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PubMed1995

A comparison of the mutagenicity of mainstream cigarette smoke condensates from a representative sample of the U.S. cigarette market with a Kentucky reference cigarette (K1R4F).

The Salmonella mutagenicity assay has been used to investigate the mutagenicity of cigarette smoke and cigarette smoke condensate. The Kentucky reference (K1R4F) cigarette is designed to be representative of full-flavor, low-tar cigarettes sold in the U.S. and to serve as a reference standard for comparative studies on the chemistry and biological activities of cigarette smoke and condensate. The objective of this study was to determine if the mutagenicity of mainstream smoke condensate from the K1R4F, as measured by the Salmonella mutagenicity assay, is representative of the mutagenic activity of U.S. cigarettes. Mainstream smoke condensates prepared in dimethyl sulfoxide from the K1R4F and 73 brand styles (representing greater than 70% of the total U.S. cigarette market) were assayed using Salmonella typhimurium TA98 and TA100 (+S9) at concentrations of 0, 25, 50, 75, 100, 125 and 250 micrograms/plate. Revertants/mg condensate were determined by calculating the slopes of the dose-response curves using linear and nonlinear regression models. Revertants/cigarette were determined by multiplying the revertants/mg condensate by the mg condensate/cigarette. No significant differences (p > 0.05) were observed between the mean mutagenicity of U.S. market and K1R4F mainstream smoke condensates in terms of revertants/mg condensate or revertants/cigarette. Increased variability in mutagenicity was observed among the U.S. brands versus that of the K1R4F. This is not surprising since variability among the U.S. brands would be expected to have both measurement error and brand style variability while the K1R4F variability contains only the measurement error portion. These results demonstrate that the K1R4F is a representative model for the U.S. cigarette market in comparative Salmonella mutagenicity studies using mainstream smoke condensates.

Mutagenicity Tests↗

PubMed1995

Condensation and cohesion of lambda DNA in cell extracts and other media: implications for the structure and function of DNA in prokaryotes.

DNA added to concentrated extracts of Escherichia coli undergoes a reversible transition to a readily-sedimentable ('condensed') form. The transition occurs over a relatively small increment in extract concentration. The extract appears to play two roles in this transition, supplying both DNA-binding protein(s) and a crowded environment that increases protein binding and favors compact DNA conformations. The two roles of the extract are suggested by properties of fractions prepared by absorption of extracts with DNA-cellulose. The DNA-binding fraction and the DNA-nonbinding fractions from these columns are separately poorer condensing agents than the original extract, but when rejoined are similar to the original extract in the amount required for condensation. The dual role for the extract is supported by model studies of condensation with combinations of purified DNA-binding materials (protein HU or spermidine) and concentrated solutions of crowding agents (albumin or polyethylene glycol 8000); in each case, crowding agents and DNA-binding materials jointly reduce the amounts of each other required for condensation. The condensation reaction as studied in extracts or in the purified systems may be a useful approach to the forces which stabilize the compact form of DNA within the bacterial nucleoid. The effect of condensation on the reactivity of the DNA was measured by changes in the rate of cohesion between duplex DNA molecules bearing the complementary single-strand termini of lambda DNA. Condensation caused large increases in the rates of cohesion of both lambda DNA and of restriction fragments of lambda DNA bearing the cohesive termini. Cohesion products of lambda DNA made in vitro are a mixture of linear and circular aggregates, whereas those made in vivo are cyclic monomers. We suggest a simple mechanism based upon condensation at the site of viral injection which may explain this discrepancy.

Bacteriophage lambda↗

PubMed2001

Chromatin condensation and sensitivity of DNA in situ to denaturation during cell cycle and apoptosis--a confocal microscopy study.

