A Pragmatic, Investigator-Driven Process for Disclosure of Amyloid PET Scan Results to ADNI-4 Research Participants.

BACKGROUND: Prior studies of Alzheimer's disease (AD) biomarker disclosure have answered important questions about individuals' safety after learning and comprehending their amyloid PET results; however, these studies have typically employed highly structured disclosure protocols and focused on the psychological impact of disclosure (e.g., anxiety, depression, and suicidality) in homogeneous populations. More work is needed to develop flexible disclosure protocols and study outcomes in ethnoculturally representative samples. METHODS: The Alzheimer's Disease Neuroimaging Initiative (ADNI) is formally incorporating amyloid PET disclosure into the newest protocol (ADNI-4). Participants across the cognitive spectrum who wish to know their amyloid PET results may learn them. The pragmatic disclosure process spans four timepoints: (1) a pre-disclosure visit, (2) the PET scan and its read, (3) a disclosure visit, and (4) a post-disclosure check-in. This process applies to all participants, with slight modifications to account for their cognitive status. In designing this process, special emphasis was placed on utilizing investigator discretion. Participant measures include perceived risk of dementia, purpose in life, and disclosure satisfaction. Investigator assessment of the disclosure visit (e.g., challenges encountered, topics discussed, etc.) is also included. RESULTS: Data collection is ongoing. Results will allow for more robust characterization of the impact of learning amyloid PET results on individuals and describe the perspectives of investigators. CONCLUSION: The pragmatic design of the disclosure process in ADNI-4 coupled with the novel participant and investigator data will inform future disclosure practices. This is especially important as disclosure of biomarker results expands in research and care.

Multi-vendor multi-site quantitative MRI analysis of cartilage degeneration 10 Years after anterior cruciate ligament reconstruction: MOON-MRI protocol and preliminary results.

OBJECTIVE: To describe the protocol of a multi-vendor, multi-site quantitative MRI study for knee post-traumatic osteoarthritis (PTOA), and to present preliminary results of cartilage degeneration using MR T1ρ and T2 imaging 10 years after anterior cruciate ligament reconstruction (ACLR). DESIGN: This study involves three sites and two MR platforms. The patients are from a nested cohort (termed as Onsite cohort) within the Multicenter Orthopaedic Outcomes Network (MOON) cohort 10 years after ACLR. Phantoms and controls were scanned for evaluating reproducibility. Cartilage was automatically segmented, and T1ρ and T2 were compared between operated, contralateral, and control knees. RESULTS: Sixty-eight ACL-reconstructed patients and 20 healthy controls were included. In phantoms, the intra-site coefficients of variation (CVs) of repeated scans ranged 1.8-2.1% for T1ρ and 1.3-1.7% for T2. The inter-site CVs ranged 1.6-2.1% for T1ρ and 1.1-1.4% for T2. In human subjects, the intra-site scan/rescan CVs ranged 2.2-3.5% for T1ρ and 2.6-4.9% for T2 for the six major compartments. In patients, operated knees showed significantly higher T1ρ and T2 values mainly in medial femoral condyle, medial tibia and trochlear cartilage compared with contralateral knees, and showed significantly higer T1ρ and T2 values in all six compartments compared to healthy control knees. The patient contralateral knees showed higher T1ρ and T2 values mainly in the lateral femoral condyle, lateral tibia, trochlear, and patellar cartilage compared to healthy control knees. CONCLUSION: A platform and workflow with rigorous quality control has been established for a multi-vendor multi-site quantitative MRI study in evaluating PTOA 10 years after ACLR. Our preliminary report suggests significant cartilage matrix changes in both operated and contralateral knees compared with healthy control knees.

Nucleation of Dislocations in 3.9 nm Nanocrystals at High Pressure.

