Synthesis And Photochemical Properties Of Some Porphyrin Derivatives

The initial step of photosynthesis involves transfer of an electron from a photo-excited porphyrin donor to a nearby ubiquinone acceptor. Compounds containing a porphyrin covalently linked to a benzoquinone have been shown to mimic this initial transfer and as such are useful models for the study of photo-induced intramolecular electron transfer. The first chapter of this thesis describes the synthesis of a linked porphyrin-benzoquinone species, designated P4Q. This compound consists of a tetra-aryl porphyrin linked cis-1,3 across a cyclobutane ring to a benzoquinone.;The photophysical properties of this compound and of two precursor species, the hydroquinone analog and the dimethoxy phenyl analog, designated as P4QH{dollar}\sb2{dollar} and P4DMB respectively, were measured in six solvents. The results indicate that intramolecular photo-induced electron transfer is possible in P4Q, and that relative to similar species it is surprisingly facile. The relatively fast observed rates for this electron transfer are rationalized in terms of the mediating role that the connecting linkage plays. The observed solvent effect on the rate of electron transfer was found to be consistent with Marcus theory.;The second and final chapter of this thesis outlines the attempted preparation of a water soluble porphyrin-cyclodextrin species and the attempts to determine complex dissociation constants for {dollar}\beta{dollar}-cyclodextrin-electron acceptor complexes in aqueous solution. This was an attempt to extend the use of {dollar}\beta{dollar}-cyclodextrin as a connecting linkage to aqueous solution. Cyclodextrins are useful as connecting linkages due to their ability to form inclusion complexes with a wide variety of species. This allows one porphyrin derivative, that linked to the cyclodextrin, to be used for screening several potential electron acceptors with a minimum of synthetic work. The attempts to measure the equilibrium dissociation constants were part of an attempt to quantify the kinetics of electron transfer in this system

The effects of rules and instructions, consultant feedback, and teacher self-monitoring on teacher and student behavior during consulting and non-consulting periods

This study investigated the effects of rules and instructions, consultant feedback, and self-monitoring on teacher approval, disapproval, and student on-task behavior. Data was collected during a consulting period, measuring changes when the consultant was present, and during a non-consulting period, assessing whether similar changes occurred with the consultant absent. Three elementary teachers who exhibited more verbal disapproval than approval participated. On-task data was collected on three randomly selected students in each classroom. Following baseline, the teachers set classroom rules and were instructed to increase their approval and decrease disapproval. During the consulting period of the feedback phase, the consultant provided feedback every five minutes to the teacher on the frequency of her approvals and disapprovals. The teachers counted their approvals on a wrist counter during both periods of the self-monitoring phase and continued receiving feedback during the consulting period. Follow-up data was collected after the fourth phase

Electrochemical models for electrode behavior in retinal prostheses

Thesis (M.Eng.)--Massachusetts Institute of Technology, Dept. of Electrical Engineering and Computer Science, 2003.Includes bibliographical references (p. 155-164).The focus of this thesis is the modeling, characterization, and improvement of microfabricated electrodes for the Retinal Implant Project. The ultimate goal of the Project is to build a retinal prosthesis able to restore a limited degree of visual function in people suffering from certain types of blindness. An important step in this process is the design and fabrication of a safe, efficient, and effective electrode array. Designing such an array will require a detailed understanding of electrode properties and accurate models for their electrical and chemical behavior. This thesis represents a few initial steps towards that goal. Besides providing useful data on the current arrays, it is hoped that this thesis will also provide a good general introduction to electrode modeling and help others in the research group better understand the devices they are using. The thesis followed four main steps. The first step was to find an appropriate circuit model for the behavior of microfabricated electrodes in an electrolyte. After some preliminary observations, the Randles model was chosen as a convenient starting point. Several aspects of this model were discussed, including its impact on electrode design, its expected behavior using different measurement techniques, and its major limitations. The second step was to calculate experimental values for the individual elements in the model. This was done for a number of different electrode designs under various physical conditions. The data was collected using several different electrochemical measurement techniques, each of which was explained in reasonable detail. The third step was to understand the physical basis of each model parameter and find chemical or physical theories to explain and predict the observed values. This modeling work focused on the series resistance and double layer capacitance. The resistance was well fit by a recessed disk model with an additional term for the oxide film. Several important aspects of the capacitance scaling were explained by a simple model involving nonuniform current density at the electrode surface, but a great deal of work remains to be done in this area. The final step of the thesis was to suggest possible improvements on the current electrode design and point out several directions for future work.by Kenneth L. Roach.M.Eng

