Differential effects of cyclosporin A and tacrolimus on magnesium influx in Caco2 cells.

PURPOSE: Hypomagnesemia with urinary magnesium (Mg) wasting is a well acknowledged side effect of cyclosporin A (CsA) and tacrolimus (FK506) treatments. TRPM6, TRPM7 and MagT1 are involved in the active transcellular Mg transport processes in intestine and kidney. Since Mg homeostasis is tightly controlled by the dynamic action of intestinal absorption of dietary Mg and renal excretion of Mg, we investigated whether CsA and FK506 in commercially available solutions for clinical use decrease the expression and the function of TRPM6, TRPM7 or MagT1 in the intestinal epithelial cell line Caco2. METHODS: Changes of intracellular free Mg concentrations were measured by Mag-fura-2 imaging in Mg-free medium after the addition of 1 mM MgCl2. TRPM6, TRPM7 and MagT1 were evidenced in cells by immunofluorescence. Proteins and mRNAs were quantified after 18 hours of treatment with CsA or FK506 by western-blot and real-time RT-PCR analyses, respectively. RESULTS: TRPM6 and MagT1 were evidenced on all cell membranes, TRPM7 only on the inner membranes. CsA was responsible for a profound decrease in Mg2+ influx in intestinal epithelial cells, which may result in a decrease of intestinal Mg absorption, whereas FK506 was responsible for a marked increase in Mg2+ influx. Neither CsA nor FK506 altered TRPM6, TRPM7 or MagT1 mRNA levels or MagT1 protein level. CONCLUSIONS: In Caco2 cells, Mg2+ influx was inhibited by CsA solutions whereas enhanced by FK506 solutions, without alteration of MagT1, TRPM6 and TRPM7 expression, leading to the conclusion that CsA and FK506 have opposite effects in the functional activity of the Mg transporters herein examined. In clinical use, FK506 should be preferred for patients at risk for hypomagnesemia.

Biodistribution and anticancer activity of a new Vinca alkaloid encapsulated into long-circulating liposomes.

S12363 is a potent therapeutic agent with a strong in vitro activity against a variety of tumor types but also a high in vivo toxicity. Loading of this drug into long-circulating liposomes is expected to enhance its therapeutic index. Pharmacokinetics of liposomal S12363 showed that circulating S12363 was entrapped into liposomes until 24 hours after intravenous injection in mice. The liposomal formulation significantly increased the plasma concentration, half-life, and AUC and decreased the plasma clearance rates and volume of distribution of S12363. Liposome extravasation was evaluated with two tumor models by both microscopic analysis and liposome radiolabeling. Liposome accumulation was much more important in the case of B16 melanoma, compared to H460 tumor, with both inoculated subcutaneously and with comparable size. H460 tumor was also inoculated into the lung. The tumor localization did not influence liposome accumulation into the tissue. The liposomal formulation injected into mice bearing B16 melanoma allowed a 10-fold accumulation of S12363 into the tumor interstitium, as compared to the solution. Bioluminescence data, supported by the survival curves of the animals, showed that S12363-liposomes were able to significantly restrict B16 melanoma progression and increase mice survival.

Threshold to N-methyl-D-aspartate-induced seizures in mice undergoing chronic nutritional magnesium deprivation is lowered in a way partly responsive to acute magnesium and antioxidant administrations.

Magnesium deficiency may be induced by a diet impoverished in magnesium. This nutritional deficit promotes chronic inflammatory and oxidative stresses, hyperexcitability and, in mice, susceptibility to audiogenic seizures. Potentiation by low-magnesium concentrations of the opening of N-methyl-D-aspartate (NMDA) receptor/calcium channel in in vitro and ex vivo studies, and responsiveness to magnesium of in vivo brain injury states are now well established. By contrast, little or no specific attention has been, however, paid to the in vivo NMDA receptor function/excitability in magnesium deficiency. The present work reports for the first time that, in mice undergoing chronic nutritional deprivation in magnesium (35 v. 930 parts per million for 27 d in OF1 mice), NMDA-induced seizure threshold is significantly decreased (38 % of normal values). The attenuation in the drop of NMDA seizure threshold (percentage of reversal) was 58 and 20 % upon acute intraperitoneal administrations of magnesium chloride hexahydrate (28 mg magnesium/kg) and the antioxidant ebselen (20 mg/kg), respectively. In nutritionally magnesium-deprived animals, audiogenic seizures are completely prevented by these compound doses. Taken as a whole, our data emphasise that chronic magnesium deprivation in mice is a nutritional in vivo model for a lowered NMDA receptor activation threshold. This nutritional model responds remarkably to acute magnesium supply and moderately to acute antioxidant administration.

