bluuebunny/crossref_metadata_2025 huggingface dataset:
NIH Stroke Scale Reliability in Ratings from a Large Sample of Clinicians
Authors: Josephson S. Andrew , Hills Nancy K. , Johnston S. Claiborne
Published: August 2006

Objective: The NIH Stroke Scale (NIHSS) is widely used in stroke clinical care and trials. Certification in its use, most commonly through rating of video vignettes, is routinely required. To investigate the reliability of the NIHSS in a representative sample of raters, we examined the results of the most frequently used certification examination. Methods: At the invitation of the National Stroke Association, we analyzed the results of all raters who completed one of two multiple patient videotaped certification examinations from 1998 to 2004. Total scores for each vignette were calculated and ratings were compared based on percentile of responses and modified kappa scores. Results: There were 7,405 unique raters with 38,148 individual NIHSS item responses; median scores for each vignette ranged from 0 to 31. Total NIHSS scores varied widely between raters; scoring for 7 of the 11 patients (64%) had a four or more point difference in NIHSS score from the 5th to 95th percentile. The aphasia (kappa = 0.60) and facial palsy (0.65) items on the test contributed most to the variance in the total NIHSS score. Nurses agreed with the most common response on scoring more frequently than physicians (p < 0.0001). Taking the certification examination multiple times did not improve agreement. Conclusions: In a large diverse sample of clinicians, inter-rater reliability for individual elements of the NIHSS on videotaped vignettes was generally good, but overall scoring was inconsistent and could impact clinical trial results. Whether additional training, modification of examination elements, or clearer definitions for scoring could improve reliability requires further study.
Effect of Carotid Artery Stenting on Ocular Circulation and Chronic Ocular Ischemic Syndrome
Authors: Kawaguchi Shoichiro , Sakaki Toshisuke , Iwahashi Hideaki , Fujimoto Kenta , Iida Jun-ichi , Mishima Hideaki , Nishikawa Norikiyo
Published: August 2006

Background: The authors evaluated the effect of carotid artery stenting (CAS) on ocular circulation and chronic ocular ischemic syndrome. Methods: We examined 38 patients with carotid artery stenosis (>80%) at its origin treated with CAS. Ocular circulation and symptoms were examined before, within 24 h, and 1 week, 1 month, and 3 months after CAS based on ophthalmic artery color Doppler flow imaging and ophthalmological examinations. Results: Ocular circulation: Before CAS, 13 patients showed reversed ophthalmic artery flow, and 25 antegrade flow. Average peak systolic flow velocity was –0.038 m/s. Within 24 h after CAS, all patients showed antegrade ophthalmic artery flow; reversed flow before CAS was thus resolved. Average peak systolic flow velocity rose significantly to 0.36 m/s (p < 0.05). One week, 1 month and 3 months after CAS, there were no significant changes compared to the findings at 1 week after CAS. Ocular symptoms: Before CAS, 8 patients showed chronic ocular ischemic syndrome. During the follow-up period (mean: 2.8 years), the visual acuity improved in 7 cases. Average retinal artery pressure and arm-to-retina circulation time improved significantly to the normal level (p < 0.05). The other 30 patients complained of recurrent and worsened visual symptoms during the follow-up period. Conclusion: CAS was effective in improving ocular circulation, and also improved the chronic ocular ischemic syndrome caused by the severe carotid artery stenosis.
Prognostic and Therapeutic Significance of Carbohydrate Antigen 19-9 as Tumor Marker in Patients with Pancreatic Cancer
Authors: Boeck Stefan , Stieber Petra , Holdenrieder Stefan , Wilkowski Ralf , Heinemann Volker
Published: August 2006

In pancreatic cancer (PC) accurate determination of treatment response by imaging often remains difficult. Various efforts have been undertaken to investigate new factors which may serve as more appropriate surrogate parameters of treatment efficacy. This review focuses on the role of carbohydrate antigen 19-9 (CA 19-9) as a prognostic tumor marker in PC and summarizes its contribution to monitoring treatment efficacy. We undertook a Medline/PubMed literature search to identify relevant trials that had analyzed the prognostic impact of CA 19-9 in patients treated with surgery, chemoradiotherapy and chemotherapy for PC. Additionally, relevant abstract publications from scientific meetings were included. In advanced PC, pretreatment CA 19-9 levels have a prognostic impact regarding overall survival. Also a CA 19-9 decline under chemotherapy can provide prognostic information for median survival. A 20% reduction of CA 19-9 baseline levels within the first 8 weeks of chemotherapy appears to be sufficient to define a prognostic relevant subgroup of patients (‘CA 19-9 responder’). It still remains to be defined whether the CA 19-9 response is a more reliable method for evaluating treatment efficacy compared to conventional imaging.
Effects of Alpha Tocopherol and Probucol Supplements on Allergen-Induced Airway Inflammation and Hyperresponsiveness in a Mouse Model of Allergic Asthma
Authors: Okamoto Nami , Murata Takuji , Tamai Hiroshi , Tanaka Hiroyuki , Nagai Hiroichi
Published: August 2006

