True reference nanosensor realized with silicon nanowires.

Conventional gate oxide layers (e.g., SiO(2), Al(2)O(3), or HfO(2)) in silicon field-effect transistors (FETs) provide highly active surfaces, which can be exploited for electronic pH sensing. Recently, great progress has been achieved in pH sensing using compact integrateable nanowire FETs. However, it has turned out to be much harder to realize a true reference electrode, which--while sensing the electrostatic potential--does not respond to the proton concentration. In this work, we demonstrate a highly effective reference sensor, a so-called reference FET, whose proton sensitivity is suppressed by as much as 2 orders of magnitude. To do so, the Al(2)O(3) surface of a nanowire FET was passivated with a self-assembled monolayer of silanes with a long alkyl chain. We have found that a full passivation can be achieved only after an extended period of self-assembling lasting several days at 80 °C. We use this slow process to measure the number of active proton binding sites as a function of time by a quantitative comparison of the measured nonlinear pH-sensitivities to a theoretical model (site-binding model). Furthermore, we have found that a partially passivated surface can sense small changes in the number of active binding sites reaching a detection limit of δN(s) ≈ 170 µm(-2) Hz(-1/2) at 10 Hz and pH 3.

Short-acting P2Y12 blockade to reduce platelet dysfunction and coagulopathy during experimental extracorporeal circulation and hypothermia.

BACKGROUND: Extracorporeal circulation (ECC) and hypothermia are routinely used in cardiac surgery to maintain stable circulatory parameters and to increase the ischaemic tolerance of the patient. However, ECC and hypothermia cause platelet activation and dysfunction possibly followed by a devastating coagulopathy. Stimulation of the adenosinediphosphate (ADP) receptor P(2)Y(12) plays a pivotal role in platelet activation. This experimental study tested P(2)Y(12) receptor blockade as an approach to protect platelets during ECC. METHODS: Human blood was treated with the short-acting P(2)Y(12) blocker cangrelor (1 µM, t(1/2)<5 min) or the P(2)Y(12) inhibitor 2-MeSAMP (100 µM) and circulated in an ex vivo ECC model at normothermia (37°C) and hypothermia (28°C). Before and after circulation, markers of platelet activation and of coagulation (thrombin-antithrombin complex generation) were analysed. During hypothermic ECC in pigs, the effect of reversible P(2)Y(12) blockade on platelet function was evaluated by cangrelor infusion (0.075 µg kg(-1) min(-1)). RESULTS: During ex vivo hypothermic ECC, P(2)Y(12) blockade inhibited platelet granule release (P<0.01), platelet-granulocyte binding (P<0.05), and platelet loss (P<0.001), whereas no effects on platelet-ECC binding, platelet CD42bα expression, glycoprotein IIb/IIIa activation, or thrombin-antithrombin complex generation were observed. During hypothermic ECC in pigs, cangrelor inhibited platelet-fibrinogen binding (P<0.05) and ADP-induced platelet aggregation (P<0.001). Platelet function was rapidly restored after termination of cangrelor infusion. CONCLUSIONS: P(2)Y(12) blockade by cangrelor prevents platelet activation during ECC and hypothermia. Owing to its short half-life, platelet inhibition can be well controlled, thus potentially reducing bleeding complications. This novel pharmacological strategy has the potential to reduce complications associated with ECC and hypothermia.

Killing of fibroblasts by dexamethasone or dibutyryl adenosine 3',5'-monophosphate is not a valid test for cystic fibrosis.

Assays based on the counting of total cells and of colony-forming cells were used to demonstrate that neither dexamethasone nor dibutyryl adenosine 3',5'-monophosphate (cyclic AMP) kills human fibroblasts under a variety of conditions. These results contradict those of previous studies showing that dexamethasone and dibutyryl cyclic AMP kill a higher percentage of fibroblasts from normal humans than from individuals with cystic fibrosis.

Pregnancy monitoring in dogs and cats using 3D and 4D ultrasonography.

