Binding of Chemotactic Collagen-Derived Peptides to Fibroblasts: THE RELATIONSHIP TO FIBROBLAST CHEMOTAXIS

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Abstract

We previously showed that collagen, α-chains, and collagen-derived peptide fragments induce chemotactic migration of human fibroblasts in vitro. We now describe biochemical and immunological evidence showing there are binding sites for collagen peptides on fibroblast membranes.

Authors

Thomas M. Chiang, Arnold E. Postlethwaite, Edwin H. Beachey, Jerome M. Seyer, Andrew H. Kang

Heterozygote Expression in Propionyl Coenzyme A Carboxylase Deficiency: DIFFERENCES BETWEEN MAJOR COMPLEMENTATION GROUPS

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Abstract

We measured propionyl coenzyme A carboxylase (PCC) activity in extracts of skin fibroblasts and peripheral blood leukocytes from controls and obligate heterozygotes for PCC deficiency. 6 heterozygotes were from the pcc A complementation group; 12 were from the other major complementation group, designated pcc C. Mean PCC activity in fibroblast extracts from pcc A heterozygotes was 52% of that in controls, whereas mean PCC activity in pcc C heterozygotes was indistinguishable from that of controls. Similar results were obtained with extracts of peripheral blood leukocytes. In none of eight families (three pcc A and five pcc C) in which PCC activity was studied in both parents of an affected child were significant intrafamilial differences observed. The activities of two other mitochondrial enzymes (β-methyl-crotonyl CoA carboxylase and glutamate dehydrogenase) were comparable in controls and both groups of heterozygotes. Whereas the data from pcc A heterozygotes are consistent with expected gene dosage effects, those from pcc C heterozygotes are not. Inasmuch as mammalian PCC is a large molecular weight tetramer, each protomer of which is probably composed of two nonidentical subunits, the latter results are most consistent with unbalanced rates of synthesis and(or) degradation of the two subunits in normal cells with compensatory balancing in pcc C heterozygotes.

Authors

Barry Wolf, Leon E. Rosenberg

Renin-Angiotensin System Inhibition in Conscious Dogs during Acute Hypoxemia: EFFECTS ON SYSTEMIC HEMODYNAMICS, REGIONAL BLOOD FLOWS, AND TISSUE METABOLISM

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Abstract

The role of the renin-angiotensin system in mediating the circulatory and metabolic responses to hypoxia was studied in three groups of conscious dogs that were infused continuously with normal saline, teprotide (10 μg/kg per min), and saralasin (1 μg/kg per min), respectively. Hypoxia was produced by switching from breathing room air to 5 or 8% oxygen-nitrogen mixture. Plasma renin activity increased from 2.3±0.4 to 4.9±0.8 ng/ml per h during 8% oxygen breathing, and from 2.8±0.4 to 8.4±1.8 ng/ml per h during 5% oxygen breathing. As expected, cardiac output, heart rate, mean aortic blood pressure, and left ventricular dP/dt and dP/dt/P increased during both 5 and 8% oxygen breathing in the saline-treated dogs; greater increases occurred during the more severe hypoxia. Teprotide and saralasin infusion diminished the hemodynamic responses to 5% oxygen breathing, but did not affect the responses to 8% oxygen breathing significantly. In addition, the increased blood flows to the myocardium, kidneys, adrenals, brain, intercostal muscle, and diaphragm that usually occur during 5% oxygen breathing were reduced by both agents. These agents also reduced the increases in plasma norepinephrine concentration during 5% oxygen breathing, but had no effects on tissue aerobic or anaerobic metabolism.

Authors

Chang-Seng Liang, Haralambos Gavras

Human Antihapten Antibodies in Trimellitic Anhydride Inhalation Reactions: IMMUNOGLOBULIN CLASSES OF ANTI-TRIMELLITIC ANHYDRIDE ANTIBODIES AND HAPTEN INHIBITION STUDIES

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Abstract

Inhalational exposure to trimellitic anhydride (TMA) produces an immediate-type asthmatic or a late respiratory systemic syndrome in certain workers after a latent period of work exposure. TMA has been found to react with proteins to produce a hapten-protein complex (trimellitate [TM] protein) or become hydrolyzed in aqueous, alkaline solutions to produce a salt, NaTM. Using a solid-phase radioimmunoassay technique, antibodies of different Ig classes were detected against TM-protein conjugates. IgE antibody was detected in three of five workers with asthma. IgG and IgA antibodies were detected in most exposed workers but higher levels of antibody were found in symptomatic workers even after long periods without direct TMA exposure. IgM antibody activity against TM-human serum albumin (TM-HSA) was detected but did not differentiate symptomatic from asymptomatic workers. NaTM served as a hapten for study because it does not react with proteins to form a hapten-protein complex as TMA does. The NaTM only partially inhibited IgG antibody activity against TM-HSA and much smaller amounts of TM-HSA than of NaTM were required to neutralize IgG antibody. A similar result was found with TM-ovalbumin. The latter results suggest that some IgG antibody is directed against a TM-protein moiety, probably a TM-amino acid determinant. In contrast to IgG, marked inhibition by NaTM of IgA and IgM antibody against TM-HSA was found in the sera studied.

