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Biomedical subjects

R M Santos

Publications and source records attributed to R M Santos.

At least 19 recordsLinked to original sources
PubMed2001

Isoform-specific inhibition of voltage-sensitive Ca(2+) channels by protein kinase C in adrenal chromaffin cells.

Selective protein kinase C (PKC) activators and inhibitors were used to investigate the involvement of specific PKC isoforms in the modulation of voltage-sensitive Ca(2+) channels (VSCCs) in bovine adrenal chromaffin cells. Exposure to the phorbol ester phorbol-12,13-dibutyrate (PDBu) inhibited the Ca(2+) currents elicited by depolarizing voltage steps. This inhibition was occluded by the PKC-specific inhibitor Ro 31-8220 but remained unaffected by Gö 6976, a selective inhibitor of conventional PKC isoforms. PDBu treatment caused the translocation of PKC-alpha and -epsilon isoforms from cytosol to membranes. PKC-iota and -zeta showed no signs of translocation. It is concluded that VSCCs are specifically inhibited by the activation of PKC-epsilon in chromaffin cells. This may be relevant to the action of phospholipase-linked receptors involved in the control of Ca(2+) influx, both in catecholaminergic cells and other cell types.

Adrenal Glands↗
PubMed2000

Glucose-mediated Ca(2+) signalling in single clonal insulin-secreting cells: evidence for a mixed model of cellular activation.

Using clonal insulin-secreting BRIN-BD11 cells, we have assessed whether the graded response of the whole cell population to glucose can be accounted for by a dose-dependent recruitment of individual cells, an amplification of the response of the recruited cells or both. Cytosolic free Ca(2+) concentration ([Ca(2+)](i)) is an established index of beta-cell function. We used fura-2 microfluorescence techniques to assess the [Ca(2+)](i) responsiveness of single BRIN-BD11 cells to glucose and other secretagogues. Glucose (1-16.7 mM) evoked oscillatory [Ca(2+)](i) rises in these cells resembling those found in parental rat pancreatic beta-cells. The percentage of glucose-responsive cells was 11% at 1 mM and increased to 40-70% at 3-16.7 mM glucose, as assessed by a single-stimulation protocol. This profile was unrelated to possible differences in the cell cycle, as inferred from experiments where the cultured cells were synchronized by a double thymidine block protocol. Individual cells exhibited variable sensitivities to glucose (threshold range: 1-5 mM) and a variable dose-dependent amplification of the [Ca(2+)](i) responses (EC(50) range: 2-10 mM), as assessed by a multiple-stimulation protocol. Glyceraldehyde and alpha-ketoisocaproic acid had glucose-like effects on [Ca(2+)](i). The data support a mixed model for the activation of insulin-secreting cells. Specifically, the graded secretory response of the whole cell population is likely to reflect both a recruitment of individual cells with different sensitivities to glucose and a dose-dependent amplification of the response of the recruited cells.

Animals↗
PubMed2000

Objective assessment of the contribution of each diagnostic test and of the ordering sequence in jaundice caused by pancreatobiliary carcinoma.

BACKGROUND: Computer-assisted diagnostic systems are not substantially more accurate than the clinician in the differential diagnosis of jaundice but may help in optimal selection and sequencing of tests. The present study aimed to assess with an electronic diagnostic tool the pattern of ordering tests and the diagnostic contribution and related financial cost of each test in jaundiced patients with pancreatobiliary carcinoma, in an effort to make the clinician's diagnostic behaviour more efficient and economical. METHODS: Clinical and diagnostic test data were prospectively gathered from 356 jaundiced patients with pancreatobiliary carcinoma and entered in a Bayesian diagnostic programme. The test results were added to the existing diagnostic evidence, and the programme calculated the diagnostic contribution of each test. RESULTS: A total of 1804 diagnostic tests were ordered. Quantitative assessment of the diagnostic contribution of each test showed that percutaneous transhepatic cholangiography and computed tomography were associated with the highest provision of information. The most cost-effective tests were ultrasonography and liver function tests. CONCLUSIONS: It is possible to determine objectively the diagnostic contribution of each test in establishing the diagnosis of pancreatobiliary carcinoma. The observed physician behaviour in ordering the various diagnostic tests might be improved with regard to its efficacy and its cost-effectiveness profile.

