Pathological hypertrophy of the heart is closely associated with endoplasmic reticulum stress (ERS), leading to maladaptations such as myocardial fibrosis, induction of apoptosis, and cardiac dysfunctions. Salubrinal is a known selective inhibitor of protein phosphatase 1 (PP1) complex involving dephosphorylation of phospho-eukaryotic translation initiation factor 2 subunit (p-eIF2)-α, the key signaling process in the ERS pathway. In this study, the effects of salubrinal were examined on cardiac hypertrophy using the mouse model of transverse aortic constriction (TAC) and cell model of neonatal rat ventricular myocytes (NRVMs). Treatment of TAC-induced mice with salubrinal (0.5 mg·kg-1·day-1) alleviated cardiac hypertrophy and tissue fibrosis. Salubrinal also alleviated hypertrophic growth in endothelin 1 (ET1)-treated NRVMs. Therefore, the present results suggest that salubrinal may be a potentially efficacious drug for treating pathological cardiac remodeling.

Human lung fibroblasts are a potential source of endothelin-1 (ET-1), a pro-fibrotic mediator. The present study explored possible muscarinic and β-adrenergic modulations of ET-1 expression in human lung fibroblasts. MRC-5 human lung fibroblasts were cultured. Expression of prepro-endothelin-1 (ppET-1) mRNA was determined by quantitative real time PCR. [(3)H]-Proline incorporation was determined as measure of collagen synthesis. The muscarinic agonist oxotremorine induced, in a tiotropium-sensitive manner, a three-fold increase in ppET-1 mRNA. The β(2)-adrenoceptor agonist olodaterol caused a reduction of ppET-1 mRNA by 45%. Olodaterol also opposed the stimulatory effect of oxotremorine. The effect of olodaterol was mimicked by the protein kinase A agonist 6-Bnz-cAMP, whereas the Epac (exchange protein activated by cAMP) agonist 8-CPT-2'-O-Me-cAMP was less effective. Transforming growth factor-β(1) (TGF-β, 0.3 and 1 ng/ml) induced a three- and eight-fold increase in pp-ET-1 mRNA, respectively. Olodaterol opposed the effect of 0.3, but not that of 1 ng/ml TGF-β. Likewise, 6-Bnz-cAMP opposed the effect of 0.3, but not that of 1 ng/ml TGF-β. TGF-β inhibited β(2)-adrenoceptor mRNA expression, maximally by 90%. Muscarinic agonist-induced stimulation of [(3)H]-proline incorporation was attenuated by the endothelin ET1 receptor antagonist bosentan. In conclusion, ET-1 expression in human lung fibroblasts is regulated by stimulatory muscarinic receptors and inhibitory β(2)-adrenoceptors. Since muscarinic up-regulation of ET-1 contributes to pro-fibrotic effects of muscarinic stimuli, inhibition of ET-1 expression could contribute to long-term beneficial effects of long-acting β(2)-adrenoceptor agonists and long-acting muscarinic antagonists. However, excessive exposure to TGF-β results in loss of β-adrenoceptor expression and function of its down-stream signaling.

Ca2+ transfer across the syncytiotrophoblast (ST) of the human placenta is essential for normal fetal development. However, the nature of Ca2+ conductance in the ST and the mechanisms by which it is regulated are poorly understood. With the major signal transduction pathway of endothelin-1 (ET1) acting via phospholipase C (PLC) and Ca2+, we used ET1 to analyse the nature of Ca2+ channels on cultured trophoblastic cells by means of cytofluorimetric analysis using the ratiometric Ca2+ indicator Indo-1. Results indicate that ET1 (10(-7) M) stimulates a biphasic (transient and sustained) increase in [Ca2+]i in trophoblastic cells. This response is mediated by the endothelin receptor B (ETB) coupled to PLC, since treatment with BQ788 (10(-6) M) or U73122 (2 microM) totally abolished the response. Persistence of the rapid transient rise in [Ca2+]i in Ca2+-free extracellular medium confirms the release of Ca2+ from intracellular stores in response to ET1 stimulation. Furthermore, abolition of the sustained increase in [Ca2+]i in Ca2+-free extracellular medium argues in favour of the entry of Ca2+ during the plateau phase. Abolition of this plateau phase by Ni2+ (1 mM) in the presence of extracellular Ca2+ confirmed the existence of an ET1-induced Ca2+ entry. No evidence for the presence of voltage-operated channels was demonstrated during ET1 action since nifedipine (10(-6) M) did not reduce the Ca2+ response and depolarization with a hyper-potassium solution had no effect. Pharmacological studies using the imidazole derivatives SK&F96365 (30 microM) and LOE 908 (10 microM) partially inhibited the ET1-evoked Ca2+ response, thus providing evidence for the presence of both store-operated Ca2+ channels and non-selective cationic channels in the human ST.

