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C D Garner

Publications and source records attributed to C D Garner.

At least 37 records · Page 2Linked to original sources
PubMed1995

Role of retinal metabolism in methanol-induced retinal toxicity.

Methanol is a toxicant that causes systemic and ocular toxicity after acute exposure. The folate-reduced (FR) rat is an excellent animal model that mimics characteristic human methanol toxic responses. The present study examines the role of the methanol metabolites formaldehyde and formate in the initiation of methanol-induced retinal toxicity. After a single oral dose of 3.0 g/kg methanol, blood methanol concentrations were not significantly different in FR rats compared with folate-sufficient (FS) (control) rats. However, FR rats treated with 3.0 g/kg methanol displayed elevated blood (14.6 mM) and vitreous humor (19.5 mM) formate levels and abnormal electroretinograms (loss of b-wave) 48 h postdose. FR rats pretreated with disulfiram (DSF) prior to 3.0 g/kg methanol treatment failed to display these symptoms. Formaldehyde was not detected in blood or vitreous humor with or without DSF treatment, suggesting that formate is the toxic metabolite in methanol-induced retinal toxicity. Additionally, creating a blood formate profile (14.2 mM at 48 h) similar to that observed in methanol-treated rats by iv infusion of pH-buffered formate does not alter the electroretinogram as is observed with methanol treatment. These data suggest that intraretinal metabolism of methanol is necessary for the formate-mediated initiation of methanol-induced retinal toxicity.

Animals↗
PubMed1994

Evaluation of methanol-induced retinotoxicity using oscillatory potential analysis.

Methanol is an ocular toxicant which causes visual dysfunction often leading to blindness after acute exposure. While the manifestation of the toxicity has been widely studied, the mechanism by which the injury is produced is still uncertain. A major unanswered question pertains to the site of action, i.e. direct retinotoxicity versus primary optic nerve toxicity with secondary retinotoxicity. In the present study, the effect of methanol on the oscillatory potentials (OPs) of the electroretinogram (ERG) were evaluated in acutely treated folate sufficient (FS) and folate reduced (FR) Long-Evans rats. The OP amplitudes of the acutely dosed FR rats displayed non-selective decreases in all OP amplitudes and non-selective increases in all OP latencies at methanol doses ranging from 1.5 to 3.0 g/kg. Comparing decreases of op2 and ERG b-wave amplitudes with blood formate concentration demonstrates that the b-wave is more sensitive than op2 in a blood formate concentration range of 6-14 mM, suggesting that retinal ischemia is not involved in methanol-induced visual system toxicity.

Animals↗
PubMed1994

A rat model manifesting methanol-induced visual dysfunction suitable for both acute and long-term exposure studies.

A toxic dose of methanol can induce visual dysfunction and metabolic acidosis in humans. However, the methanol dose range capable of inducing such toxicities and the mechanism(s) of visual dysfunction are not clearly understood. Nonprimate laboratory animals do not develop the characteristic human methanol toxicities even after a lethal dose. In the present study, we investigated whether visual dysfunction can be induced by methanol in a folate-reduced (FR) rat model which accumulates formate. Methanol was administered to rats by either the oral or inhalation route. The latencies of P1 and N1 peaks of flash-evoked potentials were significantly increased in methanol-challenged FR rats (3.5 g/kg, po), indicating that methanol administration caused an adverse effect in the retinogeniculocortical visual pathway. Effects on retinal function were then assessed by evaluating the electroretinogram (ERG). A dose-related reduction in b-wave amplitude of the ERG was manifested following oral methanol administration (1.5 to 2.5 g/kg). The b-wave amplitude reduction was also manifested in FR rats exposed to methanol vapors (2000 ppm, a concentration which was nontoxic in monkeys). These observations were consistent with reported human methanol toxicity cases. Thus, our data suggest that the FR rat model could serve as a valuable human surrogate for studying mechanisms of methanol-induced visual dysfunction and providing reliable toxicity data on the visual system under various exposure scenarios.

Administration, Inhalation↗
PubMed1994

Animal model for the study of methanol toxicity: comparison of folate-reduced rat responses with published monkey data.

