Activation Energies of Reactions Catalyzed by the Nitrate Reductase from Chlorella vulgaris

The thermal dependence of two of the reactions catalyzed bythe nitrate reductase from Chlorella vulgaris was determined.The activation energies for NADH:nitrate oxidoreductase (EC1.6.6.1 [EC] ) and NADH:Cytochrome c oxidoreductase (EC 1.6.99.3 [EC] )are 42.1 kJ?mol^–1 and 21.5 kJ?mol^–1, respectively.Since the thermal dependency of the two enzymes is different,ratios of the activities will vary with temperature. The importanceof both rigorous thermal control during nitrate reductase assaysas well as the need to specify the temperature at which theratio of activities for the enzyme are clearly established. ¹Present Address: Cropping Systems Research Laboratory, USDA-ARS,Route 3, Box 215, Lubbock, TX 79401, U.S.A. (Received November 25, 1987; Accepted March 2, 1988)     

Transferrin endocytosis and iron uptake in developing myogenic cells in culture: effects of microtubular and metabolic inhibitors, sulphydryl reagents and lysosomotrophic agents

The experiments described in this study were designed to investigate receptor-mediated endocytosis of transferrin and its role in iron uptake by cultured chick presumptive myoblasts (dividing and non-dividing) and myotubes. The effects of a variety of inhibitors on the internalization of transferrin and iron were investigated and three main effects were found: (i) sulphydryl reagents and microtubular inhibitors reduced the rate of transferrin and iron internalization to similar degrees, (ii) metabolic inhibitors reduced the rate of iron uptake more than that of transferrin endocytosis, and (iii) lysosomotrophic agents almost completely abolished iron accumulation by the cells without any effect on the rate of transferrin internalization. The results suggest that metabolic energy is required not only for the endocytosis of transferrin but also for subsequent steps in the iron uptake process, and that iron release from transferrin occurs in acidified endosomes. Overall, these experiments show that all or virtually all of the iron taken up by developing muscle cells from transferrin occurs as a consequence of receptor-mediated endocytosis of the protein.     

Induction of IL-1 secretion from human monocytes by Fc region subfragments of human IgG1

Peripheral blood-derived human monocytes and the murine P388D1-monocytes-like cell line are induced to secrete IL-1 when stimulated with Fc region but not F(ab) region subfragments obtained from the cleavage of human IgG1 with papain or pepsin. The portion of the Fc region of IgG1 responsible for stimulation of IL-1 secretion appears to be located within the C gamma 3 domain of the molecule. This hypothesis is supported by the observation that the biologically active pepsin-derived pFc' subfragment is located within the C gamma 3 domain and the long-term papain digests containing predominately Fc' are also active. In contrast, short term papain digests containing mostly intact Fc fragments were found to be unable to induce IL-1 secretion.     

Mechanism of human monocyte activation via the 40-kDa Fc receptor for IgG

It is shown that a mAb specific for the human 40-kDa FcR (FcRII) leads to activation of human monocytic cells but that extensive cross-linking of the receptor is required. Calcium mobilization can be induced in immature monocytic cells (undifferentiated U937 cells) and peripheral blood monocytes with an intact IgG1 anti-FcRII antibody (CIKM5) but not by F(ab')2 fragments of this antibody. The intact antibody can bind in a tripartite manner by its two F(ab') sites and its Fc-binding site whereas the F(ab')2 fragments of this antibody can only bind in a divalent fashion. A rise in intracellular free calcium ion concentration occurs when F(ab')2 fragments are cross-linked with F(ab')2 anti-mouse Ig indicating that more extensive cross-linking of FcRII is required rather than an obligatory requirement for an Fc-FcRII interaction. Calcium mobilization in response to intact or cross-linked F(ab')2 fragments of CIKM5 is associated with superoxide production only in IFN-gamma-primed peripheral blood monocytes and IFN-gamma differentiated U937 cells indicating that the activation signal produced via FcRII is inadequate to fully stimulate non-"primed" cells. A second mAb reactive with FcRII (2E1) does not cause calcium mobilization in monocytes or U937 cells, and partially blocks the effects of CIKM5. 2E1 also blocks CIKM5 superoxide production in IFN-gamma-primed monocytes and differentiated U937 cells. This may be explained in part by the fact that 2E1 is an IgG2a antibody and can only participate in bipartite binding with FcRII. When 2E1 is cross-linked with F(ab')2 anti-mouse Ig there is a small calcium response. This does not cause superoxide generation in IFN-primed monocytes but does do so in IFN-gamma differentiated U937 cells. FcRII is also expressed on granulocytes and some B cells but the effects of cross-linking the receptor on these cells differ from those seen in monocytes.     