Objective: We investigated the role of antioxidants in airway hyperresponsiveness to acetylcholine using young asthma model mice, which were sensitized and stimulated with ovalbumin. Methods: The mice had been fed either a normal diet, an α-tocopherol-supplemented diet or a probucol-supplemented diet 14 days before the first sensitization. They were immunized with antigen at intervals of 12 days and, starting from 10 days after the second immunization, they were exposed to antigen 3 times every 4th day using an ultrasonic nebulizer. Twenty-four hours after the last antigen inhalation, airway responsiveness to acetylcholine was measured and bronchoalveolar lavage fluid (BALF) was collected. A blood and lung tissue study was also carried out. Results: Twenty-four hours after the last antigen challenge, both IL-4 and IL-5 in the BALF of α-tocopherol-supplemented mice were significantly decreased. The IL-5 level in probucol-supplemented mice was also decreased, but there was no difference in IL-4 levels. The serum IgE level was decreased in probucol-supplemented mice. Differential cell rates of the fluid revealed a significant decrease in eosinophils due to antioxidant supplementation. Airway hyperresponsiveness to acetylcholine was also repressed in antioxidant-supplemented mice. In histological sections of lung tissue, inflammatory cells and mucus secretion were markedly reduced in antioxidant-supplemented mice. We investigated the antioxidant effect on our model mice by examining 8-isoprostane in BALF and lung tissue, and acrolein in BALF; however, our experiment gave us no evidence of the antioxidant properties of either α-tocopherol or probucol contributing to the reduction of airway inflammation. Conclusion: These findings indicate that α-tocopherol and probucol suppress allergic responses in asthma model mice, although these two drugs cause suppression in different ways that are unrelated to antioxidation.
B Cell-Activating Factor Is a Novel Diagnosis Parameter for Asthma
Authors: Kang Jong-Soon , Yoon Yeo-Dae , Ahn Joong-Hyun , Kim Seok-Chan , Kim Kwan-Hyoung , Kim Hwan-Mook , Moon Eun-Yi
Published: August 2006

Background: Asthma is a life-threatening immediate-type allergic disease. B cell-activating factor (BAFF) is a key regulator of B lymphocyte development and is required to generate and maintain the mature B cell pool. Objectives: To investigate the level of BAFF in the serum of asthma patients and the role of BAFF on T cells. Methods: The BAFF level was measured by enzyme-linked immunosorbent assay. Peripheral blood mononuclear cells (PBMC) from asthma patients were analyzed by flow cytometry. T8.1 cells were used to test the role of BAFF on T cell-antigen-presenting cell (APC) conjugate formation. Results: The BAFF level in patient serum was elevated relative to normal serum. Immunoglobulin E (IgE) concentration and the percentage of CD3+ T and CD19+ B cells vary according to the serum BAFF level. Patients with high BAFF and high IgE (group II) and those with high BAFF and low IgE (group III) show a high ratio of CD3+ T to CD19+ B cells, and the opposite is seen for patients with low BAFF and high IgE (group I) and those with low BAFF and low IgE (group IV). The addition of BAFF increased PBMC proliferation and T cell-APC conjugate formation. BAFF concentration in serum decreased after treatment with antiasthmatic drugs including glucocorticoids and immunosuppressants. Conclusion: These findings suggest that the serum BAFF level is high in both IgE-mediated asthma and non-IgE-mediated asthma and extend our knowledge about the fact that BAFF may play a stimulatory role on the proliferation of T cells. Thus, BAFF could be a parameter to monitor the severity of asthma symptoms.
Effects of Calcitriol on Type 5b Tartrate-Resistant Acid Phosphatase and Interleukin-6 in Secondary Hyperparathyroidism
Authors: Lu Kuo-Cheng , Tseng Chin-Feng , Wu Chia-Chao , Yeung Lai-King , Chen Jin-Shuen , Chao Tsu-Yi , Janckila Anthony J. , Yam Lung T. , Chu Pauling
Published: August 2006

Background/Aims: Secondary hyperparathyroidism (SHP) is characterized by high bone turnover and elevated serum bone remodeling markers. Elevation of serum interleukin-6 (IL-6) levels is also characteristic of end-stage renal disease. This study investigates the effects of intravenous calcitriol on serum bone resorptive markers, namely, type 5b tartrate-resistant acid phosphatase (TRACP5b) and IL-6 in patients with SHP. Methods: Intravenous calcitriol therapy was given for 16 weeks to 24 patients on maintenance hemodialysis with plasma intact parathyroid hormone (iPTH) levels >300 pg/ml. Blood was drawn at baseline and every 4 weeks for 16 weeks for determination of the levels of biochemical parameters, iPTH, IL-6 and bone remodeling markers, including bone-specific alkaline phosphatase (bAP) and TRACP5b. Results: Only 21 patients responded to the calcitriol therapy, with significant decrements in serum iPTH after 4 weeks of therapy and thereafter. After 16 weeks of calcitriol therapy, 21 patients had significant decrements in serum iPTH (707.9 ± 317.8 vs. 205.0 ± 63.1 pg/ml, p < 0.01). Prior to treatment, a significant correlation was found between increased levels of serum iPTH and IL-6 levels (r = 0.45, p < 0.05). After treatment, there was also a significant and parallel lowering of levels of serum iPTH, IL-6 (8.52 ± 3.59 vs. 7.24 ± 2.81 pg/ml, p < 0.01), bAP (54.68 ± 36.17 vs. 24.55 ± 13.84 U/l, p < 0.01) and TRACP5b (3.41 ± 1.89 vs. 1.80 ± 0.55 U/l, p < 0.01). Our results additionally showed significant positive correlationsbetween baseline levels of serum IL-6 and those of iPTH, bAP and TRACP5b. After 16 weeks of calcitriol treatment, the correlation between IL-6 and iPTH levels lost significance but levels of serum IL-6, bAP and TRACP5b remained significantly correlated. Conclusions: Elevated levels of serum IL-6 and bone remodeling markers, namely, bAP and TRACP5b which are common features of SHP, are effectively suppressed by calcitriol therapy. This indicates that hyperparathyroidism not only accelerates bone remodeling but may also aggravate inflammation in patients on maintenance hemodialysis.
l
-Caldesmon Regulates Proliferation and Migration of Vascular Smooth Muscle Cells and Inhibits Neointimal Formation After Angioplasty
Authors: Yokouchi Kazuhiko , Numaguchi Yasushi , Kubota Ryuji , Ishii Masakazu , Imai Hajime , Murakami Ryuichiro , Ogawa Yasuhiro , Kondo Takahisa , Okumura Kenji , Ingber Donald E. , Murohara Toyoaki
Published: August 2006

Objective—

Light-type caldesmon (
l
-CaD) is a potent cytostatic and antiangiogenic protein that regulates cell growth and survival via modulation of the cell shape and cytoskeleton. The aim of this study is to explore the potential value of
l
-CaD for use as a cytostatic agent to inhibit neointimal formation after angioplasty by suppressing vascular smooth muscle cell (VSMC) growth and migration.

