Gadopentetate meglumine activates mast cells to cause IgE-independent allergic reactions both in vitro and in vivo.

Gadopentetate meglumine (Gd-DTPA) is a commonly used magnetic resonance imaging (MRI) enhancer in the clinic for improving the clarity of MRI. Reports have shown that severe anaphylactoid reactions can occur after intravenous injections, whereas the underlying mechanisms remain undefined. In this study, the effects of Gd-DTPA in causing allergic like reactions and mast cells (MCs) activation were investigated both in vitro and in vivo. Gd-DTPA induced a severe hand paw swelling and body temperature decrease in murine topical cutaneous allergy model, and murine systemic allergy model. Meanwhile, serum IgE was not significantly changed. Histamine, tryptase, and cytokines release in mice serum and LAD2 cells supernatant were increased significantly both in mice serum and LAD2 supernatant after treated with Gd-DTPA. Subsequently, the changes in mRNA levels after Gd-DTPA activated MCs were analyzed by RNAseq and found to be related to inflammation signaling pathways. The study provides a demonstration for the adverse reaction mechanism of Gd-DTPA and its safe use in clinic.

Critical analysis of major and ancillary features of LI-RADS v2018 in the differentiation of small (≤ 2 cm) hepatocellular carcinoma from dysplastic nodules with gadobenate dimeglumine-enhanced magnetic resonance imaging.

OBJECTIVE: To evaluate the performance of major features, ancillary features, and categories of Liver Imaging Reporting and Data System (LI-RADS) version 2018 at magnetic resonance (MR) imaging in the differentiation of small hepatocellular carcinoma (HCC) from dysplastic nodules (DNs). PATIENTS AND METHODS: This retrospective study included cirrhotic patients with pathologically proven untreated HCCs and DNs (≤ 2 cm) and liver MR imaging performed with gadobenate dimeglumine contrast agent within 3 months before pathological analysis, between 2015 and 2018. 37 patients with 43 observations (17 HCCs and 26 DNs) met the inclusion criteria. Two radiologists assessed major and ancillary imaging features for each liver observation and assigned a LI-RADS v2018 category in consensus. Estimates of diagnostic performance of major features, ancillary features, and LI-RADS categories were assessed based on their sensitivity, specificity, positive (PPV), and negative predictive values (NPV). RESULTS: Major features (nonrim arterial phase hyperenhancement, nonperipheral "washout", and enhancing "capsule") had a sensitivity of 94.1%, 88.2%, and 41.2%, and a specificity of 57.7%, 42.3%, and 88.5% for HCC, respectively. Ancillary features (hepatobiliary phase hypointensity, mild-moderate T2 hyperintensity, restricted diffusion, and fat in the lesion more than adjacent liver) had a sensitivity of 94.1%, 64.7%, 58.8%, and 11.8%, and a specificity of 26.9%, 61.5%, 65.4%, and 76.9% for HCC, respectively. The LR-5 category (determined by using major features only vs. the combination of major and ancillary features) had a sensitivity of 88.2% at both evaluations and a specificity of 76.9% and 80.8% for HCC, respectively. The combination of LR-4, LR-5 categories (determined by using major features only vs. the combination of major and ancillary features) had a sensitivity of 94.1% at both interpretations and a specificity of 65.4% and 26.9% for HCC, respectively. The use of ancillary features modified LI-RADS category in 25.6% of observations (11/43), predominantly upgraded from LR-3 to LR4 (10/11), increasing the proportion of low-grade DNs and high-grade DNs categorized as LR-4 (from 15.4% to 61.5% and from 7.7% to 46.1%, respectively). CONCLUSIONS: The added value of ancillary features in combination with major features is limited for the non-invasive diagnosis of small HCC; however, their use modifies the final category in a substantial proportion of observations from LR-3 to LR-4, thus allowing possible changes in the management of patients at risk for HCC.

