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Biomedical subjects

T Stevens

Publications and source records attributed to T Stevens.

At least 37 records · Page 2Linked to original sources
PubMed1994

Human melanoma cells express functional endothelin-1 receptors.

Current evidence suggests that endothelium-derived factors enhance human melanoma vascular invasion. Therefore, we studied human melanoma cell expression of receptors to the endothelium-derived peptide, endothelin-1 (ET-1), and determined if they respond to ET-1 with proliferation and chemokinesis. Human metastatic melanoma cell lines were found to have specific, saturable, high affinity ET-1 binding. Northern analysis and competitive inhibition studies confirmed that melanoma cells express the ETB receptor isoform. Ten nanomolar ET-1 caused an 8.2 to 25.5-fold increase in intracellular free calcium. ET-1 was found to be a weak mitogen for melanoma cells, however, melanoma cell chemokinesis was significantly increased by ET-1. These data suggest that ET-1 may be involved in providing a chemokinetic and growth factor environment that enhances perivascular proliferation and invasiveness of melanoma cells.

Calcium↗
PubMed1994

Rapid modification of ribosomal S6 kinase II (S6KII) in rabbit peritoneal neutrophils stimulated with chemotactic factor fMet-Leu-Phe.

The ribosomal S6 kinase II (S6KII) in rabbit neutrophils was studied by immunoblotting with antibodies prepared against recombinant S6KII. A protein with apparent molecular weight of 80,000 Da in SDS-gel was recognized by the antibodies. A shift of the apparent molecular weight to 84,000 Da in SDS-gel was observed in cells stimulated with the chemotactic factor fMet-Leu-Phe. Cytochalasin B and phorbol 12-myristate 13-acetate, but not A23187, stimulated both the tyrosine phosphorylation of p41mapk and the change of the mobility of S6KII. Pretreatment of the cells with quin 2/AM inhibited almost completely the tyrosine phosphorylation of p41mapk induced by fMet-Leu-Phe, but only partially the change in mobility of S6KII. Under various conditions, near maximum conversion of S6KII was observed even if only about 40% of the maximum level of tyrosine phosphorylation of p41mapk was achieved. The results suggest that rapid modification of S6KII occurs in chemotactic factor-stimulated neutrophils. Furthermore, the modification of S6KII induced by fMet-Leu-Phe requires either only partial tyrosine phosphorylation of p41mapk or the activation of kinase(s) other than the p41mapk isoform.

Animals↗
PubMed1994

Reduced PO2 alters the behavior of Fura-2 and Indo-1 in bovine pulmonary artery endothelial cells.

Calcium-sensitive fluorophores are used to estimate cytosolic free Ca2+ in many cell types under various conditions. We tested the effect of reduced PO2 on the behavior of Fura-2 and Indo-1 in cultured bovine pulmonary artery endothelial cells. Reduced PO2 (PO2 25-35 mmHg) caused a significant upward shift of in vivo calibration curves for both fluorophores. The in vivo emission spectrum of Fura-2 indicated that the effect was principally due to attenuated emission at the Ca(2+)-unbound 380 nm wavelength, with no shift in position of the emission maxima for either Ca(2+)-bound or unbound forms of the fluorophore. Reduced PO2 did not directly alter the behavior of the dyes, as no shift of in vitro calibration curves was seen. Neither decreased photobleaching nor altered autofluorescence accounted for the shift. We investigated several potential indirect effects, including cellular acidification, reduced viscosity, inhibition of oxidative energy production and reductive stress. In contrast to lowered PO2, acidification in vitro produced a leftward but not an upward shift. Estimation of intracellular pH with SNAFL-calcein under reduced PO2 showed no apparent acidification in these cells, further strengthening the argument that altered intracellular pH was not causing the shift. Others have shown that decreases in viscosity in vitro may shift the calibration curve for Fura-2 upward, similar to our finding with reduced PO2. However, for Indo-1 we found that decreased viscosity in vitro attenuated fluorescence emission at the Ca(2+)-bound 405 nm wavelength, thus producing the opposite effect on fluorescence ratio and indicating that reduced PO2 was not acting through changes in cellular microviscosity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Acute reductions in PO2 depolarize pulmonary artery endothelial cells and decrease [Ca2+]i.