Three-dimensional (3D)/four-dimensional (4D) volume ultrasound is an established method in human medicine that offers various options for analysing and presenting ultrasound volume data. However, the successful application of the different 3D/4D imaging modalities in pregnant dogs and cats has not yet been reported in the literature. The main reasons for this are: (1) the high costs of 3D/4D ultrasound systems, (2) operation difficulties due to high breathing frequency in non-sedated animals and (3) the missing specific knowledge in veterinary medicine concerning how to perform high-quality volume scans. Automatically acquired ultrasound volume data sets were generated with two different ultrasound systems: the portable Voluson i and the stationary Voluson Expert 730. Different 3D/4D imaging modalities were tested in regard of their practicability in pregnancy monitoring in dogs and cats. Nine different volume imaging modalities were applied using the saved files. For the presentation of the static 3D volume data sets, we used the multiplanar, niche, surface, transparency, glass body, inversion, volume calculation and tomographic ultrasound imaging modes. For the dynamic 4D data, the surface and glass body modes were applied. By changing the human standard settings to the requirements of small animal anatomy, it was found that 3D/4D ultrasound has great potential for the characterization of pregnancy in queens and bitches. The 3D/4D technology offered advanced information about pregnancy status and birth prediction and improved the diagnostic confidence. By using standardized examination protocols, 3D/4D ultrasound will allow a reduction in examination time by generating even more relevant information. These benefits, combined with possible future cost reduction of commercial ultrasound systems, might lead to frequent utilization in routine pregnancy diagnostic and birth management in small animal practice.

Early embryo development in the elephant assessed by serial ultrasound examinations.

The elephant has an extraordinary long pregnancy, lasting 21 months. However, knowledge on embryo development is limited. To date, only single morphological observations of elephant embryo development associated with placentation are available, all lacking correlation to gestational age. The present study describes morphological characteristics of early embryo development in the elephant with exact biometric staging. Six pregnancies in five Asian and one African elephants with known conception dates were followed by 2D and 3D ultrasound, covering the embryonic period from ovulation to day 116 post-ovulation. The embryonic vesicle was earliest observed was on day 50 p.o. The proper embryo was not detected until day 62 p.o. Embryonic heartbeat was first observed on day 71 p.o. The allantois, which became visible as a single sacculation on day 71 p.o. was subdivided in four compartments on day 76 p.o. By day 95 p.o., head, rump, front and hind legs were clearly distinguished. Between days 95 and 103 p.o. the choriovitelline placenta was replaced by the chorioallantoic placenta. A physiological midgut herniation was transiently present between days 95 and 116 p.o. On the basis of the late appearance of the embryonic vesicle, delayed implantation in the elephant is discussed. The study provides a coherent description of elephant embryonic development, formation of the extraembryonic organs and their role in placenta formation, all of which are of interest for both comparative evolutionary studies and the improvement of assisted reproduction techniques.

Elevated amounts of a 7S nuclear RNA with sequence homology to a tumor virus promoter in transformed and tumorigenic cells.

A small nuclear RNA (7S-K) from cultured mammalian cells has been shown, in previous studies, to have the characteristics expected of a gene regulatory molecule, i.e., a tissue- and species-specific stimulatory activity for initiation of transcription of protein-coding genes. Moreover, in simian virus 40 (SV40)-transformed mouse cells, this RNA was shown, by S1-analysis, to bear an extensive homology to the SV40 promoter, suggesting that it acts by base-pairing to DNA in this region to facilitate the formation of the transcription-initiation complex. In order to investigate whether or not this homology is restricted to SV40-transformed cells and in any way related to transformation, a series of normal and transformed cell lines were examined for the degree of homology between their 7S-K RNAs and the SV40 promoter. Results show that the amount of 7S-K RNA hybridizable to the promoter varies as a direct function of the established degree of tumorigenic activity of the cells and is not dependent on the presence of SV 40 sequences. Taken together with the known overexpression of some cellular oncogenes in tumor tissues, these results suggest that this particular RNA may be involved in stimulating transcription of, at least, some of these genes.

Multiple facets of HIV-associated renal disease.

