Effects of post low-dose X-ray irradiation on carbon tetrachloride-induced acatalasemic mice liver damage

The catalase activities in the blood and organs of the acatalasemic (C3H/AnLCsb-Csb) mouse of the C3H strain are lower than those of the normal (C3H/AnLCSa-Csa) mouse. We examined the effects of post low-dose (0.5 Gy) X-ray irradiation which reduced the oxidative damage under carbon tetrachloride-induced hepatopathy in acatalasemic or normal mice. As a result, the 0.5 Gy irradiation after carbon tetrachloride administration decreased the glutamic oxaloacetic and glutamic pyruvic transaminase activity in the acatalasemic mouse blood to a level similar to that of the acatalasemic mouse blood not treated with carbon tetrachloride; this is in contrast to a high-dose (15 Gy) irradiation. In the same manner, pathological disorder was improved by 0.5 Gy irradiation. The fat degeneration in normal mice was quickly reduced, in contrast to acatalasemic mice. These findings suggest that low-dose irradiation after carbon tetrachloride administration accelerates the rate of recovery and that catalase plays an important role in the recovery from hepatopathy induced by carbon tetrachloride, in contrast to high-dose irradiation.     

Methanol metabolism in acatalasemic mice

The methanol metabolism in acatalasemic mice was studied by administering [14C]methanol and [14C]formic acid to acatalasemic and normal mice and determining the radioactivity of exhaled carbon dioxide. Methanol metabolism was also studied in acatalasemic and normal mice treated with 3-amino-1,2,4-triazole (AT), which is known to be an inhibitor of catalase (EC 1.11.1.6). The metabolism of methanol and formic acid was inhibited in acatalasemic mice as seen by reduced [14C]CO2 production. Similar results were obtained when AT was given prior to the methanol injection into the normal and acatalasemic mice. The results indicate the peroxidative activity of catalase plays the major role in the methanol metabolism in mice. On the other hand similar studies with [1-14C] ethanol showed that the metabolism of ethanol was not inhibited in acatalasemic mice.     

Adjustment function among antioxidant substances in acatalasemic mouse brain and its enhancement by low-dose X-ray irradiation

The catalase activities in blood and organs of the acatalasemic (C3H/AnLCsbCsb) mouse of the C3H strain are lower than those of the normal (C3H/AnLCsaCsa) mouse. We conducted a study to examine changes in the activities of antioxidant enzymes, such as catalase, superoxide dismutase (SOD) and glutathione peroxidase (GPX), the total gluathione content, and the lipid peroxide level in the brain, which is more sensitive to oxidative stress than other organs, at 3, 6, or 24 hr following X-ray irradiation at doses of 0.25, 0.5, or 5.0 Gy to the acatalasemic and the normal mice. No significant change in the lipid peroxide level in the acatalasemic mouse brain was seen under non-irradiation conditions. However, the acatalasemic mouse brain was more damaged than the normal mouse brain by excessive oxygen stress, such as a high-dose (5.0 Gy) X-ray. On the other hand, we found that, unlike 5.0 Gy X-ray, a relatively low-dose (0.5 Gy) irradiation specifically increased the activities of both catalase and GPX in the acatalasemic mouse brain making the activities closer to those in the normal mouse brain. These findings may indicate that the free radical reaction induced by the lack of catalase is more properly neutralized by low dose irradiation.     

The effect of pneumonia induced in mice with Mycoplasma pulmonis on resistance to subsequent bacterial infection and the effect of a respiratory infection with Sendai virus on the resistance of mice to Mycoplasma pulmonis.

The effect of pneumonia induced by Mycoplasma pulmonis in mice on the resistance of the lung to additional bacterial infection was examined. The effect of pneumonia induced by Sendai virus on the resistance of mice to M. pulmonis was also investigated and compared with the effect of Sendai virus on resistance to Staphylococcus aureus. Sendai virus infection decreased subsequent resistance to M. pulmonis in proportion to the virus dose. Decreased resistance to subsequent S. aureus and M. pulmonis infection was greatest at about the same time after inoculation of virus and was related to virus-induced lesions. Besides affecting the resistance of mice to subsequent mycoplasma infection, Sendai virus could enhance an existing mycoplasma infection. Pneumonia induced by M. pulmonis did not decrease resistance to subsequent bacterial infection. The mechanism whereby Sendai virus decreases host resistance is therefore similar for bacteria and mycoplasmas, but pneumonia induced by mycoplasmas does not have the same effect.     