The stimulation of phospholipase D (PLD) activity by endothelin-1 (ET1) was investigated in rabbit iris sphincter prelabelled with [3H]myristic acid. In the presence of 0.5% ethanol, ET1 caused a time- and dose-dependent increase in the production of [3H]phosphatidylethanol ([3H]PEt). Within 30 s the peptide increased PEt formation by 30% and after 5 min increased it by 140%. The EC50 value for ET1-stimulated PEt formation was found to be 30 nM. This value is appreciably lower than the EC50 we previously obtained for ET1-induced inositol trisphosphate production (45 nM), but considerably higher than that for arachidonic acid release (1 nM). PEt formation was significantly stimulated by prostaglandin F20, phorbol 12,13-dibutyrate (PDBu), chloroform, A23187 and A1F4-, but it was not affected by carbachol or the platelet-activating factor. PDBu-stimulated PEt formation was blocked by staurosporine and it was not potentiated by A23187. Staurosporine had no effect on ET1-stimulated PEt formation. Our data indicate that ET1 stimulation of PLD occurs independently of protein kinase C activation, phospholipase C activation and intracellular Ca2+ mobilization, and phospholipase A2 activation. In this tissue the ET1 receptor is probably coupled to the three phospholipases through several G-proteins, and this appears to be species and receptor type specific.

Vascular dysregulation has long been recognized as an important pathophysiological factor underlying the development of glaucomatous neuropathy. Endothelin-1 (ET1) has been shown to be a key player due to its potent vasoconstrictive properties that result in retinal ischemia and oxidative stress leading to retinal ganglion cell (RGC) apoptosis and optic nerve (ON) damage. In this study we investigated the protective effects of magnesium acetyltaurate (MgAT) against retinal cell apoptosis and ON damage. MgAT was administered intravitreally prior to, along with or after administration of ET1. Seven days post-injection, animals were euthanized and retinae were subjected to morphometric analysis, TUNEL and caspase-3 staining. ON sections were stained with toluidine blue and were graded for neurodegenerative effects. Oxidative stress was also estimated in isolated retinae. Pre-treatment with MgAT significantly lowered ET1-induced retinal cell apoptosis as measured by retinal morphometry and TUNEL staining. This group of animals also showed significantly lesser caspase-3 activation and significantly reduced retinal oxidative stress compared to the animals that received intravitreal injection of only ET1. Additionally, the axonal degeneration in ON was markedly reduced in MgAT pretreated animals. The animals that received MgAT co- or post-treatment with ET1 also showed improvement in all parameters; however, the effects were not as significant as observed in MgAT pretreated animals. The current study showed that the intravitreal pre-treatment with MgAT reduces caspase-3 activation and prevents retinal cell apoptosis and axon loss in ON induced by ET1. This protective effect of ET1 was associated with reduced retinal oxidative stress.

Estrogens have some anti-atherosclerotic properties and they influence nitric oxide (NO) production. The aim of this study was to determine NOx levels in post-menopausal women and the effect of estrogen/estrogen-progesteron therapy (ET/EPT) on plasma NO levels. Eighty postmenopausal women (M1) comprising 26 with surgically induced menopause (ET1), mean age 50.9+/-2.9 yr, and 54 with physiological menopause (EPT1), mean age 50.5+/-3.0 yr, were studied. Forty healthy pre-menopausal women, mean age 48.3+/-2.3 yr were the controls (C). The post-menopausal women were treated for 4 months: group ET1 with ET and group EPT1 with EPT. Serum estradiol (E2), FSH, NOx and lipid profile before and after therapy were measured. NOx levels were lower in group M1 than in group C (8.75+/-1.57 vs 10.27+/-2.62, p<0.01) and increased after hormonal therapy (10.65+/-2.38). NOx concentration showed significant positive correlation with E2 (r=0.25, p<0.05). Total cholesterol (240.9+/-43.2), LDL-cholesterol (155.2+/-33.6), triglycerides (124.8+/-54.1), and apolipoprotein B (1.52+/-0.33) were higher in group M1 than in group C (223.1+/-44.3, 133.0+/-38.2, 108.3+/-52.9, and 1.12+/-0.36, respectively), and after ET/EPT they decreased to the values observed in group C. There were no correlations between NO and lipids or apolipoproteins.