A microwell array cytometry system for high throughput single cell biology and bioinformatics

Thesis (Ph. D.)--Harvard-MIT Division of Health Sciences and Technology, 2009.Includes bibliographical references (p. 91-101).Recent advances in systems biology and bioinformatics have highlighted that no cell population is truly uniform and that stochastic behavior is an inherent property of many biological systems. As a result, bulk measurements can be misleading even when particular care has been taken to isolate a single cell type, and measurements averaged over multiple cell populations in a tissue can be as misleading as the average height at an elementary school. Unfortunately, there are relatively few experimental systems available at present that can provide a combination of single cell resolution, large cell populations, and the ability to track individual cells over multiple time points. Those systems that do exist are often difficult to automate and require extensive user intervention simply to generate the raw data sets for later analysis. The goal of this thesis project was to develop a powerful, inexpensive, and easy-to-use system that meets the above requirements and can serve as a platform for single cell bioinformatics. Our current system design is composed of two basic parts: 1) a customizable PDMS device consisting of one or more microwell arrays, each with associated alignment and identification features, and 2) a suite of custom software tools for automated image processing and data analysis. The system has a number of significant advantages over competing technologies such as flow cytometry and standard image cytometry. Unlike flow cytometry, the cells are not in suspension, and individual cells can be tracked across multiple time points or examined before and after a treatment.(cont.) Unlike most image cytometry approaches, the cells are arranged in a spatially defined pattern and physically separated from one another, greatly simplifying the required image analysis. The automated analysis tools require only a minimal amount of user intervention and can easily generate multi-channel fluorescence time courses for tens of thousands of individual cells in a single experiment. For visualization purposes, tools are provided to annotate the original fluorescence images or movies with the results of later analysis, and several quality control routines are available to identify improperly seeded wells or debris. The microwell array cytometry platform has allowed us to investigate a number of biological problems that would be difficult or impossible to tackle with standard techniques. Our earliest work focused on correlating pre-stress cell states with post-stress outcomes, with a major focus on the cryopreservation of primary hepatocytes. In particular, we wanted to know whether cell survival was dominated by extrinsic factors such as ice crystal nucleation, or intrinsic factors such as the energetic state of the cell. In one set of studies, we found that cells with a high initial mitochondrial content or mitochondrial membrane potential, as measured by Rh123 or JC-1 staining, were significantly less likely to survive the freezing process. This demonstrated that intrinsic cell factors do play a major role in cryopreservation survival, but perhaps more importantly demonstrated the power and versatility of the microwell system by tracking individual cells across a treatment as extreme as freezing the entire device. In another set of cryopreservation experiments, cells were transiently transfected with a GFP-tagged protective protein and the resulting cell population, with its range of expression levels, was used to generate dose response curves with single cell resolution for the protein's protective effect.(cont.) More recently, our efforts have focused on generating single cell fluorescence time courses and using bioinformatics techniques such as hierarchical and k-means clustering to visualize the data and extract interesting features. More specifically, the behavior of primary hepatocytes under oxidative stress and protective metabolic manipulation was examined using a combination of mitochondrial and free radical sensitive dyes. The resulting time courses could not only be compared between the treatment groups, but a number of distinct response patterns could be identified within each treatment group. This variation in response patterns represent potentially important information that would be missed using bulk techniques or flow cytometry. In addition, membership in each response cluster was correlated between multiple dyes and with the initial state of each cell. Using a live / dead methodology, dose response curves, survival curves, and survival time distributions were also generated for each treatment condition and further subdivided based on the initial cell state and cluster assignments. We believe that our microwell array cytometry platform will have general utility for a wide range of questions related to cell population heterogeneity, biological stochasticity, and cell behavior under stress conditions. We have really just begun exploring rich data sets of this type, and with additional work there is a great potential for groundbreaking results in many areas of biology and bioinformatics. Though we have applied techniques from gene expression analysis, there are a number of significant differences between the type of data generated by gene chips and that generated in high-throughput single cell experiments. These differences also make single cell biology a fruitful area for the development of novel bioinformatics techniques and theories.by Kenneth L. Roach.Ph.D

