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N L Baenziger

Publications and source records attributed to N L Baenziger.

At least 19 recordsLinked to original sources
PubMed2002

Bradykinin receptor modulation in cellular models of aging and Alzheimer's disease.

Human fibroblast cell culture systems have been used to model both molecular events associated with the aging process and the biochemical anomalies found in the aging-associated neurodegenerative disorder Alzheimer's disease (AD). We demonstrate modulation of bradykinin (BK) B2 receptors that results in Intermediate (I, Kd 2.5-5 nM) and Low (L, Kd 44 nM) receptor affinity states in two cellular model systems that target aging and aging-associated disorders: the human lung fibroblast cell line WI-38 model for cellular aging and a skin fibroblast cell line from a patient with early onset familial Alzheimer's disease. In both cellular models the generation of I and L BK B2 receptors is extremely rapid, occurring within 1 min of activation of protein kinase C (PKC) by phorbol ester. Blocking phosphoprotein phosphatase activity further augments the cellular content of I and L receptors in the Alzheimer's skin fibroblast cell line. These two lines of evidence suggest that a phosphorylation cascade modifying the receptors is responsible for the I and L states. The I and L receptors remain biologically active and enhance cellular responsiveness to elevated levels of BK that are found in tissue injury, one of the major risk factors for development of Alzheimer's disease. The Alzheimer's disease skin fibroblast cell line presents a cellular environment highly enriched in the amyloid Abeta1-42 peptide that is the hallmark of Alzheimer's plaque lesions in the brain. This Abeta-rich environment may serve to foster the signal transduction mechanism that generates I and L BK B2 receptors.

Aging↗
PubMed1996

Human lung fibroblasts express multiple means for enhanced activity of bradykinin receptor pathways.

Human lung fibroblasts represent important targets for the biologic activities of bradykinin (BK). We have identified multiple mechanisms in these cells which may extend their potential for BK receptor responsiveness, particularly with regard to generation of arachidonate metabolites. These fibroblasts can constitutively express B2 and B1 BK receptors concurrently, both coupled to the pathway for arachidonate metabolism resulting in generation of PGE2 and the potent vasoactive lipid mediator Thromboxane A2. Although expression patterns for B2 and B1 receptors have classically been regarded as 'constitutive' and 'inducible', respectively, we demonstrate that in human lung fibroblasts both can be expressed spontaneously at equivalent biologic activity levels without selective induction by other mediators. Concurrent B2/B1 receptor expression extends the scope of fibroblast response potential to both BK and des-Arg9-BK in the same time frame. We have identified additional short-term and long-term cellular events, involving both protein kinase pathways through which BK receptors act and those which act upon BK receptors, that result in enhanced BK receptor response potential. These properties of BK receptors may affect whether fibroblast behaviors maintain controlled activities of normal homeostasis or foster escalating cellular responses which may influence the progression of certain human disease states.

Arachidonic Acid↗
PubMed1996

Protein kinases A and C rapidly modulate expression of human lung fibroblast B2 bradykinin receptor affinity forms.

WI-38 and IMR90 human lung fibroblasts express B2 receptors for the peptide mediator bradykinin. These G-protein-coupled receptors which control cell growth, protein synthesis, and prostaglandin E2 (PGE2) production occur in three affinity forms, high (H, KD 440 pM), intermediate (I, KD 5.6 nM), and low (L, KD 42 nM). Utilizing specific monoclonal antireceptor antibodies which are able to distinguish among these B2 bradykinin receptor forms, we demonstrate regulated differential enhancement of their expression in fibroblasts. Activation of cellular second messenger regulatory pathways based on protein kinase C or protein kinase A drives B2 receptor affinity form expression in opposite directions, both of which are relevant to the levels of human bradykinin generation in vivo in the tissues of origin for these fibroblasts. On a spontaneous basis WI-38 human lung fibroblasts most frequently express the L form alone or the I+L forms concurrently. Activation of protein kinase C augments expression of both I and L affinity receptors within 30 min, increasing receptor number and enhancing PGE2 production. In contrast, activation of protein kinase A by 8-bromo-cAMP or forskolin enhances receptor expression and PGE2 production instead at the I to H types of affinity forms within 30 min. The effects of both kinase systems are blocked by serine/threonine (Ser/Thr) protein kinase inhibitors, indicating a role for phosphorylation at Ser or Thr residues in determining the cellular expression of bradykinin B2 receptor affinity forms. An increase in immunoprecipitable I form bradykinin receptors is detectable within 20 to 30 min after activation of either protein kinase C or protein kinase A. This time frame emphasizes the ability of human fibroblasts for rapid mobilization of B2 receptor affinity forms. Regulated expression of this repertoire of bradykinin B2 receptors at the level of receptor number and concurrent activity allows fibroblasts a sensitive means to adjust their responses to their cellular environment utilizing Ser/Thr phosphorylation events.

