A 31P-NMR study of tissue respiration in working dog muscle during reduced O2 delivery conditions.

To investigate the role of tissue oxygenation as one of the control factors regulating tissue respiration, 31P-nuclear magnetic resonance spectroscopy (31P-NMR) was used to estimate muscle metabolites in isolated working muscle during varied tissue oxygenation conditions. O2 delivery (muscle blood flow x arterial O2 content) was varied to isolated in situ working dog gastrocnemius (n = 6) by decreases in arterial PO2 (hypoxemia; H) and by decreases in muscle blood flow (ischemia; I). O2 uptake (VO2) was measured at rest and during work at two or three stimulation intensities (isometric twitch contractions at 3, 5, and occasionally 7 Hz) during three separate conditions: normal O2 delivery (C) and reduced O2 delivery during H and I, with blood flow controlled by pump perfusion. Biochemical metabolites were measured during the last 2 min of each 3-min work period by use of 31P-NMR, and arterial and venous blood samples were drawn and muscle blood flow measured during the last 30 s of each work period. Muscle [ATP] did not fall below resting values at any work intensity, even during O2-limited highly fatiguing work, and was never different among the three conditions. Muscle O2 delivery and VO2 were significantly less (P < 0.05) at the highest work intensities for both I and H than for C but were not different between H and I. As VO2 increased with stimulation intensity, a larger change in any of the proposed regulators of tissue respiration (ADP, P(i), ATP/ADP.P(i), and phosphocreatine) was required during H and I than during C to elicit a given VO2, but requirements were similar for H and I.(ABSTRACT TRUNCATED AT 250 WORDS)     

Intestinal synthesis of 24-keto-1,25-dihydroxyvitamin D3. A metabolite formed in vivo with high affinity for the vitamin D cytosolic receptor

24-Keto-1,25-dihydroxyvitamin D3 has been identified as an intestinal metabolite of 1,25-dihydroxyvitamin D3 by ultraviolet absorbance, mass spectroscopy, and chemical reactivity. The metabolite was produced from 1,25-dihydroxyvitamin D3 and 1,24R,25-trihydroxyvitamin D3 in rat intestinal mucosa homogenates. 24-Keto-1,25-dihydroxyvitamin D3 is present in vivo in the plasma and small intestinal mucosa of rats fed a stock diet, receiving no exogenous 1,25-dihydroxyvitamin D3, and in the plasma and small intestinal mucosa of rats dosed chronically with 1,25-dihydroxyvitamin D3. 24-Keto-1,25-dihydroxyvitamin D3 has affinity equivalent to 1,24R,25-trihydroxyvitamin D3 for the 3.7 S cytosolic receptor specific for 1,25-dihydroxyvitamin D3 in the intestine and thymus. In cytosolic preparations contaminated with the 5 S vitamin D-binding protein, both metabolites are about 7-fold less potent than 1,25-dihydroxyvitamin D3. In contrast, in cytosolic preparations largely free of the 5 S binding protein, both metabolites are equipotent with the parent compound. No evidence was obtained supporting a substantial presence of 23-keto-1,25-dihydroxyvitamin D3 in vivo; nor was the latter compound generated in detectable amounts from 1,25-dihydroxyvitamin D3 by intestinal homogenates. Thus, C-24 oxidation is a significant pathway of intestinal 1,25-dihydroxyvitamin D3 metabolism that produces metabolites with high affinity for the cytosolic receptor which mediates vitamin D action.     

Vesicle-to-cell protein transfer: insertion of band 3, the erythrocyte anion transporter, into lymphoid cells