Methods and Results—

We tested the cytostatic function of
l
-CaD in cultured VSMCs using assays for apoptosis, cell proliferation, and migration, and evaluated the expression pattern of relevant signaling proteins (focal adhesion kinase [FAK] and mitogen-activated protein kinases) in VSMCs. Transfection of adenoviral vector encoding
l
-CaD (Ad-
l
-CaD) resulted in progressive loss of actin stress fibers and cell retraction. Enzyme-linked immunosorbent assay demonstrated that Ad-
l
-CaD transfection increased the apoptosis rate by 75% and reduced BrdU uptake by 49%. Furthermore, transfection of Ad-
l
-CaD inhibited migration of VSMCs induced by platelet-derived growth factor-BB (PDGF) by 36% (
P
<0.05). Immunoblotting analysis revealed that
l
-CaD overexpression reduced PDGF-induced phosphorylation of both FAK and extracellular signal regulated-kinase (ERK). In balloon-injured rat carotid arteries, Ad-
l
-CaD transfection inhibited neointimal formation by 37% (
P
<0.05) without delaying re-endothelialization at 14 days.

Conclusions—

Overexpression of
l
-CaD suppressed cell growth and survival in VSMCs and inhibited neointimal formation after experimental angioplasty, partly by regulating the cytoskeletal tension-FAK-ERK axis.
Quantitation and Localization of Matrix Metalloproteinases and Their Inhibitors in Human Carotid Endarterectomy Tissues
Authors: Choudhary Salman , Higgins Catherine L. , Chen Iou Yih , Reardon Michael , Lawrie Gerald , Vick G. Wesley , Karmonik Christof , Via David P. , Morrisett Joel D.
Published: August 2006

Background—

Matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) play a central role in arterial wall remodeling, affecting stability of fibrous caps covering atherosclerotic plaques. The objective of this study was to determine the spatial distribution of TIMP mass and MMP mass and activity of carotid endarterectomy (CEA) tissues and relate it to the distribution of atherosclerotic lesions.

Methods and Results—

Fresh CEA tissues were imaged by multicontrast MRI to generate 3D reconstructions. Tissue segments were cut transversely from the common, bifurcation, internal, and external regions. Segments were subjected to total protein extractions and analyzed by ELISA for MMP-2 and -9 and TIMP-1 and -2 mass and by zymography for gelatinase activity. Segments at or near the bifurcation with highly calcified lesions contained higher MMP levels and activity than segments distant from the bifurcation; highly fibrotic or necrotic plaque contained lower MMP levels and activity and higher TIMP levels. Fatty streak, fibroatheroma with hemorrhage and calcification, and fully occluded lesions were enriched in MMP-2, MMP-9, and TIMP-1 and TIMP-2, respectively.

Conclusion—

The spatial distribution of MMPs and TIMPs in carotid atherosclerotic lesions is highly heterogeneous, reflecting lesion location, size, and composition. This study provides the first semi-quantitative maps of differential distribution of MMPs and TIMPs over atherosclerotic plaques.
Blockade of Angiotensin II Receptors Reduces the Expression of Receptors for Advanced Glycation End Products in Human Endothelial Cells
Authors: Fujita Masashi , Okuda Hiroko , Tsukamoto Osamu , Asano Yoshihiro , Hirata Yulin Liao Akio , Kim Jiyoong , Miyatsuka Takeshi , Takashima Seiji , Minamino Tetsuo , Tomoike Hitonobu , Kitakaze Masafumi
Published: August 2006

Objectives—

Receptors for advanced glycation end products (RAGEs) play crucial roles in atherogenesis. Because tumor necrosis factor α (TNFα) is expressed and upregulates RAGE expression in atherosclerotic lesions, the TNFα-RAGE interaction might be involved in the inflammatory process of atherogenesis. On the other hand, an angiotensin II type-1 receptor blocker (ARB), widely used as an antihypertensive drug, has been reported to have also antiatherosclerotic effects. Thus we investigated whether an ARB exerts antiatherosclerotic effects via inhibiting the TNFα-RAGE interaction.

Methods and Results—

Stimulation of human endothelial cells with candesartan as well as olmesartan decreased TNFα-induced RAGE expression in both mRNA and protein levels along with the decrease in the activity of nuclear factor κB and the expression of inflammatory mediators such as vascular cell adhesion molecule (VCAM)-1. Both candesartan and olmesartan inhibited the binding of nuclear factor κB to the RAGE gene promoter. Furthermore, gene silencing of RAGE by RNA interference decreased the expression of TNFα-induced VCAM-1 in both mRNA and protein levels.

Conclusions—

RAGE contributes at least partially to the TNFα-induced VCAM-1 expression in both mRNA and protein levels. Blockade of angiotensin II receptors might exert antiatherosclerotic effects via reducing TNFα-RAGE interaction.
Angiotensin II Type 1 Receptor–Mediated Inflammation Is Required for Choroidal Neovascularization
Authors: Nagai Norihiro , Oike Yuichi , Izumi-Nagai Kanako , Urano Takashi , Kubota Yoshiaki , Noda Kousuke , Ozawa Yoko , Inoue Makoto , Tsubota Kazuo , Suda Toshio , Ishida Susumu
Published: August 2006

Background—Choroidal neovascularization (CNV) is a critical pathogenesis in age-related macular degeneration, the most common cause of blindness in the developed countries. The aim of the current study was to determine the involvement of the renin-angiotensin system (RAS) with the development of CNV, using human surgical samples and the murine model of laser-induced CNV.