Role of Pasteurella pneumotropica and Mycoplasma pulmonis in Murine Pneumonia

Pneumonia caused by Mycoplasma pulmonis and Pasteurella pneumotropica was studied in conventional, specific pathogen-free (SPF), and germ-free mice. When P. pneumotropica was serially passed in conventional mice, M. pulmonis, as well as P. pneumotropica, was recovered from mice with gross lesions. When M. pulmonis was serially passed in conventional mice, both organisms were recovered. SPF mice given a nasal instillation of M. pulmonis alone, P. pneumotropica alone, or a combination of the two developed pneumonia when both organisms were present. These findings suggested that both organisms contribute to typical murine pneumonia. That M. pulmonis might be an L form of P. pneumotropica was suggested because some SPF mice inoculated with either organism yielded both on culture. This possibility was investigated with mole per cent guanine plus cytosine (GC) content and nucleic acid hybridization techniques. The GC content of P. pneumotropica is 42.2 mole per cent and that of M. pulmonis is 28.6 mole per cent. No specific hybrids between deoxyribonucleic acid (DNA) from M. pulmonis and DNA from P. pneumotropica were detected. This and the wide disparity in GC content showed that M. pulmonis is not an L form of P. pneumotropica. In germ-free mice, intranasal instillation with either organism alone produced pneumonia. The lesions produced when each organism was inoculated independently were characterized by areas of consolidation with perivascular and peribronchial lymphocytic infiltration. Qualitatively, the lesions produced when both organisms were inoculated simultaneously more closely resembled those seen in naturally occurring murine pneumonia. Statistical analysis indicated that the quantitative effect of the two organisms was additive. The indirect fluorescent antibody technique was used to locate organisms in lung tissue sections. M. pulmonis localized in the bronchial epithelium and P. pneumotropica localized in the alveolar lesions.     

A quantitative genetic analysis of tissue-specific catalase activity inMus musculus

Tissue-specific catalase activity in 3-week-old animals from inbred mouse strains 129/ReJ, BALB/c, C3H/HeAnl/Cas-1^b, C3H/HeSnJ, C3H/S, C57BL/6J, and Swiss-Webster was found to be highly variable by analysis of variance (P=0.01). Appropriate crosses were made among strains which were classified as normal (BALB/c, C3H/HeSnJ, C3H/S), hypocatalasemic (129/ReJ, C57BL/6J), and acatalasemic (C3H/HeAnl/Cas-1^b) with respect to blood catalase activity to study the inheritance of the blood, kidney, liver, and lung catalase activity levels in a number of generations (reciprocal F₁'s, F₂, two backcrosses —BC₁ and BC₂— and some RI lines). Segregation analysis and statistical methods which tested different models of inheritance as well as calculations of heritability were used in an effort to assess and evaluate genetic parameters that affect catalase activity. Results indicate that the inheritance of blood catalase activity in the cross involving acatalasemic and normal (BALB/c, C3H/HeSnJ) strains is compatible with the single-locus difference between the parental strains; however, the difference between the acatalasemic and the hypocatalasemic strain (C57BL/6J) would require additional genetic interaction for a satisfactory explanation. A similar pattern of generalization also applies to the inheritance of kidney catalase activity. The segregation pattern for the liver and lung catalase activity in most crosses is significantly different from the expectations of the single locus model. These results are compatible with the concept that a number of genes must affect tissue-specific catalase activity in mice. These may include previously described (e.g., Ce-1 and Ce-2) or novel genetic regulators/modifiers which interact with a single structural gene (Cas-1) or its product to produce the catalase phenotype characteristic of specific tissues in each strain.This investigation was supported by a Natural Sciences and Engineering Research Council of Canada operating grant to S.M.S.     

Immunotitration of the catalase in the blood of Japanese subjects and mice suffering from acatalasemia and hypocatalasemia.

Catalase in hemolysates of normal, heterozygous hypocatalasemic and acatalasemic Japanese was immunotitrated with an anti-human blood catalase rabbit serum. Equivalence points were calculated from the regression lines between catalase activity added and catalase activity remaining in the supernatant. Catalase activities at the equivalence points of Japanese normal, hypocatalasemia and acatalasemia were similar. The results indicate that the specific activities of catalase in the normal and of the variant bloods are identical. Catalase in hemolysates of normal and variant mice was immunotitrated with an anti-mouse liver catalase rabbit serum. In contrast to Japanese acatalasemic subject, the equivalence points of catalase in heterozygous hypocatalasemic, homozygous hypocatalasemic, acatalasemic and normal hemolysates were different, and the ratios of specific activity in these variant mice to that in normal were 0.72, 0.46 and 0.21, respectively. The differences in catalase activities at equivalence points were also supported by the statistical analysis on parameters of regression lines of catalase activities remaining in the supernatant on catalase activities added in the immunotitration. These findings suggest that the molecular properties of residual catalase of Japanese acatalasemia and those of mouse acatalasemia are entirely different.     

