Identification of highly fucosylated N-linked oligosaccharides from the human parotid gland

The glycosylation of a number of constituents of human saliva is known to modify its biological roles, such as its lubricating properties and binding of microbial flora. Gillece-Castro et al. [Gillece-Castro, B. L., Prakobphol, A., Burlingame, A. L., Leffler, H. & Fisher, S. J. (1991) J. Biol. Chem. 266, 17358-17368] have proposed that the major glycan on the salivary proline-rich glycoproteins is a trifucosylated biantennary sugar with one difucosylated and one unfucosylated antenna. Furthermore, they proposed that the non-fucosylated antenna mediated adherence to a peridontal pathogen, Fusobacterium nucleatum. The detailed structures and roles of other highly fucosylated glycans that co-exist in the parotid gland are not fully known. In view of the influence of outer-arm fucosylation on carbohydrate recognition processes in general, this paper reports the use of a combination of HPLC (normal and reversed phase), matrix-assisted laser-desorption/ionisation (MALDI) mass spectrometry and exoglycosidase digestions to dissect the detailed structures of the most abundant of these polyfucosylated glycans. For measurement of reversed-phase HPLC retention times, new calibration units were used which paralleled the glucose units used for normal-phase HPLC. These differed in that the difference in retention times were compared with those derived from a ladder of 2-aminobenzamide-labelled arabinose oligomers instead of the corresponding oligomers from partially hydrolysed dextran. Over sixty neutral sugars were identified from the parotid gland and many of these were additionally found substituted with sialic acid (both alpha2-3-linked and alpha2-6-linked) and sulphate. These glycans were mainly bi- and tri-antennary sugars with up to five and seven fucose residues respectively, containing fucose alpha1-3-linked to the outer-arm GlcNAc residues and alpha1-2-linked to the galactose. All fucosylated structures contained a core (alpha1-6-linked) fucose. The detailed structure of the trifucosylated biantennary glycan was confirmed, together with the structures of another 12 fucosylated biantennary glycans. Smaller amounts of hybrid and tetraantennary structures were also found and bisected glycans were shown to be constituents of parotid glycoproteins for the first time. Acidic glycans were mainly substituted with sialic acid. Most were monosialylated as the presence of fucose on the antennae was found to suppress the addition of extra sialic acid moieties. The possible functional significance of highly fucosylated N-glycans is discussed in relation to their modification of the availability of other non-reducing terminal monosaccharides for recognition processes.     

What can toxins tell us for drug discovery?

Toxins are of interest in drug design because the toxins provide three-dimensional templates for creating small molecular mimics with interesting pharmacological properties. Toxins are also useful in drug discovery because they can be used as pharmacological tools to uncover potential therapeutic targets. With their high potency and selectivity, toxins are often more useful in functional experiments than standard pharmacological agents. We have used two groups of neurotoxins, the dendrotoxins and the muscarinic toxins (MTs), to explore the involvement of subtypes of potassium ion channels and muscarinic receptors, respectively, in processes involved in cognition and the changes in neuronal properties with aging. From our current work, quantitative autoradiographic studies with radiolabelled dendrotoxins reveal widespread distribution of binding sites throughout rat brain sections, but few differences exist between young adult and aged rats. However, displacement studies with toxin K, which preferentially binds to the Kv1.1 subtype of cloned potassium channel, show the selective loss of such sites in regions of the hippocampus and septohippocampal pathway with aging. MTs have been tested for effects on performance of rats in memory paradigms. MT2, which activates m1 receptors, improves performance of rats in a step-down inhibitory avoidance test, whereas MT3, which blocks m4 receptors, decreases performance when given into the hippocampus. This is the first clear demonstration of a role for m4 muscarinic receptors in cognition.     

