A novel, multilayer structure of a helical peptide.

X-ray diffraction analysis at 1.5 A resolution has confirmed the helical conformation of a de novo designed 18-residue peptide. However, the crystal structure reveals the formation of continuous molecular layers of parallel-packed amphiphilic helices as a result of much more extensive helix-helix interactions than predicted. The crystal packing arrangement, by virtue of distinct antiparallel packing interactions, segregates the polar and apolar surfaces of the helices into discrete and well-defined interfacial regions. An extensive "ridges-into-grooves" interdigitation characterizes the hydrophobic interface, whereas an extensive network of salt bridges and hydrogen bonds dominates the corresponding hydrophilic interface.

Dosimetry and treatment planning for 90Y-labeled antiferritin in hepatoma.

Radiation absorbed-dose estimates and treatment planning are reported for 11 patients with hepatoma who were administered 90Y-labeled polyclonal antiferritin IgG for therapy in a Phase 1-2 trial. Dosimetric studies included quantitation of the localization and clearance of 111In-labeled antiferritin IgG in tumor and normal tissues and computer-assisted tumor and normal liver volumetrics from X ray CT scans. For the group of patients studied, hepatoma volumes at the time of treatment ranged from 135 to 3442 cm3. Quantitative 111In antiferritin imaging prior to and following 600 or 900 cGy of external-beam irradiation of the primary tumor demonstrated that tumor uptake increased 1.1 to 5.8-fold (mean 2.8) following external beam. In contrast, changes in uptake of radiolabeled antiferritin in normal liver ranged from 0.35 to 2.1-fold (mean 0.93) after external irradiation. Administered activities of 90Y antiferritin ranged from 8 to 37 mCi and were dependent on tumor volume and tumor localization of radiolabeled antiferritin. Following external-beam irradiation, tumor dose rates achieved with 90Y antiferritin ranged from 10 to 20 cGy/hr and normal liver dose rates from 1.1 to 5.7 cGy/h. The corresponding absorbed dose in hepatomas ranged from 900 to 2150 cGy and in normal liver from 80 to 650 cGy. After external-beam irradiation, tumor and normal liver uptake of 90Y antiferritin was consistent with that of 131I antiferritin.

A substituent constant analysis of the interaction of substituted naphthalene monoimides with DNA.

In a continuing analysis of substituent effects in intercalator-DNA interactions, an unsubstituted naphthalene monoimide, with a 3-(dimethylamino)propyl group on the imide nitrogen has been prepared along with 3- and 4-nitro- and 3- and 4-amino-substituted derivatives. These derivatives allow an evaluation of the importance of the Hammett substituent constant and of the substituent position on the binding of naphthalene monoimides to DNA. Viscosity and spectrophotometric analyses indicate that all five compounds bind to DNA by intercalation. The 4-nitro compound gives a smaller viscosity increase and binds only approximately one-third as strongly as the 3-nitro derivative. It is postulated that this difference is due to the significant angle that the 4-nitro group makes with the intercalated monoimide ring system. The 3-NO2 group can assume a coplanar configuration with the monoimide ring system, allowing more favorable interactions with DNA base pairs, larger viscosity increases, and stronger binding to DNA. The binding constants of the 3-substituted monoimides are in the order 2 greater than 4 greater than 1 and, thus, do not follow a substituent constant pattern. The Tm values from thermal melting of DNA, on the other hand, are in the order 2 greater than 1 greater than 4, suggesting that the enthalpy contributions are significantly different for the binding of the three compounds to DNA. van't Hoff plots support this finding and indicate that both enthalpy and entropy contribute significantly to the binding free energy of 1 and 2 while the binding of 4 is primarily an enthalpic process. Plots of Tm and 65 degrees C log K values as a function of substituent constant for 1, 2, and 4 are linear. CPK model building studies suggest that 4 can form a hydrogen bond with the 5' diester oxygen of the sugar-phosphate backbone of DNA in an intercalation complex. This would lead to more favorable energetics of binding but a loss of mobility and/or available binding configurations with a resulting enthalpy-entropy compensation in the binding free energy of 4. This series of compounds dramatically illustrates the steric and hydrogen bonding complexity that can arise in attempts to design drugs to favorably interact with a DNA intercalation site as a potential bioreceptor.

Validation of the circular harmonic transform (CHT) algorithm for quantitative SPECT.

The purpose of this study was to validate the use of the circular harmonic transform (CHT) algorithm for quantitative single-photon emission computed tomography (SPECT) with isotopes technetium-99m (99mTc) and indium-111 (111In) under clinically relevant conditions. Phantom studies were the principal tools used. Volumes of fillable organs within a tissue-equivalent anthropomorphic phantom were determined over a wide range (145-1960 ml) to within 6% by using a thresholding technique. Additionally, phantom studies with nonuniform activity distributions were made. These included a background of activity and hot as well as cold lesions. The hot lesion was computed to within 12% (111In) and 7.7% (99mTc), and contrast in the cold lesion was approximately 70% for both isotopes. The CHT algorithm incorporates the energy-distance relation (EDR) which minimizes the degrading effects of attenuation, scatter, collimator blur and poor statistics.

