Metallothionein synthesis induced by interferon alpha/beta in mice of various zinc status.

We studied the ability of interferon alpha/beta (IFN) to induce metallothionein (MT) synthesis in mice. Male mice were injected intraperitoneally with mouse IFN (5 x 10(5) IU/mouse). Plasma Zn levels were reduced at 4 hr after injection, reached a minimum value at 6 hr, and then returned to the control level at 8 hr. Hepatic MT concentrations began to increase at 4 hr and reached maximum values at 6 hr. Induction of MT gene expression and protein synthesis was confirmed by Northern blot analysis and radioimmunoassay, respectively. The induction of MT synthesis in the liver by IFN was dose-dependent. The data suggest that induction of MT-mRNA and the protein in the liver by IFN occurs rapidly but is rather transient. Furthermore, MT synthesis was not induced by IFN in the liver of mice given a Zn-deficient diet, whereas IFN induced increases in the activity of 2',5'-oligoadenylate synthetase in the spleen were unaffected by Zn status. Thus, induction of hepatic MT synthesis by IFN is influenced by Zn status.

Synergistic induction of metallothionein synthesis by interleukin-6, dexamethasone and zinc in the rat.

We investigated the reciprocal effects of interleukin-6 (IL-6), glucocorticoid and zinc (Zn) on metallothionein (MT) synthesis in rats. MT synthesis in the liver, which is a key responsible organ in acute phase responses, was induced by IL-6 or dexamethasone (Dex), and in an additive manner by a combination of IL-6 and Dex 18 h after injection. MT synthesis in the lung and heart was evaluated by immunoassay using a specific antibody to MT-1, because of its low concentration in these tissues. Heart concentrations of MT-1 were significantly increased by IL-6, and were further increased by the combination of IL-6 and Dex, although Dex by itself had no effect. This suggests a synergistic effect of IL-6 and Dex on MT-I synthesis in the heart. A similar synergism was observed in the lung. To study the effect of Zn on the induction of MT and acute phase proteins, Zn, IL-6 and Dex were administered in various concentrations. The increase in liver MT induced by the combination of IL-6 and Dex with Zn (130 micrograms MT/g of liver) was greater than the sum of the increases induced by (IL-6 + Zn) and by (Dex + Zn) (103 micrograms MT/g), suggesting a synergistic increase. The data indicate that the maximal increase in the induction of MT by a combination of IL-6 and Dex depends on an adequate liver Zn content. Thus, the in vivo synergistic induction of acute phase proteins by IL-6, glucocorticoid and Zn may be required for the maximal and rapid response, not only in liver but also in other tissues including heart and lung. This suggests that the synergistic reaction may be important for an enhancement of the radical scavenging ability of tissues in acute phase responses.

Susceptibility of metallothionein-null mice to paraquat.

Using transgenic mice in which metallothionein (MT)-I and MT-II genes, we have studied a putative role of MT as a free radical scavenger against paraquat, a free radical generator. Male mice were injected s.c. with paraquat (PQ) at a single dose of 40 or 60 mg/kg of body weight (b.w.). Two of the six MT-null mice died within 16 h at the dose of 60 mg PQ/kg. b. w. PQ administration increased hepatic MT concentration in the normal mice (C57BL/6J), but not in the MT-null mice. The lipid peroxidation (LP) determined by thiobarbituric acid-reactive substance formation was increased by PQ in the liver of normal and MT-null mice, and the enhanced level was greater in the MT-null mice than in the C57BL/6J mice. Administration of PQ significantly increased blood urea nitrogen only in the MT-null mice, indicating renal damage. Without paraquat administration, the hepatic concentration of non-protein sulphydryl compounds was less in the MT-null mice than in the C57BL/6J mice, and the basal level of LP was higher in the MT-null mice than in the C57BL/6J mice. The present results support the notion that MT plays an antioxidative role against paraquat insult under physiological conditions.

[Clinicophysiological study of multimodality evoked potentials and computed tomographic findings in persistent vegetative state].

