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Chromatin Condensation (chromatin + condensation)
Kinds of Chromatin Condensation Selected AbstractsHypoxia-induced apoptosis and tube breakdown are regulated by p38 MAPK but not by caspase cascade in an in vitro capillary model composed of human endothelial cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2007Toshiro Ohta In order to improve medical treatment of ischemic injury such as myocardial infarction, it is important to elucidate hypoxia-induced changes to endothelial cells. An in vitro blood vessel model, in which HUVECs are stimulated to form a network of capillary-like tubes, was used to analyze hypoxia-induced morphological and biochemical changes. When exposed to hypoxia, the network of capillary tubes broke down into small clusters. This tube breakdown was accompanied by chromatin condensation and cell nuclear fragmentation, morphological markers of apoptosis, and activation of two apoptotic signals, caspase-3 and p38. We investigated what roles caspase cascade and p38 play in hypoxia-induced apoptosis and tube breakdown by using zVAD-fmk and SB203580, specific inhibitors of these two apoptotic signals, respectively. Chromatin condensation and cell nuclear fragmentation and tube breakdown were effectively inhibited by SB203580, but not by zVAD-fmk. SB203580 caused dephosphorylation of p38, which indicates that p38 was autophosphorylated. Inhibition by zVAD-fmk caused slight MW increase in p17 and emergence of p19, which indicates that the inhibitor caused partial processing of caspase-3. Inhibition of p38 suppressed activation of caspase-3 but not vice versa. In addition, these two inhibitors were shown to differentially inhibit cleavage of so-called caspase substrates. SB203580 inhibited cleavage of PARP and lamin A/C, while zVAD-fmk inhibited cleavage of lamin A/C but not that of PARP. Taken together, these results show that p38 is located upstream of caspase cascade and that, although caspase-3 is activated, a p38-regulated caspase-independent pathway is crucial for the execution of hypoxia-induced apoptosis and tube breakdown. J. Cell. Physiol. 211: 673,681, 2007. © 2007 Wiley-Liss, Inc. [source] Entorhinal Cortex Lesion in the Mouse Induces Transsynaptic Death of Perforant Path Target NeuronsBRAIN PATHOLOGY, Issue 3 2004Adam D. Kovac Entorhinal cortex lesion (ECL) is a well described model of anterograde axonal degeneration, subsequent sprouting and reactive synaptogenesis in the hippocampus. Here, we show that such lesions induce transsynaptic degeneration of the target cells of the lesions pathway in the dentate gyrus. Peaking between 24 and 36 hours postlesion, dying neurons were labeled with DeOlmos silver-staining and antisera against activated caspase 3 (CCP32), a downstream inductor of programmed cell death. Within caspase 3-positive neurons, fragmented nuclei were co-localized using Hoechst 33342 staining. Chromatin condensation and nuclear fragmentation were also evident in semithin sections and at the ultrastructural level, where virtually all caspase 3-positive neurons showed these hallmarks of apoptosis. There is a well-described upregulation of the apoptosis-inducing CD95/L system within the CNS after trauma, yet a comparison of caspase 3-staining patterns between CD95 (lpr)- and CD95L (gld)-deficient with non-deficient mice (C57/bl6) provided no evidence for CD95L-mediated neuronal cell death in this setting. However, inhibition of NMD A receptors with MK-801 completely suppressed caspase 3 activation, pointing to glutamate neurotoxicity as the upstream inducer of the observed cell death. Thus, these data show that axonal injury in the CNS does not only damage the axotomized neurons themselves, but can also lethally affect their target cells, apparently by activating glutamate-mediated intracellular pathways of programmed cell death. [source] Programmed cell death of the ovarian nurse cells during oogenesis of the silkmoth Bombyx moriDEVELOPMENT GROWTH & DIFFERENTIATION, Issue 7 2006Vicky E. Mpakou In the present study, we describe the features of programmed cell death of the ovarian nurse cells occurring during vitellogenesis of the silkmoth Bombyx mori. At developmental stage 5, the nurse cells occupy one-half of the follicular volume and obtain a rather spherical shape, while the nurse cell nuclei appear large and elongated, forming impressive projections. At the following stage, stage 6, the nurse cells decrease in size and their shape becomes elliptic. The nuclei remain elongated, being also characterized by large lobes. The lobes of the ramified nurse cell nuclei seem to retain the nucleus in the center of the cell during the dumping of the nurse cell cytoplasm into the growing oocyte. At stage 7, membrane enclosed vacuoles can be easily detected into the nurse cells cytoplasm. Ultrastructural analysis and fluorescent microscopy using mono-dansyl-cadaverine staining of these vacuoles also reveal that they represent autolysosomes. Caspase activity is detected during stage 7, as it is demonstrated by using the Red-VAD-FMK staining reagent. At developmental stages 8 and 9, the nurse cells exhibit chromatin condensation, DNA fragmentation and caspase activity. Finally, during the following stage 10, the nuclear remnants are assembled into apoptotic vesicles, which, after being phagocytosed, are observed in the cytoplasm of adjacent follicle cells. We propose that apoptosis and autophagy operate synergistically during vitellogenesis of B. mori, in order to achieve an efficient and rapid clearance of the degenerated nurse cell cluster. [source] Erasure of the paternal transcription program during spermiogenesis: The first step in the reprogramming of sperm chromatin for zygotic developmentDEVELOPMENTAL DYNAMICS, Issue 5 2008Junke Zheng Abstract Male germ cells possess a unique epigenetic program and express a male-specific transcription profile. However, when its chromatin is passed onto the zygote, it expresses an transcription/epigenetic program characteristic of the zygote. The mechanism underlying this reprogramming process is not understood at present. In this study, we show that an extensive range of chromatin factors (CFs), including essential transcription factors and regulators, remodeling factors, histone deacetylases, heterochromatin-binding proteins, and topoisomerases, were removed from chromatin during spermiogenesis. This process will erase the paternal epigenetic program to generate a relatively naive chromatin, which is likely to be essential for installation of the zygotic developmental