Hypocatalasemic Mice Research Articles

Mice Lacking Catalase Develop Normally but Show Differential Sensitivity to Oxidant Tissue Injury

Catalase plays a major role in cellular antioxidant defense by decomposing hydrogen peroxide, thereby preventing the generation of hydroxyl radical by the Fenton reaction. The degree of catalase deficiency in acatalasemic and hypocatalasemic mice varies from tissue to tissue. They therefore may not be suitable for studying the function of this enzyme in certain models of oxidant-mediated tissue injury. We sought to generate a new line of catalase null mice by the gene targeting technique. The mouse catalase (Cat or Cas1) gene was disrupted by replacing parts of intron 4 and exon 5 with a neomycin resistance cassette. Homozygous Cat knockout mice, which are completely deficient in catalase expression, develop normally and show no gross abnormalities. Slices of liver and lung and lenses from the knockout mice exhibited a retarded rate in decomposing extracellular hydrogen peroxide compared with those of wild-type mice. However, mice deficient in catalase were not more vulnerable to hyperoxia-induced lung injury; nor did their lenses show any increased susceptibility to oxidative stress generated by photochemical reaction, suggesting that the antioxidant function of catalase in these two models of oxidant injury is negligible. Further studies showed that cortical injury from physical impact caused a significant decrease in NAD-linked electron transfer activities and energy coupling capacities in brain mitochondria of Cat knockout mice but not wild-type mice. The observed decrease in efficiency of mitochondrial respiration may be a direct result of an increase in mitochondrion-associated calcium, which is secondary to the increased oxidative stress. These studies suggest that the role of catalase in antioxidant defense is dependent on the type of tissue and the model of oxidant-mediated tissue injury.

Spontaneous Tumors in Hypocatalasemic C3H/ C<sup>b</sup><sub>s</sub> /Gen and C3H Mice

Hypocatalasemic mice (Cbs) were examined for their life span and compared to C3H mice with reference to oxidase enzyme activity and spontaneous tumor development. N1-methylnicotinamice oxidase, xanthine oxidase (XO), xanthine dehydrogenase (XDH), and XO+XDH activities in 8 week-old male Cbs mice were decreased as compared to the C3H counterparts. On the other hand, benzaldehyde oxidase in both sexes and xanthine dehydrogenase activity in females were significantly elevated as compared to the C3H. The total tumor incidence, number of tumors per mouse, liver tumors, and other tumor induction were significantly greater in as compared with until 499 days group in both sexes (P<0.01). The total incidence of tumors and incidence of liver tumors were not significantly different between the two strains in both sexes. The number of tumors per animal in C3H females and incidence of sarcoma in both sexes were significantly decreased as compared with the Cbs case but the numbers of liver tumor per animal and ovarian tumors were significantly greater.

Animal model for oxidative stress research—Catalase mutant mice

Catalase-deficient mouse strains was initially established by Feinstein et al. through a large scale screening of the progeny of irradiated C3H mice in 1966. Later, Feinstein provided the mice of catalase mutant strain C3H/AnICs(a)Cs(a) (wild-type), C3H/AnICs(b)Cs(b) and C3H/AnlCs(c)Cs(c) to Okayama University Medical School in Japan. It is known that a point mutation at amino acid 11 (from glutamine to histidine) of acatalasemic mouse catalase and a point mutation at amino acid 439 (from as paragine to serine) of hypocatalasemic mouse catalase are responsible for the catalase deficiency of acatalasemic and hypocatalasemic mice, respectively. Recently, a liver cell line from an acatalasemic mouse andEscherichia coli (E. coli) strains with murine normal, hypocatalasemic, or acatalasemic catalase have been established. The construction of these new systems would be useful for studying the effects of oxidative stress at the cellular level. In this review, we give a brief overview of recent findings of studies in utilizing the catalase-deficient mice and evaluate the possibility of these mouse strains as a candidate animal model for oxidative stress research.

CDNA cloning and expression of mutant catalase from the hypocatalasemic mouse: comparison with the acatalasemic mutant

Mutant catalase cDNAs from the hypocatalasemic and acatalasemic mice were cloned and expressed in bacteria. A novel missense mutation, Asp (AAT) to Ser (AGT), was identified at amino acid position 439 of the hypocatalasemic catalase. Analysis of recombinant catalase mutants revealed that the mutation is responsible for the reduced activity of hypocatalasemic catalase and the unstable tetrameric structure of acatalasemic catalase was also suggested.

Prevention of mammary tumorigenesis in acatalasemic mice by vitamin E supplementation.

