The principle of delivery of T cell epitopes to antigen-presenting cells applied to peptides from influenza virus, ovalbumin, and hen egg lysozyme: implications for peptide vaccination

Targeting of antigens to antigen-presenting cells (APCs) increases CD4(+) T cell activation, and this observation can be exploited in the development of new vaccines. We have chosen an antigen-targeting approach in which we make recombinant antibodies (Abs) with T cell epitopes in their constant region and APC-specific variable regions. Three commonly used model epitopes, amino acids 110-120 of hemagglutinin, 323-339 of ovalbumin, and 46-61 of hen egg lysozyme, were introduced as loops in the C(H)1 domain of human IgG3. For all three epitopes, we show that the recombinant molecules are secreted from transfected cells. The epitopes are presented to specific T cells, and targeting to IgD on B cells in vitro enhances the presentation efficiency by 10(4) to 10(5) compared with the free peptide. After i.v. injection, the epitopes targeted to IgD are presented by splenic APCs to activate specific T cells, whereas little or no activation could be detected without targeting, even after the amount of antigen injected was increased 100-fold or more. Because a wide variety of T cell epitopes, in terms of both length and secondary structure, can be tolerated in loops in constant domains of Abs, the Ab constant region seems to have the intrinsic stability that is needed for this fusion molecule strategy. It might thus be possible to load the Ab with several different epitopes in loops in different domains and thereby make a targeted multisubunit vaccine.     

Spontaneous regression of a monoclonal proliferation of large granular lymphocytes associated with reversal of anemia and neutropenia

A 43-year-old male with a phenotypically homogeneous, expanded subset of T cells presented in 1981 with anemia and neutropenia. The surface antigen phenotype of 99% of the peripheral blood lymphocytes was T3+, T8+, T4-, and they were morphologically large granular lymphocytes (LGL). The same cells comprised 37% of the marrow nucleated cells. Eight months after he presented, the peripheral blood T8+, LGL diminished spontaneously, and the anemia and neutropenia completely resolved. The patient remains hematologically normal as of October 1984. To determine if the T8+, LGL represented a clonal expansion, DNA from peripheral blood lymphocytes collected and cryopreserved when the patient was neutropenic and anemic, and when he was hematologically normal, was analyzed for clonal T-cell antigen receptor gene rearrangements. Using Southern blot analysis, a clonal DNA rearrangement was demonstrated, and this clone diminished but was still demonstrable in peripheral blood lymphocytes collected in 1984. The above observations implicate the expanded T8+, LGL in the pathogenesis of the neutropenia and anemia, yet the exact mechanism remains to be elucidated.     

Characterization of an antibody to factor VIII in a patient with acquired hemophilia with circulating immune complexes

A 73-year-old previously healthy woman was admitted because of severe bleeding from esophagitic lesions and intraabdominal bleeding following hysterectomy. Acquired hemophilia, probably due to an IgG antibody to factor VIII (64 inhibitor units/ml) was noticed, the VIII:C in the patient's plasma being 18% or normal. Immune complexes isolated by polyethylene glycol precipitation had only a weak factor VIII inhibiting activity whereas IgG purified from the complexes and monomeric IgG present in her plasma exerted a strong inhibition. Removal of the complexes from plasma had no effect on the inhibitor titer thus indicating that only a minor part of the antibody was circulating as immune complexes. Plasma or purified IgG from the patient decreased the VIII:C of normal plasma to 18 og 14%, respectively, total inhibition being impossible to achieve even in antibody excess, probably reflecting residual activity of factor VIII bound to the patient's antibodies. The ristocetin cofactor activity of normal plasma was unaffected by the antibodies. Transfusion of factor VIII concentrate to the patient resulted in therapeutic levels of circulating factor VIII and transfused factor VIII circulated longer than usual. Partial remission of the disease with adequate levels of VIII:C occurred after 3 months.     

