Differential Phagocytosis-Based Photothermal Ablation of Inflammatory Macrophages in Atherosclerotic Disease.

Inflammatory macrophage (Mφ)-mediated atherosclerosis is a leading cause of mortality and morbidity worldwide. Photothermal therapy (PTT) has been demonstrated as an efficient strategy in killing target cells, and its application in the treatment of inflammation in atherosclerosis is developing. However, the choice of nanomaterials, mechanisms, and side effects are seldom considered. In this study, semiconductor nanomaterials, that is, MoO2 nanoclusters, were synthesized and used for the first time in PTT for inflammatory Mφ-mediated atherosclerosis. Based on cell differential phagocytosis, the optimum amount of MoO2 and treatment time were selected to exert the maximum ablation effect on Mφ and minimal damage on endothelial cells without requiring additional target or selective groups. Moreover, MoO2-based PTT shows an excellent therapeutic effect on atherosclerosis by eliminating Mφ in animal models, with no significant side effects observed. This study explores a new method of nanotechnology and pharmaceutical development by using and optimizing cost-effective metal oxide nanostructures in the treatment of atherosclerosis and motivates further research on minimizing the side effects of related materials.

Degradable Molybdenum Oxide Nanosheets with Rapid Clearance and Efficient Tumor Homing Capabilities as a Therapeutic Nanoplatform.

Molybdenum oxide (MoOx) nanosheets with high near-infrared (NIR) absorbance and pH-dependent oxidative degradation properties were synthesized, functionalized with polyethylene glycol (PEG), and then used as a degradable photothermal agent and drug carrier. The nanosheets, which are relatively stable under acidic pH, could be degraded at physiological pH. Therefore, MoOx-PEG distributed in organs upon intravenous injection would be rapidly degraded and excreted without apparent in vivo toxicity. MoOx-PEG shows efficient accumulation in tumors, the acidic pH of which then leads to longer tumor retention of those nanosheets. Along with the capability of acting as a photothermal agent for effective tumor ablation, MoOx-PEG can load therapeutic molecules with high efficiencies. This concept of inorganic theranostic nanoagent should be relatively stable in tumors to allow imaging and treatment, while being readily degradable in normal organs to enable rapid excretion and avoid long-term retention/toxicity.

Smart MoS2/Fe3O4 Nanotheranostic for Magnetically Targeted Photothermal Therapy Guided by Magnetic Resonance/Photoacoustic Imaging.

The ability to selectively destroy cancer cells while sparing normal tissue is highly desirable during the cancer therapy. Here, magnetic targeted photothermal therapy was demonstrated by the integration of MoS2 (MS) flakes and Fe3O4 (IO) nanoparticles (NPs), where MoS2 converted near-infrared (NIR) light into heat and Fe3O4 NPs served as target moiety directed by external magnetic field to tumor site. The MoS2/Fe3O4 composite (MSIOs) functionalized by biocompatible polyethylene glycol (PEG) were prepared by a simple two-step hydrothermal method. And the as-obtained MSIOs exhibit high stability in bio-fluids and low toxicity in vitro and in vivo. Specifically, the MSIOs can be applied as a dual-modal probe for T2-weighted magnetic resonance (MR) and photoacoustic tomography (PAT) imaging due to their superparamagnetic property and strong NIR absorption. Furthermore, we demonstrate an effective result for magnetically targeted photothermal ablation of cancer. All these results show a great potential for localized photothermal ablation of cancer spatially/timely guided by the magnetic field and indicated the promise of the multifunctional MSIOs for applications in cancer theranostics.

90-Day subchronic toxicity study of sodium molybdate dihydrate in rats.

This study investigated the subchronic toxicity of molybdenum (Mo) in Sprague-Dawley rats given sodium molybdate dihydrate in the diet for 90days at dose levels of 0, 5, 17 or 60mgMo/kgbw/day. The study complied with OECD Test Guideline (TG) 408, with additional examination of estrus cycles and sperm count, motility, and morphology from OECD TG 416. The overall no-observed-adverse-effect level was 17mgMo/kgbw/day, based on effects on body weight, body weight gain, food conversion efficiency and renal histopathology (females only) at 60mgMo/kgbw/day. No treatment-related adverse effects on reproductive organ weights or histopathology, estrus cycles or sperm parameters were observed at any dose level. No adverse effects were observed in the high dose animals after the 60-day recovery period, with the exception that male rats did not fully recover from reduced body weight. Serum blood, liver and kidney samples were analyzed for molybdenum, copper, zinc, manganese, iron, cobalt and selenium; high levels of molybdenum and copper were found in the serum, blood, liver and kidneys of rats treated with 60mgMo/kgbw/day. In conclusion, the LOAEL and NOAEL for molybdenum were determined to be 60 and 17mgMo/kgbw/day, respectively.