Bradykinin↗
PubMed1994

Histamine degradative uptake by cultured human pulmonary vascular endothelial cells utilizes an inflammatory cell diamine oxidase.

Neutrophil-endothelial interactions, altered clearance properties of the lung toward vasoactive mediators, and damaging effects of histamine that target the lung represent prominent elements of the inflammatory response at the systemic level. The pulmonary vasculature is unusual in that, unlike other tissues and vascular beds, it normally does not metabolize circulating histamine in vivo, although histamine-metabolizing enzyme activities have been detected in disrupted lung tissue. We have therefore explored the capability of human pulmonary artery endothelial cells in culture to express the receptor-mediated histamine degradative uptake system we previously defined in systemic endothelial cells (Haddock, R. C., Mack, P., Leal, S., and Baenziger, N. L. (1990) J. Biol. Chem. 265, 14395-14401). Pulmonary endothelial cells display all components of this system: histamine methyltransferase generating the proximal cell-associated metabolite tele-methylhistamine and receptors binding diamine oxidase which generates the distal product methylimidazoleacetic acid that is accumulated by the cells. A diamine oxidase released from human neutrophil granules by activation with Ca2+ ionophore binds pulmonary and systemic endothelial cell and fibroblast diamine oxidase receptors and, thereby, participates in histamine degradative uptake. This enzyme utilizes cell-associated tele-methylhistamine as a substrate, preferentially generating methylimidazoleacetic acid in addition to reactive oxygen species. Thus the enzymatic and interactive cellular machinery for histamine clearance is inherently present as a functional unit in two major human pulmonary cell types. It interacts with products of inflammatory host defense cells, and pulmonary endothelial-neutrophil interactions via this pathway may influence the progression of inflammation.

Amine Oxidase (Copper-Containing)↗
PubMed1994

An environmentally regulated receptor for diamine oxidase modulates human endothelial cell/fibroblast histamine degradative uptake.

Human vascular endothelial cells and fibroblasts express a cell-surface degradative pathway for the multifunctional mediator histamine, which employs a receptor for the metabolic enzyme diamine oxidase (DAO) and results in cellular accumulation of the final metabolite methylimidazoleacetic acid. We demonstrate recognition and regulatory properties of DAO receptors as a function of cellular environmental conditions. Fast and slow ligand binding receptor populations bind DAO at 4 degrees C maximally in 1 and 7 h, respectively; upon warming cells to 37 degrees C both populations participate in degradative uptake of histamine accumulated as methylimidazoleacetic acid. Bound DAO is displaced by heparin with 24-fold greater potency than dextran sulfate, implicating structural specificity of heparin-like glycosaminoglycan moieties as a critical factor in initial receptor/enzyme interaction at fast and slow sites. Receptor-bound DAO is retained under mildly acidic conditions characteristic of early to mid endocytic intracellular compartments and thus could recycle to the plasma membrane intact after internalization. DAO initially bound to receptors in whole cells is retained through cell disruption/membrane fractionation procedures, but DAO binds poorly to isolated membrane fractions or presolubilized receptors, suggesting that the geometry of DAO binding components is not readily maintained upon cell disruption unless DAO is already bound. Cells down-regulate their complement of DAO receptors upon prolonged exposure to DAO. In cells plated at high density, half of the bound DAO becomes nondisplaceable by heparin within 15 min at 37 degrees C, a time consistent with receptor internalization, whereas cells plated at low density retain all bound DAO in a heparin-sensitive state. The protein kinase C activator phorbol 12-myristate 13-acetate modulates DAO receptor number by 35% and total histamine degradative uptake by > 2-fold. Thus this pathway is subject to regulation at the levels of DAO receptor numbers, their state of cell-surface display, and additional cellular elements of the degradative pathway with which the DAO receptors interface.