Whereas pulmonary artery endothelial cells (PAECs) are sensitive to oxygen, neither the effect of an acute reduction in PO2 on PAEC membrane potential nor its effect on intracellular free Ca2+ ([Ca2+]i) is known. We hypothesized that in confluent primary cultures of PAECs, an acute decrease in PO2 would depolarize the cell membrane, inhibit Ca2+ influx, and reduce [Ca2+]i. To test this hypothesis, the membrane-sensitive fluorophore bis (1,3-dibutylbarbituric acid) trimethine oxonol (DiBAC4, 1 microM) and [Ca2+]i-sensitive probe fura 2 (3 microM) were used. A decrease in PO2 from 125 to 35 mmHg caused membrane depolarization and a 60 +/- 8% (data are means +/- SE) reduction in Ca2+ influx, estimated by manganese quenching of fura 2 fluorescence. While basal [Ca2+]i was 79 +/- 5 nM in normoxic cells, it decreased to 31 +/- 2 nM after 15 min of hypoxia. Decreasing the electrochemical gradient for Ca2+ entry with either low extracellular Ca2+, the K+ channel blockers tetraethylammonium or charybdotoxin, or blockade of Ca2+ entry with lanthanum decreased [Ca2+]i by 54-71% of that observed during an acute reduction in PO2. These results demonstrate that an acute reduction in PO2 1) depolarizes PAECs, 2) reduces Ca2+ influx, and 3) decreases [Ca2+]i, and that a similar reduction in [Ca2+]i was observed with interventions designed to reduce the electrochemical driving force for Ca2+ entry.

Animals↗
PubMed1994

Acute hypoxia causes membrane depolarization and calcium influx in fetal pulmonary artery smooth muscle cells.

Changes in oxygen tension in the perinatal period contribute to high pulmonary vascular tone in the fetus and the decline in resistance that occurs at birth. Distal pulmonary artery smooth muscle cells (PASMC) isolated from late-gestation ovine fetuses respond to acute hypoxia with an increase in cytosolic calcium concentration ([Ca2+]i) dependent on Ca2+ entry. The purpose of this study is to determine 1) whether acute hypoxia results in PASMC membrane depolarization, 2) whether Ca2+ entry was through voltage-operated calcium channels (VOCC), 3) the contribution of Ca(2+)-induced Ca2+ release (CICR) to the hypoxic response, and 4) whether a subset of K+ channels might serve as oxygen sensors in fetal PASMC. We used microfluorimetry on subconfluent monolayers of PASMC in primary culture loaded with either a membrane potential-sensitive dye, bis(1,3-dibutylbarbituric acid) trimethine oxonol (DiBAC4; DPASMC), to estimate membrane potential, or the Ca(2+)-sensitive fluorophore, fura 2, to measure [Ca2+]i. Hypoxia increased fluorescence from PASMC loaded with DiBAC4, consistent with membrane depolarization. Verapamil (an inhibitor of VOCC) attenuated, and BAY K 8644 (a VOCC facilitator) potentiated, the hypoxia-induced increase in [Ca2+]i, respectively. The hypoxic response was transient after treatment with ryanodine (10(-7) M), a blocker of calcium release from intracellular stores. Charybdotoxin (10(-7) M), an inhibitor of Ca(2+)-activated K+ channels, almost doubled [Ca2+]i, whereas glibenclamide (10(-5) M), an ATP-sensitive K(+)-channel antagonist, had no effect.(ABSTRACT TRUNCATED AT 250 WORDS)

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1993

Acute hypoxia increases cytosolic calcium in fetal pulmonary artery smooth muscle cells.