Methods and Results—In the human and murine CNV tissues, the vascular endothelium expressed angiotensin II type 1 receptor (AT1-R), AT2-R, and angiotensin II. The CNV volume was significantly suppressed by treatment with an AT1-R blocker telmisartan, but not with an AT2-R blocker. AT1-R signaling blockade with telmisartan inhibited various inflammatory mechanisms including macrophage infiltration and upregulation of VEGF, intercellular adhesion molecule-1 (ICAM-1), MCP-1, and IL-6 in the retinal pigment epithelium-choroid complex. A PPAR-γ antagonist partially but significantly reversed the suppressive effect of telmisartan on in vivo induction of CNV and in vitro upregulation of ICAM-1 and MCP-1 in endothelial cells and IL-6 in macrophages, showing the dual contribution of PPAR-γ-agonistic and AT1-R-antagonistic actions in the telmisartan treatment.

Conclusions—AT1-R–mediated inflammation plays a pivotal role in the development of CNV, indicating the possibility of AT1-R blockade as a novel therapeutic strategy to inhibit CNV.
Basilar Artery Diameter and 5-Year Mortality in Patients With Stroke
Authors: Pico Fernando , Labreuche Julien , Gourfinkel-An Isabelle , Amarenco Pierre
Published: August 2006

Background and Purpose—

Few and conflicting data exist on the case fatality rate in stroke patients with basilar artery dolichoectasia. We analyzed basilar artery characteristics (diameter, height of bifurcation, transverse position) and 5-year mortality (all-cause, nonstroke vascular, and stroke) in patients with brain infarction.

Methods—

The basilar artery diameter was measured with a 16-diopter lens in 466 consecutively recruited patients with brain infarction confirmed by magnetic resonance imaging. The height of the bifurcation and the transverse position of the basilar artery were assessed on semiquantitative scales. Patients were followed up for a median of 5.3 years (range, 1.5 to 6.6) and were classified as having had stroke, nonstroke vascular, and nonvascular death according to the French national registry of death certificates.

Results—

Of the 157 deaths, 88 were vascular (including 54 stroke deaths). Basilar artery diameter was associated with an increased 5-year stroke mortality rate but not with all-cause or nonstroke vascular mortality. The adjusted hazard ratio (HR) of stroke mortality per 1-mm increase in basilar artery diameter was 1.23 (95% confidence interval [CI], 1.07 to 1.41). A higher risk of stroke death was associated with basilar artery diameter at the 95th percentile (diameter >4.3 mm; adjusted HR 3.69; 95% CI, 1.63 to 8.38) and the height of bifurcation (adjusted HR for score >1, 2.08; 95% CI, 0.93 to 4.68) but not with transverse position.

Conclusions—

Basilar artery diameter was independently associated with cerebrovascular mortality. A diameter >4.3 mm may be a marker for a high risk of fatal stroke.
Recent Advances of Intervention to Inhibit Progression of Carotid Intima-Media Thickness in Patients With Type 2 Diabetes Mellitus
Authors: Yokoyama Hiroki , Katakami Naoto , Yamasaki Yoshimitsu
Published: August 2006

Background and Purpose—

Type 2 diabetes is associated with a high cardiovascular morbidity and mortality. Recent advances of intervention studies in type 2 diabetes with use of carotid intima-media thickness (CIMT) measurement as a surrogate end point may allow for better understanding of the undetermined process of atherosclerosis, the effect of interventions, and the usefulness of CIMT to inhibit events of cardiovascular disease.

Summary of Review—

Data were available from 11 studies (n=1578) in subjects with type 2 diabetes (including impaired glucose tolerance, n=132) that evaluated the effect of interventions on change in CIMT. The overall weighed rate of change in mean CIMT based on data among control groups (ie, type 2 diabetes without interventions) was 0.034 mm/y (95% CI, 0.029 to 0.039; median SD, 0.054), in which mean HbA
1c
was 7.86% (95% CI, 7.72 to 8.00; median SD, 1.5). A significant close correlation of HbA
1c
with rate of CIMT change was found (
R
2
=0.35,
P
=0.01). Agents for lowering of blood glucose, platelet activation, or blood pressure significantly reduced the CIMT increase, independent of blood glucose control. This implies that other mechanisms of such agents to diminish CIMT increase should be explored.

Conclusions—

CIMT measurement may contribute to elucidating the short- and/or long-term effect of interventions on the rate of change in CIMT in relation to the levels of various risk factors. Although the method needs further standardization, pharmacological interventions are likely to inhibit progression of CIMT, leading to a reduction of cardiovascular events.
Additive Role of Plasma von Willebrand Factor Levels to Clinical Factors for Risk Stratification of Patients With Atrial Fibrillation
Authors: Lip Gregory Y.H. , Lane Deirdre , Van Walraven Carl , Hart Robert G.
Published: August 2006

Background and Purpose—

To aid decisions for thromboprophylaxis in atrial fibrillation (AF), several risk stratification schemes that predict stroke risk according to clinical and echocardiographic features have been published. von Willebrand factor (vWf) is a plasma markers of endothelial damage/dysfunction and is associated with the risk of stroke and vascular events in AF patients. This study determined the additive role of plasma vWf levels to clinical factors for risk stratification in patients with AF.

Methods—

We classified 994 AF patients who were enrolled in the SPAF III trial as being at low, moderate, or high risk of stroke and thromboembolism according to the Birmingham and CHADS
2
risk stratification schemes. vWf levels were classified as elevated when ≥158 IU /dL. Rates of ischemic stroke and vascular events within each clinical risk stratum with and without plasma vWf levels were compared.