The goal of this study was to construct high resolution 3D confocal images of regions of condensed and extended chromatin in cell nuclei and individual chromosomes. It has been shown previously that sensitivity of DNA in situ to denaturation correlates with chromatin condensation and varies during cell cycle and apoptosis. Thus, detection of DNA which was partially denatured in situ provided a means to image areas of condensed chromatin. DNA denaturation was detected using a metachromatic dye acridine orange (AO) which differentially stains single stranded (ss) and double-stranded (ds) DNA sections. Early studies of denaturability of cellular DNA utilized flow cytometry and standard fluorescence microscopy. These techniques could not reveal small local differences in DNA denaturability within cell nucleus or in individual chromosomes. For instance, it was not possible to detect the initial points of chromosome condensation in G2-phase of the division cycle or in apoptosis. In order to achieve this goal we have recently extended these studies by applying confocal microscopy. We investigated DNA denaturability in normal human fibroblasts and HL-60 leukemic cells, at different stages of cell cycle and apoptosis. Following removal of RNA and partial denaturation of DNA with acid cells were stained with AO. Green (530 nm) and red (640 nm) fluorescence (exc. 457 nm) of non-denatured and denatured DNA was imaged by confocal microscopy. Blind deconvolution was used to further improve the quality of 3D images. Photobleaching of AO fluorescence was minimized and a correction for chromatic aberration and register shift was implemented. Nuclei of interphase cells exhibited predominantly green fluorescence representing AO binding to ds DNA. Punctuate areas of red fluorescence representing AO binding to denatured DNA and most likely associated with local regions of condensed chromatin were also present in all interphase nuclei. The proportion of denatured DNA increased in cells entering mitosis. In prophase individual condensing chromosomes exhibited varied proportions of green and red fluorescence indicating different content of denatured chromatin. In some chromosomes bands of denatured and denaturation-resistant chromatin were clearly resolved. In metaphase and anaphase chromosomes exhibited red fluorescence along all length of their arms indicating the highest and uniform susceptibility to denaturation. In telophase chromosomes contained predominantly denaturation-resistant DNA again and denaturated regions were significantly less abundant. At cytokinesis some decondensing chromosomes were still resolved. At this stage almost all regions of denatured DNA were located close to nuclear envelope. These regions may correspond to pockets of heterochromatin reforming at nuclear periphery. In early apoptosis condensation of chromatin appeared to commence in several distinct regions within nucleus. Some apoptotic bodies contained condensed chromatin surrounding central regions of extended chromatin. At late stages of apoptosis the whole volume of apoptotic bodies was occupied by condensed chromatin.

Acridine Orange↗

PubMed2006

Time study of DNA condensate morphology: implications regarding the nucleation, growth, and equilibrium populations of toroids and rods.

It is well known that multivalent cations cause free DNA in solution to condense into nanometer-scale particles with toroidal and rod-like morphologies. However, it has not been shown to what degree kinetic factors (e.g., condensate nucleation) versus thermodynamic factors (e.g., DNA bending energy) determine experimentally observed relative populations of toroids and rods. It is also not clear how multimolecular DNA toroids and rods interconvert in solution. We have conducted a series of condensation studies in which DNA condensate morphology statistics were measured as a function of time and DNA structure. Here, we show that in a typical in vitro DNA condensation reaction, the relative rod population 2 min after the initiation of condensation is substantially greater than that measured after morphological equilibrium is reached (ca. 20 min). This higher population of rods at earlier time points is consistent with theoretical studies that have suggested a favorable kinetic pathway for rod nucleation. By using static DNA loops to alter the kinetics and thermodynamics of condensation, we further demonstrate that reported increases in rod populations associated with decreasing DNA length are primarily due to a change in the thermodynamics of DNA condensation, rather than a change in the kinetics of condensate nucleation or growth. The results presented also reveal that the redistribution of DNA from rods to toroids is mediated through the exchange of DNA strands with solution.

Base Sequence↗

PubMed1997

Condensation of DNA and chromatin by an SPKK-containing octapeptide repeat motif present in the C-terminus of histone H1.

Several DNA binding motifs have been described in the C-terminus of histone H1 (Churchill & Travers, 1991), of these the S/TPKK repeat (Suzuki, 1989) often occurs as a part of an octapeptide repeat of the type XTPKKXKK. We have studied in detail the DNA and chromatin condensing properties of a consensus octapeptide KSPKKAKK (8 mer) present in many histone H1 subtypes and its imperfect repeat ATPKKSTKKTPKKAKK (16 mer TPKK) as it occurs in the C-terminus of rat histone H1d. The 16 mer TPKK peptide containing two S/TPKK motifs was able to condense both rat oligonucleosomal (2-5 kbp) DNA and histone H1-depleted chromatin as revealed by circular dichroism spectroscopy. The 8 mer peptide, however, was unable to condense either the DNA or the histone H1-depleted chromatin. Both the 8 mer peptide and the 16 mer TPKK peptide displaced distamycin A from the drug-DNA complex, although with different efficiency, indicating that while these two peptides could bind DNA, only the 16 mer (TPKK) peptide could bring about condensation of DNA and histone H1-depleted chromatin. A mutant 16 mer (TAKK) peptide wherein two proline residues are replaced by alanine, was ineffective in bringing about condensation of both DNA and histone H1-depleted chromatin. These results suggest that the two beta-turn structures present in the 16 mer (TPKK) peptide could be important in facilitating binding to different regions of duplex DNA thereby bringing about close packing and condensation. The condensation property of the 16 mer (TPKK) peptide was very similar to that of histone H1 in terms of (a) its preference for AT rich DNA, (b) cooperativity of condensation, and (c) salt dependence of condensation. The 16 mer (TPKK) peptide, but not the 8 mer peptide or the 16 mer (TAKK) peptide, could form complexes with a polynucleosomal 5S DNA core resulting in retarded mobility similar to the complexes formed with histone H1 on agarose gel electrophoresis.

