Jeffrey A. Creim

In Vitro And In Vivo Evaluation Of A Novel Ferrocyanide Functionalized Nanopourous Silica Decorporation Agent For Cesium In Rats

Novel decorporation agents are being developed to protect against radiological terrorist attacks. These sorbents, known as the self-assembled monolayer on mesoporous supports (SAMMS™), are hybrid materials where differing organic moieties are grafted onto mesoporous silica (SiO2). In vitro experiments focused on the evaluation and optimization of SAMMS for capturing radiocesium (137Cs); therefore, based on these studies, a ferrocyanide copper (FC-Cu-EDA)-SAMMS was advanced for in vivo evaluation. In vivo experiments were conducted comparing the performance of the SAMMS vs. insoluble Prussian blue. Groups of jugular cannulated rats (4/treatment) were evaluated. Animals in Group I were administered 137Cs chloride (∼40 &mgr;g kg−1) by intravenous (i.v.) injection or oral gavage; Group II animals were administered pre-bound 137Cs-SAMMS or sequential 137Cs chloride + SAMMS (∼61 ng kg−1) by oral gavage; and Group III was orally administered 137Cs chloride (∼61 ng kg−1) followed by either 0.1 g of SAMMS or Prussian blue. Following dosing, the rats were maintained in metabolism cages for 72 h and blood, urine, and fecal samples were collected for 137Cs analysis (gamma counting). Rats were then humanely euthanized, and selected tissues analyzed. Orally administered 137Cs chloride was rapidly and well absorbed (∼100% relative to i.v. dose), and the pharmacokinetics (blood, urine, feces, and tissues) were very comparable to the i.v. dose group. For both exposures the urine and feces accounted for 20 and 3% of the dose, respectively. The prebound 137Cs-SAMMS was retained primarily within the feces (72% of the dose), with ∼1.4% detected in the urine, suggesting that the 137Cs remained tightly bound to SAMMS. SAMMS and Prussian blue both effectively captured available 137Cs in the gut with feces accounting for 80–88% of the administered dose, while less than 2% was detected in the urine. This study suggests that the functionalized SAMMS outperforms Prussian blue in vitro at low pH, but demonstrates comparable in vivo sequestration efficacy at low exposure concentrations. The comparable response may be the result of the low 137Cs chloride dose and high sorbent dosage that was utilized. Future studies are planned to optimize the performance of SAMMS in vivo over a broader range of doses and conditions.

Comparison of cardiac and 60 Hz magnetically induced electric fields measured in anesthetized rats

Extremely low frequency magnetic fields interact with an animal by inducing internal electric fields, which are in addition to the normal endogenous fields present in living animals. Male rats weighing about 560 g each were anesthetized with ketamine and xylazine. Small incisions were made in the ventral body wall at the chest and upper abdomen to position a miniature probe for measuring internal electric fields. The calibration constant for the probe size was 5.7 mm, with a flat response from at least 12 Hz to 20 kHz. A cardiac signal, similar to the normal electrocardiogram with a heart rate of about 250 bpm, was readily obtained at the chest. Upon analysis of its spectrum, the cardiac field detected by the probe had a broad maximum at 32-95 Hz. When the rats were exposed to a 1 mT, 60 Hz magnetic field, a spike appeared in the spectrum at 60 Hz. The peak-to-peak magnitudes of electric fields associated with normal heart function were comparable to fields induced by a 1 mT magnetic field at 60 Hz for those positions measured on the body surface (where induced fields were maximal). Within the body, or in different directions relative to the applied field, the induced fields were reduced (reaching zero at the center of the animal). The cardiac field increased near the heart, becoming much larger than the induced field. Thus, the cardiac electric field, together with the other endogenous fields, combine with induced electric fields and help to provide reference levels for the induced-field dosimetry of ELF magnetic field exposures of living animals.

Aminothiol Receptors for Decorporation of Intravenously Administered 60Co in the Rat

This report provides a comparison of the oral decorporation efficacy of L-glutathione (GSH), L-cysteine (Cys), and a liposomal GSH formulation (ReadiSorb) toward systemic 60Co to that observed following intravenous administration of GSH and Cys in F344 rats. Aminoacid L-histidine (His) containing no thiol functionality was tested intravenously to compare in vivo efficacy of the aminothiol (GSH, Cys) chelators with that of the aminoimidazole (His) chelator. In these studies, 60Co was administered to animals by intravenous injection, followed by intravenous or oral gavage doses of a chelator repeated at 24-h intervals for a total of 5 doses. The results suggest that GSH and Cys are potent decorporation agents for 60Co in the rat model, although the efficacy of treatment depends largely on the systemic availability of the chelator. The intravenous route of administration of GSH or Cys was most effective in reducing tissue 60Co levels and in increasing excretion of radioactivity compared to control animals. Liposomal encapsulation was found to markedly enhance the oral bioavailability of GSH compared to non-formulated GSH. The oral administration of liposomal GSH reduced 60Co levels in nearly all tissues by 12–43% compared to that observed for non-formulated GSH. Efficacy of oral Cys was only slightly reduced in comparison with intravenous Cys. Further studies to optimize the dosing regimen in order to maximize decorporation efficiency are warranted.

