Use of the ARPE-19 cell line as a model of RPE polarity: basolateral secretion of FGF5

PURPOSE: To determine the polarity of fibroblast growth factor 5 (FGF5) secretions from retinal pigment epithelium (RPE) cells and to examine the viability and utility of the ARPE-19 cell line as a model for the study of RPE polarity. METHODS: Influenza infection and adenovirus-mediated gene transfer were used to deliver and express genes encoding influenza hemagglutinin (HA), p75-NTR (a neurotrophin receptor), low-density lipoprotein (LDL) receptor (LDLR), and FGF5 in confluent monolayers of ARPE-19 cells. The localization of HA, p75-NTR, and LDLR was determined by confocal microscopy. Domain selective biotinylation assays were used to quantitatively determine the polarities of p75-NTR and LDLR. The secretion of FGF5 into the apical and basal media of ARPE-19 cultures was examined by immunoblot analysis of conditioned media. RESULTS: Hemagglutinin and p75-NTR were found to be localized on the apical surface of infected and transduced ARPE-19 cells. In contrast, LDLR was associated preferentially with the basolateral membrane of ARPE-19 cells. Biotinylation studies indicated that 84% of p75-NTR was present on the apical surface, and 79% of LDLR was basolaterally polarized. Over the course of 6 hours, more than 90% of the total secreted FGF5 protein accumulated in the basolateral media. CONCLUSIONS: ARPE-19 cells exhibit a polarized distribution of cell surface markers when examined by either confocal microscopy or surface-labeling assays. This indicates that the ARPE-19 cell line is a valid model for studies of RPE cell polarity. FGF5, a secreted protein normally produced by RPE cells, is accumulated preferentially in the basal media after only 6 hours, suggesting that it is vectorially secreted from the basolateral surface of ARPE-19 cells.     

Comparative studies of O,O-dialkyl-O-chloromethylchloroformimino phosphates: interaction with neuropathy target esterase and acetylcholinesterase

Acetylcholinesterase (AChE) and neuropathy target esterase (neurotoxic esterase, NTE) are two major target enzymes for organophosphorus (OP) esters. The relative potency of an OP ester to react with AChE or with NTE in vitro correlates with its relative potency in vivo to cause acute toxicity (death) or organopohosphate-induced delayed neurotoxicity (OPIDN). On this basis extrapolation from in vitro to in vivo data now seems justifiable to predict risk of OPIDN. The kinetics of NTE and AChE inhibition by experimental pesticides of the general formula (RO)2P(O)ON=CClCH2Cl, where R = methyl, ethyl, isopropyl, propyl, isobutyl, butyl, pentyl, has been studied. Compounds with short R (methyl, ethyl) were shown to be far more potent inhibitors of AChE than NTE. Both anti-NTE activity, selectivity for NTE and, correspondingly, the propensity of compounds to cause OPIDN rise with increasing their hydrophobicity. A high value of ki(NTE)/ki(AChE) for R = pentyl suggests that this compound would have the potential to cause OPIDN at doses lower than the LD50. A quantitative structure-activity relationships (QSAR) analysis indicated that NTE and AChE have different structural and electronic requirements for their respective OP inhibitors.     

Benign paroxysmal tonic upward gaze syndrome

INTRODUCTION: The benign paroxysmal tonic upward gaze syndrome (BPTUG) is a rare condition. We present two new cases analyzing the clinical, genetic, evolution and therapeutic aspects. CLINICAL CASES: Case 1. A 2 year-old girl with no family history of similar disorders started, at the age of 6 months, to have episodes of upward deviation of gaze with hyperextension of the neck and vertical nystagmus of fixation, increased by nervousness and episodes of fever. Some episodes caused the patient to fall in spite of there being no alteration of consciousness. Case 2. From the age of five months a 1 year-old girl with no significant personal or family history had episodes of ocular deviation upwards with forward inclination of the head to correct her gaze and slow motor development from the age of five months. RESULTS: Complementary studies were normal in both patients. As in the cases described in the literature, our cases had no family history and were not sensitive to Dopa. To date 11 children have been described in the literature and few familial cases seen with dominant autosomal inheritance. CONCLUSIONS: Our patients had a non-epileptic paroxystic phenomenon known as BPTUG syndrome. This condition starts during the first year of life, has a benign course and the episodes have ceased by the age of four years. We believe it is important to consider the differential diagnosis with epileptic phenomena, evaluate the response to L-Dopa and bear in mind that this syndrome may be the clinical expression of several different conditions. Although the course is usually benign, it may later be associated with other signs of neurological problems which should be taken into account.     

Long-term results of total penile reconstruction with a prefabricated lateral arm free flap

Trifluoroethanol (TFE) is often used to increase the helicity of peptides to make them usable as models of helices in proteins. We have measured helix propensities for all 20 amino acids in water and two concentrations of trifluoroethanol, 15 and 40% (v/v) using, as a model system, a peptide derived from the sequence of the alpha-helix of ribonuclease T1. There are three main conclusions from our studies. (1) TFE alters electrostatic interactions in the ribonuclease T1 helical peptide such that the dependence of the helical content on pH is lost in 40% TFE. (2) Helix propensities measured in 15% TFE correlate well with propensities measured in water, however, the correlation with propensities measured in 40% TFE is significantly worse. (3) Propensities measured in alanine-based peptides and the ribonuclease T1 peptide in TFE show very poor agreement, revealing that TFE greatly increases the effect of sequence context.     

