Anti-infective therapy of peri-implantitis with adjunctive local drug delivery or photodynamic therapy: six-month outcomes of a prospective randomized clinical trial

To compare the adjunctive clinical effects in the non-surgical treatment of peri-implantitis with either local drug delivery (LDD) or photodynamic therapy (PDT).

In vitro effects of arylimidamides against Besnoitia besnoiti infection in Vero cells

The in vitro effects of 4 arylimidamides (DB811, DB786, DB750 and DB766) against the proliferative tachyzoite stage of the apicomplexan parasite Besnoitia besnoiti were investigated. These four compounds had been shown earlier to exhibit in vitro activities in the nanomolar range against the related apicomplexans Neospora caninum and Toxoplasma gondii. Real-time-PCR was used to assess B. besnoiti intracellular proliferation in vitro. Preliminary assessment by light microscopy identified DB811 and DB750 as the most promising compounds, while DB786 and DB766 were much less effective. Three-day-growth assays and quantitative real-time PCR was used for IC50 determination of DB811 (0.079 muM) and DB750 (0.56 muM). Complete growth inhibition was observed at 1.6 muM for DB 811 and 1.7 muM for DB750. However, when infected cultures were treated for 14 days, proliferation of parasites occurred again in cultures treated with DB750 from day 4 onwards, while the proliferation of DB811-treated tachyzoites remained inhibited. Electron microscopy of B. besnoiti-infected fibroblast cultures fixed and processed at different time-points following the initiation of drug treatments revealed that DB811 exerted a much higher degree of ultrastructural alterations compared to DB750. These results show that arylimidamides such as DB811 could potentially become an important addition to the anti-parasitic arsenal for food animal production, especially in cattle.

Comparison of the effects of racemic ketamine and S-ketamine for anesthesia in Rheem gazelles (Gazella subgutturosa marica) and Subgutturosa gazelles (Gazella subgutturosa subgutturosa)

OBJECTIVE: To evaluate effects of racemic ketamine and S-ketamine in gazelles. ANIMALS: 21 male gazelles (10 Rheem gazelles [Gazella subgutturosa marica] and 11 Subgutturosa gazelles [Gazella subgutturosa subgutturosa]), 6 to 67 months old and weighing (mean+/-SD) 19 +/- 3 kg. PROCEDURES: In a randomized, blinded crossover study, a combination of medetomidine (80 mug/kg) with racemic ketamine (5 mg/kg) or S-ketamine (3 mg/kg) was administered i.m.. Heart rate, blood pressure, respiratory rate, rectal temperature, and oxygen saturation (determined by means of pulse oximetry) were measured. An evaluator timed and scored induction of, maintenance of, and recovery from anesthesia. Medetomidine was reversed with atipamezole. The alternate combination was used after a 4-day interval. Comparisons between groups were performed with Wilcoxon signed rank and paired t tests. RESULTS: Anesthesia induction was poor in 2 gazelles receiving S-ketamine, but other phases of anesthesia were uneventful. A dominant male required an additional dose of S-ketamine (0.75 mg/kg, i.m.). After administration of atipamezole, gazelles were uncoordinated for a significantly shorter period with S-ketamine than with racemic ketamine. Recovery quality was poor in 3 gazelles with racemic ketamine. No significant differences between treatments were found for any other variables. Time from drug administration to antagonism was similar between racemic ketamine (44.5 to 53.0 minutes) and S-ketamine (44.0 to 50.0 minutes). CONCLUSIONS AND CLINICAL RELEVANCE: Administration of S-ketamine at a dose 60% that of racemic ketamine resulted in poorer induction of anesthesia, an analogous degree of sedation, and better recovery from anesthesia in gazelles with unremarkable alterations in physiologic variables, compared with racemic ketamine.

