Postnatal heart development is characterized by critical periods of heart remodeling. In order to characterize the changes in the lipophilic fraction induced by free radicals, fatty acids and t heir oxidized products, lipofuscin -like pigments (LFP), were investigated. Fatty acids were analyz ed by gas chromatography and LFP were studied by fluorescence techniques. A fluorophore characterized by spectral methods was further resolved by HPLC. Major changes in the composition of fatty acids occurred immediately after birth and then during maturation. Fluorescence of LFP changed markedly on postnatal days 1, 4, 8, and 14, and differed from the adult animals. LFP comprise several fluorophores that were present since fetal state till adulthood. No new major fluorophores were formed during development, just the abundances of individual fluorophores have been modulated which produced changes in the shape of the spectral arrays. HPLC resolved the fluorophore with excitation maximum at 360 nm and emission maximum at 410 nm. New chromatographically distinct species appeared immediately on postnatal day 1, and then on days 30 and 60. Consumption of polyunsaturated fatty acids immediately after birth and subseque nt formation of LFP suggests that oxidative stress is involved in normal heart development., J. Wilhelm, J. Ivica, Z. Veselská, J. Uhlík, L. Vajner., and Obsahuje bibliografii
Chronic hypoxia results in hypoxic pulmonary hypertension characterized by fibrotization and muscularization of the walls of peripheral pulmonary arteries. This vessel remodeling is accompanied by an increase in the amount of lung mast cells (LMC) and the presence of small collagen cleavage products in the vessel walls. We hypothesize that hypoxia activates LMC, which release matrix metalloproteinases (MMPs) cleaving collagen and starting increased turnover of connective tissue proteins. This study was designed to determine whether in vitro hypoxia stimulates production of MMPs in rat LMC and increases their collagenolytic activity. The LMC were separated on the Percoll gradient and then were divided into two groups and cultivated for 24 h in 21 % O2 + 5 % CO2 or in 10 % O2 + 5 % CO2. Presence of the rat interstitial tissue collagenase (MMP-13) in LMC was visualized by immunohistological staining and confirmed by Western blot analysis. Total MMPs activity and tryptase activity were measured in both cultivation media and cellular extracts. Exposure to hypoxia in vitro increased the amount of cells positively labeled by anti-MMP-13 antibody as well as activities of all measured enzymes. The results therefore support the concept that LMC are an important source of increased collagenolytic activity in chronic hypoxia., H. Maxová, J. Novotná, L. Vajner, H. Tomášová, R. Vytášek, M. Vízek, L. Bačáková, V. Valoušková, T. Eliášová, J. Herget., and Obsahuje bibliografii a bibliografické odkazy
Prolonged cultivation of separated rat lung mast cells (LMC) in vitro is necessary to better investigate a possible role of LMC in different stages of tissue remodeling induced by hypoxia. Rat lung mast cells (LMC) were sepa rated using a protocol including an improved proteolytic extracti on and two subsequent density gradient separations on Ficoll-P aque PLUS and a new generation of Percoll, i.e. Percoll PLUS. Instead of usual isotonic stock Percoll solution, an alternative “asymptotically isotonic” stock solution was more successful in our density separation of LMC on Percoll PLUS. Separated cells were cultivated for six days in media including stem cell factor, interleu kins IL-3 and IL-6, and one of two alternative mixtures of antibi otics. These cultivations were performed without any contaminatio n and with only rare changes in cell size and morphology. Model co-cultivation of two allogenic fractions of LMC often caused considerable rapid changes in cell morphology and size. In contrast to these observations no or rare morphological changes were found after cultivation under hypoxic conditions. In conclusions, we modified separation on Percoll PLUS to be widely used, altered LMC separation with respect to purposes of long-lasti ng cultivation and observed some model morphological changes of LMC., J. Kubrycht ... [et al.]., and Obsahuje bibliografii a bibliografické odkazy