Histomorphometric, physical, and mechanical effects of spaceflight and insulin-like growth factor-I on rat long bones

Pituitary adenylate cyclase activating peptide and vasoactive intestinal peptide belong to the same neuropeptide family. Both peptides are present in nerve fibers in the gastric wall and are thought to be involved in the regulation of inflammatory processes. Experimental ulcers were induced in the rat gastric mucosa by local application of acetic acid. During the healing process we examined the PACAP and VIP innervation by means of immunohistochemistry and in situ hybridization. The ulcer area was examined from day 1 to day 15 after ulcer induction. There was a marked depletion of PACAP in nerve fibers at the ulcer margin from day 1 and onwards. On day 10 and day 15, PACAP-immunoreactive nerve fibers could again be visualized at the ulcer margin. In contrast, VIP immunoreactive nerve fibers were present at the ulcer margin at all time points studied. From day 10 following ulcer induction PACAP- and VIP- immunoreactive nerve fibers were increased in frequency in the smooth muscle beneath the ulcer. An upregulation of VIP and PACAP mRNA was also demonstrated in the myenteric ganglia adjacent to ulcer. The present results indicate that neuronal PACAP and VIP react differently to the inflammation at the ulcer margin but similarly in the smooth muscle during the ulcer healing.     

Patient education for discharge after coronary bypass surgery in the 1990s: are patients adequately prepared?

Short hospitalizations for patients undergoing coronary artery bypass grafting (CABG) require continuous nursing evaluation of patients' discharge education. Six institutions collaborated in surveying 300 postoperative patients with CABG to identify learning priorities and patients' perceptions of the effectiveness of discharge education. Data analysis from the self-administered questionnaire demonstrated consistent patient priorities across institutions. Differences in teaching methods and content did not affect perceived preparedness or importance scores. Regional experience demonstrates that variable teaching efforts meet patients' priorities and provide high overall patient preparedness for discharge. Patients with the shortest hospitalizations had higher preparedness scores.     

An analysis of program and family costs of case managed care for technology-dependent infants with bronchopulmonary dysplasia

Influenza caused low levels of mortality and morbidity in England and Wales in the 1997/98 season. Influenza viruses of the H3N2 and H1N1 subtypes were isolated in small numbers from community and hospital patients in November and December. Their numbers subsequently increased to peak in February, H1N1 a week or two before H3N2. Most of the H1N1 isolates were similar to the A/Bayern/7/95-like virus, included in the vaccine recommended for 1997-98, whereas most H3N2 isolates were A/Sydney/5/97-like, a virus not covered by the season's vaccine but recommended for 1998/99. Consultation rates in general practice for influenza-like illnesses remained low until early February, when a small increase coincided with the increase in laboratory confirmed influenza. Fewer deaths directly attributed to influenza were registered during the surveillance period than in recent years. Localised outbreaks occurred in schools and nursing homes towards the end of the winter.     

A correlation between electrochemical behaviour, composition and semiconducting properties of naturally grown oxide films on copper

Copper(I) oxide films formed under open-circuit potential in neutral aqueous solutions have been characterized, using coulometry, photocurrent spectroscopy and X-ray photo-electron spectroscopy (XPS). The reduction potential of the oxide layer was found to depend on the presence in the electrolyte of chloride ions, Cu(II) or Cu(I) ionic species or of a corrosion inhibitor. XPS analyses were performed on these oxide layers, and showed in some cases an evolution of the oxidation state of copper from + 2 to + 1 state throughout the film. Different conducting properties of the cuprous oxides could be demonstrated through photo-electrochemical measurements, and the formation of a duplex Cu₂O layer with two semiconducting components of different stoichiometries was discussed.     

Chip geometries during high-speed machining for orthogonal cutting conditions

The originality of this work consists in taking photographs of chips during the cutting process for a large range of speeds. Contrary to methods usually used such as the quick stop in which root chips are analyzed after an abrupt interruption of the cutting, the proposed process photographs the chip geometry during its elaboration. An original device reproducing perfectly orthogonal cutting conditions is used because it allows a good accessibility to the zone of machining and reduces considerably the vibrations found in conventional machining tests. A large range of cutting velocities is investigated (from 17 to 60 m/s) for a middle hard steel (French Standards XC18). The experimental measures of the root chip geometry, more specifically the tool-chip contact length and the shear angle, are obtained from an analysis of the pictures obtained with a numerical high-speed camera. These geometrical characteristics of chips are studied for various cutting speeds, at the three rake angles −5, 0, +5° and for different depths of cut reaching 0.65 mm.     

Spatial summation in simple (Fourier) and complex (non-Fourier) texture channels

Complex (non-Fourier, second-order) channels have been proposed to explain aspects of texture-based region segregation and related perceptual tasks. Complex channels contain two stages of linear filtering with an intermediate pointwise nonlinearity. The intermediate nonlinearity is crucial. Without it, a complex channel is equivalent to a single linear filter (a simple channel). Here we asked whether the intermediate nonlinearity is piecewise-linear (an ordinary rectifier), or compressive, or expansive. We measured the perceptual segregation between element-arrangement textures where the contrast and area of the individual elements were systematically varied. For solid-square elements, the tradeoff between contrast and area was approximately linear, consistent with simple linear channels. For Gabor-patch elements, however, the tradeoff was highly nonlinear, consistent with complex channels in which the intermediate nonlinearity is expansive (with an exponent somewhat higher than 2). Also, substantial individual differences in certain details were explainable by differential intrusion from "off-frequency" complex channels. Lastly, the results reported here (in conjunction with those of other studies) suggest that the strongly compressive intensive nonlinearity previously known to act in texture segregation cannot be attributed to a compressive nonlinearity acting locally and relatively early (before the spatial-frequency and orientation-selective channels) but could result from inhibition among the channels (as in a normalization network).