Amino Acid Sequence↗

PubMed1999

Comparative gene transfer efficiency of low molecular weight polylysine DNA-condensing peptides.

In a previous report (M.S. Wadhwa et al. (1997) Bioconjugate Chem. 8, 81-88), we synthesized a panel of polylysine-containing peptides and determined that a minimal repeating lysine chain of 18 residues followed by a tryptophan and an alkylated cysteine residue (AlkCWK18) resulted in the formation of optimal size (78 nm diameter) plasmid DNA condensates that mediated efficient in vitro gene transfer. Shorter polylysine chains produced larger DNA condensates and mediated much lower gene expression while longer lysine chains were equivalent to AlkCWK18. Surprisingly, AlkCWK18 (molecular weight 2672) was a much better gene transfer agent than commercially available low molecular weight polylysine (molecular weight 1000-4000), despite its similar molecular weight. Possible explanations were that the cysteine or tryptophan residue in AlkCWK18 contributed to the DNA binding and the formation of small condensates or that the homogeneity of AlkCWK18 relative to low molecular weight polylysine facilitated optimal condensation. To test these hypotheses, the present study prepared AlkCYK18 and K20 and used these to form DNA condensates and conduct in vitro gene transfer. The results established that DNA condensates prepared with either AlkCYK18 or K20 possessed identical particle size and mediated in vitro gene transfer efficiencies that were indistinguishable from AlkCWK18 DNA condensates, eliminating the possibility of contributions from cysteine or tryptophan. However, a detailed chromatographic and electrospray mass spectrometry analysis of low molecular weight polylysine revealed it to possess a much lower than anticipated average chain length of dp 6. Thus, the short chain length of low molecular weight polylysine explains its inability to form small DNA condensates and mediate efficient gene transfer relative to AlkCWK18 DNA condensates. These experiments further emphasize the need to develop homogenous low molecular weight carrier molecules for nonviral gene delivery.

Cell Line↗

PubMed2001

Dynamics of collapsing and exploding Bose-Einstein condensates.

When atoms in a gas are cooled to extremely low temperatures, they will-under the appropriate conditions-condense into a single quantum-mechanical state known as a Bose-Einstein condensate. In such systems, quantum-mechanical behaviour is evident on a macroscopic scale. Here we explore the dynamics of how a Bose-Einstein condensate collapses and subsequently explodes when the balance of forces governing its size and shape is suddenly altered. A condensate's equilibrium size and shape is strongly affected by the interatomic interactions. Our ability to induce a collapse by switching the interactions from repulsive to attractive by tuning an externally applied magnetic field yields detailed information on the violent collapse process. We observe anisotropic atom bursts that explode from the condensate, atoms leaving the condensate in undetected forms, spikes appearing in the condensate wavefunction and oscillating remnant condensates that survive the collapse. All these processes have curious dependences on time, on the strength of the interaction and on the number of condensate atoms. Although the system would seem to be simple and well characterized, our measurements reveal many phenomena that challenge theoretical models.

Journal Article↗

PubMed2002

Modeling apoptotic chromatin condensation in normal cell nuclei. Requirement for intranuclear mobility and actin involvement.