Impact of cyclosporine A on magnesium homeostasis: clinical observation in lung transplant recipients and experimental study in mice.

BACKGROUND: Hypomagnesemia is a common finding in patients receiving cyclosporine A (CsA) therapy. The relationship between CsA-induced hypomagnesemia and nephrotoxicity and the effects of oral magnesium (Mg) supplementation remain unclear. After a retrospective analysis of the time-course of plasma Mg and creatinine levels in lung allograft recipients treated with both CsA and oral Mg supplementation, we investigated the effects of CsA treatment on Mg homeostasis in mice with normal or Mg-deficient diet and the effects of oral Mg supplementation on plasma Mg levels. METHODS: Thirty lung-allograft recipients entered the retrospective study. One thousand two hundred twenty-eight blood samples were analyzed for blood and creatinine levels. Cyclosporine A (50 mg/kg/day by intraperitoneal injection) was administered to mice maintained on normal diet (1400 ppm) or Mg-deficient (50 ppm) diet. Magnesium levels were determined in plasma, urine, feces and femur, and creatinine levels were determined in plasma and urine. RESULTS: Plasma Mg concentration declines from the day of transplantation in 36.7% of the patients despite Mg supplementation, without correlation with creatinine changes. In mice, CsA induced an early moderate hypomagnesemia, which could not be ameliorated by oral Mg supplementation and was aggravated by low-Mg dietary, late increase in plasma creatinine and decrease in urine creatinine without histological signs of renal injury, decrease in intestinal Mg absorption and Mg mobilization from bone. CONCLUSION: Cyclosporine A treatment may induce moderate hypomagnesemia that is aggravated by inadequate Mg intake and is not ameliorated by Mg supplementation. Because of the clinical complications of hypomagnesemia, Mg should be monitored regularly in allograft recipients receiving CsA.

Effect of hypomagnesemia on allogeneic activation in mice.

INTRODUCTION: Magnesium (Mg) plays an essential role in a wide range of fundamental cellular reactions. It has been reported that in rodents Mg-deficient diet-induced hypomagnesemia results in an early inflammation. We have previously shown that chronic severe hypomagnesemia was associated neither with endothelial cell activation nor with an inflammatory process which are crucial in the allograft rejection process. T cell allogeneic stimulation activates the phosphatase calcineurin which triggers the signaling pathways leading to IL-2 synthesis and lymphocyte proliferation. Full activation of calcineurin requires Mg. Surveys suggest that a significant number of people consume less Mg than the international dietary reference intakes leading to hypomagnesemia in 2.5% to 15% of the general population. OBJECTIVE: The aim of the study was to investigate the effects of hypomagnesemia on lymphocyte allogeneic activation and proliferation in a murine model of dietary-induced hypomagnesemia. METHODS: C57BL/6J (H-2(b), Mls(b)) mice were given normal Mg-containing diet (1400 ppm Mg, control mice), or synthetic Mg-deficient diets containing either 50 ppm Mg or 150 ppm Mg for 28 days. Serum Mg levels were determined at days 5, 14 and 28. In parallel, complete urine and faeces were collected by using metabolic cages during a 24 h period for Mg determinations. Splenocytes from C57BL/6 mice fed either normal diet or 50 ppm Mg-diet were used as responder cells in mixed lymphocyte reaction (MLR) performed with splenocytes from C3H/He mice (H-2(k), Mls(IIa)) and C57BL/6 mice fed normal diet as stimulators for allogeneic and isogeneic conditions, respectively. TGF-beta and IL-2 productions were quantified in the supernates of mixed splenocytes cultures. 3x10(6) splenocytes from mice fed 50 ppm Mg-diet were used for calcineurin activity determination at day 28. RESULTS: In mice fed 150 ppm Mg-diet, moderate hypomagnesemia was observed from day 5 to day 28. Oral supplementation with Mg pidolate (5 or 20 mg Mg/kg/day) could not restore normal serum Mg levels. Serum Mg concentration early decreased in mice fed 50 ppm Mg-diet to achieve stabilized severe hypomagnesemia at days 14 and 28. Urine Mg concentration early dramatically fell down then stabilized in mice fed Mg-deficient diets. In MLR performed at day 28 with splenocytes from mice fed 50 ppm Mg-diet, proliferation and IL-2 production in allogeneic conditions were similar to control mice. No TGF-beta production was detected in any group. Lastly, calcineurin activity measured at day 28 was significantly lower in splenocyes from mice fed 50 ppm Mg-diet than in mice fed control diet. CONCLUSION: Mg-deficiency does not alter splenocyte allogeneic activation and proliferation and IL-2 production in vitro, although it partially inhibits calcineurin activity. We hypothesize that the remaining activity is sufficient for IL-2 gene normal activation. Alternatively, Mg-deficiency may trigger other signaling pathways leading to IL-2 production.

