Analyzing GFP-tagged cytoskeletal protein colocalization in human carcinoma cells

Thesis (S.B.)--Massachusetts Institute of Technology, Dept. of Mechanical Engineering, 2007.Includes bibliographical references.Cytoskeletal proteins function as dynamic, complex components involved in cellular structure and signaling. Characterizing the roles of such proteins would greatly benefit many research areas, including the study of cancer and protein-related diseases. There is currently no accurate, high throughput method of image analysis that clearly describes protein behavior within the cell. In addressing this problem, we chose to characterize proteins based on the colocalization parameter-the amount of overlap between two objects or signals. We aimed to create a single parameter that quantitatively defined colocalization yet complemented biological intuition about a complicated system. Cell culture techniques were used to transfect HeLa cells with four "marker" GFP-tagged protein constructs. Cells were fluorescently labeled in three channels-Hoechst for nucleus, Texas Red phalloidin for actin, and GFP for protein-and images were captured using Cellomics scanning microscopy. After collecting data and testing software applications, we analyzed our data with Definiens software and developed a flexible, comprehensible method of quantifying colocalization using minimal parameters.by Stephanie M. Reed.S.B

Level of Patient-Physician Agreement in Assessment of Change Following Conservative Rehabilitation for Shoulder Pain

Background Assessment of health-related status has been shown to vary between patients and physicians, although the degree of patient–physician discordance in the assessment of the change in status is unknown. Methods Ninety-nine patients with shoulder dysfunction underwent a standardized physician examination and completed several self-reported questionnaires. All patients were prescribed the same physical therapy intervention. Six weeks later, the patients returned to the physician, when self-report questionnaires were re-assessed and the Global Rating of Change (GROC) was completed by the patient. The physician completed the GROC retrospectively. To determine agreement between patient and physician, intra-class correlation (ICC) coefficient and Pearson’s r using the 15-point GROC and weighted kappa using a consolidated three-point GROC were calculated. Results Utilizing the 15-point GROC, complete agreement was observed in 37 of 99 patients (37%). ICC and Pearson’s r between patient and physician were 0.62 and 0.63, respectively. Utilizing a consolidated three-point GROC, complete agreement was observed in 76 of 99 patients (77%). Weighted kappa was 0.62. Conclusions Assessment of change reported by the patient demonstrates moderate to good agreement with physician assessment. These findings indicate that the GROC does reflect and represent similar assessment of change in health status by patients and physicians. This can aid discussion of both past treatment results and future treatment plans

Descriptive Analysis of Common Functional Limitations Identified by Patients with Shoulder Pain

Context: Recent establishment of G-codes by the US government requires therapists to report function limitations at initial evaluation. Limited information exists specific to the most common limitations in patients with shoulder pain. Objective: To describe the most commonly expressed shoulder limitations with activities and their severity/level of impairment from a patient’s perspective on the initial evaluation. Design: Descriptive. Setting: Patients reporting pain with overhead activity and seeking medical attention from one orthopedic surgeon were recruited as part of a cohort study. Patients: 176 with shoulder superior labral tear from anterior to posterior (SLAP), subacromial impingement, combined SLAP and rotator cuff, and nonspecific (female = 53, age = 41 ± 13 y; male = 123, age = 41 ± 12 y). Interventions: Data were obtained on the initial visit from the Patient-Specific Functional Scale (PSFS) questionnaire. Three researchers extracted meaningful concepts from the PSFS and linked them to the International Classification of Functioning (ICF) categories according to established ICF linking rules. Results: 176 participants yielded 765 meaningful concepts that were linked to the ICF with a 66% agreement between researchers before consensus. There were no differences between diagnoses. Of all patients, 88% reported functional limitations coded into meaningful concepts as represented by 10 ICF codes; 634 (83%) meaningful concepts were linked to the activities and participation domain while 129 (17%) were linked to the body function domain. Only 2 reported functional limitations that were considered nondefinable (nd). The overall average initial impairment score on the PSFS = 4 ± 2.5 out of 10 points. Conclusion: Meaningful concepts from the activities and participation domain were most commonly identified as functional limitations and were more prevalent than limitations from the body function domain. This information helps identify some of the most common limitations in patients with shoulder pain that therapists can use to efficiently document patient functional impairment

The role of taxonomic expertise in interpretation of metabarcoding studies

Abstract The performance of DNA metabarcoding approaches for characterizing biodiversity can be influenced by multiple factors. Here, we used morphological assessment of taxa in zooplankton samples to develop a large barcode database and to assess the congruence of taxonomic identification with metabarcoding under different conditions. We analysed taxonomic assignment of metabarcoded samples using two genetic markers (COI, 18S V1–2), two types of clustering into molecular operational taxonomic units (OTUs, ZOTUs), and three methods for taxonomic assignment (RDP Classifier, BLASTn to GenBank, BLASTn to a local barcode database). The local database includes 1042 COI and 1108 18S (SSU) barcode sequences, and we added new high-quality sequences to GenBank for both markers, including 109 contributions at the species level. The number of phyla detected and the number of taxa identified to phylum varied between a genetic marker and among the three methods used for taxonomic assignments. Blasting the metabarcodes to the local database generated multiple unique contributions to identify OTUs and ZOTUs. We argue that a multi-marker approach combined with taxonomic expertise to develop a curated, vouchered, local barcode database increases taxon detection with metabarcoding, and its potential as a tool for zooplankton biodiversity surveys