Results—

The accuracy of both clinical risk stratification schemes was similar for predicting event rates (Birmingham: ischemic strokes, 0.642; vascular events, 0.670; CHADS
2
: ischemic strokes, 0.672; vascular events, 0.672). Subsequent addition of categorized vWf levels to both clinical risk stratification schemes further refined risk stratification for stroke and vascular events. When added to the Birmingham and CHADS
2
clinical risk stratification, high vWf levels were independently associated with a risk of vascular events (hazard ratio, 2.05; 95% confidence interval, 1.30 to 3.22 and 2.01, 1.27 to 3.18 with Birmingham and CHADS
2
, respectively) but not ischemic stroke.

Conclusions—

When added to clinical risk stratification schemes (Birmingham; CHADS
2
), plasma vWf levels refined clinical risk stratification for stroke and vascular events among AF patients. vWf levels may aid decisions about thromboprophylaxis, particularly among AF patients at moderate risk.
Diminished proteasomal degradation results in accumulation of Gfi1 protein in monocytes
Authors: Marteijn Jurgen A. F. , van der Meer Laurens T. , Van Emst Liesbeth , de Witte Theo , Jansen Joop H. , van der Reijden Bert A.
Published: August 2006

Gfi1 is a transcriptional repressor essential during myeloid differentiation. Gfi1−/− mice exhibit a block in myeloid differentiation resulting in the accumulation of an immature myelo-monocytic cell population and the complete absence of mature neutrophils. Even though mRNA levels of Gfi1 appear to be very low in monocytes, Gfi1 might play a role in the monocytic lineage as Gfi1−/− mice exhibit diminished monocyte-derived dendritic cells and disturbed cytokine production by macrophages in response to LPS. We show here that Gfi1 protein levels are mainly regulated by the ubiquitin-proteasome system. Upon forced monocytic differentiation of U937 cells, Gfi1 mRNA levels dropped but protein levels increased due to diminished proteasomal turnover. Similarly, Gfi1 mRNA levels are low in primary monocytes whereas the protein is clearly detectable. Conversely, Gfi1 mRNA levels are high in granulocytes but the protein is swiftly degraded by the proteasome in these cells. Chromatin immunoprecipitation experiments showed that Gfi1 binds to the promoter of several granulocyte-specific genes in primary monocytes, including C/EBPα, neutrophil elastase, and Gfi1 itself. The binding of the repressor Gfi1 to these promoters correlated with low expression of these genes in monocytes compared with granulocytes. Our data fit a model in which Gfi1 protein levels are induced in primary monocytes, due to diminished proteasomal degradation, to repress genes that play a role in granulocytic differentiation.
Overlapping functions of human CD3δ and mouse CD3γ in αβ T-cell development revealed in a humanized CD3γ-deficient mouse
Authors: Fernández-Malavé Edgar , Wang Ninghai , Pulgar Manuel , Schamel Wolfgang W. A. , Alarcón Balbino , Terhorst Cox
Published: August 2006

Humans lacking the CD3γ subunit of the pre-TCR and TCR complexes exhibit a mild αβ T lymphopenia, but have normal T cells. By contrast, CD3γ-deficient mice are almost devoid of mature αβ T cells due to an early block of intrathymic development at the CD4–CD8– double-negative (DN) stage. This suggests that in humans but not in mice, the highly related CD3δ chain replaces CD3γ during αβ T-cell development. To determine whether human CD3δ (hCD3δ) functions in a similar manner in the mouse in the absence of CD3γ, we introduced an hCD3δ transgene in mice that were deficient for both CD3δ and CD3γ, in which thymocyte development is completely arrested at the DN stage. Expression of hCD3δ efficiently supported pre-TCR–mediated progression from the DN to the CD4+CD8+ double-positive (DP) stage. However, αβTCR-mediated positive and negative thymocyte selection was less efficient than in wild-type mice, which correlated with a marked attenuation of TCR-mediated signaling. Of note, murine CD3γ-deficient TCR complexes that had incorporated hCD3δ displayed abnormalities in structural stability resembling those of T cells from CD3γ-deficient humans. Taken together, these data demonstrate that CD3δ and CD3γ play a different role in humans and mice in pre-TCR and TCR function during αβ T-cell development.
Human CD4+ T lymphocytes with increased intracellular cAMP levels exert regulatory functions by releasing extracellular cAMP
Authors: Vendetti Silvia , Patrizio Mario , Riccomi Antonella , De Magistris Maria Teresa
Published: August 2006

We have previously shown that cholera toxin (CT) and other cAMP-elevating agents induce up-regulation of the inhibitory molecule CTLA-4 on human resting T lymphocytes. In this study, we evaluated the function of these cells. We found that purified human CD4+ T lymphocytes pretreated with CT were able to inhibit proliferation of autologous PBMC in a dose-dependent manner. It is interesting that this phenomenon was not mediated by inhibitory cytokines such as IL-10, IL-4, or TGF-β but was in part caused by the release of extracellular cAMP by the CD4+ T lymphocytes. Purified CD4+ T cells pretreated with forskolin, a transient cAMP inducer, or with dibutyryl cAMP, an analog of cAMP, did not exert suppressive functions, suggesting that a sustained production of cAMP, such as that induced by CT, was required to identify a novel regulatory function mediated by CD4+ T cells. Our results show that CD4+ T lymphocytes can exert regulatory functions through the release of extracellular cAMP and that the cyclic nucleotide acts as a primary messenger, which could play a biological role in the modulation of immune responses.
Suppression of experimental colitis by intestinal mononuclear phagocytes
Authors: Qualls Joseph E , Kaplan Alan M , Van Rooijen Nico , Cohen Donald A
Published: August 2006