We studied the effect of acute hypoxia on the cytosolic calcium concentration ([Ca2+]i) of fetal vascular smooth muscle cells (SMC) from late gestation fetal lambs. We tested the following hypotheses: 1) fetal pulmonary artery (PA) SMC sense hypoxia; 2) hypoxia stimulates transmembrane Ca2+ influx causing increased basal [Ca2+]i and Ca2+ responses to pharmacological vasoconstrictors; and 3) the response is unique to SMC from small (near resistance) PA. Fetal SMC were isolated from the proximal and distal pulmonary (DPA) and carotid arteries of late-gestation ovine fetuses, maintained in culture for 5-14 days prior to study, and studied with dual-excitation microfluorimetry using fura 2. Acute hypoxia caused a 233% increase in [Ca2+]i in distal PA SMC (P < 0.01), which was absent in low extracellular calcium bath. [Ca2+]i increased transiently in normoxic DPA SMC treated with angiotensin II, and oscillations in [Ca2+]i occurred (amplitude > or = 30 nM). In hypoxic DPA SMC the increase in [Ca2+]i was sustained and oscillations were attenuated or absent. [Ca2+]i in proximal PA SMC did not change with exposure to acute hypoxia and carotid artery SMC [Ca2+]i decreased by 13% (P < 0.05). We conclude that fetal SMC isolated from the DPA of late-gestation ovine fetuses directly sense decreased oxygen tension with an increase in [Ca2+]i that is dependent on the entry of extracellular Ca2+.

Acute Disease↗
PubMed1993

Pulmonary and systemic vascular responsiveness to TNF-alpha in conscious rats.

Endotoxin decreases pulmonary vascular reactivity. Because tumor necrosis factor-alpha (TNF-alpha) is a primary mediator of endotoxemia, we tested whether TNF-alpha altered pulmonary vascular reactivity in conscious adult female rats. Osmotic pumps were implanted intraperitoneally, and low-dose TNF-alpha (62 micrograms, TNF62; n = 7), high-dose TNF-alpha (> or = 250 micrograms, TNF250; n = 5), or saline (n = 5) was administered for 2 wk. Pulmonary pressor responses to 14% O2 and angiotensin II (ANG II, 0.0206 micrograms/min for 10 min) were measured without (day 13) or after (day 14) administration of nitro-L-arginine (4.4 mg/kg iv), an inhibitor of endothelium-derived relaxing factor (EDRF). TNF-alpha administration slightly decreased (P < or = 0.08) baseline pulmonary arterial pressure in TNF250 rats and decreased (P < or = 0.05) hypoxia- and ANG II-induced constrictions in TNF62 and TNF250 rats. Whereas nitro-L-arginine potentiated (P < or = 0.05) pressure responses in control rats, it had no effect on hypoxic responses in TNF-alpha-treated rats. Nitro-L-arginine increased (P < or = 0.05) ANG II-induced vasoconstriction in TNF-alpha-treated rats, but the pulmonary arterial pressure response was still lower (P < or = 0.05) in TNF250 than in control and TNF62 rats. These results suggest that chronic TNF-alpha decreases 1) pulmonary vascular reactivity in the intact rat, 2) hypoxic pulmonary vasoconstriction by a mechanism that is independent of EDRF, and 3) ANG II-induced constriction by a mechanism that is partly EDRF dependent.

Angiotensin II↗
PubMed1993

Pulmonary function and hypoxic ventilatory response in subjects susceptible to high-altitude pulmonary edema.