Hepatocyte Concentrations of Imaging Compounds Associated with Transporter Inhibition: Evidence in Perfused Rat Livers.

In the liver, several approaches are used to investigate and predict the complex issue of drug-induced transporter inhibition. These approaches include in vitro assays and pharmacokinetic models that predict how inhibitors modify the systemic and liver concentrations of the victim drugs. Imaging is another approach that shows how inhibitors might alter liver concentrations stronger than systemic concentrations. In perfused rat livers associated with a gamma counter that measures liver concentrations continuously, we previously showed how fluxes across transporters generate the hepatocyte concentrations of two clinical imaging compounds, one with a low extraction ratio [gadobenate dimeglumine (BOPTA)] and one with a high extraction ratio [mebrofenin (MEB)]. BOPTA and MEB are transported by rat organic anion transporting polypeptide and multiple resistance-associated protein 2, which are both inhibited by rifampicin. The aim of the study is to measure how rifampicin modifies the hepatocyte concentrations and membrane clearances of BOPTA and MEB and to determine whether these compounds might be used to investigate transporter-mediated drug-drug interactions in clinical studies. We show that rifampicin coperfusion greatly decreases BOPTA hepatocyte concentrations, but increases those of MEB. Rifampicin strongly decreases BOPTA hepatic clearance. In contrast, rifampicin decreases moderately MEB hepatic clearance and blocks the biliary intrinsic clearance, increasing MEB hepatocyte concentrations. In conclusion, low concentrations prevent the quantification of BOPTA biliary intrinsic clearance, while MEB is a promising imaging probe substrate to evidence transporter-mediated drug-drug interactions when inhibitors act on influx and efflux transporters.

Absence of clinical cerebellar syndrome after serial injections of more than 20 doses of gadoterate, a macrocyclic GBCA: a monocenter retrospective study.

Sound evidence of gadolinium accumulation in brain has been recently provided after repeated administrations of linear gadolinium-based contrast agents (GBCAs), especially at the cerebellum level. Although data regarding brain accumulation of macrocyclic GBCAs are more reassuring, there is now a genuine concern ("gadolinium-phobia") about possible long-term consequences of gadolinium deposits, especially in terms of cerebellar sequelae. We, therefore, questioned about the clinical impact of serial administration of gadoterate meglumine, a macrocyclic GBCA. In this retrospective study (2000-2016) of medical files of patients who received more than 20 administrations of gadoterate, we searched for cerebellar symptoms and signs developing during the regular follow-up. We reviewed medical files of ten patients (mean age 34.4 ± 20.8 years; 4 males, 6 females) who received 28.2 ± 5.3 doses of gadoterate (average total dose of GBCA 518 ± 226 ml; range 185-785 ml). Patients were examined by at least two medical specialists depending on initial diagnosis, and at least once by a neurosurgeon. Mean follow-up time was 91 months (range 49-168) and six out of ten patients experienced new symptoms or signs. No clinician reported the appearance of a rising cerebellar syndrome, nor newly appeared symptoms or signs suggested cerebellar toxicity. This retrospective clinical study shows no de novo clinical cerebellar syndrome following repeated administrations of gadoterate. Our results argue against a cerebellar toxicity of this macrocyclic agent. Still, confirmation in a larger number of subjects is required, as well as clinical studies concerning linear GBCAs whose structure and in vivo stability are distinct.

Gadolinium Brain Deposition after Macrocyclic Gadolinium Administration: A Pediatric Case-Control Study.