The contribution of innate immunity to inflammatory bowel disease (IBD) remains an area of intense interest. Macrophages (MØ) and dendritic cells (DC) are considered important factors in regulating the onset of IBD. The goal of this study was to determine if intestinal mononuclear phagocytes (iMNP) serve a pathological or protective role in dextran sulfate sodium (DSS)-induced colitis in mice. Using a conditional MØ/DC depletion transgenic mouse line—MØ Fas-induced apoptosis—to systemically deplete iMNP, DSS colitis histopathology was shown to be more severe in MØ/DC-depleted compared with MØ/DC-intact mice. Similarly, localized iMNP depletion by clodronate-encapsulated liposomes into C57BL/6, BALB/c, and CB.17/SCID mice also increased DSS colitis severity, as indicated by increased histopathology, weight loss, rectal bleeding, decreased stool consistency, and colon length compared with MØ/DC-intact, DSS-treated mice. Histology revealed that iMNP depletion during DSS treatment led to increased neutrophilic inflammation, increased epithelial injury, and enhanced mucin depletion from Goblet cells. iMNP depletion did not further elevate DSS-induced expression of TNF-α and IFN-γ mRNA but significantly increased expression of CXCL1 chemokine mRNA. Myeloperoxidase activity was increased in colons of MØ/DC-depleted, DSS-treated mice, compared with DSS alone, coincident with increased neutrophil infiltration in diseased colons. Neutrophil depletion combined with MØ/DC depletion prevented the increase in DSS colitis severity compared with MØ/DC depletion alone. This study demonstrates that iMNP can serve a protective role during development of acute colitis and that protection is associated with MØ/DC-mediated down-regulation of neutrophil infiltration.
Epigenetic Regulation of Human Trophoblastic Cell Migration and Invasion
Authors: Rahnama Fahimeh , Shafiei Farhad , Gluckman Peter D. , Mitchell Murray D. , Lobie Peter E.
Published: August 2006

Pivotal to successful mammalian reproduction is the ability of a developing embryo to implant to the uterine wall and establish a nutrient supply via placentation. Herein, we have examined the potential epigenetic regulation of human trophoblastic cell migration and invasion by use of the choriocarcinoma cell line, BeWo. Treatment of BeWo cells with a DNA methyltransferase inhibitor, 5′-aza-2′-deoxycytidine (AZA), resulted in conversion of cell morphology to a nonmigratory phenotype. This was exemplified by the ability of AZA to prevent BeWo cell migration in wound healing and transwell migration assays. AZA consequently inhibited BeWo cell invasion through reconstituted basement membrane. Examination of components of the adherens junction complex pivotal for determination of cell phenotype revealed that AZA specifically increased the mRNA level of E-cadherin and plakoglobin (γ-catenin), but not α-catenin and β-catenin. AZA also increased the gene promoter activity of both plakoglobin and E-cadherin. Protein levels of both plakoglobin and E-cadherin were increased by AZA, and AZA enhanced their localization to sites of intercellular contact. Forced expression of plakoglobin and E-cadherin abrogated BeWo cell migration, indicative that repression of these genes was required for BeWo cell migration. Small interfering RNA-mediated depletion of the individual DNA methyltransferase (DNMT) molecules did not affect plakoglobin and E-cadherin promoter activity or BeWo cell migration. However, increases in plakoglobin and E-cadherin promoter activity and inhibition of BeWo cell migration was achieved with small interfering RNA-mediated depletion of both DNMT-3a and DNMT-3b. Epigenetic regulation of plakoglobin and E-cadherin is therefore pivotal for appropriate trophoblastic invasion in vitro.
Somatostatin Regulates Aggressive Behavior in an African Cichlid Fish
Authors: Trainor Brian C. , Hofmann Hans A.
Published: August 2006

Animals respond to environmental and social change with plasticity in the neural substrates underlying particular behavioral states. In the African cichlid fish Astatotilapia burtoni, social dominance status in males is accompanied by reduced somatic growth rate as well as increased somatostatin neuron size in the preoptic area. Although somatostatin is commonly studied within the context of growth, we show here for the first time that this ancient neuropeptide also plays a role in controlling social behavior. Somatostatin antagonists increased aggressive behavior in a dose-dependent fashion and the potent somatostatin agonist octreotide decreased aggression. We cloned and sequenced the genes encoding two somatostatin receptor subtypes in this species to study transcription in the gonads. When we examined somatostatin receptor gene expression in testes, expression of the somatostatin type 3 receptor was negatively correlated with an aggressive display and androgen levels. However, octreotide treatment did not reduce plasma testosterone or 11-ketotestosterone levels, suggesting that the behavioral effects of somatostatin are not mediated by androgens. These results show that somatostatin has important effects on social behavior. In dominant male A. burtoni, somatostatin may function to contain energetically costly processes such as somatic growth and aggressive behavior.
Differential Expression of Activator Protein-1 Proteins in the Pineal Gland of Syrian Hamster and Rat May Explain Species Diversity in Arylalkylamine N-Acetyltransferase Gene Expression
Authors: Sinitskaya Natalia , Salingre Anthony , Klosen Paul , Revel Florent G. , Pévet Paul , Simonneaux Valérie
Published: August 2006

Species differences have been reported for the nighttime regulation of arylalkylamine N-acetyltransferase (AA-NAT), the melatonin rhythm-generating enzyme. In particular, de novo synthesis of stimulatory transcription factors is required for Aa-nat transcription in the Syrian hamster but not in the rat pineal gland. The present work investigated the contribution of phosphorylated cAMP-responsive element-binding protein, c-FOS, c-JUN, and JUN-B in the regulation of Aa-nat transcription in Syrian hamsters compared with rats. The nighttime pattern of cAMP-responsive element-binding protein phosphorylation and regulation by norepinephrine observed in the Syrian hamster was similar to those reported in the rat. On the contrary, strong divergences in c-FOS, c-JUN, and JUN-B expression were observed between both species. In Syrian hamster, predominant expression of c-FOS and c-JUN was observed at the beginning of night, whereas a predominant expression of c-JUN and JUN-B was observed in the late night in rat. The early peak of c-FOS and c-JUN, known to form a stimulatory transcription dimer, suggests that they are involved in the nighttime stimulation of Aa-nat transcription. Indeed, early-night administration of a protein synthesis inhibitor (cycloheximide) markedly decreased AA-NAT mRNA levels in Syrian hamster. In the rat, high levels of JUN-B and c-JUN, constituting an inhibitory transcription dimer, are probably involved in the late-night inhibition of Aa-nat transcription. Early-night administration of cycloheximide actually increased AA-NAT mRNA levels toward the late night. Therefore, composition and timing of the pineal activator protein-1 complexes differ between rat and Syrian hamster and may be an activator (Syrian hamster) or an inhibitor (rat) of Aa-nat transcription.
Inverse Relationship between Nitric Oxide Synthases and Endothelin-1 Synthesis in Bovine Corpus Luteum: Interactions at the Level of Luteal Endothelial Cell
Authors: Rosiansky-Sultan Maya , Klipper Eyal , Spanel-Borowski Katharina , Meidan Rina
Published: August 2006

