Leukocyte recruitment in the airways: an intravital microscopic study of rat tracheal microcirculation

Because of its relative inaccessibility, inflammatory cell extravasation within the airway circulation in vivo has been difficult to investigate in real time. A new method has been established using intravital microscopy in the anesthetized rat to visualize leukocytes in superficial postcapillary venules of the trachea. This technique has been validated using local superfusion of lipopolysaccharide (LPS) and N-formyl-methionyl-leucyl-phenylalanine (FMLP). Basal leukocyte rolling velocity (55.4 +/- 9.3 microm/s) and adhesion (1.4 +/- 0.3 cells/100 microm) were monitored in postcapillary venules (33.9 +/- 1.3 microm diameter). At all time points up to 90 min, these parameters were unaltered in control rats (n = 7). In contrast, vessels exposed to 1 microg/ml of LPS (n = 6) exhibited a 57% reduction in leukocyte rolling velocity and an increase in the number of adherent cells (4.7 +/- 1 cells/100 microm, P < 0.05). Superfusion with 0.1 microM of FMLP (n = 6) also resulted in a 45% reduction in rolling velocity and an increase in adherent cells (4 +/- 0.7 cells/100 microm, P < 0.05). Histological evaluation confirmed local stimulus-induced leukocyte extravasation. These results demonstrate leukocyte recruitment in the airway microvasculature and provide an important new method to study airway inflammation in real time.     

Ventilator-induced lung injury: in vivo and in vitro mechanisms

A lung-protective ventilator strategy significantly reduces mortality in patients with acute lung injury. Substantial progress has been made in understanding how mechanical stress can injure the lung, both in terms of alterations in barrier properties of the pulmonary endothelium and epithelium as well as in stimulating proinflammatory responses of macrophages and neutrophils.     

The C. elegans ric-3 gene is required for maturation of nicotinic acetylcholine receptors

Mutations in ric-3 (resistant to inhibitors of cholinesterase) suppress the neuronal degenerations caused by a gain of function mutation in the Caenorhabditis elegans DEG-3 acetylcholine receptor. RIC-3 is a novel protein with two transmembrane domains and extensive coiled-coil domains. It is expressed in both muscles and neurons, and the protein is concentrated within the cell bodies. We demonstrate that RIC-3 is required for the function of at least four nicotinic acetylcholine receptors. However, GABA and glutamate receptors expressed in the same cells are unaffected. In ric-3 mutants, the DEG-3 receptor accumulates in the cell body instead of in the cell processes. Moreover, co-expression of ric-3 in Xenopus laevis oocytes enhances the activity of the C.elegans DEG-3/DES-2 and of the rat alpha-7 acetylcholine receptors. Together, these data suggest that RIC-3 is specifically required for the maturation of acetylcholine receptors.     

DelGEF,a homologue of the Ran guanine nucleotide exchange factor RanGEF,binds to the exocyst component Sec5 and modulates secretion

In order to identify the function of deafness locus putative guanine nucleotide exchange factor (DelGEF), a protein homologous to the nucleotide exchange factor for the small GTPase Ran, a cDNA library was screened for interacting proteins using a yeast two-hybrid system. The human homologue of Sec5, a protein involved in vesicle transport and secretion, was identified as a binding partner. The interaction between DelGEF and Sec5 was found to be dependent on Mg2+ and stimulated by guanosine triphosphate (GTP) or deoxycytidine triphosphate (dCTP). Downregulation of endogenous DelGEF in HeLa cells induced increased extracellular secretion of proteoglycans indicating a possible role for DelGEF in the secretion process.     

Mutations in the extracellular domain and in the membrane-spanning domains interfere with nicotinic acetylcholine receptor maturation

The deg-3(u662) mutation is a degeneration-causing mutation in a Caenorhabditis elegans nicotinic acetylcholine receptor. In a large screen for mutations that suppress the deleterious effects of this mutation we identified 32 mutations in the deg-3 gene. Among these, 11 are missense mutations, affecting seven residues within the extracellular domain or the membrane-spanning domains. All of these mutations greatly reduce the degeneration-causing activity of deg-3(u662). All but one of these mutations cause defective localization of the DEG-3 protein, as seen in immunohistochemical analysis. Thus our screen identifies multiple residues within the nicotinic acetylcholine receptor needed for normal folding, assembly, or trafficking of this receptor. Interestingly, these mutations lead to distinct localization defects suggesting differences in their effect on DEG-3's maturation process. Specifically, mutations in the extracellular domain lead to a phenotype more severe than mutations in the membrane-spanning domains. Differences in the effects of the mutations are also predicted by homology-based modeling, showing that some mutations in the extracellular domain are likely to disrupt the native fold of the protein, while others are likely to disrupt trafficking.     

