Fluorescent speckle microscopy, a method to visualize the dynamics of protein assemblies in living cells

Fluorescence microscopic visualization of fluorophore-conjugated proteins that have been microinjected or expressed in living cells and have incorporated into cellular structures has yielded much information about protein localization and dynamics [1]. This approach has, however, been limited by high background fluorescence and the difficulty of detecting movement of fluorescent structures because of uniform labeling. These problems have been partially alleviated by the use of more cumbersome methods such as three-dimensional confocal microscopy, laser photobleaching and photoactivation of fluorescence [2]. We report here a method called fluorescent speckle microscopy (FSM) that uses a very low concentration of fluorescent subunits, conventional wide-field fluorescence light microscopy and digital imaging with a low-noise, cooled charged coupled device (CCD) camera. A unique feature of this method is that it reveals the assembly dynamics, movement and turnover of protein assemblies throughout the image field of view at diffraction-limited resolution. We found that FSM also significantly reduces out-of-focus fluorescence and greatly improves visibility of fluorescently labeled structures and their dynamics in thick regions of living cells. Our initial applications include the measurement of microtubule movements in mitotic spindles and actin retrograde flow in migrating cells.     

Central core disease (its clinical and laboratory diagnosis)]

In vivo mercury significantly inhibited ovarian maturation in the red swamp crayfish, Procambarus clarkii. 5-Hydroxytryptamine (5-HT) induced ovarian maturation in vivo. Cadmium and mercury inhibited this 5-HT-induced maturation. Ovarian explants incubated with mercury and either brain or muscle demonstrated significant inhibition of [14C]leucine incorporation into ovarian proteins compared to the corresponding groups incubated without mercury. In the absence of mercury the brain, which contains a gonad-stimulating hormone (GSH), induced significantly more incorporation of this amino acid than occurred in the ovaries incubated with muscle. These metals may have exerted their inhibitory effects by directly inhibiting protein synthesis in the ovaries, inhibiting 5-HT-stimulated GSH release, and preventing the ovaries from responding to this hormone.     

Dietary practices and lipid intake in relation to plasma lipid profile in Hong Kong Chinese

OBJECTIVE: To study dietary lipid intake and plasma lipid profile of the Hong Kong Chinese population as part of a territory wide survey on cardiovascular risk factors. DESIGN: Randomised age and sex stratified survey. SUBJECTS: 1010 subjects aged 25-74 y (500 men, 510 women). MEASUREMENTS: A food frequency method with food tables compiled for Hong Kong was used for nutrient quantitation, while a separate questionnaire was used to examine dietary practices. Plasma lipid profile was estimated using standard laboratory methods. RESULTS: Total calorie, fat, saturated fatty acid (SFA), poly- and mono-unsaturated fatty acid (PUFA and MUFA), and cholesterol intake were higher in men; however when adjusted for caloric intake no difference was observed. Men had lower intake of PUFA as percentage of total energy had a higher Hegsted Score compared with women. Subjects consuming beans twice or more per week had lower total cholesterol and LDL-cholesterol concentrations. Overall, the population dietary intake was close to the ideal for cardiovascular health: percentage fat not greater than 30% of the total calorie intake, saturated fat intake not greater than 10% of calories, and cholesterol less than 180 mg/1000 Kcal. CONCLUSION: The dietary pattern for Hong Kong Chinese appear to be satisfactory with respect to cardiovascular health.     

Patellar enthesopathy in childhood: a new clinical and radiographic observation

Enthesitis occurs commonly in patients with seronegative spondyloarthropathies. The patella is frequently affected by enthesopathy, but overgrowth has not been reported as a manifestation of enthesitis in pediatrics. We describe 3 patients with seronegative enthesopathy and arthropathy syndrome and patellar overgrowth.     

Calbindin-D28k is regulated by adrenal steroids in hypothalamic tissue during prenatal development

We report the case of a 42 year-old woman with amyotrophic lateral sclerosis (ALS). Neurological examination showed spastic paraparesis and muscular atrophy of the upper extremities. Increased signal intensity areas were present in the lateral corticospinal tract of the brain and cervical spinal cord on a T2-weighted image. Decreased signal intensity of the motor cortex on the T2-weighted image appeared during the course of the illness. SPECT showed hypoperfusion confined to the motor cortex. The area of increased signal intensity in the cervical spinal cord on the T2-weighted MR images extended to the anterolateral columns of the spinal cord. The area of hypoperfusion in SPECT extended to the fronto-parietal area with the progression of the disease. These changes in the MRI and SPECT findings may reflect progressive degeneration of the upper motor neurons in ALS.     

Treatment results in children with B-cell lymphomas using a modified BFM protocol

AIM: Trial of the modified protocol BFM in the treatment of children with B-cell lymphomas. MATERIALS AND METHODS: 26 children with B-cell lymphoma were treated. Of them 2 children were treated according to the program for risk 2 group and 24 children were treated as risk 3 group. RESULTS: Complete remission was achieved in 23 patients. Two children were resistant. There were 2 cases of early recurrence, 1 case of early death, 1 case of death in remission. 20 patients are in complete remission. The 4-year survival is 78%. CONCLUSION: The modified protocol BFM proved to be highly effective against B-cell lymphoma in children. Its drawback is high toxicity.     

Macroscopic agglutination test for rapid diagnosis of human leptospirosis

A commercially available slide agglutination test (SAT) for the diagnosis of human leptospirosis was evaluated by comparing it to an immunoglobulin M (IgM) enzyme-linked immunosorbent assay (ELISA) and to the microscopic agglutination test (MAT). For all 108 patients, leptospirosis was diagnosed on the basis of a fourfold or greater increase in titer by MAT (seroconversion), and all but 1 of 245 controls were MAT negative (titers, <1:100). Both SAT and the IgM ELISA failed to detect one case of infection (sensitivity, 99%). Only 3 of 145 blood donors and none of the 100 patients with other illnesses were SAT positive (specificity, 99%). The overall results were similar for the three tests; however, SAT and ELISA were statistically more sensitive as initial screening tests. For 22% of the patients, the diagnosis of leptospirosis was made earlier by SAT than by MAT. SAT detected 27 (44%) of 62 MAT-negative patients with the first serum sample. ELISA and SAT had very similar results. Follow-up of patients for 1 year after the onset of symptoms showed a decreasing rate of positivity by SAT from the third month on. The rate of positivity by ELISA decreased more slowly, to about 67% by the end of the study. By MAT all patients were persistently reactive. SAT and ELISA seem to be convenient methods for the rapid and early screening for leptospirosis and could replace the less sensitive MAT. ELISA gives less subjective results than SAT and provides information on IgM kinetics, but it can be performed only by the more sophisticated laboratories. SAT is inexpensive, can be performed more quickly and more easily than ELISA, and could be used by the less well equipped laboratories.