Amine Oxidase (Copper-Containing)↗
PubMed1993

Human bradykinin B2 receptors isolated by receptor-specific monoclonal antibodies are tyrosine phosphorylated.

We report the immunoaffinity isolation of bradykinin B2 receptors in a tyrosine-phosphorylated state from WI-38 human lung fibroblasts. We generated six monoclonal antibodies directed against B2 bradykinin receptor biologic activity mediating prostaglandin E2 production in WI-38. These cells express a repertoire of bradykinin receptor affinity forms with closely correlated biologic activity and [3H]bradykinin binding. Some of the monoclonal antibodies selectively recognize intermediate-affinity (Kd = 5.6 nM) or low-affinity (Kd = 42 nM) receptor forms, whereas others recognize epitopes common to both. The monoclonal antibodies block bradykinin binding and biologic activity. Immunoaffinity chromatography on an immobilized monoclonal antibody of intermediate- plus low-affinity specificity yields WI-38 B2 receptors with intact [3H]bradykinin binding activity and a molecular mass of 78 kDa. The same band is immunoblotted by all the monoclonal antibodies, indicating a similar molecular mass for receptor forms of different affinity. Anti-phosphotyrosine antibodies demonstrate that the receptors are tyrosine phosphorylated, with implications for receptor function and regulation. Genistein completely inhibits bradykinin-mediated prostaglandin E2 production with an IC50 of 8 microM, indicating that tyrosine kinase activity is critical for the signal transduction leading to arachidonic acid release.

Antibodies, Monoclonal↗
PubMed1992

Diversity of B2 bradykinin receptors with nanomolar affinity expressed in passaged IMR90 human lung fibroblasts.

IMR90 human fetal lung fibroblasts express bradykinin receptors activating the pathway for biosynthesis of PGE2. A receptor of the B2 subtype stimulates half-maximal PGE2 production at 4.8 nM bradykinin, and maximal output takes place at 25 nM bradykinin. Radioligand binding studies reveal a population of [3H]bradykinin binding sites whose affinity correlates with this B2 receptor's biologic activity, with a KD of 2.5 nM. As IMR90 cells reach 60% of their defined life span in culture, they spontaneously induce expression of a second site of lower affinity, with half-maximal binding of [3H]bradykinin at 44 nM. This second site displays a characteristic primary B2 receptor recognition profile, but differs from the 2.5 nM site on a secondary level in recognition among different B2 ligands. Bradykinin is the most potent ligand at both sites; they each preferentially recognize an N-terminal extended bradykinin peptide construct having selectivity for the rat myometrial B2 receptor, suggesting that both sites have structural features in common. However, they display diversity in their order of preference for Met-Lys-bradykinin versus Lys-Lys-bradykinin; at the 44 nM site this order is completely reversed from the order of potency exhibited at the 2.5 nM site. Expression of the second site changes the manner in which these fibroblasts control their PGE2 production; it affords a graded response of PGE2 production at bradykinin levels beyond those which would normally saturate the 2.5 nM site. The inducibility of the 44 nM site in cultured fibroblasts addresses in vivo conditions in an inflammatory environment where continuing generation of bradykinin-related peptides takes place and presents a possible mechanism for overriding constraints that would otherwise limit the progression of inflammation.

