Gas-liquid chromatography-mass spectrometry of hydroxy fatty acids as their methyl esters tert.-butyldimethylsilyl ethers

BACKGROUND: The objective of this paper is to illustrate the effect of a gap of 5 mm, an overlap of 5 mm and a perfect match on the dose distribution across the junction of tangential breast fields and adjacent supraclavicular and axillary fields. MATERIALS AND METHODS: For this purpose film dosimetry was applied to measure relative dose distributions in two sagittal planes in an anthropomorphic breast phantom having cork lungs, simulating a radiation therapy treatment of the breast and adjacent supraclavicular lymph nodes. Two different treatment techniques, an SSD match technique and a geometrically exact isocentric match technique, as routinely applied in the two institutes were examined. The three-dimensional treatment planning system of each institute was used to calculate the dose distribution in the match region of the supraclavicular fields and the two opposing tangential fields. The measured and calculated dose distributions were evaluated and compared along lines in two sagittal planes from the supraclavicular fields down to the tangential fields crossing the match planes. These dose distributions in the match region were extremely dependent on the set-up of the fields. RESULTS: Although the reproducibility of the film measurements was within 2%, it became clear that the set-up of the fields to achieve a gap of 5 mm, a perfect match or an overlap of 5 mm required a lot of attention, even when using a phantom. CONCLUSIONS: It can be concluded that in clinical practice, these set-up difficulties do influence the dose distribution in the match region much more than the systematic uncertainties in the dose calculation algorithms of the treatment planning systems and the type of treatment technique.     

Prostanoid biosynthesis by blood monocytes of children with hyperprostaglandin E syndrome

Hyperprostaglandin E syndrome (HPS), the prenatal variant of Bartter's syndrome, is characterized by a marked and selective stimulation of prostaglandin E (PGE2) synthesis. In the study group HPS patients showed increased urinary levels of PGE2, an index of renal, and of 11 alpha-hydroxy-9,15-dioxo-2,3,4,5,20-pentanor-19-carboxyprostano ic acid (PGE-M), an index of systemic PGE2 synthesis of 470% and of 570%, respectively. In addition, plasma concentration of PGE-M was also elevated 6.3-fold when compared with a control group. The urinary levels of other prostanoids were unaltered. During indomethacin treatment in both groups prostanoid excretion rates were suppressed to similar levels. To investigate the origin of stimulated prostanoid biosynthesis in HPS patients CD14+ monocytes were isolated from plasma samples, and the prostanoid synthesis was analyzed. The pattern and amounts of metabolites synthesized from endogenous arachidonic acid pools did not vary significantly between monocytes of the HPS and the control group. Thromboxane A2 (TXA2) was formed as the major prostanoid product. Using PGH2 as an exogenous substrate, again no difference in PGE2 biosynthesis was observed, indicating no difference in PGE-synthetic activity between both groups. Additionally, mRNA expression analysis of CD14+ monocytes via RT-PCR delineated the constitutive expression of cyclooxygenase-1, cyclooxygenase-2, and thromboxane synthase mRNA in cells from HPS patients and controls without statistical differences between these two groups. In conclusion, our data show that monocytes are not the source for the increased PGE2 biosynthesis in children with HPS, and a genetic defect in PGE synthesis can be excluded as the primary event in the pathogenesis in HPS.     

The Ability of the Nitric Oxide Synthases Inhibitor T1023 to Selectively Protect the Non-Malignant Tissues

Previously, we showed that a nitric oxide synthase (NOS) inhibitor, compound T1023, induces transient hypoxia and prevents acute radiation syndrome (ARS) in mice. Significant efficacy (according to various tests, dose modifying factor (DMF)—1.6–1.9 against H-ARS/G-ARS) and safety in radioprotective doses (1/5–1/4 LD10) became the reason for testing its ability to prevent complications of tumor radiation therapy (RT). Research methods included studying T1023 effects on skin acute radiation reactions (RSR) in rats and mice without tumors and in tumor-bearing animals. The effects were evaluated using clinical, morphological and histological techniques as well as RTOG classification. T1023 administration prior to irradiation significantly limited the severity of acute RSR. This was due to a decrease in radiation alteration of the skin and underlying tissues, and the preservation of the functional activity of cell populations that are critical in the pathogenesis of radiation burn. The DMF values for T1023 for skin protection were 1.4–1.7. Moreover, its radioprotective effect was fully selective to normal tissues in RT models of solid tumors—T1023 reduced the severity of acute RSR and did not modify the antitumor effects of γ-radiation. The results indicate that T1023 can selectively protect the non-malignant tissues against γ-radiation due to hypoxic mechanism of action and potentiate opportunities of NOS inhibitors in RT complications prevention.     