CONCLUSIONS

ET and EPT improve NOx synthesis and endothelial relaxation. Medroxyprogesterone acetate added to E2 does not significantly influence NOx levels.

The presence of functional endothelin converting enzyme (ECE) activity in basilar artery ring segments was investigated by measuring the contractile and relaxant effects of big endothelin (ET)-1. Under resting tension conditions cumulative application of big ET1-1 elicited a concentration-related contraction with the concentration-effect curve (CEC) shifted to the right against ET-1 by a factor of 31 and 29 in segments with the endothelium intact or mechanically removed, respectively. Preincubation with the ET(A) receptor antagonist, BQ123, induced an apparently parallel rightwards shift without affecting the maximum contraction. This shift was more pronounced for ET-1 than for big ET-1. With the putative ECE inhibitor phosphoramidon (10(-3) M) in the bath a small rightwards shift of the CEC for big ET-1 was observed in control segments and a more marked one in de-endothelialized segments. In segments precontracted with prostaglandin (PG) F(2alpha) big ET-1 induced a significant although transient relaxation whereas ET-1 did not. However, in the presence of BQ123 both ET-1 and big ET-1 elicited concentration-related relaxation with a significantly higher maximum effect obtained with big ET-1. The potency was 13 fold higher for ET-1, which is markedly less than that found for contraction. The results, therefore, suggest 1) the presence of functional ECE-activity in the rat basilar artery wall, and 2) differences in the functional ECE activity located in the endothelium and media.

We studied whether plasma endothelin (ET)-1 concentrations are altered in patients with septic shock who are undergoing hemodialysis and whether polymyxin B-immobilized fiber (PMX-F) treatment affects on these concentrations. Fifteen hemodialysis patients with septic shock treated with PMX-F (group A), 10 such patients who received conventional treatments (group B), 20 hemodialysis patients without septic shock (group C) and 20 healthy controls (group D) were included in this study. Plasma ET1 levels were measured by radioimmunoassay and endotoxin levels were determined by endospecy test. The survival rate in group A (67%) was higher than that in group B (30%). Blood endotoxin levels decreased significantly from 36.4+/-8.2 pg/mL to 10.6+/-3.8 pg/mL (p < 0.01) after PMX-F treatment in group A. The pretreatment plasma ET-1 levels in patients in group A (58.6+/-9.8 pg/mL) and group B (56.8+/-7.8 pg/mL) were significantly higher than those in group C (p < 0.01) and group D (p < 0.001). Plasma ET-1 levels in group C (11.2+/-3.2 pg/mL) were higher than those in group D (2.6+/-0.6 pg/mL) (p < 0.01). Plasma ET-1 levels following hemodialysis (10.9+/-3.0 pg/mL) were not altered significantly compared with those before hemodialysis. Plasma ET-1 levels decreased significantly in group A after PMX-F treatment (11.4+/-3.6 pg/mL) (p < 0.01); the levels in group B were not altered after conventional treatment. Our data suggest that ET-1 may be associated with septic shock in patients undergoing hemodialysis and that PMX-F is effective in reducing plasma ET-1 levels in these patients.

