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C J Meininger

Publications and source records attributed to C J Meininger.

34 records · Page 2Linked to original sources
PubMed1995

Impaired arginine metabolism and NO synthesis in coronary endothelial cells of the spontaneously diabetic BB rat.

Arginine metabolism via nitric oxide (NO) synthase and other pathways was studied in coronary endothelial cells (EC) from the spontaneously diabetic BB rat, an animal model of human type I diabetes mellitus (IDDM). EC were prepared from insulin-treated diabetic BB (BBd) and non-diabetes-prone BB (BBn) rats. Basal NO synthesis was studied in EC cultured for 48 h in medium containing 0.4 mM L-arginine. At the end of the culture period, the medium was analyzed for nitrite and nitrate (two major end stable oxidation products of NO), and the cells were used to determine arginine uptake and metabolism and the activities of some arginine-degrading enzymes. For studies of arginine metabolism, cells were incubated at 37 degrees C for 1 h in Krebs-Henseleit bicarbonate buffer (pH 7.4) containing 1 mM L(-)[1-14C]arginine or L(-)[1-14C]ornithine. The rates of production of nitrite plus nitrate by BBd EC were only 15% of those of BBn cells. This impaired NO synthesis in BBd EC was not due to alterations in arginine uptake, NO synthase activity, or intracellular arginine concentrations but might have resulted from a limited intracellular availability of cofactors of NO synthase. In addition to the arginine-NO pathway, arginine was found to be metabolized to urea, ornithine, and, to a much lesser extent, CO2 via arginase and ornithine aminotransferase. The activities of arginase and the formation of ornithine and urea from arginine were decreased by 90% in BBd compared with BBn cells. These results, coupled with the reduced NO synthesis, indicate metabolic defects in arginine metabolism in BBd EC.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Increased stem cell factor release by hemangioma-derived endothelial cells.

BACKGROUND: Capillary hemangiomas, the most common tumors in young children, consist of proliferating capillary vessels and endothelial cells. These tumors also contain large numbers of mast cells, compared with the normal surrounding skin or tissue. We have recently shown that stem cell factor (SCF), the gene product of the murine steel locus, can act as a chemoattractant for mast cells. In this study, we investigated whether SCF might be involved in the recruitment and maintenance of mast cells in hemangiomas. EXPERIMENTAL DESIGN: Cultured endothelial cells derived from a murine hemangioma were compared with normal vascular endothelial cells for the ability to produce and release SCF, a mitogen for mast cells. RESULTS: Conditioned medium from hemangioma-derived endothelial cells stimulated the proliferation of cultured mast cells. This proliferative activity was potentiated by interleukin-3. The same conditioned medium was unable to stimulate proliferation of mast cells expressing a defective receptor for SCF. The medium was also unable to stimulate proliferation when it was preincubated with neutralizing antibodies specific for SCF. Immunoprecipitation and Western blot analysis of the conditioned media from hemangioma cells and normal endothelial cells demonstrated the 31,000 molecular weight SCF in hemangioma-conditioned medium only. In addition, proliferative activity for mast cells could not be demonstrated in the conditioned medium of the normal endothelial cells, although Northern blot analysis indicated that both normal and hemangioma-derived endothelial cells express SCF mRNA. Reverse transcriptase-polymerase chain reaction techniques were used to amplify the DNA sequence coding for the proteolytic cleavage site used for release of SCF. Results indicated that both normal and hemangioma-derived endothelial cells express the same transcript for SCF. CONCLUSIONS: Our data suggest that increased release of SCF is a property of hemangioma-derived endothelial cells that may account for the high numbers of mast cells observed in hemangioma tissue. This increased release of SCF is not due to alternate splicing of SCF transcripts by hemangioma cells.

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PubMed1994

Insulin stimulates glycolysis and pentose cycle activity in bovine microvascular endothelial cells.