Double-helix coil for occlusion of large patent ductus arteriosus: evaluation in a chronic lamb model

Glyceraldehyde-3-phosphate dehydrogenase (G3PDH) is often used as a control gene for mRNA expression, however it has been proposed to be overexpressed in all hepatocellular carcinomas (HCC). Equal amounts of tumor and paired normal (T/N) RNA, based on OD260/280 nm, were compared using ethidium bromide staining, poly-T probing, gene-specific dot blot and Northern blots using control probes G3PDH, actin and histone H4. Using mRNA blots 13/20 surgical HCC pairs did not overexpress G3PDH. Those 7/20 intact samples which did appear to overexpress G3PDH on Northern blot could not be detected by poly-T probing of dot blots. The apparent overexpression was not specific for the control gene G3PDH nor for the malignancy HCC. It may represent partial mRNA degradation, or the presence of as yet unknown substances which interfere with absorption at 260/280 nm. We advise caution in selecting human T/N pairs for differential gene expression studies. For HCC, no clinicopathological variables, including cirrhosis, predicted whether a T/N sample pair was likely to be balanced or not.     

A re-examination of seasonal variation in suicides in Australia and New Zealand

BACKGROUND: To examine the seasonality of suicides in Australia and New Zealand during the period 1981 to 1993. METHODS: A chi-square test and a harmonic analysis were used to detect the seasonality of the suicide data. RESULTS: The reduced amplitude and a smaller proportion of variance accounted for by seasonality suggested the seasonal effect on suicide is greatly diminished. The absence of biseasonal distribution of female suicides was also consistently found in the two countries. The finding was contrary to the reported results in seventies in many Western countries. CONCLUSIONS: The change in living condition, roles of males and females and communication pattern resulted in the reduction of climatic and environment effect in the seasonality of suicides were suggested. LIMITATIONS: The results would be better if a longer series of suicide date were available.     

Cleavage motifs of the yeast 20S proteasome beta subunits deduced from digests of enolase 1

The 436-amino acid protein enolase 1 from yeast was degraded in vitro by purified wild-type and mutant yeast 20S proteasome particles. Analysis of the cleavage products at different times revealed a processive degradation mechanism and a length distribution of fragments ranging from 3 to 25 amino acids with an average length of 7 to 8 amino acids. Surprisingly, the average fragment length was very similar between wild-type and mutant 20S proteasomes with reduced numbers of active sites. This implies that the fragment length is not influenced by the distance between the active sites, as previously postulated. A detailed analysis of the cleavages also allowed the identification of certain amino acid characteristics in positions flanking the cleavage site that guide the selection of the P1 residues by the three active beta subunits. Because yeast and mammalian proteasomes are highly homologous, similar cleavage motifs might be used by mammalian proteasomes. Therefore, our data provide a basis for predicting proteasomal degradation products from which peptides are sampled by major histocompatibility complex class I molecules for presentation to cytotoxic T cells.     

Endothelin A-receptor blockade worsens endotoxin-induced hepatic microcirculatory changes and necrosis

BACKGROUND & AIMS: Endothelin 1 is considered to be an important regulator of sinusoidal blood flow and increases during endotoxemia. The purpose of this study was to investigate the role of endothelin 1 in hepatic microcirculation, oxygen transport, and liver injury during endotoxemia. METHODS: Male Sprague-Dawley rats were continuously infused with 2.5 mL/h of saline, 0.8 mg . kg-1 . h-1 of lipopolysaccharide (LPS), 3 mg . kg-1 . h-1 of BQ-485, an endothelin A-receptor antagonist, or LPS plus BQ-485 for 7 hours. RESULTS: BQ-485 infusion had no significant effect on hepatic microcirculation and liver injury. LPS increased the plasma levels of aspartate aminotransferase (AST) and total bilirubin and decreased the hepatic adenosine triphosphate (ATP) level and bile flow rate. LPS + BQ-485 infusion further increased the plasma levels of AST and total bilirubin and decreased the bile flow rate and the hepatic ATP level. Dual-spot microspectroscopy revealed mild decreases in sinusoidal erythrocyte velocity and oxygen transport in the LPS group and profound decreases in these parameters in the LPS + BQ-485 group. Histological examinations revealed massive necrotic changes in the pericentral regions of the LPS + BQ-485 group. CONCLUSIONS: These results suggest that blockade of endothelin A receptors disturbs hepatic microcirculation and oxygen transport and aggravates the necrotic injury induced by endotoxin.     