High-throughput single cell arrays as a novel tool in biopreservation

Microwell array cytometry is a novel high-throughput experimental technique that makes it possible to correlate pre-stress cell phenotypes and post-stress outcomes with single cell resolution. Because the cells are seeded in a high density grid of cell-sized microwells, thousands of individual cells can be tracked and imaged through manipulations as extreme as freezing or drying. Unlike flow cytometry, measurements can be made at multiple time points for the same set of cells. Unlike conventional image cytometry, image analysis is greatly simplified by arranging the cells in a spatially defined pattern and physically separating them from one another. To demonstrate the utility of microwell array cytometry in the field of biopreservation, we have used it to investigate the role of mitochondrial membrane potential in the cryopreservation of primary hepatocytes. Even with optimized cryopreservation protocols, the stress of freezing almost always leads to dysfunction or death in part of the cell population. To a large extent, cell fate is dominated by the stochastic nature of ice crystal nucleation, membrane rupture, and other biophysical processes, but natural variation in the initial cell population almost certainly plays an important and under-studied role. Understanding why some cells in a population are more likely to survive preservation will be invaluable for the development of new approaches to improve preservation yields. For this paper, primary hepatocytes were seeded in microwell array devices, imaged using the mitochondrial dyes Rh123 or JC-1, cryopreserved for up to a week, rapidly thawed, and checked for viability after a short recovery period. Cells with a high mitochondrial membrane potential before freezing were significantly less likely to survive the freezing process, though the difference in short term viability was fairly small. The results demonstrate that intrinsic cell factors do play an important role in cryopreservation survival, even in the short term where extrinsic biophysical factors would be expected to dominate. We believe that microwell array cytometry will be an important tool for a wide range of studies in biopreservation and stress biology. © 2009 Elsevier Inc. All rights reserved

Pathologic gene network rewiring implicates PPP1R3A as a central regulator in pressure overload heart failure

Heart failure is a leading cause of mortality, yet our understanding of the genetic interactions underlying this disease remains incomplete. Here, we harvest 1352 healthy and failing human hearts directly from transplant center operating rooms, and obtain genome-wide genotyping and gene expression measurements for a subset of 313. We build failing and non-failing cardiac regulatory gene networks, revealing important regulators and cardiac expression quantitative trait loci (eQTLs). PPP1R3A emerges as a regulator whose network connectivity changes significantly between health and disease. RNA sequencing after PPP1R3A knockdown validates network-based predictions, and highlights metabolic pathway regulation associated with increased cardiomyocyte size and perturbed respiratory metabolism. Mice lacking PPP1R3A are protected against pressure-overload heart failure. We present a global gene interaction map of the human heart failure transition, identify previously unreported cardiac eQTLs, and demonstrate the discovery potential of disease-specific networks through the description of PPP1R3A as a central regulator in heart failure