As circuitry approaches single nanometer length scales, it has become important to predict the stability of single nanometer-sized metals. The behavior of metals at larger scales can be predicted based on the behavior of dislocations, but it is unclear if dislocations can form and be sustained at single nanometer dimensions. Here, we report the formation of dislocations within individual 3.9 nm Au nanocrystals under nonhydrostatic pressure in a diamond anvil cell. We used a combination of x-ray diffraction, optical absorbance spectroscopy, and molecular dynamics simulation to characterize the defects that are formed, which were found to be surface-nucleated partial dislocations. These results indicate that dislocations are still active at single nanometer length scales and can lead to permanent plasticity.

Simulations on the influence of myelin water in diffusion-weighted imaging.

While myelinated axons present an important barrier to water diffusion, many models used to interpret DWI signal neglect other potential influences of myelin. In this work, Monte Carlo simulations were used to test the sensitivity of DWI results to the diffusive properties of water within myelin. Within these simulations, the apparent diffusion coefficient (D app) varied slowly over several orders of magnitude of the coefficient of myelin water diffusion (D m), but exhibited important differences compared to D app values simulated that neglect D m (=0). Compared to D app, the apparent diffusion kurtosis (K app) was generally more sensitive to D m. Simulations also tested the sensitivity of D app and K app to the amount of myelin present. Unique variations in D app and K app caused by differences in the myelin volume fraction were diminished when myelin water diffusion was included. Also, expected trends in D app and K app with experimental echo time were reduced or inverted when accounting for myelin water diffusion, and these reduced/inverted trends were seen experimentally in ex vivo rat brain DWI experiments. In general, myelin water has the potential to subtly influence DWI results and bias models of DWI that neglect these components of white matter.

What lies beneath? Assessment of leg ulcers during acute hospital admission.

Chronic leg ulceration is a common condition often noted in patients during an acute hospital admission. We present the case of a patient in whom thorough examination and investigation of an incidentally noted ulcer revealed a serious, previously unexpected diagnosis of disseminated Merkel cell carcinoma. This article illustrates how important it is that medical staff are aware of the different patterns of an ulcer disease and are alert to atypical appearances. Acute admission, regardless of cause, represents an opportunity for full examination of all ulcers with a view to further investigation or specialist referral if needed. Such assessment can support the often overburdened community services and ensure appropriate investigation and treatment, particularly in the context of detecting malignancy.

Therapeutic considerations in the treatment of obesity hypertension.

Obesity, now recognized as an independent risk factor for cardiovascular disease, is closely associated with hypertension. Complex mechanisms link increasing body weight with increasing blood pressure. Treatment of the obese patient with hypertension requires consideration of physiologic changes related to obesity hypertension. Lifestyle modification, including weight reduction and increased physical activity, can directly influence blood pressure levels and improve blood pressure control in obese, hypertensive patients. Clinical trials are needed to determine the most effective antihypertensive drugs for the obese, hypertensive patient. Antiobesity drugs offer viable adjunctive pharmacotherapy for obesity hypertension, but additional long-term studies are needed to support their safety and efficacy.

Sorting of bacteria.

This unit provides detailed instructions for the sorting of organisms using the cell sorter. These techniques are very valuable for establishing pure cultures of organisms, for example those expressing GFP. The unit provides information on the culture, sorting, reculture, and verification of the sort purity and GFP expression. For those venturing into the area of bacterial sorting, this unit is a must.

Antigen-specific proliferation of porcine CD8alphaalpha cells to an extracellular bacterial pathogen.

