Recording of mitochondrial transmembrane potential and volume in cultured rat osteoclasts by confocal laser scanning microscopy

Osteoclasts are multinuclear bone-resorbing cells which contain abundant mitochondria. Morphological studies have suggested that a correlation may exist between mitochondrial concentration and bone resorption by osteoclasts. However, investigation of mitochondrial transmembrane potential (delta psi) and volume has been hampered by the difficulty in obtaining a sufficient number of osteoclasts for assessing these characteristics by flow cytometric analysis. In this study, we have used confocal laser scanning microscopy after loading the cells with Rhodamine 123 and 10-nonyl Acridine Orange to record mitochondrial delta psi and volume, respectively, in isolated rat osteoclasts cultured on bovine bone slices. Optimal staining conditions were found to be 10 micrograms ml-1 for 40 min for Rhodamine, and 1 microM for 10 min for the 10-nonyl Acridine Orange derivative. Two osteoclast populations, whose shape seemed to reflect bone resorption and migratory functions, were identified depending on their shape and on the distribution of the two dye probes. 'Round-shaped' osteoclasts had significantly higher mitochondrial delta psi and volume in the apical regions than in the basolateral portions (p < 0.00001). In contrast, mitochondrial delta psi and volume in 'irregular-shaped' osteoclasts were rather evenly distributed in both these regions (p > 0.05). Our results indicate that there is an apical polarization of mitochondria in osteoclasts corresponding to the energy demands associated with bone resorption.     

Dependence of particle fluctuation velocity on gas flow, and particle diameter in gas fluidized beds for monodispersed spheres in the Geldart B and A fluidization regimes

In this paper we present new experimental data on the steady-state, mean squared, fluctuation velocity, or granular temperature, of Geldart B polymer, glass, nickel, and stainless steel monodispersed spheres averaged over the wall of a gas fluidized bed, as a function of gas flow and sphere diameter. The granular temperature is obtained by Acoustic Shot Noise technology—namely power spectral analysis of the steady state vibrational energy of the wall excited by random sphere impact, and calibrated by hammer excitation over the wall. The new data extends to polymer and metallic spheres the experimental discovery of a 1996 paper of Cody et al. that the fluctuation velocity of Geldart B glass spheres when scaled to the gas superficial velocity, U_s, is inversely proportional to sphere diameter, directly proportional to a fundamental length scale, D_o^B, and is a universal function of U = (U_s / U_mf). We also demonstrate that the new data is consistent with the diameter dependence of the fluctuation velocity that can be derived from both the 1997 paper of Menon and Durian, who measured random sphere motion near the wall through the spectroscopy of scattered laser light, and the 1992 paper of Rahman and Campbell, who measured the average granular pressure of random sphere impact on a porous steel membrane. While the inverse scaling of the fluctuation velocity with sphere diameter, and the existence of a fundamental length scale for gas fluidization, D_o^B, had not been a feature of any published fundamental model, or computer simulation, of the steady state granular temperature of spheres in gas fluidized beds, we show that it is a feature of two recent dense kinetic fluidization models published in 1999, by Buyevich and Kapbasov, and Koch and Sangani. Both theories implicitly define a fundamental length scale for the fluctuation velocity, D^? = (μ_f² / ρ_p²g)^1 / 3, where ρ_p is the sphere density, μ_f is the gas viscosity, and g is the laboratory gravitational field. The new data for polymer, glass, nickel and stainless steel spheres presented in this paper, defines D_o^B = (56 ± 2)D^?. We use the Anderson-Jackson stability model to show that the length scale D_o^B, also defines a stability length scale, such that for D < D_o^B(D > D_o^B), the uniform dense phase of the fluidized bed is stable (unstable), against one dimensional, first order fluctuations in sphere concentration. The length scale, D_o^B is thus the theoretical equivalent to the empirical scaling length introduced by Geldart, D_B/A, to distinguish spheres (D > D_B/A) that bubble at fluidization, from spheres (D < D_B/A) that fluidize before bubbling. Finally, we present new experimental data, on the remarkable changes in the granular temperature, bed expansion, and bed collapse time, between Geldart B and Geldart A monodispersed glass spheres, and compare that data to granular temperature, and bed expansion, for Geldart A rough, non-spherical, log-normal dispersed diameter catalytic particles.     

Modeling genetic and environmental influences in the etiology of conduct disorder: A study of 2,682 adult twin pairs.

The etiology of conduct disorder (CD) was examined retrospectively in a sample of 2,682 male, female, and unlike-sex adult twin pairs from the community-based Australian Twin Register. Model-fitting analyses indicated a substantial genetic influence on risk for CD, accounting for 71% of the variance (95% confidence interval [CI]?=?32–79%). There was not a statistically significant effect of the shared environment in the best-fitting model of CD, but a modest effect of the shared environment on the risk for CD could not be rejected (95% CI?=?0-32%). The magnitude of genetic and environmental influences for CD liability did not vary significantly for boys and girls, and the specific genetic and environmental mechanisms important for the development of CD appeared to be largely the same for both sexes. The fit of a multiple-threshold model raises the possibility that CD may not necessarily be a discrete entity but rather an extreme of the normal variation in conduct-disordered behavior found in the general population. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Interdialytic weight gain and nutritional parameters in chronic hemodialysis patients

The extent of interdialytic weight gain (IDWG) in chronic hemodialysis patients is usually attributed to the level of compliance with fluid restriction. However, in view of the substantial water content of food (and caloric content of beverages), IDWG also may be a function of calorie and protein intake and may reflect the nutritional state of patients. To investigate this theory, the relationship between 2-day IDWG and body weight, normalized protein catabolic rate (nPCR), serum albumin, and delivered Kt/V urea was assessed in a prospective, randomized study of 860 chronic hemodialysis patients in 56 dialysis units. Compared with patients having < 2 kg IDWG (n = 378), patients with > 3 kg IDWG (n = 138) weighed more (dry weight, 76.8 v 61.7 kg), had higher nPCR (1.15 v 0.96 g/kg/d), and had higher serum albumin levels (3.96 vs 3.79 g/dL) (all P < 0.001) but did not have different levels of Kt/V (1.04 v 1.06). When IDWG was assessed as a function of dry weight, patients with IDWG > 4.5% of dry weight (n = 151) had higher nPCR (1.17 v 0.94 g/kg/d) but weighed less (60.1 v 70.0 kg) and had a higher Kt/V (1.14 v 1.01) than patients with IDWG < 3% of dry weight (n = 355) (all P < 0.001). Artifactual association between IDWG and nPCR attributable to an accentuated two-pool effect from differing ultrafiltration requirements was unlikely as assessed by the relationship between modeled Kt/V and prescribed Kt/V determined using an anthropometric urea volume.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production of monoclonal antibodies using a secretion capture report web

We describe a novel method for the production of monoclonal antibodies using a secretion capture report web (SCRW). Following HAT selection in bulk culture, individual hybridomas are encapsulated in biotinylated agarose drops. Antibody secreted by the hybridoma is captured within the agarose drop using an avidin bridge and biotinylated anti-mouse immunoglobulin. The secreted antibody is detected by a fluorescent reporter which can be either a second anti-mouse antibody or an antigen. The binding of the reporter can be quantitated and the desired hybridoma directly cloned by flow cytometry. Multiparameter (i.e., two-color) reporter analysis can also be used to selectively enrich and clone rare hybridomas secreting antibodies directed to unique epitopes. The method allows the characterization of thousands of clones per second and the isolation of hundreds of clones per day.