program after fertilization. We have also showed that transcription termination in male germ cells was temporally correlated with CF dissociation. A genome-wide CF dissociation will inevitably disassemble the transcription apparatus and regulatory mechanism and lead to transcription silence. Based on data presented in this and previous studies (Sun et al., Cell Research [2007] 17:117,134), we propose that paternal-zygotic transcription reprogramming begins with a genome-wide CF dissociation to erase the existing transcription program in later stages of spermatogenesis. This will be followed by assembling of the zygotic equivalent after fertilization. The transcription/epigenetic program of the male germ cell is transformed into a zygotic one using an erase-and-rebuild strategy similar to that used in the maternal-zygotic transition. It is also noted that transcription is terminated long after meiosis is completed and before chromatin becomes highly condensed during spermatogenesis. The temporal order of these events suggests that transcription silence does not have to be coupled to meiosis or chromatin condensation. Developmental Dynamics 237:1463-1476, 2008. © 2008 Wiley-Liss, Inc. [source] Cataracts in experimentally diabetic mouse: morphological and apoptotic changesDIABETES OBESITY & METABOLISM, Issue 2 2005K. R. Hegde Aim:, The objective of these investigations was to extend our earlier study on the induction of cataracts in diabetic mice, a low aldose reductase (AR) animal model at morphological level. Previous studies were done primarily at biochemical level. Methods:, Diabetes was induced by intraperitoneal administration of streptozotocin. The lenses isolated after the establishment of diabetes were then subjected to histologic and electron microscopic studies. Results:, Morphological alterations were characterized by shrinkage, elongation and lobulization of the nuclei of the epithelial cells. This was associated with chromatin condensation and its margination. Similar structural aberrations were also observed in a significant number of the subepithelial fibre cells representing defect in fibre maturation. More interestingly, unlike that in other common animal models of diabetic cataract, such abnormally nucleated cells were also found to be prevalent in the posterior subcapsular region, a finding common in human diabetics also. Conclusion:, The present studies further affirm the suitability of the mouse model for a study of cataractogenesis induced by diabetes. Because of the findings reported herein, as well as the known biochemical similarity between the lenses of the mice and humans in respect of AR deficiency, contrary to the rat model where it is very high, use of this species is considered more useful towards understanding the basic aetiology as well as for evaluating the efficacy of various referred nutritional and metabolic antioxidants against such cataracts. [source] Caffeine mimics adenine and 2,-deoxyadenosine, both of which inhibit the guanine-nucleotide exchange activity of RCC1 and the kinase activity of ATRGENES TO CELLS, Issue 5 2003Hitoshi Nishijima Background: Both caffeine and the inactivation of RCC1, the guanine-nucleotide exchange factor (GEF) of Ran, induce premature chromatin condensation (PCC) in hamster BHK21 cells arrested in the S-phase, suggesting that RCC1 is a target for caffeine. Results: Caffeine inhibited the Ran-GEF activity of RCC1 by preventing the binary complex formation of Ran-RCC1. Inhibition of the Ran-GEF activity of RCC1 by caffeine and its derivatives was correlated with their ability to induce PCC. Since caffeine is a derivative of xanthine, the bases and nucleosides were screened for their ability to inhibit RCC1. Adenine, adenosine, and all of the 2,-deoxynucleosides inhibited the Ran-GEF activity of RCC1; however, only adenine and 2,-deoxyadenosine (2,-dA) induced PCC. A factor(s) other than RCC1, should therefore be involved in PCC-induction. We found that both adenine and 2,-dA, but none of the other 2,-deoxynucleosides, inhibited the kinase activity of ATR, similar to that of caffeine. The ATR pathway was also abrogated by the inactivation of RCC1 in tsBN2 cells. Conclusion: The effect of caffeine on cell-cycle control mimics the biological effect of adenine and 2,-dA, both of which inhibit ATR. dATP, a final metabolite of adenine and 2,-dA, is suggested to inhibit ATR, resulting in PCC. [source] Comparison between computerized slow-stage and static liquid nitrogen vapour freezing methods with respect to the deleterious effect on chromatin and morphology of spermatozoa from fertile and subfertile menINTERNATIONAL JOURNAL OF ANDROLOGY, Issue 2 2001M. E. Hammadeh The purpose of this study was to determine the negative effects (cryodamage) on human spermatozoa after freeze-thawing and to determine whether freeze-thawing of spermatozoa with a programmed slow freezer is better than freezing with liquid nitrogen vapour (rapid freezing) with regard to alterations in sperm chromatin and morphology in semen from fertile (donor) and subfertile, IVF/ICSI, patients. Ninety-five semen samples were obtained either from patients attending our IVF unit for treatment (n=34) or from donors (n=25) with proven fertility and normal sperm quality according to WHO guidelines. Each semen sample was divided into two parts after liquefaction and addition of the cryoprotectant. The first part was frozen using a programmed biological freezer and the second part was frozen by means of liquid nitrogen vapour. Smears were made before the freezing and after the thawing procedure to assess morphology (strict criteria) and chromatin condensation (Acridine Orange test). The mean percentage of chromatin condensed spermatozoa in the samples from donors (control group) was 92.4 ± 8.4% before freezing and this decreased significantly (p < 0.0001) to 88.7 ± 11.2% after freeze-thawing with the computerized slow-stage freezer and to 87.2 ± 12.3% after using static liquid nitrogen vapour (p < 0.001). The corresponding values for semen obtained from patients was 78.9 ± 10.3% before freezing which decreased to 70.7 ± 10.8 and 68.5 ± 14.8%, respectively (p < 0.001). On the other hand, the mean percentage of normal sperm morphology in the control group decreased from 26.3 ± 7.5% before freezing to 22.1 ± 6.4% (p < 0.0001) after thawing with the computerized slow-stage freezer and to 22.2 ± 6.6% (p < 0.0001) after the use of static liquid nitrogen vapour. In the patient group, the mean percentage of normal morphology decreased from 11.7 ± 6.1% after freezing with the