INSULIN PRODUCES A BIPHASIC RESPONSE INTETRAHYMENA THERMOPHILABY STIMULATING CELL SURVIVAL AND ACTIVATING PROLIFERATION IN TWO SEPARATE CONCENTRATION INTERVALS

Cells ofTetrahymenamay produce autocrine signal molecules with effects on survival and proliferation. Here we have tested the effects of human recombinant and bovine insulin, and the B22–B30 fragment of bovine insulin over a wide range of concentrations (10^?5–10^?18m ) on cell survival and proliferation in a synthetic nutrient medium. The cells were grown in conical flasks at low initial cell densities (40 and 400cells/ml). Insulin prevented rapid cell death and/or promoted cell proliferation over two separate concentration ranges: down to nanomolar levels and again in the low pico- and femtomolar range. At an initial population density of 400cells/ml the cells multiplied at both concentration intervals. At 40 or fewer organisms/ml the cells multiplied in the high concentration interval, whereas in the low interval they survived for about four times longer than those in the control cultures. B22–B30 added to cultures of 40 initial cells/ml produced a stimulation of cell survival in the low pico- and high femtomolar range. In the presence of hemin (50nm ) cells at 400 initial organisms/ml multiplied at insulin concentrations down to about 3nm and again from 300am to 10pm . In some cases, hemin plus insulin activated cell proliferation between the two concentration intervals as well. At 40cells/ml the cells not only survived but proliferated in the femtomolar range. Cells in cultures supplemented with both hemin and B22–B30 multiplied at the low concentration interval (from about 100fm to 10pm ).     

Rates and equilibria at the acetylcholine receptor of electrophorus electroplaques. A study of neurally evoked postsynaptic currents and of voltage-jump relaxations

Kinetic measurements are employed to reconstruct the steady-state activation of acetylcholine [Ach] receptor channels in electrophorus electroplaques. Neurally evoked postsynaptic currents (PSCs) decay exponentially; at 15 degrees C the rate constant, α, equals 1.2 ms(-1) at 0 mV and decreases e-fold for every 86 mV as the membrane voltage is made more negative. Voltage-jump relaxations have been measured with bath-applied ACh, decamethonium, carbachol, or suberylcholine. We interpret the reciprocal relaxation time 1/τ as the sum of the rate constant α for channel closing and a first-order rate constant for channel opening. Where measureable, the opening rate increases linearly with [agonist] and does not vary with voltage. The voltage sensitivity of small steady-state conductances (e- fold for 86 mV) equals that of the closing rate α, confirming that the opening rate has little or no additional voltage sensitivity. Exposure to α-bungarotoxin irreversibly decreases the agonist-induced conductance but does not affect the relaxation kinetics. Tubocurarine reversibly reduces both the conductance and the opening rate. In the simultaneous presence of two agonist species, voltage-jump relaxations have at least two exponential components. The data are fit by a model in which (a) the channel opens as the receptor binds the second in a sequence of two agonist molecules, with a forward rate constant to 10(7) to 2x10(8) M(-1)s(-1); and (b) the channel then closes as either agonist molecule dissociates, with a voltage-dependent rate constant of 10(2) to 3x10(3)s(-1).     

Sixty simple sequence repeat markers for use in the Festuca–Lolium complex of grasses

So far only very few simple sequence repeat (SSR) markers developed from grass species have had their primer sequences published. To make more markers available to the scientific community, we isolated and sequenced 256 microsatellite‐containing clones from four genome libraries of a Lolium multiflorum×Festuca glaucescens F₁ hybrid following enrichment in (TC)_n, (TG)_n, or both repeats. In this work, we report the primer sequences of 60 SSRs including preliminary results of polymorphism for mapping.     