Bayes Theorem↗
PubMed2000

Differential patterns of glucose-induced electrical activity and intracellular calcium responses in single mouse and rat pancreatic islets.

Although isolated rat islets are widely used to study in vitro insulin secretion and the underlying metabolic and ionic processes, knowledge on the properties of glucose-induced electrical activity (GIEA), a key step in glucose-response coupling, has been gathered almost exclusively from microdissected mouse islets. Using a modified intracellular recording technique, we have now compared the patterns of GIEA in collagenase-isolated rat and mouse islets. Resting membrane potentials of rat and mouse beta-cells were approximately -50 and -60 mV, respectively. Both rat and mouse beta-cells displayed prompt membrane depolarizations in response to glucose. However, whereas the latter exhibited a bursting pattern consisting of alternating hyperpolarized and depolarized active phases, rat beta-cells fired action potentials from a nonoscillating membrane potential at all glucose concentrations (8.4-22.0 mmol/l). This was mirrored by changes in the intracellular Ca2+ concentration ([Ca2+]i), which was oscillatory in mouse and nonoscillatory in rat islets. Stimulated rat beta-cells were strongly hyperpolarized by diazoxide, an activator of ATP-dependent K+ channels. Glucose evoked dose-dependent depolarizations and [Ca2+]i increases in both rat (EC50 5.9-6.9 mmol/l) and mouse islets (EC50 8.3-9.5 mmol/l), although it did not affect the burst plateau potential in the latter case. We conclude that there are important differences between beta-cells from both species with respect to early steps in the stimulus-secretion coupling cascade based on the following findings: 1) mouse beta-cells have a larger resting K+ conductance in 2 mmol/l glucose, 2) rat beta-cells lack the compensatory mechanism responsible for generating membrane potential oscillations and holding the depolarized plateau potential in mouse beta-cells, and 3) the electrical and [Ca2+]i dose-response curves in rat beta-cells are shifted toward lower glucose concentrations. Exploring the molecular basis of these differences may clarify several a priori assumptions on the electrophysiological properties of rat beta-cells, which could foster the development of new working models of pancreatic beta-cell function.

Animals↗
PubMed2000

Role of autonomic nervous system and hemodynamics in cardiovascular homeostasis after orthostatic stress.

Autonomic control of hemodynamics is crucial to maintaining cardiovascular homeostasis after orthostasis. Our goal was to assess the effect of passive orthostatic stress on cardiovascular hemodynamics regulation by analysis of the autonomic nervous system (ANS) and gender influence on the variables. We used a passive 70 degrees tilt test as an orthostatic maneuver. For SNA measurements we used FFT for HRV and SBPV, temporal sequences and alpha index to calculate baroreceptor gain. Hemodynamic data was calculated non-invasively by modelflow. Orthostatic stress induces a rise in heart rate (+15%), total peripheral resistance (+28%), sympathetic tonus (LF_nu: +32%; LF_pas: +64%) and a reduction in cardiac output (-28%), systolic volume (-43%), and vagal activity both tonic (HF_nu:-115%) and reflex (BRG:-103%). Females had higher vagal activity regardless of position. We concluded that the autonomic nervous system plays a crucial role in cardiovascular hemodynamics control to maintain homeostasis after orthostatic stress.

Adult↗
PubMed1999

Modulation of glucose-induced insulin secretion by cytosolic redox state in clonal beta-cells.