biological freezer to 9.3 ± 5.6% and to 8.0 ± 4.9% after freezing with static liquid nitrogen vapour. This study demonstrates that chromatin packaging and morphology of human spermatozoa decrease significantly after the freeze-thawing procedure, not only after the use of static liquid nitrogen vapour but also after the use of a computerized slow-stage freezer. However, the chromatin of semen samples with normal semen parameters (donor sperm) withstand the freeze-thaw injury better than those with low quality semen samples. Therefore, the computerized slow stage freezer could be recommended for freezing of human spermatozoa, especially for subnormal semen samples, for example, ICSI and ICSI/TESE candidates and from patients with testicular tumours or Hodgkin's disease, in order to avoid further damage to the sperm chromatin structure. [source] Signalling responses linked to betulinic acid-induced apoptosis are antagonized by MEK inhibitor U0126 in adherent or 3D spheroid melanoma irrespective of p53 statusINTERNATIONAL JOURNAL OF CANCER, Issue 5 2006Manuel Rieber Abstract MEK1/2 inhibitors like U0126 can potentiate or antagonize the antitumor activity of cytotoxic agents such as cisplatin, paclitaxel or vinblastine, depending on the drug or the target cells. We now investigated whether U0126, differentially regulates melanoma signaling in response to UV radiation or betulinic acid, a drug lethal against melanoma. This report shows that U0126 inhibits early response (ERK) kinase activation and cyclin A expression in wt p53 C8161 melanoma exposed to either UV radiation or betulinic acid. However, U0126 does not protect from UV damage, but counteracts betulinic acid-mediated apoptosis in the same cells. Protection from the latter drug by joint treatment with U0126 was also evident in wt p53 MelJuso melanoma and mutant p53 WM164 melanoma. The latter cells were the most responsive to betulinic acid, showing a selective decline in the cdk4 protein, without a comparable change in other key cell cycle proteins like cdc2, cdk2, cdk7 or cyclin A, prior to apoptosis-associated PARP fragmentation. Laser scanning cytometry also showed that betulinic acid induced a significant increase in chromatin condensation in WM164 melanoma irrespective of whether they were in adherent form or as multicellular spheroids. All these betulinic acid-induced changes were counteracted by U0126. Our data show for the first time that (a) cdk4 protein is an early target of betulinic acid-induced apoptosis and (b) unrestricted ERK signaling favours betulinic acid-induced apoptosis, but this is counteracted by U0126, partly through counteracting chromatin condensation and restoring Akt activation decreased by betulinic acid treatment. © 2005 Wiley-Liss, Inc. [source] Morphological features of Murray Valley encephalitis virus infection in the central nervous system of swiss miceINTERNATIONAL JOURNAL OF EXPERIMENTAL PATHOLOGY, Issue 1 2000Vance Matthews We have examined the histological and ultrastructural features of CNS infection with Murray Valley encephalitis (MVE) virus in mice inoculated with a virulent parental strain (BH3479). Light microscopic examination revealed neuronal necrosis in the olfactory bulb and hippocampus of MVE-infected brains by 5 days post-infection (pi). Electron microscopy of these regions showed endoplasmic reticulum membrane proliferation, and tubular and spherical structures in the cisternae of the endoplasmic reticulum, Golgi complex and nuclear envelope. At seven to eight days pi, infected neurones exhibited chromatin condensation and extrusion, nuclear fragmentation, loss of segments of the nuclear envelope, reduced surface contact with adjacent cells and loss of cytoplasmic organelles. This cell injury was particularly noticeable in the proximal CA3 and distal CA1 regions of the hippocampus. The inflammatory cell profile consisted of macrophages, lymphocytes and especially neutrophils, and many of these inflammatory cells were apoptotic. High mortality rates in the BH3479-infected population of mice correlated with the intense polymorphonuclear and mononuclear leucocyte inflammatory infiltrate in the CNS. [source] Molecular dynamics simulation on HP1 protein binding by histone H3 tail methylation and phosphorylationINTERNATIONAL JOURNAL OF QUANTUM CHEMISTRY, Issue 4 2009Yan-Ke Jiang Abstract Trimethylation of histone H3 lysine 9 is important for recruiting heterochromatin protein 1 (HP1) to discrete regions of the genome, thereby regulating gene expression, chromatin packaging, and heterochromatin formation. Phosphorylation of histone H3 has been linked with mitotic chromatin condensation. During mitosis in vivo, H3 lysine 9 methylation and serine 10 phosphorylation can occur concomitantly on the same histone tail, whereas the influence of phosphorylation to trimethylation H3 tail recruiting HP1 remains controversial. In this work, molecular dynamics simulation of HP1 complexed with both trimethylated and phosphorylated H3 tail were performed and compared with the results from the previous methylated H3-HP1 trajectory. It is clear from the 10-ns dynamics simulation that two adjacent posttranslational modifications directly increase the flexibility of the H3 tail and weaken HP1 binding to chromatin. A combinatorial readout of two adjacent posttranslational modifications,a stable methylation and a dynamic phosphorylation mark,establish a regulatory mechanism of protein,protein interactions. © 2008 Wiley Periodicals, Inc. Int J Quantum Chem, 2009 [source] Regression of blood vessels in the ventral velum of Xenopus laevis Daudin during metamorphosis: light microscopic and transmission electron microscopic studyJOURNAL OF ANATOMY, Issue 2 2000H. BARTEL Structural changes of the ventral velum of Xenopus laevis tadpoles from late prometamorphosis (stage 58) to the height of metamorphic climax (stage 62) were examined by light and transmission electron microscopy. Special emphasis was given to the blood vessel regression. Early changes of velar capillaries were formation of luminal and abluminal endothelial cell processes, vacuolation, and cytoplasmic and nuclear chromatin condensation. At the height of metamorphic climax, transmission electron microscopy revealed apoptotic endothelial cells with nuclear condensation and fragmentation, intraluminal bulging of rounded endothelial cells which narrowed or even plugged the capillary, and different stages of endothelial cell detachment (,shedding') into the vessel lumen. These changes explain the ,miniaturisation' of the velar microvascular bed as well as