Authors

Roy Patterson, C. Raymond Zeiss, Mary Roberts, Jacob J. Pruzansky, Peter Wolkonsky, R. Chacon

Cell-Cell Interaction in Erythropoiesis: ROLE OF HUMAN MONOCYTES

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Abstract

Erythroid burst forming units (BFU-E) are proliferative cells present in peripheral blood and bone marrow which may be precursors of the erythroid colony forming cell found in the bone marrow. To examine the possible role of monocyte-macrophages in the modulation of erythropoiesis, the effect of monocytes on peripheral blood BFU-E proliferation in response to erythropoietin was investigated in the plasma clot culture system. Peripheral blood mononuclear cells from normal human donors were separated into four fractions. Fraction-I cells were obtained from the interface of Ficoll-Hypaque gradients (20-30% monocytes; 60-80% lymphocytes); fraction-II cells were fraction-I cells that were nonadherent to plastic (2-10% monocytes; 90-98% lymphocytes); fraction-III cells were obtained by incubation of fraction-II cells with carbonyl iron followed by Ficoll-Hypaque centrifugation (>99% lymphocytes); and fraction-IV cells represented the adherent population of fraction-II cells released from the plastic by lidocaine (>95% monocytes). When cells from these fractions were cultured in the presence of erythropoietin, the number of BFU-E-derived colonies was inversely proportional to the number of monocytes present (r = −0.96, P < 0.001). The suppressive effect of monocytes on BFU-E proliferation was confirmed by admixing autologous purified monocytes (fraction-IV cells) with fraction-III cells. Monocyte concentrations of ≥20% completely suppressed BFU-E activity. Reduction in the number of plated BFU-E by monocyte dilution could not account for these findings: a 15% reduction in the number of fraction-III cells plated resulted in only a 15% reduction in colony formation. These results indicate that monocyte-macrophages may play a significant role in the regulation of erythropoiesis and be involved in the pathogenesis of the hypoproliferative anemias associated with infection and certain neoplasia in which increased monocyte activity and monopoiesis also occur.

Authors

John J. Rinehart, Esmail D. Zanjani, Benet Nomdedeu, Bobby J. Gormus, Manuel E. Kaplan

Prostaglandin Suppression of Mitogen-Stimulated Lymphocytes In Vitro: CHANGES WITH MITOGEN DOSE AND PREINCUBATION

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Abstract

In this study we further characterize the properties of the prostaglandin-producing suppressor cell. Overnight preincubation of peripheral blood mononuclear cells results in an increased response of the cells to phytohemagglutinin or Concanavalin A compared to the response of fresh cells. This increase in mitogen response with preincubation was similar in magnitude to the increase in mitogen response of fresh cells after the addition of indomethacin. The two manipulations were not additive; that is, after preincubation, indomethacin caused much less enhancement of mitogen stimulation of peripheral blood mononuclear cells (100 ± 12% increase before preincubation vs. 12 ± 6% after preincubation; mean±SEM, P < 0.001). Preincubated cells also lose sensitivity to inhibition by exogenous prostaglandin E2. It requires the addition of 100- to > 1,000-fold more exogenous PGE2 to produce comparable inhibition of phytohemagglutinin-stimulated preincubated cells than is required for inhibition of phytohemagglutinin-stimulated fresh cells.

Authors

James S. Goodwin, Ronald P. Messner, Glenn T. Peake

Suppressor-Cell Dysfunction in Systemic Lupus Erythematosus: CELLS INVOLVED AND IN VITRO CORRECTION

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Abstract

To characterize the cell(s) responsible for the suppressor-cell dysfunction in active systemic lupus erythematosus (SLE), we fractionated blood mononuclear cells into thymus-derived (T), bone marrow-derived (B), and monocyte-depleted populations. Various cell populations from active SLE, inactive SLE, or normals, were activated with Concanavalin A, washed, and then co-cultured with active SLE cells. Soluble immune response suppressor (SIRS) from culture supernates of the activated cells was also used for the possible correction of the suppressor-cell dysfunction. Suppression was tested by enumerating DNA-binding cells by radioautography and by quantitating anti-DNA antibody in culture supernates by radioimmunoassay; and immunoglobulin was tested in cells and supernates by the immunofluorescence and the immunofluor techniques, respectively. Except for the numbers of DNA-binding cells, which were not suppressed, all the other three parameters in co-cultures with cells from active SLE patients were suppressed by Concanavalin A-activated cells (P < 0.001), or by SIRS (P < 0.05) from normals or inactive SLE patients. Concanavalin A-activated autologous or allogeneic active SLE cells and nonactivated cells from active or inactive SLE failed to suppress the various B-cell functions. Nonactivated normal cells suppressed levels of anti-DNA and immunoglobulin in supernates (P < 0.05).