Survival and Body Condition of Raccoons at the Edge of the Range

ABSTRACT We investigated the influence of intrinsic and extrinsic variables on overwinter survival of raccoons (Procyon lotor; n = 114) at the northern edge of their distribution. A Cox proportional hazard model identified winter severity as the variable with the greatest influence on raccoon survival (β = 1.08). Autumn body condition estimates (20.5 ± 0.46% total body fat) were relatively stable across years even though we observed large differences in autumn food indices. Variations in autumn body condition did not explain heterogeneity observed in overwinter survival nor the spring condition in which raccoons emerged. Relatively constant autumn body condition suggests reliable availability of anthropogenic food resources may negate variations observed in natural food items on which raccoons rely during hyperphagia. Conversely, spring body condition did vary among years and was highly correlated with winter severity. Accordingly, we also observed a strong inverse relationship with overwinter survival and winter severity. Our findings indicate winter climatic constraints are important factors governing the northern limit of raccoon distribution and changes in winter severity could have important implications in further range expansion of this species.     

Oxygen metabolism,oxidative stress and acid-base physiology of dental plaque biofilms

Dental plaque is a natural biofilm which has been a focus of attention for many years because of its known roles in caries and periodontal diseases. Acid production by plaque bacteria leads to the erosion of tooth mineral in caries, and the cariogenicity of plaque is related to population levels of acid-tolerant organisms such as mutants streptococci. However, the biofilm character of plaque allows for survival of a diverse flora, including less acid-tolerant organisms, some of which can produce ammonia from arginine or urea to counter acidification. Plaque is often considered to be relatively anaerobic. However, evidence is presented here that both supragingival and subgingival plaque have active oxygen metabolism and that plaque bacteria, including anaerobes, have developed defenses against oxidative stress. Even in subgingival plaque associated with periodontitis, measured residual oxygen levels are sufficient to allow for oxygen metabolism by organisms considered to be extremely anaerobic such asTreponema denticola, which metabolizes oxygen by means of NADH oxidases and produces the protective enzymes superoxide dismutase and NADH peroxidase. The finding that plaque bacteria produce a variety of protective enzymes is a good indicator that oxidative stress is a part of their everyday life. The biofilm character of plaque allows for population diversity and coexistence of aerobes, anaerobes and microaerophiles. Overall, agents that affect oxidative metabolism offer possibilities for reducing the pathogenic activities of plaque.     

Chromosomal localization of a tandemly repeated DNA sequence in Trifolium repens L

INTRoDUCTIoNlYho1iumrePensL,whiteclover,isaneconomicallyimportantplantspeciesintemperatepastures.Asbrieflyreportedby[1],ithas16pairsofchromosomes(2n=32).Asyet,nodetailedcytologicalexaminationofthisspecies,suchasC-banding,hasbeenrep0rted.Inthelastdecade,thetechnique0fC-bandinghasbeenusedt0examinehighlyrepeatedsequencesinplantchrom0s0mesandhasprovidedausefultoolf0rtheanalysis0fcyt0geneticstructureincr0pplants[2-71.Inplants,thechr0m0s0mall0calizationofhighlyrepeatedDNAsequencesbyinsituhybr…     

Two new behavioral QTLs, Emo4 and Reb1, map to mouse Chromosome 1: Congenic strains and candidate gene identification studies

By use of newly developed subcongenic strains of mice from a parental B6.129-Il10^−/− knockout/congenic strain, we have narrowed the critical region for a new behavioral QTL, called Emo4, for open-field activity to a segment of Chromosome 1 between Erbb4 (68.4Mb) and B3gnt7 (86.2 Mb). We have also uncovered an additional QTL governing repetitive beam breaks in the open field. This QTL, called Reb1, maps to the interval between Asb1 (91.4 Mb) and NM_172851 (100.0 Mb) and is one of the first QTLs mapped for this type of behavior. Genome-wide microarray expression analyses were then undertaken to help to identify candidate genes that may be the cause of these genetic differences in open-field performance. In this effort, we analyzed global gene expression differences in the amygdalae by use of Affymetrix GeneChips between B6, B6.129-Il10^−/−, and B6.129R4. Several probe sets representing target Chr 1 genes were found that showed significantly differential expression in the subcongenic and congenic strains. Several candidate genes have been identified. One of these regions coincides with an homologous region in humans that has been associated with autism, a disease whose symptoms include repetitive actions. This study illustrates that the use of congenic strains combined with global gene expression analyses can produce a list of viable candidates. It further shows that caution should be observed when analyzing the effects of knockout/congenic strains because many of the gene expression differences in these comparisons could not be attributable to the ablated Il10 gene but rather to passenger gene effects.     

