Using a model-based framework for analysing genetic diversity during germination and heterotrophic growth of Medicago truncatula.

BACKGROUND AND AIMS: The framework provided by an emergence model was used: (1) for phenotyping germination and heterotrophic growth of Medicago truncatula in relation to two major environmental factors, temperature and water potential; and (2) to evaluate the extent of genetic differences in emergence-model parameters. METHODS: Eight cultivars and natural accessions of M. trunculata were studied. Germination was recorded from 5 to 30 degrees C and from 0 to -0.75 MPa, and seedling growth from 10 to 20 degrees C. KEY RESULTS: Thermal time to reach 50 % germination was very short (15 degrees Cd) and almost stable between genotypes, while base temperature (2-3 degrees C) and base water potential for germination (-0.7 to -1.3 MPa) varied between genotypes. Only 35 degrees Cd after germination were required to reach 30 mm hypocotyl length with significant differences among genotypes. Base temperature for elongation varied from 5.5 to 7.5 degrees C. Low temperatures induced a general shortening of the seedling, with some genotypes more responsive than others. No relationship with initial seed mass or seed reserve distribution was observed, which might have explained differences between genotypes and the effects of low temperatures. CONCLUSIONS: The study provides a set of reference values for M. trunculata users. The use of the ecophysiological model allows comparison of these values between such non-crop species and other crops. It has enabled phenotypic variability in response to environmental conditions related to the emergence process to be identified. The model will allow simulation of emergence differences between genotypes in a range of environments using these parameter values. Genomic tools available for the model species M. trunculata will make it possible to analyse the genetic and molecular determinants of these differences.

Development of a polymer stent with shape memory effect as a drug delivery system.

The article presents a new concept for vascular endoprothesis (stent). Almost all commercially available stents are made of metallic materials. A common after effect of stent implantation is restenosis. Several studies on metal stents coated with drug show, that the use of a drug delivery system may reduce restenosis. The purpose of this work is to develop a new stent for the drug delivery application. The shape memory properties of thermoplastic polyurethane allow to design a new fully polymeric self-expandable stent. The possibility to use the stent as a drug delivery system is described.

Changes in oligosaccharide content and antioxidant enzyme activities in developing bean seeds as related to acquisition of drying tolerance and seed quality.

Seeds of bean (Phaseolus vulgaris cv. Vernel) were collected throughout their development on the plant and dried at 15 degrees C and 75% relative humidity to a final moisture content of about 16% (fresh weight basis) to determine whether the onset of tolerance to this drying condition was related to changes in soluble sugars or the activities of the main antioxidant enzymes, namely superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR). Measurements of soluble sugars and enzyme activities were made after drying the seeds, and drying tolerance was evaluated by the ability of dried seeds to germinate and to produce normal seedlings. Seeds became tolerant to drying at 45 d after anthesis, a time marking physiological maturity. At physiological maturity, the moisture content of seeds was about 50-55% (fresh weight basis) and seed dry matter reached about 190 mg per seed. Seed vigour, evaluated by controlled deterioration and conductivity measurements, continued to increase after seed mass maturity, but decreased when seeds remained thereafter for more than 7 d on the plant. Acquisition of drying tolerance was coincident with an accumulation of raffinose and stachyose. Dried-tolerant seeds were also characterized by a high amount of sucrose, the most abundant sugar, and by a low content of monosaccharides. The (raffinose+stachyose)/sucrose ratio increased during seed filling, reaching a value close to 1 when all the seeds became tolerant to drying, and maintaining this proportion during the final stages of maturation. Acquisition of drying tolerance was also related to a reorientation of the enzymatic antioxidant defence system. Drying-tolerant dried seeds displayed high CAT and GR activities and low SOD and APX activities, while the opposite condition was observed in immature dried seeds. The shift in antioxidant enzymes corresponded to the beginning of the maturation-drying phase. These results suggest that oligosaccharide metabolism and enzymatic antioxidant defences may be involved in acquisition of drying tolerance during bean seed development, but are not related to seed vigour.

Endotracheal surfactant atomization: an alternative to bolus instillation?

