Exploring soft matter: liposomal vesicles and their tethers

The malleability of soft matter such as vesicular systems, allows their mechanical manipulation into various interesting structures. Their inherent elastic membrane properties can be utilized in fabricating micro-scaled transport machineries. In this paper, we have described several features of research carried out in our laboratory. This includes the self-healing ability of multilamellar liposome on removal of their inner core. The formation of tethers from their outer bilayers into complex networks has been demonstrated. Photo enhancement of images has revealed the apparent existence of narrow channels inside such tethers. The tethers channels were subsequently used as transport conduits to carry solid polystyrene microspheres as well as large liposomes larger than the tether diameter under controllable speeds of up to 2 microm s(- 1). The possibility of producing multi-compartment liposomal systems connected to each other directly by fusion or indirectly via tether networks has provided us with a system capable of transporting components from one vesicle entity to another.     

The plant-type ferredoxin-NADP+ reductase/ferredoxin redox system as a possible drug target against apicomplexan human parasites

Apicomplexa are unicellular, obligate intracellular parasites of great medical importance. They include human pathogens like Plasmodium spp., the causative agent of malaria, and Toxoplasma gondii, an opportunistic parasite of immunosuppressed individuals and a common cause of congenital disease (toxoplasmosis). They alone affect several hundred million people worldwide so that new drugs, especially for plasmodial infections, are urgently needed. This review will focus on a recently emerged, potential drug target, a plant-type redox system consisting of ferredoxin-NADP(+) reductase (FNR) and its redox partner, ferredoxin (Fd). Both reside in an unique organelle of these parasites, named apicoplast, which is of algal origin. The apicoplast has been shown to be required for pathogen survival. In addition to other pathways already identified in this compartment, the FNR/Fd redox system represents a promising drug target because homologous proteins are not present in host organisms. Furthermore, a wealth of structural information exists on the closely related plant proteins, which can be exploited for structure-function studies of the apicomplexan protein pair. T. gondii and P. falciparum FNRs have been cloned, and the T. gondii enzyme was shown to be a flavoprotein active as a NADPH-dependent oxidoreductase. Both phylogenetic and biochemical analyses indicate that T. gondii FNR is similar in function to the isoform present in non-photosynthetic plastids whereby electron flow is from NADPH to oxidized Fd. The resulting reduced Fd is then presumably used as a reductant for various target enzymes whose nature is just starting to emerge. Among the likely candidates is the iron-sulfur cluster biosynthesis pathway, which is also located in the apicoplast and dependent on reducing power. Furthermore, lipoic acid synthase and enzymes of the isoprenoid biosynthetic pathway may be other conceivable targets. Since all these metabolic steps are vital for the parasite, blocking electron flow from FNR to Fd by inhibition of either FNR activity or its molecular interaction with Fd should also interfere with these pathways, ultimately killing the parasite. Although the three-dimensional structure of FNR from T. gondii is not yet known, experimental and computational evidence shows that apicomplexan and plant enzymes are very similar in structure. Furthermore, single amino acid changes can have profound effects on the enzyme activity and affinity for Fd. This knowledge may be exploited for the design of inhibitors of protein-protein interaction. On the other hand, specifically tailored NAD(P) analogues or mimetics based on previously described substances might be useful lead compounds for apicomplexan FNR inhibitors.     

Can an inert sleeping pill affect sleep? Effects on polysomnographic,behavioral and subjective measures

RATIONALE: Since two recent meta-analyses on sleep changes associated with placebo in clinical trials suggested a beneficial effect of placebo treatments, pointing to a dissociation between subjective and objective measures of sleep, the current experiment was directly aimed to assess the effects of an inert compound, administered with the suggestion that it was a hypnotic substance in subjects with mild sleep complaints. OBJECTIVES: The aim of this study was to compare subjective, behavioral, polysomnographic (PSG), and quantitative electroencephalographic (EEG) changes during a night preceded or not by the intake of two 50-mg lactose capsules. METHODS: Ten female students, selected by the Pittsburgh Sleep Quality Index, slept for three consecutive nights in a sleep laboratory, with the experimental (EXP) night defined by the administration of two 50-mg lactose pills. Self-ratings of sleep quality and performance were assessed upon morning awakening of baseline (BSL) and EXP nights. RESULTS: The EXP nights were self-rated as more restful and characterized by a decreased number of nocturnal awakenings than the BSL nights. PSG measures showed that wakefulness after sleep onset significantly decreased during the EXP night as compared to the BSL night. The EXP nights also showed an increase of 0.5-4.0 Hz power during nonrapid eye movement sleep and a decrease of EEG activity in the beta frequency range during rapid eye movement sleep only at central brain sites. A specific improvement of behavioral measures was also found upon morning awakening after the EXP night compared to the BSL night. CONCLUSIONS: The administration of an inert pill improves both the subjective and objective quality of sleep. The reduced sleep fragmentation and the effects on some quantitative EEG markers of sleep homeostasis suggest that the experimental manipulation induced coherent changes in the subsequent sleep, resembling an enhancement of sleep pressure. The regional differences of EEG activity suggest the involvement of a specific physiological mechanism distinct from that of effective treatments.     