Cyclodextrin glucanotransferase from Bacillus stearothermophilus ET1 (CGTase ET1) is a potential antistaling enzyme with cyclodextrin (CD)-forming activity. To reduce cyclization activity of CGTase ET1, phenylalanine residues at 191 and 255 were replaced with a glycine (F191G-CGTase ET1) and an isoleucine (F255I-CGTase ET1), respectively. Temperature optima of both mutant enzymes were lower than that of the wild-type. Cyclization activities of both mutants decreased dramatically, but F255I-CGTase ET1 showed a 2-fold higher hydrolytic activity than the wild-type enzyme. CD content of bread loaf treated with F191G-CGTase ET1 was 28.6% of that treated with wild-type, whereas no CD was detected in the loaf treated with F255I-CGTase ET1. Loaves treated with CGTase ET1 or either of the two mutants contained more of the larger maltooligosaccharides such as maltopentaose and maltohexaose than the control and the commercial antistaling enzyme-treated loaves. Retrogradation rates decreased significantly in the loaves treated with either mutant, which indicates the applicability of CGTase ET1 in the bread industry by modulating the cyclizing and hydrolyzing activities of the enzyme.

Endothelin, encoded by ET1, is a vasoactive substance primarily synthesized in vascular endothelial cells (VECs). Elevation of endothelin levels, due to transcriptional hyperactivation, has been observed in a host of cardiovascular diseases. We have previously shown that serum response factor (SRF) is a regulator of ET1 transcription in VECs. Here we report that angiotensin II (Ang II) induced ET1 transcription paralleled activation of glycogen synthase kinase 3 (GSK3) in cultured VECs. GSK3 knockdown or pharmaceutical inhibition attenuated Ang II induced endothelin expression. Of interest, the effect of GSK3 on endothelin transcription relied on the conserved SRF motif within the ET1 promoter. Further analysis revealed that GSK3 interacted with and phosphorylated SRF at serine 224. Phosphorylation of SRF by GSK3 did not influence its recruitment to the ET1 promoter. Instead, GSK3-mediated SRF phosphorylation potentiated its interaction with MRTF-A, a key co-factor for SRF, which helped recruit the chromatin remodeling protein BRG1 to the ET1 promoter resulting in augmented histone H3 acetylation/H3K4 trimethylation. Consistently, over-expression of a constitutively active GSK enhanced Ang II-induced ET1 transcription and knockdown of either MRTF-A or BRG1 abrogated the enhancement of ET1 transcription. In conclusion, our data highlight a previously unrecognized mechanism that contributes to the transcriptional regulation of endothelin. Targeting this GSK3-SRF axis may yield novel approaches in the intervention of cardiovascular diseases.

Keywords: phosphorylation; post-translational modification; serum response factor; transcriptional regulation; vascular endothelial cell.

Aqueous (ET1) and alcoholic (ET2) extracts from Mactra veneriformis were studied for their antioxidant potentials using various in vitro assays. The ET2 was fractioned into four parts according to the polarity of the extractive medium, viz. ET2-p (petroleum ether), ET2-e (ethyl acetate), ET2-n (n-butanol) and ET2-w (water). Among the four fractions, ET2-w showed the strongest reducing power, and highest 2,2 diphenyl-1-picrylhydrazyl (DPPH) scavenging and metal chelating activities, while the ET2-p possessed higher hydroxyl radical scavenging activities. It was found that ET2-w had large amounts of free amino acids and oligosaccharides, while ET2-p contained a large amount of unsaturated fatty acids. We conclude that amino acids, oligosaccharides and unsaturated fatty acids all contribute to the antioxidant activity of M. veneriformis. In order to elucidate the above, antioxidative components involved in the antioxidant activities were also detected. It was found that amino acids and carbohydrates were the major components of ET2-w. The amino acids were Tau (0.26%), Tyr (0.38%), Met (0.31%), Cys (0.38%), Arg (1.79%) and so on, while carbohydrates were oligosaccharides that were composed of disaccharides and trisaccharides. The unsaturated fatty acids were the major components of ET2-p, which contained a high amount of docosahexaenoic acid (9.03%) and eicosapentaenoic acid (18.49%).

BACKGROUND

Alfa-interferons (IFNα2a, IFNα2b, 40KDa-PEGIFNα2a and 12KDa-PEGIFNα2b) are effective treatments for chronic hepatitis C infection. However, their usage has been associated with a variety of adverse events, including interstitial pneumonitis and pulmonary arterial hypertension. Although rare, these adverse events can be severe and potentially life-threatening, emphasizing the need for simple biomarkers of IFN-induced lung toxicity.