OBJECTIVE: To investigate the effect of an intratracheal surfactant fog on oxygenation, blood pressure, distribution, and recovery rate as a pilot study to intratracheal surfactant aerosol. DESIGN: Prospective, randomized study. SETTING: University laboratory. SUBJECTS: A total of 15 New Zealand White rabbits. INTERVENTIONS: The anesthetized ventilated rabbits were surfactant-deprived by repeated lung lavages and then received 200 mg/kg of a 99mTc-labeled porcine surfactant (Curosurf) either as bolus or as intratracheal surfactant fog. MEASUREMENTS AND MAIN RESULTS: Blood gases and pressure were measured and distribution as well as recovery rate of the surfactant determined by means of the radio label (gamma camera images and lung processing with subsequent gamma counter measurements). Respiratory function normalized immediately, no difference could be found between the two groups with regard to PaO2, PaCO2, and blood pressure. Even distribution of the radiolabel was found with both methods, the bolus group showing a higher percentage of lung pieces with label concentrations of double average or more. Recovery rates were 82.5%+/-13.1% (mean +/- SD) in the bolus group and 86.5%+/-7.7% in the fog group. CONCLUSION: Endotracheal surfactant fog application is as effective as bolus instillation and may have a role in the treatment of adult respiratory distress syndrome. It has to be considered as a first step in producing an effective aerosol.

Ultrasonic surfactant nebulization with different exciting frequencies.

Intratracheal bolus instillation of natural lung surfactant is the treatment of choice in neonatal respiratory distress syndrome and an increasing part in adults' therapy. For reasons of hemodynamics, surfactant distribution and efficiency the application mode should be improved. Nebulization seems to have some advantages but its technical realization is difficult. The aim of the present study was to investigate if ultrasonic nebulization with exciting frequencies higher than 2.8 MHz can improve the efficiency of surfactant nebulization without changing the surface-active properties of the material. Exciting frequencies of 1.7, 3.3 and 4.0 MHz were used to produce a surfactant aerosol. The phospholipid content in the liquefied aerosol and particle size distinctly dropped with higher frequencies. The surface activity was not altered in the produced aerosol and neither in the surfactant remaining in the nebulizer. Although possible, ultrasonic nebulization of surfactant suspensions is ineffective because of a striking decrease in phospholipid content.

Effect of C1-inhibitor in a rat model of necrotizing enterocolitis.

Our aim was to investigate the effect of C1-inhibitor (C1-INH) in a rat model of necrotizing enterocolitis (NEC). Twenty-four anesthetized and artificially ventilated rats received 0.1 mg/kg endotoxin. Fifteen minutes later, the animals in the study group (n = 12) were treated with 200 IU/kg C1-INH whereas the control animals (n = 12) received normal saline. In all animals, FiO(2) was reduced after 90 min from 0.21 to 0.05 and ventilation continued until 180 min or death. All animals developed shock symptoms. Drop in mean arterial blood pressure was more pronounced and survival time was shorter in the control group. Whereas the C1-INH activity increased in the study group, it decreased in the control group. The extent of macroscopic intestinal lesions did not differ between the groups. In conclusion, C1-INH did not prevent shock, but mitigated and delayed its course.

Utility of deadspace and capnometry measurements in determination of surfactant efficacy in surfactant-depleted lungs.

OBJECTIVE: To investigate if bronchoalveolar lavage leads to increased alveolar and physiologic deadspace or a deadspace/ tidal volume ratio and if surfactant replacement restores the lung to its prelavage condition. DESIGN: Prospective, animal cohort study. SETTING: An animal laboratory in a university medical center. SUBJECTS: Seven adult rabbits receiving artificial ventilation. METHODS: Our single-breath CO2 analysis station contained the following equipment: pneumotachometer Ventrak 1550, a mainstream capnometer Capnogard 1265, a signal processor, and computer software. Repeated bronchoalveolar lavage was performed in seven adult rabbits to simulate acute respiratory distress syndrome. Surfactant therapy was administered after bronchoalveolar lavage induced a 20% reduction in baseline arterial PO2. The calculated parameters of alveolar and physiologic deadspace and the deadspace/tidal volume ratio were derived from the single-breath CO2 plot by Ventrak 1550 in combination with the Capnogard 1265. The arterial end-tidal Pco2 difference, the alveolar-arterial PO2 difference, and the arterial/alveolar PO2 ratio were obtained by capnography and arterial blood gas analysis. Measurements of these parameters were performed before bronchoalveolar lavage, during bronchoalveolar lavage, and after surfactant application. MEASUREMENTS AND MAIN RESULTS: The alveolar and physiologic deadspace and the deadspace/tidal volume ratio were significantly higher in lavaged animals. After application of natural surfactant, these parameters were significantly reduced but the baseline values could not be reached. Bronchoalveolar lavage led to a significant fall in the arterial/alveolar PO2 ratio, which increased after surfactant therapy. There was a negative correlation between the arterial/alveolar PO2 ratio and the deadspace/tidal volume ratio. The alveolar and physiologic deadspace and the deadspace/tidal volume ratio correlated with the arterial end-tidal Pco2 difference. The best correlation was obtained between the arterial end-tidal Pco2 difference and the alveolar deadspace/tidal volume ratio (r = 0.98). CONCLUSIONS: Bronchoalveolar lavage elevates the alveolar and physiologic deadspace and the deadspace/tidal volume ratios and is combined with a fall in the arterial/alveolar PO2 ratio. Surfactant treatment improves the gas exchange but does not restore the lung to its prebronchoalveolar lavage condition, which indicates that the exogenous surfactant affects only partly the recruitment of the atelectatic areas.