the typical features found in resin-casts of regressing velar vessels which have been observed in a previous scanning electron microscopy study of the ventral velum. [source] 2,-Deoxyadenosine causes apoptotic cell death in a human colon carcinoma cell lineJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 6 2003Michela Giannecchini Abstract The combination of 2,-deoxyadenosine and 2,-deoxycoformycin is toxic for the human colon carcinoma cell line LoVo. In this study we investigated the mode of action of the two compounds and have found that they promote apoptosis. The examination by fluorescence microscopy of the cells treated with the combination revealed the characteristic morphology associated with apoptosis, such as chromatin condensation and nuclear fragmentation. The occurrence of apoptosis was also confirmed by the release of cytochrome c and the proteolytic processing of procaspase-3 in cells subjected to the treatment. To exert its triggering action on the apoptotic process, 2,-deoxyadenosine enters the cells through an equilibrative nitrobenzyl-thioinosine-insensitive carrier, and must be phosphorylated by intracellular kinases. Indeed, in the present work we demonstrate by analysis of the intracellular metabolic derivatives of 2,-deoxyadenosine that, as suggested by our previous findings, in the incubation performed with 2,-deoxyadenosine and 2,-deoxycoformycin, an appreciable amount of dATP was formed. Conversely, when also an inhibitor of adenosine kinase was added to the incubation mixture, dATP was not formed, and the toxic and apoptotic effect of the combination was completely reverted. © 2003 Wiley Periodicals, Inc. J Biochem Mol Toxicol 17:329,337, 2003; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.10095 [source] Hypoxia-induced apoptosis and tube breakdown are regulated by p38 MAPK but not by caspase cascade in an in vitro capillary model composed of human endothelial cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2007Toshiro Ohta In order to improve medical treatment of ischemic injury such as myocardial infarction, it is important to elucidate hypoxia-induced changes to endothelial cells. An in vitro blood vessel model, in which HUVECs are stimulated to form a network of capillary-like tubes, was used to analyze hypoxia-induced morphological and biochemical changes. When exposed to hypoxia, the network of capillary tubes broke down into small clusters. This tube breakdown was accompanied by chromatin condensation and cell nuclear fragmentation, morphological markers of apoptosis, and activation of two apoptotic signals, caspase-3 and p38. We investigated what roles caspase cascade and p38 play in hypoxia-induced apoptosis and tube breakdown by using zVAD-fmk and SB203580, specific inhibitors of these two apoptotic signals, respectively. Chromatin condensation and cell nuclear fragmentation and tube breakdown were effectively inhibited by SB203580, but not by zVAD-fmk. SB203580 caused dephosphorylation of p38, which indicates that p38 was autophosphorylated. Inhibition by zVAD-fmk caused slight MW increase in p17 and emergence of p19, which indicates that the inhibitor caused partial processing of caspase-3. Inhibition of p38 suppressed activation of caspase-3 but not vice versa. In addition, these two inhibitors were shown to differentially inhibit cleavage of so-called caspase substrates. SB203580 inhibited cleavage of PARP and lamin A/C, while zVAD-fmk inhibited cleavage of lamin A/C but not that of PARP. Taken together, these results show that p38 is located upstream of caspase cascade and that, although caspase-3 is activated, a p38-regulated caspase-independent pathway is crucial for the execution of hypoxia-induced apoptosis and tube breakdown. J. Cell. Physiol. 211: 673,681, 2007. © 2007 Wiley-Liss, Inc. [source] Mitotic catastrophe as a prestage to necrosis in mouse liver cells treated with Helicobacter pullorum sonicatesJOURNAL OF MORPHOLOGY, Issue 8 2009Liesbeth M. Ceelen Abstract Helicobacter pullorum infections have been associated with several enterohepatic diseases, but the mechanism of action is currently undefined. The present study was therefore set up to investigate possible cytotoxic effects of this pathogen on liver cells. A mouse hepatic cell line was exposed to H. pullorum sonicate and cytotoxicity was observed for all isolates after incubation for 72 h. Features characteristic for mitotic catastrophe characterized by chromatin condensation, formation of multinuclear distended cells and micronucleation, were recorded. In addition, intranuclear pseudoinclusions were seen in sonicate-treated cells. Finally, cells exposed to sonicate eventually underwent cell death with the morphological features of necrosis, which occurred without activation of caspase-3. The toxic factor involved in the cytotoxic activity proved to be soluble, trypsin,sensitive and stable at 56°C and at ,70°C with a molecular weight to be over 50 kDa. The current study shows for the first time that H. pullorum causes mitotic catastrophe resulting in primary necrosis in mouse hepatocytes. J. Morphol., 2009. © 2009 Wiley-Liss, Inc. [source] Morphological characterization of the testicular cells and seminiferous epithelium cycle in six species of Neotropical batsJOURNAL OF MORPHOLOGY, Issue 8 2009Mateus R. Beguelini Abstract We know little about the process of spermatogenesis in bats, a great and diverse clade of mammals that presents different reproductive strategies. In the present study, spermatogenesis in six species of Neotropical bats was investigated by light microscopy. On the basis of chromatin condensation, nuclear morphology, relative position to the basal membrane and formation of the flagellum, three types of spermatogonia were recognized: dark type A (Ad), pale type A (Ap), and type B; the development of spermatids was divided into seven steps. With the exception of Myotis nigricans, the seminiferous epithelium cycle of the other five species studied was similar to those of other mammals, showing gradual stages by the tubular morphology method. Asynchrony was observed in the seminiferous epithelium cycle of M. nigricans, shown by overlapping stages and undefined cycles. The frequencies found in the three phases of the cycle were variable with the greatest frequency occurring in the postmeiotic phase (>50%) and the least in the meiotic phase (<10%). The similarities observed in the five species of Phyllostomidae appeared to be related to their phylogenetic relationship and shorter divergence times, whereas the differences in M. nigricans appeared to