Search PubMed⌕ Search

Biomedical subjects

D Hess

Publications and source records attributed to D Hess.

At least 37 records · Page 2Linked to original sources
PubMed2001

Stable transformation of sunflower (Helianthus annuus L.) using a non-meristematic regeneration protocol and green fluorescent protein as a vital marker.

Stable transformation of sunflower was achieved using a non-meristematic hypocotyl explant regeneration protocol of public inbred HA300B. Uniformly transformed shoots were obtained after co-cultivation with Agrobacterium tumefaciens carrying a gfp (green fluorescent protein) gene containing an intron that blocks expression of gfp in Agrobacterium. Easily detectable, bright green fluorescence of transformed tissues was used to establish an optimal regeneration and transformation procedure. By Southern blot analysis, integration of the gfp and nptll genes was confirmed. Stable transformation efficiency was 0.1%. From 68 T1 plants analyzed, 17 showed transmission of transgene DNA and 15 of them contained the intact gfp gene. Expression of gfp was detected in 10 T1 plants carrying the intact gfp gene using a fluorimetric assay or western blot analysis. Expression of the nptll gene was confirmed in 13 T1 plants. The transformation system enables the rapid transfer of agronomically important genes.

Biomarkers↗
PubMed2001

CD40/154 blockade and rejection of islet allografts in the streptozotocin and autoimmune diabetic rat.

Islet transplantation in children with autoimmune diabetes will require immunosuppression that has minimal toxicity and side-effects, and overcomes the barrier of autoimmunity. Since antibodies directed against the CD40/154 co-stimulatory pathway may meet these criteria, we have tested the ability of hamster antirat CD154 (AH.F5, Biogen) to prevent rejection of renal subcapsular islet allografts in streptozotocin (STZ) or autoimmune (AUTO) diabetic diabetes-resistant biobreeding (DRBB) rats. STZ diabetic rats that received anti-CD154 at 15 mg/kg per dose but not 10 mg/kg per dose did not have evidence of rejection until about 80-120 d post-transplantation, by which time antibody concentrations had returned to undetectable levels. Rats retreated with anti-CD154 before recurrence of diabetes had a prolonged period of disease-free survival. Most of these rats had recurrence following a spleen cell challenge. In contrast, AUTO diabetic DRBB rats treated with anti-CD154 had recurrence of diabetes between 7 and 12 d following transplantation of the Dark Agouti (DA) islets. In a separate set, AUTO diabetic rats that received a simultaneous islet isograft, islet allograft and thyroid allograft had focal accumulation of lymphocytes at the periphery of the isograft, while the islet and thyroid allografts had diffuse infiltration with lymphocytes and destruction of tissue with no residual staining for glucagon. Therefore, autoimmunity adds an additional barrier to islet allotransplantation that is not overcome with CD40/154 blockade in an animal model that closely parallels autoimmune diabetes in humans. The results indicate the importance of testing regimen of islet transplantation in animal models of autoimmune diabetes.

Journal Article↗
PubMed2001

Repetitive high-pressure recruitment maneuvers required to maximally recruit lung in a sheep model of acute respiratory distress syndrome.

OBJECTIVE: To compare the effects of two different recruitment maneuvers repeated multiple times on gas exchange lung injury, hemodynamic, and lung mechanics. DESIGN: Randomized prospective comparison. SETTINGS: Animal research laboratory. SUBJECT: Nineteen fasted Hampshire sheep. INTERVENTIONS: In 15 27-kg sheep with saline lavage lung injury, we compared the effects of two recruitment maneuvers: 40 cm H2O continuous positive airway pressure for 60 secs and 40 cm H2O positive end-expiratory pressure with 20 cm H2O pressure control, rate 10 breaths/min, inspiratory to expiratory ratio 1:1 for 2 mins. Each recruitment maneuver was repeated four times, every 30 mins after a 30-sec ventilator disconnection. An additional group received no recruitment maneuvers. Animals were assigned randomly to the three groups and ventilated with 20 cm H2O positive end-expiratory pressure, pressure control 15 cm H2O, rate 20 breaths/min, inspiratory to expiratory ratio 1:1, and Fio2 1.0 between recruitment maneuver periods. MEASUREMENTS AND MAIN RESULTS: Significant and marked increases in Pao2 were observed in the pressure control recruitment maneuver group but only after the second recruitment maneuver. In both the control group and continuous positive airway pressure groups, Pao2 did not significantly increase after any recruitment maneuver compared with baseline injury. There was a significant decrease in cardiac output immediately after some continuous positive airway pressure recruitment maneuvers and a significant increase in mean pulmonary artery pressure in both continuous positive airway pressure and pressure control groups immediately after recruitment maneuvers, but these changes resolved within 10 mins. There were no marked histologic differences between groups and no volutrauma. CONCLUSION: In this model, maximal lung recruitment was obtained with 40 cm H2O positive end-expiratory pressure and 20 cm H2O pressure control applied repetitively every 30 mins for 2 mins without physiologic or histologic harm. Multiple recruitment maneuvers in some animals were required for maximum effect.