Purpose To determine whether signal intensity (SI) in T1 sequences as a potential indicator of gadolinium deposition increases after repeated administration of the macrocyclic gadolinium-based contrast agents (GBCAs) gadoteridol and gadoterate meglumine in a pediatric cohort. Materials and Methods This retrospective case-control study of children with brain tumors who underwent nine or more contrast material-enhanced brain magnetic resonance (MR) imaging studies from 2008 to 2015 was approved by the local ethics board. Informed consent was obtained for MR imaging. Twenty-four case patients aged 5-18 years and appropriate control patients with nonpathologic MR neuroimaging findings (and no GBCA administration), matched for age and sex, were inculded. SI was measured on unenhanced T1-weighted MR images for the following five regions of interest (ROIs): the dentate nucleus (DN), pons, substantia nigra (SN), pulvinar thalami, and globus pallidus (GP). Paired t tests were used to compare SI and SI ratios (DN to pons, GP to thalamus) between case patients and control patients. Pearson correlations between relative signal changes and the number of GBCA administrations and total GBCA dose were calculated. Results The mean number of GBCA administrations was 14.2. No significant differences in mean SI for any ROI and no group differences were found when DN-to-pons and GP-to-pulvinar ratios were compared (DN-to-pons ratio in case patients: mean, 1.0083 ± 0.0373 [standard deviation]; DN-to-pons ratio in control patients: mean, 1.0183 ± 0.01917; P = .37; GP-to-pulvinar ratio in case patients: mean, 1.1335 ± 0.04528; and GP-to-pulvinar ratio in control patients: mean, 1.1141 ± 0.07058; P = .29). No correlation was found between the number of GBCA administrations or the total amount of GBCA administered and signal change for any ROI. (Number of GBCA applications: DN: r = -0.254, P = .31; pons: r = -0.097, P = .65; SN: r = -0.194, P = .38; GP: r = -0.175, P = .41; pulvinar: r = -0.067, P = .75; total amount of administered GBCA: DN: r = 0.091, P = .72; pons: r = 0.106, P = .62; SN: r = -0.165, P = .45; GP: r = 0.111, P = .61; pulvinar: r = 0.173, P = .42.) Conclusion Multiple intravenous administrations of these macrocyclic GBCAs in children were not associated with a measurable increase in SI in T1 sequences as an indicator of brain gadolinium deposition detectable by using MR imaging. Additional imaging and pathologic studies are needed to confirm these findings. © RSNA, 2017 Online supplemental material is available for this article.

HSP90 Regulation of P2X7 Receptor Function Requires an Intact Cytoplasmic C-Terminus.

P2X7 receptors (P2X7Rs) are ATP-gated ion channels that display the unusual property of current facilitation during long applications of agonists. Here we show that facilitation disappears in chimeric P2X7Rs containing the C-terminus of the P2X2 receptor (P2X2R), and in a truncated P2X7R missing the cysteine-rich domain of the C-terminus. The chimeric and truncated receptors also show an apparent decreased permeability to N-methyl-d-glucamine(+) (NMDG(+)). The effects of genetic modification of the C-terminus on NMDG(+) permeability were mimicked by preapplication of the HSP90 antagonist geldanamycin to the wild-type receptor. Further, the geldanamycin decreased the shift in the reversal potential of the ATP-gated current measured under bi-ionic NMDG(+)/Na(+) condition without affecting the ability of the long application of agonist to facilitate current amplitude. Taken together, the results suggest that HSP90 may be essential for stabilization and function of P2X7Rs through an action on the cysteine-rich domain of the cytoplasmic the C-terminus.

Magnetic Resonance Imaging with a Weak Albumin Binding Contrast Agent can Reveal Additional Endoleaks in Patients with an Enlarging Aneurysm after EVAR.