We attempted to develop a rodent model that exhibits characteristics of human methanol toxicities such as acidosis and visual dysfunction, which are correlated with an accumulation of formate, a toxic metabolite of methanol. Initially three groups of Long-Evans rats with different levels of liver folate were prepared and examined for formate accumulation after methanol administration (3.5 g/kg). The folate-reduced (FR) rats prepared by feeding a folate-deficient diet with 1% succinylsulfathiazole yielded blood formate levels equivalent to those found in methanol-intoxicated humans and developed signs of the visual system toxicity (a manuscript on the latter aspect is in preparation). Responses of FR rats to a variety of methanol exposure scenarios were then investigated, and the results were compared with those reported in the literature for monkeys. Formate accumulation and/or lethality were used as toxic parameters for this comparative evaluation. In FR rats dosed orally with 3 g/kg, the blood formate concentration was 9.2 mmol/L at 24 h postadministration and increased to 15.6 mmol/L at 48 h. The same dose given to monkeys yielded a plateau of 7.4 mmol/L at 12 h after methanol administration, and stayed at this level for an additional 12 h. The area under the concentration vs. time curve for blood formate in FR rats was 2.5-fold greater than that in monkeys when 2.0 g/kg methanol was administered. After a 6-h exposure to 1200 ppm and 2000 ppm methanol, the blood formate concentrations in FR rats were increased by 370% and 636% above the endogenous level, respectively. However, blood formate did not accumulate above the endogenous level when monkeys were exposed to methanol up to 2000 ppm for 6 h. Under acute inhalation exposure conditions, FR rats exposed to 3000 ppm methanol, 20 h/d, could not survive more than 4 d. On the other hand, monkeys exposed to 3000 ppm, 21 h/d, out-lived 20 d. Moreover, monkeys survived for more than 4 d even after an exposure to 10,000 ppm. Thus, these results indicate that FR rats are more sensitive to methanol challenges than monkeys, and suggest that the FR rat could be a congruous animal model for evaluating the health effects of methanol in humans.

Administration, Inhalation↗
PubMed1993

X-ray absorption studies of the ferrous active site of isopenicillin N synthase and related model complexes.

Isopenicillin N synthase (IPNS) from Cephalosporium acremonium (M(r) 38,400) is an iron-containing enzyme that aerobically catalyzes the four-electron oxidative ring closure reactions of delta-(L-alpha-aminoadipoyl)-L-cysteinyl-D-valine (ACV), forming the beta-lactam and thiazolidine rings of isopenicillin N. Here, we report Fe K-edge X-ray absorption studies that provide insight into the iron coordination environment and the effect of substrate and nitric oxide binding. Our analysis reveals an iron(II) coordination environment consisting of two N/O-containing ligands at 2.01 +/- 0.02 A, three N/O ligands at 2.15 +/- 0.02 A, and one C/O scatterer at approximately 2.6-2.7 A. Three His ligands are associated with the 2.15-A shell, while an unsymmetrically chelated carboxylate is associated with a scatterer at 2.01 and at 2.6-2.7 A, a combination which is consistent with the ligand environment deduced from 1H NMR studies [Ming, L.-J., Que, L., Jr., Kriauciunas, A., Frolik, C. A., & Chen, V. J. (1991) Biochemistry 30, 11653-11659]. The remaining scatterer at 2.01 A is assigned to a coordinated solvent molecule, most likely hydroxide, which can act as the proton acceptor for the incoming substrate. ACV binding to Fe(II)IPNS evinces an Fe-S interaction at 2.35 +/- 0.02 A, indicative of the coordination of substrate cysteine thiolate to the metal center. Analysis of the Fe(II)IPNS-ACV-NO data reveals one Fe-N at 1.71 +/- 0.02 A, three Fe-(N,O) at 2.04 +/- 0.02 A, one Fe-S at 2.32 +/- 0.02 A, and one Fe-(C,O) at 2.61 +/- 0.02 A, the short Fe-N bond being derived from the binding of NO. Our EXAFS conclusions, supported by corresponding analysis of relevant model complexes, corroborate and refine the working model for the Fe(II) coordination environment developed from previous spectroscopic studies.

Absorptiometry, Photon↗
PubMed1991

Iron K-edge absorption spectroscopic investigations of the cores of ferritin and haemosiderins.

