Process and formulation variables in the preparation of injectable and biodegradable magnetic microspheres

The aim of this study was to prepare biodegradable sustained release magnetite microspheres sized between 1 to 2 μm. The microspheres with or without magnetic materials were prepared by a W/O/W double emulsion solvent evaporation technique using poly(lactide-co-glycolide) (PLGA) as the biodegradable matrix forming polymer. Effects of manufacturing and formulation variables on particle size were investigated with non-magnetic microspheres. Microsphere size could be controlled by modification of homogenization speed, PLGA concentration in the oil phase, oil phase volume, solvent composition, and polyvinyl alcohol (PVA) concentration in the outer water phase. Most influential were the agitation velocity and all parameters that influence the kinematic viscosity of oil and outer water phase, specifically the type and concentration of the oil phase. The magnetic component yielding homogeneous magnetic microspheres consisted of magnetite nanoparticles of 8 nm diameter stabilized with a polyethylene glycole/polyacrylic acid (PEG/PAA) coating and a saturation magnetization of 47.8 emu/g. Non-magnetic and magnetic microspheres had very similar size, morphology, and size distribution, as shown by scanning electron microscopy. The optimized conditions yielded microspheres with 13.7 weight% of magnetite and an average diameter of 1.37 μm. Such biodegradable magnetic microspheres seem appropriate for vascular administration followed by magnetic drug targeting.     

The microcirculation of myocutaneous island flaps in pigs studied with radioactive blood volume tracers and microspheres of different sizes.

In order to further improve the understanding of hemodynamic changes in the immediate postoperative phase after elevation of myocutaneous flaps, regional blood flow and arteriovenous (A-V) shunting were measured in rectus abdominis island flaps in 8 pigs. Radioactive microspheres of two sizes (15 and 50 micron) were used. Approximately half (53.4 +/- 6 percent) of the 15-micron microspheres and one-fourth (24.1 +/- 6 percent) of the 50-micron microspheres entering the flap appeared in the venous outflow. Compared with the control area, A-V shunting was significantly increased in muscle and substantially more pronounced in skin. Nutritional blood flow, total blood flow, and vascular volume were increased in muscle and unchanged in skin and subcutis. The lowest tissue hematocrit of 7 +/- 1 percent was found in skin as compared with a central hematocrit of 35 +/- 2 percent. Tissue hematocrit in flap muscle was decreased to 17 +/- 2 percent when compared with control muscle (22 +/- 3 percent), and the mean transit time for blood was correspondingly decreased. Thus vasodilation provided increased perfusion through muscular capillaries and through A-V shunts. Shunting of 15-micron microspheres appeared to take place not only in skin, but also in subcutis and muscle, which challenges the widespread belief that A-V shunting does not occur in muscle.     

Migration and internalization of cells and polystyrene microspheres in tumor cell spheroids

The movement and internalization of ³H-labelled cells and of inert polystyrene microspheres within multicellular spheroids has been examined through histological sectioning and autoradiography. EMT6 and RIF-1 spheroids were cultured in spinner flasks for approx. 2.5 weeks. At this time, ³H-labelled cells and/or microspheres were allowed to adhere to the spheroid surface. Microspheres, ³H-labelled RIF-1 monolayer cells and ³H-labelled EMT6 monolayer cells were observed to move centripetally as a wave into EMT6 spheroids. In contrast, ³H-labelled trypsinized RIF-1 and EMT6 spheroid cells became mixed with the other non-labelled spheroid cells in homotypic RIF-1 and EMT6 spheroids, respectively. Reduction of spheroid growth by maintaining the spheroids at room temperature and by treatment with 2500 rads irradiation did not prohibit the internalization of ³H-labelled EMT6 cells and microspheres in EMT6 spheroids.     