METHODS

Human lung microvascular endothelial cells (HLMVEC), human pulmonary artery smooth muscle (HPASM) cells and A549 cells were grown under standard conditions and plated into 96- or 6-well plates. Cells were stimulated with various concentrations of different IFNs in hydrocortisone-free medium. After 24 and 48 hours, IP10 and ET-1 were measured by ELISA in conditioned medium. In a second set of experiments, cells were pre-treated with tumour necrosis factor-α (TNF-α) (10 ng/mL).

RESULTS

IFNα2a, IFNα2b, 40KDa-PEGIFNα2a and 12KDa-PEGIFNα2b, but not IFNλ, induced IP10 (CXCL10) release and increased IP10 gene induction in HLMVEC. In addition, all four IFNα preparations induced IP10 release from HPASM cells and A549 cells pre-treated with TNFα. In each of these cell types, 40KDa-PEGIFNα2a was significantly less active than the native forms of IFNα2a, IFNα2b or 12KDa-PEGIFNα2b. Similarly, IFNα2a, IFNα2b and 12KDa-PEGIFNα2b, but not 40KDa-PEGIFNα2a, induced endothelin (ET)-1 release from HPASM cells.

CONCLUSIONS

Consistent with other interstitial pulmonary diseases, both IP10 and ET1 may serve as markers to monitor IFN-induced lung toxicity in patients. In addition, both markers may also serve to help characterize the risk associated with IFNα preparations to induce lung toxicity.

This study was to investigate the relationship between circadian blood pressure (BP) variation and circadian variation of neurohumoral factors during the acute phase of stroke. We studied 17 patients with cerebral infarction in 16 and cerebral hemorrhage in one. We performed 24-hour ambulatory BP monitoring and examined plasma renin activity (PRA), catecholamine, atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), endothelin 1 (ET1) and prothrombin fragment 1+2 (PT F1+2) and urinary catecholamine. Our result showed that the circadian variation of BP, neurohumoral and coagulation factors were diminished. There were significant relationships between BP levels and plasma BNP levels, nocturnal urinary adrenalines and ET1s. There were also significant relationships between night/day ratio of BP and plasma ET1 level. In conclusion the abnormal patterns of circadian BP rhythm were frequently observed during the acute phase of stroke. The cause of this abnormality may result from the diminished circadian rhythms of neurohumoral factors.

Background: Accurate and sensitive imaging biomarkers are required to study the progression of white matter (WM) inflammation in neurodegenerative diseases. Radioligands targeting the translocator protein (TSPO) are considered sensitive indicators of neuroinflammation, but it is not clear how well the expression of TSPO coincides with major histocompatibility complex class II (MHCII) molecules in WM. This study aimed to test the ability of TSPO to detect activated WM microglia that are immunohistochemically positive for MHCII in rat models of prodromal Alzheimer's disease and acute subcortical stroke.

Methods: Fischer 344 wild-type (n = 12) and TgAPP21 (n = 11) rats were imaged with [¹⁸F]FEPPA PET and MRI to investigate TSPO tracer uptake in the corpus callosum, a WM region known to have high levels of MHCII activated microglia in TgAPP21 rats. Wild-type rats subsequently received an endothelin-1 (ET1) subcortical stroke and were imaged at days 7 and 28 post-stroke before immunohistochemistry of TSPO, GFAP, iNOS, and the MHCII rat antigen, OX6.

Results: [¹⁸F]FEPPA PET was not significantly affected by genotype in WM and only detected increases near the ET1 infarct (P = 0.033, infarct/cerebellum uptake ratio: baseline = 0.94 ± 0.16; day 7 = 2.10 ± 0.78; day 28 = 1.77 ± 0.35). Immunohistochemistry confirmed that only the infarct (TSPO cells/mm²: day 7 = 555 ± 181; day 28 = 307 ± 153) and WM that is proximal to the infarct had TSPO expression (TSPO cells/mm²: day 7 = 113 ± 93; day 28 = 5 ± 7). TSPO and iNOS were not able to detect the chronic WM microglial activation that was detected with MHCII in the contralateral corpus callosum (day 28 OX6% area: saline = 0.62 ± 0.38; stroke = 4.30 ± 2.83; P = .029).