Hallmarks of the terminal stages of apoptosis are genomic DNA fragmentation and chromatin condensation. Here, we have studied the mechanism of condensation both in vitro and in vivo. We found that DNA fragmentation per se of isolated nuclei from non-apoptotic cells induced chromatin condensation that closely resembles the morphology seen in apoptotic cells, independent of ATP utilization, at physiological ionic strengths. Interestingly, chromatin condensation was accompanied by release of nuclear actin, and both condensation and actin release could be blocked by reversibly pretreating nuclei with Ca2+, Cu2+, diamide, or low pH, procedures shown to stabilize internal nuclear components. Moreover, specific inhibition of nuclear F-actin depolymerization or promotion of its formation also reduced chromatin condensation. Chromatin condensation could also be inhibited by exposing nuclei to reagents that bind to the DNA minor groove, disrupting native nucleosomal DNA wrapping. In addition, in cultured cells undergoing apoptosis, drugs that inhibit depolymerization of actin or bind to the minor groove also reduced chromatin condensation, but not DNA fragmentation. Therefore, the ability of chromatin fragments with intact nucleosomes to form large clumps of condensed chromatin during apoptosis requires the apparent disassembly of internal nuclear structures that may normally constrain chromosome subdomains in non-apoptotic cells.

Actins↗

PubMed2000

Atomic force microscopy analysis of intermediates in cobalt hexammine-induced DNA condensation.

The packaging pathway of cobalt hexammine-induced DNA condensation on the surface of mica was examined by varying the concentration of Co(NH3)6(3+) in a dilute DNA solution and visualizing the condensates by atomic force microscopy (AFM). Images reveal that cobalt hexammine-induced DNA condensation on mica involves well-defined structures. At 30 microM Co(NH3)6(3+), prolate ellipsoid condensates composed of relatively shorter rods with linkages between them are formed. At 80 microM Co(NH3)6(3+), the condensed features include toroids with average diameter of approximately 240 nm as well as U-shaped and rod-like condensates with nodular appearances. The results imply that the condensates, whether toroids, U-shaped or rod-like structures have similar intermediate state which includes relatively shorter rod-like segments. The average size of the condensed toroids after incubated at room temperature for 5 h (approximately 240 nm) is much larger than that incubated for 0.5 h (approximately 100 nm). The results indicate that the condensation of DNA by Co(NH3)6(3+) is a kinetic-controlled process.

Adsorption↗

PubMed2002

Modeling of DNA condensation and decondensation caused by ligand binding.

A theoretical method for computer modeling of DNA condensation caused by ligand binding is developed. In the method, starting (s) and condensed (c) states are characterized by different free energies for ligand free DNA (F(s) and F(c) respectively), ligand binding constants (K(s) and K(c)) and stoichiometry dependent parameters (c(sm) and c(cm) - maximum relative concentration of bound ligands (per base pair) for starting and condensed state respectively). The method allows computation of the dependence of the degree of condensation (the fraction of condensed DNA molecules) on ligand concentration. Calculations demonstrate that condensation transition occurs under an increase in ligand concentration if F(s) < F(c) (i.e. S(sc) = exp [- (F(c) - F(s)) / (RT)], the equilibrium constant of the s-c transition, is low (S(sc) << 1)) and K(s) < K(c). It was also found that condensation is followed by decondensation at high ligand concentration if the condensed DNA state provides the number of sites for ligand binding less than the starting state (c(sm) > c(cm)). A similar condensation-decondensation effect was found in recent experimental studies. We propose its simple explanation.

Binding Sites↗

PubMed2006

In vivo analysis of chromosome condensation in Saccharomyces cerevisiae.

Although chromosome condensation in the yeast Saccharomyces cerevisiae has been widely studied, visualization of this process in vivo has not been achieved. Using Lac operator sequences integrated at two loci on the right arm of chromosome IV and a Lac repressor-GFP fusion protein, we were able to visualize linear condensation of this chromosome arm during G2/M phase. As previously determined in fixed cells, condensation in yeast required the condensin complex. Not seen after fixation of cells, we found that topoisomerase II is required for linear condensation. Further analysis of perturbed mitoses unexpectedly revealed that condensation is a transient state that occurs before anaphase in budding yeast. Blocking anaphase progression by activation of the spindle assembly checkpoint caused a loss of condensation that was dependent on Mad2, followed by a delayed loss of cohesion between sister chromatids. Release of cells from spindle checkpoint arrest resulted in recondensation before anaphase onset. The loss of condensation in preanaphase-arrested cells was abrogated by overproduction of the aurora B kinase, Ipl1, whereas in ipl1-321 mutant cells condensation was prematurely lost in anaphase/telophase. In vivo analysis of chromosome condensation has therefore revealed unsuspected relationships between higher order chromatin structure and cell cycle control.