Glucose metabolism via the pentose cycle, glycolysis and the Krebs cycle was quantified in bovine microvascular endothelial cells. The major measured end-product of glucose was L-lactate, with relatively small amounts of glucose carbons converted to CO2 and pyruvate. The pentose cycle accounted for less than 4% of the glucose utilized. About 60-70% of the metabolized glucose carbons could not be accounted for by lactate, pyruvate and CO2. Insulin stimulated glycolysis and pentose cycle activity, but had no effect on glucose oxidation via the Krebs cycle. As the pentose cycle is a major source of NADPH which is required for the synthesis of nitric oxide (the endothelium relaxing factor), insulin may play a role in regulating NO generation in endothelial cells by modulating the pentose cycle activity.

Animals↗
PubMed1993

Regulation of L-arginine synthesis from L-citrulline by L-glutamine in endothelial cells.

L-Arginine synthesis from L-citrulline was studied in cultured bovine venular, bovine aortic, human microvascular, and cloned human microvascular endothelial cells (EC). L-Citrulline was actively recycled into L-arginine in all four EC lines, with similar rates between venular and aortic EC. L-Arginine synthesis from L-citrulline was very sensitive to extracellular L-citrulline concentrations in the range normally found in plasma (50-100 microM). L-Glutamine (0.5mM) decreased L-arginine synthesis from L-citrulline, whereas 0.5 mM L-arginine, L-alanine, L-glutamate, or L-lysine had no effect. In contrast to the findings in intact cells, 1 mM L-glutamine had no effect on L-arginine synthesis from L-citrulline in EC lysates. Similarly, L-glutamine (1 mM) had no effect on the conversion of argininosuccinate to arginine in EC lysates. L-Glutamine (0.5 and 1 mM), but not 0.5 mM L-arginine, L-alanine, L-glutamate, or L-lysine, inhibited L-citrulline transport by EC. The inhibition of L-citrulline transport by L-glutamine was dose dependent and competitive in nature. These results suggest that L-glutamine decreased L-arginine synthesis from extracellular L-citrulline by interfering with its transport. Inasmuch as nitric oxide (NO) and L-citrulline are constantly generated from L-arginine, with L-citrulline being actively converted into L-arginine in venular, microvascular, and aortic EC, our data indicate a functioning intracellular arginine-citrulline cycle in these cells. This cycle may function to efficiently scavenge the carbon and alpha-amino group of L-arginine and to maintain a sufficient cellular concentration of L-arginine during prolonged synthesis of NO in EC. (ABSTRACT TRUNCATED AT 250 WORDS)

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PubMed1992

The c-kit receptor ligand functions as a mast cell chemoattractant.

Mast cells accumulate at sites of neovascularization, solid tumors, and many immune reactions. Such accumulation requires directed migration of mature mast cells or their precursors. The nature of the chemoattractants that regulate mast cell motility and the identity of the receptors that mediate the chemotactic response are poorly understood. We have tested the ability of stem cell factor (SCF), a mast cell growth factor, to stimulate mast cell migration. Our results show that SCF is a potent mast cell attractant that stimulates directional motility of both mucosal and connective tissue-type mast cells. The activity is potentiated by costimulation with interleukin-3 (IL-3), another mast cell chemoattractant. SCF, a known ligand for the c-kit tyrosine kinase receptor, was unable to stimulate motility in W42 mutant mast cells, which have a defective c-kit tyrosine kinase. However, W42 mast cells were still able to migrate in response to IL-3. These results show that SCF is a chemotactic factor as well as a growth factor and that the c-kit receptor can transduce signals leading to both cell proliferation and increased directional cell motility.

Animals↗
PubMed1992

Mast cells and angiogenesis.

Much data exists in the literature to suggest a correlation between mast cell accumulation and angiogenesis. This correlation exists for normal blood vessel growth as well as pathological vessel growth. The recruitment of mast cells to sites of angiogenesis is not completely understood. However, once at the site, mast cell products may act directly on endothelial cells to stimulate their migration and/or proliferation or may act indirectly by degrading connective tissue matrix to provide space for neovascular sprouts to form. Understanding the role of mast cells in angiogenesis may provide avenues for intervening in and manipulating the neovascularization process.