Conclusion: TSPO was only expressed in the stroke-induced insult and proximal tissue and therefore was unable to detect remote and non-insult-related chronically activated microglia overexpressing MHCII in WM. This suggests that research in neuroinflammation, particularly in the WM, would benefit from MHCII-sensitive radiotracers.

Keywords: Alzheimer’s disease; Ischemic stroke; Major histocompatibility complex class II; TSPO; White matter inflammation.

Rhodamine-bromonaphthaleneimide (RB-NI) and rhodamine-bromonaphthalenediimide (RB-NDI) dyads were prepared for switching of the triplet excited states. Bromo-NI or bromo-NDI parts in the dyads are the spin converters, i.e., the triplet state producing modules, whereas the RB unit is the acid-activatable electron donor/energy acceptor. NI and NDI absorb at 359 and 541 nm, and the T1 state energy levels are 2.25 and 1.64 eV, respectively. RB undertakes the reversible spirolactam (RB-c) ↔ opened amide (RB-o) transformation. RB-c shows no visible light absorption, and the triplet-state energy level is ET1 = 3.36 eV. Conversely RB-o shows strong absorption at 557 nm, and ET1 is 1.73 eV. Thus, the acid-activated fluorescence-resonance-energy-transfer (FRET) competes with the ISC of NI or NDI. No triplet state was observed for the dyads with nanosecond time-resolved transient absorption spectroscopy. Upon addition of acid, strong fluorescence and long-living triplet excited states were observed. Thus, the producing of triplet state is acid-activatable. The triplet state of RB-NI is localized on RB-o part, whereas in RB-NDI the triplet state is delocalized on both the NDI and RB-o units. The ISC of spin converter was not outcompeted by RET. These studies are useful for switching of triplet excited state.

The generation of a complex microbial consortium is a promising approach for efficient biomass decomposition. An anaerobic thermophilic alkaliphilic microbial consortium with efficient degradation ability was screened from bovine manure compost using non-pretreated milling corn stover (CS) and rice straw (RS). A stable microbial consortium ISHI-3 with high degradation ability for CS and RS was isolated by the roll tube technique. ISHI-3 comprised Herbivorax saccincola and bacteria belonging to the classes Pelotomaculum, Tepidanaerobacter, and Tepidimicrobium, as determined by DGGE of the PCR-generated 16S rRNA genes. Furthermore, metagenomics analysis using a 16S rRNA library was carried out to determine the bacterial distribution during degradation of CS and RS. H. saccincola and bacteria belonging to Pelotomaculum were relatively abundant in the beginning to middle periods of culture with CS and RS whereas bacteria belonging to Tepidanaerobacter and Tepidimicrobium gradually increased in the population during the later stages. To understand the role of non-cellulolytic bacteria in the consortium, novel strains ET1 and GL4, which were most closely related to Tepidimicrobium ferriphilum and Tepidanaerobacter acetatoxydans, were isolated from ISHI-3. Based on their carbon source usage, morphology, and phylogenetic analysis, we propose that strains ET1 and GL4 should be classified as a novel genus or species. Bacteria ET1 and GL4 can utilize different organic compounds as carbon and energy sources such as organic acids, alcohols, sugars, and amino acids, showing a preference for organic acids and alcohols rather than sugars such as glucose and cellobiose. These results indicated that ET1 and GL4 help to accelerate efficient lignocellulose degradation of H. saccincola.

OBJECTIVE

Nasopharyngeal carcinoma (NPC) is one of the most commonly diagnosed cancers worldwide with significantly high prevalence in Southern China. Chemoprevention of cancer with alkylating agent compounds could potentially reverse, suppress, or prevent cancer progression. Cisplatin (CIS) is an antineoplastic or cytotoxic platinum-based drug used for chemotherapy of different types of human cancers such as NPC. Nevertheless, the effects of CIS on the migration and invasion of human NPC cells and the underlying molecular mechanisms have not yet been fully scrutinized.

METHODS

In this work, we tested the effect of CIS on the proliferation, migration and invasion of NPC cells. The results exhibited that this drug exerts remarkable inhibitory effects on the proliferation, migration and invasion of NPC cells in a dose-dependent manner. Western blotting and real time RT-PCR were used for expression analyses.