Neuroprotective gene profile in the brain of magnesium-deficient mice.

BACKGROUND: Magnesium (Mg) deficiency may lead to serious metabolic, biological and organic dysfunctions, and cause various clinical disorders. In the current study, we explore endothelial cell activation, inflammation and cell death induced in the brain of adult mice by Mg-deficient diet. METHODS AND RESULTS: Neither TNFalpha, substance P, sTNFRI, sTNFRII proteins (ELISA), nor TNFalpha, adherence molecules and prolactin mRNAs, nor NK1R (immunohistochemistry on brain sections) were up-regulated. No inflammatory infiltrates and no apoptotic cells were observed. Using cDNA assay, we showed a neuroprotective, anti-apoptotic and neurotrophic gene expression profile in the brain at early stage of hypomagnesemia. As a model for neuronal injury, mild sound stimulation of Mg-deficient mice without convulsive seizures triggers neither the release of substance P, nor the development of an inflammatory process or cell death in the brain. CONCLUSION: Our results suggest that Mg-deficiency in mice favours the development of a neuroprotective environment in the brain.

MDR1A (ABCB1)-deficient CF-1 mutant mice are susceptible to cerebral malaria induced by Plasmodium berghei ANKA.

Under experimental conditions, Plasmodium berghei infection causes cerebral malaria (CM) in susceptible strains of mice such as C57BL/6 and CBA/Ca, whereas BALB/c or DBA/2J strains serve as a model for CM-resistant mice. The aim of the present study was to investigate the susceptibility of the CF1 mouse strain, carrying a spontaneous mutation of the mdr1a gene, to infection with Plasmodium berghei ANKA (PbA). The mdr1a gene codes for P-glycoprotein (P-gp/ABCB1), an efflux pump that is one of the major components of the blood-brain barrier. P-gp effluxes a broad range of xenobiotics from the brain to blood, preventing accumulation and toxicity in the central nervous system. CFI mdr1a (-/-) mice are used to investigate drug transport by efflux pumps. Because many antimalarial agents are effluxed by P-gp (mefloquine, quinine), it was important to determine whether CF1 mice can develop cerebral malaria to predict drug toxicity during cerebral malaria. Our work showed that CF1 mdr1a (-/-) mice are susceptible to PbA. CF1 and C57BL/6N mice (the reference strain) infected with PbA have similar profiles with regard to clinical signs, brain histological lesions, and brain macrophagic activation observed by immunohistological methods.

Allogeneic activation is attenuated in a model of mouse lung perfused with magnesium-deficient blood.