Phase I study of copper-binding agent ATN-224 in patients with advanced solid tumors.

PURPOSE: Copper chelation reduces the secretion of many angiogenic factors and reduces tumor growth and microvascular density in animal models. ATN-224 is a second-generation analogue of ammonium tetrathiomolybdate. The aim of our phase I study was to reduce serum copper levels, as measured by ceruloplasmin, to 5 to 15 mg/dL (normal 16-60) in 14 to 21 days, to determine the pharmacokinetic profile of ATN-224 and to evaluate dose-limiting toxicities. PATIENTS AND METHODS: Cohorts of patients were treated with escalating oral doses of ATN-224 until copper depletion followed by a titrated maintenance dose. RESULTS: Eighteen patients received 78 cycles of ATN-224. Mean baseline ceruloplasmin was 39.6 mg/dL. The maximum administered dose was 330 mg/d where grade 3 fatigue was dose-limiting. At the maximum tolerated dose of 300 mg/d, the median time to achieve target ceruloplasmin was 21 days, and toxicities included grade 3 anemia, grade 3 neutropenia, fatigue, and sulfur eructation. ATN-224 treatment caused a significant reduction (> 90%) in RBC superoxide dismutase 1 activity and circulating endothelial cells. Pharmacokinetic data indicate greater absorption of ATN-224 and more rapid ceruloplasmin reduction when administered with a proton pump inhibitor. Stable disease of > 6 months was observed in 2 patients. CONCLUSIONS: Oral ATN-224 is a well-tolerated therapy and at a loading dose of 300 mg/d leads to a reduction of serum ceruloplasmin levels in 80% patients within 21 days. A loading dose of 300 mg/d for 2 weeks followed by a titrated maintenance dose will be the recommended starting dose for phase II study.

Tetrathiomolybdate, a copper chelator for the treatment of Wilson disease, pulmonary fibrosis and other indications.

Tetrathiomolybdate (TTM) is a copper chelator that has also demonstrated antiangiogenic, antifibrogenic and anti-inflammatory actions in preclinical studies. The drug, from the University of Michigan was licensed to Pipex Pharmaceuticals Inc for development for several indications; development of the drug for cancer was later licensed to Attenuon LLC. In a phase III clinical trial, TTM stabilized neurological function in patients with Wilson disease, causing significant recovery in 81% of patients at 3 years post initiation of therapy; a second phase III trial was ongoing at the time of publication. A phase I/II clinical trial demonstrated the efficacy of TTM in patients with idiopathic pulmonary fibrosis, and led the FDA to grant TTM Orphan Drug status for this disease. Several phase II clinical trials had also been completed in patients with various cancers, and revealed mixed efficacy. TTM was also assessed in a phase I clinical trial for age-related macular degeneration, but the results reported from the trial were negative; no further development has occurred for this indication. TTM was assessed for the treatment of psoriasis in a phase II clinical trial, but no data have been reported. At the time of publication, phase II and phase III clinical trials were ongoing in patients with Alzheimer's disease and primary biliary cirrhosis, respectively. The most common clinical side effects observed for TTM over the range of indications have been anemia, neutropenia, leukopenia and transaminase elevations. These side effects were generally resolved with either a dose adjustment or temporary suspension of the dosing regimen. TTM is predicted to most likely find a niche in the therapy of Wilson disease, for which current treatment options are limited.

Modest copper supplementation blocks molybdenosis in cattle.