Behavioural studies in different strains of mice and the problem of heterosis

Significant differences between strains were found for open field activity and emotionality, exploratory activity, initial reaction to shock, conditioned avoidance learning, and weight for three inbred strains (Balb/c, C57Bl and C3H), such that the heaviest strain (Balb/c) was the least active and most emotional in the open field, gave the lowest score for exploratory activity and longest reaction to shock times, and was the poorest at conditioned avoidance learning.The hybrids from crosses with C57Bl tended more towards complete dominance and overdominance than did the Balb/c × C3H hybrid, probably because the Balb/c and C3H strains have some common ancestry.Heterosis was most marked for traits involving exploration or learning. Variability of the hybrids for these traits was often lower than the parents, i.e. they showed behavioural homeostasis. For other traits variability of the hybrids tended towards the parent whose genotype was dominant for the trait.Of the non-genetic factors studied, age and litter size were found to have the greatest effect on the behavioural traits. Parity was found only to affect weight. Sex differences were found for weight and emotionality.     

Differences in aggressive behaviour between male mice (Mus musculus L.) in colonies of different sizes

Using cages of uniform size, colonies of three, four, five, nine and twelve unfamiliar male mice were established and observed over a 21-day period. A dominant mouse emerged in every colony, but in the two largest sizes five to six changes in dominance were recorded. The per capita aggression did not increase with colony size. There was a general linear decline in the number of attacks recorded in colony sizes nine and twelve whilst in the three smaller colony sizes the dominant showed an exponential decline in aggressiveness. Differences in hierarchical organization were interpreted in terms of the number of attacks shown by the dominant in relation to the number of subordinates present.     

Evaluating rodent vaginal and uterine histology in toxicity studies

Interpreting histopathology of the female rodent reproductive tract can be challenging in toxicity studies. However, diagnosis can be relatively uncomplicated with an understanding of the relationship between form and function. We describe this relationship for the rodent uterus and vagina and discuss some strategies to simplify diagnosis.     

Conformational studies of oxytocin analogues with different ring sizes. I. Circular dichroism.

Circular dichroic spectra were measured for three analogues of deamino-oxytocin of different ring sizes where the disulfide group of oxytocin is replaced by the (CH2)n group. Their backbone rings are composed of different numbers of atoms, i.e., they are nineteen, twenty and twenty-one for [1,6-aminopimelic acid]oxytocin (n = 1), [1,6-aminosuberic acid]oxytocin (n = 2) and [1,6-aminoazelaic acid]oxytocin (n = 3), respectively. The pH dependence of the circular dichroism spectra indicates that the conformation of [1,6-aminoazelaic acid]oxytocin is different from those of others and the temperature dependency reveals that the conformation of [1,6-aminopimelic acid]oxytocin is most rigid. [1,6-Aminosuberic acid]oxytocin is biologically most active among three derivatives and their biological activities are related to the conformation and internal motions of the peptide hormone analogues.     

Metomidate, etomidate and fentanyl as injectable anaesthetic agents in mice

Light surgical anaesthesia lasting 12-15 min was produced by metomidate at 50 mg/kg and by etomidate at 30 mg/kg after intraperitoneal injection. Full surgical anaesthesia lasting about 160 min was achieved after subcutaneous injection of a metomidate-fentanyl mixture (60 mg/kg: 0.06 mg/kg) and this proved superior to etomidate-fentanyl given subcutaneously or intraperitoneally. It was concluded that metomidate-fentanyl is superior to pentobarbitone and tribromoethanol as an injectable anaesthetic for mice.     

Mutation rate and predicted phenotypic target sizes in ethylnitrosourea-treated mice

Chemical mutagenesis of the mouse is ongoing in several centers around the world, with varying estimates of mutation rate and number of sites mutable to phenotype. To address these questions, we sequenced approximately 9.6 Mb of DNA from G1 progeny of ethylnitrosourea-treated mice in a large, broad-spectrum screen. We identified 10 mutations at eight unique sites, including six nonsynonymous coding substitutions. This calibrates the nucleotide mutation rate for two mutagenesis centers, implies significance criteria for positional cloning efforts, and provides working estimates of effective genetic target sizes for selected phenotypes.     