be related to its greater phylogenetic distance because the Vespertilionidae family diverged earlier. J. Morphol., 2009. © 2009 Wiley-Liss, Inc. [source] DNA methylation: an epigenetic pathway to cancer and a promising target for anticancer therapyJOURNAL OF ORAL PATHOLOGY & MEDICINE, Issue 8 2002Jesper Worm Abstract The unique properties of a cancer cell are acquired through a stepwise accumulation of heritable changes in the information content of proto-oncogenes and tumor suppressor genes. While gain, loss, and mutation of genetic information have long been known to contribute to tumorigenesis, it has been increasingly recognized over the past 5 years that ,epigenetic' mechanisms may play an equally important role. The main epigenetic modification of the human genome is methylation of cytosine residues within the context of the CpG dinucleotide. De novo methylation of ,CpG islands' in the promoter regions of tumor suppressor genes may lead to transcriptional silencing through a complex process involving histone deacetylation and chromatin condensation, and thus represents a tumorigenic event that is functionally equivalent to genetic changes like mutation and deletion. DNA methylation is interesting from a diagnostic viewpoint because it may be easily detected in DNA released from neoplastic and preneoplastic lesions into serum, urine or sputum, and from a therapeutic viewpoint because epigenetically silenced genes may be reactivated by inhibitors of DNA methylation and/or histone deacetylase. A better understanding of epigenetic mechanisms leading to tumor formation and chemoresistance may eventually improve current cancer treatment regimens and be instructive for a more rational use of anticancer agents. [source] Extracts of various species of Epilobium inhibit proliferation of human prostate cellsJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 5 2003Annabella Vitalone This study examined whether various species of Epilobium, a phytotherapeutic agent used in folk medicine as a treatment for benign prostatic hyperplasia, may have an antiproliferative effect in PZ-HPV-7 human prostatic epithelial cells in-vitro. The MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) test, [methyl- 3H]thymidine incorporation into DNA and flow cytometry analysis were used to evaluate cell proliferation. Ethanolic extracts of E. spicatum, E. rosmarinifolium and E. tetragonum inhibited DNA synthesis in PZ-HPV-7 cells. While at high concentrations all extracts were cytotoxic, DNA synthesis was also decreased at levels that caused no or little cytotoxicity. Treatment of cells with Epilobium extracts did not result in a formation of DNA fragments (evaluated by the TUNEL assay) or chromatin condensation (assessed by Hoechst staining). Flow cytometry analysis indicated that Epilobium extracts inhibit the progression of the cell cycle from the G0/G1 phase. These results suggest that extracts of Epilobium inhibit proliferation of human PZ-HPV-7 cells in-vitro by affecting progression of the cell cycle. This study provides some initial biological plausibility for the use of Epilobium extracts in benign prostatic hyperplasia. [source] Paederus dermatitis in Egypt: a clinicopathological and ultrastructural studyJOURNAL OF THE EUROPEAN ACADEMY OF DERMATOLOGY & VENEREOLOGY, Issue 10 2010M Assaf Abstract Background, Outbreaks of paederus dermatitis (PD) have been observed in different parts of the world, yet the histopathological and ultrastructural changes and their relationship to pederin toxin have not been described. Objective, To describe the clinical presentations of PD in Egypt and to study the effects of pederin toxin on the skin by evaluating the histopathological and ultrastructural changes of some representative cases. Methods, One hundred and thirteen patients with PD were studied clinically and epidemiologically. Skin biopsies were taken from 40 patients for histopathological examination and from 20 patients for electron microscopic (EM) examination. Results, Clinically, the most common presentation comprised erythematous plaques with micropustules. Blisters exhibited a linear configuration in 40% of the patients and kissing lesions were observed in 13%. Multiple lesions occurred in 78% of the patients and the face was the most commonly involved site (48%). The insect was identified as Paederus alfierii. Histopathological examination revealed features of acute irritant dermatitis in the upper epidermis. Mitotic figures and apoptotic changes such as chromatin condensation and DNA fragmentation were identified in the basal and suprabasal layers. These features were confirmed by EM. Conclusions, Clinical, histopathological and, for the first time, ultrastructural characteristics of paederus dermatitis are described. The pathological abnormalities of the upper epidermis are caused by the irritant effect of pederin toxin. The presence of apoptosis within the lower epidermis can be related to this toxin, a point that needs further research, hoping for its future implications in the management of hyperproliferative disorders. [source] Behaviors of ATP-dependent chromatin remodeling factors during maturation of bovine oocytes in vitroMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 2 2010Gabbine Wee The mammalian oocyte undergoes dynamic changes in chromatin structure to reach complete maturation. However, little known is about behaviors of ATP-dependent chromatin remodeling factors (ACRFs) during meiosis. Here, we found that respective ACRFs may differently behave in the process of oocyte maturation in the bovine. All ACRFs interacted with oocytic chromatin at the germinal vesicle (GV) stage. Mi-2 and hSNF2H disappeared from GV-chromatin within 1,hr of in vitro culture whereas Brg-1 and BAF-170 were retained throughout germinal vesicle break down (GVBD). Brg-1 was localized on the condensed chromatin outside, whereas BAF-170 was entirely excluded from condensed chromatin. Thereafter, Brg-1 and BAF-170 interacted with metaphase I and metaphase II chromosomes. These results imply that Mi-2 and hSNF2H may initiate the meiotic resumption, and Brg-1 and BAF-170 may support chromatin condensation during meiosis. In addition, DNA methylation and methylation of histone H3 at lysine 9 (H3K9) seem to be constantly retained in the oocyte chromatin throughout in vitro maturation. Inhibition of ACRF activity by treatment with the inhibitor apyrase led to retarded chromatin remodeling in bovine oocytes, thereby resulting in poor development of fertilized embryos. Therefore, these results indicate that precise