HIV infection has a broad spectrum of renal manifestations. This study examined the clinical and histological manifestations of HIV-associated renal disease, and predictors of renal outcomes. Sixty-one (64% male, mean age 45 years) HIV patients were retrospectively evaluated. Clinical presentation and renal histopathology were assessed, as well as CD4 T-cell count and viral load. The predictive value of histological lesion, baseline CD4 cell count and viral load for end-stage renal disease (ESRD) or death were determined using the Cox regression model. The outcomes of chronic kidney disease (CKD) and ESRD or death were evaluated by baseline CD4 cell count. The percent distribution at initial clinical presentation was non-nephrotic proteinuria (54%), acute kidney injury (28%), nephrotic syndrome (23%), and chronic kidney disease (22%). Focal segmental glomerulosclerosis (28%), mainly the collapsing form (HIVAN), acute interstitial nephritis (AIN) (26%), and immune complex-mediated glomerulonephritis (ICGN) (25%) were the predominant renal histology. Baseline CD4 cell count ≥ 200 cells/mm3 was a protective factor against CKD (hazard ratio=0.997; 95%CI=0.994-0.999; P=0.012). At last follow-up, 64% of patients with baseline CD4 ≥ 200 cells/mm3 had eGFR >60 mL·min-1·(1.73 m2)-1 compared to the other 35% of patients who presented with CD4 <200 cells/mm3 (log rank=9.043, P=0.003). In conclusion, the main histological lesion of HIV-associated renal disease was HIVAN, followed by AIN and ICGN. These findings reinforce the need to biopsy HIV patients with kidney impairment and/or proteinuria. Baseline CD4 cell count ≥ 200 cells/mm3 was associated with better renal function after 2 years of follow-up.

Ribonuclease P: a ribonucleoprotein enzyme.

The ribonucleoprotein ribonuclease P catalyzes the hydrolysis of a specific phosphodiester bond in precursor tRNA to form the mature 5' end of tRNA. Recent studies have shed light on the structures of RNase-P-RNA-P-protein and RNase-P-RNA-precursor-tRNA complexes, as well as on the positions of catalytic metal ions, emphasizing the importance of the structure to the catalytic function.

Preclinical evaluation of the thrombogenicity and endothelialization of bare metal and surface-coated neurovascular stents.

BACKGROUND AND PURPOSE: Stent-assisted coiling is routinely used for the endovascular treatment of complex or wide-neck intracranial aneurysms. However, in-stent thrombosis, thromboembolic events, and ischemic complications remain a major concern associated with stent implants. Therefore, a novel low-profile neurovascular stent with a bare metal surface was investigated with regard to thrombogenicity and endothelialization and compared with the same stent coated with albumin or heparin. MATERIALS AND METHODS: The bare metal and heparin- or albumin-coated stents were loaded in heparin-coated tubing, which was then filled with heparinized human blood (n = 5) and circulated at 150 mL/min and 37°C for 60 minutes. Before and after circulation, measurement of various inflammation and coagulation markers and scanning electron microscopy were performed. Endothelialization of the stents was investigated in an in vitro model including human umbilical vascular endothelial cells. RESULTS: Our results showed that platelet loss and platelet activation and activation of the coagulation cascade, which are induced by the bare metal stents, were significantly reduced by heparin or albumin coating. Adverse effects on erythrocytes, leukocytes, and the complement cascade were not induced by the bare metal or coated stents. Moreover, the bare metal and albumin-coated stents showed good endothelialization properties. CONCLUSIONS: Albumin and heparin coatings clearly improve the thrombogenicity of the stents and thus may represent fundamental progress in the treatment of intracranial aneurysms. Moreover, preclinical evaluation of neurovascular stents under physiologic conditions supports and accelerates the development of more biocompatible neurovascular stents.

An analysis of the origins of the cholinergic and noncholinergic septal projections to the hippocampal formation of the rat.