Acatalasemia

Summary The abnormalities in acatalasemia at the gene level as well as properties of the residual catalase in Japanese acatalasemia are historically reviewed. The replacement of the fifth nucleic acid, guanine, in the fourth intron by adenine in the acatalasemic gene causes a splicing mutation and hence a deficiency of mRNA. The guanine-to-adenine substitution was detected in two Japanese acatalasemic cases from different families. The properties of the residual catalase are similar to those of normal catalase; the exons are identical. The properties of the residual catalase and the molecular defect in the catalase gene are compared among Japanese, Swiss, and mouse acatalasemias. The physiological role of catalase, as judged from human acatalasemic blood and acatalasemic mice, is also described.     

Variation in the virulence of strains of Mycoplasma pulmonis related to susceptibility to killing by macrophages in vivo.

The virulence of five strains of Mycoplasma pulmonis, as judged by their ability to survive in the respiratory tract and induce pneumonia in CBA mice, was related to the ability of viable organisms to persist in the peritoneal cavity. This appeared to be the result of differences in the ability of the strains to resist killing by peritoneal macrophages in vivo. It is suggested that resistance to phagocytosis by macrophages is an important determinant of virulence for M. pulmonis.     

Purification and characterization of liver catalase in acatalasemic beagle dog: comparison with normal dog liver catalase

Catalase from acatalasemic dog liver was purified to homogeneity and its properties were compared with those of normal dog liver catalase. The purified acatalasemic and normal dog liver catalases were found to have the same molecular weight (230,000 Da) and isoelectric point (pI: 6.0-6.2) and both enzymes contained four hematins per molecule. The catalytic activity of catalase from acatalasemic dog was normal. Furthermore, there was no difference between the acatalasemic and normal dog catalases in the binding affinity to NADPH (apparent Kd: 0.11-0.12 microM) and in the sensitivity to oxidative stress by hydrogen peroxide, the normal substrate of catalase. The acatalasemic dog enzyme was stable only in a narrow pH range (pH 6-9) although the normal enzyme was stable in a wide pH range (pH 4-10). Acatalasemic dog liver catalase also showed a slight low thermal stability at 37 degrees C and the heat-lability was remarkable at 45 degrees C, compared to the normal dog enzyme. These results indicated that the acatalasemic dog catalase is catalytically normal although it is associated with an unstable molecular structure.     

The peroxidatic and catalatic activity of catalase in normal and acatalasemic mouse liver

Monomeric, dimeric and tetrameric forms of mouse liver catalase have been shown to express peroxidatic activity while the tetrameric form expresses the catalic activity. Autosomally inherited acatalasemia, produced by X-ray irradiation of mice results in almost complete loss of catalic activity of catalase but has no effect on the peroxidatic activity. Liver catalase from normal and acatalasemic mice was purified by following the catalic and peroxidatic activity, respectively. Antiserum produced in rabbit against catalase from normal mouse completely precipitated the catalatic and peroxidatic activity from normal liver, and peroxidatic activity from the acatalasemic liver homogenate. Similar results were obtained when antiserum against peroxidase from acatalasemic mice was used. These studies indicate that acatalasemia in mice is due to a structural gene mutation which leads to synthesis of structurally altered catalase subunits. The altered subunits express peroxidatic activity but do not combine to form a tetramer which expresses catalatic activity.     

Levels of metallic mercury and mercuric ion in the venous and arterial bloods of normal and acatalasemic mice following exposure to mercury vapor

Levels of metallic mercury and mercuric ion in the arterial and venous bloods of normal and acatalasemic mice exposed to metallic mercury vapor in vitro and in vivo were investigated. Mercury uptake in venous blood from air saturated with mercury vapor with or without hydrogen peroxide in vitro was determined. Level of mercuric ion in venous blood of normal mice was significantly higher than that of acatalasemic mice. By contrast, metallic mercury in venous blood of acatalasemic mice was elevated relative to level in normal mice. Metallic mercury level in red blood cells and plasma was also significantly higher in acatalasemic mice. The ratio of metallic mercury to total mercury (Hg degrees + Hg2+) in the arterial and venous bloods of acatalasemic mice exposed to metallic mercury vapor was increased relative to normal mice. This ratio in red blood cells and plasma in the venous bloods of acatalasemic mice in vivo was also significantly higher than those of normal mice. The significance of metallic mercury in plasma for distribution of mercury in organs is discussed.     

