Induction of a cytotoxic T lymphocyte (CTL) response to plasmid DNA delivered via Lipodine liposomes

We have previously shown that liposome-mediated plasmid DNA immunisation may be a preferred alternative to the use of naked DNA. Lipodine DNA formulations consist of liposomes containing entrapped DNA plasmid by the dehydration-rehydration (DRV) method. Such liposome formulations are distinct from liposomes with externally complexed DNA in that the majority of the DNA is "internal" to the liposome structure and hence protected from DNAase degradation. Previous studies on the immune response induced by DNA vaccines entrapped in Lipodine have focused on the humoural response. In the present study, we have expanded the analysis profile in order to include the cytotoxic T lymphocyte (CTL) component of the immune response. We have analysed the immune response induced by DNA entrapped in Lipodine compared to that induced by DNA alone when delivered subcutaneously, a route of administration not normally inducing significant plasmid DNA mediated immune activation. Our results indicate that delivery of a small dose of plasmid DNA in Lipodine results in an improved antibody response to the plasmid encoded antigen and a strong antigen specific CTL response compared to that induced by DNA delivered alone.     

Chemical and morphological variation of Mediterranean woody evergreen species: Do plants respond to ungulate browsing?

Abstract. Plant defense theories suggest that chemical or structural defences should be maximized when and where browsing is most likely to occur. We tested this hypothesis on four evergreen woody species growing in a Mediterranean area with a high density of ungulates. In this system, levels of browsing are more intense in the winter (due to the lack of annual plants) and young foliage is often preferred. Therefore we predicted that the chemical defences of these species, namely their phenolic content, would vary with leaf age, season and damage intensity. In addition, we tested whether ungulates preferentially selected species containing lower phenolic levels, and also whether browsing induced either chemical or morphological changes in damaged plants. Phenolic levels varied greatly between plant species; ungulates browsed preferentially on the species with the lowest phenolic levels. No difference in phenolic content was found between browsed and unbrowsed trees. Morphological changes in heavily browsed trees included an increase in shoot and leaf density and a net decrease in leaf size. We suggest that for Mediterranean plants, which have evolved under high browsing pressure from large mammals, the production of small leaves and dense shoots in response to browsing might decrease ungulate foraging efficiency and hence reduce the rate of further damage as effectively as high levels of chemical defence.     

Smoking and the sudden infant death syndrome: results from 1993-5 case-control study for confidential inquiry into stillbirths and deaths in infancy. Confidential Enquiry into Stillbirths and Deaths Regional Coordinators and Researchers.

OBJECTIVE--To investigate the effects of exposure to tobacco smoke and of parental consumption of alcohol and illegal drugs as risk factors for the sudden infant death syndrome after a national risk reduction campaign which included advice on prenatal and postnatal avoidance of tobacco smoke. DESIGN--Two year population based case-control study. Parental interviews were conducted for each infant who died and four controls matched for age and date of interview. SETTING--Three regions in England with a total population of 17 million people. SUBJECTS--195 babies who died and 780 matched controls. RESULTS--More index than control mothers (62.6% v 25.1%) smoked during pregnancy (multivariate odds ratio = 2.10; 95% confidence interval 1.24 to 3.54). Paternal smoking had an additional independent effect when other factors were controlled for (2.50; 1.48 to 4.22). The risk of death rose with increasing postnatal exposure to tobacco smoke, which had an additive effect among those also exposed to maternal smoking during pregnancy (2.93; 1.56 to 5.48). The population attributable risk was over 61%, which implies that the numbers of deaths from the syndrome could be reduced by almost two third if parents did not smoke. Alcohol use was higher among index than control mothers but was strongly correlated with smoking and on multivariate analysis was not found to have any additional independent effect. Illegal drug use was more common among the index parents, and paternal use of illegal drugs remained significant in the multivariate model (4.68; 1.56 to 14.05). CONCLUSIONS--This study confirms the increased risk of the sudden infant death syndrome associated with maternal smoking during pregnancy and shows evidence that household exposure to tobacco smoke has an independent additive effect. Parental drug misuse has an additional small but significant effect.     

Identification and characterization of bacterial endophytes of rice

We isolated seven different bacteria from rice seedlings grown from surface sterilized seeds. Three were associated with the rice seed husk and the other four were growing endophytically within the seed. Microscopic studies revealed that the endophytes were concentrated in the root stele region. Some of the bacteria exhibited strong anti-fungal activity against Rhizoctonia solani, Pythium myriotylum, Guamannomyces graminis and Heterobasidium annosum.     

