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At least 1,675 records · Page 93Linked to original sources
PubMed1976

Spin-labeled stearates as probes for microenvironment of murine thymocyte adenylate cyclase-cyclic adenosine 3':5'-monophosphate system.

The interaction of various spin-labeled compounds with the murine thymocyte adenylate cyclase-cyclic AMP system was investigated. Electron paramagnetic resonance spectra from spin-labeled compounds were used to calculate the order parameter, S, and indicated that the thymocyte plasma membrane is a relatively rigid structure. Increasing concentrations of spin-labeled stearates, but not their corresponding methyl esters, resulted in increased membrane fluidity, partial lysis, and concomitant complete inhibition of cholera toxin-mediated increases in cyclic AMP content. Upon subsequent isolation of plasma membranes from these cells, cholera toxin-stimulated adenylate cyclase activity was also completely inhibited. Direct addition of spin-labeled stearates, but not spin-labeled methyl stearates, to thymocyte homogenates caused a dramatic reduction of basal, cholera toxin-, isoproterenol-, NaF-, and prostaglandin E1-stimulated adenylate cyclase activity. Inhibition was complete within the first minute of addition to homogenates and required approximately 0.2 mM spin-labeled stearate I(12,3) for half-maximal inhibition. This inhibition occurred in the presence or absence of an ATP-regenerating system and was not readily reversible. Furthermore, since the membrane cyclic phosphodiesterase activity was not altered by spin-labeled stearates, their inhibition was attributed to a direct action of stearate spin labels on adenylate cyclase. Neither stearate, methyl stearate, spin-labeled methyl stearates nor 2,2,6,6,-tetramethylpiperidine-1-oxyl (Tempo) altered cell viability or enzyme activities at the concentrations studied. Spin-labeled stearates seemed to intercalate into different areas of the plasma membrane than their corresponding methyl esters. Furthermore, the action of spin-labeled stearates appeared to be on the exterior of the plasma membrane rather than the interior. These results illustrate the presence of multilipid domains and the importance of selected lipids and lipid-protein interactions in the adenylate cyclase-cyclic AMP system. Thymocyte adenylate cyclase is described in terms of a current model for membrane proteins.

Adenylyl Cyclases↗
PubMed1991

Intraglomerular thrombotic tendency and glomerular ADPase. Unilateral impairment of ADPase elicits a proaggregatory microenvironment in experimental glomerulonephritis.

It has been proposed, predominantly from ex vivo studies, that glomerular ADPase may function as an antithrombotic principle within the rat kidney. Therefore, intraglomerular platelet aggregation was studied in vivo in rats after impairment of glomerular ADPase activity using local X-irradiation (20 Gy). Biochemical assays in suspensions of glomeruli obtained from rats 24 hours after local X-irradiation (group I) demonstrated a significant reduction in ADPase activity as compared to sham treated rats (group II; p less than 0.01). Cytochemical observations at the ultrastructural level showed that this reduction in glomerular enzyme activity represents in particular ADPase activity detectable in the basement membrane. Following X-irradiation, intraglomerular platelet aggregation was quantitatively studied in two groups of rats. Both groups received X-irradiation of the left kidney (20 Gy). Twenty-four hours after X-irradiation, animals received an intravenous injection of either 0.5 ml of saline (group III; N = 6) or 0.5 ml of heterologous nephrotoxic serum (NTS; group IV; N = 6). Subsequently, 24 hours after this injection, platelet aggregation in left kidneys was compared with aggregation in contralateral non-X-irradiated kidneys. The results showed that while X-irradiation per se did not induce intraglomerular platelet aggregation as compared with the contralateral kidney (0.20 +/- 0.08% versus 0.17 +/- 0.06% platelet aggregation/glomerulus), a significant increase in platelet aggregation could be demonstrated in X-irradiated kidneys in the early phase of nephrotoxic serum nephritis as compared with the contralateral nephritic kidney (2.45 +/- 0.66% versus 1.37 +/- 0.35% platelet aggregation per glomerulus; p less than 0.005). A potential effect of altered influx of inflammatory cells after X-irradiation could be excluded since no difference in H2O2 producing cells was observed between left and right kidneys. Thus, while ADPase impairment by X-irradiation does not induce platelet aggregation per se, it is clear that in proaggregatory conditions, like in NTS nephritis, the thrombotic tendency, due to decreased glomerular ADPase, is enhanced. These results demonstrate the functional significance of glomerular ADPase activity as an antithrombotic principle following platelet activation in vivo.