Analysis of Variance↗
PubMed2001

Directed proteomics identifies a plant-specific protein rapidly phosphorylated in response to bacterial and fungal elicitors.

The perception of microbial signal molecules is part of the strategy evolved by plants to survive attacks by potential pathogens. To gain a more complete understanding of the early signaling events involved in these responses, we used radioactive orthophosphate to pulse-label suspension-cultured cells of Arabidopsis in conjunction with two-dimensional gel electrophoresis and mass spectrometry to identify proteins that are phosphorylated rapidly in response to bacterial and fungal elicitors. One of these proteins, AtPhos43, and related proteins in tomato and rice, are phosphorylated within minutes after treatment with flagellin or chitin fragments. By measuring (32)P incorporation into AtPhos43 immunoprecipitated from extracts of elicitor-treated hormone and defense-response mutants, we found that phosphorylation of AtPhos43 after flagellin treatment but not chitin treatment is dependent on FLS2, a receptor-like kinase involved in flagellin perception. Induction by both elicitors is not dependent on salicylic acid or EDS1, a putative lipase involved in defense signaling.

Amino Acid Sequence↗
PubMed2001

Ventilator modes used in weaning.

Weaning techniques include spontaneous breathing trials (SBTs), pressure-support ventilation (PSV), and synchronized intermittent mandatory ventilation (SIMV). SBTs can be conducted using one of several approaches, including T-piece breathing, low-level continuous positive airway pressure, low-level PSV, or setting the ventilator to flow-triggering with no pressure applied to the airway. The SBT can be used as a method to identify extubation readiness or as a weaning technique in which the duration of the trial is gradually increased over time. With pressure-support weaning, the level of pressure support is gradually reduced over time. With weaning using SIMV, the mandatory rate setting on the ventilator is gradually reduced. Randomized controlled trials have reported the poorest weaning outcomes using SIMV. Although new ventilator modes have been introduced to facilitate weaning, to date there is no evidence to support the use of these modes. Noninvasive positive-pressure ventilation also has been reported to facilitate weaning, but the ability to generalize these findings remains to be determined.

Humans↗
PubMed2001

Screening for retinopathy of prematurity employing the retcam 120: sensitivity and specificity.

OBJECTIVES: To compare the method of photographic screening using the RetCam 120 to the standard method of screening for retinopathy of prematurity (ROP) by ophthalmologic examination. METHODS: A total of 100 RetCam 120 photoscreening examinations of the retina were performed on 32 premature infants. These were stored in a separate file from which all identifying information was removed. At this same examination, a detailed ophthalmological evaluation, employing the indirect ophthalmoscope with scleral depression, was performed by an experienced examiner. Masked examiners performed an evaluation of the fundus photographs to identify presence or absence of ROP, the location and extent of the disease, and the presence or absence of plus disease. These data were then compared with the results of the ophthalmological examination to determine the specificity, sensitivity, and the positive and negative predictive value (PPV and NPV) of the method. RESULTS: Retinopathy of prematurity was detected in 68 of 100 subjects by ophthalmologic examinations and in 58 of 100 subjects' photoscreening examinations. No ROP was detected in 32 of 100 subjects. The sensitivity of the method was 56 (82.4%) of 68 and the specificity was 30 (93.8%) of 32. The PPV was 96.6%; NPV, 76.9%. CONCLUSIONS: The sensitivity of the method was low. The ROP that was missed was peripheral stage 1 or stage 2 disease in peripheral zone 2 or zone 3. This was largely due to the technical limitations of the speculum-camera interface preventing a better view of the periphery. The ROP cases that were missed by the photographic examination regressed spontaneously on follow-up. No disease more posterior to peripheral zone 2 was overlooked. These results detail the accuracy of the method employing the technique of photoscreening as a potential substitute for detailed ophthalmological examination. At present there are clear technical limitations to such a substitution. The study is part of an ongoing project to determine the feasibility of employing neonatal nurses trained to take digitized images of the premature infant's retina and telemeter the results to be read by an experienced ophthalmologist remote from the site.

