Ca2+-triggered peptide secretion in single cells imaged with green fluorescent protein and evanescent-wave microscopy

Green fluorescent protein fused to human chromogranin B or neuropeptide Y was expressed in PC12 cells and caused bright, punctate fluorescence. The fluorescent points colocalized with the endogenous secretory granule marker dopamine beta-hydroxylase. Stimulation of live PC12 cells with elevated [K+], or of permeabilized PC12 cells with Ca2+, led to Ca2+-dependent loss of fluorescence from neurites. Ca2+ stimulated secretion of both fusion proteins equally well. In living cells, single fluorescent granules were imaged by evanescent-wave fluorescence microscopy. Granules were seen to migrate; to stop, as if trapped by plasmalemmal docking sites; and then to disappear abruptly, as if through exocytosis. Evidently, GFP fused to secreted peptides is a fluorescent marker for dense-core secretory granules and may be used for time-resolved microscopy of single granules.     

Cloning and expression of the cDNA for rat NAD+-dependent 15-hydroxyprostaglandin dehydrogenase

Following interleukin-1 (IL1) stimulation, an IL1 receptor associated kinase (IRAK) is rapidly recruited to the receptor complex. However, it is not understood if IRAK is able to interact directly with the intracellular portion of the IL1-RI or if its recruitment is mediated by a different molecule. Using the yeast two-hybrid system, we have analysed possible protein-protein interactions between IRAK, IL1-RI and IL1-RAcP. We found that IRAK is able to interact with the equivalent cytoplasmic region of the IL1-RAcP but is unable to interact with the cytoplasmic region of the IL1-RI. Immunoprecipitation of the IL1-RAcP followed by Western blot analysis using anti-IRAK antibodies revealed that IRAK co-precipitated with the IL1-RAcP. We propose that, in non-stimulated cells, IRAK is bound to the IL1-RAcP and therefore, following IL1 stimulation, both molecules are recruited simultaneously to the ILI-RI complex.     

Uncemented total hip replacements using the Mecron acetabular cup--a prospective study

Triatoma brasiliensis is one of the most important vectors of Chagas disease in the semiarid zone of the northeast of Brazil. Intraspecific morphological and behavioural variation has been reported for different populations. Results for four distinct populations using eight isoenzymes are reported here. The literature describes three subspecies: T. brasiliensis brasiliensis Neiva, 1911; T. brasiliensis melanica Neiva & Lent, 1941 and T. brasiliensis macromelasoma Galv?o, 1956. These subspecies differ mainly in their cuticle colour pattern and were regarded as synonyms by Lent and Wygodzinsky (1979). In order to evaluate whether the chromatic pattern is a morphological variation of different melanic forms within T. brasiliensis or due to interspecific variation, field collections were performed in localities where these three subspecies have been described: Caicó (Rio Grande do Norte), the type-locality for T. b. brasiliensis; Petrolina (Pernambuco) for T. b. macromelasoma and Espinosa (Minas Gerais) for T. b. melanica. A fourth distinct chromatic pattern was found in Juazeiro (Bahia). A total of nine loci were studied. Values of Nei's genetic distance (D) were calculated. T. b. brasiliensis and T. b. macromelasoma are the closest populations with a D = 0.295. T. b. melanica had a D > or = 0.537 when compared to the others, a distance in the range of interspecific variation for other triatomine species.     

Intestinal water and solute absorption studies: comparison of in situ perfusion with chronic isolated loops in rats

The effects of lumenal glucose on jejunal water transport and the influence of glucose-induced water absorption on solute uptake from single-pass perfusions are compared in anesthetized rats in situ and isolated chronic loops in unanesthetized rats in vivo. While the magnitudes of solute membrane permeabilities are consistently higher in the chronic loop system, the effects on water transport and its promotion of jejunal solute uptake are comparable between the two experimental systems. The effect of glucose-induced water absorption on the enhanced/baseline jejunal uptake ratio of the hydrophilic drug, acetaminophen, is greater than that for the lipophilic drug, phenytoin, in both experimental systems. The fact that chronic loop effective solute permeabilities were equivalent to solute membrane permeabilities in situ is consistent with greater lumenal fluid mixing in vivo. In addition, in situ body temperature affects the uptake of phenytoin but not acetaminophen, water, or glucose. This suggests that active and paracellular solute transport is not compromised in situ, while membrane partitioning and diffusion of lipophilic species are more sensitive to experimental conditions.     

1,3-Dimethylpyrimidoheterocycles as antibacterial agents

1,3-Dimethylbarbituric acid was used as a versatile precursor for the preparation of annelated pyranopyrimidines 2 and 3a-c besides pyrrolopyrimidine 12. The isolated pyrazolopyrimidines 8a-c and 10 and fused pyrazolopyrimidines 14a-d were also synthesized. In addition the new obtained pyrimidoheterocycles were subjected to bacterial testings.     

Transdermal nitroglycerin: clinical and pharmacokinetic consequences of renewing the patch and the application site

OBJECTIVE: We examined whether nitroglycerin (glyceryl trinitrate, GTN) patch treatment for 24 h could induce local cutaneous changes that impaired drug delivery and clinical efficacy. METHODS: Twenty angina patients were exercise-tested after 2 and 24 h of treatment and then 2 h after patch renewal. The patch was either renewed on a new skin location or on the previous application site in a randomised, double-blind, cross-over protocol. GTN plasma concentrations and finger plethysmography were obtained before and after each exercise test. RESULTS AND CONCLUSIONS: The clinical efficacy, the effect seen on plethysmography and the GTN plasma concentrations tended to increase after patch renewal, regardless of the application site of the renewed patch. Hence, cutaneous changes of clinical importance could not be demonstrated.     

Anandamide and other N-acylethanolamines in mouse peritoneal macrophages

Exposure to stressful events and elevated level of stress hormones are associated with impaired spatial memory and neuronal damage in the hippocampus. These neurons are considered to be maintained by neurotrophins such as nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) and trk family of neurotrophin receptors. Male Wistar rats (6 weeks old) were exposed to immobilization stress for 8 h and their brains were processed for in situ hybridization histochemistry. Exposure to long-lasting immobilization stress reduced mRNA levels for neurotrophins and their high affinity receptors in the brain, especially in the hippocampus. Our results provide, some new information that may be relevant to the pathogenesis of stress-induced disturbances of memory and learning.     

Variability in cytotoxicity and fluoride release of resin-modified glass-ionomer cements

New-generation glass-ionomer cements contain resin to improve their restorative properties. These resin-modified glass-ionomer cements vary considerably in their chemistry, which could result in corresponding variability in their physical and biological properties. This study investigated the cytotoxicity and the fluoride release of two resin-modified glass ionomers, a conventional glass-ionomer cement, and a resin composite. Samples were prepared and extracted in distilled water for 1, 4, and 7 days; eluates were filtered and tested by means of 3T3 mouse fibroblasts. Cytotoxicity (MTT assay) values were low for all materials and extraction times, indicating minimal cytotoxicity of all materials (less than 30% inhibition). Cytotoxicity of one resin-modified glass ionomer was significantly higher than for the other materials (p < 0.001). One resin-modified glass ionomer and the conventional glass-ionomer cement released significantly more fluoride at each time interval (p < 0.001) than the other resin-modified glass-ionomer cement and the resin composite. Fluoride release and cytotoxicity were correlated (r2 = 0.60; p < 0.001), although the fluoride release does not account for the cytotoxicity observed. Cytotoxicity and fluoride release suggest that one hybrid behaved more like a conventional glass ionomer, and the other like a resin composite. These differences may have implications for material selection in specific clinical situations.