Animals↗
PubMedDate not supplied

The testicular and epididymal luminal amino acid microenvironment in the rat.

Concentrations of amino acids were measured in arterial and testicular venous blood, and in fluids from the seminiferous tubule, rete testis, and the caput, corpus, and cauda epididymidis. There were no significant differences in the concentrations of any amino acids between arterial and testicular venous blood, whereas there were significant differences between arterial/venous blood and testicular interstitial fluid. The predominant amino acids measured within seminiferous tubule fluid (STF) and rete testis fluid (RTF) were glycine, alanine, glutamate, and glutamine. RTF contained approximately equal concentrations of basic and total amino acids, but 17 times higher acidic amino acids and 1.2 and 1.3 times lower uncharged polar and nonpolar amino acids, respectively, compared to STF. The concentration of total amino acids within caput fluid reached over 50 nmol/L, but then declined to approximately 50% and 0.1% of caput for corpus and cauda, respectively. The predominant amino acids measured within epididymal luminal fluids were glutamate and taurine; glutamate contributed to approximately 90% of the total amino acids measured in caput fluid. The presence of glutamate and taurine within the epididymal lumen is due primarily to a direct contribution from the epididymal epithelium, as measured using the split-drop stopped-flow microperfusion technique. Several other amino acids within the lumen also originate from the epididymal epithelium. Amino acids contribute approximately 20%, 9%, and 2% of the total osmolality of caput, corpus, and cauda fluid, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗
PubMed1990

Dissecting the hematopoietic microenvironment. VIII. Clonal isolation and identification of cell types in murine CFU-F colonies by limiting dilution.

In this study it was found that three kinds of cells, fibroblasts, endothelial cells, and macrophages, exist in fibroblastic colony-forming unit (CFU-F) colonies, consistent with the findings of others. However, when low enough bone marrow stromal cell numbers were cultured per dish, pure fibroblast, endothelial, and macrophage colonies were observed. When the cell number cultured in each dish was increased, all colonies were of mixed cell type. This evidence strongly suggests that the macrophages and endothelial cells contained in CFU-F colonies come from locally stimulated growth resulting in contamination and not from a common progenitor with fibroblasts or each other. In some longer term experiments, adipocytes were seen to appear within the colonies of late-passage pure fibroblast cultures.

Animals↗
PubMed1990

The effect of cocaine on oocyte development and the follicular microenvironment in the rabbit.

We examined the effects of cocaine exposure in the rabbit on in vitro oocyte development and on steroidal content of follicular fluid (FF) and serum progesterone (P). Cocaine hydrochloride (0, 10, 20, 40, or 80 mg/kg) was administered daily subcutaneously for 5 days to New Zealand White female rabbits before superovulation. On the last day of cocaine administration, animals were given human chorionic gonadotropin intravenously, and laparotomy was performed 6 to 8 hours later. During laparotomy, ovaries were removed, the number of follicles recorded, oocytes retrieved, and FF was obtained. In vitro fertilization (IVF) was then performed on the oocytes and the rate of cleavage observed. For all cocaine dosage groups, no differences were observed in the number of follicles present, number of oocytes retrieved, or IVF and cleavage rates. Cocaine did, however, decrease periovulatory serum P, and FF P, whereas FF estradiol concentrations increased. This suggests that short-term cocaine exposure affects the follicular steroid milieu, possibly by delaying granulosa cell luteinization.

Animals↗
PubMedDate not supplied

[The role of the hemopoietic microenvironment on the hemopoiesis-stimulating effect of cystamine].

