U.S. lung cancer mortality and declining cigarette tobacco consumption

The usual method for estimating population exposure to cigarette tobacco has been annual per capita consumption of cigarettes, expressed as pack-years or numbers of cigarettes consumed. This technique is shown to result in an inaccurate estimate of exposure. It underestimates by 11 years the latency period from peak exposure to peak mortality. Over the years, cigarettes changed markedly. Filters came into prominence and tobacco was "fluffed". On average, tobacco content of cigarettes decreased 39.1% from 1953 to 1981. National per capita consumption of cigarette tobacco declined by 43%. Total exposure to cigarette tobacco has been declining for males for approximately 35 years; for females for 20 years. As of 1982, the secular trend for lung cancer mortality was declining for women below age 45 and for most age groups of men below 65. We appear to be at the threshold of a reversal in overall lung cancer mortality.     

Predominance of null mutations in ataxia-telangiectasia

Ataxia-telangiectasia (A-T) is an autosomal recessive disorder involving cerebellar degeneration, immunodeficiency, chromosomal instability, radiosensitivity and cancer predisposition. The responsible gene, ATM, was recently identified by positional cloning and found to encode a putative 350 kDa protein with a Pl 3-kinase-like domain, presumably involved in mediating cell cycle arrest in response to radiation-induced DNA damage. The nature and location of A-T mutations should provide insight into the function of the ATM protein and the molecular basis of this pleiotropic disease. Of 44 A-T mutations identified by us to date, 39 (89%) are expected to inactivate the ATM protein by truncating it, by abolishing correct initiation or termination of translation, or by deleting large segments. Additional mutations are four smaller in-frame deletions and insertions, and one substitution of a highly conserved amino acid at the Pl 3-kinase domain. The emerging profile of mutations causing A-T is thus dominated by those expected to completely inactivate the ATM protein. ATM mutations with milder effects may result in phenotypes related, but not identical, to A-T.      

High throughput parallel analysis of hundreds of patient samples for more than 100 mutations in multiple disease genes

As more mutations are identified in genes of known sequence, there is a crucial need in the areas of medical genetics and genome analysis for rapid, accurate and cost-effective methods of mutation detection. We have developed a multiplex allele-specific diagnostic assay (MASDA) for analysis of large numbers of samples (> 500) simultaneously for a large number of known mutations (> 100) in a single assay. MASDA utilizes oligonucleotide hybridization to interrogate DNA sequences. Multiplex DNA samples are immobilized on a solid support and a single hybridization is performed with a pool of allele-specific oligonucleotide (ASO) probes. Any probes complementary to specific mutations present in a given sample are in effect affinity purified from the pool by the target DNA. Sequence-specific band patterns (fingerprints), generated by chemical or enzymatic sequencing of the bound ASO(s), easily identify the specific mutation(s). Using this design, in a single diagnostic assay, we tested samples for 66 cystic fibrosis (CF) mutations, 14 beta-thalassemia mutations, two sickle cell anemia (SCA) mutations, three Tay-Sachs mutations, eight Gaucher mutations, four mutations in Canavan disease, four mutations in Fanconi anemia, and five mutations in BRCA1. Each mutation was correctly identified. Finally, in a blinded study of 106 of these mutations in > 500 patients, all mutations were properly identified. There were no false positives or false negatives. The MASDA assay is capable of detecting point mutations as well as small insertion or deletion mutations. This technology is amenable to automation and is suitable for immediate utilization for high-throughput genetic diagnostics in clinical and research laboratories.      

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.      

Use of negative intrathoracic pressure to obtain end-systolic pressure volume relations in dogs

Left ventricular contractility can be assessed from the end-systolic pressure-volume relationship (ESPVR). In this study we test the hypothesis that the same ESPVR can be obtained by varying LV loading with different levels of negative intrathoracic pressure as by varying LV filling. In six dogs mean aortic transmural pressure was used to approximate LV end-systolic pressure and LV volume was determined from data gathered from biplane cineradiograms of multiple markers placed in the LV midwall. In each preparation right heart bypass allowed control of cardiac output while the thoracic pressure was varied with a box surrounding a midsternal thoracotomy. Reflex effects were minimized by ganglionic blockade and bilateral vagotomy. ESPVRs were obtained by varying the cardiac output at constant thoracic pressure or by changing intrathoracic pressure at constant cardiac output. The slopes of the ESPVRs were not significantly different. This result implies that LV loading by negative intrathoracic pressure, in this highly controlled preparation, can be used to generate a systolic LV elastance similar to that obtained by varying LV filling.     

PSYCHOSOCIAL PROFILE OF PREGNANT ADOLESCENTS IN A LARGE AUSTRALIAN REGIONAL AREA

A psychosocial profile was developed of 122 pregnant adolescents attending a public hospital antenatal clinic in a large regional Australian area. Participants completed the Rosenberg Self-Esteem Scale and the Support Behaviours Inventory and were interviewed to obtain psychosocial and demographic information. Results revealed that those who were most likely to have either or a combination of poor self-esteem, lack of social support, be unemployed or smoke, were more likely to be living alone or with friends, not have a partner or have a partner for a shorter period, have a partner who was older, have previous children, have not planned their pregnancy and have less education. The degree of social support was significantly associated with self-esteem. Over half of the sample smoked, and over half were unemployed. These results underscore the importance of addressing psychosocial factors in the implementation of care for the pregnant adolescent.