Hypomagnesemia, which is frequently observed in patients treated with calcineurin inhibitors to prevent rejection after allogeneic transplantation, has been associated with a faster rate of decline in allograft function. The effect of hypomagnesemia on lung allograft has not been reported yet. In our model of isolated mouse lung, we have evaluated the early effects of allogeneic lung perfusion with blood from magnesium (Mg)-deficient mice for 3 h on lung activation and remodelling, compared to isogeneic perfusion. Hypomagnesemia (0.21+/-0.07 mmol Mg(2+)/l) was observed in blood from Mg-deficient mice, but no inflammatory pattern. The mRNA level of the intercellular adhesion molecule (ICAM)-1, but neither of the vascular cell adhesion molecule (VCAM)-1, nor of the cytokines tumor necrosis factor (TNF)alpha and interleukin (IL)-2, was enhanced (p<0.05). Although caspase-3 mRNA was transiently enhanced, no apoptotic cells were evidenced in lung tissues even after 3 h. Using cDNA array, we found that the genes encoding RANKL, RANK, TNFR2, NFATX, IL-1R2, IL-6R gp130, SOCS3, PDGFRB, P63, CSF3R, CXCL1, CXCL5, CX3CL1, CSF1, which are involved in inflammation and apoptosis regulation, were markedly up-regulated in allogeneic conditions. Our results support a limited allogeneic activation and an early stage of the inflammatory process in lung, at the time of inflammatory cell recruitment without lung tissue remodelling, as a result of hypomagnesemia. These findings suggest that cyclosporine-related hypomagnesemia, observed in most of the transplanted patients, does not constitute an additional risk for lung allograft outcome.

Role of E-selectin in cell apoptosis induced by allogeneic blood perfusion in isolated mouse lung.

BACKGROUND: In a model of mouse isolated lung, we have recently demonstrated that E-selectin is involved in the activation of endothelial cells induced by allogeneic blood perfusion. In the present study, we explored the signaling pathway of apoptosis induced by E-selectin triggering. METHODS: Lungs were perfused for 3 hours with fresh blood in the absence or presence of an anti-E-selectin monoclonal antibody, or a protein kinase C (PKC), protein tyrosine phosphatase (PTP), or protein tyrosine kinase (PTK) inhibitor. The number of apoptotic cells in lung sections was determined by a TUNEL method. mRNAs for Fas, FasL and caspase-8, and for Bad, Bax, Bcl-w, Bcl-xL and caspase-9, for the FasL and the mitochondrial cytochrome-c pathways of apoptosis, respectively, and mRNA for the effector caspase-3 were quantified in lung tissues by RT-PCR. PTP and Src-PTK activities were also measured. RESULTS: After 3 hours of allogeneic perfusion, we observed a significant increase in: 1) the number of apoptotic cells in lung sections, 2) mRNA levels of FasL, Bcl-xL, caspase-8 and caspase-3, and 3) PTP activity (P < 0.05 compared with isogeneic perfusion). Surprisingly, mRNA levels of the proapoptotic genes Bad and Bax were significantly decreased (P < 0.05). PTK activity and caspase-9 mRNA level were not affected. Blocking anti-E-selectin mAbs and inhibitors for PKC, PTP, and PTK resulted in a significant reduction of apoptosis. CONCLUSIONS: In our model, the engagement of E-selectin induced by endothelial cell allogeneic activation appeared to be a prerequisite for lung apoptosis, which involved FasL and increase of PTP activity. Blockade of apoptosis with selective inhibitors may be a promising approach to the treatment/prevention of lung graft injury.

E-selectin early overexpression induced by allogeneic activation in isolated mouse lung.

BACKGROUND: The interaction between host lymphocytes and graft endothelial cells plays an important role in graft rejection. METHODS: Using our model of isolated ventilated lung from female mouse perfused with fresh blood from either isogeneic or allogeneic male mouse for 3 hours without noticeable ischemia, we have investigated the kinetics of the early events after endothelial cell triggering by E-selectin engagement. RESULTS: Isogeneic perfusion induced nonspecific endothelial cell activation, which was characterized by up-regulation of E-selectin, intercellular adhesion molecule (ICAM)-1, and of the pro-inflammatory cytokines tumor necrosis factor (TNF)-alpha, interleukin (IL)-2, and lymphotoxin-alpha (mRNAs by real-time polymerase chain reaction). Allogeneic perfusion was characterized after 3 hours by an additional loose adhesion of lymphocytes mediated by the E-selectin and related to the allogeneic activation of endothelial cells. These in turn expressed the I-A molecule (immunostaining). ICAM-1 and lymphocyte function-associated antigen (LFA)-3 mRNA levels were significantly increased in lung extracts after 2 hours, then vascular cell adhesion molecule (VCAM)-1 and TNF-alpha mRNAs after 3 hours without evidence of TNF-alpha production (enzyme-linked immunoadsorbent assay). The major participation of the E-selectin in early allogeneic activation by way of the protein kinase (PK)C pathway was confirmed by using a neutralizing anti-CD62E monoclonal antibody or the inhibitory PKC 19-31 fragment. CONCLUSIONS: Altogether, our results demonstrate that E-selectin expression (1) is not a consequence of TNF-alpha triggering, (2) up-regulates its own expression and expression of I-A, VCAM-1, TNF-alpha, and lymphotoxin-alpha mRNAs, and (3) down-regulates expression of LFA-3 and ICAM-1 mRNAs. In conclusion, in our physiologic model, the E-selectin highly participates in the loose adhesion of allogeneic lymphocytes and in the early activation of endothelial cell and therefore in structural and functional lung alterations.

