75%). Cerebellar ataxia was present in 64%, and parkinsonism in 87%, of all cases. No significant differences in the clinical presentation were observed between the participating countries. In contrast, diagnostic work up and therapeutic strategies were heterogeneous. Less than a third of patients with documented orthostatic hypotension or neurogenic bladder disturbance were receiving treatment. This largest clinical series of MSA patients reported so far shows that the disease presents uniformly across Europe. The observed differences in diagnostic and therapeutic management including lack of therapy for dysautonomia emphasize the need for future guidelines in these areas.

Cell loss during pseudoislet formation hampers profound improvements in islet lentiviral transduction efficacy for transplantation purposes.

Islet transplantation is a promising treatment in type 1 diabetes, but the need for chronic immunosuppression is a major hurdle to broad applicability. Ex vivo introduction of agents by lentiviral vectors-improving beta-cell resistance against immune attack-is an attractive path to pursue. The aim of this study was to investigate whether dissociation of islets to single cells prior to viral infection and reaggregation before transplantation would improve viral transduction efficacy without cytotoxicity. This procedure improved transduction efficacy with a LV-pWPT-CMV-EGFP construct from 11.2 +/- 4.1% at MOI 50 in whole islets to 80.0 +/- 2.8% at MOI 5. Viability (as measured by Hoechst/PI) and functionality (as measured by glucose challenge) remained high. After transplantation, the transfected pseudoislet aggregates remained EGFP positive for more than 90 days and the expression of EGFP colocalized primarily with the insulin-positive beta-cells. No increased vulnerability to immune attack was observed in vitro or in vivo. These data demonstrate that dispersion of islets prior to lentiviral transfection and reaggregation prior to transplantation is a highly efficient way to introduce genes of interest into islets for transplantation purposes in vitro and in vivo, but the amount of beta-cells needed for normalization of glycemia was more than eightfold higher when using dispersed cell aggregates versus unmanipulated islets. The high price to pay to reach stable and strong transgene expression in islet cells is certainly an important cell loss.

Electromyographic fatigue threshold of the biceps brachii: the effect of endurance time.

Muscle fatigue can be a limiting factor to determine index as the electromyographic fatigue threshold (EMGFT) due the alterations in motivation and disconfots. This way, the purpose of this study was to identify the right biceps brachii and left biceps brachii obtained from repetitive elbow flexions at each 100% of total time. Nine healthy subjects performed the exercise named biceps curl until exhaustion with 25%, 35%, and 45%0 of one repetition maximum, in three different days. EMG amplitude (root mean square--RMS) was obtained for concentric contractions during these load levels and correlated with time to determine the slope values for each load and them determine the EMGFT. The EMGFT was obtained within of each 10% of total time and they were compared by analysis of variance. The results showed a progressive increase in RMS with time, for both muscles in all loads, characterizing the muscle fatigue process, and for the EMGFT values ware not found predominantly significant differences between the execution time, as well as between muscles (right biceps x left biceps). This protocol allowed to identify the EMGFT to both muscles during the biceps curl, which was similar at different percentage of total time, indicating the possibility to reduce the length of the contraction test without the need to maintain the contraction until exhaustion. Further studies are needed to evaluate the applicability of this method to determining the effects on performance.

Intraoperative ultrasound guidance in breast-conserving surgery shows superiority in oncological outcome, long-term cosmetic and patient-reported outcomes: Final outcomes of a randomized controlled trial (COBALT).

BACKGROUND: The multicenter randomized controlled COBALT trial demonstrated that ultrasound-guided breast-conserving surgery (USS) results in a significant reduction of margin involvement (3.1% vs. 13%) and excision volumes compared to palpation-guided surgery (PGS). The aim of the present study was to determine long term oncological and patient-reported outcomes including quality of life (QoL), together with their progress over time. METHODS: 134 patients with T1-T2 breast cancer were randomized to USS (N = 65) or PGS (N = 69). Cosmetic outcomes were assessed with the Breast Cancer Conservative Treatment cosmetic results (BCCT.core) software, panel-evaluation and patient self-evaluation on a 4-point Likert-scale. QoL was measured using the EORTC QLQ-C30/-BR23 questionnaire. RESULTS: No locoregional recurrences were reported after mean follow-up of 41 months. Seven patients (5%) developed distant metastatic disease (USS 6.3%, PGS 4.4%, p = 0.466), of whom six died of disease (95.5% overall survival). USS achieved better cosmetic outcomes compared to PGS, with poor outcomes of 11% and 21% respectively, a result mainly attributable to mastectomies due to involved margins following PGS. There was no difference after 1 and 3 years in cosmetic outcome. Dissatisfied patients included those with larger excision volumes, additional local therapies and worse QoL. Patients with poor/fair cosmetic outcomes scored significantly lower on aspects of QoL, including breast-symptoms, body image and sexual enjoyment. CONCLUSION: By significantly reducing positive margin status and lowering resection volumes, USS improves the rate of good cosmetic outcomes and increases patient-satisfaction. Considering the large impact of cosmetic outcome on QoL, USS has great potential to improve QoL following breast-conserving therapy.

