Dose calculation accuracy of lung planning with a commercial IMRT treatment planning system.

The dose calculation accuracy of a commercial pencil beam IMRT planning system is evaluated by comparison with Monte Carlo calculations and measurements in an anthropomorphic phantom. The target volume is in the right lung and mediastinum and thus significant tissue inhomogeneities are present. The Monte Carlo code is an adaptation of the MCNP code and the measurements were made with TLD and film. Both the Monte Carlo code and the measurements show very good agreement with the treatment planning system except in regions where the dose is high and the electron density is low. In these regions the commercial system shows doses up to 10% higher than Monte Carlo and film. The average calculated dose for the CTV is 5% higher with the commercial system as compared to Monte Carlo.

Mutational effects at the subunit interfaces of human hemoglobin: evidence for a unique sensitivity of the T quaternary state to changes in the hinge region of the alpha 1 beta 2 interface.

A set of variant human hemoglobins, each with an Ala or Gly substitution at a single residue, has been prepared, and the kinetics of their reactions with carbon monoxide have been measured. This reaction is rate-limited by the binding of the first CO to the deoxygenated T state of the protein. The magnitudes of the effects of the mutations on CO combination vary widely, and, with the exception of beta Y145, the residues with the most significant effects on these kinetics are found in the hinge region of the alpha 1 beta 2 interface. Mixed-metal hybrids, with zinc protoporphyrin IX in place of heme on both alpha or both beta subunits, were prepared for beta W37E, beta W37A, alpha Y140G, and alpha Y140A, hinge region variants causing large kinetic changes, and for beta Y145G. Such hybrids permit measurements of the kinetics of CO binding to only the heme-containing alpha or beta subunits within the unliganded hemoglobin tetramer. Mutations at beta 37 and alpha 140 have global effects on the T state, increasing the rates of CO binding to both types of subunits. Mutation of beta Y145 has a large effect on the beta subunits in the deoxygenated T state, but very little effect on the alpha subunits. Oxygen equilibria measurements on the crystalline T state of beta W37E also indicate large affinity increases in both subunits of this variant. The overall oxygen equilibria of the variant hemoglobins in solution are sensitive to numerous variables besides the properties of the deoxygenated T state. In contrast to CO combination kinetics, the residues whose alterations cause the largest changes in overall oxygen equilibria in solution are scattered seemingly randomly within the alpha 1 beta 2 interface.

The TNF receptor family member CD30 is not essential for negative selection.

CD30 is a member of the TNF receptor superfamily that has been implicated in negative selection and some forms of peripheral tolerance. A previous study of CD30(-/-) mice in a class I-restricted H-Y TCR-transgenic mouse model showed that CD30 is essential for removal of autoreactive thymocytes. During the course of the studies of CD30 in the class II-restricted TCR-transgenic mice, we found that the absence of CD30 has no effect on negative selection. Surprisingly, we also found that the CD30 mutation does not perturb apoptosis of the autoreactive thymocytes in the class I-restricted H-Y TCR-transgenic model. The minimal role of CD30 in negative selection and other recent data are discussed.

Effects of age on balance assessment using voluntary and involuntary step tasks.

BACKGROUND: Taking a step while standing and modifying a step while walking are two strategies often used to maintain balance when balance disturbances are encountered during activities of daily living. This study investigated whether performance on an involuntary step task, which is assumed to be a surrogate for fall recovery abilities, was comparable to performance on a voluntary step task. METHODS: The performance of a voluntary and an involuntary step task was measured in healthy young adult (mean age 21 years) and healthy elderly adult (mean age 68 years) female subjects. Subjects stepped as fast as possible in the direction of a minimally destabilizing lateral waist pull (voluntary step task), or they responded naturally to a large destabilizing lateral waist pull (involuntary step task). The effects of age, task, and their interaction on the primary outcome variables of step foot liftoff time, landing time, step length, and step height were examined. RESULTS: In the voluntary step task, the older adults, compared to the young, required significantly more time to lift their foot (Young: 307 msec; Elderly: 424 msec). In the involuntary step task, the elderly were as quick as the young in lifting their foot (Young: 322 msec; Elderly: 335 msec). The young lifted their foot at about the same time for the two tasks. The elders, on the other hand, lifted their foot significantly earlier in the involuntary step task, compared to the voluntary step task (Vol: 424 msec; Invol: 335 msec). CONCLUSIONS: A voluntary step task underestimates the ability of healthy elderly adults to respond quickly when large destabilizing balance disturbances are encountered.

Comparing comfort and wearability: custom-made vs. self-adapted mouthguards.

Several studies have shown that mouthguards can reduce orofacial injuries among athletes. Comfort and wearability factors were compared among 40 high-school athletes for custom-made and self-adapted mouthguards. Both offered adequate protection, but overall the participants preferred the custom-made mouthguard.

Isolation and characterization of the triply oxidized derivative of a cross-linked hemoglobin.