Endothelial cell early activation induced by allogeneic lymphocytes in isolated perfused mouse lung.

BACKGROUND: The early inflammatory response induced following vascularized organ allotransplantation is mediated by adhesin-ligand interactions between blood leukocytes and allogeneic endothelial cells. In the present study, we focused on the role of allogeneic blood lymphocytes in early immune alterations following lung reperfusion. METHODS: We developed an experimental model of isolated and ventilated lung in female C57BL/6 mouse perfused with either isogeneic (C57BL/6) or allogeneic (C3H/He) male mouse fresh blood for 3 hours. SRY DNA quantification by polymerase chain reaction (PCR) was used to assess the presence of perfusate cells in lung extract. Using quantitative reverse transcriptase (RT)-PCR, we measured mRNA expression both of the adhesion molecules--intercellular adhesion molecule (ICAM)-1, lymphocyte function-associated antigen (LFA)-3, vascular cell adhesion molecule (VCAM)-1, and endothelial leukocyte adhesion molecule (ELAM)-1--and of tumor necrosis factor (TNF)-alpha in lung tissue. RESULTS: The number of blood lymphocytes is significantly decreased in allogeneic versus isogeneic condition at 3 hours of perfusion, without evidence of increased apoptosis or necrosis. In parallel, SRY DNA quantity recovered in the lung was 2.5 times higher in allogeneic condition, which was related to cells loosely adherent to endothelial cells. Lastly, the levels of mRNAs of all adhesion molecules and of TNF-alpha were significantly increased in allogeneic versus isogeneic conditions. CONCLUSION: These results demonstrate that an early interaction between allogeneic blood lymphocytes and vascular endothelial cells is correlated with a high mRNA expression both of adhesion molecules and of TNF-alpha in the perfused lung. Our model of a mouse lung perfused with fresh blood appears to be a useful clinical assessment system for in-depth investigation of early cell activation and the resulting intragraft immune alterations.

Heterotopic en bloc tracheobronchial transplantation with direct revascularization in pigs.

OBJECTIVE: This article describes the application of a novel aortic tube technique for directly revascularized tracheobronchial transplantation with dual blood supply in pigs. METHODS: Eleven adult Large White pigs underwent heterotopic tracheal transplantation with a dual revascularization technique (inferior thyroid artery and bronchial artery). Seven tracheobronchial grafts were perfused ex vivo, and hemodynamic data were collected. RESULTS: At the last evaluation, 6 pigs had normally epithelialized mucus-producing allografts with correct morphologic conformation and cartilage viability. The histopathologic examination revealed homogeneous tissue regardless of biopsy site (trachea, carina, or bronchi), demonstrating the efficacy of the revascularization procedure. Four animals had early ischemic necrosis develop, 2 from acute rejection and 2 from technical mishap. One additional pig had acute rejection starting on the 14th postoperative day. The CD4(+)/CD8(+) ratio was maintained close to or above 0.8 in the subgroup with rejection and below 0.6 in the animals that were correctly immunosuppressed. Pressure-flow curves in 7 ex vivo tracheobronchial grafts showed a nonsignificant difference (P <.12) in vascular resistance between the bronchial artery territory (lower resistance) and the inferior thyroid artery territory. CONCLUSIONS: For the first time, a transplantation technique encompassing the entire trachea, carina, and stem bronchi has been made possible. By means of the dual inferior thyroid and bronchial artery axis, we were able to obtain a structurally healthy and functional graft to replace the main airway.

How to improve current therapeutic standards in upper respiratory infections: value of fusafungine.