Quantitative SPECT for indium-111-labeled antibodies in the livers of beagle dogs.

Results are presented for SPECT computations of liver volumes and 111In-labeled antibody activities in the livers of eight normal beagle dogs. Administered activities ranged from 1 to 2 mCi. SPECT studies were acquired 1 day postinjection using a rotating gamma camera system with elliptical orbits in a 360-degree rotation (128 views, 15 sec/view, 64 x 64 matrices). Uniformity-corrected images were reconstructed by use of the circular harmonic transform algorithm with computer software developed in-house. Liver volumes and activities were computed from transverse slices, 1 pixel (6.25 mm) in thickness. Comparison of SPECT and autopsy data demonstrated that absolute values of percent differences between measured and computed liver volumes ranged from 1.0% to 7.2%. Absolute values of percent differences between autopsy data and computed 111In activities in the liver ranged from 2.3% to 7.5%. These results suggest that quantitative SPECT has the potential of becoming an important tool in clinical trials for determining activities and localization volumes of radiolabeled antibodies directly from radionuclide images.

Effects of signalling transduction modulators on the transformed phenotypes in v-H-ras-transformed NIH 3T3 cells.

Several signalling transduction modulators were used to examine their effects on the morphological changes, foci formation in soft agar and cellular growth in v-H-ras-transformed NIH 3T3 cells. The results from this study showed that specific tyrosine kinase inhibitors (genistein and tyrphostin 23) and cyclic AMP-elevating agents (forskolin and 3-isobutyl-1-methyl-xanthine) could effectively induce differential flat phenotype of v-H-ras transformant at micromolar concentrations. At the same dose range, both signalling modulators also caused a significant suppression of anchorage-independent and cellular growth in the same transformant. By contrast, compound inhibitors such as protein kinase C (staurosporin and H-7), phospholipase A2 (aristolochic acid), phospholipase C (neomycin sulfate) and cyclooxygenase (indomethacin) all did not alter the cellular morphology or foci formation in soft agar, although PKC inhibitors exhibited a slight inhibition on the cellular growth. Based on these observations, we propose that the alterations of protein kinase A or tyrosine kinase-associated signal pathways is necessary and the original cause of the transformation event, but that increase of the activities of protein kinase C, phospholipase C, phospholipase A2 or cyclooxygenase probably is an indirect result of the v-H-ras-mediated transformation.

Reversion of the transformed phenotypes of v-H-ras NIH3T3 cells by flavonoids through attenuating the content of phosphotyrosine.

Fifteen flavonoids were employed to examine their effects on the morphological changes, foci formation in soft agar and cellular growth in v-H-ras-transformed NIH3T3 cells. The data presented here demonstrated that only three specific flavonoids--apigenin, kaempferol, and genistein--exhibited the reverting effect on the transformed phenotypes of ras-3T3 cells. For example, treatment with 25 microM of these flavonoids could effectively reverse the transformed morphology of ras-3T3 cells into flatter cells with contact inhibition. Colony formation in soft agar was decreased to 0.11%, 0.15%, and 0.35% by 25 microM apigenin, kaempferol, and genistein, respectively, as compared with control (0.92%). It was also found that the proliferation of ras-3T3 cells was significantly inhibited by these compounds in a dose-dependent manner. Finally, two biochemical parameters, the content of phosphotyrosine and cAMP, were examined to see whether affected by these compounds. The results showed the phosphotyrosine content in ras-3T3 cells was dramatically decreased by apigenin and kaempferol, but that was slightly reduced by genistein. By contrast, these three flavonoids all failed to significantly alter the level of cAMP within this transformant. Based on these observations, we suggest that some specific flavonoids are capable of reverting the transforming properties of v-H-ras transformed cells. The possible mechanism of this reversion is not mediated by activating the protein kinase A or its associated pathways, but rather inhibiting tyrosine kinases, subsequently leading to the blockage of p21ras-mediated signal transduction circuitry.

DHEA inhibits cell growth and induces apoptosis in BV-2 cells and the effects are inversely associated with glucose concentration in the medium.