The extended X-ray absorption fine structure (EXAFS) associated with the iron K-edge has been measured and interpreted for ferritin and haemosiderin extracted from horse spleen, and haemosiderin extracted from the livers of humans with treated primary haemochromatosis, and from the spleens of humans with treated secondary haemochromatosis. For ferritin, the data are consistent with, on average, each iron atom being in an environment comprised of approx. six oxygen atoms at 1.93 +/- 0.02 A, approx. 1.5 iron atoms at 2.95 +/- 0.02 A and approx. 1.1 iron atoms at 3.39 +/- 0.02 A, with a further shell of oxygens at approx. 3.6 A. Iron in horse spleen haemosiderin is in an essentially identical local environment to that in horse spleen ferritin. In contrast, the EXAFS data for primary haemochromatosis haemosiderin indicate that the iron-oxide core is amorphous; only a single shell of approx. six oxygen atoms at approx. 1.94 +/- 0.02 A being apparent. Secondary haemochromatosis haemosiderin shows an ordered structure with approx. 1.4 iron atoms at both 2.97 +/- 0.02 and 3.34 +/- 0.02 A. This arrangement of iron atoms is similar to that in horse spleen haemosiderin, but the first oxygen shell is split with approx. 2.9 atoms at 1.90 +/- 0.02 A and approx. 2.7 at 2.03 +/- 0.02 A, indicative of substantial structural differences between secondary haemochromatosis haemosiderin and horse spleen haemosiderin.

Animals↗
PubMed1991

Effects of benzene on DNA strand breaks in vivo versus benzene metabolite-induced DNA strand breaks in vitro in mouse bone marrow cells.

Previously, we identified p-benzoquinone (BQ) and 1,2,4-benzenetriol (BT) as toxic metabolites of benzene on the basis of their inhibitory effect on DNA synthesis. In the present study, the capability of benzene and the two metabolites to induce DNA strand breaks was investigated in either the in vivo or the in vitro system by comparing the DNA elution rate on a fine membrane filter at alkaline pH. In the in vitro system were bone marrow cells were reacted with test chemicals for 60 min, both BQ and BT induced a dose-related increase in alkali-labile DNA single-strand breaks (SSBs) of bone marrow cells. However, when glutathione (350 micrograms/ml) was added to the same reaction system, the DNA damaging effect of BQ (24 microM) and BT (24 microM) was blocked by 100 and 53%, respectively. Catalase (130 units/ml) completely blocked the DNA damaging effect of BT, while no protection was afforded with BQ. Consistent with these observations, no induction of alkali-labile DNA SSBs was observed in the in vivo system by an anesthetic dose of benzene (1760 mg/kg, ip or po) at 1, 24, and 36 hr postadministration in both male and female ICR mice. These results suggest that benzene exposure would not induce direct DNA strand breaks in vivo under realistic work-related or accidental exposure conditions and also indicate that caution should be exercised in the interpretation of in vitro data for whole-body toxicity evaluation.

Animals↗
PubMed1990

Metal ion co-ordination in the DNA binding domain of the yeast transcriptional activator GAL4.

The structure of the DNA binding domain of the yeast transcriptional activator GAL4 was investigated by extended X-ray fine structure (e.x.a.f.s.). Two samples of GAL4 were studied, one containing cadmium as a structural probe (Cd(II)GAL4) and the other containing the 'native' zinc (Zn(II)-GAL4). The results suggest that the structure of the DNA binding domain of GAL4 contains a two metal ion cluster distinguishing it from the 'zinc finger' proteins typified by the Xenopus laevis transcription factor TFIIIA.

Cadmium↗
PubMed1990

Iron K-edge X-ray-absorption spectroscopy of the iron-vanadium cofactor of the vanadium nitrogenase from Azotobacter chroococcum.

Iron K-edge e.x.a.f.s. data for the iron-vanadium cofactor (FeVaco) from Azotobacter chroococcum vanadium nitrogenase reported here provide further evidence for the structural similarity between this and the iron-molybdenum nitrogenase cofactor (FeMoco) from Klebsiella pneumoniae molybdenum nitrogenase [Arber, Flood, Garner, Gormal, Hasnain & Smith (1988) Biochem. J. 252, 421-425]. The e.x.a.f.s. data are consistent with the vanadium being present in a V-Fe-S cluster, thus confirming that the N-methylformamide extract of the VFe protein component of A. chroococcum vanadium nitrogenase does indeed contain a polynuclear metal-sulphur cluster. Additionally, a long Fe-Fe distance is observed as 0.369 nm, demonstrating the presence of a long-range order in the cluster.

