Fine structure and probable function of ring organs in the mite Histiostoma feroniarum (Acari: Actinotrichida: Acaridida: Histiostomatidae)

Histiostoma feroniarum, like other histiostomatid mites, possesses peculiar ring organs that are visible under the light microscope as ventrally located, characteristic rings of sclerotized cuticle. The ring organ is composed of three elements: a disc of modified cuticle, ring organ cells located underneath the disc, and an "empty" chamber frequently visible between the cuticular disc and the cells. The cuticle of the disc is not perforated and differs from the surrounding unmodified cuticle as revealed by special staining developed for light microscopy and by electron microscopy. The ring organ cells show a polarity, with a practically smooth apical surface and an extremely folded basal membrane. The basal invaginations reach the apical cell portion, where they form tubular canaliculi distributed beneath the apical cell membrane. The cytoplasm contains many mitochondria, which are usually in contact with the cell membrane invaginations. Structurally, the ring organ cells closely resemble the transport cells described in osmoregulatory organs both in water-inhabiting and terrestrial arthropods. Thus, our results support earlier suggestions of an osmoregulatory function performed by sclerotized rings (=ring organs), as an adaptation to aqueous environments. A possible homology with similar organs of other mites is discussed.     

Hypoxia induces mitochondrial DNA damage and stimulates expression of a DNA repair enzyme, the Escherichia coli MutY DNA glycosylase homolog (MYH), in vivo, in the rat brain

Hypoxia-associated, acutely reduced blood oxygenation can compromise energy metabolism, alter oxidant/antioxidant balance and damage cellular components, including DNA. We show in vivo, in the rat brain that respiratory hypoxia leads to formation of the oxidative DNA lesion, 8-hydroxy-2'-deoxyguanosine (oh8dG), a biomarker for oxidative DNA damage and to increased expression of a DNA repair enzyme involved in protection of the genome from the mutagenic consequences of oh8dG. The enzyme is a homolog of the Escherichia coli MutY DNA glycosylase (MYH), which excises adenine residues misincorporated opposite the oxidized base, oh8dG. We have cloned a full-length rat MYH (rMYH) cDNA, which encodes 516 amino acids, and by in situ hybridization analysis obtained expression patterns of rMYH mRNA in hippocampal, cortical and cerebellar regions. Ensuing hypoxia, mitochondrial DNA damage was induced and rMYH expression strongly elevated. This is the first evidence for a regulated expression of a DNA repair enzyme in the context of respiratory hypoxia. Our findings support the premise that oxidative DNA damage is repaired in neurons and the possibility that the hypoxia-induced expression of a DNA repair enzyme in the brain represents an adaptive mechanism for protection of neuronal DNA from injurious consequences of disrupted energy metabolism and oxidant/antioxidant homeostasis.     

Synthesis and biological activity of N(alpha)-[4-[N-[(3,4-dihydro-2-methyl-4-oxo-6-quinazolinyl)methyl]-N-propargylamino]phenylacetyl]-L-glutamic acid

2-Deamino-2-methyl-N10-propargyl-5,8-dideazafolic acid (ICI 198583) is a potent inhibitor of thymidylate synthase. Its analogue, N(alpha)-[4-[N-[(3,4-dihydro-2-methyl-4-oxo-6-quinazolinyl)methyl]-N-propargylamino]phenylacetyl]-L-glutamic acid, containing p-aminophenylacetic acid residue substituting p-aminobenzoic acid residue, was synthesized. The new analogue exhibited a moderately potent thymidylate synthase inhibition, of linear mixed type vs. the cofactor, N(5,10)-methylenetetrahydrofolate. The Ki value of 0.34 microM, determined with a purified recombinant rat hepatoma enzyme, was about 30-fold higher than that reported for inhibition of thymidylate synthase from mouse leukemia L1210 cells by ICI 198583 (Hughes et al., 1990, J. Med. Chem. 33, 3060). Growth of mouse leukemia L5178Y cells was inhibited by the analogue (IC50 = 1.26 mM) 180-fold weaker than by ICI 198583 (IC50 = 6.9 microM).     

