Genetic control of glucuronidase in mice

Two electrophoretically distinct components of glucuronidase have been resolved in extracts of murine liver. A mutation which results in a ten-fold reduction in liver glucuronidase activity leads to a reduction in intensity of both components. Since the substrate dependencies of both mutant (gg) and wild-type (GG) glucuronidase have previously been shown to be identical, it is suggested that the mutation results in a decreased concentration of liver glucuronidase molecules. Furthermore, the anodal electrophoretic component is shown to be preferentially localized in lysosomes, while the cathodal component is primarily microsomal. The difference in mobility of the two components may reflect a difference in membranous elements attached to the extracted glucuronidase molecule.This paper was presented at a symposium entitled Genetic Control of Mammalian Metabolism held at The Jackson Laboratory, Bar Harbor, Maine, June 30–July 2, 1969. The symposium was supported in part by an allocation from NIH General Research Support Grant FR 05545 from the Division of Research Resources to The Jackson Laboratory.This investigation was supported by a grant from the Pharmaceutical Manufacturers Association Foundation.     

Genetic control of glucuronidase induction in mice

The β-glucuronidase activity of mouse kidney proximal tubule cells increases rapidly after administration of dihydrotestosterone. Several inbred mouse strains show an approximately fourfold greater response in enzyme activity than the majority of strains, although both groups have similar uninduced kidney glucuronidase activity. This difference is maintained throughout a three-week induction period. It is not accounted for by a difference in any of the physiological parameters (e.g. hypertrophy, inducer specificity, enzyme secretion) associated with enzyme induction. The difference is specific to glucuronidase; assays of other androgen-indueible enzymes showed no difference between the two groups. Induction does not involve a change in enzyme structure since basal and induced glucuronidase have identical thermal stability and immunochemical reactivity.Rates of enzyme synthesis were determined by assaying the incorporation of radiolabeled leucine into antibody-purified glucuronidase. The rate of enzyme synthesis increases after induction, and the more rapidly inducing strains have a correspondingly greater increase in the rate of enzyme synthesis.The inducibility difference between the two classes of inbred strains segregates as a single Mendelian trait in both backcross and F₂ progeny. Recombination studies with a coat color mutation closely linked to the enzyme structural gene and with a mutant of the enzyme structural gene altered in electrophoretic mobility showed that the inducibility gene, called Gur, maps in the region of the glucuronidase structural gene. Tests in heterozygotes showed that the Gur locus acts cis, affecting only the rate of synthesis of glucuronidase coded by the structural gene residing on the same chromosome.     

Changes in uterine phosphatase levels in mice deprived of LH during early pregnancy.

Mice injected with normal rabbit serum on Day 4 of gestation showed a progressive increase in specific alkaline and acid phosphatase activities on Days 6 and 7. Mice injected with antiserum to LH on Day 4 showed a significant decrease in specific alkaline phosphatase activity estimated biochemically on Days 6, 7 and 8, but this decrease was histochemically evident only on Day 8. Similarly, acid phosphatase activity in antiserum-treated mice was significantly increased on Days 6, 7 and 8 when estimated biochemically, but only on Day 8 when studied histochemically.     

Relationships between levels of membrane-bound glucuronidase and the associated protein egasyn in mouse tissues

Mouse beta-glucuronidase has a dual intracellular localization, being present in both endoplasmic reticulum and lysosomes of several tissues. Previous studies demonstrated that the protein egasyn is complexed with microsomal but not lysosomal glucuronidase and that a mutant lacking egasyn is deficient in microsomal, but not lysosomal, glucuronidase. By means of a recently developed radioimmunoassay for egasyn, the relationship between microsomal glucuronidase levels and egasyn levels has been examined in various adult tissues, during postnatal development in liver, and after androgen induction of glucuronidase in kidney. The results indicate that the relative availability of egasyn determines the balance between glucuronidase incorporation into membranes and that into lysosomes.     

低氧性肺动脉高压小鼠体内脂质水平的变化

目的：观察低氧性肺动脉高压小鼠体内脂质代谢的变化,探讨脂质代谢异常在低氧性肺动脉高压发生发展中的意义。方法：SPF级雄性C57BL/6小鼠20只,随机分为2组（n=10）：常氧组和低氧组。常压连续低氧3周（9%~11% O₂,23 h/d）复制慢性低氧性肺动脉高压模型,测定小鼠右心室压（RVSP）和右心室与左心室加室间隔重量比,Elisa法检测血浆中总胆固醇（TC）、低密度脂蛋白（LDL）、高密度脂蛋白（HDL）的含量;real-time PCR法检测肝组织中3-羟基-3-甲基戊二酸单酰辅酶A还原酶（HMGCR）、低密度脂蛋白受体（LDLR）、清道夫受体B1（SR-B1）、固醇调节元件结合因子2（SREBF2）等基因的表达。结果：低氧组小鼠RVSP、RV/（LV+S）显著高于常氧组（P<0.05）,血浆中HDL含量及HDL/LDL比值较常氧组显著降低（P<0.05）,肝组织中LDLR、SR-B1基因表达较常氧组显著下调（P<0.05）;RVSP与HDL/LDL比值及LDLR、SR-B1基因表达呈显著负相关（P<0.05）。结论：脂质代谢异常参与小鼠低氧性肺动脉高压的形成。     

Enzymic composition and growth rate of human pleural mesothelioma transplants in nude mice.