Trace amines depress GABA B response in dopaminergic neurons by inhibiting G-betagamma-gated inwardly rectifying potassium channels

Trace amines (TAs) are present in the central nervous system in which they up-regulate catecholamine release and are implicated in the pathogenesis of addiction, attention-deficit/hyper-activity disorder, Parkinson's disease, and schizophrenia. By using intracellular and patch-clamp recordings from dopaminergic cells in the rat midbrain slices, we report a depressant postsynaptic action of two TAs, beta-phenylethylamine (beta-PEA) and tyramine (TYR) on the GABA(B)-mediated slow inhibitory postsynaptic potential and baclofen-activated outward currents. beta-PEA and TYR activated G-proteins, interfering with the coupling between GABA(B) receptors and G-betagamma-gated inwardly rectifying potassium channels. This is the first demonstration that beta-PEA and TYR depress inhibitory synaptic potentials in neurons of the central nervous system, supporting their emerging role as neuromodulators.     

9p21 locus analysis in high-risk gastrointestinal stromal tumors characterized for c-kit and platelet-derived growth factor receptor alpha gene alterations

BACKGROUND: Gastrointestinal stromal tumors (GISTs) are noncomplex sarcomas that often are due to c-kit-activating and platelet-derived growth factor receptor alpha gene (PDGFRalpha)-activating mutations and perturbations of their related signaling pathways. Molecular and cytogenetic findings have indicated correlations between tumor progression and high-risk GISTs with c-kit mutations, the overexpression of genes such as ezrin, and losses at 9p. In particular, it was reported recently that malignant GISTs showed alterations in the p16INK4a gene located at the 9p21 locus. METHODS: To assess the involvement of p14ARF and p15INK4b in addition to p16INK4a in GISTs, the authors undertook a molecular and cytogenetic study of the 9p21 locus. A series of 22 pre-Gleevec era, cryopreserved, high-risk GISTs that were characterized well in terms of KIT and PDGFRalpha receptors were investigated for mRNA expression, homozygous deletions, mutations, and promoter methylation of locus 9p21, in some instances complemented by fluorescent in situ hybridization studies. RESULTS: The results indicated the loss of p16INK4a mRNA expression in 41% of the GISTs, mainly due to the homozygous deletion of both the p16INK4a gene and the p14ARF gene (24%). No mutations were found, and promoter methylation (detected by means of methylation-specific polymerase chain reaction analysis in 27% of tumors) was restricted mainly to the p15INK4b gene (20%). It is noteworthy that, in all of the methylated GISTs, the epigenetic promoter alteration was coupled with mRNA expression. CONCLUSIONS: Alterations in the 9p21 locus were found cumulatively in 54% of the tumors in the current series and were represented mainly by the loss of tumor suppressor gene expression. The p16INK4a deletion, which always was coupled with p14ARF gene loss, seemed to be the most common 9p21 inactivation mechanism.     

Amplification of c-erbB2 oncogene: a major prognostic indicator in uterine serous papillary carcinoma

