A forensic case of erosive cystitis and septic pulmonary embolism: is there a correlation?

Septic pulmonary embolism (SPE) is an uncommon disease in which a microorganism- containing thrombus causes an inflammatory reaction and a mechanical obstruction in the vascular system of the lungs. Usually it is associated with tricuspid valve vegetation, septic thrombophlebitis or infected venous catheters. We present a rare and fatal case of massive septic pulmonary thromboembolism occurred in a diabetic woman, who complained in previous days dyspnoea, abdominal pain and nausea. Autopsy findings showed evidence of an erosive cystitis, while lungs examination showed total occlusion of pulmonary circulation system due to thrombo-embolic clots. In literature cases report about an association between septic pulmonary thromboembolism and erosive cystitis haven't been reported. So, we here show the first reported fatal case of a septic pulmonary thromboembolism as a lethal evolution of an erosive cystitis.

Arabidopsis thaliana expresses multiple Golgi-localised nucleotide-sugar transporters related to GONST1.

Transport of nucleotide-sugars across the Golgi membrane is required for the lumenal synthesis of a variety of essential cell surface components, and is mediated by nucleotide sugar transporters (NSTs) which are members of the large drug/metabolite superfamily of transporters. Despite the importance of these proteins in plants, so far only two have been described, GONST1 and AtUTr1 from Arabidopsis thaliana. In this work, our aim was to identify further Golgi nucleotide-sugar transporters from Arabidopsis. On the basis of their sequence similarity to GONST1, we found four additional proteins, which we named GONST2, 3, 4 and 5. These putative NSTs were grouped into three clades: GONST2 with GONST1; GONST3 with GONST4; and GONST5 with six further uncharacterized proteins. Transient expression in tobacco cells of a member of each clade, fused to the Green Fluorescent Protein (GFP), suggested that all these putative NSTs are localised in the Golgi. To obtain evidence for nucleotide sugar transport activity, we expressed these proteins, together with the previously characterised GONST1, in a GDP-mannose transport-defective yeast mutant (vrg4-2). We tested the transformants for rescue of two phenotypes associated with this mutation: sensitivity to hygromycin B and reduced glycosylation of extracellular chitinase. GONST1 and GONST2 complemented both phenotypes, indicating that GONST2, like the previously characterized GONST1, is a GDP-mannose transporter. GONST3, 4 and 5 also rescued the antibiotic sensitivity, but not the chitinase glycosylation defect, suggesting that they can also transport GDP-mannose across the yeast Golgi membrane but with a lower efficiency. RT-PCR and analysis of Affymetrix data revealed partially overlapping patterns of expression of GONST1-5 in a variety of organs. Because of the differences in ability to rescue the vrg4 - 2 phenotype, and the different expression patterns in plant organs, we speculate that GONST1 and GONST2 are both GDP-mannose transporters, whereas GONST3, GONST4 and GONST5 may transport other nucleotide-sugars in planta.

Vesicular stomatitis virus G pseudotyped retrovector mediates effective in vivo suicide gene delivery in experimental brain cancer.

Direct in vivo tumor-targeting with "suicide" viral vectors is limited by either inefficient gene transfer (i.e., retroviral vectors) or indiscriminate transfer of a conditionally toxic gene to surrounding nonmalignant tissue (i.e., adenoviral vectors). Retrovectors pseudotyped with the vesicular stomatitis virus G protein (VSVG) may serve as a remedy to this conundrum. These retroviral particles differ from standard murine retroviruses by their very broad tropism and the capacity to be concentrated by ultracentrifugation without loss of activity. We propose that a VSVG-typed retrovector can be used for efficient and tumor-specific herpes simplex virus thymidine kinase (TK) gene delivery in vivo. To test this hypothesis, we developed a bicistronic retroviral vector that expresses TK and green fluorescence protein (pTKiGFP). The 293GPG packaging cell line was used to generate vTKiGFP retroparticles. In cytotoxicity assays, vTKiGFP-transduced human glioma cell lines were sensitized to the cytotoxic effects of gangciclovir (GCV) 10,000-fold. Subsequently, virus was concentrated by ultracentrifugation to a titer of 2.3 x 10(10) cfu/ml. We tested the antitumor activity of vTKiGFP retroparticles in a rat C6 glioma model of brain cancer. Concentrated retrovector stock (9 microl volume) was injected stereotactically in preestablished intracerebral tumor. Subsequently, rats were treated with GCV for 10 days. Control rats (no GCV) had a mean survival of 38 days (range, 20-52 days). Sections performed on postmortem brain tissue revealed large tumors with evidence of high efficiency retrovector transfer and expression (as assessed by GFP fluorescence). Fluorescence was restricted to malignant tissue. In the experimental group (GCV treated), 8 of 12 remain alive and well >120 days after glioma implantation. In conclusion, vTKiGFP is very efficient at transducing human glioma cell lines in vitro and leads to significant GCV sensitization. Recombinant retroviral particles can be concentrated to titers that allow in vivo intratumoral delivery of large viral doses. The therapeutic efficiency of this reagent has been demonstrated in a preclinical model of brain cancer.