Cefalea en la disección aórtica, a propósito de un caso clínico / Headache as a symptom in aortic dissection: about a clinical case / Cefaleia em dissecção de aorta, tema de caso clínico

Se describe el caso de un paciente de 70 años que consultó por cefalea súbita, tipo trueno, sin alteración del estado de consciencia, acompañada de dolor torácico de una hora de evolución y de baja intensidad. A su ingreso fue enfocado como cefalea en trueno, que es clasificada, en cuanto a la atención, como bandera roja. La medición de troponina fue negativa y una tomografía de cráneo fue leída como normal. Desde el ingreso presentaba signos vitales normales, cuando iba a ser dado de alta se torna hipotenso (completamente asintomático) y por su síntoma cardinal (cefalea), que se asoció a dolor torácico leve y no anginoso, se solicitó angiotomografía toracoabdominal, con la que se demostró aneurisma disecante de la aorta. Con la presentación de este caso, se busca resaltar la importancia en el servicio de urgencias de la asociación de la cefalea tipo trueno, con condiciones vasculares como la disección aórtica.

We describe the case of a 70-year-old patient, who seeks medical advice due to sudden, thunder headache, without alteration of the state of consciousness, accompanied by chest pain of 1 hour of evolution and of low intensity. Upon his admission, the patient was treated as a thunderclap headache, which is considered a red flag. His troponin was negative, and his head tomography was interpreted as normal. From admission he had normal vital signs, but when he was going to be discharged, he became hypotensive (completely asymptomatic) and due to his cardinal symptom (headache) that was asso-ciated with mild non-anginal chest pain, a thoracoabdominal angioCT was requested, with which dissecting aneurysm of the aorta was evidenced. With the presentation of this case, we seek to highlight the importance of the association of thunder-type headache with possible vascular conditions such as aortic dissection in the emergency department.

Descrevemos o caso de uma paciente de 70 anos que consultou por quadro de cefaleia súbita, tipo trovão, sem alteração do estado de consciência, acompanhada de dor torácica de uma hora de evolução e de baixa intensidade. Na admissão, foi tratado como cefaleia em trovoada, que é classificada, em termos de atenção, como bandeira vermelha. A me-dição da troponina foi negativa e uma tomografia de crânio foi lida como normal. Desde a admissão apresentava sinais vitais normais, quando ia receber alta ficou hipotenso (totalmente assintomático) e devido ao seu sintoma cardinal (cefaleia), que se associou a dores torácicas ligeiras e não anginosas, foi solicitada angiografia toracoabdominal, com cujo aneurisma dissecante da aorta foi demonstrado. Com a apresentação deste caso, o objetivo é destacar a importância no pronto-socorro da associação da cefaleia do tipo trovão com afecções vasculares como a dissecção da aorta.

Identification of novel cytokine-induced genes in pancreatic beta-cells by high-density oligonucleotide arrays.

Type 1 diabetes is an autoimmune disease resulting from the selective destruction of insulin-producing beta-cells. Cytokines may contribute to pancreatic beta-cell death in type 1 diabetes. beta-cell exposure to interleukin (IL)-1beta induces functional impairment, whereas beta-cell culture for 6-9 days in the presence of IL-1beta and interferon (INF)-gamma leads to apoptosis. To clarify the mechanisms involved in these effects of cytokines, we studied the general pattern of cytokine-induced gene expression in beta-cells. Primary rat beta-cells were fluorescence-activated cell sorter-purified and exposed for 6 or 24 h to control condition, IL-1beta + INF-gamma, or IL-1beta alone (24 h only). Gene expression profile was analyzed in duplicate by oligonucleotide arrays. Nearly 3,000 transcripts were detected in controls and cytokine-treated beta-cells. Of these, 96 and 147 displayed changes in expression after 6 and 24 h, respectively, of exposure to IL-1beta + INF-gamma, whereas 105 transcripts were modified after a 24-h exposure to IL-1beta. The cytokine-responsive genes were clustered according to their biological functions. The major clusters observed were metabolism, signal transduction, transcription factors, protein synthesis/ processing, hormones, and related receptors. These modifications in gene expression may explain some of the cytokine effects in beta-cells, such as decreased protein biosynthesis and insulin release. In addition, there was induction of diverse cytokines and chemokines; this suggests that beta-cells may contribute to mononuclear cell homing during insulitis. Several of the cytokine-induced genes are potentially regulated by the transcription factor NF-kappaB. Clarification of the function of the identified cytokine-induced gene patterns may unveil some of the mechanisms involved in beta-cell damage and repair in type 1 diabetes.