BACKGROUND: Overexpression of the epidermal growth factor type II receptor HER-2/neu has been associated with resistance to chemotherapy and poor survival in several human tumors. In the current study, the authors have determined the frequency and clinical significance of HER-2/neu gene amplification in uterine serous papillary endometrial carcinoma (USPC), a highly aggressive variant of endometrial carcinoma. METHODS: Fluorescence in situ hybridization (FISH) assay was used to analyze gene amplification in paraffin blocks from 30 women harboring Stage IA-IV USPC treated at the University of Arkansas for Medical Sciences (Little Rock, AR) from 1997 to 2004. Chromosome 17 polysomy status by FISH was also assessed in all specimens. USPC patient survival in relation to HER-2/neu gene amplification was analyzed using Kaplan-Meier curves in conjunction with the log-rank test. RESULTS: Amplification of the HER-2/neu gene by FISH was observed in 14 of the 30 (47%) cases. Heterogeneity was noted in 4 of 14 cases in the amplification of the HER-2/neu gene within the same tumor samples with pockets of amplified tumor cells amidst nonamplified tumor cells. Patients with USPC harboring tumors with HER-2/neu gene amplification had a significantly shorter survival time from diagnosis to disease-related death when compared with FISH-negative patients (P = 0.0008). African-American (AA) patients were found to have a poorer prognosis compared with Caucasian (C) women (P = 0.01) and to harbor USPC with significantly higher levels of HER-2/neu gene amplification (P = 0.02). CONCLUSIONS: HER-2/neu gene amplification in USPC was found to be an important prognostic indicator for poor outcome that occurs more frequently in AA when compared with C patients. Determination of HER-2/neu gene amplification may guide clinical management of patients with USPC and may have important implications for the implementation of novel treatment strategies.     

Iatrogenic splenic injuries

Inadvertent intraoperative injuries to the spleen by the surgical team represent an underestimated complication of many abdominal procedures. Surgical reports often lack the necessary details and frequently a clear justification as why a splenectomy was indicated is not provided. The wide variability of the incidence reported in literature makes it is difficult to evaluate the morbidity and mortality associated to these injuries and to assess the early and late consequences of this complication, although it is still possible to infer some of the reasons for these inconsistencies and to roughly estimate both clinical and socio-economical effects of this injury. Given the degree of uncertainty on the incidence of iatrogenic and traumatic splenic injuries and on the immediate and long-term sequelae suffered by asplenic patients, we thought that a multicentric prospective study was warranted. We are therefore announcing the start of a study involving several Institutions within the Regione Campania, aimed at obtaining an unbiased estimate of the incidence of these injuries, together with the extent and severity of their long-term complications. We also aim to help promoting a more effective prevention.     

Achilles tendon for sternal synthesis in the treatment of mediastinitis

Surgical approaches to postoperative mediastinitis that imply wire removal achieve earlier infection recovery but leave the patient with sternal instability. In 10 patients after wound surgical debridement, my colleagues and I achieved sternal synthesis by using Achilles tendons retrieved from multiorgan donors and stored in glutaraldehyde. Three tendons were used in each patient; they were passed through the bone at the manubrium and parasternally at the midsternum and the lower sternum. Thirty-day computed tomographic scan results, infection recovery, and quality of life were satisfactory.     

Randomized study on oral administration of calcitriol to prevent symptomatic hypocalcemia after total thyroidectomy

BACKGROUND: Symptomatic hypocalcemia remains the main postoperative complication after total thyroidectomy. The aim of the present study was to evaluate the role of oral supplementation of calcitriol and calcium salts in preventing severe postoperative hypocalcemia after total thyroidectomy. METHODS: A consecutive series of patients undergoing total thyroidectomy followed by administration of 500 mg of calcium salts 3 times per day were randomized to 3 different postoperative medical treatments: in group A, .5 microg of calcitriol twice per day was administered to 104 patients; in group B, 1 mmicrog of calcitriol twice per day was administered to 111 patients; and in group C, 202 patients did not receive calcitriol. RESULTS: The rate of postoperative tetany in group A was 2.9%, in group B was 0%, and in group C was 7.4% (P=.03) and the rate of paresthesias was 28.8%, 17.1%, and 22.3%, respectively (P=.19). At discontinuation of calcitriol/calcium salts treatment, intact parathyroid hormone levels did not significantly differ from the preoperative levels. Receiver operating characteristic (ROC) curve analysis showed that the area under the curve for serum concentration of calcium in predicting postoperative tetany was .749, .858 and .862 on the first, second, and third postoperative day, respectively. The best cut-off value of calcemia for prediction tetany was 7.5 mg/dL, and the rate of severe hypocalcemia on the third postoperative day was 23.1% in group A, 9.9% in group B, and 27.2% in group C (P=.001). CONCLUSIONS: Oral administration of 1 microg of calcitriol twice per day and 500 mg of calcium salts 3 times per day after total thyroidectomy significantly decreases the risk of severe postoperative hypocalcemia.