It is widely accepted that ratios of dietary copper (Cu) to molybdenum (Mo) lower than 10:1 may produce molybdenosis in cattle, especially if sulfur concentrations are more than 3,000 ppm. Some authorities suggest that dietary Mo concentrations greater than 10 ppm are hazardous to cattle regardless of Cu concentration, but anecdotal reports suggest that this may not be the case. The original purpose of the experiment described in this report was to investigate whether supranutritional supplemental Cu could protect cattle against relatively high dietary Mo. Pregnant cows were grazed on 1 of 3 pastures: 1 with only background Mo, 1 with an average of 13 ppm Mo, and 1 that averaged 230 ppm Mo. Half the cows on the Mo pastures were supplemented with 17 ppm dietary Cu, the other half with the dietary supplement plus Cu boluses. Molybdenum effects were anticipated in the groups supplemented with 17 ppm Cu; however, despite increased tissue concentrations of Mo, only the 230 ppm Mo/17 ppm Cu group exhibited any effects. Moderate Cu supplementation permitted cows to graze a site heavily contaminated with Mo with no adverse effects on general health or reproduction.

A modified risk assessment to establish molybdenum standards for land application of biosolids.

The USEPA standards (40 CFR Part 503) for the use or disposal of sewage sludge (biosolids) derived risk-based numerical values for Mo for the biosolids --> land --> plant --> animal pathway (Pathway 6). Following legal challenge, most Mo numerical standards were withdrawn, pending additional field-generated data using modern biosolids (Mo concentrations <75 mg kg(-1) and a reassessment of this pathway. This paper presents a reevaluation of biosolids Mo data, refinement of the risk assessment algorithms, and a reassessment of Mo-induced hypocuprosis from land application of biosolids. Forage Mo uptake coefficients (UC) are derived from field studies, many of which used modern biosolids applied to numerous soil types, with varying soil pH values, and supporting various crops. Typical cattle diet scenarios are used to calculate a diet-weighted UC value that realistically represents forage Mo exposure to cattle. Recent biosolids use data are employed to estimate the fraction of animal forage (FC) likely to be affected by biosolids applications nationally. Field data are used to estimate long-term Mo leaching and a leaching correction factor (LC) is used to adjust cumulative biosolids application limits. The modified UC and new FC and LC factors are used in a new algorithm to calculate biosolids Mo Pathway 6 risk. The resulting numerical standards for Mo are cumulative limit (RPc)=40 kg Mo ha(-1), and alternate pollutant limit (APL) = 40 mg Mo kg(-1) We regard the modifications to algorithms and parameters and calculations as conservative, and believe that the risk of Mo-induced hypocuprosis from biosolids Mo is small. Providing adequate Cu mineral supplements, standard procedure in proper herd management, would augment the conservatism of the new risk assessment.

Bioavailability of biosolids molybdenum to soybean grain.

Legumes grown in biosolids-amended soils and then fed to ruminants can represent problematic sources of molybdenum (Mo), but few field data are available to quantify the risk. We used a set of fields amended to high cumulative biosolids Mo loads (>18 kg ha(-1)) over 27 yr to generate additional data. Soybean [Glycine max (L.) Merr.] was grown on 29 fields (pH values>6.8) amended to a wide range of soil Mo loads. Soybean grain harvested from each field was analyzed for Mo and the concentrations regressed against soil Mo loads estimated from actual soil Mo concentrations in the 0- to 15-cm depth. Slopes of such linear regressions represent uptake coefficients (UC values) used by the USEPA to assess risk of biosolids Mo to ruminants fed forage grown on biosolids-amended land. The UC value for all 29 fields was estimated as 1.66, which agrees with the few soybean grain data in the literature. The UC value, however, is well below a conservative UC value of 4, recently recommended for all fresh legume materials fed to cattle. Soybean grain can contain high concentrations of Mo (>10 mg kg(-1)) and have low (<2:1) Cu to Mo ratios, which can exacerbate molybdenosis problems in cattle. However, soybean grain normally constitutes only -10% of dairy cattle diet, and other constituents (e.g., corn grain, stover, mineral supplements) are sufficient, or can be manipulated, to control molybdenosis.

Excretion of copper complexed with thiomolybdate into the bile and blood in LEC rats.

