Chromium increases pancreatic metallothionein in the rat

ArticleinToxicology 142(2):111-7 · February 2000with 114 Reads
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Abstract
The ability of chromium (Cr) salts to increase metallothionein (MT) levels in rat liver, kidney and pancreas, and its relationship with the presence of toxic effects are reported here. Rats were injected subcutaneously with 0, 10, 20, 30, 40, or 50 mg K2Cr2O7/kg and sacrificed 24 h later. Total Cr accumulation followed a dose-dependent pattern, levels in kidney being higher than those in liver or pancreas, suggesting different tissue bioavailabilities and accumulation patterns. Cr(IV) administration resulted in a tissue-specific MT induction: pancreas and liver showed five- and 3.5-fold MT increases, respectively; no increase was observed in the kidney. A positive correlation was observed between zinc and MT concentrations in liver, and between total Cr and MT concentrations in pancreas. Serum alpha-amylase activity showed a dose-dependent increase starting from 20 mg/kg, whereas serum glucose levels increased at doses higher than 30 mg/kg. Serum aspartate aminotransferase and alanine aminotransferase activities were increased in a dose-dependent manner, from 20 and 30 mg/kg, respectively. Our results showed that treatment with Cr(VI) can induce MT synthesis in pancreas and suggests a subsequent binding of Cr to MT. Also, pancreas is a target organ for Cr toxicity, and the usefulness of alpha-amylase activity as a sensitive biomarker of Cr toxicity in human exposed populations merits further study.

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    Full-text available
    The present study was carried out to determine the effectiveness of N-acetylcysteine (NAC) in alleviating chromium (VI)-induced beta-cell damages in rats. Sixty male albino rats were randomly divided into four groups (n = 15/group). Group I remained untreated; Group II received single dose of potassium dichromate (50 mg/kg b.w., s.c.); Group III received NAC (200 mg/kg b.w., i.p.) before chromium (VI) injection (50 mg/kg b.w., s.c.); Group IV received NAC alone (200 mg/kg b.w., i.p.). Pancreatic tissue malondialdehyde (a marker of lipid peroxidation), pancreas antioxidant power, blood glucose level and ultrastructure of beta-cells were evaluated. Results showed that the administration of chromium (VI) resulted in a state of pancreatic injury and extensive oxidative damage in rats as manifested by the increase in lipid peroxidation and the decrease in activities of antioxidant enzymes such as glutathione peroxidase and catalase. In serum, there was significant increase in the level of glucose in these animals. Administration of NAC shortly prior to chromium (VI) significantly mitigated most of these changes to control values. Based on ultrastructural observations, the administration of NAC may effectively rescue beta-cells from oxidative damage without affecting their function and structural integrity. It can be concluded that NAC if administrated before chromium (VI) injection improves glycemic state by enhancing insulin secretion and antioxidant competence in pancreatic beta-cells.
  • Article
    Full-text available
    Hexavalent chromium as K2Cr2O7 (60mg/kgbw) was administered intraperitoneally to male Sprague Dawley rats. In the, thyroid gland, chromium concentration decreased significantly (P<0.01) as compared to the control tissue. Serum FT3 and FT4 levels decreased significantly (P<0.01 and P<0.001 respectively), while serum TSH concentration increased significantly (P<0.01) than the control. Histologically, in the pituitary gland, hypertrophy was evident in the anterior pituitary gland hypertrophied, the cellular density (number of cells/0.021mm2) decreased significantly (P<0.001) and the cellular diameter increased significantly (P<0,001). In the thyroid gland, follicle number increased significantly (P<0.001) when compared to control. However, follicular size showed a significant decrease (P<0.001). The follicles were scattered and aggregated abnormally to form small groups making large interfollicular spaces due to the disruption of the connective tissue. The height of epithelial cells also increased. The present study demonstrates that chromium in hexavalent form causes both structural and functional disturbance to the pituitary and thyroid glands and is therefore potentially toxic to these tissues.
  • Article
    Heavy metals that are harmful to humans include arsenic, cadmium, chromium, lead, mercury, and nickel. Some metals or their related compounds may even cause cancer. However, the mechanism underlying heavy metal-induced cancer remains unclear. Increasing data show a link between heavy metal exposure and aberrant changes in both genetic and epigenetic factors via non-targeted multiple toxicogenomic technologies of the transcriptome, proteome, metabolome, and epigenome. These modifications due to heavy metal exposure might provide a better understanding of environmental disorders. Such informative changes following heavy metal exposure might also be useful for screening of biomarker-monitored exposure to environmental pollutants and/or predicting the risk of disease. We summarize advances in high-throughput toxicogenomic-based technologies and studies related to exposure to individual heavy metal and/or mixtures and propose the underlying mechanism of action and toxicant signatures. Integrative multi-level expression analysis of the toxicity of heavy metals via system toxicology-based methodologies combined with statistical and computational tools might clarify the biological pathways involved in carcinogenic processes. Although standard in vitro and in vivo endpoint testing of mutagenicity and carcinogenicity are considered a complementary approach linked to disease, we also suggest that further evaluation of prominent biomarkers reflecting effects, responses, and disease susceptibility might be diagnostic. Furthermore, we discuss challenges in toxicogenomic applications for toxicological studies of metal mixtures and epidemiological research. Taken together, this review presents toxicogenomic data that will be useful for improvement of the knowledge of carcinogenesis and the development of better strategies for health risk assessment.