Predictors of rapid aortic root dilation and referral for aortic surgery in Marfan syndrome

Few data exist regarding predictors of rapid aortic root dilation and referral for aortic surgery in Marfan syndrome (MFS). To identify independent predictors of the rate of aortic root (AoR) dilation and referral for aortic surgery, we investigated the data from the Pediatric Heart Network randomized trial of atenolol versus losartan in young patients with MFS. Data were analyzed from the echocardiograms at 0, 12, 24, and 36months read in the core laboratory of 608 trial subjects, aged 6months to 25 years, who met original Ghent criteria and had an AoR z-score (AoRz)>3. Repeated measures linear and logistic regressions were used to determine multivariable predictors of AoR dilation. Receiver operator characteristic curves were used to determine cut-points in AoR dilation predicting referral for aortic surgery. Multivariable analysis showed rapid AoR dilation as defined by change in AoRz/year>90th percentile was associated with older age, higher sinotubular junction z-score, and atenolol use (R-2=0.01) or by change in AoR diameter (AoRd)/year>90th percentile with higher sinotubular junction z-score and non-white race (R-2=0.02). Referral for aortic root surgery was associated with higher AoRd, higher ascending aorta z-score, and higher sinotubular junction diameter:ascending aorta diameter ratio (R-2=0.17). Change in AoRz of 0.72 SD units/year had 42% sensitivity and 92% specificity and change in AoRd of 0.34cm/year had 38% sensitivity and 95% specificity for predicting referral for aortic surgery. In this cohort of young patients with MFS, no new robust predictors of rapid AoR dilation or referral for aortic root surgery were identified. Further investigation may determine whether generalized proximal aortic dilation and effacement of the sinotubular junction will allow for better risk stratification. Rate of AoR dilation cut-points had high specificity, but low sensitivity for predicting referral for aortic surgery, limiting their clinical use. Clinical Trial Number ClinicalTrials.gov number, NCT00429364

Optimized Acute Treatment of Migraine Is Associated With Greater Productivity in People With Migraine: Results From the Chronic Migraine Epidemiology and Outcomes (CaMEO) Study.

OBJECTIVE: This study aimed to ascertain whether level of optimization of acute treatment of migraine is related to work productivity across the spectrum of migraine. METHODS: Data were from the Chronic Migraine Epidemiology and Outcomes (CaMEO) Study, an internet-based longitudinal survey. Respondents with migraine who reported full-time employment and use of ≥1 acute prescription medication for migraine were included. We determined relationships among lost productive time (LPT; measured with the Migraine Disability Assessment Scale), acute treatment optimization (Migraine Treatment Optimization Questionnaire- ), and monthly headache days (MHDs). RESULTS: There was a direct relationship between LPT and MHD category. Greater acute treatment optimization was associated with lower total LPT, less absenteeism, and less presenteeism within each MHD category. CONCLUSIONS: Optimizing acute treatment for migraine may reduce LPT in people with migraine and reduce indirect costs

Attitudes toward cost-conscious care among U.S. physicians and medical students: analysis of national cross-sectional survey data by age and stage of training

Abstract Background The success of initiatives intended to increase the value of health care depends, in part, on the degree to which cost-conscious care is endorsed by current and future physicians. This study aimed to first analyze attitudes of U.S. physicians by age and then compare the attitudes of physicians and medical students. Methods A paper survey was mailed in mid-2012 to 3897 practicing physicians randomly selected from the American Medical Association Masterfile. An electronic survey was sent in early 2015 to all 5,992 students at 10 U.S. medical schools. Survey items measured attitudes toward cost-conscious care and perceived responsibility for reducing healthcare costs. Physician responses were first compared across age groups (30–40 years, 41–50 years, 51–60 years, and > 60 years) and then compared to student responses using Chi square tests and logistic regression analyses (controlling for sex). Results A total of 2,556 physicians (65%) and 3395 students (57%) responded. Physician attitudes generally did not differ by age, but differed significantly from those of students. Specifically, students were more likely than physicians to agree that cost to society should be important in treatment decisions (p < 0.001) and that physicians should sometimes deny beneficial but costly services (p < 0.001). Students were less likely to agree that it is unfair to ask physicians to be cost-conscious while prioritizing patient welfare (p < 0.001). Compared to physicians, students assigned more responsibility for reducing healthcare costs to hospitals and health systems (p < 0.001) and less responsibility to lawyers (p < 0.001) and patients (p < 0.001). Nearly all significant differences persisted after controlling for sex and when only the youngest physicians were compared to students. Conclusions Physician attitudes toward cost-conscious care are similar across age groups. However, physician attitudes differ significantly from medical students, even among the youngest physicians most proximate to students in age. Medical student responses suggest they are more accepting of cost-conscious care than physicians and attribute more responsibility for reducing costs to organizations and systems rather than individuals. This may be due to the combined effects of generational differences, new medical school curricula, students’ relative inexperience providing cost-conscious care within complex healthcare systems, and the rapidly evolving U.S. healthcare system.https://deepblue.lib.umich.edu/bitstream/2027.42/146517/1/12909_2018_Article_1388.pd