Dehydroepiandrosterone (DHEA), a major steroid secreted by the adrenal gland which decreases with age after adolescence, is available as a nutritional supplement. DHEA is known to have antiproliferative effects but the mechanism is unclear. In this study using BV-2 cells, a murine microglial cell line, we investigated the effect of DHEA on cell viability and the interaction between DHEA and glucose concentrations in the medium. We showed that DHEA inhibited cell viability and G6PD activity in a dose-dependent manner and that the effect of DHEA on cell viability was inversely associated with glucose concentrations in the medium, i.e. lowered glucose strongly enhanced the inhibition of cell viability by DHEA. DHEA inhibited cell growth by causing cell cycle arrest primarily in the G0--G1 phase, and the effect was more pronounced at zero glucose (no glucose added, G0) than high glucose (4.5 mg/ml of the medium, G4.5). Glucose deprivation also enhanced apoptosis induced by DHEA. At G4.5, DHEA did not induce formation of DNA ladder until it reached 200 microM. However, at G0, 100 microM DHEA was able to induce apoptosis, as evidenced by the formation of DNA ladder, elevation of histone-associated DNA fragmentation and increase in cells positively stained with annexin V-FITC and annexin V-FITC/propidium iodide. The interactions between DHEA and glucose support the contention that DHEA exerts its antiproliferative effects through alteration of glucose metabolism, possibly by inhibition of G6PD activity leading to decreased supply of ribose-5-phosphate for synthesis of DNA and RNA. Although DHEA is only antiproliferative at pharmacological levels, our results indicate that its antiproliferative effect can be enhanced by limiting the supply of glucose such as by energy restriction. In addition, the present study shows that glucose concentration is an important factor to consider when studying the antiproliferative and toxicological effects of DHEA.

Participation of Galaninergic Neurotransmission at the Paraventricular Hypothalamic Nucleus in the Suppression of Baroreceptor Reflex Response by Locus ceruleus in the Rat.

We evaluated the potential participation of galanin (GAL) at the paraventricular nucleus of hypothalamus (PVN) in the suppression of baroreceptor reflex (BRR) response by locus ceruleus (LC), using adult male Sprague-Dawley rats anesthetized with pentobarbital sodium. Microinjection of GAL (100 pmol) bilaterally into the PVN significantly depressed the BRR response. This suppressive effect was appreciably antagonized when GAL (100 pmol) and GAL antiserum (1:20) were coadministered into the bilateral PVN. Whereas bilateral microinjection of GAL antiserum into the PVN by itself elicited minimal effect, it nevertheless significantly attenuated the suppressive effect of either electrical or chemical activation of LC on the BRR response. Pretreatment with the same amount of normal rabbit serum (1:20), on the other hand, was ineffective. These results suggest that a galaninergic projection from the LC to PVN may participate in the suppression of BRR response by this dorsal pontine nucleus. Copyright 1997 S. Karger AG, Basel

Effective atomic numbers for low-energy total photon interactions in human tissues.

A new method is introduced in which the total photon interaction cross sections per electron of human tissues are used to define effective atomic numbers for blood, bone, brain, fat, heart, kidney, liver, lung, muscle, ovary, pancreas, spleen, and water. These effective atomic numbers are equal within 4% from 10 to 200 keV in each soft tissue, whereas for bones of different chemical compositions the variation ranges from 2.86% to 5.03%. This effective atomic number definition is less energy dependent than a previous definition based on the total photon interaction cross section per atom averaged over all elements in the tissue, from which the computed effective atomic numbers varied by as much as 50% (in bone) as a function of photon energy over the energy range from 10 to 200 keV.

The circular harmonic transform for SPECT reconstruction and boundary conditions on the Fourier transform of the sinogram.

The circular harmonic transform (CHT) solution of the exponential Randon transform (ERT) is applied to single-photon emission computed tomography (SPECT) for uniform attenuation within a convex boundary. An important special case also considered is the linear (unattenuated) Radon transform (LRT). The solution is on the form of an orthogonal function expansion matched to projections that are in parallel-ray geometry. This property allows for efficient and accurate processing of the projections with fast Fourier transform (FFT) without interpolation or beam matching. The algorithm is optimized by the use of boundary conditions on the 2-D Fourier transform of the sinogram. These boundary conditions imply that the signal energy of the sinogram is concentrated in well-defined sectors in transform space. The angle defining the sectors depends in a direct way on the radius of the field view. These results are also obtained for fan-beam geometry and the linear Radon transform (the Fourier-Chebyshev transform of the sinogram) to demonstrate that the boundary conditions are a more general property of the Radon transform and a not a property unique to rectangular coordinates.

A novel method for determining the blood/gas partition coefficients of inhalation anesthetics to calculate the percentage of loss at different temperatures.