  • Article
    Molybdenum (Mo) and chromium (Cr) both exist in elevated concentrations in aquatic environments primarily as oxyanions: molybdate (MoO42-) and chromate (CrO42-). These concentrations typically arise as a result of anthropogenic activity. Both metals are relatively non-toxic compared to other metals. Average 96h LC50 estimates in freshwater are at least 1000mgL-1 for Mo and approximately 100mgL-1 for Cr. Both metals are micronutrients, but there is no evidence that their internal concentrations in animals are regulated in a homeostatic manner. Molybdenum is involved in purine metabolism, and Cr is involved in fat and glucose metabolism. Very little information exists about the physiological impact of elevated aquatic concentrations of these metals, especially Mo, in fish. Chromium is taken up by the gills and distributed via the blood to a variety of tissues but the mechanism of uptake across the gills is unknown. Chromium's toxic mechanism in fish is unknown, but histopathologies, particularly of the gastrointestinal tract and kidney, play a significant role in its toxicity. Molybdenum is also transported across the gills via an unknown mechanism and accumulates internally in the liver; other sites of accumulation have not been identified. The speculation on toxicity of Mo includes a non-specific gill irritation response, but this lacks experimental evidence. The general metabolism of these two metals in fish and the factors that influence their bioavailability need future investigation, and provide limitless opportunities for future research.
  • Article
    Full-text available
    Earlier studies have demonstrated that chromium (Cr) Ⅵ compounds have been shown to be more toxic and carcinogenic than other chromium compounds. The aim of the present work was to evaluate the antioxidant effects of red ginseng against chromium Ⅵ-induced toxicity and free radical generation. Sixty adult male rats were divided into six equal groups include: control group, group received Cr Ⅵ alone (50 mg/kg b.w.), group treated with Korean ginseng (K. ginseng) alone (20 mg/kg b.w), group treated with Cr Ⅵ for 15 days then received K. ginseng for other 15 days, group treated with Cr Ⅵ and K. ginseng at the same time for 15 days, and group treated with K. ginseng for 15 days then Cr Ⅵ for other 15 days. The results revealed that Cr Ⅵ caused significant increase in ALT, AST, ALP, G-GT, urea, creatinine, and acid phosphatase. Whereas, it caused significant decrease in TP, albumin, testosterone, GPX, and SOD indicating a stress for liver, kidney and testes. K. ginseng alone caused significant increase in GPX and SOD activities in healthy animals and this result suggests a prophylactic role for this herb in protection against the damaging impact induced by free radical species. Furthermore, the other biochemical parameters measured after K. ginseng administration were comparable to the control values. Treatment with Cr Ⅵ followed by K. ginseng, Cr Ⅵ and K. ginseng or K. ginseng followed by Cr Ⅵ resulted in significant improvement in all tested parameters towards the normal values of the controls. However, this improvement was pronounced in the group pre-treated with K. ginseng for 15 days before Cr Ⅵ administration. It could be concluded that K. ginseng exhibited a protective action against the toxic effects of Cr Ⅵ and it had the ability to scavenge free radicals resulted from Cr Ⅵ intoxication.
  • Article
    Potassium dichromate (K2Cr2O7) is a soluble hexavalent chromium compound that is widely used in several industries. In the present work the effect of administration of K2Cr2O7 on rat intestinal brush border membrane(BBM) enzymes and anti-oxidant system was studied. Rats were given a single oral dose of K2Cr2O7 (100 mg/kg bodyweight) and sacrificed 6, 12, 24, 48 and 96 h after the treatment.Control animals were not given K2Cr2O7. The administration of K2Cr2O7 resulted in a reversible decline in the specific activities of several BBM enzymes. The decrease in the activities of these enzymes was due to changes in the maximum velocity while their affinities for the substrates remained unchanged. Lipid peroxidation increased while total SH groups decreased in K2Cr2O7-treated rats as compared to controls indicating increased oxidative stress in the intestinal mucosa. The activities of superoxide dismutase and glutathione-S-transferase increased while those of catalase, glutathione reductase, thioredoxin reductase and glucose-6-phosphate dehydrogenase decreased. The maximum changes in all the parameters studied above were 24 h after administration of K2Cr2O7 after which recovery took place,in most cases almost to control values after 96 h. These results show that oral administration of K2Cr2O7 to decrease in the activities of BBM enzymes, increase in oxidative stress and alters the activities of anti-oxidant enzymes in rat intestine.
  • Article
    Northern blot hybridization established that metallothionein (MT) mRNA levels were dramatically elevated in the rat pancreas following injection of Cd or Zn salts. To determine which pancreatic cell types express the MT gene, Northern blot hybridization analysis of RNA from preparations enriched for acini, in situ hybridization, and immunocytochemistry were used. RNA from pancreatic acini of Zn-treated rats contained high levels of MT mRNA. In control rats, in situ hybridization suggested very low levels of MT mRNA in both exocrine and endocrine cells in the pancreas, but these levels were dramatically increased in both these cell populations following metal injection. In contrast, levels of insulin-I mRNA in the endocrine cells were not affected by metal injection. A similar result with MT mRNA was obtained in mouse and chicken pancreas using Northern blot and in situ hybridization. Immunocytochemistry detected MT in the rat acinar cell cytoplasm following metal injection. Although inconsistent with in situ hybridization studies and immunocytochemical analysis of exocrine cells, immunocytochemistry for MT indicated a uniform staining pattern of islet cells that was unaffected by metal treatment. These results establish that metal ion induction of the MT genes in pancreas occurs in both endocrine and exocrine cells, which suggests that this protein has diverse physiologic functions in this organ.