Potential microbiological hazards in the production of refined paper products for food applications

This study sought to investigate the significance of raw materials (starch-based glues, raw material papers) at different microbiologically critical stages in the manufacturing process of refined paper products. The study examined the occurrence of microorganisms in the process and in end-product samples. Microbiological surveys verified that the production and use of pasteurized starch-based glue was the most important factor threatening the process hygiene and product safety. Subsequently, the production and use of starch-based glue was changed, and a follow-up programme targeting the microbiological quality of glue was developed as part of a hygiene and safety management system. A total of 33 spore-forming bacterial and 15 enterobacterial isolates were ribotyped, and 22 and 10 different ribogroups (ribotypes), respectively, were generated. These isolates from starch-based glue, raw material paper and end products were atypical and, thus, in many cases physiological, chemotaxonomic (FAME) and molecular (partial 16S rDNA) results did not correspond. The most common spore-forming bacteria (55% of the isolates) were Paenibacillus sp. and within this genus several new species were also proposed. The most common enterobacteria (87%) were Enterobacter cloacae and Citrobacter freundii belonging to bacteria in hazard group 2, or species closely related to them. It was demonstrated that the same spore-forming bacteria (ribotypes) were present in both the glue samples and the end products (45% of isolates). All RiboPrint patterns were saved at the VTT identification library for future use.     

Structural determinants of plant lignans for the formation of enterolactone in vivo

The quantity of mammalian lignans enterolactone (ENL) and enterodiol (END) and of plant lignans secoisolariciresinol (SECO) and 7-hydroxymatairesinol (HMR) excreted in a 24-h rat urine sample was measured after a single p.o. dose of an equivalent quantity of secoisolariciresinol diglycoside (SDG), secoisolariciresinol (SECO), matairesinol (MR), 7-hydroxymatairesinol (HMR) and ENL. Plant lignans (SECO and HMR) were partially absorbed as such. The aglycone form of SECO was more efficiently converted into mammalian lignans END and ENL than the glycosylated form, SDG. Of plant lignans, MR produced the highest quantities of ENL: the quantity was over twofold compared with HMR or SDG. The majority of the animals, which had been given SECO, excreted higher quantities of END than ENL into urine, but ENL was the main lignan metabolite after SDG. The highest quantities of ENL in urine were measured after the administration of ENL as such. The (-)SECO isolated from Araucaria angustifolia was converted into (-)ENL only. The administration of (-)SDG, which was shown to produce (+)SECO, resulted in excretion of (+)ENL only and (-)HMR was converted into (-)ENL only. This confirmed that the absolute configurations at C8 and C8' are not changed during the microbial metabolism. Whether the biological effects are enantiomer-specific, remains to be resolved.     

Effect of speed and precision demands on human shoulder muscle electromyography during a repetitive task

Effects of speed and precision on electromyography (EMG) in human shoulder muscles were studied during a hand movement task where five points were marked repeatedly with a pencil. Six female subjects performed with three precision demands and at four speeds. Three of the speeds were predefined, while the last speed was performed as fast as possible. The EMG were recorded from 13 shoulder muscles or parts of muscles. Elbow velocity, acceleration and rectified EMG were calculated for each task. The mean elbow velocity and acceleration increased with speed and precision demands. There was an increase in EMG as the speed demand increased for all three precision demands (P < 0.001), and as the precision demand increased for the two highest predefined speed demands (P < 0.05). The combination of a high speed and a high precision demand resulted in the highest EMG. Different EMG levels were attained for the 13 muscles and the supraspinatus muscle always showed the highest normalized EMG. However, analysis of variance showed the same relative increase for all muscles with speed and precision demands. The EMG changes in response to precision demand can only be explained in part by the differences in movement velocity and acceleration, and other factors such as increased co-contraction must also be taken into account. Accepted: 25 May 1998     

Effects of Differently Composed Feeds and Physical Stress on Plasma Gastrin Concentration in Horses

Plasma gastrin concentrations were determined in 6 Standardbreds (4 geldings and 2 mares) after 3 different meals consisting of unlimited amounts of hay (8-9 kg per horse), a restricted amount of hay (0.6 kg/100 kg body-weight) and grain (0.2 kg/100 kg body-weight) in combination or of grain alone (0.2 kg/100 kg body-weight). In another series of experiments the possible role of gastrin as a stress hormone was investigated. Plasma gastrin and Cortisol concentrations were determined during fasting and compared with concentrations during hay feeding. In addition, gastrin and Cortisol concentrations were determined before, during and after 2 kinds of physical exercise on a treadmill. Meal stimulation significantly increased the plasma gastrin concentration, irrespective of the meal composition. An immediate and large increase in plasma gastrin concentration was found when voluminous meals were given, whereas a small meal evoked a later onset of gastrin release, suggesting that gastric distention plays an important role in inducing gastrin release during a meal. Meals consisting of grain seem to evoke a slower onset and then a more prolonged gastrin response than a hay meal, possibly due to different emptying rates of the stomach. Nervous excitation may play a minor role in the activation of gastrin release in horses. No experimental support was obtained for the idea that gastrin acts as a stress hormone in the horse. kw|Keywords|k]Cortisol; k]exercise; k]feeding; k]fasting