The loss by blood/gas (lambda) partition of inhalation anesthetics can be estimated by an equation for the percentage of loss. However, because lambdas of inhalation anesthetics at different temperatures have not been fully determined so far, the percentage of loss at varying temperature in various headspace volumes cannot be estimated. Therefore, a novel method was developed for the determination of inhalation anesthetic lambda, in this study. The method was precise, with a relative standard deviation of less than 5%. The average of lambda from seven distinct blood samples at 4 degrees C, 25 degrees C, and 37 degrees C were determined as 6.68, 2.04, and 1.32 of isoflurane; 3.47, 1.10, and 0.65 of sevoflurane; and 2.31, 0.75, and 0.46 of desflurane, respectively. In addition, increasing temperature was found to decrease lambda profoundly by a secondary order mechanism. Using the obtained value of lambda, the percentage of loss of isoflurane, sevoflurane, and desflurane were then predicted using a 5-mL vacuum tube as a collecting container for an example. In conclusion, a novel method was developed here for lambda determination, and lambdas of isoflurane, sevoflurane, and desflurane at various temperatures were given for estimating the loss resulting from liquid/gas partitioning.

Clinical presentations and skin denervation in amyloid neuropathy due to transthyretin Ala97Ser.

OBJECTIVE: Familial amyloid polyneuropathy (FAP) due to amyloidogenic transthyretin (TTR) is often associated with impairment of thermonociceptive functions. This study investigated skin innervation and its clinical significance in genetically defined FAP due to a hot-spot Ala97Ser TTR mutation (Ala97Ser). METHODS: Skin biopsies were performed on the distal leg of patients with Ala97Ser, and intraepidermal nerve fiber (IENF) densities were quantified. RESULTS: There were 19 unrelated patients with Ala97Ser manifesting a late-onset (59.47 +/- 5.70 years) generalized neuropathy with disabling motor, sensory, and autonomic symptoms. Against a background of a slowly progressive course, 7 patients (36.8%) exhibited additional rapid declines in neurologic deficits, which were associated with elevation of the protein content in the CSF (p < 0.001). The IENF density was markedly reduced in Ala97Ser patients compared to age- and gender-matched controls (0.99 +/- 1.11 vs 8.31 +/- 2.87 fibers/mm, p < 0.001). Skin denervation was present in all patients and was lower in patients with a higher disability grade (0.17 +/- 0.26 vs 1.37 +/- 1.16 fibers/mm, p = 0.003). Albuminocytologic dissociation in the CSF was observed in 14 patients (73.7%), and the IENF density was negatively correlated with the CSF protein concentration (p = 0.015). CONCLUSIONS: Skin denervation was common in Ala97Ser, and degeneration of cutaneous nerve terminals was correlated with the severity of clinical phenotypes and the level of CSF protein.

Application of the mixed venous blood concentration equation in desflurane anesthesia.

BACKGROUND: We have previously proposed an equation derived from Fick's law and Lin's concept of effective blood concentration (EBC) to calculate the mixed venous blood concentration (MVBC) of isoflurane. Desflurane has a lower blood/air partition coefficient than isoflurane and, as such, promotes a faster induction and recovery from anesthesia. In this study, we investigated the application of the MVBC equation to predict the MVBC of desflurane. METHODS: We maintained anesthesia with a fixed inspired concentration (CI) of desflurane (10%) during cardiac anesthesia in 11 patients. In order to measure the real concentrations of desflurane in mixed venous blood, pulmonary arterial blood samples were collected at different time points via a Swan-Ganz catheter for gas chromatographic-mass spectrometric determination. The relationship between the calculated concentrations and the actual blood sample concentrations of desflurane in mixed venous blood was investigated. Lin's EBC method was also used and the results were compared with those of MVBC. RESULTS: The calculations from our derived MVBC equation and the actual blood concentrations showed a similar kinetic pattern; the concentration levels were approximately the same and correlated well (r = 0.89) during anesthesia. However, the EBC method failed to accurately estimate the actual blood concentrations. CONCLUSIONS: The results demonstrate that our equation, but not the EBC method, may be useful for estimating pulmonary blood concentrations of desflurane. The clinical significance and the importance of the method merit further investigation.

Application of MVBC equation to predict mixed venous blood concentrations of sevoflurane in cardiac anaesthesia.

We have proposed an equation for estimating the real-time mixed venous blood concentration (MVBC) of isoflurane in cardiac anaesthesia. However, information related to the application of our method to sevoflurane is lacking. We studied 12 patients undergoing cardiac surgery and anaesthetised with sevoflurane. At different time points, pulmonary arterial blood samples were collected for gas chromatography-mass spectrometry (GC-MS) to determine the real mixed venous concentrations of sevoflurane. The inspired and expired concentrations of sevoflurane, measured by a gas monitor, were used for the MVBC calculations. Using Bland-Altman analyses, we found that the calculated MVBCs accurately represent the actual concentrations of sevoflurane in pulmonary arterial blood, as shown by a near-zero percentage bias with a 0.14% precision between the two concentrations. The results demonstrated that our equation could be a useful method for estimating the pulmonary blood concentration of sevoflurane.