gp100(209-2M) peptide immunization of human lymphocyte antigen-A2+ stage I-III melanoma patients induces significant increase in antigen-specific effector and long-term memory CD8+ T cells.

Thirty-five HLA-A2(+) patients with completely resected stage I-III melanoma were vaccinated multiple times over 6 months with a modified melanoma peptide, gp100(209-2M), emulsified in Montanide adjuvant. Direct ex vivo gp100(209-2M) tetramer analysis of pre- and postvaccine peripheral blood mononuclear cells (PBMCs) demonstrated significant increases in the frequency of tetramer(+) CD8(+) T cells after immunization for 33 of 35 evaluable patients (median, 0.36%; range, 0.05-8.9%). Ex vivo IFN-gamma cytokine flow cytometry analysis of postvaccine PBMCs after brief gp100(209-2M) in vitro activation showed that for all of the patients studied tetramer(+) CD8(+) T cells produced IFN-gamma; however, some patients had significant numbers of tetramer(+) IFN-gamma(-) CD8(+)T cells suggesting functional anergy. Additionally, 8 day gp100(209-2M) in vitro stimulation (IVS) of pre- and postvaccine PBMCs resulted in significant expansion of tetramer(+) CD8(+) T cells from postvaccine cells for 34 patients, and these IVS tetramer(+) CD8(+) T cells were functionally responsive by IFN-gamma cytokine flow cytometry analysis after restimulation with either native or modified gp100 peptide. However, correlated functional and phenotype analysis of IVS-expanded postvaccine CD8(+) T cells demonstrated the proliferation of functionally anergic gp100(209-2M)- tetramer(+) CD8(+) T cells in several patients and also indicated interpatient variability of gp100(209-2M) stimulated T-cell proliferation. Flow cytometry analysis of cryopreserved postvaccine PBMCs from representative patients showed that the majority of tetramer(+) CD8+ T cells (78.1 +/- 4.2%) had either an "effector" (CD45 RA(+)/CCR7(-)) or an "effector-memory" phenotype (CD45RA(-)/CCR7(-)). Notably, analysis of PBMCs collected 12-24 months after vaccine therapy demonstrated the durable presence of gp100(209-2M)-specific memory CD8(+) T cells with high proliferation potential. Overall, this report demonstrates that after vaccination with a MHC class I-restricted melanoma peptide, resected nonmetastatic melanoma patients can mount a significant antigen-specific CD8(+) T-cell immune response with a functionally intact memory component. The data further support the combined use of tetramer binding and functional assays in correlated ex vivo and IVS settings as a standard for immunomonitoring of cancer vaccine patients.

Evolution of gene expression in fire ants: the effects of developmental stage, caste, and species.

Ants provide remarkable examples of equivalent genotypes developing into divergent and discrete phenotypes. Diploid eggs can develop either into queens, which specialize in reproduction, or workers, which participate in cooperative tasks such as building the nest, collecting food, and rearing the young. In contrast, the differentiation between males and females generally depends upon whether eggs are fertilized, with fertilized (diploid) eggs giving rise to females and unfertilized (haploid) eggs giving rise to males. To obtain a comprehensive picture of the relative contributions of gender (sex), caste, developmental stage, and species divergence to gene expression evolution, we investigated gene expression patterns in pupal and adult queens, workers, and males of two species of fire ants, Solenopsis invicta and S. richteri. Microarray hybridizations revealed that variation in gene expression profiles is influenced more by developmental stage than by caste membership, sex, or species identity. The second major contributor to variation in gene expression was the combination of sex and caste. Although workers and queens share equivalent diploid nuclear genomes, they have highly distinctive patterns of gene expression in both the pupal and the adult stages, as might be expected given their extraordinary level of phenotypic differentiation. Overall, the difference in the proportion of differentially expressed genes was greater between workers and males than between workers and queens or queens and males, consistent with the fact that workers and males share neither gender nor reproductive capability. Moreover, between-species comparisons revealed that the greatest difference in gene expression patterns occurred in adult workers, a finding consistent with the fact that adult workers most directly experience the distinct external environments characterizing the different habitats occupied by the two species. Thus, much of the evolution of gene expression in ants may occur in the worker caste, despite the fact that these individuals are largely or completely sterile. Analyses of gene expression evolution revealed a combination of positive selection and relaxation of stabilizing selection as important factors driving the evolution of such genes.

Assessment of early-stage optic nerve invasion in retinoblastoma using high-resolution 1.5 Tesla MRI with surface coils: a multicentre, prospective accuracy study with histopathological correlation.

