Expansion of the fetoplacental vasculature in late gestation is strain dependent in mice

How the fetoplacental arterial tree grows and expands during late gestational development is largely unknown. In this study, we quantified changes in arterial branching in the fetal exchange region of the mouse placenta during late gestation, when capillarization increases rapidly. We studied two commonly used mouse strains, CD1 and C57Bl/6 (B6), at embryonic days (E)13.5, 15.5, and 17.5. B6 mice differ from CD1 mice by exhibiting a blunted fetal weight gain in late gestation. We found that B6 capillarization and interhemal membrane thinning were reduced and placental hypoxia-inducible factor-1α and VEGF-A expression were higher than CD1 near term. Automated vascular segmentation of microcomputed tomography data sets revealed that the number of arterial vessels ≥50 μm remained constant during late gestation in both strains, despite large increases in downstream capillary volume quantified by stereology (+65% in B6 mice and +200% in CD1 mice). Arterial diameters expanded in both strains from E13.5 to E15.5; however, diameters continued to expand to E17.5 in B6 mice only. The diameter scaling coefficient at branch sites was near optimal (-3.0) and remained constant in CD1 mice, whereas it decreased, becoming abnormal, in B6 mice at term (-3.5 ± 0.2). Based on arterial tree geometry, resistance remained constant throughout late gestation (～0.45 mmHg·s·μl(-1)) in CD1 mice, whereas it decreased by 50% in late gestation in B6 mice. Quantification of the fetoplacental vasculature revealed significant strain-dependent differences in arterial and capillary expansion in late gestation. In both strains, enlargement of the fetoplacental arterial tree occurred primarily by increased arterial diameters with no change in segment numbers in late gestation.     

Embryonic loss due to exposure to polycyclic aromatic hydrocarbons is mediated by Bax

The high miscarriage rates observed in women smokers raises the possibility that chemicals in cigarette smoke could be detrimental to embryo development. Previous studies have established that polycyclic aromatic hydrocarbons (PAHs), transactivate the arylhydrocarbon receptor (AhR), leading to cell death. Herein we show that PAH exposure results in murine embryo cell death, acting as a potential mechanism underlying cigarette-smoking-induced pregnancy loss. Cell death was preceded by increases in Bax levels, activation of caspase-3 and decreased litter size. Chronic exposure of females to PAHs prior to conception impaired development, resulting in a higher number of resorptions. This embryonic loss could not be prevented by the disruption of Hrk, but was diminished in embryos lacking Bax. We conclude that exposure of early embryos to PAHs reduces the allocation of cells to the embryonic and placental lineages by inducing apoptosis in a Bax-dependent manner, thus compromising the developmental potential of exposed embryos.     

Zinc and cadmium 5-aminolevulinate dehydratase. Metal-dependent pH profiles

Native 5-aminolevulinic acid dehydratase contains zinc ions, which are essential for the enzymatic activity. Replacement of zinc by cadmium yielded an active enzyme whose kinetic parameters (kkat and Km) are similar to those of the zinc enzyme in the neutral pH range. However, the pH profiles of kcat and Km were different due to different pKa values. Two groups both with pKa values of 6.5 in the free zinc enzyme, but with pKa values of 7.0 in the cadmium enzyme were calculated from plots of log (kcat/Km) versus pH. On the other hand, the enzyme-substrate complex is controlled by one acidic group (zinc pKa = 6.0, cadmium pKa = 6.4) and one basis group (zinc pKa = 8.2, cadmium pKa = 7.7) as calculated from plots of log kcat versus pH. The Arrhenius plots for kcat of the two enzymes show no significant difference, the free energies of activation are 77.1 kJ/mol for the zinc and 76.8 kJ/mol for the cadmium enzyme. From this and from previous work it is concluded that the metal ions are located near the active site and influence the ionisations of essential amino acid residues. From the pH profiles of the modifying reaction and inhibition by diethylpyrocarbonate a histidinyl residue is inferred as one of the ionisable groups of the active site.     

Enhancement of UV-induced mutagenesis and sister-chromatid exchanges by nickel ions in V79 cells: evidence for inhibition of DNA repair

With regard to contradictory results concerning the mutagenicity of nickel compounds in short-term assays, especially in bacterial test systems, Chinese hamster V79 cells were used to measure mutagenicity, comutagenicity and the induction of sister-chromatid exchanges (SCEs) by NiCl2. We confirmed the induction of mutations at the HGPRT locus as well as SCEs. In addition, NiCl2 shows a pronounced comutagenic effect towards UV. When using confluent cultures or resting cells due to serum deprivation, where more time is given for repair processes, the comutagenic effect is higher compared to logarithmically growing cells (10 and 4 times, respectively, compared to twice). Hence, we attribute this enhancement in mutagenicity to inhibition of DNA repair. Also the increase in induced SCEs after combined treatment with UV and NiCl2 supports this thesis. Furthermore, NiCl2 enhances the cyto-toxicity of cis-DDP about 12-fold. Since no comutagenic effect is observed in combination with MMS, we suggest that the inhibition of DNA repair by Ni(II) applies to all DNA changes that are repaired by the 'long-patch' excision repair system. This inhibition may occur via replacement of other divalent metal ions essential in repair and regulation processes.     

Temperature dependence of activity and inhibition by lead with 5-aminolevulinic acid dehydratase

5-Aminolevulinic acid dehydratase (EC 4.2.1.24) was purified 230-fold from human erythrocytes. The effects of temperature on various properties of the isolated enzyme were studied. The maximal activity is observed at 60 degrees C. The pH optimum is shifted from pH 7.3 at 5 degrees C to pH 6.6 at 55 degrees C. The Michaelis constant increases 5-fold from 5 to 55 degrees C. Inhibition by lead is more pronounced at elevated temperatures. This effect is in contradiction to the restoration of enzyme activity by heating hemolysate after inhibition by lead in vivo, which has been described in previous reports.     

The predictive distribution of the residual variability in the linear‐fixed effects model for clinical cross‐over trials

In the linear model for cross‐over trials, with fixed subject effects and normal i.i.d. random errors, the residual variability corresponds to the intraindividual variability. While population variances are in general unknown, an estimate can be derived that follows a gamma distribution, where the scale parameter is based on the true unknown variability. This gamma distribution is often used for the sample size calculation for trial planning with the precision approach, where the aim is to achieve in the next trial a predefined precision with a given probability. But then the imprecision in the estimated residual variability or, from a Bayesian perspective, the uncertainty of the unknown variability is not taken into account. Here, we present the predictive distribution for the residual variability, and we investigate a link to the F distribution. The consequence is that in the precision approach more subjects will be necessary than with the conventional calculation. For values of the intraindividual variability that are typical of human pharmacokinetics, that is a gCV of 17–36%, we would need approximately a sixth more subjects.