Biomaterials For The Decorporation Of 85sr In The Rat

Although four stable isotopes of strontium occur naturally, 90Sr is produced by nuclear fission and is present in surface soil around the world as a result of fallout from atmospheric nuclear weapons tests. It can easily transfer to humans in the event of a nuclear/radiological emergency or through the plant-animal-human food chain causing long-term exposures. Strontium is chemically and biologically similar to calcium, and is incorporated primarily into bone following internal deposition. Alginic acid (alginate) obtained from seaweed (kelp) extract selectively binds ingested strontium in the gastrointestinal tract blocking its systemic uptake and reducing distribution to bone in rats, while other natural polysaccharides including chitosan and hyaluronic acid had little in vivo affinity for strontium. Alginate exhibits the unique ability to discriminate between strontium and calcium and has been previously shown to reduce intestinal absorption and skeletal retention of strontium without changing calcium metabolism. In our studies, the effect of commercially available alginate on intestinal absorption of strontium was examined. One problem associated with alginate treatment is its limited solubility and gel formation in water. The aqueous solubility of sodium alginate was improved in a sodium chloride/sodium bicarbonate electrolyte solution containing low molecular weight polyethylene glycol (PEG). Furthermore, oral administration of the combined alginate/electrolyte/PEG solution accelerated removal of internal strontium in rats when compared to treatment with individual sodium alginate/electrolyte or electrolyte/PEG solutions. Importantly, both alginate and PEG are nontoxic, readily available materials that can be easily administered orally in case of a national emergency when potentially large numbers of the population may require medical treatment for internal depositions. Our results suggest further studies to optimize in vivo decorporation performance of engineered alginate material via modification of its chemical and physicochemical properties are warranted.

Heating vs. cavitation in the induction of mouse hindlimb paralysis by ultrasound

Grip strength tests were performed on hairless mice before and after various ultrasound exposures in a temperature-controlled water bath at 37 degrees C. Lithotripter exposure of 800 shock waves produced no effect on hindlimb function. In contrast, 1.09-MHz exposures at 1 MPa with 10:100 ms burst mode did produce a statistically significant reduction in grip strength of about 60%. The exposure duration was important for the 1.0-MPa burst mode exposure, with grip-strength reductions appearing after 150 s or longer exposures. Continuous exposure at 3.3 W cm(-2) (0.32 MPa peak) for 200 s produced the same effect as burst mode exposure at 3.3 W cm(-2) (1 MPa peak) for 200 s, which implicates the temporal average intensity as an important factor. The temperature elevations for 1-MPa burst mode was estimated from thermocouple measurements in the spine to be 12 degrees C after 200-s exposure. Although tests of exposures in cool (32 degrees C) and warm (42 degrees C) baths produced inconclusive results in regard to the thermal mechanism, the effects observed appear to result from ultrasonic heating (rather than from cavitation). Thus, any potentially harmful consequences associated with the effects examined might be related more, for example, to ultrasonic hyperthermia therapy than to shock-wave lithotripsy.

EVALUATION OF CUPRIMINE® AND SYPRINE® FOR DECORPORATION OF 60Co AND 210Po

The acknowledged risk of deliberate release of radionuclides into local environments by terrorist activities has prompted a drive to improve novel materials and methods for removing internally deposited radionuclides. These decorporation treatments will also benefit workers in the nuclear industry, should an exposure occur. Cuprimine® and Syprine® are oral therapeutics based on the active ingredients D-penicillamine and N,N′-bis-(2-aminoethyl)-1,2-ethanediamine dihydrochloride, respectively. These therapeutic drugs have been used for several decades to treat Wilsons disease, a genetic defect leading to copper overload, by chelation and accelerated excretion of internally deposited copper. Studies were undertaken to evaluate these FDA-approved drugs for the in vivo decorporation of radioactive cobalt (60Co) and polonium (210Po) using male Wistar-Han rats. In these studies, 60Co or 210Po was administered to animals by IV injection, followed by oral gavage doses of either Cuprimine® or Syprine®. Control animals received the radionuclide alone. For 60Co studies, animals received a single dose of Cuprimine® or Syprine®, while for 210Po studies animals were repeatedly dosed at 24-h intervals for a total of 5 doses. Results show that Syprine® significantly increased urinary elimination and skeletal concentrations of 60Co compared to controls. While Cuprimine® had little effect on total excretion of 60Co, the skeletal, kidney, liver, muscle, and stomach tissues had significantly lower radioactivity compared to control animals. The low overall excretion of 210Po made it difficult to reliably measure urinary or fecal radioactivity and draw a definitive conclusion on the effect of Cuprimine® or Syprine® treatment on excretion. However, Cuprimine® treatment was effective at reducing spleen levels of 210Po compared to controls. Similarly, Syprine® treatment produced statistically significant reductions of 210Po in the spleen and skeletal tissues compared to control animals. Based on these promising findings, further studies to evaluate the dose-response pharmacokinetic profiles for decorporation are warranted.

