Morphine exposure prevents up-regulation of MR and GR binding sites in the brain of adult male and female rats due to prenatal stress

Our previous work demonstrated that the hormone response to stress and the negative feedback inhibition to these hormones are sex-dependently altered by prenatal morphine exposure in adult rats. An alteration in the glucocorticoid negative feedback inhibition is mediated by glucocorticoid receptors (GR) that are distributed throughout the brain, and mineralocorticoid receptors (MR) localized mainly in the hippocampus and involved in a tonic influence of brain functions. Therefore, the present study examined the binding characteristics of MR and GR in young adult male and female rats exposed prenatally (E11-E18) to morphine (10 mg/kg/2 x /day), saline or no treatment at all (controls). At 60-90 days of age, animals were adrenalectomized (ADX) 24 h prior to decapitation. The hippocampus and hypothalamus were dissected for saturation binding assays. The data demonstrate that prenatal stress due to maternal saline injections up-regulates MR and GR binding in the hippocampus of adult male rats and this effect is prevented by prenatal morphine exposure. There is no effect of prenatal morphine exposure on GR binding in the hypothalamus of males. In female rats, prenatal morphine exposure does not affect the binding of MR and GR in the hippocampus or GR in the hypothalamus relative to controls; however, they are affected by ovarian hormone fluctuation. Moreover, prenatal stress decreases MR binding in the hippocampus of diestrous females and GR binding in the hypothalamus of estrous females. Both decreases are prevented by prenatal morphine exposure. Thus, the present study demonstrates that: (1) prenatal stress due to maternal saline injections alters MR and GR binding of adult male and female rats and is prevented by prenatal morphine exposure; (2) the MR and GR binding in adult female rats are affected by ovarian hormone fluctuations.     

Elements of psychological support in nursing care

The paper presents the results of a pilot study which involved 50 nurses from several departments of internal medicine of The State Clinical Hospital of the Collegium Medicum, Jagiellonian University in Cracow. The results are based on the Statement Response Questionnaire. They show that the most common responses of the nurses in the face of anxiety expressed by patients are cheering up the patient, collecting information about the symptom, and offering explanation of the symptom. The least common responses included expressing one's own positive emotions and showing empathy towards the patient.     

De novo inverted duplication 9p21pter involving telomeric repeated sequences

We report on clinical and cytogenetic findings in a boy with partial 9p duplication, dup(9)(p21pter). Clinical manifestations included facial and hand anomalies and mental retardation. Fluorescence in situ hybridization (FISH) and comparative genomic hybridization (CGH) were used to characterize further and confirm the conventional banding data. Investigation by FISH using whole chromosome 9 paint probe showed that the additional material was derived from chromosome 9. Using CGH, a region of gain was found in the chromosome segment 9p21pter. YACs and telomeric probes confirmed the duplicated region. Using the all-human telomeric sequences probe, intrachromosomal telomeric signal was noted on the short arm of the abnormal chromosome 9. Mechanism of formation of the duplication, including intrachromosomal telomeric sequences, is discussed.     

Relative carriage rates of nuclear dehydrogenating clostridia in two populations of different colorectal cancer risk

Carriage of nuclear dehydrogenating clostridia has been associated with colon cancer and implicated in its aetiology. This study has compared the carriage of these organisms in a British population at high risk for the development of colon cancer with a low risk Nigerian population. Clostridia were found in all of the stools from both populations. Nuclear dehydrogenating clostridia were only found in the stools of the British subjects (32%). These results support the suggestion that the carriage rate of nuclear dehydrogenating clostridia in a population is related to the risk of colon cancer.     

Development of hatching blastocysts from immature human oocytes following in-vitro maturation and fertilization using a co-culture system

Recently, in-vitro maturation (IVM) of immature human oocytes recovered from non-stimulated follicles has been applied in the treatment of infertility. However, in previous reports, very few embryos cultured in conventional medium have reached the expanded blastocyst stage following in-vitro maturation and fertilization (IVM/IVF). The objective of this study was to investigate whether the developmental competence of human embryos following IVM/IVF could be enhanced by the use of a human ampullary cell co-culture system. Immature human oocytes were aspirated from small follicles at Caesarean section and then cultured in medium containing human menopausal gonadotrophin for 36 to 48 h, followed by insemination. Zygotes were randomly cultured either in conventional culture medium alone or in the co-culture system. Of 48 embryos cultured in conventional medium alone, all arrested at the 2-16- cell stage on day 3 after insemination. Of 46 embryos cultured in the co-culture system, 26 embryos (56.5%) arrested at the 2-16-cell stage. Six embryos (13%) developed to the morula stage. Fourteen embryos (30.4%) developed to expanded blastocysts and two blastocysts were hatching on day 7 after insemination. We conclude that co-culture significantly enhances the development of blastocysts in embryos resulting from IVM/IVF.      