Conformation of xanthene dyes in the sulfhydryl 1 binding site of myosin. 2

The fluorescent dyes 5'-(iodoacetamido)tetramethylrhodamine (5'IATR) and 5'-(iodoacetamido)-fluorescein (5'IAF) bind covalently to the reactive sulfhydryl (SH1) of myosin subfragment 1 (S1), the 5'IATR as a dimer and the 5'IAF as a monomer. The conformation of the dimer and the dye-protein complex was investigated by comparison of several spectroscopic signals of the molecules before and after their association into a complex and interpretation of any changes using a coupled dipole oscillator model adapted for this problem [Burghardt & Ajtai (1995) Biophys. Chem. (submitted for publication)]. Absorption and fluorescence spectroscopies were performed on 5'IAF, 5'IATR, and rhodamine 6G (R6G) and rhodamine B (RB) as models of dimer conformation. Absorption, fluorescence, and circular dichroism (CD) spectroscopies were performed on 5'IATR-modified S1 (5'R-S1) and 5'IAF-modified S1 (5'F-S1). Combined spectroscopic and 2-D NMR data from rhodamines in solution determined the conformations of the dimers. Xanthene rings from dimers of identical dyes (homodimers) stacked in two structures having very different spectroscopic signatures. Xanthene rings from the heterodimer of R6G and RB stacked in one conformation. The two homodimer conformations of 5'IATR are equally likely to form in solution. The other rhodamine homodimers have one dominant, but not exclusive, structure. Both conformations of the 5'IATR dimer were coupled to a tryptophan as a model of the dye-protein interaction at SH1. The calculated CD from one dimer conformer (dimer A) coupled to tryptophan is negative for the lowest energy CD absorption band. The other dimer (dimer B) gives positive CD on the two lowest energy CD absorption bands. Both dimer structures of 5'IATR contributed to the early time-dependent CD signal from 5'IATR binding to SH1, but at equilibrium the CD signal indicated only dimer B, suggesting that the SH1 binding pocket converts dimer A into dimer B. The time-dependent CD signal from 5'IAF changes amplitude but not shape during the reaction with SH1. The model calculation accounting for the spectroscopic signals of 5'R-S1 and 5'F-S1 indicates several likely conformations of the 5'IATR dimer-tryptophan and 5'IAF-tryptophan complexes embedded in S1. These structures fit to the alpha-carbon structure of the SH1 binding pocket when the 5'IATR dimer and 5'IAF interact closely with Trp510 [Rayment et al. (1993) Science 261, 50-58].(ABSTRACT TRUNCATED AT 400 WORDS)     

Substrate specificity of human prolyl-4-hydroxylase

BACKGROUND: The mechanism responsible for the forward blood flow associated with external chest compression is still controversial. Evidence for both blood flow caused by direct cardiac compression and blood flow generated by a general increase in intrathoracic pressure has been found in experimental as well as clinical studies. No data are available concerning the mechanism causing forward blood flow in hypothermic patients undergoing cardiopulmonary resuscitation. Therefore, echocardiographic findings during external chest compression in seven hypothermic arrest victims are reported. METHODS: All transesophageal echocardiographic studies performed at the Anaesthesia department between 1994 and 1997 were reviewed and seven hypothermic patients with transesophageal echocardiography performed during cardiopulmonary resuscitation were identified. RESULTS: An open mitral valve or a circumferential reduction in aortic diameter during the compression phase was found in four of seven patients, indicating that primarily an increase in intrathoracic pressure (thoracic pump mechanism) generated forward blood flow. In three patients, mitral valve closure during external chest compression indicated that direct cardiac compression (cardiac pump mechanism) contributed to forward blood flow. Two patients studied during active compression-decompression cardiopulmonary resuscitation demonstrated enhanced right ventricular filling and aortic valve opening during active decompression of the thorax. CONCLUSIONS: In contrast to normothermic arrest victims, an open mitral valve during external chest compression is a common finding during hypothermia, indicating that thoracic pump mechanism is important for forward blood flow during cardiopulmonary resuscitation in hypothermic arrest victims. Aortic valve opening in two hypothermic arrest victims suggests forward blood flow also during active decompression of the thorax with the Cardiopump.