A vaccine inducing protective immunity to a spirochaete-induced colitis of pigs predominantly stimulates expansion of CD8+ cells in vivo and in antigen-stimulated lymphocyte cultures. CD8+ cells, however, are rarely considered necessary for protection against extracellular bacterial pathogens. In the present study, pigs recovering from colitis resulting from experimental infection with Brachyspira (Serpulina) hyodysenteriae had increased percentages of peripheral blood CD4- CD8+ (alphaalpha-expressing) cells compared with non-infected pigs. CD8alphaalpha+ cells proliferated in antigen-stimulated cultures of peripheral blood mononuclear cells from B. hyodysenteriae-vaccinated pigs. Proliferating CD8alphaalpha+ cells consisted of CD4-, CD4+ and gammadelta T-cell receptor-positive cells. CD4- CD8alphabeta+ cells from vaccinated or infected pigs did not proliferate upon in vitro antigen stimulation. Of the CD8alphaalpha cells that had proliferated, flow cytometric analysis indicated that the majority of the CD4+ CD8+ cells were large (i.e. lymphoblasts) whereas the CD4- CD8+ cells were predominantly small. Addition of monoclonal antibodies (mAb) specific for either porcine major histocompatibility complex (MHC) class I or class II antigens diminished B. hyodysenteriae-specific proliferative responses whereas addition of mAb to porcine MHC II, but not porcine MHC I, reduced the CD8alphaalpha response. In vitro depletion of CD4+ cells by flow cytometric cell sorting diminished, but did not completely abrogate, the proliferative response of cells from vaccinated pigs to B. hyodysenteriae antigen stimulation. These results suggest that CD8alphaalpha cells are involved in recovery and possibly protection from a spirochaete-induced colitis of pigs; yet, this response appears to be partially dependent upon CD4+ cells.

Flow cytometric analysis of canine colonic mucosal lymphocytes from endoscopically obtained biopsy specimens.

OBJECTIVE: To validate use of canine colonic biopsy specimens obtained via endoscopy as a source of mucosal lymphocytes (ML) for flow cytometric analysis. SAMPLE POPULATION: Mucosal biopsy specimens from 10 adult dogs. PROCEDURE: Mucosal lymphocyte subsets obtained from excised colon were compared with ML subsets obtained from biopsy specimens obtained by use of an endoscopic forceps (6 dogs). Endoscopic colonic biopsy specimens from 4 other dogs were used to define whether obtained ML were predominantly of intraepithelial or lamina propria origin. Mucosal lymphocytes were isolated and labeled, using commercially available monoclonal antibodies directed against canine cell surface antigens. Lymphocyte subsets (cytotoxic or helper T cells; B cells) were determined by use of flow cytometric analysis. RESULTS: A large number of viable ML was obtained after dissociation of the colonic epithelium from excised colon (45.5 + 21.5 X 10(6)) and endoscopic (7.2+/-3.4 X 10(6)) biopsy specimens. Lymphocyte subsets obtained with both methods were identical for each dog and consisted predominantly of intraepithelial lymphocytes, with some lymphocytes from the lamina propria. Collagenase digestion of excised colon also yielded a large number of viable lymphocytes from the lamina propria (56.7+/-20.4 X 10(6)), but collagenase digestion of endoscopic biopsy specimens was less rewarding. CONCLUSION AND CLINICAL RELEVANCE: A representative sample of viable intraepithelial ML is obtainable from endoscopic biopsy specimens. Flow cytometric analysis, a minimally invasive technique, can be used to study ML of client-owned animals.

Lhcb Transcription Is Coordinated with Cell Size and Chlorophyll Accumulation (Studies on Fluorescence-Activated, Cell-Sorter-Purified Single Cells from Wild-Type and immutans Arabidopsis thaliana).

To study the mechanisms that integrate pigment and chlorophyll a/b-binding apoprotein biosynthesis during light-harvesting complex II assembly, we have examined [beta]-glucuronidase (GUS) enzyme activities, chlorophyll contents, and cell sizes in fluorescence-activated, cell-sorting-separated single cells from transgenic Arabidopsis thaliana wild-type and immutans variegation mutant plants that express an Lhcb (photosystem II chlorophyll a/b-binding polypeptide gene)/GUS promoter fusion. We found that GUS activities are positively correlated with chlorophyll content and cell size in green cells from the control and immutans plants, indicating that Lhcb gene transcription is coordinated with cell size in this species. Compared with the control plants, however, chlorophyll production is enhanced in the green cells of immutans; this may represent part of a strategy to maximize photosynthesis in the green sectors to compensate for a lack of photosynthesis in the white sectors of the mutant. Lhcb transcription is significantly higher in pure-white cells of the transgenic immutans plants than in pure-white cells from norflurazon-treated, photooxidized A. thaliana leaves. This suggests that immutans partially uncouples Lhcb transcription from its normal dependence on chlorophyll accumulation and chloroplast development. We conclude that immutans may play a role in regulating Lhcb transcription, and may be a key component in the signal transduction pathways that control chloroplast biogenesis.