These experiments were directed at determining the proportion and distribution of cholinergic septal cells which project to the rat hippocampal formation. Injections of WGA-HRP were placed into different regions of the hippocampal formation and sections through the septal complex were processed for the simultaneous demonstration of the retrogradely transported marker and for choline acetyltransferase (ChAT) immunoreactivity. Preliminary analysis of adjacent normal series prepared either for the demonstration of ChAT or stained by the Nissl method demonstrated several distinct cell groups in the classically defined medial septal nucleus and vertical limb of the nucleus of the diagonal band. The groups of cells ranged from almost entirely ChAT-positive to entirely noncholinergic. On the basis of shape and size of the constituent cells, the ChAT-positive cells of the septal complex were divided into dorsal, intermediate, and ventral subdivisions. The proportion of retrogradely labeled cells that were also ChAT positive ranged from 22.8% to 77.4% in different experiments. When only the hippocampus and dentate gyrus are considered, this variation can largely be accounted for by the topographic organization of the septohippocampal projection. The medial, noncholinergic half of the medial septal nucleus projects primarily to the rostral or septal portions of the dentate gyrus and hippocampus, whereas the lateral half, in which the dorsal ChAT group is located, projects heavily to more temporal levels. Rostral portions of the hippocampus and dentate gyrus receive most of their cholinergic input from the ventral ChAT cell group which forms a major component of the vertical limb of the nucleus of the diagnoal band. While some ChAT-positive cells in the intermediate group project to the hippocampal formation, they are generally less numerous than those from the dorsal and ventral groups. However, in a control experiment in which the WGA-HRP injection was placed into the cingulate cortex overlying the rostral hippocampal formation, the intermediate ChAT group accounted for 71.2% of the double-labeled cells.

Causes of higher in-hospital mortality in women than in men after acute myocardial infarction.

Clinical, laboratory and cardiac catheterization parameters were reviewed in 355 men and 155 women hospitalized at a tertiary care referral center between February 1987 and December 1991 to analyze why women have a higher in-hospital mortality rate than do men after acute myocardial infarction. Hospital mortality was 21.4% in women and 12.1% in men (p = 0.007). In comparison with men, women were older (63.3 +/- 11.9 vs 60.5 +/- 12.6 years; p = 0.023), had more systemic hypertension (46.5 vs 34.4%; p = 0.001) and higher serum total cholesterol levels (211 +/- 51 vs 197 +/- 49 mg/dl; p = 0.0015), sought medical care later (8.9 vs 5.3 hours; p = 0.026), were referred later (47.7 vs 43.7 hours; p = 0.063) and had more shock (34.8 vs 24.2%; p = 0.013). Logistic regression analysis revealed 5 variables predictive of hospital mortality; age > 65 years, diabetes, shock, non-Q-wave infarction, and not undergoing cardiac catheterization. Gender was of borderline significance in predicting hospital mortality. Cardiac catheterization, performed in 88% of women and 87% of men, showed similar rates of 1-, 2- and 3-vessel disease, and similar characteristics of the infarction-related artery. The differences in hospital mortality between men and women are due to a combination of pre- and in-hospitalization factors in women. The excess mortality is not due to differences in disease severity as evaluated by cardiac catheterization information.

Detection of stylet-borne and circulative potato viruses in aphids by duplex reverse transcription polymerase chain reaction.

A reverse transcription polymerase chain reaction (RT-PCR) assay was designed to amplify stylet-borne potato virus Yo (PVYo) in aphids using primers located in the viral capsid gene. A 480 bp long product was detected in aphids exposed to PVYo-infected potato plants. Approximately 40% of Myzus persicae and 15% of Aphis nasturtii exposed briefly to PVYo-infected plants acquired the virus. This rate of acquisition by both species of aphids was typical of our earlier observation of the virus transmission tests. No significant difference in virus detection was observed whether the aphids were tested immediately after exposure to virus sources or stored for up to 45 days in ethanol at room temperature. The addition of a second pair to primers located in the capsid gene of circulative potato leafroll virus (PLRV) allowed simultaneous amplification of two viruses (duplex RT-PCR) in single aphids. Acquisition of PVYo by the aphids already viruliferous with PLRV was significantly reduced, compared to aphids not carrying PLRV. Duplex RT-PCR for PVYo and PLRV could be applied to analyze aphids collected from the field to ascertain the relative presence of both viruses in a single test.

Detection of potato leafroll virus in single aphids by the reverse transcription polymerase chain reaction and its potential epidemiological application.

