Quantitative characterization of postnatal growth trends in proximal pulmonary arteries in rats by phase-contrast magnetic resonance imaging

Malformations of the pulmonary arteries can increase right heart workload and result in morbidity, heart failure, and death. With the increased use of murine models to study these malformations, there is a pressing need for an accurate and noninvasive experimental technique that is capable of characterizing pulmonary arterial hemodynamics in these animals. We describe the growth trends of pulmonary arteries in 13 male Sprague-Dawley rats at 20, 36, 52, 100, and 160 days of age with the introduction of phase-contrast MRI as such a technique. PCMRI results correlated closely with cardiac output measurements by ultrasound echocardiography and with fluorescent microspheres in right-left lung flow split (flow partition). Mean flow, average cross-sectional area, distensibility, and shear rates for the right and left pulmonary arteries (RPA and LPA) were calculated. The RPA was larger and received more flow at all times than the LPA (P < 0.0001). Right-left flow split did not change significantly with age, and arterial distensibility was not significantly different between RPA and LPA, except at 160 days (P < 0.01). Shear rates were much higher for the LPA than the RPA (P < 0.0001) throughout development. The RPA and LPA showed different structure-function relationships but obeyed similar allometric scaling laws, with scaling exponents comparable to those of the main pulmonary artery. This study is the first to quantitatively describe changes in RPA and LPA flows and sizes with development and to apply phase-contrast MRI techniques to pulmonary arteries in rats.     

Inhibition by heparin of protein kinase C activation and hydroxyl radical generation in puromycin aminonucleoside treated isolated rat hepatocytes

Heparin has been reported to have many actions similar to calcium-dependent protein kinase (PKC) inhibitors. We have found that puromycin aminonucleoside (PAN) increases hydroxyl radical generation and this was prevented by H-7, a PKC inhibitor in isolated rat hepatocytes. In this study, we investigate the effect of heparin on the increased hydroxyl radical generation as well as PKC activation by PAN in isolated rat hepatocytes. To estimate the amount of hydroxyl radical generation, we measured methylguanidine (MG) and creatol which are the products from the reaction of creatinine and hydroxyl radical. Synthetic rate of MG plus creatol in isolated rat hepatocytes incubated in Krebs-Henseleit bicarbonate buffer containing creatinine and tested reagents were recorded. This rate with or without PAN was 231 ± 11 or 112 ± 5.6 nmol/g wet cells/4 h (mean ± S.E., n = 5), respectively. Heparin concentrations of 3.3, 6.6 and 10 U/ml inhibited MG plus creatol synthesis in the presence of PAN by 30, 38 and 39%, and without PAN by 8.4, 27 and 34%, respectively. Statistical significance was observed except for 3.3 U/ml without PAN. The ratio of PKC in membrane/cytoplasmic fraction, an indicator of PKC activation, increased 2.8- and 3-fold that of the 0 time after 60 and 120 min incubation with PAN while heparin at 10 U/ml almost completely suppressed this increase in the ratio of PKC. The PKC ratio of the membrane/cytoplasmic fraction obtained from hepatocytes with heparin alone or without PAN and heparin was almost unchanged during the tested period. Variation of PKC levels in membrane fraction is similar to that of PKC ratio of the membrane/cytoplasmic fraction. Increased creatol synthesis by PAN and its inhibition by heparin were observed in the same samples as those used for the PKC study.These results indicate that heparin inhibits the increase in hydroxyl radical generation induced by PAN through inhibition of PKC activation in isolated rat hepatocytes.     

TRAPPopathies: An emerging set of disorders linked to variations in the genes encoding transport protein particle (TRAPP)‐associated proteins

The movement of proteins between cellular compartments requires the orchestrated actions of many factors including Rab family GTPases, Soluble NSF Attachment protein REceptors (SNAREs) and so‐called tethering factors. One such tethering factor is called TRAnsport Protein Particle (TRAPP), and in humans, TRAPP proteins are distributed into two related complexes called TRAPP II and III. Although thought to act as a single unit within the complex, in the past few years it has become evident that some TRAPP proteins function independently of the complex. Consistent with this, variations in the genes encoding these proteins result in a spectrum of human diseases with diverse, but partially overlapping, phenotypes. This contrasts with other tethering factors such as COG, where variations in the genes that encode its subunits all result in an identical phenotype. In this review, we present an up‐to‐date summary of all the known disease‐related variations of genes encoding TRAPP‐associated proteins and the disorders linked to these variations which we now call TRAPPopathies.      