  • Metal ions induce expression of metallothionein in pancreatic exocrine and endocrine cells A general mechanism for microsomal activation of quinone anti-cancer agents to free radicals
    • G K Andrews
    • K Kage
    • Palmiter
    • P Thomas
    • M P Sarras
    Andrews, G.K., Kage, K., Palmiter-Thomas, P., Sarras, M.P., 1990. Metal ions induce expression of metallothionein in pancreatic exocrine and endocrine cells. Pancreas 5, 458– 554. rM.J. Solis-Heredia et al. / Toxicology 142 (2000) 111–117117 Bachur, N.R., Gordon, S.L., Gee, M.V., 1978. A general mechanism for microsomal activation of quinone anti-cancer agents to free radicals. Cancer Res. 38, 1745–1755
  • Article
    Metallothioneins (MT) have been implicated in the protection of cells from oxidative stress. We studied the molecular mechanism of induction of MT-I and MT-II in response to restraint stress using a mouse model system in which the animals were restrained in well ventilated polypropylene tubes for 12 h each day (one cycle). Here, we show that MT-I and MT-II mRNA levels were elevated as much as 10-20-fold after just one cycle of this simple stress. Stress-mediated MT induction occurred at the transcriptional level. The level of MT mRNA correlated with the stress-induced increase, and not with the diurnal variation, in the level of serum glucocorticoid. Treatment of the mice with RU 486, a glucocorticoid receptor antagonist, prior to restraint stress inhibited MT induction by at least 50%. Furthermore, the glucocorticoid responsive element-binding activity in the liver nuclear extracts from the stressed mice was significantly higher than that in the control mice. The complex formations between the transcription factor Sp1, MTF1, or MLTF/ARE and the respective specific oligonucleotides were not altered in the liver from the stressed mouse. The MT mRNA levels returned to the basal level at the end of nine cycles of stress, indicating habituation of the animals to restraint stress. At this stage, exposure of the animals to another type of stress, treatment with heavy metals, resulted in further induction of MT. These data indicate that glucocorticoid is the primary physiological factor responsible for MT induction following restraint stress, and the glucocorticoid receptor is the major transcription factor involved in this process.
  • Article
    Localization of metallothionein induced by zinc or streptozotocin treatment was demonstrated in the pancreas of mice by immunohistochemical techniques.Ten hours after the administration of zinc, a small amount of metallothionein appeared in the nuclei of exocrine cells, and one day after the administration, metallothionein was found mainly in the cytoplasm and interstitium. Metallothionein was not observed in the islets of Langerhans. When streptozotocin was administered to mice, a small amount of metallothionein was found in the cytoplasm of the islets of Langerhans.These results show that zinc affected mostly exocrine cells and streptozotocin affected mainly endocrine cells, and that metallothionein was produced at locations damaged by zinc or streptozotocin.
  • Article
    Induction of hepatic metallothionein (MT) by acetaminophen was characterized in the rat and mouse. Treatment of rats with the hepatotoxin resulted in increase of liver MT in a dose-dependent manner. MT concentration was elevated by 41%, 140% and 260% following acetaminophen injection at doses of 250, 500 and 1000 mg/kg, respectively. The cadmium-binding protein was identified as MT by Sephadex G-75 gel filtration (Ve/Vo = 2.1). In the mouse the hepatotoxin was more potent i.e. maximal effect (increase of 230%) was achieved at the lowest applied dose (250 mg/kg). In both species maximal induction was observed 24 h post exposure and thereafter the hepatic MT content declined, indicating a relatively short half-life of the protein. The elevation of the intracellular concentration of a sulfhydryl-rich protein such as MT may serve as self protecting mechanism of the hepatocyte against highly reactive metabolites of toxic substances.
  • Article
    The highlyactive, quinone-containinganticancer drugs, Adriamycin, daunorubicin,carminomycin,rubidazone, no- galamycin, aclacinomycin A, and steffimycin (benzan- thraquinones); mitomycin C and streptonigrin (/V-hetero- cyclic quiñones); and lapacho! (naphthoquinone)interact with mammalian microsomes and function as free radical carriers. These quinone drugs augment the flow of elec trons from reduced nicotinamide adenine dinucleotide phosphate to molecular oxygen as measured by en hanced reduced nicotinamide adenine dinucleotide phos phate oxidation and oxygen consumption.This reaction is catalyzed by microsomal protein and produces a free radical intermediate form of the drugs as determined by electron paramagnetic resonance spectroscopy. Micro- somes from mouse and rat liver, heart, lung, and spleen and mouse L1210 and P388 tumors all catalyze the aug mented oxygen consumption. Apparent Kmvalues deter mined with normal rat liver microsomes range from 0.49 x 10~"Mfor steffimycinto 13.4 x 10~"Mfor lapacho!. Since SKF 525A and carbon monoxide have little effect on this reaction, cytochrome P-450 is probably not involved. Sev eral nonquinoneanticancer agents were tested and were found inactive in the system. Since quinone anticancer drugs are associated with chromosomal damage that appears to be dependent on metabolic activation of these drugs, we propose that the intracellular activation of these drugs to a free radical state may be primary to their cytotoxic activity. As free radicals, these drugs, because of their high affinity and selective bindingto nucleic acids, have the potential to be "site-specific free radicals" that bind to DMA or RNA and either react directly or generate oxygen-dependent free radicals such as Superoxideradi cal or hydroxyl radical to cause the damage associated with their cytotoxic actions.
  • Article
    Mouse liver and pancreatic concentrations of metallothionein were determined by gel filtration at several intervals after the ip administration of cadmium acetate. Hepatic concentrations of the protein reached peak values 48 hr after the injection, whereas the pancreatic concentrations were highest at 6 hr. The exposure of mouse isolated pancreatic islets to a concentration of 1 × 10−6m cadmium significantly reduced glucose-stimulated insulin secretion. When mice were administered cadmium 6 hr prior to islet isolation, the time required for peak concentrations of metallothionein to develop in the pancreas, subsequent exposure to the metal did not reduce glucose-stimulated insulin secretion. The results suggest that the pancreas synthesizes metallothionein and that the protein serves a protective role against the acute effects of cadmium.