Complex transgene locus structures implicate multiple mechanisms for plant transgene rearrangement

To more fully characterize the internal structure of transgene loci and to gain further understanding of mechanisms of transgene locus formation, we sequenced more than 160 kb of complex transgene loci in two unrelated transgenic oat (Avena sativa L.) lines transformed using microprojectile bombardment. The transgene locus sequences from both lines exhibited extreme scrambling of non-contiguous transgene and genomic fragments recombined via illegitimate recombination. A perfect direct repeat of the delivered DNA, and inverted and imperfect direct repeats were detected in the same transgene locus indicating that homologous recombination and synthesis-dependent mechanism(s), respectively, were also involved in transgene locus rearrangement. The most unexpected result was the small size of the fragments of delivered and genomic DNA incorporated into the transgene loci via illegitimate recombination; 50 of the 82 delivered DNA fragments were shorter than 200 bp. Eleven transgene and genomic fragments were shorter than the DNA lengths required for Ku-mediated non-homologous end joining. Detection of these small fragments provided evidence that illegitimate recombination was most likely mediated by a synthesis-dependent strand-annealing mechanism that resulted in transgene scrambling. Taken together, these results indicate that transgene locus formation involves the concerted action of several DNA break-repair mechanisms.     

Evaluation of mouse preimplantation embryos exposed to oxidative stress cultured with insulin-like growth factor I and II, epidermal growth factor, insulin, transferrin and selenium

Blastocyst culture requires strictly defined culture media to sustain its viability and quality. Although blastocyst media are commercially available, they do not meet all the needs and research focused on blastocyst-promoting agents is on the way. The aims of the study were to evaluate the significance of insulin-like growth factors I (IGF-I) and II (IGF-II); epidermal growth factor (EGF) and a mixture of insulin, transferrin and selenium (ITS) on the development of embryos exposed to oxidative stress. C3B6F1 mice were stimulated with 5 IU of pregnant mare serum gonadotropin following by administration of 5 IU of equine chorionic gonadotropin and mating with DBA males. The mice were killed 40 h after eCG injection by cervical dislocation and then the 2 cell embryos were flushed out from the fallopian tubes. To evaluate whether the growth factors may compensate the unfavorable--oxidative milieu created by hydrogen peroxide (H2O2), the embryos were transferred to 1/ control medium, 2/ control medium+0.1 mM (H2O2) or 3/ control medium+H2O2 enriched with 10(-7) g/ml of IGF-I, IGF-II, EGF or a mixture of insulin (5x10(-6) g/ml), transferrin (5 x10(-6) g/ml) and selenium (5x10(-9) g/ml; ITS). Embryos were evaluated 96-144 hours following eCG injection. In the study the dynamics of embryo development and blastocyst cell numbers (including inner cell mass) were assessed. The morphological evaluation comprised viability and apoptosis (TUNEL). In oxidative stress setting, IGF-I, IGF-II, EGF and ITS minimized the negative influence of H2O2, and embryos developed faster than in control conditions. Blastocysts cultured with hydrogen peroxide and growth factors or ITS displayed normal morphology and had more cells--also within the inner cell mass--than those treated only with H2O2. The positive TUNEL reactions were sporadically observed in embryos cultured with hydrogen peroxide supplemented with growth factors. IGF-I, IGF-II, EGF and ITS have a positive effect on pre-implantation embryo development in detrimental culture conditions of oxidative stress.     