Copper (Cu) accumulating in a form bound to metallothionein (MT) in the liver of Long-Evans rats with a cinnamon-like coat color (LEC rats), an animal model of Wilson disease, was removed with ammonium tetrathiomolybdate (TTM), and the fate of the Cu complexed with TTM and mobilized from the liver was determined. TTM was injected intravenously as a single dose of 2, 10 or 50 mg TTM/kg body weight into LEC and Wistar (normal Cu metabolism) rats, and then the concentrations of Cu and molybdenum (Mo) in the bile and plasma were monitored with time after the injection. In Wistar rats, most of the Mo was excreted into the urine, only a small quantity being excreted into the bile, while Cu excreted into the urine decreased. However, in LEC rats, Cu and Mo were excreted into the bile and blood, and the bile is recognized for the first time as the major route of excretion. The Cu excreted into both the bile and plasma was accompanied by an equimolar amount of Mo. The relative ratio of the amounts of Cu excreted into the bile and plasma was 40/60 for the low and high dose groups, and 70/30 for the medium dose group. The systemic dispositions of the Cu mobilized from the liver and the Mo complexed with the Cu were also determined for the kidneys, spleen and brain together with their urinal excretion. Although Mo in the three organs and Cu in the kidneys and spleen were increased or showed a tendency to increase, Cu in the brain was not increased at all doses of TTM.

Molybdenum and copper kinetics after tetrathiomolybdate injection in LEC rats: specific role of serum albumin.

Chelation therapy with tetrathiomolybdate (TTM) was applied to Long-Evans rats with a cinnamon coat-color (LEC rats), an animal model for Wilson disease, to remove copper (Cu) accumulated in the liver in a form bound to metallothionein (MT). Changes in molybdenum (Mo) and Cu concentrations and their biological forms in serum of LEC rats determined at different times after a single intraperitoneal injection were compared with those of Wistar (normal) rats. The change in Mo concentration in serum of normal rats was mono-phasic, whereas in LEC rats it was bi-phasic. The phase in normal rats and the first phase in LEC rats appeared to reflect the process of uptake and disappearance of TTM in the livers of Wistar and LEC rats. On the other hand, the second phase in LEC rats paralleled the changes of Cu and appeared to reflect the complex formation (Cu/thiomolybdate complex) between Mo and Cu accumulated in the liver. The complex was specifically bound to albumin as determined by high performance liquid chromatography with inductively coupled plasma-mass spectrometry (HPLC/ICP-MS). The results suggested that the changes in the Mo concentration in serum reflected the amount of Cu in the liver.

Accumulation of trace elements in sheep and the effects upon qualitative and quantitative ovarian changes.

The distribution of Cu, Zn, Fe, Mo, Se, As, Cd and Pb was determined in the organs of sheep that died of Cu intoxication from a Cu plants emissions. Simultaneously, quantitative and qualitative changes in the ovaries were evaluated. In sheep that died of copper intoxication, the highest Cu levels were in the liver (1797 +/- 946.1 mg/kg dry matter) and the kidneys (425.5 +/- 426.4 mg/kg dry matter). The liver and kidneys accumulated Zn, whereas Fe was primarily accumulated by spleen and kidneys. Selenium was mainly in the ovaries and uterus (7.72 +/- 4.88 and 5.30 +/- 5.33 mg/kg dry matter respectively). The ovaries and the uterus were the organs that accumulated As, Cd and Pb, respectively. Histological changes in the number of ovarian follicles and the increased occurrence of primary atretic follicles indicated alterations in the membrane structures and organelles of oocytes and in the follicular cells of the stratum granulosum.

Molybdenum absorption, excretion, and retention studied with stable isotopes in young men during depletion and repletion.

A study of molybdenum absorption, excretion, and balance was conducted in four young men fed a low-molybdenum diet (22 micrograms/d) for 102 d followed by 18 d of the same diet supplemented to contain 467 micrograms/d. The study was conducted to determine the minimum dietary molybdenum requirement of healthy young men. Stable isotopes of molybdenum were used as tracers. 100Mo was fed four times during the study, 97Mo was infused twice, and 94Mo was used as an isotopic diluent to quantify the molybdenum isotopes and total molybdenum in complete urine and fecal collections and in the diets. The study demonstrated that subjects could not consistently attain balance with the low-molybdenum diet, but balance improved with time, and no signs of molybdenum deficiency were observed. Molybdenum was very efficiently absorbed at both intakes of dietary molybdenum and urinary excretion increased as dietary molybdenum increased. Molybdenum turnover was significantly slower when dietary molybdenum was low. We estimate from these results that the minimum dietary molybdenum requirement is approximately 25 micrograms/d or possibly less. This suggests that the lower end of the recommended range could be less than the current recommended amount of 75 micrograms/d.

Molybdenum requirement of female rats.

