Circannual versus seasonal variations of longitudinally sampled 25-hydroxycholecalciferol serum levels

Seasonal variations in human serum levels of 25-hydroxycholecalciferol (25OHD) have been largely documented in transverse studies of population. But seasonality is not per se a demonstration that 25OHD serum levels fluctuate along the course of year according to a waveform profile with a periodic rhythm. Because of this, we attempted to investigate the possible occurrence of a circannual rhythm for 25OHD serum levels in a longitudinal design, by fitting a 365.25-day cosine curve to temporal biodata recorded in 10 clinically healthy subjects, monthly sampled for RIA determinations of 25OHD. Cosinor procedure statistically validated the occurrence of a circannual rhythm for 25OHD serum concentrations at a highly significant level of probability (P = 0.0015) for null hypothesis amplitude = 0. With 95% of probability, amplitude ranges from 5.0 to 16.5 ng/ml (mean value of oscillation = 10 ng/ml), while acrophase is temporally located from September 14 to December 3 (mean timing = October 21). Yearly, mean values for 25OHD serum concentrations is of 40.3 +/- 5.4 ng/ml as quantified by the line which transversely divides the cosine curve interpolating original biodata. By calculating the band of a complete 12 months variability which includes 90% of the distribution with 90% confidence limits, the circannual chronodesm of 25OHD serum levels has been obtained. Such a chronodesmic sinusoid has been compared to the circannual chronogram. By this comparison, a dissociation between the crest (October) and the peak (August) has been detected. The finding suggests that seasonal variations are superimposed to the circannual rhythm. Seasonal but also circannual changes, thus, characterize the yearly variability of 25OHD serum levels in man.     

Behavior of serum 25-hydroxycholecalciferol in humans after administration of vitamin D and 25-OHD3]

Serum levels of 25 hydroxycholecalciferol were evaluated following i.m. and p.o. vit. D2 and D3 and p.o. 25OHD3 administration. While no increment in 25OHD3 serum levels were observed after i. m. administration of non-hydroxylated calciferols, a marked increment of the metabolite was found following the oral administration. However the peak values were largely impredictable. Acute and chronic p.o. administration of 25OHD3 determines a rapid and dose-dependent increase of the serum levels of the metabolite. In addition considering that a lower dosage is required of 25OHD3 compared to vit. D, this metabolite is preferable in the therapeutic use.     

Kinetics and regulation of 25-hydroxycholecalciferol 1 alpha-hydroxylase from cells isolated from human term decidua

Kinetics and regulation of 25-hydroxycholecalciferol 1 alpha-hydroxylase from cells isolated from term human decidua were studied. The production of 1 alpha,25-dihydroxycholecalciferol (calcitriol) was linear with time for up to 6 h and was directly proportional to the number of cells up to 20 X 10(6)/dish at a substrate concentration of 100 nM. Under these conditions the apparent Km was 88 nM and the Vmax 3.0 pmol/10(6) cells. The production of [3H]calcitriol was inhibited by 0.1 nM (P less than 0.01) and 1 nM (P less than 0.005) unlabeled calcitriol. Unlike the kidney enzyme and for reasons that remain unclear, neither inorganic phosphate salts nor parathyroid hormone had any acute effect on the calcitriol production. Further studies are required to delineate the regulatory mechanism of this enzyme.     

Widespread, specific binding of 25-hydroxycholecalciferol in rat tissues.

In the vitamin D-depleted rat, all nucleated tissues examined (brain, lung, heart, pancreas, liver, cartilage, muscle, bone, kidney, and intestine) contained a soluble substance which bound 25-hydroxy[3H]cholecalciferol in vitro specifically and sedimented at 6.3 S in linear sucrose gradients. The serum-steroid complex sedimented a 4.1 S, and erythrocyte lysates were apparently devoid of specific binding activity. The ability of these cytosols to specifically bind the steroid was destroyed by treatment with trypsin, but not by RNase, DNase, or 1 mM p-hydroxymercuribenzoate. The sedimentation pattern was not altered in sucrose gradients containing 0.5 M KCl or following cytosol preparation and ultracentrifugation in gradients containing 0.012 M dithiothreitol. The apparent avidity for 25-hydroxycholecalciferol (KA similar to 2 times 10- M) was slightly higher in muscle and kidney cytosols than in serum, but serum contained a large number of specific binding sites. The presence of widespread, high affinity binding proteins for 25-hydroxycholecalciferol raises the possibility that tissues other than the intestine, bone, and kidney may respond directly to vitamin D metabolites.     