New drug targets in atopic dermatitis

Atopic dermatitis (AD) is a chronic inflammatory skin disorder characterized by eczematous skin lesions, pruritus and typical histopathological features. T cells are thought to play a key role, but B cells might also participate in the pathogenesis of AD. In two investigator-initiated pilot studies, we studied the effects of B cell depletion by monoclonal anti-CD20 antibody therapy or a reduction of activated T cells by LFA3-IgG fusion protein on moderate-to-severe AD. All patients treated with either rituximab or alefacept showed an improvement of their skin symptoms with a sustained effect after treatment. In both studies, histological alterations, such as spongiosis, acanthosis and dermal infiltrate, including T and B cell numbers, dramatically improved and the expression of IL-5 and IL-13 was reduced after therapy. Upon rituximab therapy, allergen-specific IgE levels were not altered and total serum IgE levels only slightly decreased. According to recent studies, neutralizing B and T cells products such as IgE or IL-5 might be effective in subgroups of patients with AD.

Phosphocreatine interacts with phospholipids, affects membrane properties and exerts membrane-protective effects

A broad spectrum of beneficial effects has been ascribed to creatine (Cr), phosphocreatine (PCr) and their cyclic analogues cyclo-(cCr) and phospho-cyclocreatine (PcCr). Cr is widely used as nutritional supplement in sports and increasingly also as adjuvant treatment for pathologies such as myopathies and a plethora of neurodegenerative diseases. Additionally, Cr and its cyclic analogues have been proposed for anti-cancer treatment. The mechanisms involved in these pleiotropic effects are still controversial and far from being understood. The reversible conversion of Cr and ATP into PCr and ADP by creatine kinase, generating highly diffusible PCr energy reserves, is certainly an important element. However, some protective effects of Cr and analogues cannot be satisfactorily explained solely by effects on the cellular energy state. Here we used mainly liposome model systems to provide evidence for interaction of PCr and PcCr with different zwitterionic phospholipids by applying four independent, complementary biochemical and biophysical assays: (i) chemical binding assay, (ii) surface plasmon resonance spectroscopy (SPR), (iii) solid-state (31)P-NMR, and (iv) differential scanning calorimetry (DSC). SPR revealed low affinity PCr/phospholipid interaction that additionally induced changes in liposome shape as indicated by NMR and SPR. Additionally, DSC revealed evidence for membrane packing effects by PCr, as seen by altered lipid phase transition. Finally, PCr efficiently protected against membrane permeabilization in two different model systems: liposome-permeabilization by the membrane-active peptide melittin, and erythrocyte hemolysis by the oxidative drug doxorubicin, hypoosmotic stress or the mild detergent saponin. These findings suggest a new molecular basis for non-energy related functions of PCr and its cyclic analogue. PCr/phospholipid interaction and alteration of membrane structure may not only protect cellular membranes against various insults, but could have more general implications for many physiological membrane-related functions that are relevant for health and disease.

Effects of doxorubicin cancer therapy on autophagy and the ubiquitin-proteasome system in long-term cultured adult rat cardiomyocytes

The clinical use of anthracyclines in cancer therapy is limited by dose-dependent cardiotoxicity that involves cardiomyocyte injury and death. We have tested the hypothesis that anthracyclines affect protein degradation pathways in adult cardiomyocytes. To this aim, we assessed the effects of doxorubicin (Doxo) on apoptosis, autophagy and the proteasome/ubiquitin system in long-term cultured adult rat cardiomyocytes. Accumulation of poly-ubiquitinated proteins, increase of cathepsin-D-positive lysosomes and myofibrillar degradation were observed in Doxo-treated cardiomyocytes. Chymotrypsin-like activity of the proteasome was initially increased and then inhibited by Doxo over a time-course of 48 h. Proteasome 20S proteins were down-regulated by higher doses of Doxo. The expression of MURF-1, an ubiquitin-ligase specifically targeting myofibrillar proteins, was suppressed by Doxo at all concentrations measured. Microtubule-associated protein 1 light chain 3B (LC3)-positive punctae and both LC3-I and -II proteins were induced by Doxo in a dose-dependent manner, as confirmed by using lentiviral expression of green fluorescence protein bound to LC3 and live imaging. The lysosomotropic drug chloroquine led to autophagosome accumulation, which increased with concomitant Doxo treatment indicating enhanced autophagic flux. We conclude that Doxo causes a downregulation of the protein degradation machinery of cardiomyocytes with a resulting accumulation of poly-ubiquitinated proteins and autophagosomes. Although autophagy is initially stimulated as a compensatory response to cytotoxic stress, it is followed by apoptosis and necrosis at higher doses and longer exposure times. This mechanism might contribute to the late cardiotoxicity of anthracyclines by accelerated aging of the postmitotic adult cardiomyocytes and to the susceptibility of the aging heart to anthracycline cancer therapy.