Sixty 3-wk-old female Sprague-Dawley rats were randomly divided into six groups of 10 animals each. Group 1 was fed for 8 wk purified AIN-76A diet (basal diet) containing 0.025 mg molybdenum/kg diet. Groups 2-6 were fed the same basal diet supplemented with sodium molybdate to provide total dietary Mo of 0.050, 0.100, 0.200, 0.400 and 0.800 mg/kg diet, respectively. Molybdenum concentration in liver and brain increased linearly up to the 0.200 mg Mo/kg diet level. Beyond this level, no further significant increase occurred. Dietary Mo of 0.100 mg/kg elevated the Mo concentration in heart to its maximal level. Supplementation with 0.025 mg Mo/kg to a total of 0.05 mg Mo/kg diet significantly increased Mo concentration in spleen and kidney; higher levels of dietary Mo did not result in further significant responses. Molybdenum supplementation significantly increased the activities of xanthine dehydrogenase/oxidase (XDH), sulfite oxidase and superoxide dismutase in the liver, and of XDH in small intestinal mucosa. Maximal activities were attained at 0.050, 0.050, 0.200 and 0.100 mg Mo/kg diet, respectively. Dietary Mo of 0.200 mg/kg diet was estimated as the Mo requirement of rats fed the AIN-76A diet.

Hepatic, placental, and fetal trace elements following molybdenum supplementation during gestation.

The effect of dietary Mo (Na2Mo(4)2H2O) added to drinking water at levels of 0, 5, 10, 50, or 100 mg on hepatic (gestating dams), placental, and fetal Mo, Cu, Zn, and Fe contents of Sprague-Dawley rats was studied. These elements were determined by a polarographic catalytic procedure for Mo and by atomic absorption spectrophotometry for Cu, Fe, and Zn. Hepatic Mo increased two to sixfold (5-100 mg Mo). There was a 1.5-fold increase in hepatic Cu, significant only at the 50 to 100 mg Mo/L treatment levels. Although the hepatic Fe content of the gestating rats significantly increased with Mo supplementation, the extent of the increase appeared to be influenced by the litter size, fetal weights, and the degree of fetal resorption. Zinc values did not differ at any of the treatment levels. Placental Mo increased 3-76-fold, Cu one to threefold. No differences were observed in placenta Fe or Zn. Fetal Mo increased two to six-fold (10-100 mg/L) and Cu increased one to fivefold. There were no differences in the Fe and Zn content although both of these elements appeared to decline as the level of supplemental Mo increased. Significant correlations were also observed between hepatic, placental, and fetal Mo, Cu, Fe, and Zn. These results suggest that changes in trace mineral status in gestation, owing to high Mo intake, do occur and such occurrences are also reflected in the fetus.

The effects of ammonium tetrathiomolybdate intake on tissue copper and molybdenum in pregnant ewes and lambs.

Pregnant ewes (d 32 of gestation) were allocated to three treatments and given intraruminal controlled-release devices designed to deliver 0, 20 or 60 mg diammonium tetrathiomolybdate (TTM) per day. Ewes given 20 or 60 mg TTM/d also received an oral drench of 120 or 360 mg TTM twice weekly commencing on d 86 of gestation. Liver and kidney samples were taken from lambs 48 h after birth and from ewes on d 18 postpartum. Trichloroacetic acid soluble Cu, ceruloplasmin and superoxide dismutase activities in the plasma of ewes were decreased (P less than .05) by TTM. Liver Cu concentrations were decreased (P less than .05), but kidney Cu concentrations increased (P less than .05) by 16-fold in ewes given the higher dose of TTM. Liver and kidney Mo concentrations were elevated (P less than .05) 9- and 30-fold, respectively, in ewes given TTM. Plasma glucose concentrations in ewes were decreased (P less than .05) by the highest level of TTM treatment. Lambs of ewes given TTM had a fivefold increase (P less than .05) in liver Mo concentration, but kidney Mo concentration was not affected (P greater than .05) and liver Cu concentration was reduced (P less than .05). In ewes, Mo apparently caused Cu to be mobilized from the liver and a Cu and Mo complex accumulated in the kidney. Some Mo crossed the placenta, but only limited Mo accumulated in the fetal livers. When given to pregnant ewes, TTM reduced liver Cu levels in the lambs but did not affect the concentration of Cu in colostrum.