False Negative Reactions↗
PubMed2000

C-mannosylation and O-fucosylation of the thrombospondin type 1 module.

Thrombospondin-1 (TSP-1) is a multidomain protein that has been implicated in cell adhesion, motility, and growth. Some of these functions have been localized to the three thrombospondin type 1 repeats (TSRs), modules of approximately 60 amino acids in length with conserved Cys and Trp residues. The Trp residues occur in WXXW patterns, which are the recognition motifs for protein C-mannosylation. This modification involves the attachment of an alpha-mannosyl residue to the C-2 atom of the first tryptophan. Analysis of human platelet TSP-1 revealed that Trp-368, -420, -423, and -480 are C-mannosylated. Mannosylation also occurred in recombinant, baculovirally expressed TSR modules from Sf9 and "High Five" cells, contradictory to earlier reports that such cells do not carry out this reaction. In the course of these studies it was appreciated that the TSRs in TSP-1 undergo a second form of unusual glycosylation. By using a novel mass spectrometric approach, it was found that Ser-377, Thr-432, and Thr-489 in the motif CSX(S/T)CG carry the O-linked disaccharide Glc-Fuc-O-Ser/Thr. This is the first protein in which such a disaccharide has been identified, although protein O-fucosylation is well described in epidermal growth factor-like modules. Both C- and O-glycosylations take place on residues that have been implicated in the interaction of TSP-1 with glycosaminoglycans or other cellular receptors.

Amino Acid Motifs↗
PubMed2000

5-methylcytosine-DNA glycosylase activity is present in a cloned G/T mismatch DNA glycosylase associated with the chicken embryo DNA demethylation complex.

We previously have shown that DNA demethylation by chicken embryo 5-methylcytosine DNA glycosylase (5-MCDG) needs both RNA and proteins. One of these proteins is a RNA helicase. Further peptides were sequenced, and three of them are identical to the mammalian G/T mismatch DNA glycosylase. A 3,233-bp cDNA coding for the chicken homologue of human G/T mismatch DNA glycosylase was isolated and sequenced. The derived amino acid sequence (408 aa) shows 80% identity with the human G/T mismatch DNA glycosylase, and both the C and N-terminal parts have about 50% identity. As for the highly purified chicken embryo DNA demethylation complex the recombinant protein expressed in Escherichia coli has both G/T mismatch and 5-MCDG activities. The recombinant protein has the same substrate specificity as the chicken embryo 5-MCDG where hemimethylated DNA is a better substrate than symmetrically methylated CpGs. The activity ratio of G/T mismatch and 5-MCDG is about 30:1 for the recombinant protein expressed in E. coli and 3:1 for the purified enzyme from chicken embryos. The incubation of a recombinant CpG-rich RNA isolated from the purified DNA demethylation complex with the recombinant enzyme strongly inhibits G/T mismatch glycosylase while slightly stimulating the activity of 5-MCDG. Deletion mutations indicate that G/T mismatch and 5-MCDG activities share the same areas of the N- and C-terminal parts of the protein. In reconstitution experiments RNA helicase in the presence of recombinant RNA and ATP potentiates the activity of 5-MCDG.

Amino Acid Sequence↗
PubMed2000

Stat5a serine phosphorylation. Serine 779 is constitutively phosphorylated in the mammary gland, and serine 725 phosphorylation influences prolactin-stimulated in vitro DNA binding activity.

The activity of transcription factors of the Stat family is controlled by phosphorylation of a conserved, carboxyl-terminal tyrosine residue. Tyrosine phosphorylation is essential for Stat dimerization, nuclear translocation, DNA binding, and transcriptional activation. Phosphorylation of Stats on specific serine residues has also been described. We have previously shown that in HC11 mammary epithelial cells Stat5a is phosphorylated on Tyr(694) in a prolactin-sensitive manner, whereas serine phosphorylation is constitutive (Wartmann, M., Cella, N., Hofer, P., Groner, B., Xiuwen, L., Hennighausen, L., and Hynes, N. E. (1996) J. Biol. Chem. 271, 31863-31868). By using mass spectrometry and site-directed mutagenesis, we have now identified Ser(779), located in a unique Stat5a SP motif, as the site of serine phosphorylation. By using phospho-Ser(779)-specific antiserum, we have determined that Ser(779) is constitutively phosphorylated in mammary glands taken from different developmental stages. Stat5a isolated from spleen, heart, brain, and lung was also found to be phosphorylated on Ser(779). Ser(725) in Stat5a has also been identified as a phosphorylation site (Yamashita, H., Xu, J., Erwin, R. A., Farrar, W. L., Kirken, R. A., and Rui, H. (1998) J. Biol. Chem. 273, 30218-30224). Here we show that mutagenesis of Ser(725), Ser(779), or a combination of Ser(725/779) to an Ala had no effect on prolactin-induced transcriptional activation of a beta-casein reporter construct. However, following prolactin induction the Ser(725) mutant displayed sustained DNA binding activity compared with that of wild type Stat5a. The results suggest that Ser(725) phosphorylation has an impact on signal duration.