Nutrient stimulation of pancreatic beta-cells increases the cellular reduced pyridine nucleotide content, but the specific role of cytosolic redox state in glucose-induced insulin release (GIIR) remains undetermined. The role of cytosolic redox state has been assessed (as reflected by the lactate/pyruvate ratio) in nutrient- and non-nutrient-induced insulin release using a recently established glucose-sensitive clonal beta-cell line (BRIN-BD11). Long-term exposure to the NAD+ precursor vitamin nicotinic acid (NA, 100 microM) was used to promote a more oxidized state in the cytosol. Glucose (2-16 mM) evoked a dose-dependent rise in the cytosolic NADH/NAD+ ratio which was linearly related to the extent of GIIR. NA suppressed the glucose-induced rise in the NADH/NAD+ ratio and concomitantly reduced GIIR by 44%. It also inhibited, by 47%, the average glucose-induced rise in cytosolic free Ca2+ concentration ([Ca2+]i, assessed by fura-2 microfluorometry from single cells). The latter effect was not accounted for by a reduction in the activity of voltage-sensitive Ca2+ channels, inasmuch as both high K+- and tolbutamide-induced [Ca2+]i rises remained insensitive to NA exposure. NA did not affect insulin release evoked by any of the depolarizing agents, indicating that steps in the stimulus-secretion coupling cascade distal to Ca2+ influx are insensitive to changes in the cytosolic redox state. It is concluded that GIIR is partially controlled by the cytosolic redox state. Moreover, the impairment in GIIR, caused by a shift toward a more oxidized state in the cytosol, originates from an attenuated [Ca2+]i response. The latter is likely mediated by the influence of cytosolic redox state on specific metabolic pathways (NADH shuttle systems and/or the malonyl-CoA pathway), leading ultimately to enhancement of the activity of ATP-sensitive K+ channels.

Animals↗
PubMed1999

Regulation of Ca2+ influx by a protein kinase C activator in chromaffin cells: differential role of P/Q- and L-type Ca2+ channels.

Phorbol esters reduce depolarization-evoked Ca2+ influx in adrenal chromaffin cells, suggesting that voltage-sensitive Ca2+ channels (VSCCs) are inhibited by protein kinase C-mediated phosphorylation. We now address the possibility that L- and P/Q-type Ca2+ channel subtypes might be differentially involved in phorbol ester action. In bovine chromaffin cells, short-term (10 min) incubations with phorbol 12-myristate 13-acetate (PMA) inhibited early high K+-evoked rises in cytosolic free Ca2+ concentration ([Ca2+]i) and the early component of the depolarization-evoked Mn2+ quenching of fura-2 fluorescence in a dose-dependent manner (IC50: 18 and 7 nM; maximal inhibitions: 45 and 48%, respectively). The protein kinase C inhibitor staurosporine (100 nM) reverted the inhibitory action of PMA. PMA (0.1-1 microM) inhibited the early and late phases of the ionomycin (2 microM)-evoked [Ca2+]i transients by 14-23%. Omega-agatoxin IVA, a blocker of P/Q-type Ca2+ channels, inhibited high K+-evoked [Ca2+]i rises in a dose-dependent fashion (IC50 = 50 nM). In contrast, 0.1 microM omega-conotoxin GVIA, a blocker of N-type channels, was without effect. A sizeable (< 45%) component of early Ca2+ influx persisted in the combined presence of omega-agatoxin IVA (100 nM) and nitrendipine (1 microM). Simultaneous exposure to omega-agatoxin IVA and PMA inhibited both the early [Ca2+]i transients and Mn2+ quenching to a much greater extent than each drug separately. Inhibition of the [Ca2+]i transients by nitrendipine and PMA did not significantly exceed that produced by PMA alone. It is concluded that phorbol ester-mediated activation of protein kinase C inhibits preferentially L-type VSCCs over P/Q type channels in adrenal chromaffin cells. However, the possibility cannot be ruled out that dihydropyridine-resistant, non-P/Q type channels might also be negatively regulated by protein kinase C. This may represent an important pathway for the specific control of VSCCs by protein kinase C-linked receptors, not only in paraneurones but presumably also in neurones and other excitable cells.