Predicting badger sett numbers: evaluating methods in East Sussex

Abstract. One way in which a species' numbers may be estimated without direct counting is to predict their dispersion and density from more readily available habitat measures, such as landscape variables measured from maps or vegetation variables measured in the field. We compare the power of ordination and regression techniques for predicting badger ( Meles meles L.) numbers at a local scale, using a land class system, map-read landscape variables and field-derived vegetation variables. Sett density was used as a surrogate of badger density. Multiple linear regression using vegetation and landscape variables together gave the most accurate prediction of sett density, while ordination techniques were of lesser value. The addition of vegetation variables to landscape variables did not substantially improve the power of ordination. Outlier Sett Density was predicted more accurately, and by different variables, to Main Sett Density. The relationship between badger ecology and habitat variables that were useful in predicting sett density is discussed.     

Ascorbate-Stimulated Lipid Peroxidation in Human Brain Is Dependent on Iron but Not on Hydroxyl Radical

Abstract: The time dependence of N -acetyl-aspartate (NAA) concentrations relative to lactate and pyruvate in the injured rat spinal cord was investigated. Segments of spinal cord from regions rostral, caudal, and at the epicenter of the injury were analyzed. NAA concentrations were determined by gas chromatography-mass spectrometry and lactate and pyruvate concentrations were determined by UV spectroscopy at 20 min, 60 min, 2 h, 8 h, 24 h, 3 days, and 1 week after injury. NAA levels fell most significantly at the epicenter of the injury, reaching 30% of basal levels within 24 h. In all segments, lactate levels increased significantly shortly after injury, peaking at two to five times normal basal levels between 20 and 60 min after injury. Rostral and caudal to the injury site, lactate elevations and NAA reductions were less dramatic. Pyruvate concentrations were not significantly altered in any of the sections after injury. The temporal and spatial relationships of NAA and lactate changes indicated that ischemic conditions due to injury in the upper thoracic rat spinal cord were distributed asymmetrically. Acute ischemia was more severe caudal to the injury site, and NAA concentrations were more severely impaired in the rostral direction. The results suggest that the extent of neuronal degeneration due to spinal cord injury does not correlate directly with acute ischemic severity as measured by the lactate/pyruvate ratio, and may be more closely related to secondary changes in the neuronal environment.     

Histocompatibility antigen(s) linked to Rfp-Y (Mhc-like) genes in the chicken

 Major histocompatibility complex (Mhc) genes influencing transplantation rejections were first described in mice within the H2 complex and secondly in chickens within the B complex. In chickens, Rfp-Y haplotypes have recently been identified which contain class I and class II Mhc-like genes that assort independently of the B complex. Three Rfp-Y haplotypes have been defined in a closed breeding flock of line N chickens. In this study, progeny were obtained from line N Rfp-Y heterozygous matings to establish the role of Rfp-Y in transplantation immunity. Rfp-Y incompatibility did not induce significant one-way mixed lymphocyte responses. However, Rfp-Y-incompatible skin grafts were rejected more frequently and at a faster rate than Rfp-Y-compatible grafts by two-week-old chicks. The control Mhc B-incompatible grafts were rejected faster than the Rfp-Y-incompatible grafts; the latter were rejected at speeds that resemble rejection of minor histocompatibility antigens. We conclude that Rfp-Y class I and II Mhc-like genes are linked to the expression of minor histocompatibility antigens in chickens. Received: 21 June 1996 / Revised: 23 July 1996     

Structure and Bioactivity of Recombinant Human CTAP-III and NAP-2

Connective tissue-activating peptide III (CTAP-III) and neutrophil-activating peptide-2 (NAP-2) are both derived from a common precursor, platelet basic protein (PBP), which is stored in the -granules of platelets and released upon their activation. CTAP-III is an 85-residue peptide which is converted to NAP-2 by enzymic removal of the 15 amino-terminal residues. Both peptides play a role in the early stages of wound healing and inflammation through different activities. We have cloned the cDNA for PBP and expressed constructs coding for the CTAP-III and NAP-2 polypeptides in Escherichia coli. We have purified and renatured these recombinant proteins. The integrity of the recombinant proteins has been ascertained by in vitro bioassays. CTAP-III causes 51% histamine release from the basophilic cell lin KU812 at 10^–7 M, whereas NAP-2 only causes 28% release at the same concentration. In assays on human neutrophils, NAP-2 had an EC ₅₀ of 2×10^–8 M in chemotaxis, an EC ₅₀ of 3×10^–8 M for shape change, and could displace IL-8 from neutrophils with a K _d of 7.5×10^–9 M. CTAP-III had no activity in these assays. The disulfide bonds have been identified by peptide mapping and sequence analysis, and are in the positions predicted by homology to interleukin-8 and platelet factor 4. Measurement of the molecular mass at physiologic concentrations by gel permeation chromatography has shown that CTAP-III forms predominantly tetramers and dimers, whereas NAP-2 is only dimetric. SDS/PAGE analysis of samples cross-linked with disuccinimidyl suberate support these topologies. We postulate a mechanism for tetramer formation based on the interaction of the amino-terminal extension in CTAP-III involving a helix–helix interaction that could stabilize the association of two CTAP-III dimers.