OBJECTIVES: To assess the accuracy of high-resolution (HR) magnetic resonance imaging (MRI) in diagnosing early-stage optic nerve (ON) invasion in a retinoblastoma cohort. METHODS: This IRB-approved, prospective multicenter study included 95 patients (55 boys, 40 girls; mean age, 29 months). 1.5-T MRI was performed using surface coils before enucleation, including spin-echo unenhanced and contrast-enhanced (CE) T1-weighted sequences (slice thickness, 2 mm; pixel size <0.3 × 0.3 mm(2)). Images were read by five neuroradiologists blinded to histopathologic findings. ROC curves were constructed with AUC assessment using a bootstrap method. RESULTS: Histopathology identified 41 eyes without ON invasion and 25 with prelaminar, 18 with intralaminar and 12 with postlaminar invasion. All but one were postoperatively classified as stage I by the International Retinoblastoma Staging System. The accuracy of CE-T1 sequences in identifying ON invasion was limited (AUC = 0.64; 95 % CI, 0.55 - 0.72) and not confirmed for postlaminar invasion diagnosis (AUC = 0.64; 95 % CI, 0.47 - 0.82); high specificities (range, 0.64 - 1) and negative predictive values (range, 0.81 - 0.97) were confirmed. CONCLUSION: HR-MRI with surface coils is recommended to appropriately select retinoblastoma patients eligible for primary enucleation without the risk of IRSS stage II but cannot substitute for pathology in differentiating the first degrees of ON invasion. KEY POINTS: • HR-MRI excludes advanced optic nerve invasion with high negative predictive value. • HR-MRI accurately selects patients eligible for primary enucleation. • Diagnosis of early stages of optic nerve invasion still relies on pathology. • Several physiological MR patterns may mimic optic nerve invasion.

Stage of change of cigarette smoking in alcohol-dependent patients.

Despite the heavy burden of tobacco-related problems in alcohol-dependent patients, little effort has been directed toward reducing the prevalence of smoking in these patients. It seems reasonable to develop nicotine addiction treatments for alcohol-dependent patients based on the smoker's stage of change. To assess the stage of change for tobacco consumption and possible quitting barriers in alcohol-dependent patients, 88 consecutively admitted inpatients of a Swiss university-affiliated alcohol withdrawal clinic were interviewed with a semistructured schedule. More than half of the alcohol-dependent smokers (50.7%) considered the possibility of smoking cessation or had already decided to stop, although the majority (83.1%) were highly dependent smokers. Positive reinforcers were factors influencing motivation both to stop smoking as well as to continue smoking, whereas negative reinforcers had no influence. As recovering alcoholic patients are often interested in smoking cessation and the introduction of nicotine treatment interventions has been shown not to jeopardize the outcome of alcohol treatment, alcohol treatment programs should include counseling for smoking cessation. Education and training for staff is essential, as their beliefs and habits remain an important barrier.

Artificial muscle for end-stage heart failure.

We describe a device made of artificial muscle for the treatment of end-stage heart failure as an alternative to current heart assist devices. The key component is a matrix of nitinol wires and aramidic fibers called Biometal muscle (BM). When heated electrically, it produces a motorless, smooth, and lifelike motion. The BM is connected to a carbon fiber scaffold, tightening the heart and providing simultaneous assistance to the left and right ventricles. A pacemaker-like microprocessor drives the contraction of the BM. We tested the device in a dedicated bench model of diseased heart. It generated a systolic pressure of 75 mm Hg and ejected a maximum of 330 ml/min, with an ejection fraction of 12%. The device required a power supply of 6 V, 250 mA. This could be the beginning of an era in which BMs integrate or replace the mechanical function of natural muscles.

Automatic Tracer-Dilution Method Used for Stage-Discharge Ratings and Streamflow Hydrographs on Small Iowa Streams, HR-285, 1990

An automatic system was designed to concurrently measure stage and discharge for the purpose of developing stage-discharge ratings and high flow hydrographs on small streams. Stage, or gage height, is recorded by an analog-to-digital recorder and discharge is determined by the constant-rate tracer-dilution method. The system measures flow above a base stage set by the user. To test the effectiveness of the system and its components, eight systems, with a variety of equipment, were installed at crest-stage gaging stations across Iowa. A fluorescent dye, rhodamine-WT, was used as the tracer. Tracer-dilution discharge measurements were made during 14 flow periods at six stations from 1986 through 1988 water years. Ratings were developed at three stations with the aid of these measurements. A loop rating was identified at one station during rapidly-changing flow conditions. Incomplete mixing and dye loss to sediment apparently were problems at some stations. Stage hydrographs were recorded for 38 flows at seven stations. Limited data on background fluorescence during high flows were also obtained.

bHLH class transcription factors take centre stage in phytochrome signalling.