Mobilization of arachidonic acid in thrombin-stimulated human platelets.

In the present work we investigated the effect of serine esterase inhibitors such as 2-nitro-4-carboxyphenyl N,N-diphenylcarbamate (NCDC) and phenylmethylsulfonyl fluoride (PMSF), as well as the effect of mepacrine on thrombin-induced mobilization of arachidonic acid (AA) in human platelets. The inhibitor NCDC (0.6 mM) completely abolished the thrombin-induced activation of phospholipase C, phospholipase A2, and transacylase enzymes, whereas the pretreatment of platelets with PMSF (2 mM) resulted in a highly selective inhibition of phospholipase A2 and transacylase activities, with no marked effect on thrombin-induced activation of phospholipase C. The thrombin-induced release of [3H]AA from phosphatidylcholine and phosphatidylinositol was reduced by 90 and 56%, respectively, in the presence of PMSF. This inhibitor also caused a parallel inhibition in the accumulation of [3H]AA (85%) with little effect on thrombin-induced formation of [3H]phosphatidic acid (5%), whereas mepacrine (0.4 mM) caused a selective inhibition of phospholipase A2 and transacylase activities with concomitant stimulation of [3H]phosphatidic acid formation in intact human platelets. These results demonstrate that NCDC and PMSF (serine esterase inhibitors) do not affect agonist-induced activation of phospholipases that mobilize arachidonic acid through a common site. Our results further demonstrate that the inhibition of [3H]AA release observed in the presence of NCDC, PMSF, and mepacrine is primarily due to their direct effects on enzyme activities, rather than due to their indirect effects through formation of complexes between inhibitors and membrane phospholipids. Based upon these results, we also conclude that the combined hydrolysis of phosphatidylcholine and phosphatidylinositol by phospholipase A2 serves as a major source for eicosanoid biosynthesis in thrombin-stimulated human platelets.

The inhibition of arachidonic acid mobilization in human platelets by R59 022, a diacylglycerol kinase inhibitor.

R59 022 (6-[2-[4-[(4-fluorophenyl)phenylmethylene]-1- piperidinyl]ethyl]-7-methyl-5H-thiazolo[3,2-a]pyrimidin-5-one) has been suggested as an inhibitor of diacylglycerol kinase in erythrocyte membranes and intact platelets. In the present study, we have investigated the effects of this drug on arachidonic acid mobilization occurring in response to thrombin in intact human platelets. Our results indicate that release of arachidonic acid from membrane phospholipids such as phosphatidylcholine and phosphatidylinositol was severely impaired by R59 022 and the extent of inhibition amounted to 77% and 84%, respectively, as compared to controls. This resulted in a dramatic decrease in the accumulation of free arachidonic acid (labeled/unlabeled) and the percent inhibition of free arachidonic acid accumulation amounted to 80-90% as compared to controls. Furthermore, the drug caused a significant accumulation of thrombin-induced diacylglycerol (labeled) without affecting the formation of labeled phosphatidic acid (PA). We found no significant changes in the radioactivity of either phosphatidylethanolamine or phosphatidylserine following stimulation with thrombin in the presence or absence of R59 022. We conclude that the observed inhibition of thrombin-induced arachidonic acid mobilization by R59 022 may be due to its effects on the activities of diacylglycerol lipase/phospholipase A2. In addition, the failure of further stimulation of thrombin-induced PA by R59 022 may indicate that PA-specific phospholipase A2 is either not involved in the release of arachidonic acid or is not a major source for arachidonic acid release in thrombin-stimulated human platelets. These findings may prove to be important when this drug is used as a selective inhibitor of diacylglycerol kinase.