Amino Acid Sequence↗
PubMed2000

Ventilators and weaning modes.

Although new ventilator modes have become available to facilitate weaning, there is little evidence that these have improved weaning outcomes. Knowledge based computer weaning systems have also been described, but these are in their infancy, and their role is unclear. Prospective, randomized clinical studies are required to examine whether such modalities are superior to existing approaches.

Fuzzy Logic↗
PubMed2000

Epidoxorubicin and docetaxel as first-line chemotherapy in patients with advanced breast cancer: a multicentric phase I-II study.

BACKGROUND: The combination of anthracyclines and taxanes is currently considered the first choice chemotherapy in advanced breast cancer (ABC) and considerable emphasis has been placed on programs exploring the safest and most efficient way to integrate these classes of drugs in both the metastatic and, more recently, the adjuvant setting. We report here the overall results of the combination of epidoxorubicin (E) 90 mg/m2 and docetaxel (D) 75 mg/m2 as first-line chemotherapy in ABC. PATIENTS AND METHODS: A total of 70 patients were entered in the initial dose-finding study (20 patients) and in the subsequent extended phase II trial (50 patients). Overall 54% of patients had dominant visceral disease and 57% had at least two metastatic sites. Adjuvant anthracyclines were allowed in the phase II part of the study based on the lack of cardiac toxicity observed in the phase I study at a median cumulative E dose of 480 mg/m2. A maximum of eight cycles of the combination was allowed, and cardiac function was monitored at baseline and after every second course by echocardiography. RESULTS: Overall, the median number of cycles administered with the combination was 4 (range 3-8). Neutropenia was confirmed to be the main haematological toxicity, with granulocyte colony-stimulating factor (G-CSF) support required in 44% of the cycles. Febrile neutropenia occurred in 12% of cycles of the combination but 52% of the episodes could be managed on an outpatient basis with oral antibiotics. Overall, the median cumulative dose of E, including prior adjuvant anthracyclines, was 495 mg/m2 (range 270-1020 mg/m2). One patient who received adjuvant E together with radiotherapy to the left chest wall developed fully reversible clinical signs of cardiotoxicity and a significant decrease of LVEF to 35% after a cumulative E dose of 870 mg/m2, with four additional patients (6%) developing asymptomatic and transient decline of resting LVEF. The overall response rate (ORR) in 68 evaluable patients was 66% (95% confidence interval (95% CI): 54%-73%). A comparable antitumour activity of 71% was reported in the group of patients with a prior adjuvant chemotherapy with anthracyclines. After an overall median follow-up time of 22 months (range 4-39+), the median time to progression (TTP) was 4.5 months and the median duration of response was 8 months (range 3-16). No pharmacokinetic (Pk) interaction could be demonstrated between E and D when given simultaneously and sequentially with a one-hour interval. CONCLUSIONS: The combination of E and D in a multiinstitutional setting is an active and safe regimen in poor-prognosis patients with ABC. New combinations and schedules are worth considering in an attempt to further improve disease response and long-term control of the disease.

Adult↗
PubMed2000

Inhaled nitric oxide delivery by anesthesia machines.