Complement and contact activation related to surfactant response in respiratory distress syndrome.

The activation of inflammation and coagulation cascades is part of the pathogenesis of adult respiratory distress syndrome (RDS). Previous studies have demonstrated contact activation in preterm infants with RDS, whereas no concordant results have been found with complement activation. In this study, both systems were investigated in preterm infants with severe RDS and related to surfactant response. Thirty preterm newborns with severe respiratory distress (FiO2 > 0.5), but with no evidence of infection or fetal acidosis, were studied. Eighteen healthy preterm newborns of similar gestational age and birth weight served as controls. The study group was divided into two subgroups, according to their response to a porcine natural surfactant 6 h after administration: responders (FiO2 reduction > 50%) and poor responders (FiO2 reduction < or = 50%). C1q, C4, factor B, C3a, C5a, complement, and C1-inhibitor activity, as well as factor XIIa, were determined in blood samples, drawn 24 h after birth. Except for C1-inhibitor concentration and C1-inhibitor activity, all parameters for infants with severe RDS were different from controls. Complement precursor proteins were lower, and activated split products of the complement and contact system were higher. Infants with a poor response after application of surfactant showed higher amounts of C3a, C5a, and factor XIIa but lower C1q and C4 levels compared with infants with a good response to surfactant. Activation of the complement and the contact system was demonstrated in all respiratory distress patients. This activation was more pronounced in poor responders to exogenous surfactant.

In vitro and in vivo assessment of the Ventrak 1550/Capnogard 1265 for single breath carbon dioxide analysis in neonates.

The Ventrak 1550/Capnogard 1265 (V&C) enables deadspace (VD) measurements to be made in neonates. The aim of our studies was to validate the V&C device for VD measurement in vitro (lung model) and in vivo (adult rabbits). Methods of measurement of VD using the V&C (automatic computation, interactive carbon dioxide-volume plot analysis, Bohr equation) were tested by comparing known added deadspace volumes (VDadd) with calculated VDadd. After producing a change in alveolar (VDalv) and physiological (VDphys) deadspace by in vivo broncho-alveolar lavage, VDalv and VDphys computed automatically were compared with values calculated by the Bohr-Enghoff equations. VDadd was slightly underestimated (absolute error in mean: automatically -0.61 ml; interactively -0.55 ml; Bohr -0.54 ml). The higher the VDadd, the lower the absolute errors and coefficients of variation (cv). The highest cv occurred for automatic analysis (approximately 11%) compared with < 6% for interactive analysis or the Bohr equation. Average differences between results calculated automatically and by the Bohr-Enghoff equation were -0.79 ml for VDalv (95% confidence interval -2.02 to 0.44 ml) and -0.23 ml for VDphys (-0.6 to 0.14 ml). We conclude that the V&C can be used in newborn infants undergoing mechanical ventilation, if changes in VD are < 5 ml, interactive analysis or the Bohr equation should be used.

Complement and contact activation in term neonates after fetal acidosis.

AIMS: To evaluate complement and contact activation after fetal acidosis. METHODS: Fifteen term neonates with hypoxic-ischaemic encephalopathy after umbilical arterial pH < 7.10 were compared with 15 healthy neonates with umbilical arterial pH > 7.20. Determinations of the complement function and C1-inhibitor activity were performed as kinetic tests 22-28 hours after birth. C1q, C1-inhibitor, and factor B concentrations were determined by radial immunodiffusion and those of C3a, C5a, and factor XIIa by enzyme immunoabsorbent assay. RESULTS: Median complement function (46 vs 73%), C1q (4.3 vs 9.1 mg/dl), and factor B (5.2 vs 7.7 mg/dl) decreased after fetal acidosis. The activated split products C3a (260 vs 185 micrograms/l), C5a (5.0 vs 0.6 micrograms/l), and factor XIIa (3.2 vs 1.3 micrograms/l) increased in the neonates after fetal acidosis. No differences were found in the concentration and activity of C1-inhibitor. CONCLUSIONS: Complement and contact activation occurred in the newborns with hypoxic-ischaemic encephalopathy. Activation of these systems generates mediators which can trigger inflammation and tissue injury.