OBJECTIVES/BACKGROUND: To examine the additional diagnostic value of magnetic resonance imaging (MRI) after administration of a weak albumin binding contrast agent in post-endovascular aneurysm repair (EVAR) patients with aneurysm growth with no or uncertain endoleak after computed tomography angiography (CTA). METHODS: This was a prospective diagnostic cross sectional study carried out between April 2011 and August 2013. MRI was performed in all patients with aneurysm growth≥5 mm after EVAR implantation and no or uncertain endoleak on CTA, or the inability, on CTA, to identify the source of a visible endoleak. All MRI scans were performed on a 1.5 T clinical MRI scanner after administration of a weak albumin binding contrast agent. The presence of endoleaks was assessed by visually comparing pre- and post-contrast T1-weighted images with fat suppression. Post-contrast images were acquired 5 and 15 minutes after contrast administration. RESULTS: Twenty-nine patients (26 men; 90%) with a median age of 74 years (interquartile range [IQR] 67-76) were included. The median interval between EVAR and MRI was 39 months (IQR 20-50). The median increase in maximum aneurysm diameter during total follow up after EVAR was 11 mm (IQR 6-17). At CTA, 16 patients (55%) had no detectable endoleak, five patients (17%) had suspected but uncertain endoleak, and eight patients had a definite endoleak (28%). On the post-contrast MRI images, endoleak was observed in 24 patients (83%). In all patients with uncertain endoleak on CTA, endoleak was detected with MRI. For type II endoleaks, feeding vessels were detected in 22/23 patients (96%) and these were all, except one, lumbar arteries. CONCLUSION: In patients with enlarging aneurysms of unknown origin after EVAR, MRI with a weak albumin binding contrast agent has additional value for both the detection and determination of the origin of the endoleak.

Quantitative and qualitative comparison of 0.025 mmol/kg gadobenate dimeglumine for abdominal MRI at 1.5T and 3T MRI in patients with low estimated glomerular filtration rate.

PURPOSE: To investigate the efficacy and adequacy of enhancement employing 0.025 mmol/kg of gadobenate dimeglumine at 1.5 Tesla (T), and to compare the extent of enhancement of this dosage between 1.5T and 3T systems. MATERIALS AND METHODS: Our final population included 116 consecutive patients who underwent 0.025 mmol/kg gadobenate dimeglumine-enhanced abdominal MRI (78 men and 38 women; age, 64.1 ± 13.6 years). Sixty patients underwent imaging at 1.5T and 56 patients underwent imaging at 3T. Abdominal enhancement was evaluated qualitatively and quantitatively. The quality of enhancement was compared using Mann-Whitney U test. The percentage of enhancement of each organ was compared using Student t-test. RESULTS: The mean quality rating of enhancement was at least "good" in all phases of enhancement for both 1.5T and 3T. There was a non-significant trend to higher mean ratings at 3T. The liver showed a 1.3-fold higher arterial-phase percentage of enhancement at 3T (p=0.0138). There were no differences between the mean relative enhancement of the pancreas and aorta throughout all phases of enhancement. The percentage of enhancement of the renal cortex was significantly higher at 3T (p<0.0001 to p=0.0293). CONCLUSION: A dose of 0.025 mmol/kg of gadobenate dimeglumine demonstrates diagnostic enhancement in the majority of patients at 1.5T, without significant differences on qualitative evaluation compared to 3T.

Impact of precontrast T10 relaxation times on dynamic contrast-enhanced MRI pharmacokinetic parameters: T10 mapping versus a fixed T10 reference value.

PURPOSE: To investigate variation in dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) pharmacokinetic parameter measurements between different methods of precontrast tissue relaxation (T10) estimation: pixel-based mapping versus a fixed reference value. MATERIALS AND METHODS: In 15 DCE-MRI studies the female pelvis, uterine fibroids, the left psoas muscle, and the fifth lumbar vertebral body were chosen to represent tissues with varying perfusion characteristics. All DCEMRI studies were processed using a variable flip angle T10 map and a fixed T10 reference value of 1000 msec. A subset of five DCE-MRI studies were each processed multiple times using the fixed T10 method with the reference T10 ranging from 0­2000 msec in 100-msec increments. Pharmacokinetic measurements of Ktrans, kep, ve, and initial area under the gadolinium curve (iAUGC) were performed maintaining the identical position for region of interest placement on each structure. RESULTS: The mean difference in pharmacokinetic output between the pixel-based T10 map and the fixed T10 reference value ranged from 6.6% for kep in the muscle to 54.9% for iAUGC in the vertebral body. At lower T10 (<1000 msec) aberrations in T10 estimation resulted in a larger error. Accurate measurement of T10 for each structure subsequently incorporated as a fixed T10 reference value yielded relative differences from 41.8% to 22.3% compared to the pixel-based T10 map. CONCLUSION: Direct comparison of pharmacokinetic parameters derived from a pixel-based approach versus a reference value uniformly applied to all pixels for T10 estimation is impeded by the inherent spatial heterogeneity of T10 within tissues.