Adult male and female acatalasemic (C3H/AnLCsbCsb), hypocatalasemic (C3H/AnLCscCsc) and normal mice of C3H strain fed on regular laboratory chow for 15 months showed an increased incidence of spontaneous mammary tumor in the decreasing order of female acatalasemic, male acatalasemic, female hypocatalasemic and male hypocatalasemic mice. Normal mice did not develop mammary tumor. We conducted a prospective study with female acatalasemic mice, which showed the highest incidence of mammary tumor, to examine the preventive effect of vitamin E on mammary tumor. Female acatalasemic mice were fed on vitamin E‐deficient (28 animals) and vitamin E‐supplemented diet (25 animals) for 29 months. The incidence of mammary tumor in mice given the vitamin E‐supplemented diet was 47%, while that in mice given vitamin E‐deficient diet was 82% (P<0.002). Mammary tumors were apparent after 9 months of vitamin E deprivation and after 14 months of vitamin E supplementation. Female normal mice did not develop mammary tumor during a comparable period of time. The mean catalase activity of mammary gland in acatalasemic mice was 18.8% of that in normal mice. The results indicate that vitamin E protects acatalasemic mice against the development of mammary tumor.

Invasions of Stomach Tumors in Hypocatalasemic Mice (C3H/Cbs/Gen) Treated with N-Methyl-N-Nitrosourea but Not N-Methyl-N'-Nitro-N-Nitrosoguanidine.

Hypocatalasemic mice were treated with N-methyl-N-nitrosourea (MNU) in their drinking water or gastric intubation for 16 weeks or were given N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) in their drinking water for 6 months. With MNU treatment in the drinking water, well differentiated tumors were observed in the glandular stomachs of one male (4%) and 5 female (15%) animals. Sarcomas of the dermis also appeared in 10 males (40%). Adrenal lesions developed in I I females (35%) along with kidney, ovary, and uterus tumors. In the Group given MNU intubation, squamous cell carcinomas of the forestomach appeared from 122 days after the first treatment, reaching incidences of 81% in males, 94% in females, with invasion of the diaphragm in 3 male cases and the diaphragm, liver, pancreas, or adrenal in I I female cases. Glandular stomach tumors occurred in 12 males (52%), with 3 being signet ring cell carcinomas, and in 12 females (35%). In a sub-Group undergoing forestomach removal after MNU intubation esophagectasis appeared from 80 days after the operation at incidences of 63% in males and 74% in females. The glandular stomach tumors including 3 signet ring cell carcinomas were found in 17 of 46 (40%) males, 2 of 18 (11%) females demonstrated lesion, one being a signet ring cell carcinoma. In the Group receiving MNNG treatment, anemia appeared from the 5.5 month time point and no gastric tumors were observed. The present results do not support the conclusion that MNU induction of glandular stomach lesions in hypocatalasemic mice may provide a useful model of signet ring cell carcinomas

STRAIN DIFFERENCES IN MOUSE SKIN TUMORIGENESIS BY THE PROMOTION OF 12-O-TETRADECANOYLPHORBOL-13-ACETATE OR BENZOYL PEROXIDE

Strain difference of mouse skin tumorigenesis was studied with 12-O-tetradecanoylphorbol-13-acetate (TPA) and benzoyl peroxide (BzP) both in 8-week-old hypocatalasemic C3H mice (C-s(b)) and in normal C3H/HeNCrj mice (C3H). Two weeks after initiation by 4 mu mole MNNG, both strains of female mice were promoted by 10 nmole TPA or 10 mg, 20 mg BzP twice weekly for 30 and 52 weeks. The incidence of skin tumors in C-b(s) was significantly increased as compared to that in C3H mice promoted by TPA for 30 weeks but after 52 weeks of TPA promotion there was no difference between the 2 strains. By the promotion with 10 mg or 20 mg BzP, the incidence of skin tumor in C3H was 5% at both levels after 30 weeks of promotion and 19% and 38%, respectively, after 52 weeks of promotion. No skin tumors appeared in C-b(s) mice with BzP treatment after 30 weeks of promotion. After 52 weeks of promotion by 10 mg BzP, skin tumors were induced in only 15% of C-b(s) mice. The results show that genetic factors, especially radical scavenging enzymes, controlled susceptibility to skin tumorigenesis by promotion with either TPA or BzP.

Exhaled metallic mercury in acatalasemic, hypocatalasemic and normal mice injected with mercury (II) chloride.