The effect of thidiazuron level on in vitro regeneration type and peroxidase profile in <Emphasis Type="Italic">Eucalyptus microtheca</Emphasis> F. Muell

The green twigs of 1-year-old Eucalyptus microtheca F. Muell seedlings were cultured on modified MS medium, supplemented with α-naphthalene acetic acid (NAA) and kinetin (Kin) hormones at 12 different concentrations. After 4 weeks, the combination of 1 mg l⁻¹ NAA + 1 mg l⁻¹ Kin induced the highest number of axillary shoots. Meanwhile, embryogenic calli were observed in media containing 4 mg l⁻¹ NAA + 0.5 mg l⁻¹ Kin, without any regeneration. The hormone treatments were followed by subculturing the twigs in different levels of thidiazuron (TDZ). The combination of 1 mg l⁻¹ NAA + 1 mg l⁻¹ Kin together with 0.01 mg l⁻¹ TDZ resulted in an increase of direct shoot, while higher amounts of TDZ led to adventitious shoot induction. Somatic embryogenesis was observed in the treatment containing 0.01 mg l⁻¹ TDZ + 4 mg l⁻¹ NAA + 0.5 mg l⁻¹Kin. The peroxidase (POD) band patterns in regenerated plantlets were investigated in order to determine the effect of different levels of TDZ on loci synthesis. A dimer locus, a tetramer locus and two epigenetic bands (a new band for NAA + Kin and the other for TDZ) were observed in the POD profiles. In case of low (0.01 mg l⁻¹ and 0.1 mg l⁻¹) levels of TDZ, one heterozygote allele was disappeared from dimer locus, while at higher TDZ levels, the dimer locus lost its stability and tetramer locus showed a high activity. Thus, POD allele patterns seems to be a feasible marker for different types of regeneration.     

Investigation of the Effects of Hydroalcoholic Solutions on Textural and Rheological Properties of Various Controlled Release Grades of Hypromellose

Hypromellose (hydroxypropyl methylcellulose, HPMC) matrices are widely used in the formulation of sustained release dosage forms. The integrity and performance of an HPMC matrix formulation depends on rapid hydration and gel formation upon ingestion. Due to the recent alert issued by the Food and Drug Administration regarding the potential negative influence of alcoholic beverages on extended release (ER) formulations, several researchers have evaluated the potential influence of hydroalcoholic media on drug release from ER dosage forms. It has been reported that HPMC matrix formulations do not show “dose dumping” in hydroalcoholic media. The purpose of this study was a fundamental investigation on the effect of hydroalcoholic solutions (0–40% v/v ethanol) on textural and rheological properties of different viscosity grades of neat HPMC, as the functional ingredient within a hydrophilic matrix. In general, hydroalcoholic solutions had little effect on gel formation and mechanical properties of hydrated compacts, while the rheological behavior of HPMC showed dependency on the ethanol content of such solutions.     

Enantioselectivity of inhibition of cytochrome P450 3A4 (CYP3A4) by ketoconazole: Testosterone and methadone as substrates

Racemic ketoconazole (KTZ) was the first orally active azole antifungal agent used in clinical practice and has become widely used in the treatment of mucosal fungal infections associated with AIDS immunosuppression and cancer chemotherapy. However, the use of KTZ has been limited because of adverse drug-drug interactions. KTZ blocks ergosterol biosynthesis by inhibiting the fungal cytochrome P450 (CYP51). KTZ is also a potent inhibitor of human cytochrome P450 3A4 (CYP3A4) enzyme, the major drug-metabolizing CYP isozyme in the human liver. We examined the enantioselective differences of KTZ in the inhibition of human CYP3A4 and in antifungal action. Dextro- and levo-KTZ exhibited modest enantioselective differences with respect to CYP3A4 inhibition of testosterone and methadone metabolism. For both substrates levo-KTZ was approximately a 2-fold more potent inhibitor. We examined the enantioselective differences in the in vitro activity of KTZ against medically relevant species of Candida and Aspergillus, as well as Cryptococcus neoformans. Overall, levo-KTZ was 2-4-fold more active than dextro-KTZ. Therefore, levo-KTZ is a more potent inhibitor of CYP3A4 and has stronger in vitro antifungal activity. Chirality 16:79-85, 2004.