Biomedical subjects
Publications and source records attributed to M A Morcillo.
The scientific basis for the treatment of the contamination of the human body by plutonium, americium and other actinides is reviewed. Guidance Notes are presented for the assistance of physicians and others who may be called upon to treat workers or members of the public who may become contaminated internally with inhaled plutonium nitrate, plutonium tributyl phosphate, americium nitrate or americium oxide.
Transgenic mice expressing IL-3 and IFN-alpha under the regulatory control of the GFAP gene promoter (GFAP-IL3 and GFAP-IFNalpha mice) exhibit a cytokine-specific, late-onset chronic-progressive neurological disorder which resemble many of the features of human diseases such as multiple sclerosis, Aicardi-Goutières syndrome, and some viral encephalopathies including HIV leukoencephalopathy. In this report we show that the metallothionein-I+II (MT-I+II) isoforms were upregulated in the brain of both GFAP-IL3 and GFAP-IFNalpha mice in accordance with the site and amount of expression of the cytokines. In the GFAP-IL3 mice, in situ hybridization analysis for MT-I RNA and radioimmunoassay results for MT-I+II protein revealed that a significant upregulation was observed in the cerebellum and medulla plus pons at the two ages studied, 1-3 and 6-10 months. Increased MT-I RNA levels occurred in the Purkinje and granular layers of the cerebellum, as well as in its white matter tracts. In contrast to the cerebellum and brain stem, MT-I+II were downregulated by IL-3 in the hippocampus and the remaining brain in the older mice. In situ hybridization for MT-III RNA revealed a modest increase in the cerebellum, which was confirmed by immunohistochemistry. MT-III immunoreactivity was present in cells that were mainly round or amoeboid monocytes/macrophages and in astrocytes. MT-I+II induction was more generalized in the GFAP-IFNalpha (GIFN12 and GIFN39 lines) mice, with significant increases in the cerebellum, thalamus, hippocampus, and cortex. In the high expressor line GIFN39, MT-III RNA levels were significantly increased in the cerebellum (Purkinje, granular, and molecular layers), thalamus, and hippocampus (CA2/CA3 and especially lacunosum molecular layers). Reactive astrocytes, activated rod-like microglia, and macrophages, but not the perivenular infiltrating cells, were identified as the cellular sources of the MT-I+II and MT-III proteins. The pattern of expression of the different MT isoforms in these transgenic mice differed substantially, demonstrating unique effects associated with the expression of each cytokine. The results indicate that the MT expression in the CNS is significantly affected by the cytokine-induced inflammatory response and support a major role of these proteins during CNS injury.
Several studies have recently shown that metallothionein (MT), a protein characterized by a high thiol content and that binds Zn2+ and Cu+, might be involved in the protection against oxidative stress and can act as a free radical scavenger. Oxidative stresses, such as irradiation, increase lipid peroxidation (LP) and subsequent tissue damage through free radical production. The induction of hepatic MT synthesis by gamma-irradiation (20 Gy) at 8, 24, 30 and 48 hrs. post-irradiation in two different age groups of Sprague-Dawley rats (39-40 and 48-49 days old) was studied. LP measured by the thiobarbituric acid reactive substances (TBARS) assay and Cu and Zn levels in liver have also been determined. In the younger group, the gamma-irradiation induced hepatic MT synthesis and increased LP that peaked 24 hrs. after irradiation. During the first 30 hrs. post-irradiation, a positive and statistically significant correlation between hepatic MT content and LP level in liver was found. In the older group, liver MT synthesis was only increased 1.7-fold and LP levels were not altered at 24 hrs. post-irradiation compared with sham-irradiated rats. Therefore it appears that LP is not necessary for induction of MT synthesis by gamma-irradiation.
The effects of long-term daily intake of low and high levels of mercury on its organ distribution and binding to renal metallothionein (MT) in male rats were studied. The animals were exposed to mercuric chloride labelled with 203Hg via drinking water for 8 weeks (5, 50 and 500 microM Hg). The greatest concentration of mercury was found in the kidneys. Similar levels of radioactivity in the buccal cavity and oesophagus were also observed by whole-body autoradiography. In the kidneys, the mercury was accumulated in the outer stripe of the outer zone of the medulla and, to a minor degree, in the renal cortex. Almost 50% the total renal mercury was associated to MT. The binding capacity of the renal MT for mercury tends to saturate with increasing doses, thus this means that the capacity of the kidneys to accumulate mercury is limited.
The effects of long-term daily intake of mercury on its urinary and fecal excretion, whole-body retention, and blood concentration in male rats were observed. The animals were exposed to mercuric, chloride labeled with 203Hg via drinking water for 8 weeks (5, 50 and 500 microM Hg). 203Hg in urine, feces and blood was quantified. The blood mercury concentration did not keep a linear relationship with the increasing dose. The percentage of the total amount of mercury intake which is excreted by the fecal route in rats exposed to 500 microM Hg was significantly lower than in those exposed to 5 and 50 microM. The daily dose percentage of mercury excreted in urine increased with dose size. The results show that the absorption fraction of mercury through the gastrointestinal tract (30-40%) was higher than values previously reported.
High-performance liquid chromatography (HPLC) was applied to the separation of metallothionein (MT) isoforms from different tissues from a variety of eukaryotic species. Recently we reported an analytical method for 203Hg-metallothionein, which detects the radioisotope bound to each iso-MT after separation by HPLC on a size-exclusion column coupled with on-line radioactivity flow detection. The MTs can be separated as distinct isoprotein peaks by elution with alkaline buffer solution owing to cation-exchange chromatographic action. In the present work, renal MT from rats exposed to inorganic mercury was separated into four peaks by UV and 203Hg detection. Moreover, it was resolved into four components by non-denaturing polyacrylamide gel electrophoresis. The two major components correspond to MT-1 and MT-2, which were characterized by amino acid analysis. Finally, Hg induces and binds to both iso-MTs.
Rat kidney 203Hg-induced metallothionein (HgMT) was separated on a high performance liquid chromatograph equipped with a gel permeation column and an on-line beta radioactivity detector, in order to obtain the simultaneous measurements of renal MT by UV detection and MT-associated 203Hg by a beta radioactivity detector. Metallothionein was separated in three major species by both UV detection at 254 nm and 203Hg detection, probably due to the presence of mercury and copper. A standard curve was prepared which demonstrated excellent linear correlation between the integrated HgMT peaks area and the quantity of HgMT injected into the column. In contrast to the results with the gel permeation column above mentioned, rat kidney HgMT was separated in four peaks by reversed-phase height performance liquid chromatography.
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