UNLABELLED: Inhaled nitric oxide (NO) is a selective pulmonary vasodilator used to treat intraoperative pulmonary hypertension and hypoxemia. In contrast to NO delivered by critical care ventilators, NO delivered by anesthesia machines can be complicated by rebreathing. We evaluated two methods of administering NO intraoperatively: via the nitrous oxide (N(2)O) flowmeter and via the INOvent (Datex-Ohmeda, Madison, WI). We hypothesized that both systems would deliver NO accurately when the fresh gas flow (FGF) rate was higher than the minute ventilation (VE). Each system was set to deliver NO to a lung model. Rebreathing of NO was obtained by decreasing FGF and by simulating partial NO uptake by the lung. At FGF > or = VE (6 L/min), both systems delivered an inspired NO concentration ([NO]) within approximately 10% of the [NO] set. At FGF < VE and complete NO uptake, the N(2)O flowmeter delivered a lower [NO] (70 and 40% of the [NO] set at 4 and 2 L/min, respectively) and the INOvent delivered a higher [NO] (10 and 23% higher than the [NO] set at 4 and 2 L/min, respectively). Decreasing the NO uptake increased the inspired [NO] similarly with both systems. At 4 L/min FGF, [NO] increased by 10%-20% with 60% uptake and by 18%-23% with 30% uptake. At 2 L/min, [NO] increased by 30%-33% with 60% uptake and by 60%-69% with 30% uptake. We conclude that intraoperative NO inhalation is accurate when administered either by the N(2)O flowmeter of an anesthesia machine or by the INOvent when FGF > or = VE. IMPLICATIONS: Inhaled nitric oxide (NO) is a selective pulmonary vasodilator. In a lung model, we demonstrated that NO can be delivered accurately by a N(2)O flowmeter or by a commercial device. We provide guidelines for intraoperative NO delivery.

Administration, Inhalation↗
PubMed2000

Use of recruitment maneuvers and high-positive end-expiratory pressure in a patient with acute respiratory distress syndrome.

OBJECTIVE: To present the use of a novel high-pressure recruitment maneuver followed by high levels of positive end-expiratory pressure in a patient with the acute respiratory distress syndrome (ARDS). DESIGN: Observations in one patient. SETTING: The medical intensive care unit at a tertiary care university teaching hospital. PATIENT: A 32-yr-old woman with severe ARDS secondary to streptococcal sepsis. INTERVENTIONS: The patient had severe gas exchange abnormalities because of acute lung injury and marked lung collapse. Attempts to optimize recruitment based on the inflation pressure-volume (PV) curve were not sufficient to avoid dependent lung collapse. We used a recruitment maneuver using 40 cm H2O of positive end-expiratory pressure (PEEP) and 20 cm H2O of pressure controlled ventilation above PEEP for 2 mins to successfully recruit the lung. The recruitment was maintained with 25 cm H2O of PEEP, which was much higher than the PEEP predicted by the lower inflection point (P(Flex)) of the PV curve. MEASUREMENTS AND MAIN RESULTS: Recruitment was assessed by improvements in oxygenation and by computed tomography of the chest. With the recruitment maneuvers, the patient had a dramatic improvement in gas exchange and we were able to demonstrate nearly complete recruitment of the lung by computed tomography. A PV curve was measured that demonstrated a P(Flex) of 16-18 cm H2O. CONCLUSION: Accumulating data suggest that the maximization and maintenance of lung recruitment may reduce lung parenchymal injury from positive pressure ventilation in ARDS. We demonstrate that in this case PEEP alone was not adequate to recruit the injured lung and that a high-pressure recruitment maneuver was required. After recruitment, high-level PEEP was needed to prevent derecruitment and this level of PEEP was not adequately predicted by the P(Flex) of the PV curve.

Adult↗
PubMed2000

Techniques to avoid intubation: noninvasive positive pressure ventilation and heliox therapy.

NPPV is useful in decreasing the intubation rate in carefully selected patients with acute respiratory failure--particularly in patients with COPD. The results of some studies also suggest a survival benefit for use of NPPV with acute respiratory failure associated with COPD. Heliox therapy may be beneficial for selected patients with large airway obstruction or asthma. The use of heliox for other indications is unclear. Heliox may adversely affect the function of respiratory care equipment such as flow meters, ventilators, nebulizers, and pulmonary function monitors.

Airway Obstruction↗
PubMed2000

Association of betaine-homocysteine S-methyltransferase with microtubules.

In mammals, betaine of the mitochondrial matrix is used in the cytosol by betaine-homocysteine S-methyltransferase for methionine synthesis. The resulting dimethylglycine is shuttled back into the mitochondrial matrix for further degradation. Nanospray tandem mass spectrometry and N-terminal amino acid sequencing of microtubule-associated proteins from rat liver tubulin revealed that betaine-homocysteine S-methyltransferase is microtubule associated. This was confirmed by confocal laser scanning microscopy of HepG2 cells labeled with betaine-homocysteine S-methyltransferase- and alpha-tubulin-specific monoclonal antibodies. The association of betaine-homocysteine S-methyltransferase with the cytoskeleton may functionally integrate the mitochondrial and cytoplasmic compartments of choline degradation.

Amino Acid Sequence↗