Despite guidelines and educational programs, systemic antibiotics and anti-inflammatory drugs are often inappropriately prescribed in upper respiratory tract infections (URTIs), although they are most often of viral origin, generally benign, and self-limiting with spontaneous recovery in more than 80% of cases. Reduced use of systemic antibiotics is crucial in the current context of concern about emerging antibiotic resistance and reducing unnecessary costs associated both with drug over-consumption and with the management of the consequences of antibiotic resistance. Local bacterial or viral infection of the airways induces an early inflammatory reaction. Although this inflammatory reaction has a beneficial effect in the capture and destruction of the pathogens, it can be responsible for deleterious tissue damage and vascular alterations leading to a self-perpetuating cycle of events. A wide array of medicines is available for symptomatic relief of URTIs: many of them are partially effective in reducing symptoms, but none is curative. Local administration of antibiotics and anti-inflammatory drugs allows drug delivery directly to the target site of infection and inflammation, i.e., the respiratory mucosa, thus enabling a higher concentration of the drug, which results in smaller doses to be given, decreased potential for systemic toxicity, fewer side effects, protection of other flora, and rapid relief. Fusafungine is a naturally occurring peptide antibiotic with anti-inflammatory properties, which selectively targets the tissue reaction and preserves the natural antibacterial and antiviral defences. It is indicated for topical use in nose and throat infections. A recent analysis of French general practitioners' (GPs) prescribing pattern in the field of URTIs has demonstrated that prescription of fusafungine has achieved what many educational programs have failed to do: a significant reduction in the 'real life' prescription of systemic antibiotics and antiinflammatory drugs, without the side effects of corticosteroids and vasoconstrictive agents, and without impact on microbial ecology.

Influence of the pro-inflammatory cytokines on P-glycoprotein expression and functionality.

PURPOSE: P-glycoprotein (P-gp) is involved in the transport of many drugs at different barriers with consequence in terms of drug distribution and elimination. The expression and activity of P-gp can be modulated by different factors and pathologies. The present article reviews the knowledge regarding the effect of pro-inflammatory cytokines (TNFalpha, IL-1beta, IL-6, IL-2, IFNgamma) on the expression and the functionality of P-gp at three major sites of drug absorption and disposition: the liver, the blood-brain barrier, and the intestine. METHODS: The various methods used to study the effect of pro-inflammatory cytokines include in vivo models (i.e. animals infected with Staphylococcus sp, animals injected with bacterial lipopolysaccharide or directly with cytokines, ...) and in vitro models (i.e. primary rat hepatocytes, human brain endothelial cells, ...). RESULTS: The data on P-gp expression and/or function may differ according to the compound used to induce inflammation. However, there is a general trend towards a decrease in both the expression of P-gp (mRNA and protein) and its functionality. Transcription factors and nuclear receptors are probably involved in this regulation. CONCLUSION: Cytokines may interfere with P-gp. Hence, in pathological conditions (inflammation, infection, ...), the expression and functionality of P-glycoprotein may be modulated with consequences for drug disposition and, consequently treatment efficacy.

[The impact of fusafungine on the prescription of antibiotics in the treatment of rhinopharyngitis]. / Impact de la fusafungine sur la prescription d'antibiotiques dans le traitement de la rhinopharyngite en France.

OBJECTIVE: The analysis in France, during the period 01/12/99 to 30/11/2000, of the prescription of systemic antibiotics in patients with rhinopharyngitis and of the variables statistically related to such prescriptions and the potential role of fusafungine in the form of a rhinopharyngeal spray. METHODS: A retrospective study, based on a panel of 1,010 general practitioners, in a cohort of 30,568 patients presenting with rhinopharyngitis. The fusafungine group consisted of 16,076 patients who had rhinopharyngitis and in whom fusafungine was prescribed. The control group consisted of 14,492 patients with rhinopharyngitis without prescription of fusafungine. The overall rate of antibiotic prescription was documented. A stepwise statistical analysis was conducted to specify the variables statistically associated with the prescription of a systemic antibiotic. The rate of prescription of a systemic antibiotic and the cost of the treatment were also compared within both groups. RESULTS: The overall rate of systemic antibiotic prescription was 52.9%, falling from 60.4% in the group without fusafungine down to 46.2% in the group with fusafungine (p<0.01) whichever the systemic antibiotic prescribed. The stepwise analysis documented various variables that appear to be related to the systemic antibiotic prescription. A saving of 0.7 euros per prescription was noted in the fusafungine group. CONCLUSION: Although various variables appear to influence systemic antibiotic prescription in patients with rhinopharyngitis, our study shows that prescription of fusafungine in spray from led to statistically significant reduction in systemic antibiotic prescription.