  • Article
    The isolation of two forms of hepatic zinc-thioneins after either zinc injection into rats or partial restriction of their food intake is described. The proteins differed slightly in their amino acid composition and electrophoretic mobilities. Increases in liver zinc content after both treatments were synchronous with, and associated almost completely with, increased zinc-binding to these proteins. The time-course for the appearance and disappearance of the zinc proteins is shown. It is suggested that metallothionein is involved in the normal metabolism of zinc, perhaps in some temporary storage or detoxication capacity.
  • Article
    Metallothionein genes (MT) are inducible by a variety of agents, including heavy metals. We report the induction of MT expression by arsenite (As3+) in rat liver in vivo. As3+ (but not arsenate [As5+]) injection increased MT protein and MT-1 and MT-2 mRNA accumulation in liver only, but not in kidney or pancreas. In addition, As3+ enhanced zinc-induced MT protein accumulation in liver without any increase in MT mRNA levels. These data indicate that arsenic may increase MT expression either directly (by inducing MT mRNA accumulation), or indirectly by altering post-transcriptional events. This constitutes an unusual mechanism of enhancement of MT gene expression and appears to be mediated by processes not specifically associated with binding of arsenite to MT in vivo.
  • Article
    A comparison of the effects of intraperitoneal and subcutaneous routes of administration of sodium dichromate on nephrotoxicity in rats was studied. Dichromate when injected subcutaneously (SC group) produced a higher degree of nephrotoxicity than when administered intraperitoneally (IP group). It caused severe progressive proteinuria followed by polyuria and glucosuria, reaching maximum levels at 3 days after treatment in the SC group, whereas it produced mild proteinuria without glucosuria in the IP group. The dose-dependent increases in blood urea nitrogen (BUN) and creatinine concentrations, shown in the SC group, were not observed in the IP group. However, between the two groups, there were no great differences in either the urinary excretion rate of chromium or the electrophoretic patterns of urinary protein in the day 1 urine specimens. Pretreatment of phenobarbital (PB) had no remarkable effect on the dichromate-induced nephrotoxicity. In contrast, it potentiated dichromate-induced hepatotoxicity, the indices of which were the elevation in serum alanine aminotransferase (ALT) activity and hepatic lipid peroxide formation. These results suggest that the dependence of dichromate-induced nephrotoxicity on the route of administration is related to the chemical forms of chromium reaching the kidney, and the necrotizing property of dichromate results from its metabolic fate in vivo.
  • Article
    Effects of metallothionein (MT) on pancreatic endocrine cells of mice, injected with alloxan and at different zinc status were studied. Mice were given drinking water containing four different concentrations of zinc (0, 0.05, 0.1 or 0.5%) for 18 days, and alloxan was injected once on the 14th day. When zinc was added to the drinking water, pancreatic zinc and MT contents increased after injection of alloxan, but did not change with injection of vehicle alone, except in the group of mice drinking 0.5% zinc in water. However, plasma glucose level was increased in all the alloxan injected groups, and was independent of their zinc status. In mice given water with 0.5% of zinc, both pancreatic zinc and MT contents were higher than control mice given water alone. There was no difference in zinc and MT contents of the pancreas in mice drinking 0.5% zinc in groups injected with either alloxan or vehicle. The increase in plasma alpha-amylase activity, an indicator of pancreatic exocrine toxicity, was observed only in mice drinking water with 0.5% zinc after injection of both alloxan and vehicle. Histochemically, degranulation of zymogen and duct-like structures of exocrine cells and atrophy and disappearance of islet cells were observed in alloxan-injected mice drinking 0.5% zinc in water. The zymogen degranulation was observed on the vehicle-injected mice drinking 0.5% zinc in water. MT was immunohistochemically detected in the exocrine cells of both alloxan- and vehicle-injected mice given 0.5% zinc in water. No MT was detected in islet cells of mice in any group. The results show that an increase of zinc content may be followed by induction of MT synthesis in the pancreas of mice given increasing amounts of zinc in drinking water. However, MT dose not provide any protection against damage caused by alloxan to endocrine cells of the pancreas.
  • Article
    Metallothionein (MT) is a low-molecular-weight protein with a high cysteine content that has been proposed to play a role in protecting against oxidative stress. For example, MT has been shown to be a scavenger of hydroxyl radicals in vitro, and cells with high levels of MT are resistant to radiation. However, it is not known if compounds that cause oxidative stress affect MT levels. Therefore, mice were injected subcutaneously with 11 chemicals (t-butyl hydroperoxide, paraquat, diquat, menadione, metronidazole, adriamycin, 3-methylindole, cisplatin, diamide, diethyl maleate, and phorone) that produce oxidative stress by four main mechanisms. MT was quantitated in the cytosol of major organs (liver, pancreas, spleen, kidney, intestine, heart, and lung) by the Cd/hemoglobin radioassay 24 hr after administration of the chemicals. All agents significantly increased MT levels in at least one organ. Liver was the most responsive to these agents in that all 11 chemicals increased MT concentrations in liver, with diethyl maleate, paraquat, and diamide producing 20- to 30-fold increases. Pancreas and kidney were the next most responsive organs to these chemicals. The organ least responsive to these agents was the heart, as only 3 compounds caused significant increases in MT concentrations in heart. Diethyl maleate and diquat were the most general inducers of MT in that they increased MT in six of the seven organs examined. No treatment resulted in a significant decrease in MT concentration in any organ. In conclusion, chemicals that produce oxidative stress by one of four distinct mechanisms are very effective at increasing MT concentrations in a variety of organs. This suggests that MT might be involved in protecting against oxidative stress.
