Three-dimensional electron beam dose calculations

The MDAH pencil-beam algorithm developed by Hogstrom et al (1981) has been widely used in clinics for electron beam dose calculations for radiotherapy treatment planning. The primary objective of this research was to address several deficiencies of that algorithm and to develop an enhanced version. Two enhancements have been incorporated into the pencil-beam algorithm; one models fluence rather than planar fluence, and the other models the bremsstrahlung dose using measured beam data. Comparisons of the resulting calculated dose distributions with measured dose distributions for several test phantoms have been made. From these results it is concluded (1) that the fluence-based algorithm is more accurate to use for the dose calculation in an inhomogeneous slab phantom, and (2) the fluence-based calculation provides only a limited improvement to the accuracy the calculated dose in the region just downstream of the lateral edge of an inhomogeneity. The source of the latter inaccuracy is believed primarily due to assumptions made in the pencil beam's modeling of the complex phantom or patient geometry.^ A pencil-beam redefinition model was developed for the calculation of electron beam dose distributions in three dimensions. The primary aim of this redefinition model was to solve the dosimetry problem presented by deep inhomogeneities, which was the major deficiency of the enhanced version of the MDAH pencil-beam algorithm. The pencil-beam redefinition model is based on the theory of electron transport by redefining the pencil beams at each layer of the medium. The unique approach of this model is that all the physical parameters of a given pencil beam are characterized for multiple energy bins. Comparisons of the calculated dose distributions with measured dose distributions for a homogeneous water phantom and for phantoms with deep inhomogeneities have been made. From these results it is concluded that the redefinition algorithm is superior to the conventional, fluence-based, pencil-beam algorithm, especially in predicting the dose distribution downstream of a local inhomogeneity. The accuracy of this algorithm appears sufficient for clinical use, and the algorithm is structured for future expansion of the physical model if required for site specific treatment planning problems. ^

Three-dimensional structure of tropism-switching Bordetella bacteriophage.

Bacteriophage BPP-1, which infects Bordetella species, can switch its specificity by mutations to the ligand-binding surface of its major tropism-determinant protein, Mtd. This targeted mutagenesis results from the activity of a phage-encoded diversity-generating retroelement. Purified Mtd binds its receptor with low affinity, yet BPP-1 binding and infection of Bordettella cells are efficient because of high-avidity binding between phage-associated Mtd and its receptor. Here, using an integrative approach of three-dimensional (3D) structural analyses of the entire phage by cryo-electron tomography and single-prticle cryo-electron microscopy, we provide direct localization of Mtd in the phage and the structural basis of the high-avidity binding of the BPP-1 phage. Our structure shows that each BPP-1 particle has a T = 7 icosahedral head and an unusual tail apparatus consisting of a short central tail "hub," six short tail spikes, and six extended tail fibers. Subtomographic averaging of the tail fiber maps revealed a two-lobed globular structure at the distal end of each long tail fiber. Tomographic reconstructions of immuno-gold-labeled BPP-1 directly localized Mtd to these globular structures. Finally, our icosahedral reconstruction of the BPP-1 head at 7A resolution reveals an HK97-like major capsid protein stabilized by a smaller cementing protein. Our structure represents a unique bacteriophage reconstruction with its tail fibers and ligand-binding domains shown in relation to its tail apparatus. The localization of Mtd at the distal ends of the six tail fibers explains the high avidity binding of Mtd molecules to cell surfaces for initiation of infection.

Development of a SPECT -based three-dimensional treatment planner for radionuclide therapy with iodine -131

