Human retinal pigment epithelial cell interleukin-8 and monocyte chemotactic protein-1 modulation by T-lymphocyte products

PURPOSE: The purpose of the study was to examine the effect of T-lymphocyte products on human retinal pigment epithelial (HRPE) cell interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) secretion and gene expression. METHODS: HRPE cells were stimulated for 2, 4, 8, or 24 hours with 20% conditioned media (CM) from T-lymphocytes stimulated with CD3 or CD28 monoclonal antibodies (mAbs) or phorbol myristic acid. In some experiments, CM from CD3 mAb-stimulated T-lymphocytes was preincubated with neutralizing anti-(alpha)-tumor necrosis factor (TNF), alpha-interferon-gamma (IFN-gamma), or alpha-interleukin-1 (IL-1) mAb (control) to determine the contributions of each of these cytokines to HRPE chemokine induction by stimulated T-lymphocyte CM. HRPE cells were stimulated for 8 and 24 hours with IL-1 beta (0.2 to 20.0 ng/ml) (positive control), TNF-alpha (0.2 to 20.0 ng/ml) (positive control), IFN-gamma (1 to 1000 U/ml), IFN-gamma + IL-1 beta, IFN-gamma + TNF-alpha. Interleukin-2 (IL-2; 100 ng/ml) alone or in combination with IL-1 beta, TNF-alpha, or IFN-gamma also was tested. Enzyme-linked immunosorbent assay (ELISA) and Northern blot analyses were performed to determine secreted IL-8 and MCP-1 and their steady state mRNA expression, respectively. RESULTS: ELISA showed significant increases in HRPE IL-8 and MCP-1 secretion by CM from T-lymphocytes stimulated with CD3 or CD3 + CD28 mAb. Smaller, but significant, increases in IL-8 and MCP-1 resulted from CM phorbol myristic acid-stimulated T-lymphocytes. CM preincubated with neutralizing alpha-TNF or alpha-IFN-gamma mAb induced significantly less HRPE IL-8 and MCP-1, whereas preincubation of CM with neutralizing alpha-IL-1 mAb failed to inhibit CM-induced IL-8 or MCP-1. Northern blot analysis showed increased HRPE IL-8 and MCP-1 mRNA expression within 2 hours of stimulation and was maintained up to 24 hours. CM from T-lymphocytes stimulated with CD3 mAb or CD3 + CD28 mAb produced the greatest increases in IL-8 and MCP-1 mRNA. IFN-gamma induced dose-dependent increases in HRPE MCP-1, but not IL-8, IFN-gamma potentiated IL-1 beta and TNF-alpha-induced MCP-1 production, but showed little modulation of IL-1 beta and TNF-alpha-induced IL-8 production. IL-2 did not induce HRPE IL-8 or MCP-1, nor did it modulate the effects of the other cytokines. Northern blot analysis confirmed the ELISA results. CONCLUSIONS: T-lymphocyte secretions induce HRPE IL-8 and MCP-1 gene expression and secretion. TNF and IFN-gamma appear to be necessary components of T-lymphocyte CM for the induction of HRPE IL-8 and MCP-1. IFN-gamma alone induces HRPE MCP-1, albeit to a lesser extent than would IL-1 beta or TNF-alpha, and potentiates IL-1 beta- and TNF-alpha-induced HRPE MCP-1. IL-2 does not appear to modulate cytokine-induced HRPE IL-8 or MCP-1.     

Reverse micellar extraction of antibiotics from aqueous solutions

PURPOSE: We reviewed 1,360 EEG reports for all patients studied in two different neurophysiology laboratories during 1 calendar year to determine whether epileptiform discharges have a hemispheric dominance. METHODS: Both inpatients and outpatients, with or without epilepsy, were included. RESULTS: Ninety-four records (6.9%) demonstrated generalized epileptiform activity. Of 95 EEG reports indicating spikes solely from one hemisphere, spikes arose from the left in 61 and from the right in 34. Among 50 other records with bilateral independent spikes with lateralization, 40 were left hemisphere dominant and 10 were right hemisphere dominant. CONCLUSIONS: These findings raise the possibility that the left cerebral hemisphere may generate focal epilepsy more frequently than the right.     