Endothelin-1 (ET-1) and nitric oxide (NO) play pivotal roles in corpus luteum (CL) function. The present study examined the interplay between NO and ET-1 synthesis in the bovine CL. We found similar inducible and endothelial NO synthase (iNOS and eNOS, respectively) activities in the young CL (d 1–5) expressing the highest levels of both eNOS and iNOS mRNA. These values later declined at mid-cycle (d 8–15) and remained low at later stages (d 16–18). Luteolysis, initiated by prostaglandin F2α analog administration, further reduced NOS mRNA and by 24 h, NOS values dropped to approximately 15% of those at mid-cycle. eNOS protein levels followed a similar pattern to its mRNA. Because endothelial cells (ECs) are the main site for ET-1 and NO production in the CL, we examined the direct effects of the NO donor, NONOate on luteal ECs (LECs). Elevated NO levels markedly decreased ET-1 mRNA, and peptide concentrations in cultured and freshly isolated LECs in a dose-dependent manner. In agreement, NOS inhibitor, NG-nitro-l-arginine methyl ester, stimulated ET-1 mRNA expression in these cells. Interestingly, NO also up-regulated prostaglandin F2α receptors in LECs. These data show that there is an inverse relationship between NOS and ET-1 throughout the CL life span, and imply that this pattern may be the result of their interaction within the resident LECs. NOS are expressed in a physiologically relevant manner: elevated NO at an early luteal stage is likely to play an important role in angiogenesis, whereas reduced levels of NO during luteal regression may facilitate the sustained up-regulation of ET-1 levels during luteolysis.
A Novel Pathway Involving Progesterone Receptor, Endothelin-2, and Endothelin Receptor B Controls Ovulation in Mice
Authors: Palanisamy Gopinath S. , Cheon Yong-Pil , Kim Jaeyeon , Kannan Athilakshmi , Li Quanxi , Sato Marcey , Mantena Srinivasa R. , Sitruk-Ware Regine L. , Bagchi Milan K. , Bagchi Indrani C.
Published: August 2006

The steroid hormone progesterone (P) plays a pivotal role during ovulation. Mice lacking P receptor (Pgr) gene fail to ovulate due to a defect in follicular rupture. The P receptor (PGR)-regulated pathways that modulate ovulation, however, remain poorly understood. To identify these pathways, we performed gene expression profiling using ovaries from mice subjected to gonadotropin-induced superovulation in the presence and in the absence of CDB-2914, a synthetic PGR antagonist. Prominent among the genes that were down-regulated in response to CDB-2914 was endothelin (ET)-2, a potent vasoactive molecule. ET-2 mRNA was transiently induced in mural granulosa cells of the preovulatory follicles immediately preceding ovulation. This induction was absent in the ovaries of PGR null mice, indicating a critical role of this receptor in ET-2 expression. To investigate the functional role of ET-2 during ovulation, we employed selective antagonists of endothelin receptors, ETR-A and ETR-B. Mice treated with an ETR-B antagonist exhibited a dramatic (>85%) decline in the number of released oocytes. Strong expression of ETR-B was observed in the mural and cumulus granulosa cells of the preovulatory follicles as well as in the capillaries lining the inner border of the theca interna. We also identified cGMP-dependent protein kinase II, a previously reported PGR-regulated gene, as a downstream target of ET-2 during ovulation. Collectively, our studies uncovered a unique pathway in which ET-2, produced by PGR in mural granulosa cells, acts in a paracrine or autocrine manner on multiple cell types within the preovulatory follicle to control the final events leading to its rupture.
Peroxisome Proliferator-Activated Receptor-γ Activates p53 Gene Promoter Binding to the Nuclear Factor-κB Sequence in Human MCF7 Breast Cancer Cells
Authors: Bonofiglio Daniela , Aquila Saveria , Catalano Stefania , Gabriele Sabrina , Belmonte Maria , Middea Emilia , Qi Hongyan , Morelli Catia , Gentile Mariaelena , Maggiolini Marcello , Andò Sebastiano
Published: August 2006

The aim of the present study was to provide new mechanistic insight into the growth arrest and apoptosis elicited by peroxisome proliferator-activated receptor (PPAR)γ in breast cancer cells. We ascertained that PPARγ mediates the inhibition of cycle progression in MCF7 cells exerted by the specific PPARγ agonist rosiglitazone [BRL4653 (BRL)], because this response was no longer notable in the presence of the receptor antagonist GW9662. We also provided evidence that BRL is able to up-regulate mRNA and protein levels of the tumor suppressor gene p53 and its effector p21WAF1/Cip1 in a time- and dose-dependent manner. Moreover, in transfection experiments with deletion mutants of the p53 gene promoter, we documented that the nuclear factor-κB sequence is required for the transcriptional response to BRL. Interestingly, EMSA showed that PPARγ binds directly to the nuclear factor-κB site located in the promoter region of p53, and chromatin immunoprecipitation experiments demonstrated that BRL increases the recruitment of PPARγ on the p53 promoter sequence. Next, both PPARγ and p53 were involved in the cleavage of caspases-9 and DNA fragmentation induced by BRL, given that GW9662 and an expression vector for p53 antisense blunted these effects. Our findings provide evidence that the PPARγ agonist BRL promotes the growth arrest and apoptosis in MCF7 cells, at least in part, through a cross talk between p53 and PPARγ, which may be considered an additional target for novel therapeutic interventions in breast cancer patients.
Cooperative activities ofDrosophilaDE-Cadherin and DN-Cadherin regulate the cell motility process of ommatidial rotation
Authors: Mirkovic Ivana , Mlodzik Marek
Published: August 2006