Animals↗
PubMed1992

[The effect of vasoactive factors on the growth of of coronary endothelial cells].

The aim of this study was to evaluate the effect of adenosine and bradykinin on endothelial cell growth at capillary level. DNA synthesis was measured in subconfluent capillary endothelial cells obtained from coronary venules (CVEC) after exposure to adenosine or bradykinin. Our results indicate that adenosine and bradykinin induced a dose-dependent proliferation of CVEC. Maximal effect was observed at the dose 100 microM concentration for adenosine and 0.1 microM concentration for bradykinin.

Adenosine↗
PubMed1990

An in vitro model of ischemia/reperfusion-induced microvascular injury.

The major objective of this study was to develop an in vitro model of ischemia/reperfusion (I/R)-induced microvascular injury. Cultured venular endothelial cells were grown to confluency, labeled with 51Cr, and exposed to different durations of anoxia (0.5, 1, 2, 3, and 4 h). 51Cr release and cell detachment (indexes of cell injury) were determined at different times after reoxygenation (1, 2, 4, 6, 8, and 18 h). Because in vivo studies have implicated neutrophils in I/R injury, in some experiments human neutrophils were added to the endothelial cells upon reoxygenation. Periods of anoxia greater than or equal to 2 h resulted in 70-80% 51Cr release and 80-95% cell detachment upon reoxygenation. Under these conditions (near maximal injury), the addition of neutrophils produced negligible effects. Periods of anoxia less than or equal to 1 h resulted in 30-40% 51Cr release and 50-60% cell detachment. Under these conditions (moderate cell injury), addition of neutrophils enhanced endothelial cell injury. Using a 30-min period of anoxia, we also assessed the effects of superoxide dismutase (SOD; 300 U/ml) and allopurinol (20 microM) on anoxia/reoxygenation (A/R)-induced injury in the presence or absence of neutrophils. In the absence of neutrophils, SOD or allopurinol did not protect against A/R-induced injury. However, in the presence of neutrophils, both SOD and allopurinol attenuated the increases in 51Cr release. The results derived using this in vitro model of I/R injury are largely consistent with published in vivo studies. Thus this in vitro model may provide further insights regarding the mechanisms involved in I/R injury.

Allopurinol↗
PubMed1990

Mechanisms leading to adenosine-stimulated proliferation of microvascular endothelial cells.

This study investigated the mechanisms by which adenosine stimulates proliferation of microvascular endothelial cells. The metabolic byproducts of adenosine, inosine and hypoxanthine were unable to stimulate proliferation. When adenosine uptake was prevented, the stimulation of proliferation was unchanged, suggesting that uptake of adenosine with subsequent incorporation into the nucleotide pool is not the mechanism for increasing proliferation. Treatment of endothelial cells with adenosine analogues, presumably selective for either the A1 or A2 receptor, stimulated proliferation equally. This suggested that adenosine 3', 5'-cyclic monophosphate (cAMP) might not mediate the proliferative response to adenosine. However, radioimmunoassay of cell extracts after treatment with either analogue showed an increase in cAMP. In addition, adenylate cyclase blockade with 2', 5'-dideoxyadenosine prevented the proliferative response brought about by these analogues. These data suggest that the proliferative response to adenosine depends on an increase in cAMP. A 2-h pulse of cholera toxin stimulated endothelial cell proliferation, further supporting a role for cAMP. Pretreatment of endothelial cells with pertussis toxin blocked the stimulation of proliferation, indicating that a Gi or similar G protein is also involved in proliferation. We conclude that the proliferative response to adenosine involves a pertussis toxin-sensitive substrate as well as an increase in cAMP.

Adenosine↗
PubMed1990

Anoxia-reoxygenation-induced, neutrophil-mediated endothelial cell injury: role of elastase.