Histochemical localization of IGF-I and IGF-II mRNA in the rat between birth and adulthood

We describe the postnatal ontogeny and localization of insulin-like growth factors I and II (IGF-I and -II) in the rat. We have used oligodeoxyribonucleotide probes for in situ hybridization (hybridization histochemistry) and for Northern blotting. IGF-II mRNA is strongly expressed in liver, skeletal muscle, perichondrium, leptomeninges and choroid plexus of the newborn. Demonstrable levels fall dramatically in the liver at 18-20 days postnatally but persist for longer periods in muscle and remain undiminished throughout life in the pia/choroid plexus, indicating that different control mechanisms operate in these tissues. IGF-I mRNA is predominantly found in the liver. Its level in this organ rises well before levels of IGF-II fall. This suggests that distinct factors govern the expression of IGF-I and -II genes.     

Blood lipid concentrations and other cardiovascular risk factors: distribution,prevalence, and detection in Britain

To establish the distribution of blood lipid concentrations and the prevalences of other risk factors for cardiovascular disease in Britain 12 092 men and women aged 25-59 in Glasgow, Leicester, London, and Oxford were studied. Subjects were selected by opportunistic case finding, in which patients consulting their general practitioner for any reason were offered a health check by appointment, or random selection from age-sex registers, in which an invitation for a health check was posted. The overall rate of response was 73%, being 91-94% by opportunistic case finding and 36-63% by random selection. At the health check subjects answered a brief questionnaire about risk factors for cardiovascular disease, and their height, weight, and blood pressure were recorded; a blood sample was taken for measuring plasma concentrations of cholesterol, triglyceride, high density lipoprotein cholesterol, and glucose.The mean cholesterol concentrations were 5·9 (SD 1·2) and 5·8 (1·2) mmol/l in men and women, respectively. In London the mean value was 5·5 (1·2) mmol/l for both men and women and was significantly lower than mean values in the three other centres, among which there were no significant differences. In men and women aged 25-29 concentrations were similar but they increased in men until the age of 45-49, after which they showed no further increase; in women concentrations did not increase until the age of 40-44 and by the age of 50-59 values were higher than in men. Mean triglyceride concentrations were significantly higher in men than in women (1·8 (1·4) v 1·3 (0·9) mmol/l, respectively), and trends with age were similar to those for cholesterol concentrations, except that at no age were values higher in women than in men. Mean triglyceride values overall were higher in Glasgow and London than in Oxford and Leicester. Body mass index was higher in Glasgow and London than in the other two centres and correlated with systolic and diastolic blood pressures and triglyceride concentration. In addition, subjects in Glasgow smoked significantly more than those in the other centres. These observations could contribute to the higher rate of coronary heart disease in Glasgow. Plasma lipid concentrations and the prevalences of other risk factors for cardiovascular disease were similar in subjects selected by opportunistic case finding and by random selection.In Britain cholesterol values have changed little during the past 12 years despite dietary recommendations and health education. Identifying subjects at particularly high risk of coronary heart disease is required to supplement advice to the general population to reduce the prevalence of this disease. Opportunistic case finding would be an appropriate method of identifying such subjects in general practice, although none of the potential markers for hyperlipidaemia was particularly useful in identifying all subjects at high risk.     

Bowel sound biofeedback as a treatment for irritable bowel syndrome

Using an electronic stethoscope placed on subjects' abdomens, bowel sound biofeedback was administered to five subjects suffering from irritable bowel syndrome (functional diarrhea). They were instructed to alternately increase and decrease colonic sounds in an attempt to gain control over bowel activity. Using daily ratings of diarrhea as the primary dependent measure, three of five subjects reduced mean ratings enough at posttreatment to meet our 50% criterion for success (100%, 94%, and 54%). At 1-year follow-up, two of the three short-term successes had maintained their level of improvement — each had ratings 75% below those of pretreatment.