In vitro chondrocyte toxicity following long-term, high-dose exposure to Gd-DTPA and a novel cartilage-targeted MR contrast agent.

OBJECTIVE: To determine the concentrations exhibiting toxicity of a cartilage-targeted magnetic resonance imaging contrast agent compared with gadopentetate dimeglumine (Gd-DT-PA) in chondrocyte cultures. MATERIALS AND METHODS: A long-term Swarm rat chondrosarcoma chondrocyte-like cell line was exposed for 48 h to 1.0-20 mM concentrations of diaminobutyl-linked nitroxide (DAB4-DLN) citrate, 1.0-20 mM Gd-DTPA, 1.0 µM staurosporine (positive control), or left untreated. Cell appearance, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays of metabolic activity, quantitative PicoGreen assays of DNA content, and calcein-AM viability assays were compared. RESULTS: At 1.0-7.5 mM, minimal decrease in cell proliferation was found for both agents. At all doses of both agents, cell culture appearances were similar after 24 h of treatment. At the higher doses, differences in cell culture appearance were found after 48 h of treatment, with dose-dependent declines in chondrocyte populations for both agents. Concentration-dependent declines in DNA content and calcein fluorescence were found after 48 h of treatment, but beginning at a lower dose of DAB4-DLN citrate than Gd-DTPA. Dose-dependent decreases in MTT staining (cell metabolism) were apparent for both agents, but larger effects were evident at a lower dose for DAB-DLN citrate. Poor MTT staining of cells exposed for 48 h to 20 mM DAB4-DLN citrate probably indicates dead or dying cells. CONCLUSION: The minimal effect of the long-term exposure of model chondrocyte cell cultures to DAB4-DLN citrate and Gd-DTPA concentrations up to 7.5 mM (3x typical arthrographic administration) is supporting evidence that these doses are acceptable for MR arthrography. The findings are reassuring given that the experimental exposure to the contrast agents at sustained concentrations was much longer than when used clinically.

In vitro toxicity in long-term cell culture of MR contrast agents targeted to cartilage evaluation.

OBJECTIVE: Contrast-enhanced magnetic resonance (MR) imaging methods have been proposed for non-invasive evaluation of osteoarthritis (OA). We measured cell toxicities of cartilage-targeted low-generation dendrimer-linked nitroxide MR contrast agents and gadopentetate dimeglumine (Gd-DTPA) on cultured chondrocytes. DESIGN: A long-term Swarm rat chondrosarcoma chondrocyte-like cell line was exposed for 48-h to different salts (citrate, maleate, tartrate) and concentrations of generation one or two diaminobutyl-linked nitroxides (DAB4-DLN or DAB8-DLN), Gd-DTPA, or staurosporine (positive control). Impact on microscopic cell appearance, MTT spectrophotometric assays of metabolic activity, and quantitative PicoGreen assays of DNA content (cell proliferation) were measured and compared to untreated cultures. RESULTS: Chondrocyte cultures treated with up to 7.5 mM Gd-DTPA for 48-h had no statistical differences in DNA content or MTT reaction compared to untreated cultures. At all doses, DAB4-DLN citrate treated cultures had results similar to untreated and Gd-DTPA-treated cultures. At doses >1 mM, DAB4-DLN citrate treated cultures showed statistically greater DNA and MTT reaction than maleate and tartrate DAB4-DLN salts. Cultures exposed to 5 mM or 7.5 mM DAB8-DLN citrate exhibited rounded cells, poor cell proliferation, and barely detectable MTT reaction. Treatment with 0.1 µM staurosporine caused chondrocyte death. CONCLUSION: Long-term exposure, greater than clinically expected, to either DAB4-DLN citrate or Gd-DTPA had no detectable toxicity with results equivalent to untreated cultures. DAB4-DLN citrate was more biocompatible than either the maleate or tartrate salts. Cells exposed for 48-h to 5 mM or 7.5 mM DAB8-DLN salts demonstrated significant cell toxicity. Further evaluation of DAB8-DLN with clinically appropriate exposure times is required to determine the maximum useful concentration.

Nephrogenic systemic fibrosis in rats treated with erythropoietin and intravenous iron.

