Cryopreservation of human semen and prepared sperm: effects on motility parameters and DNA integrity

Objective: To investigate effects of cryopreservation on sperm motility and DNA integrity. Design: Pre-cryopreservation and post-cryopreservation analysis of motility and DNA integrity of semen and prepared sperm samples. Setting: A hospital andrology laboratory. Patient(s): Forty men attending the Regional Fertility Centre, Belfast, Northern Ireland. Intervention(s): Each sample was divided, and an aliquot was frozen unprepared. Remaining aliquots were prepared by Percoll density centrifugation (95.0:47.5) or direct swim-up procedure and divided into aliquots to allow direct comparison of fresh and frozen semen and prepared sperm (frozen with or without the addition of seminal plasma) from the same ejaculate. Samples were frozen by static-phase vapor cooling and being plunged into liquid nitrogen. Thawing was carried out at room temperature. Main Outcome Measure(s): Sperm DNA integrity was determined using a modified alkaline single cell gel electrophoresis (comet) assay, and motility was determined using computer-assisted semen analysis. Result(s): Sperm frozen unprepared in seminal fluid appeared more resistant to freezing damage than frozen prepared sperm. Further improvements can be achieved by selecting out the subpopulation of sperm with best motility and DNA integrity and freezing these sperm in seminal plasma, making this the optimal procedure. Conclusion(s): Freezing sperm in seminal plasma improves postthaw motility and DNA integrity.

ATR-dependent radiation-induced gammaH2AX foci in bystander primary human astrocytes and glioma cells.

Radiotherapy is an important treatment for patients suffering from high-grade malignant gliomas. Non-targeted (bystander) effects may influence these cells' response to radiation and the investigation of these effects may therefore provide new insights into mechanisms of radiosensitivity and responses to radiotherapy as well as define new targets for therapeutic approaches. Normal primary human astrocytes (NHA) and T98G glioma cells were irradiated with helium ions using the Gray Cancer Institute microbeam facility targeting individual cells. Irradiated NHA and T98G glioma cells generated signals that induced gammaH2AX foci in neighbouring non-targeted bystander cells up to 48 h after irradiation. gammaH2AX bystander foci were also observed in co-cultures targeting either NHA or T98G cells and in medium transfer experiments. Dimethyl sulphoxide, Filipin and anti-transforming growth factor (TGF)-beta 1 could suppress gammaH2AX foci in bystander cells, confirming that reactive oxygen species (ROS) and membrane-mediated signals are involved in the bystander signalling pathways. Also, TGF-beta 1 induced gammaH2AX in an ROS-dependent manner similar to bystander foci. ROS and membrane signalling-dependent differences in bystander foci induction between T98G glioma cells and normal human astrocytes have been observed. Inhibition of ataxia telangiectasia mutated (ATM) protein and DNA-PK could not suppress the induction of bystander gammaH2AX foci whereas the mutation of ATM- and rad3-related (ATR) abrogated bystander foci induction. Furthermore, ATR-dependent bystander foci induction was restricted to S-phase cells. These observations may provide additional therapeutic targets for the exploitation of the bystander effect.

A Holocene record of human induced and natural environmental change from Lake Forsyth (Te Wairewa), New Zealand

A 1.2 m sediment core from Lake Forsyth, Canterbury, New Zealand, records the development of the catchment/lake system over the last 7000 years, and its response to anthropogenic disturbance following European settlement c. 1840 AD. Pollen was used to reconstruct catchment vegetation history, while foraminifera, chironomids, Trichoptera, and the abundance of Pediastrum simplex colonies were used to infer past environmental conditions within the lake. The basal 30 cm of core records the transition of the Lake Forsyth Basin from a tidal embayment to a brackish coastal lake. Timing of closure of the lake mouth could not be accurately determined, but it appears that Lake Forsyth had stabilised as a slightly brackish, oligo mesotrophic shallow lake by about 500 years BP. Major deforestation occurred on Banks Peninsula between 1860 AD and 1890 AD. This deforestation is marked by the rapid decline in the main canopy trees (Prumnopitys taxifolia (matai) and Podocarpus totara/hallii (totara/mountain totara), an increase in charcoal, and the appearance of grasses. At around 1895 AD, pine appears in the record while a willow (Salix spp.) appears somewhat later. Redundancy analysis (RDA) of the pollen and aquatic species data revealed a significant relationship between regional vegetation and the abundance of aquatic taxa, with the percentage if disturbance pollen explaining most (14.8%) of the constrained variation in the aquatic species data. Principle components analysis (PCA) of aquatic species data revealed that the most significant period of rapid biological change in the lakes history corresponded to the main period of human disturbance in the catchment. Deforestation led to increased sediment and nutrient input into the lake which was accompanied by a major reduction in salinity. These changes are inferred from the appearance and proliferation of freshwater algae (Pediastrum simplex), an increase in abundance and diversity of chironomids, and the abundance of cases and remains from the larvae of the caddisfly, Oecetis unicolor. Eutrophication accompanied by increasing salinity of the lake is inferred from a significant peak and then decline of P. simplex, and a reduction in the abundance and diversity of aquatic invertebrates. The artificial opening of the lake to the Pacific Ocean, which began in the late 1800s, is the likely cause of the recent increase in salinity. An increase in salinity may have also encouraged blooms of the halotolerant and hepatotoxic cyanobacteria Nodularia spumigena.