Evidence of drug-drug interactions through uptake and efflux transport systems in rat hepatocytes: implications for cellular concentrations of competing drugs.

For drugs with hepatobiliary transport across hepatocytes, the interplay between uptake and efflux transporters determines hepatic concentrations of drugs, but the evolution over time of these concentrations is difficult to measure in humans other than with magnetic resonance imaging contrast agents in the liver. Gadobenate dimeglumine (BOPTA) is a contrast agent used in liver magnetic resonance imaging that enters into human hepatocytes through organic anion transporting polypeptides (OATP) and exits unchanged into bile through the multiple resistance-associated protein 2 (MRP2). Rifampicin (RIF) is transported by the same membrane proteins and may compete with BOPTA for hepatic uptake. Simultaneous drug-drug interactions through uptake and efflux transport systems in hepatocytes according to the cellular concentrations of competing drugs were never investigated. In perfused rat liver preparations, we demonstrate how the drug-drug interactions through transporters determine cellular concentrations of the competing drugs BOPTA and RIF, and we show that the cellular concentrations by modulating transport through membranes regulate the rat Oatp-Mrp2 interplay. Moreover, drug interactions through transporters change greatly over time.

Uptake and antileishmanial activity of meglumine antimoniate-containing liposomes in Leishmania (Leishmania) major-infected macrophages.

Leishmaniasis is a parasitic disease caused by the intramacrophage protozoa Leishmania spp. and may be fatal if left untreated. Although pentavalent antimonials are toxic and their mechanism of action is unclear, they remain the first-line drugs for treatment of leishmaniasis. An effective therapy could be achieved by delivering antileishmanial drugs to the site of infection. Compared with free drugs, antileishmanial agent-containing liposomes are more effective, less toxic and have fewer adverse side effects. The aim of this study was to develop novel meglumine antimoniate (MA)-containing liposome formulations and to analyse their antileishmanial activity and uptake by macrophages. Determination of the 50% inhibitory concentration (IC(50)) values showed that MA-containing liposomes were ≥10-fold more effective than the free drug, with a 5-fold increase in selectivity index, higher activity and reduced macrophage toxicity. The concentration required to kill 100% of intracellular amastigotes was ≥40-fold lower when MA was encapsulated in liposomes containing phosphatidylserine compared with the free drug. Fluorescence microscopy analysis revealed increased uptake of fluorescent liposomes in infected macrophages after short incubation times compared with non-infected macrophages. In conclusion, these data suggest that MA encapsulated in liposome formulations is more effective against Leishmania-infected macrophages than the non-liposomal drug. Development of liposome formulations is a valuable approach to the treatment of infectious diseases involving the mononuclear phagocyte system.

How organic anions accumulate in hepatocytes lacking Mrp2: evidence in rat liver.