Scale Up of PLA Nanoparticle Synthesis by Solvent Displacement and Functionalization of the Particles with a Silica Shell for Targeted Drug Delivery Applications

Biodegradable polymer nanoparticles have the properties necessary to address many of the issues associated with current drug delivery techniques including targeted and controlled delivery. A novel drug delivery vehicle is proposed consisting of a poly(lactic acid) nanoparticle core, with a functionalized, mesoporous silica shell. In this study, the production of PLA nanoparticles is investigated using solvent displacement in both a batch and continuous manner, and the effects of various system parameters are examined. Using Pluronic F-127 as the stabilization agent throughout the study, PLA nanoparticles are produced through solvent displacement with diameters ranging from 200 to 250 nm using two different methods: dropwise addition and in an impinging jet mixer. The impinging jet mixer allows for easy scale-up of particle production. The concentration of surfactant and volume of quench solution is found to have minimal impact on particle diameter; however, the concentration of PLA is found to significantly impact the diameter mean and polydispersity. In addition, the stability of the PLA nanoparticles is observed to increase as residual THF is evaporated. Lastly, the isolated PLA nanoparticles are coated with a silica shell using the Stöber Process. It is found that functionalizing the silica with a phosphonic silane in the presence of excess Pluronic F-127 decreases coalescence of the particles during the coating process. Future work should be conducted to fine-tune the PLA nanoparticle synthesis process by understanding the effect of other system parameters and in synthesizing mesoporous silica shells.

Drug interaction potential of resveratrol

Resveratrol is a naturally occurring polyphenol that is often used as a food supplement. Many positive health effects, including cardio protection, tumor suppression, and immune modulation, are associated with the intake of resveratrol. Resveratrol is well tolerated in healthy subjects without any comedication. However, supplemental doses of resveratrol in the range of 1 g/day or above by far exceed the natural intake through food. Whether resveratrol-drug interactions can be harmful in patients taking additional medications remains unknown. Recent in vivo studies and clinical trials indicate a possible drug-drug interaction potential using high-dosage formulations. In this review, the known in vitro and in vivo effects of resveratrol on various cytochrome P450 (CYP) isoenzymes are summarized. They are discussed in relation to clinically relevant plasma concentrations in humans. We conclude that resveratrol may lead to interactions with various CYPs, especially when taken in high doses. Aside from systemic CYP inhibition, intestinal interactions must also be considered. They can potentially lead to reduced first-pass metabolism, resulting in higher systemic exposure to certain coadministrated CYP substrates. Therefore, patients who ingest high doses of this food supplement combined with additional medications may be at risk of experiencing clinically relevant drug-drug interactions.

A novel steroid-like compound F90927 exerting positive-inotropic effects in cardiac muscle

Here we report a novel steroid-like compound F90363, exhibiting positive inotropy in vivo and in vitro in various cardiac muscle preparations. F90363 is a racemic mixture composed of the stereoisomers (-)-F90926 and (+)-F90927. Only F90927 exerted positive inotropy, while F90926 induced a weak negative inotropy, but only at concentrations 10(3) times higher than F90927 and most likely resulting from an unspecific interaction. The rapid time course of the action of F90927 suggested a direct interaction with a cellular target rather than a genomic alteration. We could identify the L-type Ca2+ current I(Ca(L)) as a main target of F90927, while excluding other components of cardiac Ca2+ signalling as potential contributors. In addition, several other signaling pathways known to lead to positive inotropy (e.g. alpha- and beta-adrenergic stimulation, cAMP pathways) could be excluded as targets of F90927. However, vessel contraction and stiffening of the cardiac muscle at high doses (>30 microM, 0.36 mg kg(-1), respectively) prevent the use of F90927 as a candidate for drug development. Since the compound may still find valuable applications in research, the aim of the present study was to identify the cellular target and the mechanism of inotropy of F90927.