Investigation of bone developmental and histopathologic changes from intraosseous infusion.

STUDY OBJECTIVE: To evaluate clinical and cellular changes of bone through the rapid growth phase of development after intraosseous infusion of hypertonic or isotonic solutions at slow or fast infusion rates in a pig model. METHODS: This was a prospective, randomized, partially blinded, comparative study using a porcine model in an urban teaching hospital laboratory with further development in a local farm environment. Sixty pigs weighing 12 to 30 kg were anesthetized and endotracheally intubated, and a no. 15 Jamshidi bone marrow needle was inserted into a front forelimb. Hypertonic (mannitol) or isotonic (saline) solutions of 8 mL/kg were infused through the intraosseous site at a rapid or slow infusion rate. Animals were observed for approximately 6 months, after which they were killed and the front forelimbs harvested for gross pathologic and histologic evaluation. RESULTS: No clinical complications were noted in any of the animal groups. No substantial histologic differences were found between the hypertonic and isotonic groups. Although gross pathologic lesions were found in 32% of the hypertonic groups and in fewer than 5% of the isotonic groups, this difference was not statistically significant. Equal bone changes were found in the slow- and rapid-infusion groups. CONCLUSION: The rate of intraosseous infusion and the osmolarity of the infused fluid did not appear to be related to any gross pathologic or histologic cellular or marrow changes or to any clinical complications in animal development in this study.

Mimosine differentially inhibits DNA replication and cell cycle progression in somatic cells compared to embryonic cells of Xenopus laevis.

The plant amino acid mimosine has been reported to block cell cycle progression and DNA replication in cultured mammalian cells, perhaps by blocking initiation. In this study, we show that mimosine does not block initiation or any other step in DNA replication in embryonic cells of Xenopus laevis. Mimosine does not block DNA replication in cell-free "cycling" extracts of Xenopus eggs, nor does it block M to S phase transition in cell-free egg extracts released from metaphase arrest. Microinjection of mimosine into 4-cell embryos had no visible effect on development during the first 3 days after fertilization. Prior to the midblastula transition, when the cell cycle consists of alternating S and M phases, neither chromosomal DNA replication nor replication of microinjected plasmid DNA were inhibited by mimosine microinjected into cleaving Xenopus embryos. Microinjection of mimosine after the midblastula transition, when large endogenous stockpiles of DNA replication components have begun to be depleted and Xenopus embryonic cells have acquired G1 and G2 phases, still did not inhibit cell cycle progression or DNA replication. In marked contrast, mimosine arrested the growth of proliferating cultured Xenopus kidney epithelial A6 cells near the G1/S boundary. We conclude that mimosine appears to block DNA replication and cell cycle progression in somatic cells, but has no apparent effect in rapidly dividing Xenopus embryonic cells.

In situ screening assay for cell viability using a dimeric cyanine nucleic acid stain.

A rapid and sensitive assay is described for the determination of cell viability of adherent and nonadherent cells that can be performed in situ in 96-well microtiter plates using fluorescence plate scanners. The assay, based on dye exclusion, utilizes a plasma membrane-impermeable, dimeric cyanine dye (YOYO-1). YOYO-1 fluoresces brightly only when bound to nucleic acids. Cells are incubated with YOYO-1, and fluorescence is measured before and after the addition of detergent, which allows the dye to enter the cells. The fluorescence before detergent treatment originates from nonviable cells that have membrane damage and take up YOYO-1. The fluorescence after detergent treatment originates from all cells in the sample. The ratio of the two fluorescence values is used as an indicator of cell viability. The cell viability results of this microplate assay closely resemble those of dye exclusion studies by flow cytometry and are similar but not identical to those of the thiazolyl blue assay, which uses a metabolic indicator of cell death. Because the assay can be performed in situ, without removing the medium, disintegrated cells, cell aggregates, and cells that stick to culture vessel walls are all included in the measurement.