Magnesium-deficiency does not alter calcineurin inhibitors activity in mice.

INTRODUCTION: Tacrolimus (TAC) and cyclosporin (CsA) are commonly responsible for hypomagnesemia that predisposes in turn for hypertension, renal impairment and encephalopathy. OBJECTIVE: The effects of TAC on Mg(2+)-homeostasis and of pre-existing Mg(2+)-deficiency on TAC immunosuppressive activity were compared to CsA in mice. METHODS: Mg(2+) was quantified in plasma, erythrocytes, urine, feces, and femurs from mice treated with TAC 5mg/kg/day. Immunosuppression was assessed in splenocytes by mixed lymphocyte reaction, IL-2 quantification and CN activity determination. RESULTS: Plasma and urine Mg(2+) levels in TAC-treated mice were significantly lower from day 7 until day 21 (p<0.05 versus control) and returned to control value at day 28. Mg(2+) levels were unchanged in erythrocyte, feces and femur. Inhibition of allogeneic proliferation, IL-2 production and CN activity were 68, 56 and 30% lower (p<0.01) after 7 days of TAC-treatment, and 72, 68 and 51% lower (p<0.01) after 7 days of CsA-treatment with a dose of 50mg/kg/day. Dietary-induced hypomagnesemia resulted in significant inhibition of CN activity (p<0.01) without alteration of IL-2 production or allogeneic proliferation. However, it did not alter the effects observed with CsA- or TAC-treatment on allogeneic proliferation, IL-2 production and CN activity. CONCLUSION: By contrast with CsA, long-course TAC-treatment induced an early, but transient, and moderate hypomagnesemia without alteration of bone or erythrocyte stocks, intestinal absorption or renal function. Therefore, in clinical use, TAC should be preferred to CsA in patients with pathological or pharmacological conditions which favor Mg(2+)-deficiency. However, dietary-induced hypomagnesemia did not alter the immunosuppressive effects of TAC and CsA.

A study of magnesium deficiency in human and experimental pulmonary hypertension.

Pulmonary hypertension (PH) is defined as an increase in mean pulmonary arterial pressure above 25 mmHg. Pulmonary vasoconstriction, cellular proliferation, inflammation, and oxidative stress are involved in the pathophysiology of PH. Since hypomagnesemia was reported to promote endothelial cell dysfunction leading to inflammation and oxidative stress, we investigated the potential involvement of magnesium (Mg) deficiency in experimental and human PH. Our results indicate that Mg deficiency has no impact on hypoxia-induced PH development or severity, and that no reduction in Mg plasma concentration was observed in patients with severe pulmonary arterial hypertension. Thus, hypomagnesemia does not appear to play a role in the pathophysiology of experimental and human pulmonary hypertension.

Impact of cerebral malaria on brain distribution of mefloquine.

Cerebral malaria (CM) is the most severe complication of Plasmodium falciparum malaria. The aim of this study was to investigate the influence of CM on the cerebral uptake of mefloquine (MQ), in an experimental model of mice infected with Plasmodium berghei ANKA (PbA). Drug diffusion in brain is closely related to efflux pumps such as P-glycoprotein (P-gp/ABCB1/MDR1) and Breast Cancer Resistant Protein (BCRP/ABCG2), two major components of the blood-brain barrier (BBB) which can be modified by inflammation and/or infection. After a single IP dose, MQ concentrations were measured by liquid chromatography in blood and brains of mice infected with Plasmodium berghei ANKA and compared with that of non-infected mice. Our results show that MQ brain concentrations were decreased in CM mice versus healthy mice (0.77 versus 1.31 for brain/plasma concentrations). Although MQ is transported out of endothelial cells by P-glycoprotein, this result cannot be related to this transporter as we have previously shown that CM does not alter P-gp function (personal data). CM induces a reduction of MQ brain transport and, therefore, an increase of central toxicity due to MQ should not be expected during CM.