The influence of cystamine delivered in a radioprotective dose before and after irradiation of mouse-recipients (8 Gy) on the effectiveness of exogenous bone marrow cloning has been investigated. Cystamine administered prior to irradiation exerts a protective effect on CFUs and also causes an increase in the number of splenic colonies grown from CFUs of the transplanted bone marrow. With cystamine administered after irradiation the protective effect is absent, but the CFUs number in the femur increases in recipients transplanted with intact bone marrow in comparison with those transplanted without cystamine. It is believed, that in addition to the specific protective mechanism of action of radioprotectors, there is a nonspecific mechanism of increasing the proliferation of protected stem cells that is connected with the stimulatory effect of radioprotective agents on the haemopoietic stroma elements.

Animals↗
PubMed1990

Physical microenvironment of platelet membrane in acute myocardial infarction.

Patients with acute myocardial infarction have more reactive platelets than those from normal population. These pathological platelets had more viscous plasma membrane, as inferred from fluorescence polarisation studies, and a lower fusion activation energy (delta E) of the membrane, reflecting a higher degree of order within lipid-lipid interactions.

Blood Platelets↗
PubMed1990

Dissecting the hematopoietic microenvironment. VI. The effects of several growth factors on the in vitro growth of murine bone marrow CFU-F.

The effects of several growth factors on the proliferation of fibroblastic colony-forming units (CFU-F) were studied. In the present study CFU-F colonies were found to consist of fibroblasts, macrophages, and endothelial cells. Growth factors, including interleukin 3 (IL-3), interleukin 1 alpha (IL-1 alpha), epidermal growth factor (EGF), fibroblast growth factor (FGF), platelet-derived growth factor (PDGF), macrophage colony-stimulating factor (M-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), and buffalo rat liver cell-conditioned medium (BRL-CM) were tested for stimulation of the proliferation of CFU-F in a standard culture in both 2% and 15% serum. Overall, the colony numbers produced in 15% serum were much higher than in 2% serum with or without growth factors. However, the influence of several growth factors on CFU-F cultured in 2% serum was relatively greater than in 15% serum when compared to controls. The stimulation of CFU-F by FGF only occurred in culture with 15% serum, and the stimulation by PDGF only occurred with 2% serum. Overall, the strongest stimulations were produced by PDGF, IL-3, and BRL-CM. Combining the other growth factors with IL-3, PDGF, or IL-1 alpha enhanced their effects only modestly. The stimulation by growth factors included increases of the cell numbers between and within colonies as well as an increase in the number of colonies. The study produced results that suggest a complex interaction mediated by growth factors between fibroblasts and other stromal cells within the CFU-F colonies and within the bone marrow itself.

Animals↗
PubMed1990

Dissecting the hematopoietic microenvironment. VII. The production of an autostimulatory factor as well as a CSF by unstimulated murine marrow fibroblasts.

Purified normal murine bone marrow-derived fibroblasts were shown to produce a factor that stimulates the in vitro growth of fibroblastic colony-forming unit (CFU-F) colonies. Conditioned medium from the purified fibroblasts (F-CM) also stimulated pure marrow fibroblasts themselves. Analysis of the F-CM detected the presence of macrophage colony-stimulating factor (M-CSF), and low levels of interleukin 1 (IL-1) and interleukin 6 (IL-6), but no detectable levels of interleukin 3 (IL-3), interleukin 5 (IL-5), granulocyte-macrophage colony-stimulating factor (GM-CSF), or granulocyte colony-stimulating factor (G-CSF). Macrophages and endothelial cells, freed from other bone marrow components, required the F-CM if no other growth factors were added. We conclude that F-CM contains an autocrine factor, which the evidence suggests is IL-1, for bone marrow fibroblasts, and a paracrine factor (CSF-1) for macrophages and/or endothelial cells.

Animals↗
PubMed1990

Hemopoietic progenitor cell binding to the stromal microenvironment in vitro.