Resistance to wheat streak mosaic virus in transgenic wheat engineered with the viral coat protein gene

Wheat (Triticum aestivum) plants were stably transformed with the coat protein (CP) gene of wheat streak mosaic virus (WSMV) by the biolistic method. Eleven independently transformed plant lines were obtained and five were analyzed for gene expression and resistance to WSMV. One line showed high resistance to inoculations of two WSMV strains. This line had milder symptoms and lower virus titer than control plants after inoculation. After infection, new growth did not show symptoms. The observed resistance was similar to the recovery type resistance described previously using WSMV NIb transgene and in other systems. This line looked morphologically normal but had an unusually high transgene copy number (approximately 90 copies per 2C homozygous genome). Northern hybridization analysis indicated a high level of degraded CP mRNA expression. However, no coat protein expression was detected.     

The disturbances of the zinc concentration in the blood in selected neoplasms

A study was carried out on 92 patients (58 males and 34 females) aged 42–76 treated for malignant neoplasm of the gastrointestinal tract (54 patients with colorectal carcinoma, 38 with gastric carcinoma). In all patients, the zinc serum concentration was measured and the results obtained were referred to some epidemiological-clinical factors (sex, age, primary cause of cancer, the stage of clinical progression, and histological type). The results showed that the most pronounced hypozincemia occurred in male patients with mucous membrane carcinoma of the stomach.     

An approach to diagnosis of T-cell lymphoproliferative disorders by flow cytometry

T-cell lymphoproliferative disorders are among the most challenging diagnoses in hematopathology. Unlike the more common B-cell disorders, in which clonality is often readily discernible by surface immunoglobulin light chain restriction, there is no specific immunophenotypic signature that is diagnostic of a clonal T-cell population. Immunophenotypic criteria that are helpful in the diagnosis of T-cell neoplasms include T-cell subset antigen restriction, anomalous T-cell subset antigen expression, deletion or diminution of one of the pan T-cell antigens, a precursor T-cell phenotype, and expression of additional markers (e.g., CD30, CD20, major myeloid antigens, and TCRgammadelta). Analysis of the inherent forward and orthogonal light scatter properties of the cell can also provide important diagnostic clues. None of these features is 100% specific, however, for aberrant expression of pan-T antigens may be seen in viral infections, B-cell malignancies, or in reactive changes following administration of certain medications. An increased CD4:CD8 ratio is often observed in Hodgkin's lymphoma. Based on the analysis of 87 neoplastic and 80 control cases, we conclude that flow cytometric features that are most suspicious for malignancy include the loss or markedly dim expression of CD45; complete loss of one or more pan-T antigens; diminished expression of more than two pan-T antigens in conjunction with altered light scatter properties; and CD4/CD8 dual-positive or dual-negative expression (except thymic lesions).     

The Roman high- and low-avoidance rats respond differently to novelty in a familiarized environment

The purpose of the present study was to investigate the response to novelty in Roman high- and low-avoidance rats under non-stressful conditions. To reduce fear, a procedure of repetitive placing in the experimental chamber consisting of start, screen, and tunnel zones was applied. Each animal was placed in the experimental chamber daily for a 6min period. The first 11 sessions were the habituation sessions. In the 12th session, the novelty was introduced into the screen and tunnel zones. The subsequent two sessions were conducted under novelty conditions. Behavioral activities such as walking, object contacts, time spent in given zones, and entering the tunnels were measured. All the comparisons were made for two 3-min intervals, using a three-factor MANOVA, involving 2(sex)x2(subline)x8(3-mininterval). All subjects increased time spent in the tunnel zone, but RHA/Verh rats responded to a greater extent, especially the males. All subjects spent shorter times in the screen zone, but the RHA/Verh rats responded to a much greater extent. RHA/Verh rats, especially males responded with a substantial increase of time spent inside the tunnels. All subjects responded with an increased amount of object contacts. In general the RHA/Verh subjects showed a more pronounced response to novelty, as evidenced by a significant shift toward the tunnel zone. They spent more time in this zone than their RLA/Verh counterparts. Among the RLA/Verh rats, males tended to behave similarly to RHA/Verh rats, especially during the second 3min interval of session "12." The differences between the rat lines obtained in this study may be attributed to mechanisms specific to exploration, making them promising subjects to study the relationships between reactivity, novelty detection, adaptation, and environmental information processing.