不同粒径口蹄疫病毒样颗粒-ZIF-8复合物诱导小鼠的体液免疫效果分析

为进一步提高口蹄疫(foot-and-mouthdisease,FMD)病毒样颗粒(virus-likeparticles,VLPs)疫苗的免疫效果,本研究采用仿生矿化方法,将Zn2+和2-甲基咪唑按照不同浓度配比制备了不同粒径的FMDV VLPs-沸石咪唑骨架-8 (zeolitic imidazolate framework-8, ZIF-8)复合物,以探究尺寸效应对免疫效果的影响。结果显示,成功制备出3种不同粒径的FMDV VLPs-ZIF-8,粒径分别约为70、100、1 000 nm。细胞毒性和组织病理学试验表明,3种复合物均具有良好的生物安全性。小鼠免疫试验表明,3种复合物均能明显提高中和抗体和特异性抗体水平,并且随着复合物体积的减小,其免疫效果也随之增强。本研究表明,ZIF-8包封FMDV VLPs可显著增强其免疫效果,且具有尺寸依赖性。     

Intestinal uptake of fluorescent microspheres in young and aged mice

Rhodamine B-labeled synthetic latex particles (microspheres), 1.8 micron in diameter, were administered by gavage 5 days per week to young (24 days) and aged (18 months) mice. After 25 days (19 gavages), the particles were assayed in solubilized tissues by depositing them on filters and counting under fluorescence microscopy. Aged mice exhibited significantly more fluorescent particle accumulation in Peyer's patches but significantly less in lungs than young mice. Mesenteric lymph nodes and Peyer's patch-free intestinal segments contained measurable latex, but differences between young and aged animals were not significant. Liver contained only trace amounts of latex, and spleen and kidney were latex free in both young and aged animals. Nonquantitative observations on KOH-glycerol-cleared whole Peyer's patches and slices of liver, lung, and mesenteric lymph node were similar.     

Repeatability of adaptation in experimental populations of different sizes

The degree to which evolutionary trajectories and outcomes are repeatable across independent populations depends on the relative contribution of selection, chance and history. Population size has been shown theoretically and empirically to affect the amount of variation that arises among independent populations adapting to the same environment. Here, we measure the contribution of selection, chance and history in different-sized experimental populations of the unicellular alga Chlamydomonas reinhardtii adapting to a high salt environment to determine which component of evolution is affected by population size. We find that adaptation to salt is repeatable at the fitness level in medium (N_e = 5 × 10⁴) and large (N_e = 4 × 10⁵) populations because of the large contribution of selection. Adaptation is not repeatable in small (N_e = 5 × 10³) populations because of large constraints from history. The threshold between stochastic and deterministic evolution in this case is therefore between effective population sizes of 10³ and 10⁴. Our results indicate that diversity across populations is more likely to be maintained if they are small. Experimental outcomes in large populations are likely to be robust and can inform our predictions about outcomes in similar situations.     

Competition between rotifers and cladocerans of different body sizes

Summary We conducted laboratory experiments to test the hypothesis that rotifers could coexist with small (<1.2 mm) but not large (>1.2 mm) cladocerans. Keratella cochlearis was excluded in <8 days by the large cladocerans Daphnia pulex and D. magna, probably through both interference and exploitative competition. On the other hand, K. cochlearis persisted for 8 weeks with two small cladocerans (Bosmina longirostris and Ceriodaphnia dubia) and excluded a third small cladoceran (D. ambigua). Similarly, Synchaeta oblonga coexisted with B. longirostris for >7 weeks, and K. testudo coexisted with D. ambigua for >4 weeks. Coexistence of small cladocerans and rotifers was always accompanied by suppression of one or both populations, probably primarily if not exclusively by exploitative competition for limiting food resources. These results indicate that the competitive dominance of cladocerans over rotifers decreases markedly with cladoceran body size and that factors other than body size may determine the competitive outcome between rotifers and small cladocerans. Our study provides a mechanistic explanation for a commonly observed pattern in natural zooplankton communities: planktonic rotifers often are abundant when only small cladocerans occur but typically are rare when large cladocerans are present.     

