Paternally transmitted chromosomal aberrations in mouse zygotes determine their embryonic fate

The developmental consequences of chromosomal aberrations in embryos include spontaneous abortions, morphological defects, inborn abnormalities, and genetic/chromosomal diseases. Six germ-cell mutagens with different modes of action and spermatogenic stage sensitivities were used to investigate the relationship between the types of cytogenetic damage in zygotes with their subsequent risk of postimplantation death and of birth as a translocation carrier. Independent of the mutagen used, over 98% of paternally transmitted aberrations were chromosome type, rather than chromatid type, indicating that they were formed during the period between exposure of male germ cells and initiation of the first S phase after fertilization. There were consistent one-to-one agreements between the proportions of a) zygotes with unstable aberrations and the frequencies of dead embryos after implantation (slope = 0.87, confidence interval [CI]: 0.74, 1.16) and b) zygotes with reciprocal translocations and the frequency of translocation carriers at birth (slope = 0.74, CI: 0.48, 2.11). These findings suggest that chromosomal aberrations in zygotes are highly predictive of subsequent abnormal embryonic development and that development appears to proceed to implantation regardless of the presence of chromosomal abnormalities. Our findings support the hypothesis that, for paternally transmitted chromosomal aberrations, the fate of the embryo is already set by the end of G1 of the first cell cycle of development.     

Quarantine strategies for olive fruit fly (Diptera: Tephritidae): low-temperature storage, brine, and host relations

A dose-response relationship was not observed in olive fruit fly, Bactrocera oleae (Gmelin), larvae exposed to acetic acid concentrations (0-2.5%) used in commercial brine solutions to cure olives. Immersion in a 1% acetic acid brine solution impeded emergence of the immature stages. A 1-wk exposure of olives infested with olive fruit fly larvae to low-temperature storage as a postharvest treatment at 0-1 degree C resulted in 8% survival of the population, and exposures of 2 through 5 wk further reduced pupal and adult emergence to <1.0%. One- to 2-wk exposures at 2-3 degrees C resulted in a significant decrease in survival from 20 to 3%, respectively, and longer durations of 3-5 wk reduced survival to <1.0%. Mean daily fruit pulp temperatures in olives in the top, middle, and bottom of plastic bins stored at 2-3 degrees C decreased by 5-8 degrees C from the first to the second day. Lowest temperatures were observed in the top, and highest temperatures were observed in the middle layer of fruit, which attained a mean temperature of 3.8 degrees C on day 5. Laboratory choice tests showed that olive fruit fly oviposited at a higher rate in late season Mission olives that were green than in fruit that were in the red blush maturity stage in tests with 1- and 3-4-d exposure periods, and an increase in duration of exposure was related to an increase in the total number of ovipositional sites. Higher percentages of olive fruit fly third instars, pupae, and adults were reared from green fruit than from fruit in the red blush stage after a 1-d exposure to oviposition. Manzanillo olives were more attractive for oviposition by olive fruit fly than Mission olives, and significantly more third instars, pupae, and adults developed in Manzanillo fruit than in Mission fruit in the red blush stage. These differences were related to the better quality and higher flesh content of the Manzanillo versus Mission olives used in the tests.     

Functional ability of the elderly in institutional and non-institutional care in Croatia

Gerontology-public health indicators of functional ability of the elderly in institutional and non-institutional health care in Croatia were determined by use of expert methodology developed at Department of Gerontology, Dr. Andrija Stampar Institute of Public Health in Zagreb, with the aim to upgrade the Program of Health Care Measures and Procedures in Health Care of the Elderly. Comparison of functional ability between the users of selected Old People's Homes (institutional care; N = 5030) and Gerontology Centers (non-institutional care; N = 2112) yielded highest between-group difference in the proportion of "fully movable" and "fully independent" categories in favor of the latter, thus steering the program of health care for the elderly accordingly. In addition, study results showed greater difference in the proportion of categories describing mental status of institutional and non-institutional care users as compared with the categories describing their physical status, suggesting that mental status plays a more important role than physical status in the geriatric user's stay in non-institutional care versus institutional care. This issue requires additional studies. The results obtained by this indicator analysis pointed to the preventive and geroprophylactic measures to ensure efficient health care for the elderly and to develop the program of mental health promotion and preservation. According to 2007 estimate, there were 759,318 (16.9%) persons aged > or = 65 in Croatia. Data collected at gerontology database kept at Department of Gerontology, Dr. Andrija Stampar Institute of Public Health (September 2008) showed 2% of the elderly (N = 14807) to be accommodated at Old People's Homes, which is below the European average of 4%.     

Pannexin1 and Pannexin2 Channels Show Quaternary Similarities to Connexons and Different Oligomerization Numbers from Each Other

Pannexins are homologous to innexins, the invertebrate gap junction family. However, mammalian pannexin1 does not form canonical gap junctions, instead forming hexameric oligomers in single plasma membranes and intracellularly. Pannexin1 acts as an ATP release channel, whereas less is known about the function of Pannexin2. We purified cellular membranes isolated from MDCK cells stably expressing rat Pannexin1 or Pannexin2 and identified pannexin channels (pannexons) in single membranes by negative stain and immunogold labeling. Protein gel and Western blot analysis confirmed Pannexin1 (Panx1) or Pannexin2 (Panx2) as the channel-forming proteins. We expressed and purified Panx1 and Panx2 using a baculovirus Sf9 expression system and obtained doughnut-like structures similar to those seen previously in purified connexin hemichannels (connexons) and mammalian membranes. Purified pannexons were comparable in size and overall appearance to Connexin46 and Connexin50 connexons. Pannexons and connexons were further analyzed by single-particle averaging for oligomer and pore diameters. The oligomer diameter increased with increasing monomer molecular mass, and we found that the measured oligomeric pore diameter for Panxs was larger than for Connexin26. Panx1 and Panx2 formed active homomeric channels in Xenopus oocytes and in vitro vesicle assays. Cross-linking and native gels of purified homomeric full-length and a C-terminal Panx2 truncation mutant showed a banding pattern more consistent with an octamer. We purified Panx1/Panx2 heteromeric channels and found that they were unstable over time, possibly because Panx1 and Panx2 homomeric pannexons have different monomer sizes and oligomeric symmetry from each other.     