The aim of this study was to assess the role of neutrophilic elastase in anoxia-reoxygenation-induced, neutrophil-mediated injury to microvascular endothelium. Cultured bovine microvascular endothelial cells were grown to confluence and labeled with 51Cr. The endothelial cells were exposed to a 30-min period of anoxia and subsequently reoxygenated. Endothelial cell injury, quantitated as 51Cr release and cell detachment, was determined 8 h after reoxygenation. Addition of neutrophils upon reoxygenation enhanced the anoxia-reoxygenation-induced increase in 51Cr release and cell detachment. The neutrophil-mediated injury was associated with elastase release from the neutrophils. Four agents were used to inhibit neutrophilic elastase activity: Eglin C, methoxysuccunyl-Ala2-Pro-Val-CH2Cl, L658,758, and a monoclonal antibody against neutrophilic elastase. All elastase inhibitors attenuated the neutrophil-mediated endothelial cell detachment but not 51Cr release. Addition of purified human neutrophilic elastase, at a level that mimicked the release from neutrophils, increased cell detachment in endothelial cells exposed to anoxia-reoxygenation but did not affect 51Cr release. Our results indicate that elastase plays an important role in anoxia-reoxygenation-induced, neutrophil-mediated endothelial cell dysfunction.

Animals↗
PubMed1988

Venular endothelial cells from bovine heart.

Coronary venular endothelial cells were isolated by a bead-perfusion technique that allowed the selection of endothelial cells from venules of a specific size. Culture conditions for the microvascular cells were established. Cells grew well in supplemented Dulbecco's modified Eagle's medium. The effect of various substrata on the proliferation of the venular endothelial cells was determined. Matrigel, gelatin, and fibronectin supported high levels of proliferation. Cell shape was correlated with ability of the substratum to support cell proliferation. Cells exhibiting a broad, flattened morphology achieved high levels of proliferation. The formation of vessel meshworks by the coronary venular endothelial cells provides an in vitro model for the study of coronary angiogenesis. Confluent monolayers of these cells can be utilized to examine mechanisms of water and protein transport across coronary venules.

Animals↗
PubMed1988

Adenosine and hypoxia stimulate proliferation and migration of endothelial cells.

The proliferation of bovine aortic or coronary venular endothelial cells (EC) in vitro was stimulated by the addition of adenosine (0.5 or 5.0 microM) to the culture medium. Cell counts of adenosine-treated aortic EC were 23-76% and coronary venular EC 19-52% greater than nontreated controls. Because adenosine is known to be released by hypoxic tissues, cell proliferation was quantitated when aortic EC were grown at 2% O2. Cell counts were 41-102% greater under hypoxic conditions than when cells were grown at standard tissue culture conditions (approximately 20% O2). When culture medium conditioned by coronary EC grown at 2% O2 was added to EC growing at standard conditions, cell counts were 24-69% greater than controls with medium conditioned by coronary EC grown at 20% O2. This suggests that hypoxia causes endothelial cells to release a factor(s) into the medium that can stimulate cell proliferation. The addition of the adenosine receptor blocker 8-phenyltheophylline (10(-5) M) prevented the stimulation of proliferation caused by hypoxia-conditioned medium, 2% O2 or 5.0 microM adenosine, suggesting that adenosine mediates its effect via an external membrane receptor. Adenosine also stimulated EC chemotaxis. Taken together, these results suggest that adenosine, released as a result of tissue hypoxia, may act as an angiogenic stimulus for the growth of new blood vessels.

Adenosine↗
PubMedDate not supplied

Physiology and pathobiology of the microcirculation.

The delivery of nutrients to the tissues and the removal of waste products from the tissues is made possible by forcing a stream of blood through an arborizing network of microscopic blood vessels that comprise the microcirculation. The rapidity of the flow stream and, therefore, the rate of nutrient delivery to the tissue, is regulated by the automatic adjustment of the caliber of the precapillary arterioles that serve as the primary loci of vascular resistance. Exchange between the blood stream and the parenchymal cells occurs in capillaries and pericytic venules. Pathologic processes such as inflammation, diabetes, ischemia, and hypertension are characterized by abnormalities in microvascular structure and function.

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