Hypoxia-inducible factor 1-alpha acts as a bridge factor for crosstalk between ERK1/2 and caspases in hypoxia-induced apoptosis of cementoblasts

Hypoxia-induced apoptosis of cementoblasts (OCCM-30) may be harmful to orthodontic treatment. Hypoxia-inducible factor 1-alpha (HIF-1α) mediates the biological effects during hypoxia. Little is known about the survival mechanism capable to counteract cementoblast apoptosis. We aimed to investigate the potential roles of HIF-1α, as well as the protein-protein interactions with ERK1/2, using an in-vitro model of chemical-mimicked hypoxia and adipokines. Here, OCCM-30 were co-stimulated with resistin, visfatin or ghrelin under CoCl₂-mimicked hypoxia. In-vitro investigations revealed that CoCl₂-induced hypoxia triggered activation of caspases, resulting in apoptosis dysfunction in cementoblasts. Resistin, visfatin and ghrelin promoted the phosphorylated ERK1/2 expression in OCCM-30 cells. Furthermore, these adipokines inhibited hypoxia-induced apoptosis at different degrees. These effects were reversed by pre-treatment with ERK inhibitor (FR180204). In cells treated with FR180204, HIF-1α expression was inhibited despite the presence of three adipokines. Using dominant-negative mutants of HIF-1α, we found that siHIF-1α negatively regulated the caspase-8, caspase-9 and caspase-3 gene expression. We concluded that HIF-1α acts as a bridge factor in lengthy hypoxia-induced apoptosis in an ERK1/2-dependent pathway. Gene expressions of the caspases-3, caspase-8 and caspase-9 were shown to be differentially regulated by adipokines (resistin, visfatin and ghrelin). Our study, therefore, provides evidence for the role of ERK1/2 and HIF-1α in the apoptotic response of OCCM-30 cells exposed to CoCl₂-mimicked hypoxia, providing potential new possibilities for molecular intervention in obese patients undergoing orthodontic treatment.     

Effect of various parameters on viability and growth of bacteria immobilized in sol–gel-derived silica matrices

Immobilized bacteria are being extensively used for metabolite production, biocatalysts, and biosensor construction. However, long-term viability and metabolic activity of entrapped bacteria is affected by several conditions such as their physiological state, the presence of high-osmolarity environments, porous structure and shrinkage of the matrix. The aim of this work was to evaluate the effect of various parameters on bacteria immobilized in sol–gel-derived silica matrices. With this purpose, we evaluated the stress of immobilization over bacteria cultures obtained from different growing states, the effect of cell density and bacteria capability to proliferate inside matrices. Best results to attain longer preservation times were obtained when we immobilized suspensions with an optimized bacterial number of 1 × 10⁷ cfu/gel in the presence of LB medium using aqueous silica precursors. Furthermore, the impact of osmotic stress with the subsequent intracellular trehalose accumulation and the addition of osmolites were investigated. Shorter preservation times were found for bacteria immobilized in the presence of osmolites while trehalose accumulation in stressed cells did not produce changes on entrapped bacteria viability. Finally, nutrient addition in silica matrices was studied indicating that the presence of a carbon source without the simultaneous addition of nitrogen was detrimental for immobilized E. coli. However, when both carbon and nitrogen sources were present, bacteria were able to survive longer periods of time.     

Biomonitoring of concurrent exposure to ochratoxin A and citrinin in pregnant women in Bangladesh

Ochratoxin A (OTA) and citrinin (CIT) are both nephrotoxic and teratogenic in animals, and the occurrence of these mycotoxins in food may cause adverse health effects in humans. Data on the combined exposure to these food contaminants are still scarce, especially in pregnancy. Therefore, a biomonitoring study was conducted to determine the presence of urinary biomarkers of exposure to OTA and CIT in pregnant women in Bangladesh. In total, 54 spot urine samples were collected from residents of a rural and a suburban area of the Savar region in Dhaka district for analysis of OTA and CIT urinary biomarkers by previously validated HPLC-FD and LC-MS/MS methods. Most urines were positive for OTA and CIT biomarkers, with OTA being detected in 93 % (range 0.01–0.84 ng/mL) and CIT biomarkers in 87 % (range 0.02–6.93 ng/mL) of all samples. The mean levels of OTA were different between the rural (0.06?±?0.07 ng/mL) and suburban (0.15?±?0.19 ng/mL) study participants. CIT and its metabolite dihydrocitrinone (HO-CIT) were more than twofold higher in the rural (0.42?±?1.20 and 0.55?±?1.04 ng/mL, respectively) than the suburban (CIT 0.15?±?0.13 ng/mL; HO-CIT 0.23?±?0.18 ng/mL) participants. When a provisional daily intake for CIT was calculated, it exceeded the preliminary tolerable value set by European Food Safety Authority (0.2 μg/kg/day) in 9 % of the rural participants but in none of the urban participants. Urinary biomarker levels for OTA and CIT did not show significant association with intake of certain types of food consumed by the pregnant women, although total CIT biomarker levels were considerably higher among participants who consumed more rice in a day. Overall, this study indicates a frequent co-exposure to OTA and CIT among pregnant women in Bangladesh, at levels similar to those determined recently in the general population of this country.     

Analysis of protein patterns from different organs and cell fractions of trisomy 19 mice

Summary Proteins were extracted from liver, brain, and skin of 6-day-old mice with trisomy (Ts) 19 and fractionated into solubilized cell proteins and structure-bound cell proteins. The proteins were separated by two-dimensional electrophoresis, and protein patterns were compared in the combinations Ts/normal and normal/normal. Analysis of the protein patterns revealed protein spots (variants) with densities higher (h-type) or lower (l-type) in trisomies than in normal mice. Some of these variants were found in all Ts individuals investigated for a particular protein class. These variants, termed regular Ts-variants, constituted 0.8%–1.6% of the total number of spots. The proteins of the regular Ts variants were in most cases organ-nonspecific. However, in almost all cases a given quantitative variation was expressed in only one of the three organs investigated. To explain our results, we have presented models for the control of protein levels on the basis of gene regulation. New aspects in the conception of studies on trisomies in man could be gained.     

A Novel In Vivo Vascular Imaging Approach for Hierarchical Quantification of Vasculature Using Contrast Enhanced Micro-Computed Tomography

The vasculature of body tissues is continuously subject to remodeling processes originating at the micro-vascular level. The formation of new blood vessels (angiogenesis) is essential for a number of physiological and pathophysiological processes such as tissue regeneration, tumor development and the integration of artificial tissues. There are currently no time-lapsed in vivo imaging techniques providing information on the vascular network at the capillary level in a non-destructive, three-dimensional and high-resolution fashion. This paper presents a novel imaging framework based on contrast enhanced micro-computed tomography (micro-CT) for hierarchical in vivo quantification of blood vessels in mice, ranging from largest to smallest structures. The framework combines for the first time a standard morphometric approach with densitometric analysis. Validation tests showed that the method is precise and robust. Furthermore, the framework is sensitive in detecting different perfusion levels after the implementation of a murine ischemia-reperfusion model. Correlation with both histological data and micro-CT analysis of vascular corrosion casts confirmed accuracy of the method. The newly developed time-lapsed imaging approach shows high potential for in vivo monitoring of a number of different physiological and pathological conditions in angiogenesis and vascular development.     

Über Stoffverluste bei Formol-Fixierung von menschlichem Gehirngewebe

Ohne ZusammenfassungVorgetragen auf dem II. Internationalen Kongreß für Histo- und Cytochemie, Frankfurt a. Main, vom 16.-21. 8. 64.Mit Unterstützung der Deutschen Forschungsgemeinschaft.