In vitro and in silico analysis of the vascular effects of asymmetrical <Emphasis Type="Italic">N</Emphasis>,<Emphasis Type="Italic">N</Emphasis>-bis(alkanol)amine aryl esters,novel multidrug resistance-reverting agents

Asymmetrical N,N-bis(alkanol)amine aryl esters (FRA77, GDE6, and GDE19) are potent multidrug resistance (MDR) reversers. Their structures loosely remind that of the Ca²⁺ antagonist verapamil. Therefore, the aim of this study was to investigate their vascular activity in vitro. Their effects on the mechanical activity of fresh and cultured rat aorta rings on Ca_v1.2 channel current (I _Ca1.2) of A7r5 cells and their cytotoxicity on A7r5 and EA.hy926 cells were analyzed. Docking at the rat α_1C subunit of the Ca_v1.2 channel was simulated in silico. Compounds tested were cytotoxic at concentrations >1 μM (FRA77, GDE6, GDE19) and >10 μM (verapamil) in EA.hy926 cells, or >10 μM (FRA77, GDE6, GDE19) and at 100 μM (verapamil) in A7r5 cells. In fresh rings, the three compounds partly antagonized phenylephrine and 60 mM K⁺ (K60)-induced contraction at concentrations ≥1 and ≥3 μM, respectively. On the contrary, verapamil fully relaxed rings pre-contracted with both agents. In cultured rings, 10 μM GDE6, GDE19, FRA77, and verapamil significantly reduced the contractile response to both phenylephrine and K60. Similarly to verapamil, the three compounds docked at the α_1C subunit, interacting with the same amino acids residues. FRA77, GDE6, and GDE19 inhibited I _Ca1.2 with IC₅₀ values 1 order of magnitude higher than that of verapamil. FRA77-, GDE6-, and GDE19-induced vascular effects occurred at concentrations that are at least 1 order of magnitude higher than those effectively reverting MDR. Though an unambiguous divergence between MDR reverting and vascular activity is of overwhelming importance, these findings consistently contribute to the design and synthesis of novel and potent chemosensitizers.     

Assessing early to late stage dementia: the TSI and BANS-S scales in the nursing-home

BACKGROUND: The traditional assessment tools for dementia, such as the MMSE, have only limited ability to follow subjects with severe dementia because they show a floor effect. Specific observational and performance-based instruments were recently developed. OBJECTIVES: To directly compare an observational scale to a performance-based instrument in moderate to severe dementia. METHODS: We compared a slightly modified version of the performance-based Test for Severe Impairment (mTSI) to the observer-based Bedford Alzheimer Nursing Severity Scale (BANS-S). Both scales were administered, together with the Mini-Mental State Examination (MMSE) and the Clinical Dementia Rating scale (CDR), to a nursing-home sample of 130 women suffering from different types of dementia (CDR range:1-4; MMSE range:0-18), defined according to DSM-IV criteria. Mean age was 86.9 +/- 7.3 years and mean education was 2.7 +/- 1.1 years. RESULTS: The BANS-S could be applied to all patients, the mTSI to 87 subjects (66.9%). Mean mTSI score decreased progressively from CDR stage 2 to CDR stage 4, whereas no difference was detectable between CDR stages 1 and 2. By contrast, the BANS-S was not significantly different for CDR stages 1 to 3, and the mean BANS-S score worsened only in CDR stage 4. Results were similar for AD and non AD dementia. Both scales were independent from age and education and their test-retest and inter-rater reliabilities were satisfactory. CONCLUSION: The mTSI looks promising in the moderate-to-severe range, whereas the BANS-S seems more useful in the very late stage of dementia. However, neither scale was optimal and additional instruments should be tested in future studies.     

Methadone and nicotine self-administration in humans: a behavioral economic analysis

Rationale Prior research has revealed inconsistencies in the behavioral relations between nicotine and opiates among methadone-maintained patients.Objectives The current study examined whether the drug reinforcers cigarette puffs and methadone were economic complements or substitutes.Methods Five methadone-maintained, nicotine-dependent participants were trained to self-administer methadone, cigarette puffs, or concurrently available methadone and puffs. Following training, the fixed ratio (FR) value (price) was increased across sessions (FR 32, 64, 128, 256, and 512), first for methadone and then for puffs. Subsequently, methadone and puffs were concurrently available, and the price of each drug was increased independently, while the price of the alternative (puffs or methadone) remained constant at FR 32.Results Demand for methadone and cigarette puffs decreased as a function of increases in methadone and cigarette puff prices, respectively. When methadone and puffs were concurrently available, an increase in methadones price decreased puff consumption, and demand for methadone was less elastic than when puffs were not concurrently available. An increase in puff price decreased puff and methadone demand, but the elasticity of puff demand was unaffected. The concurrent presence of methadone had no effect on the elasticity of demand for cigarette puffs.Conclusions Methadone and cigarette puffs appear to be asymmetric economic complements.     