behaviors of ACRFs during meiosis are critical for nuclear maturation and subsequent embryonic development in the bovine. Mol. Reprod. Dev. 77: 126,135, 2010. © 2009 Wiley-Liss, Inc. [source] Immunolocalization of the High-Mobility Group N2 protein and acetylated histone H3K14 in early developing parthenogenetic bovine embryos derived from oocytes of high and low developmental competenceMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 2 2008Guilherme M. Bastos Abstract This study investigated differences in the distribution of acetylated histone H3 at Lysine 14 (H3K14ac) and the High-Mobility Group N2 (HMGN2) protein in the chromatin of early- (before 24 hr) and late-cleaved (after 24 hr) bovine embryos derived from small- (1,2 mm) and large-follicles (4,8 mm). The presence of HMGN2 and H3K14ac has been associated with different nuclear functions including chromatin condensation, transcription, DNA replication and repair. In vitro matured oocytes were parthenogenetically activated (PA) and cultured in synthetic oviduct fluid medium. Early- and late-cleaved embryos were fixed at 36, 50, 60, 70 and 80 hr after PA to detect the presence of H3K14ac and HMGN2. The rates of nuclear maturation (81.1% vs. 58.7%), early cleavage (46.9% vs. 38.9%), and development to blastocyst stage (34.3% vs. 18.9%) were higher (P,<,0.05) in oocytes derived from large- compared to small follicles. The proportion of positively stained nuclei at 50 and 60 hr after PA was higher for both H3K14ac (27.2% vs. 4.8% and 64.3% vs. 30%) and HMGN2 (47% vs. 21.3% and 60.6% vs. 46%) in early versus late cleaved embryos derived from small- versus large-follicles, respectively. However, the rate of positive nuclei in early-cleaved embryos from small-versus large-follicles was similar for HMGN2 (87% vs. 93%) but lower for H3K14ac (51% vs. 64.4%) at 80 hr after PA. These data suggest that less developmentally competent embryos derived from small follicles had an altered chromatin remodeling process at the early stages of development compared to those derived from large follicles that are more competent to support development to blastocyst stage. Mol. Reprod. Dev. 75: 282,290, 2008. © 2007 Wiley-Liss, Inc. [source] Large-scale chromatin remodeling in germinal vesicle bovine oocytes: Interplay with gap junction functionality and developmental competenceMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 6 2007Valentina Lodde Abstract In mammals, oocyte acquires a series of competencies sequentially during folliculogenesis that play critical roles at fertilization and early stages of embryonic development. In mouse, chromatin in germinal vesicle (GV) undergoes dynamic changes during oocyte growth and its progressive condensation has been related to the achievement of developmental potential. Cumulus cells are essential for the acquisition of meiotic competence and play a role in chromatin remodeling during oocyte growth. This study is aimed to characterize the chromatin configuration of growing and fully grown bovine oocytes, the status of communications between oocyte and cumulus cells and oocyte developmental potential. Following nuclear staining, we identified four discrete stages of GV, characterized by an increase of chromatin condensation. GV0 stage represented 82% of growing oocytes and it was absent in fully grown oocytes. GV1, GV2, and GV3 represented, respectively, 24, 31, and 45% of fully grown oocytes. Our data indicated a moderate but significant increase in oocyte diameter between GV0 and GV3 stage. By dye coupling assay the 98% of GV0 oocytes showed fully open communications while the number of oocytes with functionally closed communications with cumulus cells was significantly higher in GV3 group than GV1 and GV2. However, GV0 oocytes were unable to progress through metaphase II while GV2 and GV3 showed the highest developmental capability. We conclude that in bovine, the progressive chromatin condensation is related to the sequential achievement of meiotic and embryonic developmental competencies during oocyte growth and differentiation. Moreover, gap-junction-mediated communications between oocyte and cumulus cells could be implicated in modulating the chromatin remodeling process. Mol. Reprod. Dev. 74: 740,749, 2007. © 2006 Wiley-Liss, Inc. [source] Premature translation of transition protein 2 mRNA causes sperm abnormalities and male infertilityMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 3 2007Khailun Tseden Abstract During mammalian spermiogenesis somatic histones are replaced at first by transition proteins, which are in turn replaced by the protamines, forming the sperm nucleoprotamines. It is believed that transition protein 2 (Tnp2) is necessary for maintaining the normal processing of protamines and, consequently, the completion of chromatin condensation. The transition protein mRNAs are stored in translationally inert messenger ribonucleoprotein particles for up to 7 days until translational activation in elongated spermatids. Substantial evidence suggests an involvement of 3,untranslated region (UTR) in the translational regulation of the Tnp2 mRNAs. In order to determine the role of Tnp2 3,UTR in translational regulation and to study whether the translational repression of Tnp2 mRNA is necessary for normal spermatid differentiation in mice, we generated transgenic mice that carry a Tnp2-hGH transgene. In this transgene, 3,UTR of Tnp2 gene was replaced by 3, 3,UTR of human growth hormone gene. In these transgenic animals, transcription and translation of Tnp2 occur simultaneously in round spermatids which is an evidence for involvement of Tnp2 3,UTR in its translation repression. Premature translation of Tnp2 mRNA caused abnormal head morphogenesis, reduced sperm motility and male infertility. These results show clearly that a strict temporal and stage-specific Tnp2 translation is necessary for the correct differentiation of round spermatids into mature spermatozoa and for male fertility. Mol. Reprod. Dev. © 2006 Wiley-Liss, Inc. [source] Transition of nuclear proteins and chromatin structure in spermiogenesis of Sepia officinalis,MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 3 2007F. MartÍnez-Soler Abstract During spermiogenesis of Sepia officinalis histones are directly substituted by a molecule of precursor protamine, which is later transformed into the protamine through a deletion of the amino terminal end. In the present work, it is shown that the pattern of spermiogenic chromatin condensation consists of a phase of "patterning" and a phase of "condensation." In the phase