Efficacy of various inorganic sorbents to reduce the toxicity of aflatoxin and T-2 toxin in broiler chickens

Experiments were conducted to determine the efficacy of three inorganic sorbents, S1, S2, and S3, to reduce the toxicity of aflatoxins (AF) and T-2 toxin in male broiler chickens from day of hatch to 21 d of age. The compounds had been reported to bind to AF and T-2 toxin in vitro. S1 and S2 were the same basic compound that had been stored for different lengths of time following activation. In Experiments 1, 2, and 3, the appropriate diets were produced to contain no mycotoxins, the specific adsorbent at 0.5% of diet, AF alone at 5 mg/kg of diet, T-2 alone at 8 mg/kg of diet, AF at 5 mg/kg of diet plus the specific sorbent at 0.5% of diet, or T-2 at 8 mg/kg of diet plus the specific sorbent at 0.5% of diet. The specific sorbents used were: 1) Experiment 1, S1; 2) Experiment 2, S1 and S2; and 3) Experiment 3, S3. In Experiments 1 and 3, S1 and S3, respectively, showed no protection against AF or T-2 toxin as measured by BW gain, when compared to AF alone group. In Experiment 2, S1 showed no protection; however S2 reduced the effects of AF on BW gain by 25% as compared to AF alone diet. The data demonstrate that under the conditions of our experiment: 1) one of the sorbents provided some protection against aflatoxicosis; 2) there was variability in protection against aflatoxicosis between two different samples of the same sorbent that had been stored for different lengths of time following activation; 3) protection by the sorbents against the effects of T-2 toxin was not observed.     

Analysis of coccal cell formation by Campylobacter jejuni using continuous culture techniques, and the importance of oxidative stress

OBJECTIVES: We tested the hypothesis that patients with incomplete systolic mitral leaflet closure (IMLC: apically displaced coaptation) also have restricted diastolic leaflet opening that is independent of mitral inflow volume and provides evidence supporting increased leaflet tethering. BACKGROUND: Competing hypotheses for functional mitral regurgitation (MR) with IMLC include global left ventricular (LV) dysfunction per se (reduced leaflet closing force) versus geometric distortion of the mitral apparatus by LV dilation (augmented leaflet tethering). These are inseparable in systole, but restricted leaflet motion has also been observed in diastole, and attributed to reduced mitral inflow. METHODS: Diastolic mitral leaflet excursion and orifice area were measured by two-dimensional echocardiography in 58 patients with global LV dysfunction, 36 with and 22 without IMLC, compared with 21 normal subjects. The biplane Simpson's method was used to calculate LV ejection volume, which equals mitral inflow volume in the absence of aortic regurgitation. RESULTS: The diastolic mitral leaflet excursion angle was markedly reduced in patients with IMLC compared with those without IMLC, whose ventricles were smaller, and normal subjects (17 +/- 10 degrees vs. 58 +/- 13 degrees vs. 67 +/- 8 degrees, p < 0.0001). Excursion angle was dissociated from mitral inflow volume (r2 = 0.04); excursion was reduced in patients with IMLC despite a normal inflow volume in the larger ventricles with MR (60 +/- 25 vs. 61 +/- 12 ml in normal subjects, p = NS), and excursion was nearly normal in patients without IMLC despite reduced inflow volume (40 +/- 10 ml, p < 0.001 vs. normal subjects). The anterior leaflet when maximally open coincided well with the line connecting its attachments to the anterior annulus and papillary muscle tip (angular difference = 3 +/- 7 degrees vs. 25 +/- 9 degrees vs. 32 +/- 10 degrees in patients with and without IMLC vs. normal subjects, p < 0.0001). In patients with IMLC, the leaflet tip orifice was smaller in an anteroposterior direction but wider than in the other groups, giving a normal total area (6.8 +/- 1.8 vs. 7.1 +/- 1.2 vs. 6.9 +/- 0.8 cm2, p = NS). CONCLUSIONS: Patients with LV dysfunction and systolic IMLC also have restricted diastolic leaflet excursion that is independent of inflow volume, coincides with the tethering line connecting the annulus and papillary muscle and reflects limitation of anterior motion relative to the posteriorly placed papillary muscles without a decrease in total orifice area. These observations are consistent with increased tethering by displaced mitral leaflet attachments in the dilated ventricles of patients with IMLC that can restrict both diastolic opening and systolic closure.     

