Bone markers during a 6-month space flight: effects of vitamin K supplementation

Rapid bone loss is a serious health problem for astronauts during long lasting missions in space. We have recorded the changes of biochemical markers for bone metabolism in one of the astronauts during the 6-month space flight of the EUROMIR-95 mission. Immediately after launch both bone resorption markers and urinary calcium excretion increased about two fold, whereas bone formation markers remained unchanged. After 12 1/2 weeks the astronaut received vitamin K1 (10 mg/day for 6 weeks). Vitamin K is known to be involved in the formation of gamma-carboxyglutamate (Gla) in proteins, such as the calcium-binding bone Gla-proteins osteocalcin and matrix Gla-protein. Concomitant with the start of vitamin K treatment, the calcium-binding capacity of osteocalcin increased, and so did the urinary excretion of free Gla. This is suggestive for a subclinical vitamin K-deficiency in the astronaut before vitamin K-supplementation. During periods of high vitamin K status markers for bone formation (osteocalcin and bone alkaline phosphatase) had increased as compared to the first part of the flight. The mean increases were 14 and 23%, respectively. Our data suggest that increased intake of vitamin K may contribute to counteracting microgravity-induced loss of bone mass during long lasting space missions, but need confirmation in more astronauts.     

The rhizosphere in Zea: new insight into its structure and development

Some of the nodal roots of field-grown Zea mays L. bear a persistent soil sheath along their entire length underground except for a glistening white soil-free zone which extends approximately 25 mm behind the root cap. These roots are generally unbranched. The histology of the surface and the rhizosphere of the sheathed roots has been examined by correlated light and electron microscopy. All mature peripheral tissues including root hairs, are largely intact and apparently alive where enclosed by the soil sheath. The sheath is permeated by extracellular mucilage which is histochemically distinct from the mucilage at the epidermal surface, but similar to that produced by the root cap. Isolated cells resembling those sloughed from the sides of the root cap persist in the soil sheath along the length of these roots. Fresh whole mounts of the sheath show that these detached cells may be alive and streaming vigorously even at some distance from the root cap. Rhizosphere mucilage is associated with the isolated cells.To whom correspondence should be addressed     

The formation of thromboxane B2, leukotriene B4 and 12-hydroxyeicosatetraenoic acid by alveolar macrophages after activation during tumor growth in the rat

Pieces of tumor tissue were implanted subcutaneously in the right flank of BN female rats. After 3, 7, 10, 12, 14 and 17 days the lungs were lavaged and the alveolar macrophages collected. The cells were activated with the calcium ionophore A23187 and the formation of thromboxane B2 (TxB2), leukotriene B4 (LTB4) and 12-hydroxyeicosatetraenoic acid (12-HETE) determined. The formation of TxB2 decreased considerably until day 7. Thereafter, no changes occurred. The formation of LTB4 increased after the tumor implantation until day 10 and remained stable for the rest of the period, 12-HETE formation was approximately similar, with a decrease at day 12 but continued to increase after day 14. These results suggest that during tumor growth an inhibition of the cyclo-oxygenase or thromboxane synthase occurs and an activation of the C5- and C12-lipoxygenases of the alveolar macrophages.     

Vitamin K-dependent carboxylase: effect of ammonium sulfate on substrate carboxylation and on inhibition by stereospecific substrate analogs

(1) High concentrations of ammonium sulfate may stimulate the carboxylase activity of bovine liver microsomes about 10-fold. This effect results from an increase of the Vmax, whereas neither the apparent Km for a number of substrates nor the Ki for substrate analogs is affected. (2) The effect of ammonium sulfate was only found in substrates lacking the pro-sequence. No effect was measurable on the carboxylation of pro-PT28 and endogenous precursor proteins. (3) If the pro-fragment was added as a peptide not covalently bound to a carboxylatable substrate, the carboxylation thereof was only slightly affected and ammonium sulfate remained active as a stimulator of carboxylase activity. (4) S-MeTPT is a much stronger inhibitor of carboxylase activity than is R-MeTPT. (5) The inhibition of carboxylase by the methylated tripeptides is competitive and independent of the type of substrate. Also pro-PT28, which contains the full pro-sequence, could be inhibited completely. (6) On the other hand the carboxylation of endogenous protein precursors could only be partly inhibited by the substrate analogs: even at high concentrations of S-MeTPT a residual endogenous substrate carboxylation of about 30% was left.     

Comparison of the vitamins K1, K2 and K3 as cofactors for the hepatic vitamin K-dependent carboxylase

The vitamins phylloquinone (K1), menadione (K3) and various menaquinones (K2) were compared for their ability to serve as a cofactor for the hepatic vitamin K-dependent carboxylase. It was found that the cofactor activity of the menaquinones varied with the length of the aliphatic side-chain and showed an optimum at MK-3. Menadione was not active at all. The concentration required for half-maximal reaction velocity (K 1/2) was determined for the various menaquinones and decreased at increasing chain length. The K 1/2 value for MK-4 was 3-times lower than that for vitamin K1. Under our in vitro conditions both vitamin K1 and the K2 vitamins were rapidly metabolized into a mixture of the quinone, the hydroquinone and the epoxide form. The fact that at equilibrium the level of these three metabolites was independent of the starting material shows that the vitamin K cycle is operational for vitamin K1 as well as for K2.     

Purification and properties of two lipases from pig adipose tissue.

Two lipases were purified from pig adipose tissue after delipidation by a mild and effective procedure using mixtures of chloroform and butanol. This was followed by hydrophobic adsorption chromatography on aminohexyl-Sepharose 4B coupled with octanoic acid, gel filtration on Sephadex G-100, and isoelectric focusing. Two electrophoretically and chromatographically pure enzymes were obtained, which had the same molecular weight (60 000 +/- 3000) and specific activity, and almost identical amino acid compositions; the isoelectric points, i.e. 5.2 and 5.5, differed.     

Visualization of binding and receptor-mediated uptake of low density lipoproteins by human endothelial cells

Low density lipoproteins (LDL) were conjugated to colloidal gold for investigation of the ultrastructural aspects of binding and receptor-mediated internalization of LDL by cultured endothelial cells from the human umbilical artery and vein. The number of LDL receptors was increased by preincubation in lipoprotein-depleted serum. When the cells were incubated with LDL-gold particles for 2 h at 4 degrees C, the complexes were found in coated pits as well as in clusters attached to the plasma membrane. Small vesicles containing a few LDL-gold complexes appeared in the cytoplasm close to the plasma membrane when the cells were incubated with the conjugate for 5 min at 37 degrees C. After 15 min at 37 degrees C, larger vesicles with a pale matrix and membrane-orientated LDL-gold complexes were seen. After incubation for 30 min at 37 degrees C, colloidal gold particles were present in dense bodies. Quantification of the binding of LDL-gold complexes to the plasma membrane at 4 degrees C showed no differences between arterial and venous endothelial cells.     

Isoenzymes of vitamin-K-dependent carboxylase

Vitamin-K-dependent carboxylase was prepared from bovine liver, kidney, lung and testis and it was checked that these systems obeyed the laws of normal enzyme kinetics. Four carboxylatable substrates were obtained from different sources and the apparent Michaelis constants of the various carboxylases for these four substrates were measured. From the results thus obtained we concluded that carboxylase is a group name for a number of isoenzymes which are present in hepatic as well as in various non-hepatic tissues.