PURPOSE: To use a rat model for nephrogenic systemic fibrosis (NSF) that was administered high-dose gadodiamide to determine whether the co-administration of erythropoietin (Epo) and intravenous iron potentiated development of skin lesions that are thought to be a marker for the development of NSF. MATERIALS AND METHODS: The local committee for animal research approved this study. High-dose gadodiamide was administered, 2.5 mmol per kilogram of body weight for 20 days, or 500 times the U.S. Food and Drug Administration-approved dose, to four groups of Hannover-Wistar rats: group A, gadodiamide only; B, gadodiamide and Epo; C, gadodiamide and intravenous iron; and D, gadodiamide, Epo, and intravenous iron. The animals were sacrificed 7 days after final injection, and the authors examined dermal histologic findings from each animal and measured metal deposition by using inductively coupled plasma mass spectrometry. To compare the effect of metal deposition and cellularity, a linear mixed effects model was used to fit the data within PROC MIXED modeled with rat-specific random effects, and subsequently a Dunnett adjustment was performed. RESULTS: Rats treated with gadodiamide and both Epo and intravenous iron (group D) had significantly worse skin lesions at gross and histologic analysis (P = .004) compared with the rate treated with gadodiamide only (group A). Group D also had increased levels of deposited gadolinium as measured by means of mass spectrometry (P = .012). CONCLUSION: With a rat model similar to those already existing in the literature, skin changes were more marked in animals exposed to gadodiamide, Epo, and intravenous iron, as opposed to those animals exposed to gadodiamide alone; this experiment suggests that great caution may be warranted when prescribing gadolinium-based contrast agents to patients receiving Epo and intravenous iron.

Evaluating the role of zinc in the occurrence of fibrosis of the skin: a preclinical study.

PURPOSE: To investigate the possible role of Zn as a trigger for NSF we were using a previously established preclinical model. The depletion of endogenous Zinc ions (Zn) caused by the administration of gadolinium-based contrast agents (GBCAs) has been suggested as a possible pathomechanism for nephrogenic systemic fibrosis (NSF). MATERIALS AND METHODS: In the Zn supplementation study, rats were injected with Gadodiamide, Omniscan, and Magnevist with or without Zn supplementation. In the Zn depletion study, animals were kept on a Zn-deficient diet or a special control diet and received injections of Omniscan, OptiMARK, Magnevist, Gadovist, and Gd-EDTA. Gd, Zn, and Cu concentrations in tissue were measured and histology of the skin was performed. RESULTS: As seen in earlier studies, a difference in Gd concentration in the skin was observed following treatment with the different GBCAs. High Gd concentration in the skin correlated with the occurrence of NSF-like skin lesions. We observed no differences in the occurrence of skin lesions between the Zn supplementation and the Zn-deficient groups compared to their respective control groups. CONCLUSION: We found no significant effect of Zn on the initiation of NSF-like skin lesions. The results further support data from previous studies highlighting the importance of complex stability of the investigated GBCAs.

Preclinical safety evaluation of Gd-EOB-DTPA (Primovist).

OBJECTIVES: Gadoxetic acid [gadolinium-ethoxybenzyl-diethylenetriamine penta-acetic acid (Gd-EOB-DTPA); Primovist] is a liver specific contrast agent for magnetic resonance imaging. For risk assessment of the single diagnostic use the toxicity of this compound was assessed. MATERIALS AND METHODS: Studies into acute, repeated-dose, reproductive and developmental toxicity, and local tolerance, contact sensitizing, and genotoxic potential were performed. RESULTS: Lethality was observed after a single intravenous administration at doses 2 orders of magnitude higher than the clinical dose. The no observed adverse effect levels after repeated administration markedly exceeds the single diagnostic dose in humans and no unexpected organ toxicity was observed. No indications of reproductive and developmental toxicity, potential contact allergenic, and genotoxic effects were observed. Gd-EOB-DTPA was well tolerated after intravenous administration. CONCLUSIONS: Gd-EOB-DTPA was well tolerated with high safety margins between the single diagnostic dose and the doses showing adverse effects in animal studies.