Modification of tandem mass spectrometric method to permit simultaneous quantification of 17 anti-HIV drugs which include atazanavir and tipranavir.
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Anti-HIV Agents↗
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Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
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Radiofrequency ablation permits an effective treatment for colorectal liver metastasis.
BACKGROUND: Radiofrequency ablation (RFA) has become an important adjunct to modern liver surgery. However, scant knowledge on long-term outcome of RFA for colorectal liver metastasis is available, nowadays. METHODS: This is a prospective clinical study of patients with liver metastasis of colorectal cancer who were treated by RFA between April 1, 1998, and November 30, 2004. Forty-seven patients with 147 liver metastases were treated with RFA in a total of 70 interventions. A metastasis resection was not feasible in 80% of the interventions. All the patients were followed up at regular intervals with contrast-enhanced computed tomography (CT) and laboratory tests including carcinoembryonic antigen (CEA). RESULTS: No RFA-related mortality occurred. The median follow-up time after the diagnosis of liver metastasis was 33 months. The RFA-related morbidity was 7%. After the RFA, the expected median overall survival rate is, to date, 39 months. Overall survival rates at 1, 2 and 3 years were 88%, 80% and 57%, respectively. Local recurrence rates reached 8.8% overall and 1.6% for metastasis smaller than 3cm in diameter. No local recurrence occurred for metastasis smaller than 3cm in diameter if treated with the newest RFA device. CONCLUSIONS: Excellent local tumour control was achieved with radiofrequency ablation of small liver metastasis. The expected overall survival rate of patients with RFA for unresectable or non-resected colorectal liver metastasis improved in comparison with the survival rate reported following the natural course (best supportive care) or chemotherapy. The low local recurrence rate of metastases of less than 3cm challenges the results obtained by the more invasive treatment of conventional liver surgery.
Adult↗
Local down-regulation of myelin-associated glycoprotein permits axonal sprouting with chronic nerve compression injury.
Chronic nerve compression (CNC) injuries induce a robust Schwann cell proliferation in a distinct spatial and temporal pattern, which is accompanied by an increase in the number of small un-myelinated axons in the area of the injury. These findings suggest that this local proliferation of Schwann cells may induce local axonal sprouting. Here, we use quantitative electron microscopic techniques to define the nature of this sprouting response, and explore whether the local sprouting is in response to down-regulation of expression of myelin-associated glycoprotein (MAG) by proliferating Schwann cells. Axonal sprouting was observed without evidence of Wallerian degeneration in the outer region of CNC-injured nerves with a noticeable increase in Remak bundles within this region of injury. Immunolabeling of teased nerve fibers and Western blot analysis of nerves from CNC-injured animals revealed a local down-regulation of MAG protein within the zone of injury. Moreover, local delivery of purified MAG protein intraneurally at the time of CNC model creation abrogates the axonal sprouting response. These data demonstrate that CNC injury triggers axonal sprouting and suggests that a local down-regulation of MAG within the peripheral nerve secondary to CNC injury is the critical signal for the sprouting response.
Animals↗
Laparoscopic hepatic resection using saline-enhanced electrocautery permits short hospital stays.
Laparoscopic hepatic resection has been reported to yield lower morbidity and shorter hospital stays than open resection. However, few studies have evaluated patient and technical factors associated with short hospital stays. We conducted a retrospective review of patients undergoing laparoscopic hepatic resection at our institution from May 2002 to February 2004. Patient and operative factors were analyzed with respect to time to discharge. Seventeen patients underwent 10 wedge resections and seven segmentectomies or bisegmentectomies. There were no mortalities, conversions to open procedure, clinically evident bile leaks, or transfusion requirements. Eleven patients were discharged within 24 hours. When compared with those discharged later than 24 hours, there were fewer patients with advanced ASA classification (0 versus 3 in ASA class 3, p < 0.05). With appropriate patient selection, laparoscopic hepatic resections may be safely performed, result in short hospital stays, and are facilitated by technologies such as saline-enhanced electrocautery and endoscopic ultrasound. Information reflected in advanced ASA class may predict patients unlikely to be discharged within 24 hours.
Adult↗
Cell surface recycling of internalized antigen permits dendritic cell priming of B cells.
Dendritic cells process internalized antigens to present degradative products on MHC for TCR recognition. Because antigen-exposed DCs also induce humoral immunity, DCs must also retain antigen in its native state for the engagement of BCR on B cells. Here, we demonstrate that antigen endocytosed by the inhibitory Fc receptor, FcgammaRIIB, accesses a non-degradative intracellular vesicular compartment that recycles to the cell surface, enabling interaction of native antigen with BCR on B cells. Immunization with IgG-opsonized, T independent antigens leads to enhanced humoral responses in a FcgammaRIIB and complement dependent manner. IC-loaded DCs trafficking to the splenic marginal zone can prime a T independent response in an FcgammaRIIB-dependent manner. Thus dendritic cells are equipped with both non-degradative and degradative antigen uptake pathways to facilitate antigen presentation to both B and T cells.