Accurate calculation of absorbed dose to target tumors and normal tissues in the body is an important requirement for establishing fundamental dose-response relationships for radioimmunotherapy. Two major obstacles have been the difficulty in obtaining an accurate patient-specific 3-D activity map in-vivo and calculating the resulting absorbed dose. This study investigated a methodology for 3-D internal dosimetry, which integrates the 3-D biodistribution of the radionuclide acquired from SPECT with a dose-point kernel convolution technique to provide the 3-D distribution of absorbed dose. Accurate SPECT images were reconstructed with appropriate methods for noise filtering, attenuation correction, and Compton scatter correction. The SPECT images were converted into activity maps using a calibration phantom. The activity map was convolved with an $\sp{131}$I dose-point kernel using a 3-D fast Fourier transform to yield a 3-D distribution of absorbed dose. The 3-D absorbed dose map was then processed to provide the absorbed dose distribution in regions of interest. This methodology can provide heterogeneous distributions of absorbed dose in volumes of any size and shape with nonuniform distributions of activity. Comparison of the activities quantitated by our SPECT methodology to true activities in an Alderson abdominal phantom (with spleen, liver, and spherical tumor) yielded errors of $-$16.3% to 4.4%. Volume quantitation errors ranged from $-$4.0 to 5.9% for volumes greater than 88 ml. The percentage differences of the average absorbed dose rates calculated by this methodology and the MIRD S-values were 9.1% for liver, 13.7% for spleen, and 0.9% for the tumor. Good agreement (percent differences were less than 8%) was found between the absorbed dose due to penetrating radiation calculated from this methodology and TLD measurement. More accurate estimates of the 3-D distribution of absorbed dose can be used as a guide in specifying the minimum activity to be administered to patients to deliver a prescribed absorbed dose to tumor without exceeding the toxicity limits of normal tissues. ^

Intact flagellar motor of Borrelia burgdorferi revealed by cryo-electron tomography: evidence for stator ring curvature and rotor/C-ring assembly flexion.

The bacterial flagellar motor is a remarkable nanomachine that provides motility through flagellar rotation. Prior structural studies have revealed the stunning complexity of the purified rotor and C-ring assemblies from flagellar motors. In this study, we used high-throughput cryo-electron tomography and image analysis of intact Borrelia burgdorferi to produce a three-dimensional (3-D) model of the in situ flagellar motor without imposing rotational symmetry. Structural details of B. burgdorferi, including a layer of outer surface proteins, were clearly visible in the resulting 3-D reconstructions. By averaging the 3-D images of approximately 1,280 flagellar motors, a approximately 3.5-nm-resolution model of the stator and rotor structures was obtained. flgI transposon mutants lacked a torus-shaped structure attached to the flagellar rod, establishing the structural location of the spirochetal P ring. Treatment of intact organisms with the nonionic detergent NP-40 resulted in dissolution of the outermost portion of the motor structure and the C ring, providing insight into the in situ arrangement of the stator and rotor structures. Structural elements associated with the stator followed the curvature of the cytoplasmic membrane. The rotor and the C ring also exhibited angular flexion, resulting in a slight narrowing of both structures in the direction perpendicular to the cell axis. These results indicate an inherent flexibility in the rotor-stator interaction. The FliG switching and energizing component likely provides much of the flexibility needed to maintain the interaction between the curved stator and the relatively symmetrical rotor/C-ring assembly during flagellar rotation.

14-3-3 zeta overexpression in early stage breast disease and tumor progression

Recent progress in diagnostic tools allows many breast cancers to be detected at an early pre-invasive stage. Thus, a better understanding of the molecular basis of early breast cancer progression is essential. 14-3-3 is a family of highly conserved and ubiquitously expressed proteins that are expressed in all eukaryotic organisms. In mammals there are seven isoforms, which bind to phosphor-serine/threonine residues regulating essential cellular processes such as signal transduction, cell cycle progression, and apoptosis. Our laboratory has discovered that a particular 14-3-3 family member, Zeta, is overexpressed in over 40% of breast tumor tissues. Furthermore, I examined the stage of breast disease in which 14-3-3ζ overexpression occurs and found that increased expression of 14-3-3ζ begins at the stage of atypical ductal hyperplasia, a very early stage of breast disease that confers increased risk for progress toward breast cancer. To determine whether 14-3-3ζ overexpression is a decisive early event in breast cancer, I overexpressed 14-3-3ζ in MCF10A cells, a non-transformed mammary epithelial cell (MEC) line and examined its impact on acini formation in a three dimensional (3D) culture model which simulates a basic unit of structure in the mammary gland. I discovered that 14-3-3ζ overexpression severely disrupted the acini architecture resulting in the disruption of polarity and luminal filling. Both are critical morphological events in the pre-neoplastic breast disease. This thesis focuses on the molecular mechanism of luminal filling. Proper lumen formation is a result of anoikis, a specific type apoptosis of cells not attached to the basement membrane. I found that 14-3-3ζ overexpression conferred a resistance to anoikis. Additionally, 14-3-3ζ overexpression in MCF10A cells and in MECs from 14-3-3ζ transgenic mice reduced expression of p53, which is known to mediate anoikis. Mechanistically, 14-3-3ζ induced hyperactivation of the PI3K/Akt pathway which led to phosphorylation and translocation of the MDM2 to the nucleus resulting in increased p53 degradation. Ectopic expression of p53 restored luminal apoptosis in 14-3-3ζ overexpressing MCF10A acini in 3D cultures. These data suggest that 14-3-3ζ overexpression is a critical event in early breast disease and down-regulation of p53 is one of the mechanisms by which 14-3-3ζ alters MEC acini structure and may increase the risk of progression to breast cancer. ^

