Spinal cord neuronal precursors generate multiple neuronal phenotypes in culture

Neuronal restricted precursors (NRPs) () can generate multiple neurotransmitter phenotypes during maturation in culture. Undifferentiated E-NCAM+ (embryonic neural cell adhesion molecule) immunoreactive NRPs are mitotically active and electrically immature, and they express only a subset of neuronal markers. Fully mature cells are postmitotic, process-bearing cells that are neurofilament-M and synaptophysin immunoreactive, and they synthesize and respond to different subsets of neurotransmitter molecules. Mature neurons that synthesize and respond to glycine, glutamate, GABA, dopamine, and acetylcholine can be identified by immunocytochemistry, RT-PCR, and calcium imaging in mass cultures. Individual NRPs also generate heterogeneous progeny as assessed by neurotransmitter response and synthesis, demonstrating the multipotent nature of the precursor cells. Differentiation can be modulated by sonic hedgehog (Shh) and bone morphogenetic protein (BMP)-2/4 molecules. Shh acts as a mitogen and inhibits differentiation (including cholinergic differentiation). BMP-2 and BMP-4, in contrast, inhibit cell division and promote differentiation (including cholinergic differentiation). Thus, a single neuronal precursor cell can differentiate into multiple classes of neurons, and this differentiation can be modulated by environmental signals.     

Association of HLA profiles with early plasma viral load, CD4+ cell count and rate of progression to AIDS following acute HIV-1 infection. Multicenter AIDS Cohort Study

The products of a growing number of genes have been shown to display seemingly contradictory functions; namely, the induction of tumorigenesis and the induction of apoptosis. Heregulin's involvement in oncogenesis occurs through its interactions with members of the EGF receptor tyrosine kinase family. Recently one isoform of heregulin, beta2b, was isolated in an in vitro screen for dominant, apoptosis-inducing genes in kidney epithelial cells. Here we show that heregulin is also capable of mediating apoptosis in human and murine mammary tumor cell lines and murine tumors. Furthermore, through transfection of the human breast cancer cell line MCF-7 with the truncated transmembrane/cytoplasmic segment of the heregulin gene, we show that the intracellular region of the heregulin precursor is sufficient for induction of apoptosis. Through the use of DNA fragmentation assays we also show that apoptosis occurs in cell lines established from heregulin-induced mammary gland tumors. TdT addition of digoxigenin labeled nucleotides to 3' OH ends of DNA breaks recapitulated these results in the actual tumors. Finally, over-expression of heregulin is shown to lead to the down-regulation of Bcl-2, an inhibitor of apoptosis. Conversely, the transfection of Bcl-2 into MCF-7 cells inhibits heregulin-mediated programmed cell death.     

T1/ST2 is preferentially expressed on murine Th2 cells, independent of interleukin 4, interleukin 5, and interleukin 10, and important for Th2 effector function

T helper (Th) cells can be categorized according to their cytokine expression. The differential induction of Th cells expressing Th1 and/or Th2 cytokines is key to the regulation of both protective and pathological immune responses. Cytokines are expressed transiently and there is a lack of stably expressed surface molecules, significant for functionally different types of Th cells. Such molecules are of utmost importance for the analysis and selective functional modulation of Th subsets and will provide new therapeutic strategies for the treatment of allergic or autoimmune diseases. To this end, we have identified potential target genes preferentially expressed in Th2 cells, expressing interleukin (IL)-4, IL-5, and/or IL-10, but not interferon-gamma. One such gene, T1/ST2, is expressed stably on both Th2 clones and Th2-polarized cells activated in vivo or in vitro. T1/ST2 expression is independent of induction by IL-4, IL-5, or IL-10. T1/ST2 plays a critical role in Th2 effector function. Administration of either a mAb against T1/ST2 or recombinant T1/ST2 fusion protein attenuates eosinophilic inflammation of the airways and suppresses IL-4 and IL-5 production in vivo following adoptive transfer of Th2 cells.     