Multivariate analysis of roentgenologic characteristics and risk factors for nonpalpable carcinoma of the breast.

Screening mammography improves carcinoma of the breast survival through early detection and treatment of nonpalpable, often noninvasive, carcinomas. Consideration of the roentgenologic characteristics of mammographic masses and calcifications in combination with risk factors of patients may improve the yield of spot localization breast biopsy. Risk factors solicited by questionnaire were correlated with the roentgenologic appearance and histopathologic factors of 482 specimens from spot localization breast biopsies (39 percent malignant). Masses with irregular contour (p < 0.001) and high density (p < 0.005) were associated with malignant tumors. Packed, coarse and round calcifications were associated with benignity (p < 0.001), whereas scattered, fine and mixed calcifications were associated with malignant tumors (p < 0.05). Malignant masses with smooth contour were significantly more likely to be associated with noninvasive histology. Patients found to have malignant tumors were significantly older than patients with benign lesions (58 versus 52 years, p < 0.001). Stepwise logistic regression was used to evaluate the relative usefulness of demographics and roentgenologic features in predicting benign and malignant pathology among masses and calcifications. Age and irregular contour were independent significant predictors of malignancy for masses. Age, round, coarse, packed or scattered calcifications were significant independent predictors of benignity among calcifications. The radiologist correctly predicted malignant pathology in 60 percent of the patients and benign pathology in 78 percent. Parity, age at menarche, age at first pregnancy, age at menopause, history of benign breast disease, history of carcinoma of the breast, family history of carcinoma of the breast, birth control pill exposure and smoking were not significantly associated with the diagnosis of malignant tumor. These results suggest that risk factors for carcinoma of the breast are insignificant for nonpalpable lesions because patients referred for needle localization are already selected for high risk. Roentgenologic patterns of masses are useful for predicting which lesions are invasive.

A comparative study of adolescents' perceptions of normal-speaking and dysarthric children.

The purpose of this investigation was to compare adolescents' perceptions of the nonspeech characteristics of dysarthric and normal speakers. Recordings of six three-word phrases produced by 16 speakers, eight cerebral palsied and eight normal-speaking children, were presented to 19 students for judgments of nonspeech characteristics on a semantic differential scale containing 22 bipolar adjective pairs. Results of their ratings indicated that for all 22 adjective pairs the normal speakers were judged more positively than the dysarthric speakers. Moreover, the differences in listeners' ratings between the two speaker groups were statistically significant for all adjective pairs. Implications of these findings and their comparison to the judgments of peer and adult listeners are discussed.

Flow Cytometric Analysis and Sorting of Heterodera glycines Eggs.

A nondestructive technique was developed to characterize and separate eggs of soybean cyst nematode, Heterodera glycines, by developmental stage using flow cytometry. Eggs from cysts cultured on susceptible soybean roots were suspended in 0.1% xanthan gum or 59% sucrose and loaded into either a Coulter EPICS 752 or EPICS 753 flow cytometer. Eggs were analyzed and sorted according to forward angle and 90 degrees light scatter, flow cytometric parameters that are relative measures of object size and granularity, respectively. Mature eggs containing vermiform juveniles were less granular and slightly larger than eggs in earlier stages of embryogeny, allowing for separation of mature eggs from immature eggs. The effectiveness of flow cytometric sorting was evaluated by comparing the developmental stages of subpopulations of unsorted and sorted eggs. Of a subpopulation of unsorted eggs, 62% contained vermiform juveniles, whereas 85 to 95% of sorted subpopulations of larger, less granular eggs contained vermiform juveniles. Suspending H. glycines eggs in 0.1% xanthan gum or 59% sucrose for flow cytometric analysis had no effect on subsequent egg hatch in vitro. This technique is an efficient and effective means to collect large, relatively homogeneous quantities of H. glycines eggs in early or late embryogeny, and would likely be useful for analyzing and sorting eggs of other nematode species for use in developmental, genetic, or physiological research, or for identification and collection of parasitized eggs.