Band 3, the erythrocyte anion transporter, transfers spontaneously between human red cells and model membranes. During incubation of intact erythrocytes with sonicated dimyristoylphosphatidylcholine vesicles, the transporter inserts in functional form and native orientation into the liposome bilayer, with the cytoplasmic segment of the protein contacting the lumen of the vesicle [Newton, A. C., Cook, S. L., & Huestis, W. H. (1983) Biochemistry 22, 6110-6117; Huestis, W. H., & Newton, A. C. (1986) J. Biol. Chem. 261, 16274-16278]. When band 3-vesicle complexes are incubated with erythrocytes whose native band 3 has been inhibited irreversibly, reverse transfer of the protein restores anion transport capacity to the cells [Newton, A. C., Cook, S. L., & Huestis, W. H. (1983) Biochemistry 22, 6110-6117]. Here we report the vesicle-mediated transfer of band 3 to human peripheral blood lymphocytes and to cultured murine lymphoma cells (BL/VL3). Subsequent to incubation with protein-vesicle complexes, both lymphoid cell types exhibit a 2-4-fold increase in the rate of chloride uptake. This enhanced permeability is inhibited greater than or equal to 98% by the exofacial band 3 inhibitor 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid, consistent with right-side-out insertion of functional band 3 into the lymphoid cell membrane.     

Functional anatomy of the musculature of the trachea.

The smooth musculature of the human trachea was studied and compared with earlier observations in the rabbit. The results may be summarized as follows: 1. The annular m. constrictor tracheae, previously observed in the rabbit, has also been identified in the human trachea. 2. Longitudinal muscle fibers outside the constrictor musculature were observed in man. These fibers are rudimentary and appear to be of no functional importance. 3. From a functional point of view, it appears justified to regard the outer tracheal musculature largely as a constrictor musculature. 4. The main function of the outer musculature of the trachea and the elastic cartilaginous arches is to maintain the stability of the tracheal wall. 5. The variation of the lumen of the trachea is mainly controlled by the m. trachealis in the pars membranacea.     

Altered patterns of protein phosphorylation in articular chondrocytes treated with interleukin-1 or synovial cytokines

Cultures of lapine articular chondrocytes were exposed to purified, human, recombinant interleukin-1 alpha or partially purified preparations of lapine, synovial, cytokines in the presence of [32P]orthophosphate. After 30 min incubation, phosphoproteins were extracted from the cells, separated by two-dimensional gel electrophoresis and visualized autoradiographically. Analysis of the autoradiograms revealed that interleukin-1 and the synovial factors produced marked changes in the pattern of protein phosphorylation. The synovial cytokines induced many of the same changes as interleukin-1, as well as a number of unique changes. This finding is consistent with the notion that, in addition to interleukin-1, synoviocytes secrete other cytokines which modulate the metabolism of chondrocytes. These data support the idea that signal transduction in chondrocytes responding to interleukin-1 involves the activation of one or more protein kinases.     

Collagen crosslinking and cartilage glycosaminoglycan composition in normal and scoliotic chickens

The amounts of lysine-derived crosslinks in collagens from tendon, cartilage, intervertebral disc, and bone and changes in the composition of sternal cartilage glycosaminoglycans were estimated in two lines of chickens, a control-isogenic line and a line that develops scoliosis. In the scoliotic line, scoliosis first appears at 3-4 weeks and progressively increases in severity and incidence so that 90% of the birds express the lesion by week 10. We have reported previously that cartilage, tendon, and bone collagens from scoliotic birds are more soluble than corresponding collagens from normal birds. Herein, collagen crosslinking and altered proteoglycan metabolism are examined as possible mechanisms for the differences in collagen solubility. At 1 week of age there were fewer reducible crosslinking amino acids (hydroxylysinonorleucine, dihydroxylysinonorleucine, and lysinonorleucine) in collagens from sternal cartilage and tendon in the scoliotic line than in the isogenic line. However, by week 3 and at weeks 5 or 7 values were similar in both groups. The amounts of hydroxypyridinium in vertebral bone and intervertebral disc collagen were also similar in both groups of birds. Consequently, differences in collagen crosslinking do not appear to be a persistent developmental defect underlying the expression of scoliosis in the model. However, differences were observed in cartilage proteoglycans and glycosaminoglycans from the scoliotic line that were not present in cartilage from the isogenic line. The average molecular weight of the uronide-containing glycosaminoglycans was 30% less in the scoliotic line than in the isogenic line, i.e., 12,000 compared to 18,000. The size distribution of cartilage proteoglycans from the scoliotic line also differed from that of proteoglycans from the isogenic line.(ABSTRACT TRUNCATED AT 250 WORDS)