Illuminated near card assessment of potential acuity in eyes with cataract

OBJECTIVE: This study aimed to determine the accuracy and potential role of the illuminated near card (INC) for predicting visual outcome after cataract surgery in eyes with and without comorbid disease. STUDY DESIGN: A consecutive case series. PARTICIPANTS: A total of 101 preoperative patients with cataracts participated. MAIN OUTCOME MEASURES: Accuracy of predicted postoperative distance acuity was measured. METHOD: The preoperative acuity obtained with the INC was compared by linear regression to the postoperative INC acuity and the postoperative distance acuity for 100 consecutive eyes undergoing cataract surgery. Variables analyzed were preoperative distance acuity and the presence or absence of comorbid disease. RESULTS: The preoperative INC acuity was significantly predictive of postoperative INC (P = 0.0005) and postoperative distance (P = 0.0007) acuities for the 100 eyes studied. For the subgroup of 15 eyes with 20/200 or worse, the preoperative INC acuity was not predictive of postoperative INC acuity (P = 0.8673) or postoperative distance acuity (P = 0.8789). For the 21 eyes with comorbid disease, the predictions were more accurate for postoperative INC acuity (P < 0.0001) and postoperative distance acuities (P < 0.0001) than for 64 eyes without comorbid disease: postoperative INC acuity (P = 0.0051), and postoperative distance acuity (P = 0.0046). The INC predicted postoperative distance acuity to within two lines in 98% of eyes when preoperative distance acuity was 20/100 or better. When the preoperative distance acuity was 20/200 or worse, the postoperative distance vision was predicted to within two lines in only 53% of the eyes. CONCLUSION: The INC can be a useful adjunct for predicting postoperative distance acuity in eyes with cataract that have preoperative distance acuity of 20/100 or better, particularly in eyes with comorbid disease, in which the clinical judgment of vision potential may be difficult.     

Outcome of transfusion of K:11 erythrocytes in a patient with anti-K11 antibody

Arteriovenous (AV) malformations are rare and may occur at various locations, like in the brain, liver, lungs, or in the subcutaneous tissue. This case report describes an AV malformation from the right occipital artery. The value of combined B-mode ultrasonography and color Doppler mode imaging of masses in the neck region is discussed.     

Morphology of protein-protein interfaces

This study investigated the usefulness of collagen plugging with VasoSeal in patients after PTCA compared to a control group having identical sheath dwell times and therefore comparable levels of anticoagulation. A total of 150 patients were enrolled in this prospective and randomized study. Sheaths were pulled at exactly 5 h after arterial puncture. Time to hemostasis and local complications were determined. There were no statistical differences in baseline characteristics. The mean time to hemostasis in the collagen group was significantly shorter (3 +/- 3 min) than that of the control group (17.4 +/- 7 min). At 24 h, 23% of the collagen group patients had a small, 1% a medium and 4% a large hematoma. In the control group, 32% had a small, 4% a medium sized, but no patient a large hematoma. After collagen, one patient developed a pseudoaneurysm needing vascular surgery. In the control group, no major complication occurred. Compared to patients with manual compression at an identical sheath dwell time and an identical level of anticoagulation, there was a significant reduction in time to hemostasis but no statistical difference regarding local complications. Although the incidence of medium or large hematoma was low, the trend towards a decreased risk of smaller hematomas seemed to be counterbalanced by an increased risk of larger hematomas.     

Identification of high-affinity binding sites for the insulin sensitizer rosiglitazone (BRL-49653) in rodent and human adipocytes using a radioiodinated ligand for peroxisomal proliferator-activated receptor gamma

A radioiodinated ligand, [125I]SB-236636 [(S)-(-)3-[4-[2-[N-(2-benzoxazolyl)-N-methylamino]ethoxy]3-[125I]i odo phenyl]2-ethoxy propanoic acid], which is specific for the gamma isoform of the peroxisomal proliferator activated receptor (PPARgamma), was developed. [125I]SB-236636 binds with high affinity to full-length human recombinant PPARgamma1 and to a GST (glutathione S-transferase) fusion protein containing the ligand binding domain of human PPARgamma1 (KD = 70 nM). Using this ligand, we characterized binding sites in adipose-derived cells from rat, mouse and humans. In competition experiments, rosiglitazone (BRL-49653), a potent antihyperglycemic agent, binds with high affinity to sites in intact adipocytes (IC50 = 12, 4 and 9 nM for rat, 3T3-L1 and human adipocytes, respectively). Binding affinities (IC50) of other thiazolidinediones for the ligand binding domain of PPARgamma1 were comparable with those determined in adipocytes and reflected the rank order of potencies of these agents as stimulants of glucose transport in 3T3-L1 adipocytes and antihyperglycemic agents in vivo: rosiglitazone > pioglitazone > troglitazone. Competition of [125I]SB-236636 binding was stereoselective in that the IC50 value of SB-219994, the (S)-enantiomer of an alpha-trifluoroethoxy propanoic acid insulin sensitizer, was 770-fold lower than that of SB-219993 [(R)-enantiomer] at recombinant human PPARgamma1. The higher binding affinity of SB-219994 also was evident in intact adipocytes and reflected its 100-fold greater potency as an antidiabetic agent. The results strongly suggest that the high-affinity binding site for [125I]SB-236636 in intact adipocytes is PPARgamma and that the pharmacology of insulin-sensitizer binding in rodent and human adipocytes is very similar and, moreover, predictive of antihyperglycemic activity in vivo.     