The enzymic composition of 7 human mesothelioma lines propagated in nude mice was compared with 4 of the original and 15 additional mesotheliomas sampled during the patients' surgery. The xenografts exhibited several-fold higher thymidine kinase (TK), uridine kinase (UK), phosphoserine phosphatase (PSP) and peptidyl proline hydroxylase (PPH) concentrations than the fresh human samples, while their DNA, gamma-glutamyl transpeptidase (GGT) and beta-galactosidase (Bgal) contents remained similar. The volume growth rate of the xenografts (doubling time, DT = 9.23 +/- 1.25 days) was much faster than that of tumors in the human host, and the decline of this rate with increasing nodule size was accompanied by decreases in TK and PSP concentrations. This first quantitative biochemical study of xenografted human neoplasms indicates that 1) pleural mesotheliomas, though preserving their histological characteristics after heterotransplantation, show considerable increases of enzymes in nucleic acid, collagen, and nonessential amino acid synthesis, and that 2) the concentration of TK is a good indicator of the different growth properties of tumors in a mouse rather than in the human host.     

Hysteresis of plant cell-wall beta-glucosidase.

A transient-kinetic study of beta-glucosidase from soyabean cell walls was performed with the use of a stopped-flow apparatus. The progress curve of the reaction exhibits a 'slow' burst of about 1 s before the steady state is reached. In the time scale investigated this burst may be accounted for by only one exponential, whose time constant varies with the substrate concentration. As this concentration is increased the value of the time constant increases at first, then decreases. Premixing the enzyme with glucose, the last product of the reaction sequence, reverses the 'slow' burst into a 'slow' lag. Taken together, these results are only compatible with a model that involves the existence of a 'slow' conformational transition of the enzyme.     

Deficiency of steriod beta-glucosidase in Gaucher disease.

A deficiency in the activity of steroid: β-glucosidase has been observed in the particulate fraction of Gauchers tissues. There was no diminution of the “soluble” form of this enzyme in adult tissue samples. In contrast, there was a marked reduction in the soluble steroid β-glucoside hydrolytic activity in the brain and spleen, and not liver from the infantile form of the disease.     

Changes in hemopoietic and regulator levels in mice during fatal or nonfatal malarial infections. II. Nonerythroid populations

Levels of mature lymphocytes, granulocytes, macrophages, platelets, their progenitor cells, and cytokines were monitored in the blood, marrow, and spleen during fatal or nonfatal murine malarial infections. In all four malaria models, before anemia developed, there was a lymphopenia, a rapid lymphocyte depletion in the marrow with a compensating rise in spleen lymphocytes, thrombocytopenia with increased megakaryocytic progenitor cell numbers, and monocyte increases in the bone marrow and later the spleen. The development of anemia was associated with a monocytosis and neutropenia, an increase in granulomonocytic progenitor cells in the spleen, and a reduction of spleen lymphocytes. Spleen granulocytes, monocytes, and their progenitor cells increased two- to threefold more in nonfatal than in fatal malaria and the spleen lymphocyte pool became severely depleted in fatal malaria. The data suggest that a defective effector cell response was of importance for the fatal outcome of the disease. Other than an early rise in serum macrophage colony stimulating factor levels in fatal infections, changes in levels of the regulators of these effector cells did not correlate well with the outcome of the infection.     

Changes in hemopoietic and regulator levels in mice during fatal or nonfatal malarial infections. I. Erythropoietic populations

Erythroid precursors BFU-E and CFU-E and erythroblasts (ERB) were monitored in the marrow and spleen of mice during fatal or nonfatal malaria. Transient depletions of marrow CFU-E and ERB without modification of BFU-E or erythropoietin (Epo) levels were found as early events in fatal infections. Before anemia development, erythropoiesis was reduced in the bone marrow but increased in the spleen. During the anemic phase, for comparable levels of anemia, plasma Epo levels were elevated to a similar degree in fatal and nonfatal malaria. In the bone marrow, CFU-E increased twofold and BFU-E were usually reduced as expected in severe anemia. ERB populations increased but remained below or within normal values, suggesting an impairment of marrow erythropoiesis related to early events following infection. In contrast, in the spleen, ERB production was strongly simulated but amplification of ERB, CFU-E, and BFU-E populations was 2.5-fold lower in fatal than in nonfatal malaria. The results suggest that a defect in amplification of splenic erythropoiesis is a crucial determinant of the fatal outcome of malarial infection. This may have been mediated by a defective stem cell migration or multiplication. Some evidence obtained during recovery stages suggested that a factor(s) other than Epo may control splenic erythropoiesis during the anemia associated with malaria.     