Efavirenz does not interact with the ABCB1 transporter at the blood-brain barrier.

PURPOSE: This work characterizes the interactions between efavirenz (EFV) and P-glycoprotein (P-gp/ABCB1) at the blood-brain barrier (BBB) and predicts the possible consequences on the brain uptake of coadministered P-gp substrates. METHODS: The uptake of EFV was measured in whole brains of rat and mdr1a-/- and mdr1a+/+ mice, and in GPNT cells (rat brain endothelial cell line) with and without P-gp inhibitors (PSC833, S9788, Quinidine). The effect of a single dose or multiple doses of EFV on the P-gp functionality was evaluated in vivo and in vitro by measuring the brain and cell uptake of digoxin, completed by the analysis of the P-gp expression at the rat BBB after repeated administrations of EFV. RESULTS: Inhibition of P-gp did not alter the uptake of EFV in rat brain and GPNT cells. The EFV brain/plasma ratio in mdr1a-/- mice, lacking the expression of P-gp, was not different from that in mdr1a+/+ mice. Moreover, a single dose of EFV did not modify the uptake of digoxin in rat brain and GPNT cells. Finally, the 3-day exposure of GPNT cells to EFV did not have any effect on the uptake of digoxin. Similarly, the 7-day treatment with EFV did not change the uptake of digoxin in rat brain nor the expression of P-gp at the BBB. CONCLUSION: EFV is strongly distributed in the brain, but is neither a substrate nor an inhibitor of the P-gp at the blood-brain barrier. On the other hand, EFV did not induce P-gp, allowing to sustain the brain accumulation of associated P-gp substrates such as protease inhibitors. These findings make EFV suitable for combinations circumventing the brain HIV-1 residency.

Antibiotic prescribing patterns of French GPs for upper respiratory tract infections: impact of fusafungine on rates of prescription of systemic antibiotics.

INTRODUCTION: Despite attempts to limit their use, systemic antibiotics are extensively prescribed for respiratory infections in France. This survey analyzed data from the Thales database, which contains information from 1010 representative French general practitioners (GPs). The objective was to assess French GP prescribing patterns in upper respiratory tract infections (URTIs) including the rate of prescription of systemic antibiotics and anti-inflammatory drugs in the presence or absence of prescribing fusafungine (Locabiotal) an antibiotic with anti-inflammatory activity indicated for local use in URTIs. Drug costs to the French National Sickness Fund were also assessed. METHODS: This was a retrospective, longitudinal, case-control analysis. Prescribing patterns and costs were compared between patients who did and patients who did not receive fusafungine for a URTI (rhinopharyngitis, tonsillitis, or an influenza-like condition). The fusafungine group consisted of all patients in the database who were prescribed fusafungine at least once between 1 December 1999 and 30 November 2000. The control group was made up of randomly selected patients, matched for age and sex with the study group, who received at least one drug prescription (but not fusafungine) for a URTI during the same period. Patients were selected at the time of their first prescription, and their records for 1 year were analyzed. RESULTS: Each group contained 22 164 patients. For URTIs overall, systemic antibiotics were widely prescribed (at a rate of 54.6% and 67.8% in the fusafungine and control groups, respectively; p < 0.01). The rate of prescription of systemic antibiotics, NSAIDs and corticosteroids per prescription and per episode was significantly lower in the fusafungine group than in the control group. The mean cost per prescription for the French National Sickness Fund was significantly lower for the three URTIs overall when fusafungine was prescribed (9.21 euros [euro] vs euro9.67; p < 0.01). The mean cost to the National Sickness Fund per prescription of systemic antibiotics, NSAIDs, and corticosteroids was also significantly lower in the fusafungine group compared with the control group. The cost of nasal preparations was higher in the fusafungine group because Locabiotal is classified as a nasal preparation. The cost per prescription to the National Sickness Fund was increased by the presence of systemic antibiotics, NSAIDs, or corticosteroids among the prescribed drugs and decreased with the prescription of fusafungine. CONCLUSION: When fusafungine was prescribed for URTIs, fewer systemic antibiotics were prescribed, an important result in the current context of concern about emerging antibiotic resistance. The use of fusafungine was associated with a lower mean cost per prescription to the French National Sickness Fund.