The phytochrome family of photoreceptors (there are five phytochromes in Arabidopsis, named phyA to phyE) maximally absorbs red and far-red light and plays important functions throughout the life cycle of plants. Several recent studies have shown that multiple related bHLH (basic helix-loop-helix) class transcription factors play key roles in phytochrome signal transduction. Somewhat surprisingly these transcription factors primarily act as negative regulators of phytochrome signalling. Moreover, in some cases, the phytochromes inhibit those negative regulators.

Neoadjuvant chemotherapy and radiotherapy followed by surgery in selected patients with stage IIIB non-small-cell lung cancer: a multicentre phase II trial.

BACKGROUND: Stage IIIB non-small-cell lung cancer (NSCLC) is usually thought to be unresectable, and is managed with chemotherapy with or without radiotherapy. However, selected patients might benefit from surgical resection after neoadjuvant chemotherapy and radiotherapy. The aim of this multicentre, phase II trial was to assess the efficacy and toxicity of a neoadjuvant chemotherapy and radiotherapy followed by surgery in patients with technically operable stage IIIB NSCLC. METHODS: Between September, 2001, and May, 2006, patients with pathologically proven and technically resectable stage IIIB NSCLC were sequentially treated with three cycles of neoadjuvant chemotherapy (cisplatin with docetaxel), immediately followed by accelerated concomitant boost radiotherapy (44 Gy in 22 fractions) and definitive surgery. The primary endpoint was event-free survival at 12 months. Efficacy analyses were done by intention to treat. This trial is registered with ClinicalTrials.gov, number NCT00030810. FINDINGS: 46 patients were enrolled, with a median age of 60 years (range 28-70). 13 (28%) patients had N3 disease, 36 (78%) had T4 disease. All patients received chemotherapy; 35 (76%) patients received radiotherapy. The main toxicities during chemotherapy were neutropenia (25 patients [54%] at grade 3 or 4) and febrile neutropenia (nine [20%]); the main toxicity after radiotherapy was oesophagitis (ten patients [29%]; nine grade 2, one grade 3). 35 patients (76%) underwent surgery, with pneumonectomy in 17 patients. A complete (R0) resection was achieved in 27 patients. Peri-operative complications occurred in 14 patients, including two deaths (30-day mortality 5.7%). Seven patients required a second surgical intervention. Pathological mediastinal downstaging was seen in 11 of the 28 patients who had lymph-node involvement at enrolment, a complete pathological response was seen in six patients. Event-free survival at 12 months was 54% (95% CI 39-67). After a median follow-up of 58 months, the median overall survival was 29 months (95% CI 16.1-NA), with survival at 1, 3, and 5 years of 67% (95% CI 52-79), 47% (32-61), and 40% (24-55). INTERPRETATION: A treatment strategy of neoadjuvant chemotherapy and radiotherapy followed by surgery is feasible in selected patients. Toxicity is considerable, but manageable. Survival compares favourably with historical results of combined treatment for less advanced stage IIIA disease. FUNDING: Swiss Group for Clinical Cancer Research (SAKK) and an unrestricted educational grant by Sanofi-Aventis (Switzerland).

Prospective randomized study of two cryopreservation policies avoiding embryo selection: the pronucleate stage leads to a higher cumulative delivery rate than the early cleavage stage.

OBJECTIVE: To compare the cumulative live birth rates obtained after cryopreservation of either pronucleate (PN) zygotes or early-cleavage (EC) embryos. DESIGN: Prospective randomized study. SETTING: University hospital. PATIENT(S): Three hundred eighty-two patients, involved in an IVF/ICSI program from January 1993 to December 1995, who had their supernumerary embryos cryopreserved either at the PN (group I) or EC (group II) stage. For 89 patients, cryopreservation of EC embryos was canceled because of poor embryo development (group III). Frozen-thawed embryo transfers performed up to December 1998 were considered. MAIN OUTCOME MEASURE(S): Age, oocytes, zygotes, cryopreserved and transferred embryos, damage after thawing, cumulative embryo scores, implantation, and cumulative live birth rates. RESULT(S): The clinical pregnancy and live birth rates were similar in all groups after fresh embryo transfers. Significantly higher implantation (10.5% vs. 5.9%) and pregnancy rates (19.5% vs. 10.9%; P< or = .02 per transfer after cryopreserved embryo transfers were obtained in group I versus group II, leading to higher cumulative pregnancy (55.5% vs. 38.6%; P < or = .002 and live birth rates (46.9% vs. 27.7%; P< or = .0001.Conclusion(s): The transfer of a maximum of three unselected embryos and freezing of all supernumerary PN zygotes can be safely done with significantly higher cumulative pregnancy chances than cryopreserving at a later EC stage.