In the liver, the accumulation of hepatobiliary contrast agents is a crucial issue to understand the images of liver scintigraphy or magnetic resonance (MR) imaging. Thus, depending on the regulation of uptake and exit membrane systems in normal and injured hepatocytes, these contrast agents will accumulate differently within cells. Gadobenate dimeglumine (Gd-BOPTA) is a hepatobiliary MR contrast agent that distributes to the extracellular space and enters into rat hepatocytes through the sinusoidal transporters, organic anion-transporting polypeptides. Gd-BOPTA is not metabolized during its transport to the canalicular membrane where it is excreted into bile through multiple resistance protein-2 (Mrp2). It is not well known how Gd-BOPTA accumulates in normal livers and in livers lacking Mrp2. We perfused livers from normal rats and from rats lacking Mrp2 with (153)Gd-BOPTA at increasing concentrations and assessed the hepatic accumulation of this agent using a gamma probe placed above the livers. By use of a pharmacokinetic model that best described the amounts of Gd-BOPTA in perfusate, bile, and hepatic tissue over time, we showed how increasing concentrations and the absence of Mrp2 modify the hepatic accumulation of the contrast agent. It is noteworthy that despite the absence of Gd-BOPTA bile excretion and a similar efflux back to sinusoids in livers lacking Mrp2, the maximal hepatic accumulation of contrast agent was similar to normal rats. We also showed how hepatic accumulation relies on the concomitant entry into and exit from hepatocytes. Such information improves our understanding of liver imaging associated with the perfusion of hepatobiliary contrast agents, which was recently introduced in clinical practice.

Solid hypervascular liver lesions: accurate identification of true benign lesions on enhanced dynamic and hepatobiliary phase magnetic resonance imaging after gadobenate dimeglumine administration.

PURPOSE: To evaluate hepatobiliary phase magnetic resonance imaging with gadobenate dimeglumine for differentiation of benign hypervascular liver lesions from malignant or high-risk lesions. METHODS AND MATERIALS: Retrospective assessment was performed of 550 patients with 910 hypervascular lesions (302 focal nodular hyperplasia [FNH], 82 nodular regenerative hyperplasia [NRH], 59 hepatic adenoma or liver adenomatosis [HA/LA], 329 hepatocellular carcinomas [HCC], 12 fibrolamellar-HCC [FL-HCC], 21 peripheral cholangiocarcinomas [PCC], 105 metastases). Imaging was performed before and during the arterial, portal-venous, equilibrium, and hepatobiliary phases after gadobenate dimeglumine administration (0.05 mmol/kg). Histologic confirmation was available for ≥1 lesion per patient, except for patients with suspected FNH (diagnosis based on characteristic enhancement/follow-up). Lesion differentiation (benign/malignant) on the basis of contrast washout and lesion enhancement (hypo-/iso-/hyperintensity) was assessed (sensitivity, specificity, accuracy, PPV, and NPV) relative to histology or final diagnosis. RESULTS: On portal-venous or equilibrium phase images, washout was not seen for 208 of 526 (39.5%) malignant (HCC, FL-HCC, PCC, metastases) and high-risk (HA/LA) lesions. Conversely, only 5 of 384 (1.3%) true benign lesions (FNH/NRH) showed washout. Taking washout as indicating malignancy, the sensitivity, specificity, and accuracy for malignant lesion identification during these phases was 61.8%, 98.7%, and 77.4%. On hepatobiliary phase images, 289 of 302 FNH, 82 of 82 NRH, 1 of 59 HA or LA, 62 of 341 HCC or FL-HCC, and 2 of 105 metastases were hyperintense or isointense. Taking iso- or hyperintensity as an indication for lesion benignity, the sensitivity, specificity, accuracy, PPV, and NPV for benign lesion identification was 96.6%, 87.6%, 91.4%, 85.1%, and 97.3%, respectively. CONCLUSIONS: Hepatobiliary phase imaging with gadobenate dimeglumine is accurate for distinguishing benign lesions from malignant or high-risk lesions. Biopsy should be considered for hypointense lesions on hepatobiliary phase images after gadobenate dimeglumine.

Dynamic changes in the TRPA1 selectivity filter lead to progressive but reversible pore dilation.