A reverse transcription and polymerase chain reaction (RT-PCR) system was developed using two 20-mer primers located in the potato leafroll virus (PLRV) capsid gene. A 336-bp PCR product was detected from aphids (Myzus persicae) which had been fed on PLRV-infected plants. The PCR band was specific to PLRV as determined by Southern blots and detection by a PLRV-specific probe. As little as 5 min exposure of aphids to PLRV-infected leaves resulted in the presence of PLRV-specific bands in 13% of aphids. However, the percentage of PLRV-positive aphids increased with longer exposure to infected sources and reached 90% after 3-4 days of feeding. PLRV can be detected from a single viruliferous aphid or a single viruliferous aphid combined with up to 29 non-viruliferous aphids. PLRV can be detected from freshly collected aphids, those stored at -70 degrees C, or those stored in 70% ethanol at room temperature for extended periods. This method is applicable to assess the viruliferous nature of aphids caught in yellow-pan traps during the growing season or stored for over a year.

The time of origin of cells demonstrating glutamic acid decarboxylase-like immunoreactivity in the hippocampal formation of the rat.

Using the combination of [3H]thymidine autoradiography and immunohistochemistry for the detection of glutamic acid decarboxylase (GAD), we have determined that most of the cells demonstrating GAD-like immunoreactivity in the hippocampal formation of the rat become postmitotic on fetal day 14, though some cease dividing on fetal days 13 and 15-18. GAD-positive cells in all fields of the hippocampal formation appear to differentiate during roughly the same period though there is evidence for both septotemporal and regional variations in the pattern of development.

Singlet oxygen inactivates fibrinogen, factor V, factor VIII, factor X, and platelet aggregation of human blood.

Activated polymorphonuclear leukocytes participate in hemostasis. These phagocytes generate up to 5 mmol/l of oxidants of the HOCl- and chloramine-type. The present study shows, for the first time, that physiological concentrations of NaOCl or chloramines act as anticoagulants in human plasma. Prothrombin time, activated partial thromboplastin time, and thrombin time at chloramine concentrations greater than 1 mmol/l are prolonged proportional to the oxidant concentration. Plasmatic coagulation factors sensible to oxidation are fibrinogen, factor V, factor VIII, and factor X with a 50% effective dose of 2-3 mmol/l NaOCl or taurine-chloramine. Chloramines or chloramine-like agents (e.g., chloramine T(R) or vancomycin) also inactivate platelet aggregation (in whole blood or platelet-rich plasma) at an 50% effective dose of about 1.0 mmol. This irreversible oxidation of the hemostasis components is inhibited by addition of methionine, cysteine, ascorbic acid, or azide in 10-fold molar excess prior to oxidation. The oxy-radical inhibitors mannitol, superoxide dismutase, or catalase do not antagonize the action of NaOCl or chloramines. Therefore, the oxidant here involved has reaction characteristics of singlet oxygen (1O(2)), a nonradical, excited (i.e., light-emitting) oxidant. The hemostasis factors sensible to oxidation might dispose of oxidizable, for their function critical, methionine or cysteine residues. In conclusion, blood coagulation factors I, V, VIII, X and thrombocytes are sensible to nonradical oxidants of activated phagocytes. Via 1O(2) generation, polymorphonuclear leukocytes can generate a local pericellular zone of anticoagulation. The data suggest that the cell signal 1O(2) in physiological amounts is an antithrombotic agent.

A simple screening assay for certain fibrinolysis parameters (FIPA).