RESULTS

We found that CIS treatment led to a dose-dependent inhibition of Endothelin-1 (ET1) expression, at protein as well as mRNA levels in NPC cells. CIS was also found to activate the expression of BTG1 in NPC cells. Moreover, mechanistic analyses revealed that CIS increased the expression of B cell translocation gene 1 (BTG1) to suppress the expression of ET1. Furthermore, we show that ET1 could not be induced in CIS-resistant cells with suppressed BTG1 expression, and subsequently demote the proliferation, migration and invasion of NPC cells.

CONCLUSIONS

These findings provided compelling evidence of the role of CIS in suppressing NPC metastasis and its underlying molecular mechanisms.

Increased bone formation resulting from mechanical loading is well documented; however, the interactions of the mechanotransduction pathways are less well understood. Endothelin-1, a ubiquitous autocrine/paracrine signaling molecule promotes osteogenesis in metastatic disease. In the present study, it was hypothesized that exposure to big endothelin-1 (big ET1) and/or mechanical loading would promote osteogenesis in ex vivo trabecular bone cores. In a 2×2 factorial trial of daily mechanical loading (-2000με, 120cycles daily, "jump" waveform) and big ET1 (25ng/mL), 48 bovine sternal trabecular bone cores were maintained in bioreactor chambers for 23days. The bone cores' response to the treatment stimuli was assessed with percent change in core apparent elastic modulus (ΔEapp), static and dynamic histomorphometry, and prostaglandin E2 (PGE2) secretion. Two-way ANOVA with a post hoc Fisher's LSD test found no significant treatment effects on ΔEapp (p=0.25 and 0.51 for load and big ET1, respectively). The ΔEapp in the "no load + big ET1" (CE, 13±12.2%, p=0.56), "load + no big ET1" (LC, 17±3.9%, p=0.14) and "load + big ET1" (LE, 19±4.2%, p=0.13) treatment groups were not statistically different than the control group (CC, 3.3%±8.6%). Mineralizing surface (MS/BS), mineral apposition (MAR) and bone formation rates (BFR/BS) were significantly greater in LE than CC (p=0.037, 0.0040 and 0.019, respectively). While the histological bone formation markers in LC trended to be greater than CC (p=0.055, 0.11 and 0.074, respectively) there was no difference between CE and CC (p=0.61, 0.50 and 0.72, respectively). Cores in LE and LC had more than 50% greater MS/BS (p=0.037, p=0.055 respectively) and MAR (p=0.0040, p=0.11 respectively) than CC. The BFR/BS was more than two times greater in LE (p=0.019) and LC (p=0.074) than CC. The PGE2 levels were elevated at 8days post-osteotomy in all groups and the treatment groups remained elevated compared to the CC group on days 15, 19 and 23. The data suggest that combined exposure to big ET1 and mechanical loading results in increased osteogenesis as measured in biomechanical, histomorphometric and biochemical responses.

BACKGROUND

Edema is a common condition in most venous and lymphatic diseases. The ACI edema testers (ET) have been developed to objectively evaluate the presence of edema. Two types of testers have been developed. ET1 is a soft plastic plate (5 x 2 cm) characterised by two parallel protrusions while the ET2 is characterised by two lines of 7 holes.

METHODS

The ETs are applied onto the internal perimalleolar region with the protrusions/holes in contact with the skin. A blood pressure cuff is applied over the area (pressure maintained at 50 mmHg for a period between 1-3 minutes). When the cuff is removed, with the ET1 skin marks are usually just visible in normal limbs (they disappear in a few minutes). We studied 22 normal limbs, 19 with varicose veins, 22 with chronic venous insufficiency, 5 with primary lymphedema and 8 with severe chronic lymphedema.

RESULTS

In limbs with severe edema the whole length of the protrusions is visible; with moderate edema only a part of the protrusions is visible. With the ET2 skin marks are just visible in normal limbs (only the larger holes). Marks disappear in a few minutes in normal limbs while in limbs with edema the number of visible holes is increased (in severe edema all holes are visible). There were significant differences between normals and patients (considering skin mark length, number of visible holes and disappearance times).