of patterning, three structural remodelings are produced in the chromatin structure: [somatic-like chromatin,,,18 nm granules,,,25 nm fibers,,,44 nm fibers]. The first remodeling of the chromatin into granules of 18 nm takes place without the entrance of specific proteins in the spermiogenic nuclei. The second remodeling [granules of 18 nm,,,fibers of 25 nm] is due to the entrance of the precursor protamine and its interaction with the DNA,histone complex. The third remodeling [fibers of 25 nm,,,fibers of 44 nm] occurs simultaneously with the disappearance of histones from the chromatin. In the phase of condensation, the fibers of 44 nm coalesce among themselves to form progressively larger aggregates of chromatin. In this phase there are no substantial variations in the nuclear proteins, so that the condensation of the chromatin must respond to posttranscriptional changes of the precursor protamine (dephosphorylation, deletion of the amino-terminal end). Mol. Reprod. Dev. © 2006 Wiley-Liss, Inc. [source] XY chromosomal bivalent: Nucleolar attractionMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 1 2005Laura L. Tres Abstract Nucleolar organization by autosomal bivalents occurs during male meiotic prophase in mammalian species. During late leptotene,early zygotene stages, several autosomal bivalents are engaged in ribosomal RNA synthesis. At pachytene stage, nucleolar masses detach from the sites of primary autosomal origin, relocate close to the XY chromosomal pair, and nucleolar components become segregated. In early pachytene, an extensive synaptonemal complex at the pseudoautosomal region, links X and Y chromosomes in close juxtaposition along most of the length of the Y chromosome, except for a terminal region of the Y that diverges from the pairing region. As meiotic prophase advances, X and Y chromosomes progressively desynapse and, at diplotene, the XY pair is associated end-to-end. Xmr (Xlr-related, meiosis regulated) is a protein component of the nucleolus associated to the XY pair and of the asynapsed portions of the X and Y axial cores. Xmr, like SCP3, is a component of the lateral element of the synaptonemal complex. Both share structural homology in their C-terminal region. This region contains several putative coiled-coil domains known to mediate heterodimeric protein,protein interactions and to provide binding sites to regulatory proteins. Like Xmr, the tumor repressor protein BRCA1 is present along the unsynapsed cores of the XY bivalent. Both Xmr and BRCA1 have been implicated in a mechanism leading to chromatin condensation and transcription inactivation of the XY bivalent. The BRCA1-ATR kinase complex, as recent research suggests, triggers the phosphorylation of histone H2AX, which predominates in the condensed chromatin of the XY chromosomal pair. Xmr is not present in the XY bivalent when the expression of histone H2AX is deficient. The role of Xmr in chromatin condensation of the XY bivalent has not been determined. The partial structural homology of SCP3 and Xmr, their distribution along the unsynapsed axial cores of the X and Y chromosomes, and the presence of Xmr in the XY pair-associated nucleolus raises the possibility that Xmr, and other proteins including protein kinases, may be recruited to the nucleolus to perform functions related to chromosomal synapsis, chromatin condensation and recombination processes, as well as cell cycle progression. Mol. Reprod. Dev. © 2005 Wiley-Liss, Inc. [source] Configurations of germinal vesicle (GV) chromatin in the goat differ from those of other speciesMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 2 2005Hong-Shu Sui Abstract Configuration of germinal vesicle (GV) chromatin has been studied and found correlated with the developmental competence of oocytes in several mammalian species. A common feature in the configuration of GV chromatin in the species studied so far is that the diffuse chromatin (the so called "NSN" pattern) condenses into a perinucleolar ring (the so called "SN" configuration) with follicular growth. However, no study has been published on the configuration of GV chromatin in the goat. Nor is it known whether the perinucleolar ring of condensed chromatin (CC) in an oocyte represents a step toward final maturation or atresia. Changes in configurations of GV chromatin and RNA synthesis during goat oocyte growth, atresia and maturation in vivo and in vitro were investigated in this study. Based on both the size of nucleoli and the degree of chromatin condensation, the GV chromatin of goat oocytes was classified into GV1 characterized by large nucleoli and diffuse chromatin, GV2 with medium-sized nucleoli and condensed net-like (GV2n) or clumped (GV2c) chromatin, GV3 with small nucleoli and net-like (GV3n) or clumped (GV3c) chromatin, and GV4 with no nucleolus but clumped chromatin. The results showed that (i) the configurations of GV chromatin in the goat differ from those of other species in that the chromatin did not condense into a perinucleolar ring; (ii) most of the goat oocytes are synchronized at the GV3n configuration before GVBD; (iii) the GVn pattern might represent a healthy state, but the GVc an atretic state; (iv) in both goats and mice, the GC-specific (Chromomycin A3, CMA3) and the AT-specific (Hoechst 33342) fluorochromes followed the same pattern of distribution in GV chromatin; (v) the nucleolar size decreased significantly with oocyte growth and maturation in vivo and in vitro; and (vi) goat oocytes began GVBD at 8 hr and had completed it by 20 hr after onset of estrus. The peculiar configuration of GV chromatin of goat oocytes can be a useful model for studies of morphological and functional changes of different nuclear compartments during the cell cycle and cell differentiation, and the functional differentiation between GV3n and GV3c might be used for reference to the question whether the "SN" configuration in other species inclines toward ovulation or atresia. Mol. Reprod. Dev. 71: 227,236, 2005. © 2005 Wiley-Liss, Inc. [source] Several signaling pathways are involved in the control of cattle oocyte maturationMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 4 2004Céline Vigneron Abstract The main limit of in vitro production of domestic mammal embryos comes from the low capacity of in vitro matured oocytes to develop after fertilization. As soon as they are separated from follicular environment, oocytes spontaneously resume meiosis without completion of their terminal differentiation. Roscovitine (ROS), an inhibitor of M-phase promoting factor (MPF) kinase activity