Animals↗
PubMed1999

Local drug delivery of argatroban from a polymeric-metallic composite stent reduces platelet deposition in a swine coronary model.

Thrombus formation after intracoronary stent implantation provides a stimulus for neointimal hyperplasia and if excessive can result in stent thrombosis. We tested the hypothesis that local delivery of an antithrombin drug from a polymeric-metallic stent inhibits platelet thrombus formation. An uncoated metal slotted tube, a jellyroll slotted metal stent with an Argatroban-loaded polymeric sleeve, and a jellyroll slotted metal stent with a drug-leached polymeric sleeve were randomly deployed into the coronary arteries of eight juvenile farm swine. Platelet deposition in the stented segments was determined at 2 hr using autologous 111Indium oxime-labeled platelets. Platelet deposition was significantly less in the Argatroban-loaded stents compared to the Argatroban-leached stents (1.40 x 10(8) platelets/cm2 vs. 26.8 x 10(8) platelets/cm2; P=0.005). When corrected for differences in the metal surface area exposed to blood, platelet deposition was significantly lower in the Argatroban-loaded stent (1.74+/-1.95 x 10(8)/cm2) compared to the Argatroban-leached stent (33.5+/-39.1 x 10(8)/cm2; P=0.005) and the uncoated metal stent (36.2+/-73.3 x 10(8)/cm2; P=0.006). In this coronary stent thrombosis model Argatroban has local antithrombotic properties when delivered with a polymer-metallic stent. Improved polymeric designs may reduce risk of thrombus deposition at the site of stent implantation.

Animals↗
PubMed1999

Interaction between substance P and gastrin-releasing peptide on thyrotropin secretion by rat pituitary in vitro.

The effect of substance P (SP) on thyrotropin (TSH) secretion is controversial. In this study we evaluated the effect of SP on TSH secretion by hemipituitaries of 3-month-old Wistar rats in vitro and its interaction with gastrin-releasing peptide (GRP) at equimolar concentrations (1 microM and 10 microM). TSH release was measured under basal conditions and 30 min after incubation in the absence or presence of SP, GRP or both peptides. Pituitary TSH content was also measured in the pituitary homogenate after incubation. SP at both concentrations caused a significant (P<0.05) increase in TSH secretion compared with all other groups, which was approximately 60% (1 microM) and 85% (10 microM) higher than that of the control group (23.3 +/- 3.0 ng/ml). GRP at the lower concentration did not produce a statistically significant change in TSH secretion, whereas at the concentration of 10 microM it produced a 50% reduction in TSH. GRP co-incubated with substance P completely blocked the stimulatory effect of SP at both concentrations. Pituitary TSH content decreased in the SP-treated group compared to controls (0.75 +/- 0.03 microg/hemipituitary) at the same proportion as the increase in TSH secretion, and this effect was also blocked when GRP and SP were co-incubated. In conclusion, in an in vitro system, SP increased TSH secretion acting directly at the pituitary level and this effect was blocked by GRP, suggesting that GRP is more potent than SP on TSH secretion, and that this inhibitory effect could be the predominant effect in vivo.

Analysis of Variance↗
PubMed1998

Local administration of L-703,081 using a composite polymeric stent reduces platelet deposition in canine coronary arteries.

We compared the effect on platelet deposition of the glycoprotein IIb/IIIa receptor antagonist L-703,081, administered locally via a drug delivery stent, with that of a standard metal stent in a canine coronary model. There was a significant reduction in platelet deposition using the L-703,081-impregnated stent compared with the bare metal stent. This study demonstrates an alternative route of delivery of GPIIb/IIIa antagonists with potential advantages over systemic administration.

