A radioiodinated, intracellularly trapped ligand for determining the sites of plasma protein degradation in vivo.

We recently developed a general method for determining tissue sites of degradation of plasma proteins in vivo that made use of covalently attached radioactive sucrose. On degradation of the protein, the sucrose remained trapped in the cells as a cumulative marker of protein degradation. The method described here depends on the same principles, but uses an adduct of cellobiose and tyramine that is radioiodinated to high specific radioactivity and then covalently attached to protein. Use of the radioiodinated ligand increases the sensitivity of the method at least 100-fold and allows simplified tissue analysis. Proteins derivatized with the radioiodinated ligand were recognized as underivatized proteins both in vitro and in vivo. On degradation of derivatized low-density lipoprotein, the rate of leakage from cultured fibroblasts was only 5% during 24 h. Similarly, on injection of labelled proteins into rats and rabbits, urinary excretion of the label was in all cases less than 10% of total labelled catabolic products recovered 24 h after injection. Examination of the tissue contents of label at two times after injection of labelled asialofetuin or apolipoprotein A1 in rats, and asialotransferrin in rabbits showed that the label did not detectably redistribute between tissues after initial uptake and catabolism; a significant leakage from liver was quantitatively accounted for by label appearing in gut contents and faeces. A simple double-label method was devised to provide a correction for intact protein in trapped plasma, the extravascular spaces, and within cells. By using this method it becomes unnecessary to fractionate tissue samples.     

The stimulation of glycogenolysis and gluconeogenesis in isolated hepatocytes by opioid peptides.

The opioid agonists [leucine]enkephalin, [D-Ala2,D-Leu5]enkephalin and dynorphin-(1-13)-peptide, but not morphine, stimulated the conversion of [2-14C]pyruvate into glucose and glycogenolysis when added directly to isolated hepatocytes. Naloxone produced a small but significant inhibition of both the basal and stimulated rate of incorporation of label into glucose but had no effect on the total glucose output by the cells. The effects of the opioid peptides were mediated by a cyclic AMP-independent mechanism.     

Transitory hematologic effects of moderate exercise are not influenced by iron supplementation

A young women's exercise/fitness class tested the idea that administration of supplemental iron would prevent "sports anemia" that may develop during exercise and training and improve iron status of exercising females of menstrual age. Fifteen women (aged 18-37) were selected for each of three treatment groups: (1) no supplemental iron; (2) 9 mg X d-1 of Fe; and (3) 18 mg X d-1 of Fe (1 US Recommended Daily Allowance). Women exercised at approximately 85% of maximal heartrate for progressively increasing lengths of time in a jogging program and worked up to 45 min of exercise 4 d X week-1 for 8 weeks. Hematologic analysis was performed in weeks 1, 5, and 8. A significant decline in hemoglobin (Hb) concentration and hematocrit (Hct) was observed at week 5 when all data were examined without regard for iron intake; these red cell indices returned to pre-exercise levels by week 8. Reduction of mean cell hemoglobin concentration (MCHC) indicated that the midpoint decline was not caused by simple hemodilution during exercise. Serum ferritin (SF) concentration changed in parallel with Hb and Hct. Although the midpoint decline in SF was not statistically significant, it ruled out the possibility that turnover of red cell iron was directed to storage. Lowered MCHC and SF suggested lower availability of iron during the synthesis of a new generation of red cells. Few iron treatment effects of magnitude were observed. Iron did not prevent the midpoint decline in Hb concentration. Iron intake did not affect SF, serum iron, transferrin saturation, or final Hb, and Hct.(ABSTRACT TRUNCATED AT 250 WORDS)     

Metabolic and physical control of cell elongation rate: in vivo studies in nitella

Several levels of control of elongation rate are revealed through the detailed study of responses of the Nitella internode to abrupt shifts in turgor. The immediate response, which apparently reflects the physical state of the cell, is approximately described by the equation r = (P — Y)m where r is rate, P is pressure, Y is the wall's yielding threshold, and m is related to the wall's apparent fluidity (reciprocal viscosity). Because P and Y are in the range 5 to 6 atmospheres, and (P — Y) is roughly 0.2 atmosphere, elongation rate is initially extremely sensitive to changes in P. A small step-down in turgor (0.7 atmosphere) stops growth, and a similar rise greatly accelerates it. These initial responses are, however, soon (15 minutes) compensated by changes in Y. An apparent metabolism-dependent reaction (azide-sensitive) lowers Y; strain hardening (azide-insensitive) raises it. These two opposing processes, acting on Y, serve as a governor on (P — Y), tending to maintain it at a given value despite changes in P. This ability to compensate is itself a function of turgor. Turgor step-downs are less and less well compensated, leading to lower rate, as turgor falls from 5 atmospheres to about 2 atmospheres where growth appears not to resume. This is the lowest attainable yield value, Y₁. The turgor dependency of compensation reflects a turgor requirement of the Y-lowering (“wall-softening”) process. Thus the relation between steady state, r_s, and turgor is an indirect one, derived from time-dependent alterations of the cell wall. This relationship superficially resembles the instantaneously valid one in that, roughly, r_s = (P — Y₁)m_s. Y₁ and m_s, however, have much lower values than Y and m. The duality of the elongation rate versus turgor relation and the prominent role of Y in regulating rate are the major features of growth control in Nitella.     

On the structure of biological membranes: The double-tiered pattern

A set of six biological membranes has been examined electron microscopically in positively-stained sections at both low and high resolution. At low resolution, all six membranes exhibit a railroad track pattern, while at high resolution all six membranes exhibit a pattern of a double-tier of globular particles. The correspondence between the railroad track pattern and the double-tiered pattern is considered. The relationship of the double-tiered pattern to the bimodal properties of the constituent protein and phospholipid molecules is discussed.     

Induction of a deoxycytidineless state in cultured mammalian cells by bromodeoxyuridine

Bromodeoxyuridine triphosphate is an allosteric inhibitor of ribonucleotide reductase, and bromodeoxyuridine can therefore kill cells by starving them for deoxycytidine nucleotides. The toxicity of bromodeoxyuridine for some cell lines is reduced many fold when deoxycytidine is also present. For example, wild type 3T6 cells can be grown serially in 1.5 × 10^?4 Molar bromodeoxyuridine and 2 × 10^?4 Molar deoxycytidine, remain healthy, and incorporate bromodeoxyuridine extensively into cellular DNA. Some of the numerous effects of this drug on the behavior of cells and viruses may be due to a deoxycytidineless state, rather than to the incorporation of the bromodeoxyuridine into DNA.