Metallic mercury in the arterial blood of normal and acatalasemic mice exposed to metallic mercury vapor

Concentration of metallic mercury in the arterial blood was higher in acatalasemic mice after exposure to 3.45 mg/m3 for 10 minutes in comparison with normal mice, whereas concentration of mercuric ion was lower in acatalasemic mice than in normal mice. Thus, the ratio of metallic mercury to total mercury in the arterial blood of acatalasemic mice was 5.86 +/- 3.61%, which was statistically significantly higher than the value (1.36 +/- 0.65%) of corresponding normal mice. The mercury concentration and distribution in the brain and liver of acatalasemic mice were higher than those in normal mice. Data indicate that the concentration of metallic mercury in the arterial blood of acatalasemic mice was higher than that of normal mice and that metallic mercury soluble in lipids is likely transferred to the brain and liver from the blood. Conclusively, metallic mercury in the arterial blood is the biologically active form for transferring mercury from blood to organs.     

Exhalation of metallic mercury by acatalasemic, hypocatalasemic and normal mice exposed to metallic mercury vapor.

In order to elucidate the amounts of metallic mercury exhaled from mice having different amounts of catalase activity, normal, homozygous hypocatalasemic and acatalasemic mice were exposed to radioactive metallic mercury vapor at three levels of concentrations (0.072, 0.144, and 0.297 mg/m3). The timed and cumulative amounts of metallic mercury exhaled from the mice were used to classify them, in the descending order, as being acatalasemic, hypocatalasemic, and normal at the three environmental mercury concentrations. The statistical differences in the mean values among the acatalasemic, hypocatalasemic, and normal mice were calculated by the use of one-way ANOVA and multiple comparison of the mean values by Tukey's method. The mean values of the timed amounts of exhaled metallic mercury were found to be significantly different (P less than 0.05) among the three kinds of mice several hours after exposure, and those of the cumulative amounts of exhaled metallic mercury were in the descending order of acatalasemic, hypocatalasemic and normal mice, and were significantly different among the three types of mice almost over the entire time course. Thus, negative correlation coefficients were obtained between the logarithms of the catalase activity in the lungs and the blood, and the logarithms of the cumulative amounts of exhaled metallic mercury.     

Characterization of acatalasemia detected in two Hungarian sisters.

Acatalasemia was detected in 2 sisters of a Hungarian family. The pedigree of the family showed hypocatalasemia in the children of the patients and in 1 of their brothers, while the other members of the family had normal blood catalase activity. The biochemical characterization (catalase activity, electrophoretic migration, isoelectric point and enzyme stability) of the blood as well as tissue catalase of the acatalasemic patients yielded a catalase form which did not differ from normal.     

Mycoplasma pulmonis depresses humoral and cell-mediated responses in mice

Humoral and cell-mediated immune responses to sheep red blood cells (SRBC) were studied in mice infected experimentally with Mycoplasma pulmonis. The hemagglutinating (HA) antibody against SRBC was evaluated at 0, 3, 5, 7, 14, 21 and 28 days postinfection (PI). Antibody tiers during all days PI were depressed significantly (p less than 0.05) in infected mice as compared to noninfected controls. The HA antibody, which is of the IgM class, peaks at day 5 PI. There is no shift in the kinetics of the humoral response in M. pulmonis infected mice. Cellular immune responses were evaluated by a delayed-type hypersensitivity (DTH) reaction and the lymphocyte transformation technique. Mice were sensitized at 0,3,5,7,14, 21 and 28 days PI with SRBC and challenged by footpad injection of SRBC 7 days later. The DTH reaction measured at 24 hours after challenge was depressed significantly (p less than 0.05) in all infected animals. After a transient enhancement on day 3 PI, the DTH responses remained depressed through day 28 PI. The lymphocyte transformation test showed a significantly (p less than 0.05) depressed response except on days 5 and 7 PI. These results indicate that M. pulmonis infection in mice suppresses the humoral antibody and cell-mediated immune responses.     