Quantitative peptide bioanalysis using column-switching nano liquid chromatography/mass spectrometry

Many endogenous peptides are circulating in bodily fluids at the low pmol l-1 range, placing high demands on the bioanalytical procedure. In order to analyze these minute concentrations in complex matrices, a miniaturized liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) bioanalysis method was developed using custom-made nanoLC columns (75 microns i.d.) and a micro-electrospray interface (micro ESI). To be able to analyze large sample volumes in order to cope with low biological analyte concentrations, the nanoLC/ESI-MS method was coupled to an on-line preconcentration (PC) system based on a strong anion-exchange material. This method was used to analyze endothelin peptides (ETs) in complex matrices, which are potent vasoconstrictors of M(r) approximately 2500 Da. The ET isoforms could be simultaneously analyzed with detection limits down to 30 pmol l-1 in cell supernatants (1.5 fmol on column). The method was linear from 50 to 2000 pmol l-1 with correlation coefficients of 0.99 for two of the three endothelin isoforms. Several other parameters, such as matrix effects and recovery, were also investigated.     

Enhancement of 5-aminolaevulinic acid-induced photodynamic therapy in normal rat colon using hydroxypyridinone iron-chelating agents

Currently, the clinical use of 5-aminolaevulinic acid (ALA)-induced protoporphyrin IX (PPIX) for photodynamic therapy (PDT) is limited by the maximum tolerated oral ALA dose (60 mg kg(-1)). This study investigates whether hydroxypyridinone iron-chelating agents can be used to enhance the tissue levels of PPIX, without increasing the administered dose of ALA. Quantitative charge-coupled device (CCD) fluorescence microscopy was employed to study PPIX fluorescence pharmacokinetics in the colon of normal Wistar rats. The iron chelator, CP94, when administered with ALA was found to produce double the PPIX fluorescence in the colonic mucosa, compared with the same dose of ALA given alone and to be more effective than the other iron chelator studied, CP20. Microspectrofluorimetric studies demonstrated that PPIX was the predominant porphyrin species present. PDT studies conducted on the colonic mucosa showed that the simultaneous administration of 100 mg kg(-1) CP94 i.v. and 50 mg kg(-1) ALA i.v. produced an area of necrosis three times larger than similar parameters without the iron-chelating agent with the same light dose. It is possible, therefore, to increase the amount of necrosis produced by ALA-induced PDT substantially, without increasing the administered dose of ALA, through the simultaneous administration of the iron-chelating agent, CP94.     

Chaos under canvas: a Salmonella enteritidis PT 6B outbreak

Triphenylselenonium chloride, a novel synthetic organic selenium compound in which selenium is bonded to three unsubstituted benzene rings, possesses significant chemopreventive activity against chemically-induced mammary carcinogenesis. The effects of triphenylselenonium chloride on a mammary tumor cell line (MOD) were compared to selenite, a reference compound in selenium chemoprevention research. It was observed that triphenylselenonium chloride treatment exerted a cytostatic effect in the absence of membrane damage or DNA strand breaks. The observed cytostasis was associated with a selenium concentration-dependent inhibition of cell proliferation, measured by [3H]thymidine incorporation into DNA, and delayed cell cycle progression. In contrast, selenite treatment rapidly induced DNA damage and cell death. These marked differences were observed across the same levels of cellular selenium. In addition, triphenylselenonium chloride treatment increased glucose consumption and lactate production, indicating an effect of the compound on cellular energy metabolism. Collectively these observations demonstrate that the toxic activities associated with selenite treatment do not occur when cells are treated with triphenylselenonium chloride. This compound represents a new type of selenium compound that exerts significant cellular effects through mechanisms distinct from those induced by selenite.