Age alters expression and inducibility of heme oxygenase isozymes in mice.

Heme oxygenase (HO) performs the rate limiting step in heme degradation and is induced by cell injury or stress. We wished to determine if dietary fatty acid composition, increased age and/or an induced oxidative stress would alter the expression of HO-1 (constitutive and inducible isozyme) or of HO-2 (constitutive isozyme), in mouse liver, spleen and brain. Six-and 24-month-old male B6C3F1 mice were fed AIN-76A diets containing either 5% corn oil (CO, moderately unsaturated, n=5 per age group) or 19% menhaden fish oil plus 1% corn oil (FO, highly polyunsaturated, n=20 per age group). After 2 weeks, 5 CO and 5 FO fed mice in each age group were sacrificed. The remaining FO diet mice (n=15 per age group) were then challenged with a systemic oxidative stress by intraperitoneal injection of 125 mg iron/kg body weight as iron dextran. Five stressed mice from each age group were sacrificed 1, 5, and 24 hours post injection; liver, spleen and brain were removed. Part of each tissue was fixed in formalin, and microsomal protein isolated from the remaining tissue. HO-1 and HO-2 were detected by immunoblot of microsomal protein and by immunohistochemical staining of fixed tissue in the liver and spleen, but only HO-2 was detected in the brain. There was no significant difference in HO-1 or HO-2 expression due to diet. The liver of old unstressed mice had significantly more HO-1 than young mice. However, HO-1 was significantly induced in the livers of young mice, but not of old mice, following oxidative stress. Spleen HO-1 expression was not significantly altered by age or oxidative stress. HO-2 expression was not significantly altered by age or induced oxidative stress in any tissue examined. Age-related alterations in liver HO-1 isozyme expression and inducibility may contribute to increased susceptibility to exogenous stress and disease.

Cytoplasmic domain of rhodopsin is essential for post-Golgi vesicle formation in a retinal cell-free system.

In retinal photoreceptors, highly polarized organization of the light-sensitive organelle, the rod outer segment, is maintained by the sorting of rhodopsin and its associated proteins into distinct post-Golgi vesicles that bud from the trans-Golgi network (TGN) and by their vectorial transport toward the rod outer segment. We have developed an assay that reconstitutes the formation of these vesicles in a retinal cell-free system. Vesicle formation in this cell-free assay is ATP-, GTP-, and cytosol-dependent. In frog retinas vesicle budding also proceeds at 0 degrees C, both in vivo and in vitro. Vesicles formed in vitro are indistinguishable from the vesicles formed in vivo by their buoyant density, protein composition, topology, and morphology. In addition to the previously identified G-proteins, these vesicles also contain rab11. Concurrently with vesicle budding, resident proteins are retained in the TGN. Collectively these data suggest that rhodopsin and its associated proteins are sorted upon exit from the TGN in this cell-free system. Removal of membrane-bound GTP-binding proteins of the rab family by rab GDP dissociation inhibitor completely abolishes formation of these vesicles and results in the retention of rhodopsin in the Golgi. A monoclonal antibody to the cytoplasmic (carboxy-terminal) domain of rhodopsin and its Fab fragments strongly inhibit vesicle formation and arrest newly synthesized rhodopsin in the TGN rather than the Golgi. Therefore rhodopsin sorting at the exit from the TGN is mediated by the interaction of its cytoplasmic domain with the intracellular sorting machinery.

Effect of uncomplicated pregnancy on body water content and distribution in baboons.

Body water content and distribution were estimated before, during, or after 32 pregnancies in baboons. Water content of the various compartments (in liters) correlated with body weight in both nonpregnant and pregnant baboons, and with length of gestation in pregnant baboons. In proportion to body weight (in ml/kg), most water compartments did not change significantly with length of gestation. Mean plasma volume and blood volume were higher during pregnancy than before or after.

Tolerance of fat emulsions in very-low-birth-weight neonates.

Fluid and energy intakes and plasma concentrations of various lipid fractions were studied in 45 newborn infants, weighing between 820 and 1550 g, randomly assigned to one of three regimens for infusion of a fat emulsion: group 1, stepwise increase in daily dosage (1 to 4 g/kg) at a steady rate over 24 hours; group 2, stepwise increase in daily dosage (1 to 4 g/kg) at a steady rate over 16 hours, followed by a rest period of eight hours; and group 3, full dosage (4 g/kg) at a steady rate over 24 hours. At a given daily rate of infusion, mean plasma concentrations of the various lipid fractions tended to be higher in group 2 than in either group 1 or group 3. Plasma lipid concentrations correlated best with the hourly rate of lipid infusion, regardless of whether the infusion was continuous over 24 hours or intermittent.

Tolerance of fat emulsions in very-low-birthweight neonates. Monitoring of plasma lipid concentrations.

Tolerance to parenterally administered fat emulsions was studied in 45 normally grown neonates ranging from 820 to 1550 gm in birthweight, from 27 to 34 weeks in gestational age, and from 2 to 10 days postnatal age. Concentrations of total lipids, free glycerol, true triglycerides, free fatty acids, and cholesterol in plasma were studied over an 8-day period. The aim was to determine whether the concentrations of any one lipid fraction could be used to predict those of other fractions. With 650 pairs of data being correlated, the best correlation coefficients were obtained between total lipids and triglycerides (r = 0.67), total lipids and cholesterol (r = 0.63), free glycerol and free fatty acids (r = 0.55), total lipids and free fatty acids (r = 0.54), and triglycerides and free fatty acids (r = 0.50). Although all correlation coefficients were highly significant statistically (P less than 0.001), the very large standard errors precluded using any of these relationships to make clinical predictions. Problems and limitations of currently available techniques are discussed in order to provide a lead for further research.

Identification of a 32-kilodalton protein of Mycoplasma pneumoniae associated with hemadsorption.

Monoclonal antibodies directed against a 32-kilodalton (kDa) protein of Mycoplasma pneumoniae have been used to characterize a hemadsorption-negative (HA-) mutant class whose protein profile was previously indistinguishable from the wild-type, hemadsorbing (HA+) strain. Electron microscopy and colloidal gold labeling techniques were applied for ultrastructural analysis of the 32-kDa protein. Results indicate that this protein clusters in the tip structure of M. pneumoniae (HA+) wild-type organisms. Additionally, the protein is precipitated by infected hamster sera.