Multisystem organ failure and capillary leak syndrome in severe necrotizing enterocolitis of very low birth weight infants.

BACKGROUND: Classification systems for necrotizing enterocolitis (NEC) in preterm infants have been developed to define severity grades relevant for treatment and prognosis. Multisystem organ failure (MSOF) and capillary leak syndrome (CLS) also have prognostic value in these patients. The aim of this retrospective study was to investigate the incidence and predictive value of MSOF and CLS according to the classification criteria. METHODS: The records of 1,022 very low birth weight infants admitted from 1982 to 1996 were reviewed for diagnosis of NEC stage IIA or higher (classification of Walsh and Kliegman). Among those patients (n = 50) the incidence of MSOF and CLS was determined, separately for surgical or conservative treatment. RESULTS: Twelve patients were assigned to stage II, 22 to stage IIIa, and 16 to stage IIIb; 31 infants underwent operation. Mortality rate was not influenced by the grade. In eight patients only gastrointestinal symptoms were found, whereas in 23 patients, up to three organ systems and in 19 patients, four or more organ systems were affected. Mortality depended on the number of involved organ systems. CLS occurred postoperatively in 10 of the 31 infants; eight of them died. CONCLUSION: The prognostic values of MSOF and CLS are higher than that of classification criteria in NEC of VLBW infants.

Circulatory changes after surfactant bolus instillation in lung-lavaged adult rabbits.

Following surfactant instillation in infants treated for respiratory distress syndrome, a mean arterial blood pressure (MABP) decrease is often observed. Its etiology and pathogenesis are still unknown. In this study various circulatory parameters were recorded continuously after surfactant instillation to elucidate the role of pulmonary vascular resistance as one possible cause for the MABP drop. Seven anesthetized adult New Zealand white rabbits were artificially ventilated after tracheotomy. Arterial and right atrial pressure were recorded continuously. Pulmonary artery pressure and cardiac output were determined by means of a thermodilution catheter. After inducing surfactant deficiency by repeated saline lavages, 200 mg/kg body weight of a natural surfactant preparation was administered by tracheal bolus instillation. PaO2 increased rapidly from 8.0 +/- 1.3 kPa to 51.2 +/- 8.8 kPa (mean +/- standard deviation) within 2 min (p < .05). MABP dropped from 12.1 +/- 1.9 kPa to 8.9 +/- 2.3 kPa within 2 min (p < .05). Pulmonary artery pressure, cardiac output, and right atrial pressure did not change during the observation period of 60 min. The results suggest that a peripheral vasodilatation is the most likely cause for the drop in MABP.

Rapid tracheal infusion of surfactant versus bolus instillation in rabbits: effects on oxygenation, blood pressure and surfactant distribution.

UNLABELLED: Surfactant bolus instillation may be associated with a drop in blood pressure. Platelet-activating factor (PAF) has been found in surfactant preparations. The aim of this study was to evaluate rapid tracheal infusion of surfactant during 5 min as an alternative to bolus instillation and to examine whether a PAF receptor antagonist is able to prevent the decrease in blood pressure. METHODS: Surfactant deficiency was induced in 16 adult rabbits by lung lavages with saline. Six animals received a bolus of a porcine surfactant preparation (Curosurf (CS); 200 mg/kg), labeled with red microspheres to assess pulmonary distribution. In another 5 rabbits, the same amount of labelled CS was instilled by tracheal infusion within 5 min. A third group of 5 animals received 3 mg/kg body weight of the PAF antagonist WEB 2170 before CS bolus instillation. RESULTS: After CS bolus administration, mean PaO2 increased by 44.7 +/- 8.3 kPa (mean +/- SD) within 2 min and remained at this level. Mean arterial blood pressure dropped transiently by 2.3 +/- 2 kPa within 5 min. Pulmonary distribution of surfactant was even. After infusion, mean PaO2 rose by 22.4 +/- 16.3 kPa within 15 min. Blood pressure dropped by 1.8 +/- 1.1 kPa within 15 min. The distribution was extremely uneven. Blood pressure decreases also occurred after pretreatment with PAF receptor antagonist. CONCLUSION: Rapid tracheal infusion of surfactant results in poorer oxygenation, an inhomogeneous distribution and a similar decrease in blood pressure compared to the bolus instillation method. Blood pressure changes could not be prevented by a PAF receptor-specific antagonist.