CONCLUSIONS

The two testers separated patients with different severity of edema due to chronic venous or lymphatic problems. In severe lymphatic problems all parameters were different (p < 0.02) from those observed in venous disease. A reproducibility study indicated that the ET tests have minimal variations in mark visibility or length or hole numbers (for the ET2).

The work presented relates to developing a stochastic version of the ICRP 66 respiratory tract deposition model and applying the stochastic model to characterize the variability/uncertainty associated with inhaled PuO2 for a hypothetical population of nuclear workers engaged in light work-related exercise. The parameter uncertainty/variability distributions used are essentially the same as the FORTRAN-based stochastic deposition model of Bolch et al. known as LUDUC (LUng Dose Uncertainty Code). Based on Crystal Ball software, this stochastic deposition model includes particle polydispersity, which Bolch et al. did not discuss. This paper first compares model-simulated regional deposition probability distributions to deterministic results based on LUDEP (LUng Dose Evaluation Program) software, which implements the ICRP 66 deterministic deposition model. For these comparisons, a particle density of 3 g cm(-3) (for hypothetical radioactive particles) was used. The range of possible depositions generated by LUDUC and the Crystal Ball program results revealed LUDEP's limitations. Even though LUDEP tends to use parameters that represent average parameter values for adult males, it overestimates deposition in the lower regions of the lung for most of the population. The Crystal Ball program was then used to generate radioactivity intake distributions for single and multiple PuO2 particle intakes by a hypothetical population of nuclear workers for the stochastic intake (STI) paradigm. These distributions of radioactivity intake are evaluated for the five primary regions of the respiratory tract as defined in the ICRP Publication 66. The results reveal that when a particle has been deposited, the radioactivity is likely to be low if it is in the lower regions (< 10 Bq for the bb and AI regions), but it may be quite large in the upper regions (as much as 600 Bq for the ET1, and ET2 regions), and the distributions for radioactivity become less and less skewed to the right, as particles penetrate deeper within the respiratory tract.

We have studied epsilon PKC-mediated phosphorylation events in neonatal cardiac myocytes using back phosphorylation. 3 nM 4-beta 12-myristate-13-acetate (PMA)-intact cell treatment preferentially activates epsilon PKC in these cells (Circ. Res. 76 (1995) 654) and caused decreased 32P incorporation (back phosphorylation) into an approximately 18-kDa protein. This response required physiological levels of free Mg(2+) and short (3-5 min) incubation periods in back phosphorylation assays. Introduction of a selective epsilon PKC translocation inhibitor (epsilon V1) into these cells attenuated the 3 nM PMA-induced back phosphorylation response while translocation inhibitors to the classical PKC or deltaPKC isozymes were without effect. Pretreatment of our cells with endothelin-1 (ET1) had similar effects to 3 nM PMA albeit the magnitude of the ET1 back phosphorylation response was about one-half that of 3 nM PMA. Our results suggest that epsilon PKC phosphorylates an approximately 18-kDa protein found in the particulate cell fraction of neonatal cardiac myocytes. Epsilon PKC modulates diverse cardiac responses including contraction, ion channel functions, hypertrophy, and ischemic preconditioning. Characterization of epsilon PKC-selective phosphotransferase events may reveal novel regulatory mechanisms for this enzyme in neonatal cardiac myocytes.

Objective. The purpose of this prospective study was to record Endothelin 1 (ET1) concentrations in the second trimester amniotic fluid and in women who develop premature rupture of membranes (PROM), preterm premature rupture of the membranes (PPROM) and in women with uneventful pregnancies. Method. Amniotic fluid was retrieved by amniocentesis from 125 women in the second trimester of pregnancy. The levels of Endothelin were measured by a sensitive and specific radioimmunoassay. Results. From the 125 women included in the study 20 developed PROM and preterm PROM (13 PPROM and 7 PROM). The ET1 concentration was significantly higher (P<0.001) in PROM and PPROM than in normal pregnancy (96.4 vs. 43 pg/ml). The sub-analysis of the two rupture of membranes groups found that the concentration of ET1 was higher in the PPROM than in PROM (118 vs. 72 pg/ml). Conclusion. The amniotic fluid concentration of ET1 is elevated by the second trimester in women who later develop preterm PROM or term PROM.