Animals↗
Use of anti-CD3/CD28 mAb coupled magnetic beads permitting subsequent phenotypic analysis of activated human T cells by indirect immunofluorescence.
Functional analysis of T lymphocytes requires in vitro stimulation of these cells under experimental conditions that mimic as closely as possible physiological in vivo stimulation and that involve antigen/T cell receptor (TCR)-mediated activation. Because of the low frequency of antigen-specific T cells in human clinical samples, stimulation with a combination of anti-CD3 and anti-CD28 monoclonal antibodies (mAbs) is a preferred method. Interaction of these mAbs with their ligand results in modulation of the mAb-ligand complex from the cell surface. However, as a result of incomplete modulation, CD3/CD28 mAb complexes often remain at the cell surface, thereby precluding subsequent indirect immunofluorescence and flow cytometry analysis using mouse immunoglobulin (Ig)-specific antibodies. This is of importance in situations in which no specific fluorochrome-conjugated mAbs are available, such as in screening procedures of Ig-containing hybridoma culture supernatants. We propose here the use of CD3/CD28 mAbs, linked to magnetic beads allowing standardization of the activation conditions, optimal activation of T cells and complete modulation of antigen-antibody complexes from the cell surface.
ADP-ribosyl Cyclase↗
Novel assay format permitting the prolonged use of regeneration-based sensor chip technology.
A polyclonal antibody raised against morphine-3-glucuronide (M3G, the main metabolite of heroin and morphine) was used in the development of a novel assay format using a surface plasmon resonance (SPR)-based biosensor. Previously developed assays have generated calibration curves based on differences in the quantity of response units binding to the surface of a chip coated with the analyte. The novel assay described here was based on the development of a standard curve using the slope of a series of consecutive binding interactions. Using this format, regeneration between each assay cycle was no longer required. This increased the useable life span of the chip surface and, as a result, decreased the cost associated with the assay. Thus, at least 15 binding interactions could be carried out before the saturation of antibody on the surface of the chip caused the response to deviate significantly from linearity. After 15 nonregenerated binding interactions, the slope still remained within 1.5% of the slope after a single binding event. Analysis time, and the sample volumes required were also markedly decreased while sensitivity was enhanced. The inhibition assay developed had a detection range of 270 to 17,500 pg ml(-1).
Antibodies↗
Ex vivo expansion of human CD4+ CD25high regulatory T cells from transplant recipients permits functional analysis of small blood samples.
Regulatory T cell (Treg) function is associated with immune tolerance and could serve as a biomarker for optimization of immunosuppressive regimens. This is hampered by the limited number of Treg in the blood circulation; functional Treg analysis requires large volumes of blood or is dependent on indirect analysis. A more attractive strategy is the ex vivo expansion of Treg, provided the original T cell pool remains unaltered. Here, we show that it is possible to ex vivo expand Treg from limited amounts of blood, preserving the original TCR Vbeta repertoire and suppressive capacity. The protocol proved successful in selected renal transplant recipients and leukopenic patients. Sampling of 10-20 ml blood sufficed; Treg numbers increased over 100-fold during the 2-3 week expansion period, easily reaching cell numbers required for functional analysis (>10(6) cells). This protocol will facilitate the monitoring of Treg function in patients in order to address the role of Treg in transplantation tolerance.
CD4-Positive T-Lymphocytes↗
DNA-looping by RXR tetramers permits transcriptional regulation "at a distance".
RXR, a member of the superfamily of nuclear hormone receptors, regulates gene transcription in response to 9-cis-retinoic acid. We previously showed that, among nuclear receptors, RXR is unique in that it self-associates into homotetramers, and that these tetramers dissociate rapidly upon ligation. Here, we report that binding of RXR tetramers to DNA containing two RXR response elements results in a dramatic DNA-looping. RXR can thus juxtapose distant DNA sequences, enabling transcriptional regulation by far-upstream factors. We show that RXR functions as a DNA architectural factor and that, while this activity is regulated by 9-cis-retinoic acid, it is distinct from and independent of the receptor's intrinsic transcriptional activity. The data establish RXR as the first identified architectural factor whose activity is regulated by a small ligand, and demonstrate a novel mechanism of transcriptional regulation by retinoids.
Animals↗
Quantitative no-net-flux microdialysis permits detection of increases and decreases in dopamine uptake in mouse nucleus accumbens.