Young patients with colorectal cancer have poor survival in the first twenty months after operation and predictable survival in the medium and long-term: Analysis of survival and prognostic markers

Introduction

Male breast cancer (MBC) is a rare, yet potentially aggressive disease. Although literature regarding female breast cancer (FBC) is extensive, little is known about the etiopathogenesis of male breast cancer. Studies from our laboratory show that MBCs have a distinct immunophenotypic profile, suggesting that the etiopathogenesis of MBC is different from FBCs. The aim of this study was to evaluate and correlate the immunohistochemical expression of cell cycle proteins in male breast carcinoma to significant clinico-biological endpoints.

Methods

75 cases of MBC were identified using the records of the Saskatchewan Cancer Agency over 26 years (1970-1996). Cases were reviewed and analyzed for the immunohistochemical expression of PCNA, Ki67, p27, p16, p57, p21, cyclin-D1 and c-myc and correlated to clinico-biological endpoints of tumor size, node status, stage of the disease, and disease free survival (DFS).

Results

Decreased DFS was observed in the majority of tumors that overexpressed PCNA (98%, p = 0.004). The overexpression of PCNA was inversely correlated to the expression of Ki67 which was predominantly negative (78.3%). Cyclin D1 was overexpressed in 83.7% of cases. Cyclin D1 positive tumors were smaller than 2 cm (55.6%, p = 0.005), had a low incidence of lymph node metastasis (38.2%, p = 0.04) and were associated with increased DFS of >150 months (p = 0.04). Overexpression of c-myc (90%) was linked with a higher incidence of node negativity (58.3%, p = 0.006) and increased DFS (p = 0.04). p27 over expression was associated with decreased lymph node metastasis (p = 0.04). P21 and p57 positive tumors were related to decreased DFS (p = 0.04). Though p16 was overexpressed in 76.6%, this did not reach statistical significance with DFS (p = 0.06) or nodal status (p = 0.07).

Conclusion

Aberrant cell cycle protein expression supports our view that these are important pathways involved in the etiopathogenesis of MBC. Tumors with overexpression of Cyclin D1 and c-myc had better outcomes, in contrast to tumors with overexpression of p21, p57, and PCNA with significantly worse outcomes. P27 appears to be a predictive marker for lymph nodal status. Such observation strongly suggests that dysregulation of cell cycle proteins may play a unique role in the initiation and progression of disease in male breast cancer. Such findings open up new avenues for the treatment of MBC as a suitable candidate for novel CDK-based anticancer therapies in the future.     

Site selection by swans wintering in Britain and Ireland; the importance of habitat and geographic location

Monthly surveys of Bewick's Swans Cygnus columbianus bewickii , Whooper Swans Cygnus cygnus and Mute Swans Cygnus olor in Britain and Ireland were made during the 1990–1991 winter to determine factors affecting the swans' selection of feeding sites. Geographic location and habitat both influenced site selection. Whooper Swans occurred in greatest numbers at sites in Scotland, northeastern England and Northern Ireland, whereas Bewick's Swans had a more southerly distribution, reflecting differences in the migratory routes used by these two species. The resident Mute Swans were more widespread, with large flocks occurring in southeastern England and in parts of Scotland. Whooper and Mute Swans were found mainly on permanent inland waters (68% and 61%, respectively), but the majority of Bewick's Swans (60%) were on arable land. The percentage of Bewick's Swan flocks found on permanent inland waters (42%) was higher than that found on arable fields (23%), indicating that the large number recorded on arable land was a result of the birds congregating at a comparatively small number of sites. Overall, less than 15% of Whooper Swans and 3% of Mute Swans were on arable crops during the winter, but the largest flocks were associated with arable land for all three species. Thus, although the occurrence of large flocks at particular arable sites may give an impression that swans feed mainly on farmland, the swans are in fact more widely dispersed. Regional variation in the percentage of juveniles present was recorded for all three species. Changes during the winter in the distribution of juveniles, and of the swans as a whole, are considered in relation to food supply and to migratory routes for the Bewick's and Whooper Swans.