Ommatidial rotation is a cell motility read-out of planar cell polarity(PCP) signaling in the Drosophila eye. Although the signaling aspects of PCP establishment are beginning to be unraveled, the mechanistic aspects of the associated ommatidial rotation process remain unknown. Here, we demonstrate that the Drosophila DE- and DN-cadherins have opposing effects on rotation. DE-cadherin promotes rotation, as DE-cad mutant ommatidia rotate less than wild type or not at all. By contrast, the two DN-cadherins act to restrict this movement, with ommatidia rotating too fast in the mutants. The opposing effects of DE- and DN-cadherins result in a coordinated cellular movement, enabling ommatidia of the same stage to rotate simultaneously. Genetic interactions, phenotypic analysis and localization studies indicate that EGF-receptor and Frizzled-PCP signaling feed into the regulation of cadherin activity and localization in this context. Thus, DE-and DN-cadherins integrate inputs from at least two signaling pathways,resulting in a coordinated cell movement. A similar input into mammalian E-and N-cadherins might function in the progression of diseases such as metastatic ovarian cancer.
Smad5 determines murine amnion fate through the control of bone morphogenetic protein expression and signalling levels
Authors: Bosman Erika A. , Lawson Kirstie A. , Debruyn Joke , Beek Lisette , Francis Annick , Schoonjans Luc , Huylebroeck Danny , Zwijsen An
Published: August 2006

Smad5 is an intracellular mediator of bone morphogenetic protein (Bmp)signalling. It is essential for primordial germ cell (PGC) development, for the development of the allantois and for amnion closure, as demonstrated by loss of Bmp signalling. By contrast, the appearance of ectopic PGC-like cells and regionalized ectopic vasculogenesis and haematopoiesis in thickened Smad5m1/m1 amnion are amnion defects that have not been associated with loss of Bmp signalling components. We show that defects in amnion and allantois can already be detected at embryonic day (E) 7.5 in Smad5 mutant mice. However, ectopic Oct4-positive (Oct4+)and alkaline phosphatase-positive (AP+) cells appear suddenly in thickened amnion at E8.5, and at a remote distance from the allantois and posterior primitive streak, suggesting a change of fate in situ. These ectopic Oct4+, AP+ cells appear to be Stella negative and hence cannot be called bona fide PGCs. We demonstrate a robust upregulation of Bmp2 and Bmp4 expression, as well as of Erk and Smad activity, in the Smad5 mutant amnion. The ectopic expression of several Bmp target genes in different domains and the regionalized presence of cells of several Bmp-sensitive lineages in the mutant amnion suggest that different levels of Bmp signalling may determine cell fate. Injection of rBMP4 in the exocoelom of wild-type embryos can induce thickening of amnion,mimicking the early amnion phenotype in Smad5 mutants. These results support a model in which loss of Smad5 results paradoxically in gain of Bmp function defects in the amnion.
Bone morphogenetic protein receptor 1A signaling is dispensable for hematopoietic development but essential for vessel and atrioventricular endocardial cushion formation
Authors: Park Changwon , Lavine Kory , Mishina Yuji , Deng Chu-Xia , Ornitz David M. , Choi Kyunghee
Published: August 2006

Bone morphogenetic protein 4 (BMP4) is crucial for the formation of FLK1-expressing (FLK1+) mesodermal cells. To further define the requirement for BMP signaling in the differentiation of blood, endothelial and smooth muscle cells from FLK1+ mesoderm, we inactivated Alk3 (Bmpr1a) in FLK1+ cells by crossing Alk3floxed/floxed and Flk1+/CreAlk3+/floxed mice. Alk3conditional knockout (CKO) mice died between E10.5 and E11.5. Unexpectedly, Alk3 CKO embryos did not show any hematopoietic defects. However, Alk3 CKO embryos displayed multiple abnormalities in vascular development, including vessel remodeling and maturation, which contributed to severe abdominal hemorrhage. Alk3 CKO embryos also displayed defects in atrioventricular canal (AVC) endocardial cushion formation in the heart. Collectively, our studies indicate a crucial role for ALK3 in vessel remodeling, vessel integrity and endocardial cushion formation during the development of the circulation system.
Disruption of early proximodistal patterning and AVE formation inApcmutants
Authors: Chazaud Claire , Rossant Janet
Published: August 2006

In the postimplantation mouse embryo, axial patterning begins with the restriction of expression of a set of genes to the distal visceral endoderm(DVE). This proximodistal (PD) axis is subsequently transformed into an anteroposterior axis as the VE migrates anteriorly to form the anterior visceral endoderm (AVE). Both Nodal and Wnt signaling pathways are involved in these events. We show here that loss of function in the adenomatous polyposis coli gene (Apc) leads to constitutive β-catenin activity that induces a proximalization of the epiblast with the activation of a subset of posterior mesendodermal genes, and loss of ability to induce the DVE. The loss of some DVE genes such as Hex and goosecoid is rescued in chimeras where only the epiblast was wild type; however, these DVE markers were no longer restricted distally but covered the entire epiblast. Thus, the Apc gene is needed in both embryonic and extraembryonic lineages for normal PD patterning around implantation, suggesting that early restricted activation of the Wnt pathway may be important for initiating axial asymmetries. In addition, we found that nuclear β-catenin and other molecular markers are asymmetrically expressed by 4.5 days.