Orosomucoid (alpha1-acid glycoprotein) plasma concentration and genetic variants: effects on human immunodeficiency virus protease inhibitor clearance and cellular accumulation

BACKGROUND AND OBJECTIVE: Protease inhibitors are highly bound to orosomucoid (ORM) (alpha1-acid glycoprotein), an acute-phase plasma protein encoded by 2 polymorphic genes, which may modulate their disposition. Our objective was to determine the influence of ORM concentration and phenotype on indinavir, lopinavir, and nelfinavir apparent clearance (CL(app)) and cellular accumulation. Efavirenz, mainly bound to albumin, was included as a control drug. METHODS: Plasma and cells samples were collected from 434 human immunodeficiency virus-infected patients. Total plasma and cellular drug concentrations and ORM concentrations and phenotypes were determined. RESULTS: Indinavir CL(app) was strongly influenced by ORM concentration (n = 36) (r2 = 0.47 [P = .00004]), particularly in the presence of ritonavir (r2 = 0.54 [P = .004]). Lopinavir CL(app) was weakly influenced by ORM concentration (n = 81) (r2 = 0.18 [P = .0001]). For both drugs, the ORM1 S variant concentration mainly explained this influence (r2 = 0.55 [P = .00004] and r2 = 0.23 [P = .0002], respectively). Indinavir CL(app) was significantly higher in F1F1 individuals than in F1S and SS patients (41.3, 23.4, and 10.3 L/h [P = .0004] without ritonavir and 21.1, 13.2, and 10.1 L/h [P = .05] with ritonavir, respectively). Lopinavir cellular exposure was not influenced by ORM abundance and phenotype. Finally, ORM concentration or phenotype did not influence nelfinavir (n = 153) or efavirenz (n = 198) pharmacokinetics. CONCLUSION: ORM concentration and phenotype modulate indinavir pharmacokinetics and, to a lesser extent, lopinavir pharmacokinetics but without influencing their cellular exposure. This confounding influence of ORM should be taken into account for appropriate interpretation of therapeutic drug monitoring results. Further studies are needed to investigate whether the measure of unbound drug plasma concentration gives more meaningful information than total drug concentration for indinavir and lopinavir.

Effects of oxcarbazepine on cognitive function in children and adolescents with partial seizures

The authors investigated the effect of oxcarbazepine on cognitive function in children and adolescents (6 to younger than 17 years of age) with newly diagnosed partial seizures in an open-label comparison with standard antiepileptic drug therapy (carbamazepine and valproate). No differences in cognitive tests were observed between oxcarbazepine and carbamazepine/valproate over a 6-month treatment period.

Long-lasting reactivity and high frequency of drug-specific T cells after severe systemic drug hypersensitivity reactions

BACKGROUND: Drug-reactive T cells are involved in most drug-induced hypersensitivity reactions. The frequency of such cells in peripheral blood of patients with drug allergy after remission is unclear. OBJECTIVE: We determined the frequency of drug-reactive T cells in the peripheral blood of patients 4 months to 12 years after severe delayed-type drug hypersensitivity reactions, and whether the frequency of these cell differs from the frequency of tetanus toxoid-reactive T cells. METHODS: We analyzed 5 patients with delayed-type drug hypersensitivity reactions, applying 2 methods: quantification of cytokine-secreting T cells by enzyme-linked immunospot (ELISpot), and fluorescent dye 5,6-carboxylfluorescein diacetate succinimidyl ester (CFSE) intensity distribution analysis of drug-reactive T cells. RESULTS: Frequencies found were between 0.02% and 0.4% of CD4(+) T cells reacting to the respective drugs measured by CFSE analysis, and between 0.01% and 0.08% of T cells as determined by ELISpot. Reactivity was seen neither to drugs to which the patients were not sensitized nor in healthy individuals after stimulation with any of the drugs used. CONCLUSION: About 1:250 to 1:10,000 of T cells of patients with drug allergy are reactive to the relevant drugs. This frequency of drug-reactive T cells is higher than the frequency of T cells able to recognize recall antigens like tetanus toxoid in the same subjects. A substantial frequency could be observed as long as 12 years later in 1 patient even after strict drug avoidance. Patients with severe delayed drug hypersensitivity reactions are therefore potentially prone to react again to the incriminated drug even years after strict drug avoidance.