Development of Dietary Patterns Spanning Infancy and Toddlerhood: Relation to Body Size,Composition and Metabolic Risk Markers at Three Years

Little is known about the development of dietary patterns during toddlerhood and the relation to growth and health. The study objective was to characterise the development of dietary patterns from 9–36 mo of age and investigate the association to body size, body composition and metabolic risk markers at 36 mo. Food records were filled out at 9, 18 and 36 mo of age (n = 229). Dietary patterns were identified by principal component analysis (PCA). Three dietary patterns were identified: Transition Food, Healthy Food and Traditional Food. The course of development in dietary patterns from 9–36 mo indicated tracking for a relatively large group of participants in the three patterns. Transition Food and Healthy Food were associated with some of the investigated outcomes. Children with lower adherence to the Transition Food pattern than average at 18 and 36 mo irrespectively of intake at 9 mo had higher BMI z-scores at 36 mo. Similar trend was identified for higher fat mass indices. Children with lower adherence to the Healthy Food pattern than average at all three ages compared to children with higher adherence to the Healthy Food pattern at the first two registrations, 9 and 18 mo had higher total cholesterol and LDL. Hence, this could represent undesirable development of dietary patterns in toddlers. In conclusion, development of dietary patterns can be exploratory characterised by PCA and related to potential cardiovascular risk markers in toddlers even within a relatively homogeneous population with a high socioeconomic status. The tracking of dietary patterns from 9 mo of age indicates a need for early and sustained promotion of healthy diets.     

Bilateral and multifocal phyllodes tumours of the breast: A case report

Phyllodes tumours are rare breast neoplasms that present as painless breast masses. They are classified as benign, malignant and borderline. More rare presentations of these tumours include bilateral asynchronous disease and unilateral multifocal disease. Surgical excision with clear margins remains the treatment of choice for these tumours. The present case report is the first to be discussed in the literature. It describes a patient presenting with synchronous bilateral, multifocal breast phyllodes tumours who underwent immediate reconstruction with tissue expanders at the time of her mastectomies.     

Streptococcus pyogenes Isolates Causing Severe Infections in Norway in 2006 to 2007: emm Types,Multilocus Sequence Types,and Superantigen Profiles