reversibly blocks the meiotic resumption in vitro. However, in cattle maturing oocytes several cellular events such as protein synthesis and phosphorylation, chromatin condensation and nuclear envelope folding escape ROS inhibition suggesting the alternative pathways in oocyte maturation. We compared the level of synthesis and phosphorylation of several protein kinases during bovine cumulus oocyte complex (COC) maturation in vitro in the presence or not of epidermal growth factor (EGF) and ROS. We showed that during the EGF-stimulated maturation, ROS neither affected the decrease of EGF receptor (EGFR) nor did inhibit totally its phosphorylation in cumulus cells and also did not totally eliminate tyrosine phosphorylation in oocytes. However, ROS did inhibit the Phosphoinositide 3-kinase (PI3) activity when oocytes mature without EGF. Accumulation of Akt/PKB (protein kinase B), JNK1/2 (jun N-terminal kinases) and Aurora-A in oocytes during maturation was not affected by ROS. However, the phosphorylation of Akt but not JNKs was diminished in ROS-treated oocytes. Thus, PI3 kinase/Akt, JNK1/2 and Aurora-A are likely to be involved in the regulation of bovine oocyte maturation and some of these pathways seem to be independent to MPF activity and meiotic resumption. This complex regulation may explain the partial meiotic arrest of ROS-treated oocytes and the accelerated maturation observed after such treatment. Mol. Reprod. Dev. 69: 466,474, 2004. © 2004 Wiley-Liss, Inc. [source] Caspase activity in newt spermatogonial apoptosis induced by prolactin and cycloheximideMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 2 2001Takashi Yazawa Abstract We previously showed in vivo and in vitro, that among the spermatogenic stages of the newt, prolactin (PRL) induces apoptosis specifically in the penultimate stage of secondary spermatogonia. In the current report, we demonstrate in vitro that cycloheximide (CHX), an inhibitor of protein synthesis, induces morphological apoptotic changes similar to those caused by PRL, such as chromatin condensation and apoptotic body formation. Next, we found that Z-VAD-fmk, an inhibitor of various caspases, suppressed the apoptosis induced by PRL and CHX, but ICE inhibitor Ac-YVAD-CHO or caspase-3 inhibitor Ac-DEVD-CHO did not. As high caspase activity was present in extracts of testes treated with CHX, we suggest that an unidentified caspase induces the morphological changes of apoptosis in newt spermatogonia. Mol. Reprod. Dev. 59:209,214, 2001. © 2001 Wiley-Liss, Inc. [source] The Binding Characteristics and Intracellular Localization of Temoporfin (mTHPC) in Myeloid Leukemia Cells: Phototoxicity and Mitochondrial Damage,PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 4 2000J. Y. Chen ABSTRACT The state of aggregation of the photosensitizer meso -tetrahydroxyphenylchlorin (mTHPC) in both cell free and intracellular environment was elucidated by comparing its absorption and excitation spectra. In methanol, mTHPC existed as monomers and strongly fluoresced. In aqueous solutions such as phosphate-buffered saline (PBS), mTHPC formed nonfluorescent aggregates. Some portion of mTHPC monomerized in the presence of 10% fetal calf serum PBS. In murine myeloid leukemia M1 and WEHI-3B (JCS) cells, cytoplasmic mTHPC were monomeric. By using organelle-specific fluorescent probes, it was found that mTHPC localized preferentially at the mitochondria and the perinuclear region. Photodynamic treatment of mTHPC-sensitized leukemia cells caused rapid appearance of the apoptogenic protein cytochrome c in the cytosol. Results from flow cytometric analysis showed that the release of cytochrome c was especially pronounced in JCS cells, and well correlated with the extent of apoptotic cell death as reported earlier. Electron microscopy revealed the loss of integrity of the mitochondrial membrane and the appearance of chromatin condensation as early as 1 h after light irradiation. We conclude that rapid release of cytochrome c from photodamaged mitochondria is responsible for the mTHPC-induced apoptosis in the myeloid leukemia JCS and M1 cells. [source] Morphological changes of sperm nuclei during spermatogenesis in the brown alga Cystoseira hakodatensis (Fucales, Phaeophyceae)PHYCOLOGICAL RESEARCH, Issue 2 2003Shinya Yoshikawa SUMMARY Morphological changes and chromatin condensation of sperm nuclei were observed during spermatogenesis in the fucalean brown alga Cystoseira hakodatensis (Yendo) Fensholt. Ultrastructural studies have shown that the mature spermatozoid has an elongated and concave nucleus with condensed chromatin. The morphological changes and the chromatin condensation process during spermatogenesis was observed. Nuclear size decreased in two stages during spermatogenesis. During the first stage, spherical nuclei decreased in size as they were undergoing meiotic divisions and the subsequent mitoses within the antheridium. During the second stage, the morphological transformation from a spherical into an elongated nucleus occurred. Afterwards, chromatin condensed at the periphery in each nucleus, and chromatin-free regions were observed in the center of the nucleus. These chromatin-free regions in the center of nucleus were compressed by the peripheral chromatin-condensed region. As the result, the elongated and concave nucleus of the mature sperm consisted of uniformly well-condensed chromatin. [source] Apoptosis of BGC823 cell line induced by p -hydroxymethoxybenzobijuglone, a novel compound from Juglans mandshuricaPHYTOTHERAPY RESEARCH, Issue 4 2009ZhiBo Li Abstract p -Hydroxymethoxybenzobijuglone (HMBBJ), a new quinone compound isolated from Juglans mandshurica (by bioassay-guided fractionation), showed cytotoxic activity in the gastric carcinoma cell line BGC823. The growth of BGC823 cells was inhibited as demonstrated by MTT assay and several cellular characteristic changes, such as cell shrinkage, chromatin condensation and apoptotic body formation with programmed cell death. Flow cytometry analysis revealed that the BGC823 cell cycle was arrested at G2/M phase by HMBBJ, and the apoptotic rate of BGC823 cells increased with respect to HMBBJ in a dose-dependent manner. HMBBJ also activated caspase-3, decreased the expression of Bcl-2 and caused a decrease in the mitochondrial membrane potential (,,m). These findings suggest that HMBBJ could significantly induce apoptosis in BGC823 cells and should be considered as a potential candidate for a chemotherapeutic drug against cancer. Copyright © 2008 John Wiley & Sons, Ltd. [source] | |||||||||||||||||||||||||||||||