To determine if spirometric changes reflect early high-altitude pulmonary edema (HAPE) formation, we measured the FVC, FEV1, and FEF25-75 serially during the short-term period following simulated altitude exposure (4,400 m) in eight male subjects, four with a history of HAPE and four control subjects who had never experienced HAPE. Three of the four HAPE-susceptible subjects developed acute mountain sickness (AMS), based on their positive Environmental Symptom Questionnaire (AMS-C) scores. Clinical signs and symptoms of mild pulmonary edema developed in two of the three subjects with AMS after 4 h of exposure, which prompted their removal from the chamber. Their spirometry showed small decreases in FVC and greater decreases in FEV1 and FEF25-75 after arrival at high altitude in the presence of rales or wheezing on clinical examination and normal chest radiographs. One of the two subjects had desaturation (59 percent) and tachycardia during mild exercise, and excessive fatigue and inability to complete the exercise protocol developed in the other at 4 h. The six other subjects had minimal changes in spirometry and did not develop signs of lung edema. Further, we measured each subject's ventilatory response to hypoxia (HVR) prior to decompression to determine whether the HVR would predict the development of altitude illness in susceptible subjects. In contrast to anticipated results, high ventilatory responses to acute hypoxia, supported by increased ventilation during exposure to high altitude, occurred in the two subjects in whom symptoms of HAPE developed. The results confirm that HAPE can occur in susceptible individuals despite the presence of a normal or high ventilatory response to hypoxia.

Adult↗
PubMed1992

Acute and long-term TNF-alpha administration increases pulmonary vascular reactivity in isolated rat lungs.

Tumor necrosis factor-alpha (TNF-alpha) causes pulmonary hypertension and arterial hypoxemia, but the mechanisms are unknown. We conducted two experiments to test the hypothesis that TNF-alpha alters pulmonary vascular reactivity, which in turn could cause either pulmonary hypertension or arterial hypoxemia. In experiment 1, rats were given acute or long-term injections of TNF-alpha (recombinant human) in vivo. Rats treated acutely received either saline or TNF-alpha (40 micrograms/kg iv in saline) 3 min (TNF-3 min; n = 8), 20 min (TNF-20 min; n = 8), or 24 h (TNF-24 h; n = 5) before the lungs were isolated. Rats treated chronically received injections of either saline or TNF-alpha (250 micrograms/kg ip in saline) two times per day for 7 days (TNF-7 days; n = 9). Lungs were isolated and perfused with Earle's salt solution (+2 g/l NaHCO3 + 4 g/100 ml Ficoll), and vascular reactivity was tested with acute hypoxia (3 min; 3% O2) and angiotensin II (ANG II; 0.025-0.40 micrograms). Pulmonary pressor responses to hypoxia were greater (P less than 0.05) in TNF-20 min and TNF-7 day groups. ANG II responses were increased (P less than 0.05) in TNF-7 day rats. In experiment 2, lungs were isolated and perfused and received direct pulmonary arterial injections of TNF-alpha (0.2, 2.0, and 20 micrograms) or saline, after stable responses to hypoxia and ANG II (0.10 microgram) were attained. Reactivity was not different between control and TNF-alpha rats before the injections, but TNF-alpha increased (P less than 0.05) responses to hypoxia and ANG II.(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensin II↗
PubMed1991

Safety and efficacy of percutaneous transluminal coronary angioplasty in patients with left ventricular dysfunction.

The risks and long-term outcome after 845 elective percutaneous transluminal coronary angioplasties (PTCA) in patients with left ventricular (LV) dysfunction (ejection fraction less than or equal to 40%) were examined. Procedural results were compared with 8,117 consecutive procedures in patients with ejection fractions greater than 40%. The patients with LV dysfunction were older (63 vs 60 years, p less than 0.01), had a greater incidence of prior myocardial infarction (84 vs 45%, p less than 0.001), prior bypass surgery (39 vs 21%, p less than 0.001), 3-vessel disease (62 vs 33%, p less than 0.001), and class IV angina (48 vs 41%, p less than 0.01) than the control group. Angiographic success was lower (93 vs 95%, p less than 0.01), and overall procedural mortality was increased ( 4 vs 1%, p less than 0.001) in the study group. Emergency surgery rates were identical (2%). No significant difference was found in rates of nonfatal Q-wave myocardial infarction (2 vs 1%). At mean follow-up of 33.5 months, 15% of the patients with LV dysfunction required late bypass surgery, 27% underwent repeat PTCA, and 59% were angina free. Actuarial survival at 1 and 4 years was 87 and 69%, respectively. Cox regression analysis identified 3-vessel disease, age greater than or equal to 70 years, class IV angina and incomplete revascularization as correlates of long-term mortality. These data suggest that PTCA may be an effective treatment for coronary artery disease in patients with LV dysfunction.