Adenosine Triphosphatases↗

PubMed1999

Comparison between chromatin condensation and morphology from testis biopsy extracted and ejaculated spermatozoa and their relationship to ICSI outcome.

A significant association between male subfertility, imperfect spermiation and abnormal nuclear condensation has been suggested. The DNA content of spermatozoa might be responsible for inducing alterations in sperm morphology. The final nuclear shape, which is species-specific, depends on chromatin condensation during spermatogenesis as well as a precise organization of DNA within the nucleus. Many reports have described the association between disturbances in sperm chromatin condensation, morphology and male infertility. Chromatin condensation is achieved by gradual substitution of lysinerich somatic histones by testis-specific histone and finally by protamine. In this study two groups of patients were compared: the first consisted of 63 patients who had undergone intracytoplasmic sperm injection (ICSI) with freshly ejaculated spermatozoa whereas the second included 47 patients assigned to ICSI with testes biopsy-extracted spermatozoa. In both groups chromatin condensation was assessed by aniline blue staining and morphology evaluated according to strict criteria. The condensed chromatin and morphology of spermatozoa were significantly (P < 0.0001) less in the second group compared to the first. However the fertilization, cleavage, implantation and pregnancy rates were almost the same in both investigated groups. There was no significant difference between the two groups with respect to ICSI outcome. The percentage of chromatin condensation (nuclear maturity) and morphologically-normal spermatozoa were significantly higher (P < 0.0001) in the ejaculated spermatozoa than in those from testis biopsy but the ICSI outcome (fertilization, cleavage, implantation and pregnancy rates) was the same. In view of these results the fertilization capability and the embryo quality obtained using testis biopsy extracted spermatozoa is not influenced by chromatin condensation and sperm morphology in testicular sperm extraction (TESE)-ICSI programmes. Therefore, it could be said that neither chromatin condensation nor morphology of testis extracted sperm could predict the fertilization, implantation and pregnancy rate in TESE-ICSI programmes.

Adult↗

PubMed1988

Comparative bone marrow clastogenicity of cigarette sidestream, mainstream and recombined smoke condensates in mice.

The chromosome-damaging effects of cigarette sidestream (SS) and mainstream (MS) smoke condensates and a mixture of these were compared in 8-week-old NMRI mice by intraperitoneal administration. Each filtered commercial brand of cigarette was smoked by a smoking machine under the standard conditions, and the separately collected SS and MS smoke condensates were extracted with acetone/methanol as described elsewhere. The extracts were tested before and after treatment of animals with an enzyme inducer (Aroclor 1254) or inhibitor (Metyrapone). Increased formation of micronuclei within polychromatic erythrocytes (PCEs) of femural bone marrow 30 h after injection of the extracts was regarded as being due to a clastogenic effect. Regardless of the type of smoke extract injected, the increased formation of micronuclei was found to be dose dependent. The SS smoke condensate induced approximately 29% more micronuclei than the MS smoke condensate, the difference being significant (P less than 0.01). The overall clastogenicity of a 1:1 mixture of SS and MS smoke condensates was not substantially different from the activity of either SS or MS smoke condensate alone. Pretreatment of animals with Aroclor clearly enhanced the differences between the number of micronucleated PCEs caused by SS versus MS smoke condensate; SS smoke condensate induced 50% more micronuclei than did MS smoke condensate (P less than 0.001). Pretreatment of mice with Metyrapone did not modify appreciably the induction of micronuclei by either type of smoke. These results are discussed with reference to our previous data involving inhalation experiments and the recent issue of passive smoking.

Animals↗

PubMed2004

Involvement of histone H3 (Ser10) phosphorylation in chromosome condensation without Cdc2 kinase and mitogen-activated protein kinase activation in pig oocytes.