Primitive clonogenic progenitor cells in human bone marrow bind to preformed marrow-derived stromal layers in vitro and generate colonies of blast cells. The binding interaction does not require calcium or magnesium ions and occurs equally well in serum-free and serum-supplemented culture medium. It does not appear to involve known cell adhesion molecules (CAMs) for which monoclonal antibodies are available (integrins, N-CAM, LFA-1, and ICAM-1), and we were unable to demonstrate a role for the progenitor cell antigen CD34 in progenitor cell adhesion to cultured stroma. The CAM expressed by the blast colony-forming cells may exist in transmembrane or phosphatidylinositol (PI)-linked forms because it is only partially degraded by exposure to trypsin or to PI-specific phospholipase C. However, binding of these cells to stroma is not prevented in the presence of monoclonal antibodies reacting with known PI-linked structures (Thy-1, CD14, and CD16). It is either masked by neuraminidase-sensitive residues or is no longer expressed as cells mature, respectively, along the granulocytic or erythroid lineages. The properties of the hemopoietic progenitor CAM are discussed with reference to the properties of other CAMs and of hemopoietic progenitor cell markers.

Antibodies, Monoclonal↗
PubMed1978

The effect of evolution on homologous proteins: a comparison between the chromophore microenvironments of Italian water buffalo (Bos bubalus, L.) and sperm whale apomyoglobin.

The perturbing effect of guanidium hydrochloride and pH on the molecular structure of water buffalo apomyoglobin has been investigated by circular dichroism in the far and near ultraviolet and by fluorescence. In the wavelength region between 320 and 260 nm the circular dichroic spectrum of the globin is highly structured and the contributions of the aromatic chromophores have been resolved. Buffalo apomyoglobin undergoes a structural transition at neutral pH which involves elements of the secondary and tertiary structure, as indicated by changes of dichroic activity of the peptide and aromatic chromophores and the fluorescence of the two tryptophanyl residues. The possibility of charge-transfer complex between indole and imidazole is discussed. A major structural transition with abrupt unfolding takes place in the pH region between 5.6 and 4.3. Below pH 4.3 the peptide helical residues, which survive the acid transition, appear to be resistent to further acidification to pH 2.0 while tryptophanyl emission is quenched and shifted to longer wavelengths. A structural transition occurs also in alkali above pH 10, which has been detected by the same techniques. The relationships between buffalo and sperm whale apomyoglobin are discussed.

Animals↗
PubMed1986

The bursal microenvironment: phenotypic characterization of the epithelial component of the bursa of Fabricius with the use of monoclonal antibodies.

Monoclonal antibodies were raised against newborn chick bursa of Fabricius, and here we describe two antibodies, BEP-1 and BEP-2, which react selectively with the epithelial component of the bursa of Fabricius. In previous studies, using quail chick chimeric bursas, we have demonstrated that the epithelium of the bursal rudiment, presumably of endodermal origin, gives rise to the epithelium lining the bursal lumen, the basement membrane-associated epithelium and the network of reticular cells of the medulla, while the interfollicular connective cells are derived from the mesoderm. When tested in indirect immunofluorescence assay on bursa tissue sections or cell suspensions, BEP-1 reacts with a surface antigen present on all the epithelial cells of the bursa and could be used as a marker for this cell lineage. BEP-2 binds to an intracytoplasmic antigen that is present in about 5% of cells, representing the epithelial cells, and which is excreted in the medulla. BEP-2 also reacts with the epithelial cells of the thymic medulla and with the mucin-secreting goblet cells of the intestinal villi. A rabbit antiserum raised against human cytokeratin gives a different pattern of reactivity on bursal tissue compared to BEP-1 and BEP-2, tentatively suggesting that these two antibodies do not bind to keratin-like molecules. During ontogeny, BEP-1 reactivity appears in bursal epithelium from the early stages of bursal ontogeny (8 days). BEP-2 reactivity is detected around hatching time. BEP-1 and BEP-2 do not show any antigenic heterogeneity among the epithelial cells of the bursa.

Aging↗