To investigate the epidemiological patterns and genetic characteristics of disease caused by group A Streptococcus (GAS), all available isolates from invasive cases in Norway during 2006 to 2007 (262 isolates) were subjected to antimicrobial susceptibility testing, T serotyping, emm typing, and multilocus sequence typing and screened for known streptococcal pyrogenic exotoxin (Spe) genes, smeZ, and ssa. The average incidence rate was 3.1 cases per 100,000 individuals. The most prevalent sequence types (STs) were STs 52, 28, and 334. In association with emm types 28, 77, and 87, the serotype T-28 comprised 24.8% of the strains. emm types 28, 1, and 82 were dominating. In 2007, a sharp increase in the number of emm-6 strains was noted. All strains were sensitive to penicillin and quinupristin-dalfopristin, while 3.4% and 6.1% of the strains were resistant to macrolides and tetracycline, respectively. Furthermore, the emm-6 strains had intermediate susceptibility to ofloxacin. Isolates displayed a wide variety of gene profiles, as shown by the presence or absence of the Spe genes, smeZ, and ssa, but 48% of the isolates fell into one of three profiles. In most cases, an emm type was restricted to one gene profile. Although the incidence decreased during this study, invasive GAS disease still has a high endemic rate, with involvement of both established and emerging emm types displaying variability in virulence gene profiles as well as differences in gender and age group preferences.Group A streptococcus (GAS), Streptococcus pyogenes, is a highly prevalent Gram-positive human pathogen with a worldwide distribution. Most often, it causes superficial infections of the upper respiratory tract and of the skin, leading to pharyngitis and impetigo, respectively. Invasive GAS infections, on the other hand, can be life-threatening due to conditions such as bacteremia, cellulitis, erysipelas, meningitis, and pneumonia, including the severe manifestations of necrotizing fasciitis (NF) and streptococcal toxic shock syndrome (STSS) (8). In a historical perspective, GAS has been associated with high fatality rates due to severe scarlet fever, puerperal sepsis, and systemic disease (24). With the introduction of antibiotics in the 1940s, incidence rates of severe GAS infections dropped in developed countries and stayed low until the 1980s. Increased virulence and invasiveness then resulted in an increased number of reported septicemia, NF, and STSS cases in previously healthy children and adults in the United States and Europe (14, 33, 49).S. pyogenes harbors a large number of virulence factors that contribute to its complex pathogenicity (7, 12). One of the major virulence factors, the M protein, encoded by the emm gene, confers antiphagocytic properties and induces a type-specific host immune response. Another important group of virulence factors, targeted by anti-T sera, are the pilin proteins, producing pilus-like structures (40) involved in adhesion and invasion of eukaryotic cells and in biofilm formation (1, 31). The streptococcal pyrogenic exotoxins (Spe proteins), a family of bacterial superantigens, are potent immunostimulators associated with disease conditions such as acute rheumatic fever, scarlet fever, and STSS (12, 54). In total, 11 superantigens have been identified in GAS to date, including SpeA, SpeC, SpeG to -M, streptococcal mitogenic exotoxin Z (SmeZ), and streptococcal superantigen (SSA). Because of an issue with the naming of some of the more recently discovered superantigens and for the sake of simplicity, SpeK/L (4, 21, 45), SpeL/M (45, 52), and SpeM (52) are here referred to as SpeK, SpeL, and SpeM, respectively. The proteins SpeB, a cysteine protease, and SpeF, a DNase, were previously considered to be superantigens due to contamination with the potent SmeZ protein (7, 44). Except for speG, speJ, and smeZ, most genes encoding superantigens are associated with bacteriophages (4, 17, 18). Phages are believed to be the major contributors to genetic variation in GAS, both between strains and within strains of the same type (2, 4). Isolates of the same emm type usually share a superantigen profile. Variants differing by the presence or absence of one or a few genes may occur, however, and geographic and temporal differences in the superantigen content of strains of a given emm type have been reported (11).The M protein has traditionally been targeted for serotyping of GAS strains because of its importance as a virulence determinant. However, sequencing of the emm gene (3) is now becoming the standard method, and to date, more than 150 emm types have been described (36). Another method, which has been used for the last 50 years and is still an important alternative to serological M typing, is T typing using slide agglutination tests (39). Multilocus sequence typing (MLST), a widely used method for genetic characterization of organisms of a bacterial species, which is based on the nucleotide sequence variation in seven housekeeping genes, provides unambiguous results that are easily comparable between laboratories (15). Although geographical and temporal variation has been described for GAS populations (15, 35, 47), strains with the same emm type isolated as much as 50 years apart may harbor identical allelic profiles (15) and share the same T type (23). Due to the clonal population structure of S. pyogenes strains, results obtained by T typing, emm typing, and MLST correlate with each other (15, 23, 53).Norway experienced relatively low incidence rates of severe GAS disease after the introduction of antibiotics, but in the mid-1980s there was an increased occurrence of severe invasive disease, especially in otherwise healthy young adults, largely caused by M-1 strains (9, 33). Thereafter, until the early 2000s, there was a significant decrease in the frequency of emm-1 strains and, at the same time, an increase in diversity among the Norwegian GAS strains (37). Antibiotic resistance levels were generally very low in Norway during this period (37).In more recent reports from the United States (2000 to 2004) and the United Kingdom (2003 to 2004), emm-1 and emm-3 strains were still among the most frequent strains found in invasive GAS disease (29, 43). The overall distributions of the most prevalent emm types in Europe and the United States during this period were in congruence, but there were marked differences in the emm type distributions between countries such as Norway''s neighbors, Denmark, Finland, and Sweden (29). The most prominent difference was seen in Finland, where 45% of the strains were emm-28 strains (29). Recently, Finland also reported a rapid change in genotype prevalence caused by the previously uncommon emm type 84 during 2005 to 2007 (50).The distribution of GAS strains and the virulence factors associated with the different strains are not stable over time. Therefore, epidemiological studies targeting genetic types, important virulence factors, and the antimicrobial susceptibility status of these microorganisms are of basic importance for detection of new emerging clones, determination of their potential to cause disease, and development and refinement of vaccines. To provide better insight into the current epidemiological situation for severe GAS infections in Norway, we characterized all available isolates from invasive GAS disease obtained in 2006 to 2007, using emm typing, MLST, spe gene profiling, including smeZ and ssa, and antibiotic resistance screening using selected antibiotics.     