Value-added services that increase physicians' intent to refer patients to an outpatient nutrition clinic.

We surveyed 378 practicing physicians to identify value-added services that would increase their patient referrals to registered dietitians at an outpatient nutrition clinic. One hundred thirty-nine completed surveys were used in compiling the results. In the survey we presented 16 value-added services specifically designed to overcome physicians' barriers to referral as noted in the literature, and we requested demographic information. Frequency distributions revealed that the services physicians most strongly agreed would increase their referrals were help patients receive reimbursement from third-party graphic information. Frequency distributions revealed that the services physicians most strongly agreed would increase their referrals were help patients receive reimbursement from third-party payers (92.1%), provide a free initial meeting with the patient (82.4%), and offer group cooking classes (81.3%). The services physicians most strongly disagreed would increase their referrals were provide a list of physicians currently using the outpatient nutrition clinic who can provide recommendations (33.1%) and provide a free consultation for the physician (30.2%). Our findings indicate that (a) value-added services most likely to stimulate referrals are those that moderate the cost of nutrition counseling, document its effectiveness, and provide skills training in the form of cooking classes, and (b) value-added services least likely to stimulate referrals are those that require physicians' time.

Modulation of ion transport in cultured rabbit tracheal epithelium by lipoxygenase metabolites.

Lipoxygenase metabolites influence ion movement and fluid balance in the airways. We studied the effects of nordihydroguaiaretic acid (NDGA), a general inhibitor of the lipoxygenase pathway, on Na+ and Cl- secretion in cultured tracheal epithelial cells from adult rabbits through short-circuit current (Isc) and radioactive tracer flux experiments. NDGA inhibition of leukotriene release in freshly isolated rabbit tracheal epithelial cells was assayed by a 3H peptidyl-leukotriene radioimmunoassay. 3 microM NDGA resulted in a 91% reduction of leukotriene release. In unstimulated cultures, Cl- secretion (furosemide-inhibited fraction of Isc) was 11.1 +/- 2.8 muamp/cm2 (n = 10) and was unchanged in the presence of NDGA (n = 10). Epinephrine-stimulated Cl- secretion increased Isc by 12.2 +/- 2 muamp/cm2 (n = 10). This stimulation was unchanged by pretreatment with NDGA (n = 10), suggesting that inhibition of the lipoxygenase pathway did not affect Cl- secretion. In unstimulated cultures, Na+ absorption (amiloride-inhibited portion of Isc) was 10.7 +/- 3.3. muamp/cm2 (n = 10) and was reduced by 79% in the presence of NDGA (n = 10), suggesting that inhibition of the lipoxygenase pathway was associated with inhibition of Na+ absorption. Radioactive tracer flux experiments supported these findings. Exogenous LTD4 (n = 7) and LTC4 (n = 7) were added to cells pretreated with NDGA, and Na+ absorption was restored to 76% and 70% of control, respectively. In addition, LTD4 (n = 4) and LTC4 (n = 4) were added to cells without prior inhibition of the lipoxygenase pathway to NDGA and resulted in an increase in Cl- secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Subacute effects of respiratory syncytial virus infection on lung function in lambs.

We examined the effects of ovine respiratory syncytial virus (RSV) infection on lung mechanics, lung histology, and airway reactivity in lambs. Nine lambs were inoculated with ovine RSV and seven control lambs with normal saline or viral media. Serum neutralization titers were obtained prior to and 3 weeks post-inoculation (PI). Open lung biopsies were performed 1 and 3 weeks PI. Lung mechanics including dynamic compliance (Cdyn), resistance of the lung (RL), and functional residual capacity (FRC) were measured 2 and 6 weeks PI using a plethysmograph. Airway reactivity to aerosolized carbachol, citric acid, and histamine was determined 2 and 6 weeks PI. Most RSV and control lambs were asymptomatic after inoculation. Control lambs had significantly greater average daily weight gain by the third week after inoculation. Seven RSV lambs tested had a fourfold or greater rise in serum neutralization titers, while two control lambs had a fourfold increase. At 2 weeks PI, RSV lambs had significantly lower FRC and higher RL. At 6 weeks RL remained significantly elevated in the RSV lambs. Airway reactivity was not increased in the RSV group. This animal model is useful for studying the effects of RSV infection on lung growth and lung function over time.