When oocytes resume meiosis, chromosomes start to condense and Cdc2 kinase becomes activated. However, recent findings show that the chromosome condensation does not always correlate with the Cdc2 kinase activity in pig oocytes. The objectives of this study were to examine 1) the correlation between chromosome condensation and histone H3 phosphorylation at serine 10 (Ser10) during the meiotic maturation of pig oocytes and 2) the effects of protein phosphatase 1/2A (PP1/ PP2A) inhibitors on the chromosome condensation and the involvement of Cdc2 kinase, MAP kinase, and histone H3 kinase in this process. The phosphorylation of histone H3 (Ser10) was first detected in the clump of condensed chromosomes at the diakinesis stage and was maintained until metaphase II. The kinase assay showed that histone H3 kinase activity was low in oocytes at the germinal vesicle stage (GV) and increased at the diakinesis stage and that high activity was maintained until metaphase II. Treatment of GV-oocytes with okadaic acid (OA) or calyculin-A (CL-A), the PP1/PP2A inhibitors, induced rapid chromosome condensation with histone H3 (Ser10) phosphorylation after 2 h. Both histone H3 kinase and MAP kinase were activated in the treated oocytes, although Cdc2 kinase was not activated. In the oocytes treated with CL-A and the MEK inhibitor U0126, neither Cdc2 kinase nor MAP kinase were activated and no oocytes underwent germinal vesicle breakdown (GVBD), although histone H3 kinase was still activated and the chromosomes condensed with histone H3 (Ser10) phosphorylation. These results suggest that the phosphorylation of histone H3 (Ser10) occurs in condensed chromosomes during maturation in pig oocytes. Furthermore, the chromosome condensation is correlated with histone H3 kinase activity but not with Cdc2 kinase and MAP kinase activities.

Animals↗

PubMed2002

Lotus corniculatus condensed tannins decrease in vivo populations of proteolytic bacteria and affect nitrogen metabolism in the rumen of sheep.

Condensed tannins in forage legumes improve the nutrition of sheep by reducing ruminal degradation of plant protein and increasing crude protein flow to the intestine. However, the effects of condensed tannins in forage legumes on rumen bacterial populations in vivo are poorly understood. The aim of this study was to investigate the specific effects of condensed tannins from Lotus corniculatus on four proteolytic rumen bacteria in sheep during and after transition from a ryegrass (Lolium perenne)-white clover (Trifolium repens) diet (i.e., low condensed tannins) to a Lotus corniculatus diet (i.e., higher condensed tannins). The bacterial populations were quantified using a competitive polymerase chain reaction. Lotus corniculatus was fed with or without ruminal infusions of polyethylene glycol (PEG), which binds to and inactivates condensed tannins, enabling the effect of condensed tannins on bacterial populations to be examined. When sheep fed on ryegrass-white clover, populations of Clostridium proteoclasticum B316T, Butyrivibrio fibrisolvens C211a, Eubacterium sp. C12b, and Streptococcus bovis B315 were 1.5 x 10(8), 1.1 x 10(6), 4.6 x 10(8), and 7.1 x 10(6) mL(-1), respectively. When the diet was changed to Lotus corniculatus, the average populations (after 8-120 h) of C. proteoclasticum, B. fibrisolvens, Eubacterium sp., and S. bovis decreased (P < 0.001) to 2.4 x 10(7), 1.1 x 10(5), 1.1 x 10(8), and 2.5 x 10(5) mL(-1), respectively. When PEG was infused into the rumen of sheep fed Lotus corniculatus, the populations of C. proteoclasticum, B. fibrisolvens, Eubacterium sp., and S. bovis were higher (P < 0.01-0.001) than in sheep fed Lotus corniculatus without the PEG infusion, with average populations (after 8-120 h) of 4.9 x 10(7), 3.8 x 10(5), 1.9 x 10(8), and 1.0 x 10(6), respectively. Sheep fed the Lotus corniculatus diet had lower rumen proteinase activity, ammonia, and soluble nitrogen (P < 0.05-0.001) than sheep that were fed Lotus corniculatus plus PEG. The Lotus corniculatus diet reduced rumen nitrogen digestibility (P < 0.05) and ammonia pool size and increased the flow of undegraded feed nitrogen to the abomasum. The nitrogen intake, rumen non-ammonia nitrogen pool size, rumen microbial non-ammonia nitrogen pool size, and abomasal microbial non-ammonia nitrogen fluxes were similar both in sheep fed only Lotus corniculatus and in sheep fed Lotus corniculatus plus PEG, but nonmicrobial non-ammonia nitrogen flux to the abomasum was higher (P < 0.01) for the sheep fed only Lotus corniculatus. Although condensed tannins in Lotus corniculatus reduced the populations of some proteolytic bacteria, total ruminal microbial protein and microbial protein outflow to the abomasum were unchanged, suggesting a species-specific effect of condensed tannins on bacteria in the rumen.