  • Article
    Subcutaneous injection of sodium dichromate into male Sprague-Dawley rats immediately produced a variety of metabolic changes in a dose-dependent manner. Serum lactate and glucose were significantly increased after dichromate treatment, reaching maximum levels at 15 and 30 min, respectively. Then, the toxicity progressively diminished. In contrast, a steady increase in blood urea nitrogen (BUN) concentration was caused by dichromate, reaching maximum levels at 60 min after the administration; elevated BUN levels were sustained for several hours thereafter. Unlike KCN (5 mg/kg, ip) and As2O3 (5 mg/kg, ip), dichromate rapidly decreased serum insulin within 15 min after intoxication in doses of 20 and 40 mg/kg; hypoinsulinemia lasted 60 min. However, insulin levels returned to the normal range at 120 min after treatment. Dichromate-induced metabolic disturbance was also observed in the 24 hr-fasted rats, the response of which was similar to normal rats except for later hyperglycemia. In both cases, the duration time was short (30 to 60 min). Adrenalectomy and insulin pretreatment had no effect on dichromate-induced hyperglycemia. These results suggest that dichromate-induced metabolic disturbance results from the concomitant effects of a sudden decrease in serum insulin level and its direct inhibitory effect on carbohydrate metabolism. In addition, the characteristic biphasic pattern of metabolic disturbance might be related to metabolic fate of dichromate in vivo.
  • Article
    The ability of zinc (Zn) salts to induce the synthesis of metallothionein (MT) in liver, kidney and pancreas of rats pretreated with cadmium (Cd) salts was investigated. Twenty-four hours after either CdCl2 (2.0 mg Cd/kg, s.c.) or saline pretreatment, rats were injected with saline, CdCl2 (2.0 mg Cd/kg, s.c.) or ZnSO4 (20 mg Zn/kg, s.c.) and the concentrations of MT and MT-1 mRNA in tissues subsequently measured. After a single injection of Cd salts, concentrations of MT and MT-1 mRNA were significantly increased in liver as compared to control. With two injections of Cd, the accumulation of MT in liver was approximately twice the levels of MT following a single injection of Cd. In kidney, MT and MT-1 mRNA expression were significantly increased only after two injections of Cd and in the pancreas, Cd injections did not alter either MT content or MT-1 mRNA expression. Treatment with Zn salts increased MT concentrations in both liver and pancreas. However, the pancreas was the most responsive to injections of Zn salts as compared to the liver in terms of increases in both protein concentration and MT-1 mRNA expression. When Zn injection was preceded by a Cd injection, induction as measured by MT-1 mRNA and MT concentrations were approximately additive in liver. In kidney, although Cd or Zn treatment separately had no effect on MT or MT-1 mRNA content, injection of Cd followed by Zn resulted in significantly increased levels of renal MT and MT-1 mRNA. Fractionation of liver cytosols on a Sephadex G-75 column revealed that in animals receiving two injections of Cd, virtually all the Cd was associated with MT whereas Zn was distributed between both high molecular weight (HMW) proteins and MT. In animals receiving both Cd and Zn injections, cytosolic Cd was still bound predominantly to the MT fraction, while the proportion of cytosolic Zn associated with MT increased. The results of this study suggest that, treatment with Cd salts followed by Zn salt injection can induce further synthesis of MT in liver, kidney and pancreas with subsequent binding of both Zn and Cd to the intracellular MT.
  • Metal-lothionein synthessis by trivalent or hexavalnt chromium in mice
    • H Ohta
    • Y Seki
    • S Imamiya
    • H Yoshikawa
    Ohta, H., Seki, Y., Imamiya, S., Yoshikawa, H., 1993. Metal-lothionein synthessis by trivalent or hexavalnt chromium in mice. In: Anke, M., Meissner, D. (Eds), Trace Elements in Man and Animals. pp. 178 – 179.
  • Article
    The nature of hepatic metallothionein (MT) induction by several metals and its relationship to an inflammatory response was studied in chicks. Intraperitoneal (ip) injection of chromium (Cr), managanese, and iron (Fe) caused a much greater increase in hepatic MT (10.2-, 9.0-, and 6.8-fold) compared with cobalt and nickel (2.5- and 2.9-fold); thus not all transition metals are effective. Cr3+ caused markedly greater hepatic MT accumulation than Cr6+, suggesting that the ionic nature of the metal is an important factor. Small organic complexes of Fe (ferrous gluconate or lactate, 6.2-fold) caused significantly greater accumulation of hepatic MT than ferric dextran (1.4-fold), a large organic aggregate. In vitro data from chick hepatocytes and/or fibroblasts clearly indicated that Fe does not effect the induction of MT directly. The role of inflammation, as measured by recruitment of peritoneal exudate cells (PEC), was examined. Endotoxin (LPS), Sephadex (S), and Fe elicited significant elevations in PEC number at 24 h posttreatment (S), and Fe elicited significant elevations in PEC number at 24 h posttreatment (S = Fe greater than LPS much greater than control). The percentage of heterophils but not macrophages was significantly correlated with the accumulation and induction of hepatic MT. In a similar experiment with Cr, we demonstrated that Cr3+ but not Cr6+ stimulated MT messenger RNA accumulation and concomitant hetereophil infiltration at 3 h after injection. Our results indicate that the induction of hepatic MT by the parenteral administration of a number of metals is dependent on the chemical nature of the metal and is associated with an inflammatory response.