Resident attrition from family practice residencies: United States versus international medical graduates

BACKGROUND: Attrition of residents from family practice residency programs may cause significant problems for faculty, residents, and patients. The objective of this study was to determine international medical graduates' attrition rate from family practice residencies, compared with US medical school graduates. METHODS: Surveys were sent to all family practice residency program directors asking them to calculate their attrition rate for a 10-year period. RESULTS: The overall response rate was 56.6%, but interpretable responses were received from 45% of all civilian, continental US family practice residencies. Responding programs did not differ from all family practice programs with respect to program overall. Of those residents leaving, 63% did so to enter other specialties. The attrition rate was 18.5% for international graduates, compared with 7.8% for US graduates (P < .0001). International graduates enrolled outside of the National Resident Matching Program (NRMP) were most likely to leave programs before completion. CONCLUSIONS: Attrition rates from family practice residency programs are higher for international medical graduates than for US graduates. International graduates enrolled outside of the NRMP were most likely to leave a program.     

Sequencing of the alpha-synuclein gene in a large series of cases of familial Parkinson''s disease fails to reveal any further mutations. The European Consortium on Genetic Susceptibility in Parkinson''s Disease (GSPD)

PURPOSE: Tumor hypoxia may be an important factor predicting relapse following radiation therapy. This study was designed to determine the relationship between the oxygenation parameters measured using a polarographic oxygen electrode, prior to and during treatment in patients with cervix cancer, and to assess these results with regard to patient survival. MATERIALS AND METHODS: Forty-three patients had pretreatment oxygen assays performed and measurements repeated following external beam radiation to a median dose of 50 Gy (range 26-52 Gy). Stage distribution showed 15 patients in Stage IB, 17 in Stage II, and 11 in Stage III. The median tumor size was 5 cm (range 3-10 cm). RESULTS: The median proportion of pO2 values <5 mm Hg (the HP5) was 41% following radiation, and the median pO2 was 12 mm Hg. These results were not significantly different from the pretreatment HP5 or pO2 of 37% and 12 mm Hg, respectively. Disease-free survival at 2 years was 50% in patients with posttreatment HP5 < or =50%, compared to 60% when posttreatment HP5 was >50% (p = 0.35). CONCLUSIONS: Unlike pretreatment results, tumour oxygenation measured following external radiation does not appear to be a useful predictive assay in patients with cervical cancer.     

Endocytosis and recycling of neurokinin 1 receptors in enteric neurons

Neurotransmission depends on the availability of transmitter and on the presence of functional, high-affinity receptors at the plasma membrane that are capable of binding ligand. The pathway, mechanism and function of endocytosis and recycling of the substance P or neurokinin 1 receptor in enteric neurons were studied using fluorescent substance P, receptor antibodies and confocal microscopy. In both the soma and neurites, substance P induced rapid, clathrin-mediated internalization of the neurokinin 1 receptor into early endosomes, which also contained the transferrin receptor. After 4-8 h, there was a return in surface neurokinin 1 receptor immunoreactivity in the soma, which was not prevented by cycloheximide, and was thus independent of new protein synthesis. This return was prevented by acidotropic agents, therefore required endosomal acidification. This suggests that the neurokinin 1 receptor recycles in the soma. In contrast, in neurites, substance P and the neurokinin 1 receptor remained in endosomes and recycling was not detected. Neurons of the myenteric plexus were heavily innervated by substance P-containing nerve fibers, and K(+)-stimulated release of endogenous substance P from cultured neurons induced internalization of the neurokinin 1-receptor. Therefore, endogenous substance P may induce endocytosis of the neurokinin 1 receptor. In the soma, endocytosis and recycling correlated with loss and recovery of functional binding sites for substance P. suggesting that this process contributes to the regulation of peptidergic neurotransmission. Thus, ligand-induced endocytosis of the neurokinin 1 receptor in myenteric neurons is associated with a loss of surface receptors and functional binding sites. Since release of endogenous substance P induces neurokinin 1 receptor internalization, and neurokinin 1 receptor neurons are innervated by substance P-containing fibers, endocytosis of neuropeptide receptors may regulate neurotransmission.     