Hemoglobin A, cross-linked between Lys 99 alpha 1 and Lys 99 alpha 2, was used to obtain a partially oxidized tetramer in which only one of the four hemes remains reduced. Because of the absence of dimerization, asymmetric, partially oxidized derivatives are stable. This is evidenced by the fact that eight of the ten possible oxidation states could be resolved by analytical isoelectric focusing. A triply oxidized hemoglobin population HbXL+3 was isolated whose predominant component was (alpha + alpha +, beta + beta 0). This triferric preparation was examined as a possible model for the triliganded state of ferrous HbA. The aquomet and cyanomet derivatives were characterized by their CD spectra and their kinetic reactions with carbon monoxide. CD spectra in the region of 287 nm showed no apparent change in quaternary structure upon binding ligand to the fourth, ferrous heme. The spectra of the oxy and deoxy forms of the cyanomet and aquomet derivatives of HbXL+3 differed insignificantly and were characteristic of the normal liganded state. Upon addition of inositol hexaphosphate (IHP), both the oxy and deoxy derivatives of the high-spin triaquomet species converted to the native deoxy conformation. In contrast, IHP had no such effect on the conformation of the low-spin cyanomet derivatives of HbXL+3. The kinetics of CO combination as measured by stopped-flow and flash photolysis techniques present a more complex picture. In the presence of IHP the triaquomet derivative does bind CO with rate constants indicative of the T state whether these are measured by the stopped-flow technique or by flash photolysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Spin equilibria in human methemoglobin: effects of bezafibrate and inositol hexaphosphate as measured by susceptometry and visible spectroscopy.

The effects of inositol hexaphosphate (IHP) and a second allosteric effector, bezafibrate, on the spin-state equilibria of the mixed-spin derivatives of ferric human hemoglobin A are examined. Changes in spin-state equilibrium are monitored by measuring absorption spectra in the visible region (460-700 nm) as well as by direct measurements of magnetic susceptibility by means of a superconducting fluxmeter. The addition of IHP at pH 6.5 results in a measurable shift in the spin equilibria of these derivatives toward higher spin. However, the addition of bezafibrate in the presence of IHP results in still larger shifts toward the high-spin form. The changes in the free energies of the spin-state equilibria resulting from the combination of these two effectors are similar in magnitude to that which results from the R-state to T-state transition in carp hemoglobin.

Spin equilibrium in human methemoglobin: effects of inositol hexaphosphate and bezafibrate as measured by resonance Raman spectroscopy.

The spin equilibria of several derivatives of human methemoglobin were probed by resonance Raman scattering. The intensity of lines in the Raman spectrum gives a measure of the high-spin (S = 5/2) to low-spin (S = 1/2) ratio which agrees well with the spin equilibria determined from direct magnetic susceptibility measurements. The addition of bezafibrate (BZF) to methemoglobin in the absence of organic phosphate, IHP, has very little effect on the spin equilibrium, whereas in the presence of IHP it augments the change in spin significantly. When both IHP and BZF are added to the mixed-spin derivatives (H2O, SCN-, OCN-, and NO2-) of human methemoglobin, the spin equilibrium is shifted toward higher spin by about 700 cal/mol, similar to the spin change detected in derivatives of carp methemoglobin upon addition of IHP alone. These data support a general mechanism for the allosteric transition in which a constant fraction of the cooperative energy (approximately 20%) is detected at the heme of the ferric ligand-bound forms.

Reexamination of the evidence for paramagnetism in oxy- and carbonmonoxyhemoglobins.

Experiments have been carried out in an attempt to confirm previous reports of paramagnetism in the oxy- and carbonmonoxy derivatives of human and carp hemoglobin. When care is taken to ensure complete saturation of the hemoglobins with ligand and the diamagnetic contributions of all of the buffers are carefully evaluated, these hemoglobin derivatives are found to have the same gram susceptibilities as exhibited by a set of metal free proteins, suggesting that they are fully diamagnetic.

Ligand-induced conformational changes in spin label-modified human hemoglobins and chains and their carboxypeptidase A-digested derivatives.

The reactive sulfhydryls of human adult and fetal hemoglobin and the single sulfhydryl of isolated gamma chains have been spin labeled with N-(1-oxyl-2,2,5,5-tetramethyl-3-pyrrolidinyl) iodoacetamide. Similar electron paramagnetic spectral differences between oxy- and deoxy-modified hemoglobins were observed for both these hemoglobins and for the isolated chains, indicating that ligand-induced conformational changes occur in isolated hemoglobin subunits as well as intact hemoglobin tetramers. Ligand induced changes in the reactivity of p-hydroxymercuribenzoate with the sulfhydryl groups of both intact hemoglobins and isolated subunits, observed by McDonald and Noble (1974) J. Biol. Chem. 249, 3161-3165), led them to draw a similar conclusion. Following carboxypeptidase A digestion of these modified hemoglobins and gamma chains, a procedure which specifically removes the two C-terminal residues of the beta or gamma chains, spectral differences between the liganded and unliganded spin-labeled derivatives still persisted. However, the magnitude of this difference was not only more reduced in the case of the hemoglobins than in that of the subunits but the spectra of both the oxy and deoxy derivatives of the hemoglobins were characteristic of the oxy derivative of a cooperative tetrameric hemoglobin. These findings support the premise that the COOH-terminal end of the beta or gamma chain contributes, although possibly to different extents, to the spectral differences exhibited by both the spin-labeled hemoglobins and chains.

Kinetic studies on the binding affinity of human hemoglobin for the 4th carbon monoxide molecule, L4.

L4, the affinity of hemoglobin for the 4th CO molecule, has been determined for human adult hemoglobin (HbA) as a function of pH and the presence of organic phosphates by measuring the kinetic parameters for the reaction. l'4, the rate of combination of CO with the triliganded molecule, was measured by flash photolysis while l4, the rate of CO dissociation for the ligand-saturated molecule, was measured by ligand replacement. L4 is pH-dependent and affected by 2,3-diphosphoglycerate. Additionally, this pH dependence of the high affinity state is largely eliminated by carboxypeptidase A digestion. L4 for human fetal hemoglobin (HbF) in phosphate buffers was also determined and found to be pH-dependent. These results cannot be reconciled within the framework of the two-state allosteric model. Additional structures in the conformational equilibrium due to either intermediates in the T to R transition or two or more R states must exist.