Inhibition of cytokine-induced NF-kappaB activation by adenovirus-mediated expression of a NF-kappaB super-repressor prevents beta-cell apoptosis.

Cytokine-induced beta-cell death is an important event in the pathogenesis of type 1 diabetes. The transcription factor nuclear factor-kappaB (NF-kappaB) is activated by interleukin-1beta (IL-1beta), and its activity promotes the expression of several beta-cell genes, including pro- and anti-apoptotic genes. To elucidate the role of cytokine (IL-1beta + gamma-interferon [IFN-gamma])-induced expression of NF-kappaB in beta-cell apoptosis, rat beta-cells were infected with the recombinant adenovirus AdIkappaB((SA)2), which contained a nondegradable mutant form of inhibitory kappaB (IkappaB((SA)2), with S32A and S36A) that locks NF-kappaB in a cytosolic protein complex, preventing its nuclear action. Expression of IkappaB((SA)2) inhibited cytokine-stimulated nuclear translocation and DNA-binding of NF-kappaB. Cytokine-induced gene expression of several NF-kappaB targets, namely inducible nitric oxide synthase, Fas, and manganese superoxide dismutase, was prevented by AdIkappaB((SA)2), as established by reverse transcriptase-polymerase chain reaction, protein blot, and measurement of nitrite in the medium. Finally, beta-cell survival after IL-1beta + IFN-gamma treatment was significantly improved by IkappaB((SA)2) expression, mostly through inhibition of the apoptotic pathway. Based on these findings, we conclude that NF-kappaB activation, under in vitro conditions, has primarily a pro-apoptotic function in beta-cells.

Electromyographic fatigue threshold of the biceps brachii muscle during dynamic contraction.

The purpose of this study was to identify the Electromyographic Fatigue Threshold (EMGFT) of the biceps brachii muscle bilaterally during the elbow flexion in tests performed in different times. 30 second test, 1 minute test and fatiguing test, in concentric (CC) and eccentric (EC) phases. Nine healthy young men performed the elbow flexion with loads corresponding at 25%, 35% and 45% of the one repetition maximum (1-RM) in separate days. The results indicated that the test applied for the biceps brachii muscle during elbow flexion induced a progressive increment of EMG activity with time indicating muscle fatigue and allowed the identification of the EMGFT. The three tests presented no difference of EMGFT between CC and EC phases bilaterally.

Neuronal changes in the substantia nigra with aging: a Golgi study.

In order to recognize substantia nigra neuronal changes occurring in aging, 20 human control brains from 13 males and 7 females with a mean age of 61 years (range 20 to 93 years) without neurological disease were examined using the Golgi method. A quantitative study of dendrites and dendritic spines was performed as well as a statistical analysis of obtained data. Parallel sections to the impregnated material were histologically and immunohistologically studied with the aim to identify possible neuronal cytoskeletal abnormalities. Results were compared to changes of substantia nigra reported in other conditions such as Parkinson's disease (PD) and methyl-4-phenylpyridine (MPTP) experimental toxicity. Three different substantia nigra neuronal types were observed. Morphological changes during aging consisted of distorted profile of the cell body and swelling and beading of dendritic branches. The quantitative assessment of changes observed in neuronal types showed a significant loss of dendrites and dendritic spines, especially in the oldest cases. These findings were similar to those previously described in other cerebral areas during aging, but a specific vulnerability of the largest substantia nigra neuronal type could be observed. Nodulations and beaded aspects of dendrites are reminiscent of those changes previously described in MPTP toxicity. Dendritic varicosities found in the oldest cases have also been found in dendrites of large substantia nigra neurons in PD. Cytoskeletal abnormalities have been described in PD but were not found in the present study. Therefore, other pathophysiological mechanisms different from the cytoskeletal compromise occurring in some neurodegenerative diseases should be involved in aging.