Anti-ulcer polysaccharides from Cola cordifolia bark and leaves

Ethnopharmacological relevance

Aqueous extracts of bark and leaves of C. cordifolia are traditionally used in Mali (West Africa) in the treatment of wounds and gastric ailments like abdominal pain, gastritis and gastric ulcers.

Aim of the study

To evaluate and compare the anti-ulcer and immunological activities, as well as the toxicity of polysaccharide rich water extracts from the bark and leaves of C. cordifolia.

Materials and methods

Gastric ulcers were induced in rats and the inhibition of ulcer formation was calculated based on lesion index. Immunological activities were measured by complement fixation and macrophage activation. Toxicity was tested on brine shrimps. The two extracts were characterised by GC, Yariv-precipitation and quantification of phenolic compounds. An ethnomedical survey on C. cordifolia was carried out in Siby (Mali, West-Africa) to generate more knowledge about the traditional use.

Results

Bark and leaf extracts from C. cordifolia significantly inhibited the formation of gastric lesions in rodents in a dose depending manner. CCbark50 showed a high complement fixation activity in vitro. No toxicity was found. The ethnomedical survey showed that C. cordifolia was mainly used for treating pain and wounds.

Conclusions

Our results shows that the bark and the leaves comprise a dose dependant anti-ulcer activity in an experimental rat model (no statistical difference between the plant parts). Clinical studies should be performed to evaluate the effect of both bark and leaves of C. cordifolia as a remedy against gastric ulcer in human.     

Neurotrophin receptors TrkB.T1 and p75NTR cooperate in modulating both functional and structural plasticity in mature hippocampal neurons

Tropomyosin-related kinase (Trk) receptors modulate neuronal structure and function both during development and in the mature nervous system. Interestingly, TrkB and TrkC are expressed as full-length and as truncated splice variants. The cellular function of the kinase-lacking isoforms remains so far unclear. We investigated the role of the truncated receptor TrkB.T1 in the hippocampus of transgenic mice overexpressing this splice variant by analyzing both neuronal structure and function. We observed an impairment in activity-dependent synaptic plasticity as indicated by deficits in long-term potentiation and long-term depression in acute hippocampal slices of transgenic TrkB.T1 mice. In addition, dendritic complexity and spine density were significantly altered in TrkB.T1-overexpressing CA1 neurons. We found that the effect of TrkB.T1 overexpression differs between subgroups of CA1 neurons. Remarkably, overexpression of p75(NTR) and its activation by chemical induction of long-term depression in slice cultures rescued the TrkB.T1-dependent morphological alterations specifically in one of the two subgroups observed. These findings suggest that the TrkB.T1 and p75(NTR) receptor signaling systems might be cross-linked. Our findings demonstrate that TrkB.T1 regulates the function and the structure of mature pyramidal neurons. In addition, we showed that the ratio of expression levels of p75(NTR) and TrkB.T1 plays an important role in modulating dendritic architecture and synaptic plasticity in the adult rodent hippocampus, and, indeed, that the endogenous expression patterns of both receptors change reciprocally over time. We therefore propose a new function of TrkB.T1 as being dominant-negative to p75(NTR).