Influence of CD4+/CD25+ regulatory T cells on atherogenesis in patients with end-stage kidney disease.

Atherosclerosis, which is influenced by both traditional and nontraditional cardiovascular risk factors and has been characterized as an inflammatory process, is considered to be the main cause of the elevated cardiovascular risk associated with chronic kidney disease. The inflammatory component of atherosclerosis can be separated into an innate immune response involving monocytes and macrophages that respond to the excessive uptake of lipoproteins and an adaptive immune response that involves antigen-specific T cells. Concurrent with the influx of immune cells to the site of atherosclerotic lesion, the role of the adaptive immune response gradually increases. One of those cells are represented by the CD4+/CD25+ Tregs, which play indispensable roles in the maintenance of natural self-tolerance and negative control of pathological, as well as physiological, immune responses. Altered self-antigens such as oxidized LDLs may induce the development of CD4+/CD25+ Tregs with atheroprotective properties. However, atherosclerosis may be promoted by an imbalance between regulatory and pathogenic immunity that may be represented by the low expression of the forkhead box transcription factor (Foxp3) in CD4+/CD25+ Tregs. Such a defect may break immunologic tolerance and alter both specific and bystander immune suppression, leading to exacerbation of plaque development. Patients with end-stage kidney disease (ESKD) display a cellular immune dysfunction and accelerated atherosclerosis. Uremic solutes that accumulate during ESKD may be involved in these processes. In patients with ESKD and especially in those that are chronically hemodialyzed, oxidative stress induced by oxidized LDLs may increase CD4+/CD25+ Treg sensitivity to Fas-mediated apoptosis as a consequence of specific dysregulation of IL-2 expression. This review will focus on the current state of knowledge regarding the influence of CD4+/CD25+ Tregs on atherogenesis in patients with ESKD, and the potential effect of statins on the circulating number and the functional properties of these cells.

Protection against Plasmodium falciparum challenge induced in Aotus monkeys by liver-stage antigen-3-derived long synthetic peptides.

The vaccine potential of Plasmodium falciparum liver stage antigen-3 (LSA3) was investigated in Aotus monkeys using two long synthetic peptides corresponding respectively to an N-terminal non-repeat peptide (NRP) and repeat 2 (R2) region of the LSA3, adjuvanted by ASO2. Both 100-222 (NRP) and 501-596 repeat peptides induced effector B- and T-cell responses in terms of antigen-driven antibodies and/or specific IFN-gamma secretion. Animals challenged with P. falciparum sporozoites were protected following immunization with either the NRP region alone or the NRP combined with the R2 repeat region, as compared with controls receiving the adjuvant alone. These results indicate that the NRP may be sufficient to induce full, sterile protection and confirm the vaccine potential of LSA3 previously demonstrated in chimpanzees and in Aotus.

Emergence, longevity and fecundity of Trissolcus basalis and Telenomus podisi after cold storage in the pupal stage

Pupae of Trissolcus basalis (Wollaston) and Telenomus podisi Ashmead (Hymenoptera: Scelionidae) were stored at 12ºC and 15ºC for 120-210 days, after different periods of parasitism at 18ºC in order to evaluate adult emergence, longevity and ovipositional capacity. There was no emergence of adults at 12ºC. The rate of emergence of parasitoids transferred to 15ºC at the beginning of the pupal stage was 1.5% and 26.3%, for T. basalis and T. podisi respectively, whereas those parasitoids transferred one day before the expected date of emergence at 18ºC showed 86.4% of emergence for T. basalis and 59.9% for T. podisi. Mean adult longevity was also significantly lower when pupae were transferred to 15ºC at the beginning of the pupal stage. Females emerged after storage and maintained for 120 to 210 days at 15ºC parasitized host eggs after transference to 25ºC; however, fecundity of T. podisi was reduced in about 80% after cold storage.