Hemostasis, the system of generation and degradation of thrombi, consists of coagulation and fibrinolysis. Whereas global assays to study coagulation have existed for many years, there has been no simple, rapid, and economic routine test for the plasmatic fibrinolysis parameters plasminogen activator inhibitor-1, alpha2-antiplasmin, plasminogen, and aprotinin. Here a fast functional global assay for these plasmatic fibrinolytic parameters is presented. However, the present assay is not sensitive to physiological concentrations of prourokinase or tissue-type plasminogen activator. The following assay conditions have been found to be optimal: 50 microL of citrated plasma is incubated with 50 microL of 10 IU urinary-type plasminogen activator (urokinase)/mL, 1.1 mmol/L tranexamic acid, 1% polygelin, 0.1% Triton X-100, phosphate-buffered saline, pH 7.4, for 20 min at 37 degrees C (plasmin generation phase). Then 50 microL of 3 mmol/L HD-Nva-CHA-Lys-pNA, 1.05 mol/L KCl is added, and deltaA (405 nm)/10 min (37 degrees C) is determined, by using a microtiterplate reader (plasmin detection phase). The results are calibrated against pooled normal plasma (100% plasmatic fibrinolytic parameters activity). The intra- and interassay coefficients of variation have been found to be less than 5%. The detection limit (sensitivity) of the functional fibrinolysis assay is 5 % of the normal plasmatic fibrinolysis parameters activity. The normal plasmatic fibrinolysis parameters activity is 100%, sigma = 25%. The plasmatic fibrinolysis parameters activity correlates negatively (r = -0.684) with the plasminogen activator inhibitor-1 activity of patient samples. The plasmatic fibrinolysis parameters assay is a simple, rapid, and economic functional test for several clinical relevant fibrinolysis parameters.

Viriplanin A, a new anthracycline antibiotic of the nogalamycin group. I. Isolation, characterization, degradation reactions and biological properties.

Viriplanin A, a new anthracycline antibiotic produced by Ampullariella regularis strain SE 47, was isolated from a raw product that demonstrated activity against Herpes simplex viruses. Based on spectroscopic data, the structure of the aglycone, viriplanol, was determined, and the antibiotic was found to contain the sugar moieties 2-deoxy-L-fucose, 4-O-mesaconoyl-L-diginose and decilonitrose. In solution viriplanin A is very unstable to light. The antibiotic belongs to the nogalamycin group and is related to arugomycin and decilorubicin.

Intraerythrocytic parasites in rodent populations of Connecticut: Babesia and Grahamella species.

A total of 612 Peromyscus leucopus, 11 Microtus pennsylvanicus, 21 Clethrionomys gapperi, and 4 Tamias striatus was collected in Connecticut and examined for Babesia and Grahamella during 1976 and 1977. Babesia antibodies were detected in sera of 9 P. leucopus collected from 4 sites. Babesia parasites were not detected in the blood smears of captured rodents. Subsequent splenectomy and subinoculation of blood from these rodents into susceptible animals failed to induce disease and no Babesia was isolated. Six of 10 P. leucopus inoculated with a Shelter Island, New York strain of B. microti remained infected for 3 1/2 months. Indirect fluorescent antibody titers of experimentally infected P. leucopus ranged from 1:8 to 1:256. Prevalence of Grahamella peromysci infection, as determined from examinations of blood smears of P. leucopus, was 13%. This infection rate is a conservative estimate because parasitemia is difficult to detect in intact animals. Twenty of 58 P. leucopus, taken at 2 sites and with negative blood smears for G. peromysci, developed parasitemia after splenectomy.

Desire for control, coping, and quality of life in heart and lung transplant candidates, recipients, and spouses: a pilot study.

To understand the interdependence of couples involved in transplantation, both partners need to be included in the same study. This pilot study explored the desire for control, coping, and quality of life in heart and lung transplant candidates, recipients, and spouses, using McCubbin's Double ABCX Model as the conceptual basis. Sixty letters were mailed to randomly selected candidates and recipients at 2 transplant centers. Twenty-five couples completed a demographic sheet and the following surveys: the Desire for Control Scale, the Family Crisis Oriented Personal Evaluation Scale, and the Quality of Life Index. There were 9 pretransplant and 16 posttransplant couples. Analyses included measures of central tendency, analysis of variance, t tests, and correlations. There were statistically significant differences between pretransplant patients' quality of life and all other groups. There were moderate correlations between several patient and spouse variables. It was recommended that healthcare professionals include both patients and spouses when planning interventions.

Q fever in cattle and sheep in Southern Africa. A preliminary report.

In the course of a study initiated to elucidate the role of known abortifacient in ruminants in Southern Africa, Coxiella burnetti was identified in smears prepared from bovine and ovine placental tissues obtained from 12 farms. The organism was also isolated in embryonated eggs, and specific antibodies were demonstrated in sera of animals originating from these farms. In 7 of the 12 herds in which Q fever was diagnosed, other pathogens were identified concomitantly.