Clinical Experiences Calcium and phosphate balance in children on home nocturnal hemodialysis (NHD)

Objective: To evaluate and describe biochemical indices of bone metabolism in 4 children on NHD. Method: The children, aged 12, 13, 14, and 16 yrs, have been treated exclusively on NHD for 6, 9, 9, and 15 mos. Subsequently, Pt 1 converted to a hybrid program of 4 nights on home nocturnal plus 1 session of in center conventional HD per week. Biochemical indices of bone metabolism have been collected prospectively. Results: All baseline pre‐dialysis calcium levels were within normal ranges and each patient was started on a dialysis calcium concentration of 3.0 mEq/L. However, over time the number of asymptomatic biochemical hypocalcaemic episodes increased. The dialysate calcium concentration was increased to 3.5 mEq/L in one and decreased to 2.0 mEq/L in another who was hypercalcemic and receiving concurrent calcitonin for bone pain related to osteoporosis. In Pt 1, the dialysate calcium was increased to 3.5 mEq/L during nocturnal and continued on hybrid therapy. Including an evaluation of dietary intake, all 4 patients had a net positive calcium balance, ranging between 9.8 to 23.5 mmol (393–942 mg). A significant reduction in the predialysis phosphate level was observed in all 4 patients, and none required dietary restrictions or the use of phosphate binders within 2 months or vitamin D within 6 months of HND. In addition, phosphate was added to provide a dialysate concentration of 2.4–6.1 mEq/L to prevent hypophosphatemia. This is reflected by significant reductions in intact PTH levels to the desired range (twice the normal range) in all 4, but the level continued to drop to the normal range and below in 2. In Pt 1, after introduction of hybrid therapy, both levels of phosphate and PTH rose, necessitating recommencement of phosphate binders and vitamin D. Likewise, the (Ca × PO₄) dropped and remained <55 in all 4 patients exclusively on NHD, but started to climb in Pt 1 during hybrid therapy. Conclusion: In our cohort of patients, NHD rapidly lowered plasma phosphate and PTH levels. With NHD, additional dialysate phosphate and possibly calcium may be necessary to prevent chronic losses and development of renal osteodystrophy, and caution is required to prevent either oversuppression of PTH and extraskeletal calcification.     

Dual-task information processing in schizotypal personality disorder: Evidence of impaired processing capacity.

Working memory theories heavily rely on the concept of processing resources and the their efficient deployment. Some recent work with schizophrenia-spectrum patients has suggested that many associated cognitive impairments may be reduced to deficits in working memory, possibly related to reductions in information-processing capacity resources. In this study, 38 patients with schizotypal personality disorder (SPD), 22 patients with other personality disorders, and 14 healthy comparison participants performed a dual-task processing assessment that was designed specifically for use in this type of study. Participants recalled lists of digits at their predetermined maximum digit span and performed box-checking tests, first alone and then in a dual-task format. Instructions included equal prioritization of both tasks. SPD patients had significantly shorter digit spans, and they also showed more deterioration on both tasks. Performance operating characteristics curves indicated that SPD patients' reduced performance was not due to abnormal resource allocation strategies leading to strategic failures. The authors discuss the implications of these processing capacity limitations for understanding both the signature of cognitive impairment within the schizophrenia spectrum and general abnormalities in working memory. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Development of a Work Sampling Methodology for Behavioral Observations: Application to Teamwork

Engineering programs must assess students' abilities to master “criteria 3 a‐k.” Skills such as teamwork, problem solving, design, and ethical understanding entail learning various processes; hence, assessing these outcomes is better accomplished by focusing on the process rather than the result. Methods for observing students' performance, such as 100 percent behavioral observation, are ideal but expensive. We extend work sampling, an economic industry‐based alternative, to observe cognitive and behavioral processes. Specifically, we describe a work sampling methodology to assess students engaged in teamwork. We then determine attributes of teamwork, establish target time proportions using 100 percent observation, and statistically compare the targets to proportions obtained from work sampling intervals to determine the effective interval. The robustness of work sampling is tested in four learning environments. Results indicate that sampling provides a statistically valid alternative for assessing teamwork. However, when observing design and ethical understanding processes, additional research is needed to make work sampling viable.     

Optimizing Digital Hardware Perceptrons for Multi-Spectral Image Classification

We propose a system for solving pixel-based multi-spectral image classification problems with high throughput pipelined hardware. We introduce a new shared weight network architecture that contains both neural network and morphological network functionality. We then describe its implementation on Reconfigurable Computers. The implementation provides speed-up for our system in two ways. (1) In the optimization of our network, using Evolutionary Algorithms, for new features and data sets of interest. (2) In the application of an optimized network to large image databases, or directly at the sensor as required. We apply our system to 4 feature identification problems of practical interest, and compare its performance to two advanced software systems designed specifically for multi-spectral image classification. We achieve comparable performance in both training and testing. We estimate speed-up of two orders of magnitude compared to a Pentium III 500 MHz software implementation.