  • Article
    Chromium is an essential nutrient required for normal sugar and fat metabolism. Insufficient dietary chromium is associated with maturity-onset diabetes and/or cardiovascular diseases. Dietary chromium intake in the U.S. and other developed countries is roughly half of the minimum suggested intake of 50 micrograms. Well controlled studies involving human subjects have demonstrated beneficial effects of supplemental chromium on fasting glucose, glucose tolerance, blood lipids, insulin binding, and hypoglycemic blood glucose values and symptoms. Since chromium is a nutrient and not a therapeutic agent, it will only benefit those people whose signs and symptoms are due to marginal or overt chromium deficiency. Stresses including high sugar diets, strenuous exercise, physical trauma, infection and certain diseases exacerbate the signs and symptoms associated with marginal intakes of dietary chromium. While excessive levels of chromium are usually limited to industrial settings, marginal dietary chromium intake is widespread in the general population and may lead to serious health problems.
  • Article
    Previous results regarding the two metalloforms of MT which accumulate in chick liver following the parenteral administration (ip) of copper were discussed. One metalloform, which is exclusively zinc, was suggested to reflect the marked accumulation of hepatic zinc following copper injection. The present report shows that there is a marked difference in hepatic zinc accumulation if copper is administered iv. Under these conditions there is virtually no change in hepatic zinc and thus MT produced under these conditions appears to contain only copper. We suggest the the changes in zinc metabolism as effected by copper when given intraperitoneally reflect a secondary response analogous to that observed when iron is similarly administered.
  • Article
    Insulin has been reacted with five chromium(III) complexes that are capable of relatively facile substitution of aquo ligands. The new Cr(III) insulin derivatives have been characterized by means of electronic and infrared spectra, and evidence for major changes in the protein structure, including the state of aggregation, has been presented. Supporting evidence for the arguments favoring the beneficiary role of chromium(III) in glucose metabolism has been obtained using in vivo studies, and it has been shown that insulin derived with Cr(salen) (H2O)2+ is capable of reversing the blood sugar, serum cholesterol, and phospholipids levels to those of normal rats. The results emphasize the dependence of biopotency on the structure of Cr(III) complexes used for derivation of insulin and discount the postulates that Cr(III) serves to assemble insulin and receptor units through metal-sulphur bonding. The influence of Cr(III) on the structural stability and state of aggregation of insulin and their possible role in glucose metabolism is discussed.
  • Article
    The influence of maternal dietary zinc intake and recombinant human interleukin-1 alpha (rhIL-1 alpha) administration on metallothionein gene expression and the distribution of 65Zn were investigated. Pregnant rats were fed diets containing 1, 5, 30 or 180 mg Zn/kg diet in an equalized regime from d 13-20 of gestation. Metallothionein gene expression was examined by Northern blot and dot blot hybridization using combined 60-mer oligonucleotides specific for rat metallothionein-1 and -2 genes. Expression was progressively depressed in the fetal livers and livers and kidneys of dams fed diets marginal (5 mg/kg) and deficient (1 mg/kg) in zinc content. Administration of rhIL-1 alpha increased expression in maternal liver, placenta and in fetal liver of dams fed adequate or deficient diets. Kinetics of intravenously administered 65Zn showed that in response to rhIL-1 alpha, there was a higher uptake by the maternal liver and bone marrow with less 65Zn uptake by bone, intestine and plasma activity compared to controls. No change was observed in 65Zn taken up by the placenta or transferred to the fetus. Alteration of metallothionein gene expression could represent, in part, the mechanism whereby altered effects of zinc metabolism and function are mediated during fetal development.
  • Article
    Full-text available
    Induction of metallothionein-I (MT-I) and metallothionein-II (MT-II) by glucocorticoids was determined by h.p.l.c. analysis of proteins and Northern-blot analysis of MT mRNAs. Rats were injected with dexamethasone (0.03-10 mumol/kg) and hepatic concentrations of MTs were determined 24 h later. In control rats, only MT-II was detected (9.4 +/- 2.5 micrograms/g of liver), whereas the hepatic concentration of MT-I was below the detection limit (5 micrograms of MT/g). Dexamethasone did not increase MT-I above the detection limit at any dosage tested, but MT-II increased to 2.5 times control values at dosages of 0.30 mumol/kg and higher. Time-course experiments indicated that MT-II reached a maximum at 24 h after a single dosage of dexamethasone and returned to control values by 48 h. To determine whether dexamethasone increased MT-I in liver, samples were saturated with 109Cd, after which the amount of 109Cd in MT-I and MT-II was determined. Results indicated that, by this approach, MT-I and MT-II could be detected in control rats, and there was approx. 1.8 times more 109Cd in MT-II than in MT-I. At 24 h after administration of dexamethasone (1 mumol/kg), there was a small increase in the amount of 109Cd bound to MT-I, whereas the amount of 109Cd bound to MT-II increased to more than 2 times control values. Northern-blot hybridization with mouse cRNA probes indicated that MT-I and MT-II mRNAs increased co-ordinately after administration of dexamethasone. Thus, although glucocorticoids increase both MT-I and MT-II mRNAs, MT-II preferentially accumulates after administration of dexamethasone.
  • Article
    Chromium(III) is thought to be an essential element in mammals, its toxicity being very low. On the contrary, chromium (VI) is highly toxic to man, even if its effects are generally local, involving the respiratory tract and the skin. Once absorbed, chromium(VI) is quickly reduced to the trivalent form which accounts for all of this element present in the blood stream or taken up by tissues. As a result, any differences in systemic toxicity can only be attributed to differential solubilities and absorption rates. The kidney should be regarded as the critical organ, although tubular damage following occupational exposure is mostly due to acute absorption and transient in nature. Sensitive immunochemical techniques for the measurement of specific proteins in the urine have been used for the early detection of kidney damage, a possible threshold having been indicated at exposure levels yielding 15 micrograms/g creatinine in urine. Such a threshold has been confirmed by using monoclonal antibodies to reveal antigens from the brush-border of proximal tubules. Two main features of kidney damage were, however, apparent. The first one is the lack of dose-effect/response relationships, i.e. the lack of any progression toward more severe impairments when the exposure intensity increases. The second one is that the recent absorption rate more than the cumulated dose is responsible for the observed nephrotoxic effects.