New Zealand chironomids as proxies for human-induced and natural environmental change: Transfer functionsfor temperature and lake production (chlorophyll a)

The analysis of chironomid taxa and environmental datasets from 46 New Zealand lakes identified temperature (February mean air temperature) and lake production (chlorophyll a (Chl a)) as the main drivers of chironomid distribution. Temperature was the strongest driver of chironomid distribution and consequently produced the most robust inference models. We present two possible temperature transfer functions from this dataset. The most robust model (weighted averaging-partial least squares (WA-PLS), n = 36) was based on a dataset with the most productive (Chl a > 10 lg l)1) lakes removed. This model produced a coefficient of determination (r2 jack) of 0.77, and a root mean squared error of prediction (RMSEPjack) of 1.31C. The Chl a transfer function (partial least squares (PLS), n = 37) was far less reliable, with an r2 jack of 0.49 and an RMSEPjack of 0.46 Log10lg l)1. Both of these transfer functions could be improved by a revision of the taxonomy for the New Zealand chironomid taxa, particularly the genus Chironomus. The Chironomus morphotype was common in high altitude, cool, oligotrophic lakes and lowland, warm, eutrophic lakes. This could reflect the widespread distribution of one eurythermic species, or the collective distribution of a number of different Chironomus species with more limited tolerances. The Chl a transfer function could also be improved by inputting mean Chl a values into the inference model rather than the spot measurements that were available for this study.

Human Papillomavirus Type 16 E6 and E7 Cause Polyploidy in Human Keratinocytes and Up-Regulation of G2-M-phase Proteins

Human papillomavirus type 16 proteins E6 and E7 have been shown to cause centrosome amplification and lagging chromosomes during mitosis. These abnormalities during mitosis can result in missegregation of the chromosomes, leading to chromosomal instability. Genomic instability is thought to be an essential part of the conversion of a normal cell to a cancer cell. We now show that E6 and E7 together cause polyploidy in primary human keratinocytes soon after these genes are introduced into the cells. Polyploidy seems to result from a spindle checkpoint failure arising from abrogation of the normal functions of p53 and retinoblastoma family members by E6 and E7, respectively. In addition, E6 and E7 cause deregulation of cellular genes such as Plk1, Aurora-A, cdk1, and Nek2, which are known to control the G2-M-phase transition and the ordered progression through mitosis.

Area factors and suicide: 5-year follow-up of the Northern Ireland population

Background: Suicide rates vary markedly between areas but it is unclear whether this is due to differences in population composition or to contextual factors operating at an area level.
Aims: To determine if area factors are independently related to suicide risk after adjustment for individual and family characteristics.
Method: A 5-year record linkage study was conducted of 1 116 748 non-institutionalised individuals aged 16-74 years, enumerated at the 2001 Northern Ireland census.
Results: The cohort experienced 566 suicides during follow-up. Suicide risks were lowest for women and for those who were married or cohabiting. Indicators of individual and household disadvantage and economic and health status at the time of the census were also strongly related to risk of suicide. The higher rates of suicide in the more deprived and socially fragmented areas disappeared after adjustment for individual and household factors. There was no significant relationship between population density and risk of suicide.
Conclusions: Differences in rates of suicide between areas are predominantly due to population characteristics rather than to area-level factors, which suggests that policies targeted at area-level factors are unlikely to significantly influence suicides rates.