Studies on the mode of action of calciferol. X. 24-Nor-25-hydroxyvitamin D3, an analog of 25-hydroxyvitamin D3 having "anti-vitamin" activity.

24-Nor-25-hydroxyvitamin D₃, an analog of 25-hydroxyvitamin D₃, has been chemically synthesized in six steps. This steroid was tested in chicks, $\text{in vivo}$, for its ability to generate the classic vitamin D mediated responses of stimulation of intestinal calcium transport and bone calcium mobilization. Although the 24-nor-25-OH-vitamin D₃ itself exhibited no biological activity in these assays, the analog was found to inhibit the normal responses produced by a physiological dose of vitamin D₃. These results suggest that 24-nor-25-OH-vitamin D₃ may satisfy certain requirements expected of a calciferol “anti-vitamin.”     

Structural requirements for the interaction of 1 alpha, 25-(OH) 2- vitiamin D3 with its chick interestinal receptor system.

The mechanism by which 1 alpha,25-dihydroxycholecalciferol (1alpha,25-(OH)2D3), the biologically active metabolite of cholecalciferol (vitamin D3), stimulates intestinal calcium absorption has been shown to involve an interaction of the steroid with a specific cytosol-chromatin receptor system in this target organ. Thus, 1alpha,12(OH)2D3 binds to a specific cytoplasmic receptor protein and then, following a temperature-dependent step, becomes associated with a finite number of chromatin acceptor sites prior to the initiation of the physiological response. In this respect, 1alpha,25(OH) 2D3 is similar to a number of other steroid hormones. In this investigation, studies were performed to help define the essential structural features required for interaction of 1alpha,25-(OH)2D3 with its intestinal receptor system, and presumably, for biological activity. To this end, competition studies utilizing a series of closely related structural analogs of cholecalciferol were carried out by means of a competitive binding assay highly specific for 1alpha,25(OH)2D3. The competitive binding assay employed in this study is dependent upon the ability to duplicate, in vitro, the conditions which permit the saturable binding of 1alpha,25-(OH)2[3H]D3 to chick intestinal chromatin, in vivo. Optimal conditions for this assay were achieved by the incubation of a reconstituted intestinal receptor system consisting of separately isolated cytosol and Triton X-100 chromatin fractions at 25 degrees for 45 min with 2.0 X 10-8 M 1alpha,25-(OH)2[3H]D3. Maximal binding of about 21 to 24 pmol of radioactive steroid bound per chick intestinal chromatin occurred under these conditions. The ability of the various analogs to compete with the radioactive hormone was assessed by virtue of a decrease in the amount of radioactive steroid bound to the chromatin in the presence of increasing concentrations of nonradioactive analog.     

Nineteen Paracentric Chromosomal Inversions in Mice

Using mutagens on sperm and spermatids we have produced nineteen chromosomal inversions in mice. The levels of radiation and chemical mutagen we used induced inversions in about one per cent of the animals screened. Nine inversions have been transmitted through successive generations, and the particular frequency of first meiotic anaphase bridges manifested by each inversion remained constant. The cytological properties of first meiotic anaphases varied considerably among the inversions. Chromosomal locations of five inversions are known. Four of the five are fully viable in homozygous condition.     

Ontogenetic changes in haemoglobins in roach, Rutilus rutilus (L.) and rudd, Scardinius erythrophthalmus (L.)

The haemoglobins and globins of the roach, Rutilus rutilus (L.) and the rudd, Scardinius erythrophthalmus (L.) have been studied by starch gel electrophoresis. At all stages of development studied, the roach and rudd haemoglobins give a similar electrophoretic pattern, the young fish of about 2 cm standard length possessing three separate haemoglobins, of which two bands only persist in the adult. The evolutionary significance of multiple haemoglobins in fishes is briefly discussed.     

Reduction of ferricytochrome c by hydrogen atoms: Evidence for intramolecular transfer of reducing equivalent

Direct evidence obtained by means of the technique of pulse radiolysis-kinetic spectrometry, with measurements in the time range 10⁻⁶ to 1 s, is presented that, consequent upon reaction of a single H-atom with a single molecule of ferricytochrome c, a reducing equivalent is transmitted via the protein structure to the ferriheme moiety. Such transmission accounts for at least 70% of the total reduction of the ferri to the ferro state of cytochrome c. The remainder of the total reduction takes place without stages resolvable on the time scale of these experiments. Reduction brought about by H atoms appears to follow a different course than reduction by hydrated electrons. In the latter case, intramolecular transmission of reducing equivalents could not be demonstrated (Lichtin, N. N., Shafferman, A. and Stein, G. (1973) Biochim. Biophys. Acta 314, 117–135).

Not every H-atom reacts with ferricytochrome c at a site which results in conversion of the Fe(III) state to the Fe(II) state. Approximately half of reacting H-atoms do not produce reduction.

The following second order rate constants have been determined in solutions of low ionic strength at 20±2 °C: k[H+ferricytochrome c] = (1.0±0.2) · 10¹⁰ M⁻¹ · s⁻¹ at pH 3.0 and 6.7; k[H+ferrocytochrome c] = (1.3±0.2) · 10¹⁰ M⁻¹ · s⁻¹ at pH 3.0; k[e⁻_aq + ferrocytochrome c] = (1.9±0.4) · 10¹⁰ M⁻¹ · s⁻¹ at pH 6.7.     

Heparin fractionation by electrofocusing: Presence of 21 components of different molecular weights

Electrofocalization of heparin with ampholyte mixtures pH 3.0 to 5.0 has shown the presence of at least 21 components in several commercial heparin preparations. The difference between these 21 components resides exclusively in their molecular weights which range from 3 000 to 37 500.     

Parathyroid carcinoma: location of pelvic metastases by parathyroid hormone assay

A metastasis from a functioning parathyroid carcinoma was located by PTH radioimmunoassay and selective venous catheterization. The site of the metastasis, verified at autopsy, was in the right side of the pelvis. This is the most distant reported location for metastatic parathyroid carcinoma. The patient''s plasma immunoreactive PTH rose more than twofold in response to induced hypocalcemia. This suggests that relative hypocalcemia, induced therapeutically in such patients, may result in a higher chronic level of PTH secretion.