Animals↗
PubMed1998

Control of pulsatile 5-HT/insulin secretion from single mouse pancreatic islets by intracellular calcium dynamics.

1. Glucose-induced insulin release from single islets of Langerhans is pulsatile. We have investigated the correlation between changes in cytosolic free calcium concentration ([Ca2+]i) and oscillatory insulin secretion from single mouse islets, in particular examining the basis for differences in secretory responses to intermediate and high glucose concentrations. Insulin release was monitored in real time through the amperometric detection of the surrogate insulin marker 5-hydroxytryptamine (5-HT) via carbon fibre microelectrodes. The [Ca2+]i was simultaneously recorded by whole-islet fura-2 microfluorometry. 2. In 82 % of the experiments, exposure to 11 mM glucose evoked regular high-frequency (average, 3.4 min-1) synchronous oscillations in amperometric current and [Ca2+]i. In the remaining experiments (18 %), 11 mM glucose induced an oscillatory pattern consisting of high-frequency [Ca2+]i oscillations that were superimposed on low-frequency (average, 0.32 min-1) [Ca2+]i waves. Intermittent high-frequency [Ca2+]i oscillations gave rise to a similar pattern of pulsatile 5-HT release. 3. Raising the glucose concentration from 11 to 20 mM increased the duration of the steady-state [Ca2+]i oscillations without increasing their amplitude. In contrast, both the duration and amplitude of the associated 5-HT transients were increased by glucose stimulation. The amount of 5-HT released per secretion cycle was linearly related to the duration of the underlying [Ca2+]i oscillations in both 11 and 20 mM glucose. The slopes of the straight lines were identical, indicating that there is no significant difference between the ability of calcium oscillations to elicit 5-HT/insulin release in 11 and 20 mM glucose. 4. In situ 5-HT microamperometry has the potential to resolve the high-frequency oscillatory component of the second phase of glucose-induced insulin secretion. This component appears to reflect primarily the duration of the underlying [Ca2+]i oscillations, suggesting that glucose metabolism and/or access to glucose metabolites is not rate limiting to fast pulsatile insulin release.

Animals↗
PubMed1998

TGF-beta isoforms in alcoholic liver disease.

The increased deposition of extracellular matrix proteins in the liver is a key factor in the morbidity and mortality of alcoholic liver disease (ALD). This increased fibrosis may be due to a superabundance of profibrogenic factors such as transforming growth factor-beta (TGF-beta). The original peptide is now called TGF-beta 1, and two other isoforms have been recognized in humans (TGF-beta 2 and TGF-beta 3). It was the aim of the present study to determine the expression of the TGF-beta isoforms in different stages of ALD. Thirty patients with ALD had percutaneous liver biopsies performed for diagnostic purposes. They were grouped by clinical findings and by liver histology into four groups: I, steatosis; II, fibrosis; III, hepatitis; and IV, cirrhosis. An unused portion of each biopsy sample was used to evaluate the gene expression of TGF-beta 1, TGF-beta 2, and TGF-beta 3 by reverse transcription polymerase chain reaction (RT-PCR). The expression of all isoforms from patients was significantly greater than their expression in controls. No significant correlation was determined between TGF-beta isoform expression and liver function test results. When the different isoforms were grouped by histology, increased expression with more severe disease was found; however, differences existed among the isoforms. In ALD, all TGF-beta isoforms were increased and their expression was significantly greater in patients with more active and advanced disease. RT-PCR is an effective method for evaluating gene expression in clinical samples which often provide a limited amount of tissue.

Adult↗
PubMed1998

Kissing stents in the aortic bifurcation.