Control of beta-glucosidase synthesis in Mucor racemosus.

The beta-glucosidase of Mucor racemosus was shown to be synthesized when the organism was grown in the presence of such diverse carbon sources as glycerol, lactate, xylose, ribose, alpha-methylglucoside, alpha-phenylglucoside, maltose, and cellobiose. Enzyme synthesis was strongly repressed in the presence of hexoses. In addition, exogenous cyclic adenosine 3',5'-monophosphate (cAMP) resulted in enzyme repression. When cAMP was added exogenously after enzyme activity had accumulated, a reversible enzyme inactivation occurred. Growth on disaccharides (maltose or cellobiose) was severely retarded in the presence of cAMP, whereas that on glucose remained unaffected. The results indicate a probable role for cAMP in control of glucosidase synthesis in Mucor.     

Changes in the ascorbic acid levels of peritoneal lymphocytes and macrophages of mice with endotoxin-induced oxidative stress.

Ascorbic acid (AA) is an important cytoplasmic antioxidant that mice synthesize in the liver, the intracellular levels of which decrease in an oxidative stress situation such as endotoxic shock. The present work deals with the changes in AA levels, that modulate the immune function, in the two main immune cells, namely macrophages and lymphocytes, from female BALB/c mice suffering endotoxic shock caused by intraperitoneal injection of Escherichia coli lipopolysaccharide (LPS) (100 mg/kg). The intake by cells of this antioxidant present in vitro at different concentrations was also studied. The animals show an oxidative stress, standardized in previous studies, that causes mortality at 30 h after LPS injection. The cells were obtained from the peritoneum at 2, 4, 12 and 24 h after LPS or PBS (control) injections and were incubated without or with AA at 0.01, 0.1 and 1 mM for 10, 30, 60, 120 or 180 min. The hepatic AA levels were also studied at 0, 2, 4, 12 and 24 h after LPS injection. The peritoneal cells obtained from animals injected with LPS showed increased AA levels in relation to the control cells at all times after LPS injection, with maximal effect at 12h. The AA levels decreased after this time, in agreement with changes in the AA hepatic levels. The increase was due to the AA of lymphocytes since macrophages showed a decrease in AA at different times after LPS injection. Both cells showed an increase in the intracellular levels of AA when this antioxidant was added in vitro. This takes place mainly at 30-60 min of incubation in cells from controls and at 10 min in cells from treated mice 12-24 h after LPS injection. The incorporation decreased at these times of endotoxic shock, a few hours before death. In all cases AA levels were higher in lymphocytes than in macrophages, and 1 mM was the most effective concentration. These results suggest that the immune cells need appropriate levels of antioxidants, such as AA, under oxidative stress conditions, and that while lymphocytes take and accumulate AA, macrophages use it.     

The effects of air ions on brain levels of serotonin in mice

Mice were maintained in a controlled pollutant-free microenvironment and were exposed for 12, 24, 48 and 72 hr to 3 different concentrations of small positive or negative air ions: 2–4 × 10³ ions/cm³, 3–4 × 10⁴ ions/cm³ or 3.5–5 × 10⁵ ions/cm³. Spectrophotofluorometric assays of brain serotonin levels of air ion-treated mice showed statistically significant differences as early as 12 hours from those of mice kept in untreated pollutant-free air. Essentially no deviation from control values were observed at 24 and 48 hours. After 72 hours of exposure sharp decreases took place in all groups with the single exception of the animals exposed to 3–4 × 10⁴ positive ions/cm³. The hypothesis that alterations in mood and affect associated with certain meteorological conditions, e.g. winds such as the foehn, sirocco, etc. might depend upon air ion-induced changes in brain levels of serotonin was examined in the light of recent advances in neurophysiology and neuropharmacology.     

Immobilization of Aspergillus beta-glucosidase on chitosan.

beta-Glucosidase of Aspergillus phoenicis QM 329 was immobilized on chitosan, using the bifunctional agent glutaraldehyde. The most active preparation based on the amount of support contained a 1:2.5 enzyme-to-chitosan ratio (wt/wt). However, the specific activity of the bound enzyme decreased from 10 to 1% with increasing enzyme-to-chitosan ratio. Compared with free beta-glucosidase, the immobilized enzyme exhibited: (i) a similar pH optimum but more activity at lower pH values; (ii) improved thermal stability; (iii) a similar response to inhibition by glucose; and (iv) mass transfer limitations as reflected by higher apparent Km and lower energy of activation.