A number of investigators are using the quantitative no-net-flux microdialysis technique to monitor basal neurotransmitter dynamics in discrete brain regions of behaving animals. The predictive validity of the probe extraction fraction (Ed) for quantifying decreases in the rate of dopamine (DA) clearance from the extracellular space is well documented. It was recently suggested, however, that Ed may be insensitive to increases in DA clearance. Here we report that the Ed for DA in the nucleus accumbens (NAc) of the behaving mouse is increased following pharmacological inactivation of kappa-opioid receptors, a treatment previously shown to augment DA uptake. The Ed obtained in control mice and those that received the long-acting kappa-opioid receptor antagonist, nor-binaltorphimine (nor-BNI), satisfied the requirement that the mean values of each were lower than the mean value in vitro for the same probes immersed in well-stirred artificial cerebrospinal fluid. The Ed was increased in the NAc of nor-BNI-treated mice as compared to saline-treated control animals. The corresponding increase in the DA uptake rate was quantified by using the Ed values to calculate a change in the apparent clearance rate constant. Nor-BNI treatment did not alter the apparent extracellular dopamine concentration represented by the point of no-net-flux indicating that the rates of DA uptake and release were both increased.
Animals↗
A new role for androgen in testicular descent: permitting gubernacular cell proliferation in response to the neuropeptide, calcitonin gene-related peptide.
BACKGROUND/PURPOSE: Cell proliferation at the gubernacular tip increases in response to exogenous calcitonin gene-related peptide (CGRP) during migration into the scrotum. Calcitonin gene-related peptide is contained in the masculinized sensory branches of the genitofemoral nerve. We tested the independent effects of chemical sensory nerve disruption and prenatal androgen blockade on the in vitro gubernacular proliferative response to CGRP. METHODS: Neonatal Sprague-Dawley rats were injected with capsaicin, a sensory nerve toxin, and gubernacula dissected 2 days later (D2). Sprague-Dawley dams were injected with flutamide, an androgen receptor antagonist, between days 15 and 19 of gestation. Flutamide pretreated males, and normal neonatal rats, were dissected at D0 and D2. Gubernacula were cultured for 24 hours +/- CGRP, pulse-labelled for the last 4 hours of culture with bromodeoxyuridine, a thymidine analogue marker for DNA replication, sectioned, and stained using immunohistochemistry. The percentage of positively staining cells in the gubernacular tip was calculated from three separate counts by a blinded observer and compared using analysis of variance. RESULTS: Normal D0 gubernacular tips showed a significant response of cell proliferation to exogenous CGRP (34% vs 9% in controls, P < .001), which resolved by day 2 (16% vs 12%, P > .05). Calcitonin gene-related peptide markedly increased cell proliferation in D2 capsaicin pretreated gubernacula compared with controls (25% vs 14%, P < .01) and normal D2 gubernacula cultured with CGRP (P < .01). D0 flutamide pretreated cultured with CGRP showed no increase in cell proliferation compared with controls (16% vs 11%), but a small response was seen by D2 (19% vs 9%, P < .05). There was no significant difference between proliferation rates in the control groups. CONCLUSIONS: Sensory innervation interruption sensitises the gubernaculum to exogenous CGRP, suggesting upregulation of CGRP receptors. In contrast, androgen blockade abolishes the increased rate of cell proliferation within the gubernacular tip. We conclude that androgens are necessary to "preprogramme" the proliferative response of the gubernaculum to CGRP.
Animals↗
Intravascular administration of tumor tropic neural progenitor cells permits targeted delivery of interferon-beta and restricts tumor growth in a murine model of disseminated neuroblastoma.
BACKGROUND: Interferon-beta (IFN-beta) has potent antitumor activity; however, systemic toxicity has limited its clinical use. We investigated the potential of targeted delivery using tumor-tropic neural progenitor cells (NPCs) transduced to express human IFN-beta (hIFN-beta). METHODS: Disseminated neuroblastoma was established in SCID mice by tail vein injection of tumor cells. Fourteen days after tumor cell inoculation, systemic disease was confirmed with bioluminescence imaging (BLI). Mice were then treated by intravenous injection of human F3.C1 NPCs that had been transduced with a replication deficient adenovirus to overexpress hIFN-beta (F3-IFN-beta). Two injections were given: the first at 14 days and the second at 28 days following tumor cell injection. Control mice received NPCs transduced with empty vector adenovirus at the same time points. Progression of disease was monitored using BLI. At sacrifice, organ weights and histology further evaluated tumor burden. RESULTS: After initiation of therapy, BLI demonstrated a significant decrease in the rate of disease progression in mice receiving F3-IFN-beta. At necropsy, control mice had bulky tumor replacing the liver and kidneys, as well as extensive retroperitoneal and mediastinal adenopathy. Impressively, these sites within mice receiving F3-IFN-beta therapy appeared grossly normal with the exception of small nodules within the kidneys of some of the F3-IFN-beta-treated mice. The accumulation of F3.C1 cells within sites of tumor growth was confirmed by fluorescence imaging. Importantly, systemic levels of hIFN-beta in the treated mice remained below detectable levels. CONCLUSIONS: These data indicate that in this model of disseminated neuroblastoma, the tumor-tropic property of F3.C1 NPCs was exploited to target delivery of IFN-beta to disseminated tissue foci, resulting in significant tumor growth delay. The described novel approach for effective IFN-beta therapy may circumvent limitations associated with the systemic toxicity of IFN-beta.