Amino Acid Sequence↗
PubMed1990

The histamine degradative uptake pathway in human vascular endothelial cells and skin fibroblasts is dependent on extracellular Na+ and Cl-.

We have previously reported that human vascular endothelial cells and skin fibroblasts carry out degradation of [3H]histamine by a mechanism involving two successive enzymatic steps: imidazole ring tele-methylation by the cells' endogenous methyltransferase and subsequent amine oxidation by an exogenous diamine oxidase. Both histamine and the exogenous second enzyme in the pathway associate with the cells via separate binding sites or receptors. The enzymatic degradation process results in cellular accumulation of the proximal and distal metabolites tele-methylhistamine and 1-methyl-4-imidazoleacetic acid (MIAA) (Haddock, R. C., Mack, P., Fogerty, F. J., and Baenziger, N. L. (1987) J. Biol. Chem. 262, 10220-10228). We have now demonstrated that this two-stage histamine degradative pathway is dependent on Na+ and Cl- in the extracellular environment. Accumulation of [3H] histamine-derived products is partially inhibited under conditions of Na+ deprivation and more substantially when Cl- is also withdrawn. The individual tele-methylation and amine oxidation enzymatic reactions themselves are unaffected or actually facilitated under these conditions. This indicates that it is the cellular mechanism for uptake coupled to the degradative pathway which reflects the cation and anion dependency. Restoration of degradative uptake displays a biphasic Na+ concentration curve, suggesting that the uptake process may be driven by multiple components. These findings indicate a role for both inward Na+ and Cl- ion movement in this cellular degradative uptake mechanism.

Biological Transport↗
PubMed1989

Heterogeneity of cellular expression of arachidonate 15-lipoxygenase: implications for biological activity.

Comparison of relative arachidonate 15-lipoxygenase activities in different types of human cell reveals striking differences in levels of product generation. Among available human cell types, the tracheal epithelial cell and the eosinophil have markedly higher levels of activity than other cell types with reported activity. Theories of functional significance for this enzymatic pathway will need to account for the selective expression of high levels of activity in a highly limited number of cell types.

Arachidonate 15-Lipoxygenase↗
PubMed1987

Role of receptors in metabolic interaction of histamine with human vascular endothelial cells and skin fibroblasts. An ordered sequence of enzyme action.

The interaction of histamine with an H1 receptor on human endothelial cells evokes production of the lipid mediator prostaglandin I2 (PGI2) and is accompanied by tachyphylaxis of this H1 receptor response (Baenziger, N. L., Fogerty, F. J., Mertz, L. F., and Chernuta, L. F. (1981) Cell 24, 915-923). We have explored the affected cells' capability for subsequent metabolic degradation of histamine molecules. Human vascular endothelial cells and skin fibroblasts exhibit a two-stage histamine degradation sequence whose participants are an enzyme native to the cells themselves and one provided from an extracellular source. Initially, the cells' endogenous histamine N-methyltransferase activity mediates conversion of cell-associated [3H]histamine to tele-methylhistamine with retention of this intermediate metabolite. Subsequently, in the presence of exogenous diamine oxidase derived from fetal calf serum or human placenta, cell-associated tele-methyl-histamine is further converted to the end product methylimidazoleacetic acid. After an initial lag phase lasting 3-6 min, the cell-associated radioactivity accumulates as methylimidazoleacetic acid at a linear rate substantially enhanced over that without diamine oxidase. The entire sequence is blocked by the histamine methyltransferase inhibitor homodimaprit. Accumulation of [3H]histamine metabolites by endothelial cells is saturable both with respect to exogenous diamine oxidase and to histamine. Thus this metabolic pathway is carried out at the level of the individual cell by means of binding sites or receptors for the substrate and for the distal degradative enzyme, diamine oxidase.

Amine Oxidase (Copper-Containing)↗
PubMed1985

Exaggerated human vascular cell prostaglandin biosynthesis mediated by monocytes: role of monokines and interleukin 1.