The muscarinic M1 agonist xanomeline increases soluble amyloid precursor protein release from Chinese hamster ovary-m1 cells

The functionally selective M1 agonist xanomeline, which is currently undergoing clinical trials as a therapy for Alzheimer's disease, was compared to the muscarinic agonist carbachol for effects on secretion of soluble amyloid precursor protein (APPs) from Chinese hamster ovary cells transfected with the human m1 receptor (CHO-m1). Release of APPs from CHO-m1 cells was increased maximally (4-10 fold) by 100 microM carbachol (EC50 = 11 microM) and by 100 nM xanomeline (EC50 = 10 nM). Stimulation of APPs secretion by xanomeline and carbachol was blocked by preincubation with 1 microM atropine. Carbachol did not stimulate APPs secretion from non-transfected CHO cells. Pilocarpine at 1 mM also increased APPs release. The efficacy of carbachol, xanomeline and pilocarpine for stimulating APPs secretion did not differ significantly. Activation of protein kinase C (PKC) in m1 transfected cell lines by 1 microM phorbol dibutyrate (PDBu) increased APPs release, and this was inhibited 97% by the PKC inhibitor bisindolemalemide. The PKC inhibitor decreased xanomeline and carbachol-stimulated APPs secretion by only 25-30%. These results demonstrate that xanomeline increased APPs release by activation of m1 muscarinic receptors and support the possibility that cholinergic replacement therapy for Alzheimer's Disease may reduce amyloid deposition.     

Looking for the answer

Metabotropic glutamate receptors (mGluRs) were identified in olfactory receptor neurons of the channel catfish, Ictalurus punctatus, by polymerase chain reaction. DNA sequence analysis confirmed the presence of two subtypes, mGluR1 and mGluR3, that were coexpressed with each other and with the putative odorant receptors within single olfactory receptor neurons. Immunocytochemical data showed that both mGluR subtypes were expressed in the apical dendrites and some cilia of olfactory neurons. Pharmacological analysis showed that antagonists to each mGluR subtype significantly decreased the electrophysiological response to odorant amino acids. alpha-Methyl-L-CCG1/(2S,3S,4S)-2-methyl-2-(carboxycyclopropyl++ +)glycine (MCCG), a known antagonist to mGluR3, and (S)-4-carboxyphenylglycine (S-4CPG), a specific antagonist to mGluR1, each significantly reduced olfactory receptor responses to L-glutamate. S-4CPG and MCCG reduced the glutamate response to 54% and 56% of control, respectively, which was significantly greater than their effect on a neutral amino acid odorant, methionine. These significant reductions of odorant response by the antagonists, taken with the expression of these receptors throughout the dendritic and ciliated portions of some olfactory receptor neurons, suggest that these mGluRs may be involved in olfactory reception and signal transduction.     

Institutional arrangements for flood hazard management in Malaysia: an evaluation using the criteria approach

Institutional aspects of flood hazards significantly affect their outcomes in Malaysia. Institutional arrangements to deal with floods include: legislative activity, organisational structures, attitudes and sub-culture, and policies and instruments. When assessed in terms of four specific criteria, institutional aspects of flood hazards are found to be largely inadequate. Disaster reduction programmes are over-dependent on a reactive approach based largely on technology and not even aimed at floods specifically. Structural flood reduction measures are the predominant management tool and, although the importance of non-structural measures is recognised, thus far they have been under-employed. Current laws and regulations with regard to flood management are also insufficient and both the financial and human resources of flood hazard organisations are generally found to be wanting. Finally, economic efficiency, equity and public accountability issues are not adequately addressed by institutional arrangements for flood hazards.