Metabolism of 25-hydroxycholecalciferol in human promyelocytic leukemia cells (HL-60). Isolation and identification of (5Z)- and (5E)-19-nor-10-oxo-25-hydroxycholecalciferol

Human promyelocytic leukemia cells incubated with 25-hydroxy[26,27-methyl-3H] cholecalciferol (1 microCi) or non-radioactive 25-hydroxycholecalciferol (550 micrograms) produced significant quantities of two vitamin D3 metabolites. The two metabolites were isolated and purified by methanol chloroform extraction and a series of chromatographic procedures. The metabolite purification and elution positions on these columns were followed by radioactivity and their ultraviolet absorption at 310 nm. The two metabolites have been unequivocally identified as (5Z)- and (5E)-19-nor-10-oxo-25-hydroxycholecalciferol by ultraviolet absorption spectrophotometry, mass spectrometry, Fourier-transform infrared spectrophotometry and co-chromatography with synthetic compounds on a high-performance liquid chromatograph. (5E)- but not (5Z)-19-nor-10-oxo-25-hydroxycholecalciferol was able to induce HL-60 cell phenotypic and functional differentiation. However, these two metabolites of 25-hydroxycholecalciferol did not bind specifically to the chick intestinal 3.7 S. receptor protein for 1 alpha,25-dihydroxycholecalciferol. The precise biological role of these metabolites is as yet unclear.     

High-pressure liquid chromatographic method for the determination of 25-hydroxycholecalciferol in cow plasma

A high-pressure liquid chromatographic (hplc) procedure was developed for the determination of 25-hydroxycholecalciferol (25-OH-D₃) in cow plasma or serum. The procedure involved extraction with an ethanol-ethyl ether mixture, separation of the aqueous phase, solvent partitions, column chromatography on silica gel, and, finally, determination by reversed phase hplc on a C₁₈-bonded microparticulate silica column. The identity of the drug in the extract was confirmed by comparison with a standard by liquid, thin-layer, and gas-liquid chromatography as the free steroid and the heptafluorobutyrate and by the uv spectra and also from the mass spectrum of the heptafluorobutyrate. Twenty-four samples from cows on normal diet (dry, lactating, and pregnant) were analyzed. The normal circulating levels of 25-OH-D₃ ranged from 40 to 58 ng/g; mean 48 ± 5.0 ng/g. The procedure was used to analyze a limited number of human and hog samples. Human serum contained 10–20 ng/g which was in agreement with literature values. Hog serum contained 18 ng/g.     

Regional and seasonal differences in the fatty acid composition of human subcutaneous fat

The fatty acid composition of human subcutaneous fat obtained at a hospital in the north of Japan, was determined by means of gas chromatography. In the fat of chest and abdominal walls percentages of saturated fatty acids were higher and those of mono-unsaturated ones were less in comparison with respective values in the fat of forearm and leg. The unsaturation of the fat of extremities was more pronounced in the winter than in the summer, while no seasonal variations were observed in the fatty acid composition of the fat of trunk.

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Zusammenfassung Die Verteilung der Fettsäuren im subcutanen Fett des Menschen wurde gaschromatographisch bestimmt. Die Fettproben wurden in einer Klinik im Norden Japans bei Operationen entnommen. Der prozentuale Anteil an gesättigten Fettsäuren war höher im Fett der Brust und Bauchwand und der an einfach ungesättigten Fettsäuren im Fett des Vorderarms und Beines. Die Anreicherung ungesättigter Fettsäuren in den Extremitäten war im Winter ausgeprägter als im Sommer. In der Zusammensetzung der Fett des Rumpfes fehlte eine jahreszeitenabhängige Variation.

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Resume Par analyse chromatographique des gaz, on a déterminé la répartition des acides gras dans les tissus adipeux sous-cutanés de l'homme. Les échantillons de graisse ont été prélevés lors d'opérations dans une clinique du nord du Japon. La proportion d'acides gras saturés était plus élevée dans la graisse de la poitrine et du ventre, alors que les acides gras mono-nonsatures l'emportaient dans l'avant-bras et dans les jambes. L'accumulation d'acides gras non saturés dans les extrémités était plus accentuée en hiver qu'en été. On n'a par contre pas pu établir de relation saisonnière dans la composition de la graisse du tronc.

     

Effect of experimental diabetes on levels of 25-hydroxycholecalciferol in pregnant rats and fetuses

Induction of diabetes in female rats by streptozotocin administration 7 days before mating led to an increase in maternal and fetal calcemia at day 21 of gestation. The plasma levels of 25-hydroxycholecalciferol (25 OH D3) were increased in the diabetic mother whereas the 25 OHD3 contents in entire fetuses were greatly decreased in comparison with control values obtained in both normal pregnant rat and normal fetuses. Our results obtained in pregnant rat were different from those found in the literature concerning non pregnant animals in which only 1,25-dihydroxycholecalciferol was affected by diabetes.