A convolution model for energy transport in a therapeutic fast neutron beam

A three-dimensional model has been proposed that uses Monte Carlo and fast Fourier transform convolution techniques to calculate the dose distribution from a fast neutron beam. This method transports scattered neutrons and photons in the forward, lateral, and backward directions and protons, electrons, and positrons in the forward and lateral directions by convolving energy spread kernels with initial interaction available energy distributions. The primary neutron and photon spectrums have been derived from narrow beam attenuation measurements. The positions and strengths of the effective primary neutron, scattered neutron, and photon sources have been derived from dual ion chamber measurements. The size of the effective primary neutron source has been measured using a copper activation technique. Heterogeneous tissue calculations require a weighted sum of two convolutions for each component since the kernels must be invariant for FFT convolution. Comparisons between calculations and measurements were performed for several water and heterogeneous phantom geometries. ^

Quantitative MRI of spinal cord injury in a rat model: Correlative studies

Serial quantitative and correlative studies of experimental spinal cord injury (SCI) in rats were conducted using three-dimensional magnetic resonance imaging (MRI). Correlative measures included morphological histopathology, neurobehavioral measures of functional deficit, and biochemical assays for N-acetyl-aspartate (NAA), lactate, pyruvate, and ATP. A spinal cord injury device was characterized and provided a reproducible injury severity. Injuries were moderate and consistent to within $\pm$20% (standard deviation). For MRI, a three-dimensional implementation of the single spin-echo FATE (Fast optimum angle, short TE) pulse sequence was used for rapid acquisition, with a 128 x 128 x 32 (x,y,z) matrix size and a 0.21 x 0.21 x 1.5 mm resolution. These serial studies revealed a bimodal characteristic in the evolution in MRI pathology with time. Early and late phases of SCI pathology were clearly visualized in $T\sb2$-weighted MRI, and these corresponded to specific histopathological changes in the spinal cord. Centralized hypointense MRI regions correlated with evidence of hemorrhagic and necrotic tissue, while surrounding hyperintense regions represented edema or myelomalacia. Unexpectedly, $T\sb2$-weighted MRI pathology contrast at 24 hours after injury appeared to subside before peaking at 72 hours after injury. This change is likely attributable to ongoing secondary injury processes, which may alter local $T\sb2$ values or reduce the natural anisotropy of the spinal cord. MRI, functional, and histological measures all indicated that 72 hours after injury was the temporal maximum for quantitative measures of spinal cord pathology. Thereafter, significant improvement was seen only in neurobehavioral scores. Significant correlations were found between quantitated MRI pathology and histopathology. Also, NAA and lactate levels correlated with behavioral measures of the level of function deficit. Asymmetric (rostral/caudal) changes in NAA and lactate due to injury indicate that rostral and caudal segments from the injury site are affected differently by the injury. These studies indicate that volumetric quantitation of MRI pathology from $T\sb2$-weighted images may play an important role in early prediction of neurologic deficit and spinal cord pathology. The loss of $T\sb2$ contrast at 24 hours suggests MR may be able to detect certain delayed mechanisms of secondary injury which are not resolved by histopathology or other radiological modalities. Furthermore, in vivo proton magnetic resonance spectroscopy (MRS) studies of SCI may provide a valuable addition source of information about changes in regional spinal cord lactate and NAA levels, which are indicative of local metabolic and pathological changes. ^