The auditory brainstem response (ABR), short latency somatosensory evoked potential (SSEP) and visual evoked potential (VEP) of patients in the persistent vegetative state (PVS) are reported, and the correlations between the electrophysiological findings and the CT scan findings with the three clinical grades of the PVS (transitional, incomplete and complete vegetative syndromes) are discussed. Twenty two patients in a vegetative state caused by subarachnoid hemorrhage (3), hypertensive intracerebral hemorrhage (5), cerebral infarction (6), head injury (3), cerebral anoxia (4) and brain tumor (1). Each evoked response was evaluated for the presence or absence of abnormalities and assigned a grade ranked I to III. Briefly an evoked response was assigned a grade I, II, III if it satisfied the respective criteria of normal, moderately abnormal and severely abnormal or absent electrical activity. On the other hand CT scan findings in the PVS were evaluated for abnormal low density areas, ventricular dilatation and enlargement of the sulci and cisterns indicative of atrophy of the brain parenchyma. SSEP and VEP were better correlated with the clinical grade than ABR, and upper brainstem atrophy and abnormal low density area in CT scan findings were more valuable as an index to expresses the clinical features than ventricular dilatation. On the basis of these results, it is concluded that studies of ABR, SSEP and VEP associated with CT scan findings in the PVS could be a useful diagnostic aid to evaluate the lesions of these patients.

Aromatization of androstenedione by normal and neoplastic endometrium of the uterus.

The ability of human uterine endometrium to aromatize androstenedione to estrogens was investigated using 10 normal and neoplastic tissues. Normal and neoplastic endometrial homogenates were incubated with [6,7-3H]androstenedione (A) and NADPH. Estrone (E1) and estradiol (E2) were subsequently isolated in amounts ranging from 0-17600 fmol/h/g and 0-377 fmol/h/g, respectively, from the incubates after purifications by using Bio-Rad AG1-X2 column, thin layer chromatographies and co-crystallization. The conversion of A to E1 and E2 was significantly higher in neoplastic tissues.

Uptake of estrone and estrone-3-sulfate by lung of macaca fuscata.

An equimolar mixture of 3H-E1-S2 and 14C-E1 was injected in one shot into the inferior vena cava near the heart of female Japanese monkey. Following the injection, blood was collected from the aortic arch at intervals of 15 s over a period of 10 min. The concentration of radioactivities in the whole blood and serum was measured. The metabolites were analyzed by DEAE-Sephadex A-25 column chromatography, enzyme hydrolysis, thin layer chromatography and paper chromatography. Both radioactivities of 3H-E1-S and 14C-E1 rapidly decreased in the first 90-s serum sample. The 3H/14C ratio in the 0-15-s serum sample was 5 times higher than the initial ratio of injected compounds. The radioactivities in the serum gradually decreased after 90 s of injection. The 3H/14C ratio in the pulmonary tissue was very low after collecting the final blood sample. This result shows that the most of 3H-E1-S passed through the lung and the larger part of 14C-E1 remained in the lung following injection of these materials. So, it is probable that E1-S is in a form to be carried in the general circulation.

Estrogen biosynthesis in human liver--a comparison of aromatase activity for C-19 steroids in fetal liver, adult liver and hepatoma tissues of human subjects.

After incubation of various tritiated C-19 steroids (androstenedione, testosterone, dehydroepiandrosterone, dehydroepiandrosterone sulfate) with human fetal liver, adult liver and hepatoma tissue homogenates, estrone, estradiol and estriol were analysed after a series of purification steps involving column chromatography, thin layer chromatography and co-crystallization. The findings indicated that the human fetal liver extensively aromatized various C-19 steroids to estrogens, whereas human adult liver and hepatoma tissues exhibited little or no aromatase activities. The formation of estradiol from androstenedione in human fetal liver indicated the presence of 17 beta-hydroxysteroid dehydrogenase in this tissue. It was therefore concluded that although the liver participated in the aromatization process during the fetal stage, extensive aromatization did not take place in the adult liver.

[Estrogen biosynthesis in human female adipose tissue in vitro].

The level of androstenedione aromatase activity in 10 cases of abdominal adipose tissues of premenopausal patients with benign gynecological disease (myoma uteri 7 cases, ovarian cyst 1 case, intersex 1 case, and hydatidiform mole 1 case) was examined. The adipose tissue homogenate (500mg) was incubated with [1,2,6, 7-3H]-androstenedione (10 microCi, 110 pmol), and NADPH (0.5mg) at 37 degrees C for 2h in air. After stopping the enzyme reaction, [14C]-estrone, estradiol (1 X 10(4) dpm) and cold estrone, estradiol (250 micrograms) were added as tracers. Ethyl acetate extract was submitted to Bio-Rad AG1-X2 column chromatography, TLC, and co-crystallization to constant specific activity and 3H/14C ratio. Estrone was formed in all samples (29-65 fmol/g/h), and estradiol was formed in 7 samples. The estradiol level was extremely low compared to estrone. Control samples (no tissue blank and zero time incubated samples) were under 1 fmol/g/h for estrone and estradiol through this procedure, respectively. This result indicates that adipose tissue may contribute to estrogen formation as one of the extragonadal tissues. The enzyme activity did not differ significantly according to gynecological disease, age, and superficial area.