Incubation of cultured human umbilical vein endothelial cells with factors derived from human peripheral blood mononuclear cells (MNCF) or adherent monocytes (AMF) resulted in concentration-and time-dependent increases in prostacyclin and prostaglandin E2 (PGE2) production. MNCF and AMF also stimulated prostacyclin and PGE2 biosynthesis in cultured human arterial smooth muscle cells and human dermal fibroblasts. The effect of these monokines on endothelial cells and fibroblasts was mimicked by treatment with purified human interleukin 1 (IL 1). Mononuclear cell-conditioned medium subjected to gel filtration yielded fractions (Mr 12,000 to 18,000 daltons) which simultaneously contained endothelial cell and fibroblast prostaglandin-stimulating activity and IL 1 activity. Therefore, monokines, specifically IL 1, appear to serve as chemical mediators of the interaction between monocytes and vascular cells as would occur in blood vessel injury, inflammation, and atherosclerosis.

6-Ketoprostaglandin F1 alpha↗
PubMed1983

A rapid miniature thin layer chromatography system for analysis of prostaglandins and lipoxygenase products.

We have developed a miniature thin layer chromatography system for rapidly identifying the major arachidonate metabolites in a radiolabeled form elaborated by cells or tissues. This system separates the total spectrum of cyclooxygenase products and several of the most commonly found lipoxygenase pathway metabolites, while retaining the fine resolution of larger-scale and more time-consuming procedures. It requires less than one hour for extraction of metabolites, chromatography, and counting.

Animals↗
PubMed1982

Regulation of prostaglandin synthesis mediated by thrombin and B2 bradykinin receptors in a fibrosarcoma cell line.

The murine fibrosarcoma cell line HSDM1C1 synthesizes prostaglandin E2 in response to thrombin and bradykinin, two products of the coagulation pathway. These physiologic effectors interact with two independent cell-surface receptor systems whose properties we have characterized. HSDM1C1 cells possess a B2 bradykinin receptor, a type more sensitive to native bradykinin than to related peptides, including Met-Lys- and desArg9-bradykinin. A period of bradykinin desensitization follows the initial response. Recovery occurs within 1 hr by a process independent of serum factors. The thrombin-mediated pathway differs in several respects. The maximum amount of prostaglandin E2 synthesized is 40% lower. Prolonged desensitization of the thrombin response occurs after an initial exposure; recovery requires at least 3 hr and depends strictly on the presence of serum. Antithrombin III and hirudin, two proteins that specifically inactivate thrombin, act in serum-free medium to relieve thrombin desensitization. Thrombin's prolonged desensitization thus suggests a persistent ligand-receptor association. The expression of receptor-mediated prostaglandin synthesis governed by multiple physiologic effectors in the same cell may reflect environmental conditions, such as the relative proportions of each effector and the presence of exogenous factors that modulate the ligand-receptor interaction.

Animals↗
PubMed1981

Regulation of histamine-mediated prostacyclin synthesis in cultured human vascular endothelial cells.

Histamine stimulates the production of prostacyclin (PGI2) in cultured human endothelial cells. We have examined the cell specificity of histamine-mediated PGI2 synthesis in primary and subcultured human cells. Venous and arterial smooth muscle cells and skin fibroblasts synthesized PGI2 from exogenous arachidonic acid, but they did not synthesize a significant amount of PGI2 when treated with histamine. Endothelial cells, however, produced similar amounts of PGI2 in response to histamine and arachidonic acid. Thrombin also stimulates PGI2 production in endothelial cells. Histamine and thrombin yielded an additive production of PGI2 when added simultaneously to endothelial cells. When histamine and thrombin were added sequentially, the amount of PGI2 produced was not additive but equaled the amount characteristic of the first agonist alone. Following an initial treatment with histamine, endothelial cells were unable to respond to histamine for 3 hr, after which the PGI2 biosynthetic response rapidly returned to normal by 4 1/2 hr. When the initial histamine treatment was carried out under mildly alkaline conditions, the complete return of activity was delayed to 8 hr after treatment. The synthesis of PGI2 from exogenous arachidonic acid was unaffected by prior treatment with histamine. Recovery of histamine-mediated PGI2 production was not dependent on protein synthesis but required a component of fetal calf serum that is nondialyzable and moderately heat stable. Thus endothelial cell PGI2 synthesis in responses to a physiologic agonist is subject to several levels of regulation, reflecting not only intracellular events but also the extracellular environment.