TRPA1 is a nonselective cation channel belonging to the transient receptor potential (TRP) family that is expressed in peripheral sensory neurons and may play important roles in pain perception and inflammation. We found that agonist stimulation of TRPA1, along with other members of the TRP family (TRPV1-4 and TRPM8), can induce the appearance of a large pore permeable to large organic cations such as Yo-Pro (YP) and N-methyl-d-glucamine, in an agonist and divalent cation-dependent manner. YP uptake was not inhibited by a panel of putative gap junction/pannexin blockers, suggesting that gap junction proteins are not required in this process. Our data suggest that changes in the TRP channel selectivity filter itself result in a progressive but reversible pore dilation process, a process that is under strong regulation by external calcium ions. Our data suggest that calcium plays a novel role in setting the amount of time TRPA1 channels spend in a dilated state providing a mechanism that may limit sensory neuron activation by painful or irritating substances.

Genotoxic effects of the antileishmanial drug Glucantime.

Leishmaniasis is caused by species of the protozoan parasite Leishmania. It is the third most important vector-borne disease and is widely distributed throughout the world. The World Health Organization recommends pentavalent antimonials as drugs of first choice in its treatment. Although Glucantime has traditionally been used to treat leishmaniasis, there are still many questions about its structure, mechanisms of action and ability to induce damage in DNA. In this study, the genotoxic activity of this drug was evaluated in vitro using human lymphocytes treated for 3 and 24 h (comet assay) and 48 h (apoptosis assay) with 3.25, 7.5 and 15 mg/ml of Glucantime, respectively, corresponding to 1.06, 2.12 and 4.25 mg/ml of pentavalent antimony. In the in vivo tests, Swiss mice received acute treatment with three doses (212.5, 425 and 850 mg/kg) of pentavalent antimony. All the treatments were administered intraperitoneally in the volumes of 0.1 ml/10 g of body weight, adapting human exposure to murine conditions. The animals were treated for 3 h in the comet assay using resident peritoneal exudate macrophages, for 24 h in the comet assay using peripheral blood leukocytes and for 24 h in the bone marrow erythrocyte micronucleus test. While no genotoxic effect was observed in the in vitro tests, the in vivo tests showed that Glucantime induces DNA damage. These findings indicate that Glucantime is a promutagenic compound that causes damage to DNA after reduction of pentavalent antimony (SbV) into the more toxic trivalent antimony (SbIII) in the antimonial drug meglumine antimoniate.

Antimony in plasma and skin of patients with cutaneous leishmaniasis--relationship with side effects after treatment with meglumine antimoniate.

OBJECTIVE: To evaluate the levels of antimony in plasma and skin of patients being treated with pentavalent antimonials (Glucantime) and their relationship with side effects. METHODS: We evaluated 19 patients treated endovenously at the conventional dose (20 mg Sb(v)/kg/day), two at a smaller dose (5 mg Sb(v)/kg/day) and three treated intralesionally (up to 4.0 ml/week). During treatment, patients underwent periodic blood exams and were interviewed weekly about the incidence of adverse symptoms. The levels of antimony in plasma and skin samples were determined by Inductively Coupled Plasma with Mass Spectrometry (ICP-MS). RESULTS: The patients under conventional treatment presented a mean initial antimony plasma concentration of 3.39 microg/l; at the end of treatment, these levels were 0.21 before Glucantime application and 125.8 mg after Glucantime application. The mean antimony level in their skin at the end of the treatment was 9.24 microg/g. The main adverse symptoms were arthralgia and myalgia; laboratory results showed mainly lymphocytosis and eosinophilia. CONCLUSIONS: We found some significant correlations between antimony concentrations, adverse symptoms and laboratory alterations, strengthening the hypothesis of a dose-dependent relationship between antimony concentration in plasma and skin and side effects.

Control of voltage-gated K+ channel permeability to NMDG+ by a residue at the outer pore.