  • Article
    Early research on metallothionein centered on aspects related to a detoxification role. As our understanding of the complex endocrine control that regulates metallothionein gene expression increases, a wider appreciation of a functional role(s) is emerging. Medical implications of control of metallothionein turnover include diagnosis of specific diseases and regulation of its expression as a host defense component.
  • Article
    The binding of Ag+ to metallothionein (MT) was investigated, and a Ag-saturation assay was developed for the measurement of MT in tissues. When samples of purified hepatic Zn-MT or Cd-MT were titrated with Ag+ followed by hemolysate-heat treatment, it was found that saturation of MT occurred at 17 to 18 g-atoms Ag+/mol protein. The rank order of potencies of metals to displace Ag+ from 110mAg-labeled Ag-MT was Ag+ greater than Cu2+ greater than Cd2+ greater than Hg2+ greater than Zn2+ at pH 8.5 in 0.5 M glycine buffer. When a Ag-saturation (Ag-hem) assay was compared with a Cd-saturation (Cd-hem) technique, excellent correspondence was obtained in the measurement of MT from various sources including purified Zn-MT (II), MT in human kidney and liver tissue samples, hepatic MT in Cd-injected adult rats, and renal MT in Hg-injected adult rats. However, in cases where significant amounts of Cu-MT were present, such as in the livers of Cu-injected rats or in the kidneys of Cd-injected rats, the Cd-hem assay consistently underestimated the MT concentrations compared to the Ag-hem method. This is attributed to the inability of Cd to displace Cu effectively from MT at pH 8.5. Thus the Cd-hem assay is not recommended for the measurement of metallothioneins containing a high Cu content. Under such conditions, the Ag-hem method seems superior.
  • Article
    The ability of selected metals (Ag, Al, As, Ca, Cd, Co, Cr, Cu, Fe, Hg, Mg, Mn, Mo, Ni, Pb, Tl and V) to displace Zn from Zn-metallothionein (Zn-MT) was quantitated. Of the metals tested Cd had the highest affinity for MT, with 1.33 microM displacing 50% of the 65Zn bound to MT (EC50), followed by Pb (1.46 microM), Cu (1.93 microM), Hg (3.93 microM), Zn (8.06 microM), Ag (10.4 microM), Ni (474 microM) and Co (880 microM). As, Ca and Mo had a limited ability to displace Zn from MT while Al, Cr, Fe, Mg, Mn, Tl and V had no effect on Zn binding even at 1.0 mM.
  • Article
    Although the induced synthesis of metallothionein (MT) after exposure to certain metals has been known for some time, there is little information on the quantitation of MT in various tissues. In this study, tissue MT concentrations were measured by a modified Cd-saturation method in tissues of adult male rats after injection of different metal salts. There were differences in tissue levels of MT, depending on the injected metals. Of all the metals studied, Cd2+ was the most effective element in increasing MT concentrations in liver, kidney, pancreas and small intestine. The highest increase in tissue MT concentration after CdCl2 injection was found in the liver, while the pancreas contained the highest MT level after ZnSO4 injection. Co and Ni salts increased MT levels in both liver and kidney, while Mn and Ca increased MT levels only in liver. A direct correlation between tissue MT levels and Cd or Zn concentration was observed in most of the tissues after injection of CdCl2 or ZnSO4. Although there was no positive relationship between tissue levels of MT and tissue Mn and Ni concentrations, the increase in hepatic Zn after injection of these metals was related to hepatic MT levels. The tissue distribution of injected Cd2+ in control adult rats and Zn-deficient rats was similar. However, there was no increase in pancreatic MT levels in Zn-deficient rats after injection of CdCl2. The high concentration of MT in pancreas after ZnSO4 injection in adult rats and the inability of the pancreas to synthesize MT in Zn-deficient rats suggest that the induction of pancreatic MT synthesis is sensitive to Zn status.(ABSTRACT TRUNCATED AT 250 WORDS)
  • Article
    CrCl3, 0.25, 0.5, 1.0, and 1.5 mmol/l inhibited glucose-induced insulin secretion in a reversible and dose dependent manner. Cr also inhibited basal secretion of insulin in the presence of 5.5 mmol/l glucose and insulin secretion stimulated by 50 mmol/l K+ or 15 mmol/l L-leucine. When 2 mmol/l theophylline was employed to potentiate the stimulatory effect of 16.5 mmol/l glucose, the inhibitory effect of 1.5 mmol/l Cr was reduced and that of 0.5 mmol/l virtually abolished. A similar reduction in the inhibitory effect of Cr was observed when the medium calcium concentration was increased from 2.5 to 5, 7.5 and 12.5 mmol/l. Cr did not alter the conversion of 14C-glucose to 14CO2 or 45Ca uptake by isolated islets. It is concluded that the inhibitory effect of Cr on insulin secretion may be mediated through interference with an intracellular function of Ca++ in the beta cell.