Azotobacter↗
PubMed1990

Unilateral degeneration of retina and optic nerve in Fischer-344 rats.

Unilateral degeneration of the retina and optic nerve was observed among Fischer-344 (F-344) rats fed a semi-purified synthetic feed. Further studies were conducted using standard cereal-based and synthetic diets. Beginning at 4 weeks of age, all experimental rats (169 F-344 rats) were fed various diets and were examined for morphologic and functional changes in the retina and optic nerve. No ocular lesions were observed in any F-344 rats prior to 21 weeks of age, whether fed a synthetic diet or a standard diet; however, approximately 16% (13/86) of the F-344 rats examined between 57 and 64 weeks of age developed unilateral degeneration of the retina and optic nerve. On the other hand, the F-344 rats fed the synthetic diet developed the degenerative lesions by 30 weeks of age, while the F-344 rats fed the standard diet did not develop lesions over this shorter time period. Degenerative changes of the affected retinas and optic nerves were closely related with functional abnormalities evaluated by electroretinogram and visual evoked potentials. In contrast with the F-344 rats, Long-Evans rats that were fed either the synthetic or standard diet up to the age of 68 weeks (77 rats) did not develop the ocular lesions. There was no apparent relationship of the development of the lesions with dietary modification, toxicity or trauma; thus, these observations appear to indicate that spontaneous unilateral degeneration of the retina and optic nerve occurs in F-344 rats and that these ocular lesions may be accelerated by the feeding of certain semi-purified synthetic diets.

Animals↗
PubMed1989

Vanadium K-edge X-ray absorption spectroscopy of bromoperoxidase from Ascophyllum nodosum.

Bromoperoxidase from Ascophyllum nodusum was the first vanadium-containing enzyme to be isolated. X-ray absorption spectra have now been collected in order to investigate the coordination of vanadium in the native, native plus bromide, native plus hydrogen peroxide, and dithionite-reduced forms of the enzyme. The edge and X-ray absorption near-edge structures show that, in the four samples studied, it is only on reduction of the native enzyme that the metal site is substantially altered. In addition, these data are consistent with the presence of vanadium(IV) in the reduced enzyme and vanadium(V) in the other samples. Extended X-ray absorption fine structure data confirm that there are structural changes at the metal site on reduction of the native enzyme, notably a lengthening of the average inner-shell distance, and the presence of terminal oxygen together with histidine and oxygen-donating residues.

Eukaryota↗
PubMed1989

Vanadium K-edge X-ray-absorption spectroscopy of the functioning and thionine-oxidized forms of the VFe-protein of the vanadium nitrogenase from Azotobacter chroococcum.

Vanadium K-edge X-ray-absorption spectra were collected for samples of thionine-oxidized, super-reduced (during enzyme turnover) and dithionite-reduced VFe-protein of the vanadium nitrogenase of Azotobacter chroococcum (Acl*). Both the e.x.a.f.s and the x.a.n.e.s. (X-ray-absorption near-edge structure) are consistent with the vanadium being present as part of a VFeS cluster; the environment of the vanadium is not changed significantly in different oxidation states of the protein. The vanadium atom is bound to three oxygen (or nitrogen), three sulphur and three iron atoms at 0.215(3), 0.231(3) and 0.275(3) nm respectively.

Azotobacter↗
PubMed1989

Manganese catalyzed auto-oxidation of dopamine to 6-hydroxydopamine in vitro.

Manganese (Mn) is an essential trace element which, upon excessive exposure, produces a neurological syndrome similar to chronic Parkinson's disease in animals and humans. Previous work demonstrated that Mn was more potent than other transition metals in stimulating dopamine (DA) auto-oxidation. In these experiments, DA was incubated under physiological conditions in the presence and absence of Mn for up to 60 min. 6-Hydroxydopamine (6-OHDA) was produced in the presence of Mn, while the incubation mixture without Mn showed no DA oxidation. 6-Hydroxydopamine is a neurotoxicant which exerts its effects by destroying DA nerve terminals in the CNS. Therefore, this work suggests that the Mn catalyzed increase in DA auto-oxidation could be linked mechanistically to the appearance of Mn-induced neurotoxic effects.