Back to basics: managing gunshot injuries in East Timor

BACKGROUND: The management of patients following severe trauma requires a combination of skills. Among these are accurate clinical assessment, basic resuscitation measures, basic general surgical skills and techniques, good decision making and team leadership. More technically advanced imaging and investigations are often useful and can enhance clinical assessment but should not replace the basic elements. In situations where more advanced investigations are not available good outcomes can be achieved by application of the basic principles in trauma management and surgery. This article describes a mass casualty situation of gunshot wounds managed with limited resources and the outcomes. METHODS: A series of gunshot wounds managed in a single day at Dili National Hospital, Dili, East Timor is described. The presentation of the patients, initial assessment of injuries and haemodynamic stability are detailed in addition to the subsequent surgical treatment, complications and outcomes. The information was obtained from hospital records and personal recollection of the treating doctors. All injuries were documented with digital photography. All patients were managed by two surgeons who also supervised their postoperative course and reported on their outcomes. Surgical procedures on all of the patients were performed by the named two consultant surgeons and one basic surgical trainee. RESULTS: Fourteen patients were treated for gunshot injuries. All were single bullet injuries. Six trajectories involved the thoracic or abdominal cavity (one trajectory traversed both). Eight injuries were to limbs or soft tissue of the pelvis and included four compound fractures. Surgical intervention was needed for all patients ranging from emergency laparotomy for haemodynamic instability through to wound debridements. Thirteen patients survived and were discharged from hospital within 3 weeks. One patient died of uncontrolled haemorrhage. CONCLUSIONS: In the present series of gunshot injuries managed with limited resources a good outcome was achieved with the application of basic principles. The present series emphasizes the importance of basic trauma management as outlined in courses such as Emergency Management of Severe Trauma and the use of basic surgical techniques as taught during the Advanced Surgical training program.     

Preferential increase of extracellular dopamine in the rat nucleus accumbens shell as compared to that in the core during acquisition and maintenance of intravenous nicotine self-administration

Rationale It has been reported that passive administration of nicotine increases preferentially extracellular dopamine (DA) release in the shell as compared to that in the core of the nucleus accumbens (NAc). To date, no information is available if this also applies to active, response-contingent nicotine administration. Objective This study was aimed to monitor the changes of extracellular DA in the NAc shell and core during active intravenous nicotine self-administration (SA). Methods Rats were bilaterally implanted with chronic cannulae and were trained to self-administer nicotine (0.03 mg/kg, i.v.) in single daily 1-h session for 6 weeks, with an initial fixed ratio (FR) 1 schedule increased to FR 2. Dialysate DA from the NAc shell and core was monitored before and for 90 min after the start of SA. Results Significant increases of active nose-pokes over inactive ones were found starting from the 16th SA session. No differences were found in basal extracellular DA in the NAc subdivisions. Data analysis showed (1) significant increases over basal of dialysate DA in the NAc subdivisions during nicotine SA, starting from the first week in the shell and from the second week in the core, (2) preferential increase of extracellular DA during nicotine SA in the shell (24–43%) compared to that in the core (10–23%) and (3) no change in dialysate DA in NAc subdivisions during extinction. Conclusions Response-contingent nicotine SA preferentially increases the DA output in the NAc shell as compared to that in the core, independently from the duration of the nicotine exposure. Increase in NAc DA is strictly related to nicotine action since is not observed during extinction in spite of active responding.     