Detection of chromosomes and estimation of aneuploidy in human spermatozoa using fluorescence in-situ hybridization

The development and application of fluorescence in-situ hybridization (FISH) has opened the way for comprehensive studies on numerical chromosome abnormalities in human spermatozoa. FISH can be rapidly applied to large numbers of spermatozoa and thus overcomes the major limitation of karyotyping spermatozoa after penetration of zona-free hamster oocytes. The simultaneous hybridization of two or more chromosome-specific probes to spermatozoa and subsequent detection of the bound probes using different fluorescent detection systems enables two or more chromosomes to be localized simultaneously in the same spermatozoon and provides a technique for undertaking reasonable estimates of aneuploidy. The most commonly used probes are those which bind to the centromeric region of specific chromosomes. Most studies to date have concentrated on estimating aneuploidy in spermatozoa from normospermic men, although reports are beginning to appear on aneuploidy in spermatozoa from subfertile and infertile men. Multi- probe FISH studies have generally reported disomy (hyperhaploidy) estimates of 0.05-0.2% per chromosome. There is preliminary evidence that some chromosomes such as X, Y and 21 are predisposed towards higher rates of non-disjunction during spermatogenesis. There are also suggestions of inter-donor variability in aneuploidy frequencies for specific chromosomes, although this requires confirmation in larger studies. While FISH is clearly a powerful technique that has many applications in reproductive medicine, it must also be realized that it does have limitations and the technology itself is still evolving and has yet to be fully validated on spermatozoa.      

Identical abnormality of the short arm of chromosome 18 in two Philadelphia-positive chronic myelocytic leukemia patients with erythroblastic transformation,resulting in duplication of BCR-ABL1 fusion

Two patients with Ph-positive chronic myelocytic leukemia in erythroblastic transformation and rearrangement of the short arm of chromosome 18 are reported. Fluorescence in situ hybridization studies showed that the 18p rearrangement resulted from translocation of the main part of chromosome 22 long arm to 18p, including BCR-ABL1 fusion. The 18p abnormality resulted, thus, in loss of 18p and duplication of BCR-ABL1 in both patients. The possible relation to the erythroblastic type of blastic phase is briefly discussed. In addition an apparently intact germline ABL1 gene was duplicated and inserted into chromosome 6 at band p21 in one of these patients.     

Preliminary observations on polar body extrusion and pronuclear formation in human oocytes using time-lapse video cinematography

In this study, we have used time-lapse video cinematography to study fertilization in 50 human oocytes that had undergone intracytoplasmic sperm injection (ICSI). Time-lapse recording commenced shortly after ICSI and proceeded for 17-20 h. Oocytes were cultured in an environmental chamber which was maintained under standard culture conditions. Overall, 38 oocytes (76%) were fertilized normally, and the fertilization rate and embryo quality were not significantly different from 487 sibling oocytes cultured in a conventional incubator. Normal fertilization followed a defined course of events, although the timing of these events varied markedly between oocytes. In 35 of the 38 fertilized oocytes (92%), there were circular waves of granulation within the ooplasm which had a periodicity of 20-53 min. The sperm head decondensed during this granulation phase. The second polar body was then extruded, and this was followed by the central formation of the male pronucleus. The female pronucleus formed in the cytoplasm adjacent to the second polar body at the same time as, or slightly after, the male pronucleus, and was subsequently drawn towards the male pronucleus until the two abutted. Both pronuclei then increased in size, the nucleoli moved around within the pronuclei and some nucleoli coalesced. During pronuclear growth, the organelles contracted from the cortex towards the centre of the oocyte, leaving a clear cortical zone. The oocyte decreased in diameter from 112 to 106 microm (P < 0.0001) during the course of the observation period. The female pronucleus was significantly smaller in diameter than the male pronucleus (24.1 and 22.4 microm respectively, P = 0.008) and contained fewer nucleoli (4.2 and 7.0 respectively, P < 0.0001). After time-lapse recording, oocytes were cultured for 48 h prior to embryo transfer or cryopreservation. Embryo quality was related to fertilization events and periodicity of the cytoplasmic wave, and it was found that good quality embryos arose from oocytes that had more uniform timing from injection to pronuclear abuttal and tended to have a longer cytoplasmic wave. In conclusion, we have shown that time-lapse video cinematography is an excellent tool for studying fertilization and early embryo development, and have demonstrated that human fertilization comprises numerous complex dynamic events.