Chlorides↗
PubMed1989

Inhibitory effect of benzene metabolites on nuclear DNA synthesis in bone marrow cells.

Effects of endogenously produced and exogenously added benzene metabolites on the nuclear DNA synthetic activity were investigated using a culture system of mouse bone marrow cells. Effects of the metabolites were evaluated by a 30-min incorporation of [3H]thymidine into DNA following a 30-min interaction with the cells in McCoy's 5a medium with 10% fetal calf serum. Phenol and muconic acid did not inhibit nuclear DNA synthesis. However, catechol, 1,2,4-benzenetriol, hydroquinone, and p-benzoquinone were able to inhibit 52, 64, 79, and 98% of the nuclear DNA synthetic activity, respectively, at 24 microM. In a cell-free DNA synthetic system, catechol and hydroquinone did not inhibit the incorporation of [3H]thymidine triphosphate into DNA up to 24 microM but 1,2,4-benzenetriol and p-benzoquinone did. The effect of the latter two benzene metabolites was completely blocked in the presence of 1,4-dithiothreitol (1 mM) in the cell-free assay system. Furthermore, when DNA polymerase alpha, which requires a sulfhydryl (SH) group as an active site, was replaced by DNA polymerase I, which does not require an SH group for its catalytic activity, p-benzoquinone and 1,2,4-benzenetriol were unable to inhibit DNA synthesis. Thus, the data imply that p-benzoquinone and 1,2,4-benzenetriol inhibited DNA polymerase alpha, consequently resulting in inhibition of DNA synthesis in both cellular and cell-free DNA synthetic systems. The present study identifies catechol, hydroquinone, p-benzoquinone, and 1,2,4-benzenetriol as toxic benzene metabolites in bone marrow cells and also suggests that their inhibitory action on DNA synthesis is mediated by mechanism(s) other than that involving DNA damage as a primary cause.

Animals↗
PubMed1988

Iron K-edge X-ray absorption spectroscopy of the iron-molybdenum cofactor of nitrogenase from Klebsiella pneumoniae.

Iron K-edge X-ray absorption data for the iron-molybdenum cofactor ('FeMoco') from Klebsiella pneumoniae reported here provide the first evidence for long-range structural order in the cofactor [Fe...Fe(Mo) = 0.368 nm in addition to Fe...S = 0.22 nm and Fe...Fe(Mo) = 0.27 nm] and, in contrast with previously published data [Antonio, Teo, Orme-Johnson, Nelson, Groh, Lindahl, Kauzlarich & Averill (1982) J. Am. Chem. Soc. 104, 4703-4705], indicate that most of the iron centres are not co-ordinated to light (oxygen, nitrogen) atoms. This demonstrates that presently available chemical models for FeMoco are inadequate.

Ferredoxins↗
PubMed1988

A proposed role played by benzene itself in the induction of acute cytopenia: inhibition of DNA synthesis.

A single intraperitoneal dose of benzene (880 mg/kg) in mice inhibited DNA synthesis of bone marrow cells within one hour postinjection. However, there was no inhibitory effect on the synthesis of heme and protein at that dosage. Dose-dependent inhibition of DNA synthesis by benzene was observed over the range of 440 to 1760 mg/kg, supporting the idea that cytopenia which was observed by others following multiple doses of benzene (e.g., 440 or 880 mg/kg) might be due to the inhibitory effect of benzene on DNA synthesis. In our studies, benzene concentrations above 81 micrograms/g wet bone marrow resulted in inhibition of DNA synthesis, regardless of whether it was given ip or by inhalation. The effect of benzene itself, rather than its toxic metabolites, on DNA synthesis was further seen in experiments using a bone marrow cell culture system and cell-free DNA synthetic system. Experimental results demonstrated that benzene alone was capable of inhibiting the DNA synthesis of bone marrow cells and that the reduced DNA synthesis resulted from the inhibitory effect of benzene on DNA polymerase alpha, the enzyme that catalyzes the last step of the DNA synthetic pathway. Thus, benzene itself could play a significant role in inducing myelotoxicity in the case of acute or subacute toxicity by exerting its inhibitory effect on DNA synthesis.

Administration, Inhalation↗