To clarify the relationship between the catalase activity in mouse organs and the amounts of metallic mercury exhaled, normal, homozygous hypocatalasemic and acatalasemic mice were injected with mercuric chloride. The cumulative amount of metallic mercury exhaled by mice was evidently expressed in the descending order of acatalasemic, hypocatalasemic, and normal mice. Statistically significant differences in the cumulative exhaled metallic mercury levels were observed between acatalasemic and hypocatalasemic mice, between normal and hypocatalasemic mice, and between acatalasemic and normal mice using the method of one way analysis of variance (ANOVA). A linear relationship was obtained through logarithm of catalase activity in the lungs or the blood, and logarithm of the cumulative amount of the exhaled mercury.

無および低カタラーゼ血症マウスのカタラーゼ分子の性質に関する研究

無および低カタラーゼ血症マウスのカタラーゼ分子の性質に関する研究

変異マウス血液中のカタラーゼ活性度とメトヘモグロビン濃度の関係

変異マウス血液中のカタラーゼ活性度とメトヘモグロビン濃度の関係

Erythrocyte catalase. A somatic oxidant defense?

Mammalian erythrocytes have large amounts of catalase, an enzyme which catabolizes hydrogen peroxide (H2O2). Because catalase has a low affinity for H2O2, others have suggested that glutathione peroxidase clears most H2O2 within the erythrocyte and that catalase is of little import. We hypothesized that erythrocyte catalase might function to protect heterologous somatic cells against challenge by high levels of exogenous H2O2 (e.g., in areas of inflammation). We find that, whereas nucleated cells (L1210 murine leukemia) are readily killed by an enzymatically generated flux of superoxide (and, therefore, H2O2), the addition of human and murine erythrocytes blocks lethal damage to the target cells. Inhibition of erythrocyte superoxide dismutase, depletion of glutathione, and lysis of the erythrocytes do not diminish this protection. However, inhibition of erythrocyte catalase abrogates the protective effect and the addition of purified catalase (but not superoxide dismutase) restores it. Furthermore, erythrocytes derived from congenitally hypocatalasemic mice (in which other antioxidant systems are intact) do not protect L1210 cells. Our results raise the possibility that the erythrocyte may serve as protection against by-products of its own cargo, oxygen.

無および低カタラーゼ血症マウスのカタラーゼ分子の性質に関する研究

無および低カタラーゼ血症マウスのカタラーゼ分子の性質に関する研究

On the turnover and proteolysis of catalase in tissues of the guinea pig and acatalasemic mice

Turnover characteristics (half-lives and rate constants for synthesis and degradation) have been determined for the catalases of guinea pig and three different strains of mice by means of the kinetics of return of enzyme activity after inhibition with 3-amino-1,2,4-triazole. The catalase of hypocatalasemic mice (strain C sD) did not display an appreciably different half-life to that of the wild-type mice, but catalase in the tissues of acatalasemic mice (strain C sB) exhibited a half-life which was only half that of the wild type, while the half-life of guinea pig catalase was more than twice that of wild-type mice. Significant differences were also noticed in regard to the in vitro susceptibility of the catalases of these animals to protease inactivation. Large-granule (lysosomal, mitochondrial and peroxisomal) extracts proved far more susceptible to protease inactivation than cytosol extracts, and marked changes in the heteromorph pattern of mouse liver cytosol catalase were observed to accompany limited proteolysis. These results support the conclusion that the in vitro susceptibility of proteases may be an important determining factor in the rate of degradation of an enzyme in vivo.

Nature of the heterozygote blood catalase in a hypocatalasemic mouse mutant

The blood catalase of a hypocatalasemic mouse mutant has been compared with that of the wild-type (normal) animal and with that of the heterozygote. Comparison is on the basis of stability to heat and to urea. Electrophoretic evidence is of no value, because all forms tested show the same mobility. Because the heterozygote heat and urea inactivation curves differ from those of the two parental forms, and because the curves are smoothly S-shaped, with no shoulders or other irregularities, it is suggested that this heterozygote produces only a single molecular form of the enzyme.

Acatalasemic and hypocatalasemic mouse mutants: II. Mutational variations in blood and solid tissue catalases

In a series of one acatalasemic and four hypocatalasemic mouse strains, the catalase of blood, liver, and kidney has been compared with regard to sensitivity to heat, pH extremes, ionizing radiation, and a variety of chemical reagents. It is concluded that the catalase of the normal, or wild type, strain differs qualitatively from all the mutants, and that the catalase of the five mutants all differ amongst themselves. The catalase of one pair of hypocatalasemic mutants shows so little difference in behavior that the two molecular species are possibly identical.