Portfolio Vol. IV N 1

Phillips, Alison. Reflection. Poetry. 2. Rood, John. The Accused. Picture. 2. Lay, Mary Virginia. Irony. Poetry. 6. Lay, Mary Virginia. Not Know God? Poetry. 6. Shields, Margaret. Hope. Poetry. 6. Rogers, Tom. Football and Education. Prose. 7. Hammer, John. Red? Prose. 3-4. Brown, Kenneth I. The Christmas Guest. Prose. 5. Price III, Ira. Thomas Carlyle-Political Reactionary. Prose. 8-10. Tinnerman, Betty. Greater Love Hath No Man. Prose. 11. Hall, Jim. Gentlement--To Arms! Prose. 12-13. Benson, Virginia. Drop That Hammer! Prose. 14-15. Wilson, William. Thomas Wolfe--Volcano. Prose. 16. Seagrave, Leslie. Trelawney. Prose. 16. Chester, Bob. Keeping the Records Straight. Prose. 17. Wurdman, Audrey. I, II, XLIX. Poetry. 18. Auslander, Joseph. These Are the Wounds. Poetry. 19. Auslander, Joseph. Christmas Encyclical. Poetry. 19. Auslander, Joseph. Encounter With Keats. Poetry. 19. Koncana, Jean. Backstage at the Opera House. Prose. 20. Timrud, David. Ward Seventy Tonight. Prose. 21-22. Roach, Margaret. Wedding Bells. Prose. 23-26. Close, Dean. Back Country. Picture. 6

The qualification of an enrichment biomarker for clinical trials targeting early stages of Parkinson’s disease

As therapeutic trials target early stages of Parkinson’s disease (PD), appropriate patient selection based purely on clinical criteria poses significant challenges. Members of the Critical Path for Parkinson’s Consortium formally submitted documentation to the European Medicines Agency (EMA) supporting the use of Dopamine Transporter (DAT) neuroimaging in early PD. Regulatory documents included a comprehensive literature review, a proposed analysis plan of both observational and clinical trial data, and an assessment of biomarker reproducibility and reliability. The research plan included longitudinal analysis of the Parkinson Research Examination of CEP-1347 Trial (PRECEPT) and the Parkinson’s Progression Markers Initiative (PPMI) study to estimate the degree of enrichment achieved and impact on future trials in subjects with early motor PD. The presence of reduced striatal DAT binding based on visual reads of single photon emission tomography (SPECT) scans in early motor PD subjects was an independent predictor of faster decline in UPDRS Parts II and III as compared to subjects with scans without evidence of dopaminergic deficit (SWEDD) over 24 months. The EMA issued in 2018 a full Qualification Opinion for the use of DAT as an enrichment biomarker in PD trials targeting subjects with early motor symptoms. Exclusion of SWEDD subjects in future clinical trials targeting early motor PD subjects aims to enrich clinical trial populations with idiopathic PD patients, improve statistical power, and exclude subjects who are unlikely to progress clinically from being exposed to novel test therapeutics

Risk factors for late bowel and bladder toxicities in NRG Oncology prostate cancer trials of high-risk patients: A meta-analysis of physician-rated toxicities

Purpose: A meta-analysis of sociodemographic variables and their association with late (\u3e180 days from start of radiation therapy[RT]) bowel, bladder, and clustered bowel and bladder toxicities was conducted in patients with high-risk (clinical stages T2c-T4b or Gleason score 8-10 or prostate-specific antigen level \u3e20) prostate cancer. Methods and materials: Three NRG trials (RTOG 9202, RTOG 9413, and RTOG 9406) that accrued from 1992 to 2000 were used. Late toxicities were measured with the Radiation Therapy Oncology Group Late Radiation Morbidity Scale. After controlling for study, age, Karnofsky Performance Status, and year of accrual, sociodemographic variables were added to the model for each outcome variable of interest in a stepwise fashion using the Fine-Gray regression models with an entry criterion of 0.05. Results: A total of 2432 patients were analyzed of whom most were Caucasian (76%), had a KPS score of 90 to 100 (92%), and received whole-pelvic RT+HT (67%). Of these patients, 13 % and 16% experienced late grade ≥2 bowel and bladder toxicities, respectively, and 2% and 3% experienced late grade ≥3 bowel and bladder toxicities, respectively. Late grade ≥2 clustered bowel and bladder toxicities were seen in approximately 1% of patients and late grade ≥3 clustered toxicities were seen in 2 patients ( Conclusions: Patients with high-risk prostate cancer who receive whole-pelvic RT+LT HT are more likely to have a grade ≥2 bowel toxicity than those who receive prostate-only RT. LT bowel and bladder toxicities were infrequent. Future studies will need to confirm these findings utilizing current radiation technology and patient-reported outcomes