Arachidonic Acids↗
PubMed1980

Bradykinin-stimulated release of arachidonate from phosphatidyl inositol in mouse fibrosarcoma cells.

We recently proposed a new pathway by which arachidonate is released from platelet phosphatidyl inositol after stimulation by either thrombin or calcium ionophore A23187. The initial step in arachidonate liberation involves hydrolysis of phosphatidyl inositol to form 1,2-diacyglycerol which is subsequently hydrolyzed by a diacyglycerol lipase to liberate arachidonate for the prostaglandin and lipoxygenase pathways. Whether this pathway is unique to platelets or accounts for arachidonate release from other tissues has not been previously studied. Thus we have now investigated arachidonate metabolism in mouse fibrosarcoma cells (HSDM1C1) grown in culture. These cells contain approximately 7.6% of their total phospholipid as phosphatidyl inositol in the resting state (range 6.5-8.3%). When bradykinin (12 microM) is added to the fibrosarcoma cells, there is a rapid depletion of membrane phosphatidyl inositol reaching 62 +/- 8% S.D. of baseline values by 15 seconds, falling to 36 +/- 6% by 15 minutes. The drop in membrane phosphatidyl inositol is accompanied by release of arachidonate and PGE2 into the culture medium. The time course of phosphatidyl inositol breakdown and PGE2 formation supports the idea that phosphatidyl inositol breakdown provides he arachidonate for prostaglandin synthesis in mouse fibrosarcoma cells. Crude extracts of HSDM1C1 cells contained sufficient phosphatidyl inositol-specific phospholipase C activity and diacylglycerol lipase activity to account for arachidonate release in these cells.

Arachidonic Acids↗
PubMed1980

Isolation and quantitation of the platelet membrane glycoprotein deficient in thrombasthenia using a monoclonal hybridoma antibody.

We used the hybridoma technique to characterize further the platelet glycoprotein abnormality in Glanzmann's thrombasthenia. Spleen cells from Balb/c mice immunized with human platelets were fused to mouse myeloma cell line Sp2/0-Ag14. Hybridoma lines producing a variety of antiplatelet antibodies were isolated by hypoxanthine-aminopterin-thymidine selection and cloned, and purified monoclonal IgG from six lines was prepared. One of these lines, 8aB5-9, produced an antibody, Tab, that binds to a protein on normal but not thrombasthenic platelets. We isolated this protein from Triton X-100 solubilized normal platelet membranes by affinity chromatography on Tab-Sepharose. As determined by SDS polyacrylamide gel electrophoresis, the isolated protein is a complex of glycoproteins IIb and IIIa, because the two subunits comigrate with glycoproteins IIb and IIIa of whole platelets and show identical changes in mobility after disulfide bond reduction. We prepared (125)I-Tab to determine the number of glycoprotein IIb-IIIa complexes on normal and thrombasthenic platelets by a direct binding assay. Platelets from 17 normal donors bound 39,000+/-4,600 (SD) Tab molecules/platelet. Platelets from four patients with thrombasthenia lacked Tab binding sites (<5%). Five obligate and four presumed heterozygotes for thrombasthenia bound 24,500+/-5,800 Tab molecules/platelet. The platelet alloantigen, Pl(Al), is not that recognized by Tab, because platelets from three Pl(Al)-negative subjects bound Tab normally. Studies with the Tab antibody have (a) enabled quantitation of the number of glycoprotein IIb-IIIa complexes on normal platelet membranes, (b) demonstrated that thrombasthenic homozygotes lack and heterozygotes have a partial deficiency of this complex, and (c) made possible the isolation of this membrane protein which may be required for normal platelet aggregation and clot retraction.

Antibodies↗