Adenoviridae↗
An immunocytometric assay based on dengue infection via DC-SIGN permits rapid measurement of anti-dengue neutralizing antibodies.
Dengue remains a global public health threat and development of a safe and effective vaccine is a principal public health goal. The primary correlate of immunity is thought to be neutralizing antibodies. Currently, the plaque reduction neutralization test (PRNT) is the gold standard measure of dengue neutralizing antibody responses, but this test is limited by time-consuming performance. In addition, some feel that use of viral strains adapted to grow in Vero or BHK cells may not accurately reflect protective responses. A human cell line transfected to express a putative natural dengue receptor, DC-SIGN (CD209), was used to measure antibody-mediated dengue neutralization. Using neutralizing monoclonal antibodies, immune sera, and laboratory adapted dengue viruses, serotype-specific neutralizing activity was demonstrated similar to that seen in the Vero PRNT. Importantly, serotype-specific neutralizing activity against recently isolated dengue strains with less heterotypic cross-neutralization than laboratory adapted viruses was also demonstrated.
Animals↗
The CDK inhibitor p27 enhances neural differentiation in pluripotent NTERA2 human EC cells, but does not permit differentiation of 2102Ep nullipotent human EC cells.
Embryonal carcinoma (EC) cells, the stem cells of teratocarcinomas, are the malignant counterparts of pluripotent embryonic stem (ES) cells, but commonly exhibit a reduced ability to differentiate, presumably because of continual selection for genetic changes that alter the balance between self-renewal, differentiation and apoptosis in favour of self-renewal. To explore the nature of the genetic changes that promote nullipotency, we have compared two human EC cell lines, a 'nullipotent' line, 2102Ep, and a 'pluripotent' line, NTERA2. A hybrid derived by fusion of these cells differentiates in response to retinoic acid but, unlike the parental NTERA2 line, does not form terminally differentiated neurons. This implies that the nullipotent EC cell line, 2102Ep, differs in expression of at least two functions in comparison with the NTERA2 pluripotent line, one affecting commitment to differentiation, and one affecting terminal neural differentiation. We have now investigated the possible role of the CDK inhibitor, p27kip1 (p27) in commitment and terminal differentiation. In NTERA2, but not in 2102Ep cells, retinoic acid induces up-regulation of p27 expression, suggesting that 2102Ep cells lack this capacity. However, constitutive expression of a p27 transgene does not overcome the block to differentiation in the 2102Ep parental cells; commitment to differentiation must be blocked elsewhere. On the other hand, constitutive over-expression of p27 from a transgene enhances the neural differentiation of NTERA2 cells. Our results suggest that p27 plays a role in terminal neuronal differentiation of human EC cells, but not in their initial commitment to differentiation, and that other factors, possibly Cyclin D2, specifically limit its ability to promote neural differentiation.
Base Sequence↗
State laws permitting intoxication exclusions in insurance contracts: implications for public health policy and practice.
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Alcohol Drinking↗
The HOPE-technique permits Northern blot and microarray analyses in paraffin-embedded tissues.
There is an increasing demand for tissue samples that, after having been used for conventional histologic examination, are also suited for molecular analyses. As to formalin-fixed, paraffin embedded (FFPE) tissue, the latter applications are very limited. The HOPE (Hepes-Glutamic acid buffer mediated Organic solvent Protection Effect) technique comprises a new protection-solution with an organic buffer, with acetone as the only dehydrating agent, and pure paraffin of 52-54 degrees C melting temperature, allowing for all pathologic routine investigations. In contrast to FFPE tissue, the HOPE-technique allows for the application of molecular methods, such as high molecular DNA and RNA isolation, which can be used for PCR and reverse transcription PCR (RT-PCR). In this study, we investigated whether RNA from HOPE-fixed tissue samples is suitable for Northern blot and microarray analyses. RNAs of two HOPE-fixed breast cancer specimens of different histologic grade were used to carry out an array experiment. It turned out that RNA from HOPE-fixed tissue is of high quality and can be successfully used for array experiments. In addition, by detecting GAPDH and high mobility group protein gene B1 (HMGB1)-specific transcripts, we were able to demonstrate that RNA from HOPE-fixed tissue can also be used for Northern blot hybridization.
Blotting, Northern↗
Four-dimensional computed tomography permits preoperative planning for directed parathyroidectomy.
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Humans↗
"In two minds"--consideration of evolutionary precursors permits a more integrative theory.
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Behavior↗