Alterations in lung prostaglandin synthesis and release in murine respiratory mycoplasmosis

Pathogenetic mechanisms in murine respiratory mycoplasmosis are poorly understood; however, non-specific immune responses appear to be important in controlling the growth of Mycoplasma pulmonis in vitro. To date, no study has examined the role of pulmonary prostaglandin production during the development of M. pulmonis infection. The present study was designed to determine if alterations in pulmonary prostaglandin synthesis and release occur in M. pulmonis infection and the possible role for prostaglandins in the modulation/pathogenesis of murine respiratory mycoplasmosis. Ten to 20 days after intranasal inoculation of pathogen-fee F344 rats with M. pulmonis, lung lavage concentrations of prostaglandin E (PGE) and thromboxane A2 (TxA2) were significantly elevated. To confirm a role for prostaglandins in the pathogenesis of murine mycoplasmosis we blocked the cyclo-oxygenase pathway with indomethacin. Indomethacin-treated rats had significantly lower lavage levels of PGE and TxA2 and significantly increased numbers of M. pulmonis in the lung. These data indicate that prostaglandins may be involved in the pathogenesis of murine respiratory mycoplasmosis, possibly through alteration of mycoplasmacidal and/or mycoplasmastatic mechanisms.     

Role of Cu,Zn-SOD in the synthesis of endogenous vasodilator hydrogen peroxide during reactive hyperemia in mouse mesenteric microcirculation in vivo

We have recently demonstrated that endothelium-derived hydrogen peroxide (H2O2) is an endothelium-derived hyperpolarizing factor and that endothelial Cu/Zn-superoxide dismutase (SOD) plays an important role in the synthesis of endogenous H2O2 in both animals and humans. We examined whether SOD plays a role in the synthesis of endogenous H2O2 during in vivo reactive hyperemia (RH), an important regulatory mechanism. Mesenteric arterioles from wild-type and Cu,Zn-SOD(-/-) mice were continuously observed by a pencil-type charge-coupled device (CCD) intravital microscope during RH (reperfusion after 20 and 60 s of mesenteric artery occlusion) in the cyclooxygenase blockade under the following four conditions: control, catalase alone, N(G)-monomethyl-L-arginine (L-NMMA) alone, and L-NMMA + catalase. Vasodilatation during RH was significantly decreased by catalase or L-NMMA alone and was almost completely inhibited by L-NMMA + catalase in wild-type mice, whereas it was inhibited by L-NMMA and L-NMMA + catalase in the Cu,Zn-SOD(-/-) mice. RH-induced increase in blood flow after L-NMMA was significantly increased in the wild-type mice, whereas it was significantly reduced in the Cu,Zn-SOD(-/-) mice. In mesenteric arterioles of the Cu,Zn-SOD(-/-) mice, Tempol, an SOD mimetic, significantly increased the ACh-induced vasodilatation, and the enhancing effect of Tempol was decreased by catalase. Vascular H(2)O(2) production by fluorescent microscopy in mesenteric arterioles after RH was significantly increased in response to ACh in wild-type mice but markedly impaired in Cu,Zn-SOD(-/-) mice. Endothelial Cu,Zn-SOD plays an important role in the synthesis of endogenous H(2)O(2) that contributes to RH in mouse mesenteric smaller arterioles.     

Sensitization to alloxan-induced diabetes and pancreatic cell apoptosis in acatalasemic mice

Human acatalasemia may be a risk factor for the development of diabetes mellitus. However, the mechanism by which diabetes is induced is still poorly understood. The impact of catalase deficiency on the onset of diabetes has been studied in homozygous acatalasemic mutant mice or control wild-type mice by intraperitoneal injection of diabetogenic alloxan. The incidence of diabetes was higher in acatalasemic mice treated with a high dose (180 mg/kg body weight) of alloxan. A higher dose of alloxan accelerated severe atrophy of pancreatic islets and induced pancreatic β cell apoptosis in acatalasemic mice in comparison to wild-type mice. Catalase activity remained low in the acatalasemic pancreas without the significant compensatory up-regulation of glutathione peroxidase or superoxide dismutase. Furthermore, daily intraperitoneal injection of angiotensin II type 1 (AT1) receptor antagonist telmisartan (0.1 mg/kg body weight) prevented the development of alloxan-induced hyperglycemia in acatalasemic mice. This study suggests that catalase plays a crucial role in the defense against oxidative-stress-mediated pancreatic β cell death in an alloxan-induced diabetes mouse model. Treatment with telmisartan may prevent the onset of alloxan-induced diabetes even under acatalasemic conditions.