Authors

Akira Sagawa, Nabih I. Abdou

Chronic Myelocytic Leukemia: ORIGIN OF SOME LYMPHOCYTES FROM LEUKEMIC STEM CELLS

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Abstract

In three patients with chronic myelocytic leukemia who were heterozygous at the X-linked glucose-6-phospháte dehydrogenase locus, lymphocytes were studied to determine if they had the same stem cell origin as the leukemic myeloid cells. Normal tissues such as skin had both B and A glucose-6-phosphate dehydrogenase isoenzymes, but the leukemic myelogenous cells displayed only one isoenzyme type, consistent with their clonal origin. A population of cells with undoubted thymus-derived (T)-lymphocyte characteristics had both isoenzymes. Presumably, then, these T cells did not arise from the leukemic stem cell, either because they antedated the development of leukemia in that stem cell or, more likely, because they arose from progenitors not involved by the disease. In contrast, another population of lymphocytes showed only one isoenzyme type, suggesting that it arose from the chronic myelocytic leukemia stem cell. However, although this population contained many cells with the characteristics of bone marrow-derived (B) lymphocytes, it is not certain that the single enzyme produced by the cells over all can be attributed to B lymphocytes rather than to contaminating non-B-lymphoid cells.

Authors

Philip J. Fialkow

Biosynthesis of Chenodeoxycholic Acid in Man: STEREOSPECIFIC SIDE-CHAIN HYDROXYLATIONS OF 5β-CHOLESTANE-3α, 7α-DIOL

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Abstract

Stereospecific side-chain hydroxylations of 5β-cholestane-3α, 7α-diol were studied in mitochondrial and microsomal fractions of human liver. Incubation of 5β-cholestane-3α, 7α-diol resulted in hydroxylations at C-12, C-24, C-25, and C-26. Hydroxylations at C-24 and C-26 were accompanied by the introduction of additional asymmetric carbon atoms at C-24 and C-25 respectively, that led to the formation of two distinct pairs of diastereoisomers, namely 5β-cholestane-3α, 7α,24-triols (24R and 24S) and 5β-cholestane-3α, 7α,26-triols (25R and 25S). A sensitive and reproducible radioactive assay to measure the formation of the different biosynthetic 5β-cholestanetriols was developed. Optimal assay conditions for human mitochondrial and microsomal systems were tentatively established.

Authors

S. Shefer, F. W. Cheng, A. K. Batta, B. Dayal, G. S. Tint, G. Salen

Tissue Guanosine-3′,5′-Cyclic Monophosphate Levels and Soluble Guanylate Cyclase Activity: A POSITIVE CORRELATION DURING UNILATERAL CRYPTORCHIDISM IN THE RAT TESTIS

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Abstract

The relationship between the subcellular distribution of guanylate cyclase and tissue guanosine-3′,5′-cyclic monophosphate (cGMP) levels was investigated in rat testes after surgically induced unilateral cryptorchidism. Placement of one of a testis pair in the abdominal cavity results in loss of testicular weight and function in the abdominal testis whereas the remaining scrotal testis appears to be functionally normal. Within 5 days after surgery, tissue cGMP levels were increased by twofold in the abdominal testis. A fourfold elevation was noted from 10 to 30 days after surgery. Whereas the homogenate guanylate cyclase activity was only slightly elevated 10 and 20 days postoperatively, a 200% increase in the soluble guanylate cyclase activity was seen at 5 days. Between 10 and 30 days, the rise in activity was >250% (P < 0.01). An increase in soluble guanylate cyclase activity was noted when the data were expressed as per milligram protein, per milligram DNA or per whole testis. Conversely, particulate guanylate cyclase activity was reduced by 40% in the cryptorchid testis. Kinetic analysis of the soluble enzyme prepared from abdominal and scrotal testes yielded linear Line-weaver-Burke plots for both enzyme preparations with an identical Km for guanosine triphosphate, but a three-fold higher maximal velocity for the abdominal enzyme. When the soluble guanylate cyclases from both testes were mixed and assayed together, the activities were additive rather than exhibiting synergism or inhibition. These experiments indicate that the altered Vmax is not due to a transferable activator or inhibitor.

Authors

W. Austin Spruill, Alton L. Steiner, H. Shelton Earp