Astroglial in vivo response to cocaine in mouse dentate gyrus: a quantitative and qualitative analysis by confocal microscopy

Astrocytes have been proved to play a critical role in neuromodulation, neuroprotection, pH maintenance, axon guidance control during development, homeostasis preservation and blood brain barrier maintenance in the CNS (Kimmelberg and Norenberg, 1989). Quantitative changes in the expression of glial fibrillary acidic protein (GFAP), a cytoskeletal intermediate filament protein exclusively expressed in astrocytes (Bignami et al, 1972), have been observed after administration of alcohol (Framke, 1995), morphine (Beitner-Johnson et al., 1993), amphetamine and its derivates (Aguirre et al., 1999), cannabinoids (Suarez et al., 2000), nicotine (Janson and Moller, 1993), caffeine (Marret et al., 1993) and prenatal exposure to cocaine (Clarke et al., 1996; Nassogne et al., 1998). However, the general astrocytic response to drugs of abuse is still far from being defined. In the present study we examined the in vivo astroglial response to cocaine in mouse dentate gyrus, the hippocampus being a common target of neurotoxic agents (Walsh and Emerich, 1988) which has a prominent effect on learning and memory processes (Eichenbaum et al., 1992). Quantitative changes in immunoreactivity of GFAP were investigated 24 h after acute and repeated daily administration of intraperitoneal cocaine (20 mg/kg). Drug-induced morphological alterations and spatial distribution of astrocytes were evaluated by means of confocal microscope. The results show that, compared to control animals, GFAP expression is two-fold enhanced after a single cocaine injection, still significantly higher after seven consecutive daily administrations, but not statistically different after prolonged (14 days) drug treatment. Moreover, morphological and morphometric analyses reveal significant modifications in astrocytic numbers, cell size and shape complexity. These data demonstrate that in mouse dentate gyrus, cocaine exposure differently affects the expression of GFAP and induces strong changes in astrocytes proliferation rate and cell morphology. Taken together, our findings provide the first in vivo quantitative and qualitative evaluation of astrocytic response to several regimens of cocaine in adult animals brain.     

Human methadone self-administration: effects of dose and ratio requirement

The effects of response requirement and small doses of methadone on human oral self-administration of methadone were examined. Three methadone maintenance patients stabilized at a dose of 80mg methadone per day were recruited as subjects. Completing a response requirement, fixed ratio (FR) of 32, 64 or 128 responses (FR32, FR64, FR128) on one button dispensed 10ml of drug solution. Completing the equivalent response requirement on a second concurrently available response button dispensed 10ml of vehicle. The opportunity to respond was unavailable until the drug or vehicle had been consumed. Each 10ml of drug solution contained methadone doses of 0.027, 0.054 or 0.108mg/ml. The frequency of deliveries was limited so that subjects could not ingest more than 54mg of methadone; the difference between the 80mg daily methadone dose and the methadone consumed in session was administered 30min post-session. At FR64 and FR128 the frequency of deliveries decreased, at the 0.054 and 0.027mg/ml doses, relative to the frequency of deliveries at FR32. The amount of methadone consumed increased with increases in methadone dose and decreased with increases in FR size. These results demonstrate the reinforcing effects of small unit doses of methadone. This procedure provides a sensitive baseline for examining effects of other pharmacological interventions on methadone ingestion in humans.     

Synthesis of sericin-based conjugates by click chemistry: enhancement of sunitinib bioavailability and cell membrane permeation

Sericin is a natural protein that has been used in biomedical and pharmaceutical fields as raw material for polypeptide-based drug delivery systems (DDSs). In this paper, it has been employed as pharmaceutical biopolymer for the production of sunitinib–polypeptide conjugate. The synthesis has been carried out by simple click reaction in water, using the redox couple l-ascorbic acid/hydrogen peroxide as a free radical grafting initiator. The bioconjugate molecular weight (50?kDa?In vitro studies demonstrated that bioavailability has been increased to almost 74%, compared with commercial formulation. Also cell membrane permeability has been augmented in in vitro tests, in which membrane models have been used to determine the lipid membrane/physiological fluid partition coefficient (Kp). The log(Kp) value of the bioconjugate was increased to over 4. This effect resulted in a three-fold decrease of IC₅₀ value against MCF-7 cells.     

Endotoxin tolerance and cross-tolerance in mast cells involves TLR4, TLR2 and FcεR1 interactions and SOCS expression: perspectives on immunomodulation in infectious and allergic diseases

Background  

The study of the endotoxin tolerance phenomenon in light of the recently defined roles of mast cells and toll-like receptors as essential components of the innate immune response and as orchestrators of acquired immunity may reveal potentially useful mechanisms of immunomodulation of infectious and allergic inflammatory responses, such as sepsis or asthma. Here we evaluated the phenomenon of direct tolerance of endotoxins, as well as the induction of cross-tolerance and synergism by stimulation with toll-like receptor-2 (TLR2) and FcεR1 agonists, in murine mast cells prestimulated with lipopolysaccharide (LPS). Additionally, we evaluated some stimulatory and inhibitory signaling molecules potentially involved in these phenomena.