Adolescent↗
PubMed1991

Modulation of leukotriene B4 and platelet-activating factor binding to neutrophils.

Preincubation of human neutrophils with the human hormone granulocyte-macrophage colony-stimulating factor (GM-CSF) inhibits the specific binding of leukotriene B4 ([3H]LTB4) but not the nonmetabolizable bioactive platelet-activating factor ([3H]C-PAF) to intact cells. This inhibition requires that the GM-CSF interacts with intact cells. The action of GM-CSF is not prevented by pertussis toxin. Moreover, the rise in calcium produced by LTB4 but not by PAF is also inhibited in human neutrophils pretreated with GM-CSF. Interestingly, neither the inhibitory action of GM-CSF on [3H]LTB4 binding or LTB4-induced calcium rise nor the potentiation of superoxide production by GM-CSF is reduced by inhibitors of arachidonic acid metabolism by the lipoxygenase pathway. In contrast, preincubation of human neutrophils with either the chemotactic factor formyl-methionyl-leucyl-phenylalanine (fMet-Leu-Phe) or the active phorbol ester, phorbol 12-myristate 13-acetate (PMA), inhibits the binding of both [3H]LTB4 and [3H]C-PAF to intact cells. The inhibitory actions of GM-CSF, PMA, and fMet-Leu-Phe require that they interact with the intact cells; their actions cannot be reproduced in plasma membrane preparations. The effects of both GM-CSF and fMet-Leu-Phe cannot be prevented by the protein kinase C inhibitor staurosporine. The mechanisms of fMet-Leu-Phe and GM-CSF actions are probably not mediated through the release of LTB4 by the cells. Interestingly, this new action, unlike other reported effects of GM-CSF, is not mediated through a pertussis toxin-sensitive G protein (Gi alpha 2). This indicates that not all GM-CSF receptors are coupled to Gi alpha 2.

Alkaloids↗
PubMed1986

The performance of medical malpractice review panels.

In this paper we present the results of an empirical study of Arizona's medical malpractice review panels. We compare insurance company claim files data from before and after the implementation of panels to analyze the effects of the panel system. The data indicate that although the frequency and average amount of recovery are not affected by the panel system, the system leads to an increase in the number of disputes seeking formal adjudication, an increase in the cost of the process, and a lengthening of the time within which disputes are resolved.

Arizona↗
PubMed1982

Early stages in the yeast secretory pathway are required for transport of carboxypeptidase Y to the vacuole.

Temperature-sensitive secretory mutants (sec) of S. cerevisiae have been used to evaluate the organelles and cellular functions involved in transport of the vacuolar glycoprotein, carboxypeptidase Y (CPY). Others have shown that CPY (61 kd) is synthesized as an inactive proenzyme (69 kd) that is matured by cleavage of an 8 kd amino-terminal propeptide. sec mutants that are blocked in either of two early stages in the secretory process and accumulate endoplasmic reticulum or Golgi bodies also accumulate precursor forms of CPY when cells are incubated at the nonpermissive temperature (37 degrees C). These forms are converted to a proper size when cells are returned to a permissive temperature (25 degrees C). Vacuoles isolated from sec mutant cells do not contain the proCPY produced at 37 degrees C. These results suggest that vacuolar and secretory glycoproteins require the same cellular functions for transport from the endoplasmic reticulum and from the Golgi body. The Golgi body represents a branch point in the pathway: from this organelle, vacuolar proenzymes are transported to the vacuole for proteolytic processing and secretory proteins are packaged into vesicles.

Biological Transport↗