  • Article
    Pancreatic metallothionein (MT) and zinc levels were measured after the administration of 4-aminopyrazolo(3,4-d)pyrimidine (4APP) to mice to determine the role of pancreatic MT. 4APP was found to increase the pancreatic MT level dose-dependently; the pancreatic MT level in mice administered 175 mg/kg of 4APP was 3.9 fold higher than that in control mice. When 40 mg/kg or less of 4APP was administered, the zinc concentration of the pancreatic cytosol fraction did not differ from that in control mice, while after 175 mg/kg of 4APP it was 1.7-fold higher. There was no difference in plasma glucose and insulin levels between the controls and mice administered 175 mg/kg of 4APP, although the plasma alpha-amylase activity was reduced in the latter. Moreover, a linear negative relationship was observed between pancreatic MT level and plasma alpha-amylase activity (r = -0.795, p < 0.01). These findings suggest that pancreatic MT is affected by the disruption of the exocrine function in 4APP-administered mouse pancreas.
  • Article
    Metallothionein (MT) concentrations were determined in the cytosol of isolated pancreatic islets of mice, using both the cadmium (Cd)-heme and the Cd-Chelex assay. Both constitutive MT levels and significant MT induction were detected in islet cells. For MT induction, mice were injected intraperitoneally (i.p.) with either zinc sulfate (Zn) or the diabetogen streptozotocin (STZ). Following an i.p. injection of 15 mg Zn.kg-1 body weight (body wt), the mean index of MT induction found in islets was comparable to that found in liver tissue, which was used as control. After i.p. injection of 30 mg Zn.kg-1 or a single high dose of STZ (100 or 200 mg.kg-1), the indices of MT induction in islets exceeded those in liver by a factor of 1.3, 2.5, or 1.5. After multiple low doses of STZ (3 or 5 x 40 mg.kg-1 given on consecutive days), in contrast, the MT induction indices in islets exceeded those in liver by a factor of 3.5 and 3.9 for 3 x 40 respectively 5 x 40 mg STZ.kg-1. In conclusion, our results demonstrate constitutive MT levels in isolated pancreatic islets and significant MT induction after i.p. injection with Zn or STZ, in particular after repeated low doses of STZ.
  • Article
    Metallothionein (MT)-bound zinc accumulates when animals are exposed to excess zinc and is depleted under conditions of zinc deficiency, suggesting that MT serves as a means of sequestering excess zinc as well as a zinc reservoir that can be utilized when zinc is deficient. To examine the importance of MT for these processes, mice with null alleles of both MT I and MT II genes were created and the zinc concentration and histological appearance of multiple organs assessed. At birth, the hepatic zinc concentration of these MT-null mice was lower than that of wild-type controls (0.27 +/- 0.02 vs. 0.65 +/- 0.11 micromol zinc/g tissue, P < 0.05). During the next 3 wk of suckling zinc-replete (95 micrograms zinc/g diet) dams, the hepatic zinc concentration of controls fell to 0.42 +/- 0.04 micromol/g but was unchanged in the MT-null mice (0.28 +/- 0.04 micromol/g). The most prominent histological anomaly observed at 3 wk of age was the presence of swollen Bowman's capsules in the kidneys of MT-null mice. When nursing MT-null dams were fed a severely zinc-deficient (1.5 microg/g) diet, kidney development in the MT-null pups was retarded as indicated by the retention of the nephrogenic zone and incomplete tubule development. We suggest that the lack of a hepatic reservoir of zinc jeopardizes the developing kidney in the MT-null mice. In addition to being more sensitive to dietary zinc restriction, MT-null mice are more sensitive to zinc toxicity. When adult mice were challenged with a ramping dose of zinc up to a total of 3700 micromol zinc/kg body weight, MT-null mice had a greater incidence of pancreatic acinar cell degeneration compared with control mice despite accumulating less zinc (2.72 +/- 0.46 vs. 1.23 +/- 0.52 micromol zinc/g pancreas, control and MT-null, respectively, P < 0.05). The results of these experiments suggest that MT I and MT II can protect against both zinc deficiency and zinc toxicity.
  • Article
    Increased pancreatic beta-cell secretory activity usually is associated with decreased alpha-cell activity; stimulated beta-cells release gamma-aminobutyric acid, which hyperpolarizes alpha-cells, inhibiting glucagon release. Thus, insulin secretion and glucagon secretion are usually inversely coupled. This suggests that chromium and other insulin-sensitizing modalities, by down-regulating beta-cell activity, may increase glucagon secretion. Such an effect might play a role in the documented therapeutic activity of supplemental chromium and biguanides in reactive hypoglycemia, and might also be of benefit to dieters.
  • Article
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    Using transgenic mice that overexpress metallothionein-I (MT-I) and zinc-induced normal and transgenic animals, we have explored the localization of MT in the pancreas. Light-level immunocytochemistry demonstrated MT in acinar cells but not islet cells. Immunolabeling also revealed the presence of MT in pancreatic ducts, suggesting that it is released from acinar cells. Ultrastructural immunolocalization showed that MT was cytoplasmic, and no MT immunoreactivity was detected in lumens of the vesicular secretory pathway. Secreted pancreatic juice was collected from pilocarpine-stimulated mice and assayed for MT by a 109Cd-labeled hemoglobin-exchange assay and by radioimmunoassay. Both methods revealed high (> 1,000 ng/ml) levels of MT in the stimulated secretion. The level of MT in pancreatic juice from transgenic mice was only slightly (2-fold) increased despite dramatic overexpression of MT-I in the pancreas (> 20-fold). In contrast, zinc induction of MT significantly increased MT by 5- to 10-fold in the pancreatic juice, in normal and transgenic mice. These data indicate that MT is released from pancreatic acinar cells but not by the classical vesicular secretory pathway. In addition, MT levels in pancreatic juice are regulated by zinc, suggesting a physiological role of the pancreas in metal homeostasis.