Phosphorylated map kinase (ERK1, ERK2) expression is associated with early tau deposition in neurones and glial cells, but not with increased nuclear DNA vulnerability and cell death, in Alzheimer disease, Pick's disease, progressive supranuclear palsy and corticobasal degeneration.

Abnormal tau phosphorylation and deposition in neurones and glial cells is one of the major features in taupathies. The present study examines the involvement of the Ras/MEK/ERK pathway of tau phosphorylation in Alzheimer disease (AD), Pick's disease (PiD), progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD), by Western blotting, single and double-labelling immunohistochemistry, and p21Ras activation assay. Since this pathway is also activated in several paradigms of cell death and cell survival, activated ERK expression is also analysed with double-labelling immunohistochemistry and in situ end-labelling of nuclear DNA fragmentation to visualise activated ERK in cells with increased nuclear DNA vulnerability. The MEK1 antibody recognises one band of 45 kD that identifies phosphorylation-independent MEK1, whose expression levels are not modified in diseased brains. The ERK antibody recognises one band of 42 kD corresponding to the molecular weight of phosphorylation-independent ERK2; the expression levels, as well as the immunoreactivity of ERK in individual cells, is not changed in AD, PiD, PSP and CBD. The antibody MAPK-P distinguishes two bands of 44 kD and 42 kD that detect phosphorylated ERK1 and ERK2. MAPK-P expression levels, as seen with Western blotting, are markedly increased in AD, PiD, PSP and CBD. Moreover, immunohistochemistry discloses granular precipitates in the cytoplasm of neurones in AD, mainly in a subpopulation of neurones exhibiting early tau deposition, whereas neurones with developed neurofibrillary tangles are less commonly immunostained. MAPK-P also decorates neurones with Pick bodies in PiD, early tau deposition in neurones in PSP and CBD, and cortical achromatic neurones in CBD. In addition, strong MAPK-P immunoreactivity is found in large numbers of tau-positive glial cells in PSP and CBD, as seen with double-labelling immunohistochemistry. Yet no co-localisation of enhanced phosphorylated ERK immunoreactivity and nuclear DNA fragmentation is found in AD, PiD, PSP and CBD. Finally, activated Ras expression levels are increased in AD cases when compared with controls. These results demonstrate increased phosphorylated (active) ERK expression in association with early tau deposition in neurones and glial cells in taupathies, and suggest activated Ras as the upstream activator of the MEK/ERK pathway of tau phosphorylation in AD.

Influence of endothelin ET(A) and ET(B) receptor antagonists on endothelin-induced contractions of the guinea pig isolated gall bladder.

The receptors mediating guinea pig gall bladder (GPGB) contractions induced by endothelin-1 (ET-1) and related peptides were characterized using various ET receptor antagonists. As all ET-receptor agonists used, except sarafotoxin S6c (SRTX), failed to induce a clear-cut maximal response at the highest concentration tested (i.e. 100 nM), their potencies are expressed in terms of a CK50 (i.e. the concentration causing 50% of the response to 80 mM KCl). ET-1 (CK50 0.8 nM) was equipotent to ET-2 and SRTX (selective ET(B) receptor agonist), but more potent than ET-3 (5-fold) or IRL 1620 (selective ET(B) receptor agonist). BQ-123 (0.3 microM, peptidic ET(A) receptor antagonist) did not alter responses to ET-1, ET-3 or SRTX. BQ-788 (1 microM, peptidic ET(B) receptor antagonist) reduced the potency of ET-3 (9-fold at the CK50 level) and SRTX ( > 20-fold), but not ET-1. SRTX responses were unaffected by RES-701-1 (3 microM, peptidic ET(B) receptor antagonist). The combination BQ-123 (0.3 microM) plus BQ-788 (1 microM) did not modify responses to ET-1, inhibited SRTX responses similarly to BQ-788 alone and abolished ET-3 responses. Bosentan (1 microM, non-peptidic ET(A)/ET(B) receptor antagonist) reduced the potency of ET-1 (15-fold). ET-3 (9-fold) and SRTX (4-fold). In rat aorta, the antagonists blocked ET-1-induced contractions (BQ-123 and bosentan) or SRTX-induced endothelium-dependent relaxations (BQ-788, RES-701-1 and bosentan). Thus, the GPGB expresses both ET(A) and ET(B) receptors. As BQ-123 only blocked responses to ET-3 in the presence of BQ-788, there appears to be cross-talk between both receptor types. Also, the binding sites of ET-1 and ET-3 on the ET(A) receptor may not coincide entirely, as BQ-123, even in presence of BQ-788, did not affect ET-1-induced contractions.