Crystal structures of potassium (K(+)) channels reveal that the selectivity filter, the narrow portion of the pore, is only approximately 3-A wide and buttressed from behind, so that its ability to expand is highly constrained, and the permeation of molecules larger than Rb(+) (2.96 A in diameter) is prevented. N-methyl-d-glucamine (NMDG(+)), an organic monovalent cation, is thought to be a blocker of Kv channels, as it is much larger (approximately 7.3 A in mean diameter) than K(+) (2.66 A in diameter). However, in the absence of K(+), significant NMDG(+) currents could be recorded from human embryonic kidney cells expressing Kv3.1 or Kv3.2b channels and Kv1.5 R487Y/V, but not wild-type channels. Inward currents were much larger than outward currents due to the presence of intracellular Mg(2+) (1 mM), which blocked the outward NMDG(+) current, resulting in a strong inward rectification. The NMDG(+) current was inhibited by extracellular 4-aminopyridine (5 mM) or tetraethylammonium (10 mM), and largely eliminated in Kv3.2b by an S6 mutation that prevents the channel from opening (P468W) and by a pore helix mutation in Kv1.5 R487Y (W472F) that inactivates the channel at rest. These data indicate that NMDG(+) passes through the open ion-conducting pore and suggest a very flexible nature of the selectivity filter itself. 0.3 or 1 mM K(+) added to the external NMDG(+) solution positively shifted the reversal potential by approximately 16 or 31 mV, respectively, giving a permeability ratio for K(+) over NMDG(+) (P(K)(+)/P(NMDG)(+)) of approximately 240. Reversal potential shifts in mixtures of K(+) and NMDG(+) are in accordance with P(K)(+)/P(NMDG)(+), indicating that the ions compete for permeation and suggesting that NMDG(+) passes through the open state. Comparison of the outer pore regions of Kv3 and Kv1.5 channels identified an Arg residue in Kv1.5 that is replaced by a Tyr in Kv3 channels. Substituting R with Y or V allowed Kv1.5 channels to conduct NMDG(+), suggesting a regulation by this outer pore residue of Kv channel flexibility and, as a result, permeability.

Glibenclamide modulates glucantime activity and disposition in Leishmania major.

A source of chemotherapeutic failure in anti-infective therapies is the active movement of drugs across membranes, through ATP-binding cassette (ABC) transporters. In fact, simultaneous administration of therapeutic drugs with ABC transporter blockers has been invoked to be the way to actively prevent the emergence of drug resistance. Herein, we demonstrate that glucantime's efficacy in decreasing the infection rate of Leishmania-infected macrophages is strongly enhanced when used in combination with glibenclamide, a specific blocker of ABC transporters. Intracellular ABC transporters mediate glucantime sequestration in intracellular organelles. Their selective inhibition may effectively increase the cytoplasmic concentration of glucantime and its leishmanicidal activity. Our results reveal for the first time that glibenclamide targets in Leishmania major a compartment associated with a multivesicular system that is simultaneously labeled by the acidic marker LysoTracker-red and may represent the organelle where antimonials are sequestered. These results constitute a proof of concept that conclusively demonstrates the potential value that combination therapy with an ABC transporter blocker may have for leishmaniasis therapy.

The presence of halide salts influences the non-covalent interaction of MRI contrast agents and human serum albumin.

The rationale and objectives of the study were to evaluate the influence of the experimental conditions (buffer, salt, etc.) on the data characterizing the non-covalent interaction between MRI contrast agents and human serum albumin and hence their in vivo relaxivity. The interaction of three gadolinium contrast agents (Gd-EOB-DTPA, Gd-BOPTA and MP-2269) with human serum albumin was assessed through the measurement of proton relaxation rate enhancement in various experimental conditions. The data show the negative effect of halide salts on the paramagnetic relaxation enhancement of the three contrast agents. The presence of halide salts can thus have a negative effect on the efficacy of MRI contrast agents interacting with HSA. In addition, careful attention must be paid to comparisons of the binding parameters of various contrast agents reported in different studies since the composition of the medium can greatly influence the non-covalent interaction.