Effects of epinephrine, norepinephrine, and phenylephrine on microcirculatory blood flow in the gastrointestinal tract in sepsis

OBJECTIVE: The use of vasopressors for treatment of hypotension in sepsis may have adverse effects on microcirculatory blood flow in the gastrointestinal tract. The aim of this study was to measure the effects of three vasopressors, commonly used in clinical practice, on microcirculatory blood flow in multiple abdominal organs in sepsis. DESIGN: Random order, cross-over design. SETTING: University laboratory. SUBJECTS: Eight sedated and mechanically ventilated pigs. INTERVENTIONS: Pigs were exposed to fecal peritonitis-induced septic shock. Mesenteric artery flow was measured using ultrasound transit time flowmetry. Microcirculatory flow was measured in gastric, jejunal, and colon mucosa; jejunal muscularis; and pancreas, liver, and kidney using multiple-channel laser Doppler flowmetry. Each animal received a continuous intravenous infusion of epinephrine, norepinephrine, and phenylephrine in a dose increasing mean arterial pressure by 20%. The animals were allowed to recover for 60 mins after each drug before the next was started. MEASUREMENTS AND MAIN RESULTS: During infusion of epinephrine (0.8 +/- 0.2 mug/kg/hr), mean arterial pressure increased from 66 +/- 5 to 83 +/- 5 mm Hg and cardiac index increased by 43 +/- 9%. Norepinephrine (0.7 +/- 0.3 mug/kg/hr) increased mean arterial pressure from 70 +/- 4 to 87 +/- 5 mm Hg and cardiac index by 41 +/- 8%. Both agents caused a significant reduction in superior mesenteric artery flow (11 +/- 4%, p < .05, and 26 +/- 6%, p < .01, respectively) and in microcirculatory blood flow in the jejunal mucosa (21 +/- 5%, p < .01, and 23 +/- 3%, p < .01, respectively) and in the pancreas (16 +/- 3%, p < .05, and 8 +/- 3%, not significant, respectively). Infusion of phenylephrine (3.1 +/- 1.0 mug/kg/min) increased mean arterial pressure from 69 +/- 5 to 85 +/- 6 mm Hg but had no effects on systemic, regional, or microcirculatory flow except for a 30% increase in jejunal muscularis flow (p < .01). CONCLUSIONS: Administration of the vasopressors phenylephrine, epinephrine, and norepinephrine failed to increase microcirculatory blood flow in most abdominal organs despite increased perfusion pressure and-in the case of epinephrine and norepinephrine-increased systemic blood flow. In fact, norepinephrine and epinephrine appeared to divert blood flow away from the mesenteric circulation and decrease microcirculatory blood flow in the jejunal mucosa and pancreas. Phenylephrine, on the other hand, appeared to increase blood pressure without affecting quantitative blood flow or distribution of blood flow.