Changes in paw oedema triggered via bradykinin B(1) and B(2) receptors in streptozotocin-diabetic rats.

The present study investigated hind paw oedema mediated by bradykinin B(1) and B(2) receptors in streptozotocin-diabetic rats. Paw oedema induced by intraplantar (i.pl.) injection of bradykinin or the selective bradykinin B(2) receptor agonist, Tyrosine(8)-bradykinin ([Tyr(8)]bradykinin) (both 3 nmol/paw), was significantly reduced at 4 weeks after streptozotocin treatment (34 +/- 8% and 40 +/- 7%). At 6 weeks after streptozotocin, when paw oedema caused by substance P or prostaglandin E(2) (both 10 nmol/paw) was unchanged, inhibition of bradykinin B(2) receptor-mediated oedema was maximal (66 +/- 6% and 72 +/ -2%, for bradykinin and [Tyr(8)]bradykinin, respectively). The selective bradykinin B(1) receptor agonist, [des-Arg(9)]bradykinin (100 nmol/paw), induced only slight paw oedema in non-diabetic controls. Responses to [des-Arg(9)]bradykinin were markedly enhanced 8 weeks after streptozotocin (from 0.09 +/- 0.01 to 0.38 +/- 0.05 ml), less so at 10 weeks (0.22 +/- 0.03 ml), and returning to basal values at 12 weeks (0.11 +/- 0.03 ml). Treatment with insulin protamine zinc (1-3 U/day/7 weeks, s.c.) did not reverse the inhibition of responses to [Tyr(8)]bradykinin or the potentiation of responses to [des-Arg(9)]bradykinin seen at 8 weeks. Thus, streptozotocin-induced diabetes induces long-lasting alterations in oedematogenic responsiveness to kinins in the rat, characterized by marked reduction of oedema involving activation of bradykinin B(2) receptors, associated with enhancement of bradykinin B(1) receptor-mediated oedema.

Clinical and pathological study of two patients with progressive supranuclear palsy and Alzheimer's changes. Antigenic determinants that distinguish cortical and subcortical neurofibrillary tangles.

Two cases with classical clinical manifestations of progressive supranuclear palsy (PSP) showed severe progressive dementia as an additional clinical feature. Neuropathological study demonstrated typical features of PSP in the brainstem. Additionally, histological criteria of Alzheimer's disease (AD) were observed. A topographic and immunohistological study (with neurofilament subunit and Tau and Ubiquitin antibodies) of the distribution of neurofibrillary tangles (NFTs) was performed in order to compare the characteristics of NFTs from cortex and brainstem. NFTs from cortex were positive with all antibodies used and were predominantly distributed in cortical layers III and V and affected medium size neurons. Brainstem NFTs were positive only for neurofilament subunits and Tau. Cortical and brainstem NFTs showed immunohistological differences. Cortical NFTs in our two cases had a similar distribution as in control AD cases. On the basis of our observations we believe (1) that cortical tangles in our PSP cases are related to Alzheimer's disease and (2) that the cortical NFTs of PSP and AD are morphologically and immunohistologically distinct. Mechanisms concerned with the production of cortical and brainstem NFTs in PSP and AD are discussed.

Ubiquitin in cerebral amyloid angiopathy.

Immunohistological findings in cerebral blood vessels of 4 cases with cerebral amyloid angiopathy (CAA) were compared with those of 4 Alzheimer's (AD) cases. A panel of antibodies against 2 neurofilament subunits (BF10 and RT97), a microtubule-associated protein (TAU) and ubiquitin were used. CAA cases showed a strong immunoreactivity for ubiquitin in blood vessel wall. Senile plaques (SPs) in CAA cases showed strong ubiquitin positivity but the central amyloid core was negative. AD brains showed immunoreactivity with all antibodies in SPs and neurofibrillary tangles (NFTs); blood vessels were consistently negative for ubiquitin. Control brains showed few SPs and NFTs; these were positive for ubiquitin, but blood vessels were negative. These results indicate that vascular amyloid deposition in CAA and AD may have different pathophysiological mechanisms.