Determination of birch pollen allergens in different aerosol sizes

Allergens in fine particles may cause symptoms inallergic asthmatics. In order to assess the exposureof susceptible persons, a method to measure theallergen load in fine and coarse particles wasdeveloped.Aerosols are collected with a high-volume air samplerby multistage impaction. They are separated into fivesize classes, ranging from >10 m to <1 mand sampled on glass fibre filters. After sampling,filters are crushed into a fine powder using ahydraulic press. Allergens are then eluted on a shakerinto Tween-20-containing phosphate buffered saline.After microfiltration, the eluate is ready foranalysis with ELISA-techniques (Enzyme Linked ImmunoSorbens Assay).Two different methods are used for the analysis ofallergens: One is a sandwich-ELISA using monoclonalIgG-antibodies, the other is a competitive ELISA basedon polyclonal IgE-antibodies obtained from patientsallergic to birch pollen. Using the monoclonalantibodies information on the amount of one particularallergen (the major allergen Bet v1) is obtained. Onthe other hand the competitive ELISA using thepolyclonal IgE is much more sensitive and indicatesthe total birch pollen allergens. Data obtained duringspring 1998 show good correlation of pollen counts andallergen content in the coarse particle fractioncontaining intact pollen (>10 m). In smallersized fractions, the allergen load is often close tothe detection limit. When clearly detectable amountsof allergen are present, in the fine size fraction theallergen load shows only a weak correlation to thepollen counts and the allergen concentrations in thecoarse particle fraction.     

A comparison of rooting environments in containers of different sizes

Experiments on plants are often carried out in growth chambers or greenhouses which necessitate the use of an artificial rooting environment, though this is seldom characterized in detail. Measurements were made to compare the rooting environment in large boxes (0.25 m³) with that in small pots (0.19, 0.55 and 1.90 dm³) in naturally lit chambers. Diurnal temperature fluctuations of 14.6, 11.6 and 7.7°C occurred in the post compared with only 1.9°C in the boxes. Soil drying to a matric potential of-50 kPa was approximately 25 times faster in the pots. The mean heights of 2 year old Sitka spruce (Picea sitchensis (Bong.) Carr.) seedlings grown throughout their second growing season in the three sizes of pots were 38, 62 and 92% of the mean height of those grown in the boxes. Soil solution nutrient concentrations in the boxes were considerably increased by soil drying, an aspect which seems to have received little attention in experiments involving artificially imposed drought. An alternative system of constraining the roots of individual plants within nylon fabric bags, embedded in larger volumes of soil, to facilitate harvesting of complete root systems is described. The importance of the rooting environment in determining the outcome of physiological experiments is also briefly discussed.     

Recycling of surfactant in black and beige mice: pool sizes and kinetics

Lung disaturated phosphatidylcholine (DSPC) turnover was investigated in normal C57 black and mutant beige mice; the latter have been postulated to have microtubular defects. Turnover experiments were performed on 117 black and 74 beige mice that were assayed for lamellar body-rich fraction (LB) and alveolar lavage fluid (AF) DSPC from 0.5 to 100 h after injection of [3H]glycerol. The data were analyzed by a program that derived the best-fit rate constants for an operator-chosen compartmental model. For black mice, a simple model with bidirectional exchange of DSPC between LB and AF compartments fitted the data almost as well as more complex models. This model yielded a turnover time of 5.9 h, a biological half-life of 16 h, and recycling of AF DSPC into LB of 47%. There was some evidence to suggest that DSPC might be degraded rather than recycled as a unit. For beige mice, the DSPC turnover time was 4.8 h, and its biological half-life was 40 h. The AF DSPC pool was smaller than in black mice, but the LB pool was larger. The bidirectional flux of DSPC between AF and LB was much greater than in black mice, the percent of recycling being 85. These data do not support a microtubular defect in beige mice, but the calculations for beige mice are based on a model of questionable validity.