Optical Spectroscopy and Multivariate Analysis for Biodosimetry and Monitoring of Radiation Injury to the Skin

Strategy, Management and Health Policy Enabling Technology, Genomics, Proteomics Preclinical Research Preclinical Development Toxicology, Formulation Drug Delivery, Pharmacokinetics Clinical Development Phases I‐III Regulatory, Quality, Manufacturing Postmarketing Phase IV

Detecting Radiation-Induced Injury Using Rapid 3D Variogram Analysis of CT Images of Rat Lungs

RATIONALE AND OBJECTIVES To investigate the ability of variogram analysis of octree-decomposed computed tomography (CT) images and volume change maps to detect radiation-induced damage in rat lungs. MATERIALS AND METHODS The lungs of female Sprague-Dawley rats were exposed to one of five absorbed doses (0, 6, 9, 12, or 15 Gy) of gamma radiation from a Co-60 source. At 6 months postexposure, pulmonary function tests were performed and four-dimensional (4D) CT images were acquired using a respiratory-gated microCT scanner. Volume change maps were then calculated from the 4DCT images. Octree decomposition was performed on CT images and volume change maps, and variogram analysis was applied to the decomposed images. Correlations of measured parameters with dose were evaluated. RESULTS The effects of irradiation were not detectable from measured parameters, indicating only mild lung damage. Additionally, there were no significant correlations of pulmonary function results or CT densitometry with radiation dose. However, the variogram analysis did detect a significant correlation with dose in both the CT images (r = -0.57, P = .003) and the volume change maps (r = -0.53, P = .008). CONCLUSION This is the first study to use variogram analysis of lung images to assess pulmonary damage in a model of radiation injury. Results show that this approach is more sensitive to detecting radiation damage than conventional measures such as pulmonary function tests or CT densitometry.

Evaluation of Cuprimine® and Syprine® for decorporation of radioisotopes of cesium, cobalt, iridium and strontium.

Cuprimine® and Syprine® are therapeutics approved by the USFDA to treat copper overload in Wilson Disease (a genetic defect in copper transport) by chelation and accelerated excretion of internally-deposited copper. These oral therapeutics are based on the respective active ingredients D-penicillamine (DPA) and N,N′-bis (2-aminoethyl) -1,2-ethanediamine dihydrochloride (Trien). Cuprimine is considered the primary treatment, although physicians are increasingly turning to Syprine as a first-line therapy. Both drugs exhibit oral systemic activity and low toxicity; their biological effects and safety are established. Previous in vivo studies using a rodent animal model established the decorporation potential of Cuprimine and Syprine for 60Co and 210Po. Currently these studies are being expanded to evaluate the in vivo decorporation efficacy of these drugs for several additional radionuclides. In this report, results of this investigation are discussed using the radionuclides 137Cs, 60Co, 192Ir and 85Sr. Short-term 48-h pilot studies were undertaken to evaluate DPA and Trien for their in vivo decorporation potential using male Wistar-Han rats. In these studies, a radionuclide solution was administered to the animals by intravenous (IV) injection, followed by a single IV dose of either DPA or Trien. Control animals received the radionuclide alone. Results show effective decorporation of 60Co by DPA within the time frame evaluated. DPA and Trien were also modestly effective in decorporation of 137Cs and 85Sr, respectively. The study did not find DPA or Trien effective for decorporation of 192Ir. Based on these encouraging findings, further studies to evaluate the dose-response profiles and timing of the chelator administration post exposure to radionuclides are warranted.

Investigation of Chitosan for Decorporation of 60Co in the Rat

With the increased threat of terrorist release of radioactive materials, there is a need for non-toxic decorporation agents to treat internal contamination with radionuclides. In this study, low molecular weight chitosan was evaluated for decorporation of radioactive cobalt (60Co). The affinity of chitosan for Co(II) was tested in vitro using spectrophotometric and potentiometric titration techniques. For in vivo studies, the effect of chitosan on ingested 60Co was evaluated using F344 rats administered a single dose followed by oral chitosan. Chitosan was also evaluated for systemic decorporation of 60Co following intravenous injection with repeated chitosan administration over 5 d. Control animals received 60Co without chelation treatment. Excreta and tissues were collected for analysis using gamma-counting techniques. Results from in vitro experiments confirmed the binding of Co(II) to chitosan, with the postulated formation of a mixed cobalt-chitosan-hydroxide complex species; a stability constant was calculated for this complex. For in vivo studies, oral administration of chitosan significantly reduced systemic absorption of orally administered 60Co as evidenced by an increase in fecal elimination and decrease in urinary elimination. However, oral administration of chitosan lactate slightly decreased fecal excretion of 60Co. Further, oral administration of chitosan significantly reduced 60Co levels in kidney, liver, and skeleton compared to control animals receiving 60Co alone. By the IV route, chitosan slightly reduced levels of 60Co in tissues compared to controls, although statistically significant reductions were only observed for blood and kidney. Overall, this commercially available chitosan oligosaccharide exhibited promising potential; further studies are warranted to evaluate the optimal dosing regimen and chemical modifications to increase effectiveness.