Physicochemical properties and nuclease resistance of antisense-oligodeoxynucleotides entrapped in the core of polyion complex micelles composed of poly(ethylene glycol)-poly(L-lysine) block copolymers.

In this study, the physicochemical properties of polyion complex (PIC) micelles formed from antisense-oligodeoxynucleotides (antisense-ODN) and poly(ethylene glycol)-poly(L-lysine) block copolymers (PEG-PLL) were investigated to utilize them as a novel formulation for antisense-ODN delivery. Angular and concentration dependences of the diffusion coefficient of PIC micelles were evaluated by dynamic light scattering. Results suggested that the formed PIC micelles may have spherical shape with core-shell structure, in which the PIC core formed from antisense-ODN and PLL segment was surrounded by a PEG shell. The average radius of PIC micelles was dependent on the chain length of the PLL segment and was not influenced by the change in the length of ODN molecules at least in the range between 15 and 20 base pairs. Critical association concentration (cac) of PIC micelles was then determined from a profile of light scattering intensity versus concentration (Debye plots). Cac is ca. 0.20 mg/ml, which is low enough to ensure the micelle stability in very diluted condition as is the case with systemic injection into the blood compartment for antisense-ODN therapy. Furthermore, the stability of antisense-ODN against deoxyribonuclease I (DNase I) attack was evaluated using capillary gel electrophoresis, revealing that the complexation of antisense-ODN with PEG-PLL effectively prohibited DNase I attack. These characteristics of the PIC micelle system highlight its promising feature as ODN carrier used in the field of targeting therapy.

Correlations between bone mineral density and circulating bone metabolic markers in diabetic patients.

Diabetic osteopenia, a known chronic complication of diabetes, has been demonstrated with alterations in the calcium-vitamin D endocrine system. In order to investigate the relationship between the decrease of bone density and the altered mineral metabolism in non-insulin-dependent diabetes mellitus (NIDDM), 104 male clinical-proven NIDDM patients were studied. All patients were examined on metacarpal bone mineral density (m-BMD) by means of computed X-ray densitometry (CXD) methods. The values of the Z-score of m-BMD were significantly lower than those of age-matched controls (P<0.01). There was a positive correlation between m-BMD and serum calcium levels (P<0.01), but a negative correlation was conversely observed between m-BMD and serum intact parathyroid hormone (PTH) (P<0.01), indicating that the negative calcium balance under diabetic conditions could cause the decrease of m-BMD in NIDDM. Interestingly, since a significantly positive correlation was found between circulating levels of calcium and parathyroid hormone-related peptide (PTHrP) (P<0.05) but not PTH, it seems likely that PTHrP plays a more compensatory role on the maintenance of calcium homeostasis than PTH under diabetic conditions. Based on these observations, the CXD method would be useful in measuring the mineral density of cortical bone, and would also be beneficial to investigate underlying pathogenetic mechanism of diabetic osteopenia.

A fundamental study of a computed X-ray densitometry system for bone mineral measurement in the second metacarpal bone.

The effect of radiographic exposure factors such as the photographic film density, X-ray tube voltage, and focus-film distance on the indices of bone mineral obtained by computed X-ray densitometry (CXD), a newly developed X-ray microdensitometric system, was investigated. sigma GS/D, which corresponded to the bone mineral density divided the integrated bone mineral by the bone width of the second metacarpal bone, was influenced by these exposure factors, whereas MCI, which represented the ratio of the cortical bone to whole bone, was independent of them. Present results indicated that the focus-film distance between 85 and 115 cm, the X-ray tube voltage in 45-60 kV, and a photographic density of lower than 1.82-1.86 of the screen-film combination should be used to assess quantitatively bone mineral of the second metacarpal bone using the CXD system.

Meso-alpha,epsilon-diaminopimelate D-dehydrogenase: distribution and the reaction product.

A high activity of meso-alpha-epsilon-diaminopimelate dehydrogenase was found in extracts of Bacillus sphaericus, Brevibacterium sp., Corynebacterium glutamicum, and Proteus vulgaris among bacteria tested. B. sphaericus IFO 3525, in which the enzyme is most abundant, was chosen to study the enzyme reaction. The enzyme was not induced by the addition of meso-alpha-epsilon-diaminopimelate to the growth medium. The reaction product was isolated and identified as alpha-amino-epsilon-ketopimelate by a comparison of the properties of its 2,4-dinitrophenylhydrazone with those of an authentic sample in silica gel thin-layer chromatography, absorption, infrared and proton nuclear magnetic resonance spectrometry, and elemental analyses. The alpha-amino-epsilon-ketopimelate formed enzymatically was decarboxylated by H2O2 to yield L-alpha-aminoadipate. This suggests that the amino group with D-configuration in the substrate is oxidatively deaminated; the enzyme is a D-amino acid dehydrogenase. L-alpha-Amino-epsilon-ketopimelate undergoes spontaneous dehydration to the cyclic delta1-piperideine-2,6-dicarboxylate. The enzyme reaction is reversible, and meso-alpha-epsilon-diaminopimelate was formed in the reductive amination of L-alpha-epsilon-ketopimelate.