Molecular survival strategies of Echinococcus multilocularis in the murine host

Larval infection with Echinococcus multilocularis starts with the intrahepatic postoncospheral development of a metacestode that-at its mature stage-consists of an inner germinal and an outer laminated layer (GL ; LL). In certain cases, an appropriate host immune response may inhibit parasite proliferation. Several lines of evidence obtained in vivo and in vitro indicate the important bio-protective role of the LL. For instance, the LL has been proposed to protect the GL from nitric oxide produced by periparasitic macrophages and dendritic cells, and also to prevent immune recognition by surrounding T cells. On the other hand, the high periparasitic NO production by peritoneal exsudate cells contributes to periparasitic immunosuppression, explaining why iNOS deficienct mice exhibit a significantly lower susceptibility towards experimental infection. The intense periparasitic granulomatous infiltration indicates a strong host-parasite interaction, and the involvement of cellular immunity in control of the metacestode growth kinetics is strongly suggested by experiments carried out in T cell deficient mouse strains. Carbohydrate components of the LL, such as Em2(G11) and Em492, as well as other parasite metabolites yield immunomodulatory effects that allow the parasite to survive in the host. I.e., the IgG response to the Em2(G11)-antigen takes place independently of alpha-beta+CD4+T cells, and in the absence of interactions between CD40 and CD40 ligand. Such parasite molecules also interfere with antigen presentation and cell activation, leading to a mixed Th1/Th2-type response at the later stage of infection. Furthermore, Em492 and other (not yet published) purified parasite metabolites suppress ConA and antigen-stimulated splenocyte proliferation. Infected mouse macrophages (AE-MØ) as antigen presenting cells (APC) exhibited a reduced ability to present a conventional antigen (chicken ovalbumin, C-Ova) to specific responder lymph node T cells when compared to normal MØ. As AE-MØ fully maintain their capacity to appropriately process antigens, a failure in T cell receptor occupancy by antigen-Ia complex or/and altered co-stimulatory signals can be excluded. Studying the status of accessory molecules implicated in T cell stimulation by MØ, it could be shown that B7-1 (CD80) and B7-2 (CD86) remained unchanged, whereas CD40 was down-regulated and CD54 (=ICAM-1) slightly up-regulated. FACS analysis of peritoneal cells revealed a decrease in the percentage of CD4+ and CD8+T cells in AE-infected mice. Taken together the obstructed presenting-activity of AE-MØ appeared to trigger an unresponsiveness of T cells leading to the suppression of their clonal expansion during the chronic phase of AE infection. Interesting information on the parasite survival strategy and potential can be obtained upon in vitro and in vivo treatment. Hence, we provided very innovative results by showing that nitazoxanide, and now also, respectively, new modified compounds may represent a useful alternative to albendazole. In the context of chemotherapeutical repression of parasite growth, we searched also for parasite molecules, whose expression levels correlate with the viability and growth activity of E. multilocularis metacestode. Expression levels of 14-3-3 and II/3-10, relatively quantified by realtime reverse transcription-PCR using a housekeeping gene beta-actin, were studied in permissive nu/nu and in low-permissive wild type BALB/c mice. At 2 months p.i., the transcription level of 14-3-3 was significantly higher in parasites actively proliferating in nu/nu mice compared to parasites moderately growing in wild type mice. Immunoblotting experiments confirmed at the protein level that 14-3-3 was over-expressed in parasites derived from nu/nu mice at 2 months p.i. In vitro-treatment of E. multilocularis with an anti-echinococcal drug nitazoxanide for a period of 8 days resulted in a significant decrease of both 14-3-3 and II/3-10 transcription levels,

The effects of dexmedetomidine/remifentanil and midazolam/remifentanil on auditory-evoked potentials and electroencephalogram at light-to-moderate sedation levels in healthy subjects

Avoidance of excessively deep sedation levels is problematic in intensive care patients. Electrophysiologic monitoring may offer an approach to solving this problem. Since electroencephalogram (EEG) responses to different sedation regimens vary, we assessed electrophysiologic responses to two sedative drug regimens in 10 healthy volunteers. Dexmedetomidine/remifentanil (dex/remi group) and midazolam/remifentanil (mida/remi group) were infused 7 days apart. Each combination of medications was given at stepwise intervals to reach Ramsay scores (RS) 2, 3, and 4. Resting EEG, bispectral index (BIS), and the N100 amplitudes of long-latency auditory-evoked potentials (ERP) were recorded at each level of sedation. During dex/remi, resting EEG was characterized by a recurrent high-power low-frequency pattern which became more pronounced at deeper levels of sedation. BIS Index decreased uniformly in only the dex/remi group (from 94 +/- 3 at baseline to 58 +/- 14 at RS 4) compared to the mida/remi group (from 94 +/- 2 to 76 +/- 10; P = 0.029 between groups). The ERP amplitudes decreased from 5.3 +/- 1.3 at baseline to 0.4 +/- 1.1 at RS 4 (P = 0.003) in only the mida/remi group. We conclude that ERPs in volunteers sedated with dex/remi, in contrast to mida/remi, indicate a cortical response to acoustic stimuli, even when sedation reaches deeper levels. Consequently, ERP can monitor sedation with midazolam but not with dexmedetomidine. The reverse is true for BIS.