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1.  Spatiotemporal Pattern of Neuroinflammation After Impact-Acceleration Closed Head Injury in the Rat 
Mediators of Inflammation  2006;2006(1):90123.
Inflammatory processes have been implicated in the pathogenesis of traumatic brain damage. We analyzed the spatiotemporal expression pattern of the proinflammatory key molecules: interleukin-1β, interleukin-6, tumor necrosis factor-α, and inducible nitric oxide synthase in a rat closed head injury (CHI) paradigm. 51 rats were used for RT-PCR analysis after CHI, and 18 for immunocytochemistry. We found an early upregulation of IL-1β, IL-6, and TNF-α mRNA between 1 h and 7 h after injury; the expression of iNOS mRNA only revealed a significant increase at 4 h. After 24 h, the expression decreased towards baseline levels, and remained low until 7 d after injury. Immunocytochemically, IL-1β induction was localized to ramified microglia in areas surrounding the primary impact place as well as deeper brain structures. Our study shows rapid induction of inflammatory gene expression that exceeds by far the primary impact site and might therefore contribute to tissue damage at remote sites.
PMCID: PMC1570383  PMID: 16864909
2.  The Novel Azole R126638 Is a Selective Inhibitor of Ergosterol Synthesis in Candida albicans, Trichophyton spp., and Microsporum canis 
R126638 is a novel triazole with in vitro activity similar to that of itraconazole against dermatophytes, Candida spp., and Malassezia spp. In animal models of dermatophyte infections, R126638 showed superior antifungal activity. R126638 inhibits ergosterol synthesis in Candida albicans, Trichophyton mentagrophytes, Trichophyton rubrum, and Microsporum canis at nanomolar concentrations, with 50% inhibitory concentrations (IC50s) similar to those of itraconazole. The decreased synthesis of ergosterol and the concomitant accumulation of 14α-methylsterols provide indirect evidence that R126638 inhibits the activity of CYP51 that catalyzes the oxidative removal of the 14α-methyl group of lanosterol or eburicol. The IC50s for cholesterol synthesis from acetate in human hepatoma cells were 1.4 μM for itraconazole and 3.1 μM for R126638. Compared to itraconazole (IC50 = 3.5 μM), R126638 is a poor inhibitor of the 1α-hydroxylation of 25-hydroxyvitamin D3 (IC50 > 10 μM). Micromolar concentrations of R126638 and itraconazole inhibited the 24-hydroxylation of 25-hydroxyvitamin D3 and the conversion of 1,25-dihydroxyvitamin D3 into polar metabolites. At concentrations up to 10 μM, R126638 had almost no effect on cholesterol side chain cleavage (CYP11A1), 11β-hydroxylase (CYP11B1), 17-hydroxylase and 17,20-lyase (CYP17), aromatase (CYP19), or 4-hydroxylation of all-trans retinoic acid (CYP26). At 10 μM, R126638 did not show clear inhibition of CYP1A2, CYP2A6, CYP2D6, CYP2C8, CYP2C9, CYP2C10, CYP2C19, or CYP2E1. Compared to itraconazole, R126638 had a lower interaction potential with testosterone 6β hydroxylation and cyclosporine hydroxylation, both of which are catalyzed by CYP3A4, whereas both antifungals inhibited the CYP3A4-catalyzed hydroxylation of midazolam similarly. The results suggest that R126638 has promising properties and merits further in vivo investigations for the treatment of dermatophyte and yeast infections.
PMCID: PMC514767  PMID: 15328084
3.  Hibernating myocardium: Programmed cell survival or programmed cell death? 
Evidence that programmed cell death contributes to cardiomyocyte loss is substantial for some cardiac pathologies such as myocardial infarction and a variety of cardiomyopathies. For others, such as chronic hibernating and stunned myocardium, its involvement is still debated. Recent studies have indicated that the heart remodels its structure in a rather stereotypical way when subjected to unfavourable conditions such as ischemia and pressure or volume overload. This stereotypical response is characterized by subcellular adaptations in cardiomyocytes whereby the cells switch from an adult (functional) to a fetal (survival) phenotype, a process akin to dedifferentiation. Structural hallmarks of dedifferentiation are reduction of contractile filaments, accumulation of glycogen in the cytosol, dispersion of nuclear heterochromatin, changes in mitochondrial shape and size, and loss of sarcoplasmic reticulum and T-tubules. The changes are accompanied by important alterations in the expression and distribution of structural proteins in these organelles. Today, there is only circumstantial evidence that cardiomyocyte dedifferentiation is an adaptive and reversible phenomenon instead of a degenerative event leading to apoptotic cell death. Indeed, some research groups consider the switch to a fetal phenotype to be a rescue reaction and therefore coined the name ‘programmed cell survival’, whereas others interpret this as an event on the ‘programmed cell death’ pathway. It is obvious that resolving this controversial issue is of direct clinical importance as far as prognosis and therapy are concerned.
PMCID: PMC2719170  PMID: 19649226
Apoptosis; Cell survival; Hibernating myocardium
4.  Accumulation of 3-Ketosteroids Induced by Itraconazole in Azole-Resistant Clinical Candida albicans Isolates 
Antimicrobial Agents and Chemotherapy  1999;43(11):2663-2670.
The effects of itraconazole on ergosterol biosynthesis were investigated in a series of 16 matched clinical Candida albicans isolates which had been previously analyzed for mechanisms of resistance to azoles (D. Sanglard, K. Kuchler, F. Ischer, J. L. Pagani, M. Monod, and J. Bille, Antimicrob. Agents Chemother., 39:2378–2386, 1995). Under control conditions, all isolates contained ergosterol as the predominant sterol, except two strains (C48 and C56). In isolates C48 and C56, both less susceptible to azoles than their parent, C43, substantial concentrations (20 to 30%) of 14α-methyl-ergosta-8,24(28)-diene-3β,6α-diol (3,6-diol) were found. Itraconazole treatment of C43 resulted in a dose-dependent inhibition of ergosterol biosynthesis (50% inhibitory concentration, 2 nM) and accumulation of 3,6-diol (up to 60% of the total sterols) together with eburicol, lanosterol, obtusifoliol, 14α-methyl-ergosta-5,7,22,24(28)-tetraene-3βol, and 14α-methyl-fecosterol. In strains C48 and C56, no further increase of 3,6-diol was observed after exposure to itraconazole. Ergosterol synthesis was less sensitive to itraconazole inhibition, as was expected for these azole-resistant isolates which overexpress ATP-binding cassette transporter genes CDR1 and CDR2. In addition to 3,6-diol, substantial amounts of obtusifolione were found after exposure to itraconazole. This toxic 3-ketosteroid was demonstrated previously to accumulate after itraconazole treatment in Cryptococcus neoformans and Histoplasma capsulatum but has not been reported in Candida isolates. Accumulation of obtusifolione correlated with nearly complete growth inhibition in these azole-resistant strains compared to that found in the susceptible parent strain, although the onset of growth inhibition only occurred at higher concentrations of itraconazole. ERG25 and ERG26 are the only genes assigned to the 4-demethylation process, of which the 3-ketoreductase is part. To verify whether mutations in these ERG25 genes contributed to obtusifolione accumulation, their nucleotide sequences were determined in all three related isolates. No mutations in ERG25 alleles of isolates C48 and C56 were found, suggesting that this gene is not involved in obtusifolione accumulation. The molecular basis for the accumulation of this sterol in these two strains remains to be established.
PMCID: PMC89540  PMID: 10543744

Results 1-4 (4)