Oxytocin receptor binding in the hypothalamus during gestation in rats

Central oxytocin receptors (OTR) may be involved in adaptations of the brain oxytocin (OT) system during gestation, which are critical for systemic release of OT during parturition and lactation. We used quantitative autoradiography to determine changes in OTR binding in numerous brain sites during the course of gestation in the rat. Furthermore, to evaluate the importance of ovarian steroids in mediating pregnancy-related changes in OTR binding, we measured binding in ovariectomized animals treated with progesterone and/or estrogen, and in pregnant animals treated with exogenous progesterone during late gestation. We found that OTR binding was significantly increased in the paraventricular nucleus (PVN) and supraoptic nucleus (SON) by midgestation (day 15) compared with control. In addition, there was a further significant increase in OTR binding in these nuclei by late gestation (day 20). The bed nucleus of the stria terminalis (BNST) and the medial preoptic area (MPOA) also showed significant gestation-associated increases in OTR binding, which were similar during mid- and late pregnancy. Treatment with exogenous progesterone throughout pregnancy did not alter the increase in OTR binding characteristic of late gestation in any of these brain sites. Finally, estrogen treatment in ovariectomized animals resulted in increased OTR binding in the SON, BNST, and MPOA, but not the PVN. These data demonstrate that OTR binding in the hypothalamus is increased during mid- and late-gestation, compared with ovariectomized control animals, which may be mediated by increased estradiol.     

Identification of a novel human granzyme B inhibitor secreted by cultured sertoli cells

Sertoli cells have long since been recognized for their ability to suppress the immune system and protect themselves as well as other cell types from harmful immune reaction. However, the exact mechanism or product produced by Sertoli cells that affords this immunoprotection has never been fully elucidated. We examined the effect of mouse Sertoli cell-conditioned medium on human granzyme B-mediated killing and found that there was an inhibitory effect. We subsequently found that a factor secreted by Sertoli cells inhibited killing through the inhibition of granzyme B enzymatic activity. SDS-PAGE analysis revealed that this factor formed an SDS-insoluble complex with granzyme B. Immunoprecipitation and mass spectroscopic analysis of the complex identified a proteinase inhibitor, serpina3n, as a novel inhibitor of human granzyme B. We cloned serpina3n cDNA, expressed it in Jurkat cells, and confirmed its inhibitory action on granzyme B activity. Our studies have led to the discovery of a new inhibitor of granzyme B and have uncovered a new mechanism used by Sertoli cells for immunoprotection.     

GenePro: a Cytoscape plug-in for advanced visualization and analysis of interaction networks

MOTIVATION: Analyzing the networks of interactions between genes and proteins has become a central theme in systems biology. Versatile software tools for interactively displaying and analyzing these networks are therefore very much in demand. The public-domain open software environment Cytoscape has been developed with the goal of facilitating the design and development of such software tools by the scientific community. RESULTS: We present GenePro, a plugin to Cytoscape featuring a set of versatile tools that greatly facilitates the visualization and analysis of protein networks derived from high-throughput interactions data and the validation of various methods for parsing these networks into meaningful functional modules. AVAILABILITY: The GenePro plugin is available at the website http://genepro.ccb.sickkids.ca.     

Incision of trivalent chromium [Cr(III)]-induced DNA damage by Bacillus caldotenax UvrABC endonuclease

Some hexavalent chromium [Cr(VI)]-containing compounds are lung carcinogens. Once within cells, Cr(VI) is reduced to trivalent chromium [Cr(III)] which displays an affinity for both DNA bases and the phosphate backbone. A diverse array of genetic lesions is produced by Cr including Cr–DNA monoadducts, DNA interstrand crosslinks (ICLs), DNA–Cr–protein crosslinks (DPCs), abasic sites, DNA strand breaks and oxidized bases. Despite the large amount of information available on the genotoxicity of Cr, little is known regarding the molecular mechanisms involved in the removal of these lesions from damaged DNA. Recent work indicates that nucleotide excision repair (NER) is involved in the processing of Cr–DNA adducts in human and rodent cells. In order to better understand this process at the molecular level and begin to identify the Cr–DNA adducts processed by NER, the incision of CrCl₃ [Cr(III)]-damaged plasmid DNA was studied using a thermal-resistant UvrABC NER endonuclease from Bacillus caldotenax (Bca). Treatment of plasmid DNA with Cr(III) (as CrCl₃) increased DNA binding as a function of dose. For example, at a Cr(III) concentration of 1 μM we observed 2 Cr(III)–DNA adducts per plasmid. At this same concentration of Cr(III) we found that 17% of the plasmid DNA contained ICLs (0.2 ICLs/plasmid). When plasmid DNA treated with Cr(III) (1 μM) was incubated with Bca UvrABC we observed 0.8 incisions/plasmid. The formation of endonuclease IV-sensitive abasic lesions or Fpg-sensitive oxidized DNA bases was not detected suggesting that the incision of Cr(III)-damaged plasmid DNA by UvrABC was not related to the generation of oxidized DNA damage. Taken together, our data suggest that a sub-fraction of Cr(III)–DNA adducts is recognized and processed by the prokaryotic NER machinery and that ICLs are not necessarily the sole lesions generated by Cr(III) that are substrates for NER.