Animals↗

PubMed2000

Experimental condensation inhibition in constitutive and facultative heterochromatin of mammalian chromosomes.

What drives the dramatic changes in chromosome structure during the cell cycle is one of the oldest questions in genetics. During mitosis, all chromosomes become highly condensed and, as the cell completes mitosis, most of the chromatin decondenses again. Only chromosome regions containing constitutive or facultative heterochromatin remain in a more condensed state throughout interphase. One approach to understanding chromosome condensation is to experimentally induce condensation defects. 5-Azacytidine (5-aza-C) and 5-azadeoxycytidine (5-aza-dC) drastically inhibit condensation in mammalian constitutive heterochromatin, in particular in human chromosomes 1, 9, 15, 16, and Y, as well as in facultative heterochromatin (inactive X chromosome), when incorporated into late-replicating DNA during the last hours of cell culture. The decondensing effects of 5-aza-C analogs, which do not interfere with normal base pairing in substituted duplex DNA, have been correlated with global DNA hypomethylation. In contrast, decondensation of constitutive heterochromatin by incorporation of 5-iododeoxyuridine (IdU) or other non-demethylating base analogs, or binding of AT-specific DNA ligands, such as berenil and Hoechst 33258, may reflect an altered steric configuration of substituted or minor-groove-bound duplex DNA. Consequently, these compounds exert relatively specific effects on certain subsets of AT-rich constitutive heterochromatin, i.e. IdU on human chromosome 9, berenil on human Y, and Hoechst 33258 on mouse chromosomes, which provide high local concentrations of IdU incorporation sites or DNA-ligand-binding sites. None of these non-demethylating compounds affect the inactive X chromosome condensation. Structural features of chromosomes are largely determined by chromosome-associated proteins. In this light, we propose that both DNA hypomethylation and steric alterations in chromosomal DNA may interfere with the binding of specific proteins or multi-protein complexes that are required for chromosome condensation. The association between chromosome condensation defects, genomic instability, and epigenetic reprogramming is discussed. Chromosome condensation may represent a key ancestral mechanism for modulating chromatin structure that has since been realloted to other nuclear processes.

Animals↗

PubMed1991

Chromosome condensation from prophase to late metaphase: relationship to chromosome bands and their replication time.

As chromosomes condense during early mitosis, their subbands fuse in a highly coordinated fashion. Subband fusion occurs when two large subbands flanking one minor subband come together to form one band, which takes on the cytological characteristics of the original flanking subbands. Using four different banding techniques--GTG (G-bands obtained with trypsin and Giemsa), GBG (G-bands obtained with BrdU and Giemsa), RHG (R-bands obtained by heating and Giemsa), and RBG (R-bands obtained with BrdU and Giemsa)--we studied subband fusion from prophase (1,250 bands per haploid set) to late metaphase (300 bands). To quantify the condensation process, a fusion index was established. We found that chromosomes contain preferential zones of condensation. From prophase to late metaphase, the early replicating subbands (R-subbands) fuse more readily with each other than do the late-replicating subbands (G-subbands). R-bands usually replicate early and condense late independently of the adjacent G-bands, which replicate late but condense early. Therefore, chromosome bands can undergo DNA replication and chromatin condensation relatively autonomously. Our data suggest that (1) chromosome replication and condensation are closely connected in time, (2) the metaphase bands represent independent units of chromatin condensation, and (3) the condensation process is an important feature of chromosome organization.

Cells, Cultured↗

PubMed2005

Influence of condensation temperature on selected exhaled breath parameters.

BACKGROUND: The effects of changes in cooling temperature on biomarker levels in exhaled breath condensate have been little investigated. The aim of the study was to test the effect of condensation temperature on the parameters of exhaled breath condensate and the levels of selected biomarkers. METHODS: Exhaled breath condensate was collected from 24 healthy subjects at temperatures of -10, -5, 0 and +5 C degrees. Selected parameters (condensed volume and conductivity) and biomarkers (hydrogen peroxide, malondialdehyde) were measured. RESULTS: There was a progressive increase in hydrogen peroxide and malondialdehyde concentrations, and condensate conductivity as the cooling temperature increased; total condensate volume increased as the cooling temperature decreased. CONCLUSION: The cooling temperature of exhaled breath condensate collection influenced selected biomarkers and potential normalizing factors (particularly conductivity) in different ways ex vivo. The temperature of exhaled breath condensate collection should be controlled and reported.

Adult↗