BACKGROUND: We report the first series of simultaneously delivered stents used to treat stenosis of the aortic bifurcation. Surgical treatment of aortoiliac occlusive disease carries up to a 3% mortality rate. Percutaneous balloon techniques to treat aortic bifurcation stenosis, although safer, are still associated with up to a 9% incidence of dissection, thrombosis, or significant residual stenosis. Kissing stent insertion should decrease the incidence of these complications. METHODS: Twenty patients underwent kissing stent insertion. Suitable candidates included patients with symptoms of lower limb ischemia and significant atherosclerotic lesions in both ostial common iliac arteries (n = 15) or with extremely complex single ostial iliac stenoses (n = 5). Palmaz stents were delivered simultaneously to both limbs of the aortic bifurcation. RESULTS: Kissing stent insertion was successfully performed in all 20 patients without acute complications. Mean percent stenosis decreased from 46.2%+/-24.8% to -6.8%+/-13.3% (P = .0001) in the right iliac artery, 42.3%+/-22.8% to -1.6% +/-18.1% (P = .0001) in the left iliac artery, and 19.1%+/-16.6% to 2.3%+/-16.4% (P= .0008) in the distal aorta. Intermittent claudication symptoms were improved in 18 (95%) of 19 patients with 12 (63%) of 19 patients becoming totally asymptomatic. The strongest predictor of clinical outcome after kissing stent insertion was the preprocedural extent of femoropopliteal disease: 8 (89%) of 9 patients with femoropopliteal narrowing <75% bilaterally became completely asymptomatic at follow-up compared with only 3 (30%) of 10 patients with more severe stenoses (P = .02). CONCLUSIONS: We have demonstrated in 20 patients that stenoses of the aortic bifurcation can be treated effectively with kissing stents with few serious adverse events.

Aged↗
PubMed1998

Progression of renal artery stenosis in patients undergoing cardiac catheterization.

BACKGROUND: Renal artery stenosis is potentially correctable by either revascularization surgery or percutaneous methods. However, appropriate use of these techniques has been hampered by a lack of data on the natural history of this disease. This study assesses the prevalence, risk factors for progression, and effect on renal function of angiographically demonstrated renal artery disease in patients undergoing cardiac catheterization. METHODS: The severity of renal artery stenosis was quantified in all patients who underwent abdominal aortography as part of a diagnostic cardiac catheterization study at Duke University Medical Center between January 1989 and February 1996. RESULTS: There were 14,152 patients in the study (mean age 61+/-12 years, 62% male). Normal renal arteries were identified in 12,543 (88.7%) patients, insignificant disease (<50% stenosis) in 1 or more vessels in 726 patients (5.1 %), and significant stenosis in 883 patients (6.3%). Significant bilateral renal artery stenosis was present in 178 patients (1.3%). By multivariate logistic regression, elevated serum creatinine level, coronary artery disease, peripheral vascular disease, hypertension, cerebrovascular disease, older age, female sex, and family history of coronary artery disease were identified as independent predictors of significant renal arterial disease. Disease progression was assessed in 1189 patients. Mean time between cardiac catheterizations was 2.6+/-1.6 years. Significant disease progression occurred in 133 patients (11.1 %). Independent predictors of disease progression were female sex, age, coronary artery disease at baseline, and time between baseline and follow-up. At follow-up, serum creatinine level was significantly higher in patients who demonstrated > or =75% stenosis in 1 or more vessels (mean creatinine level 141+114 micromol/L compared with those with insignificant disease (mean creatinine level 97+/-44 micromol/L (P= .01). CONCLUSIONS: Renal artery disease is frequently progressive in patients who undergo cardiac catheterization for investigation of coronary artery disease. Significant stenotic disease may develop over a short period despite evidence of normal renal arteries at prior catheterization.

Age Factors↗
PubMed1998

[Changes of the complement system in myelodysplastic syndromes].

PURPOSE: To search complement system activity and detection of circulating immune complexes in a group of patients with myelodysplastic syndromes (MDS) classified as with relatively favourable and unfavourable prognosis. PATIENTS AND METHODS: 28 patients with MDS were examined, 12 with refractory anaemia and 4 with refractory ringed sideroblastic anaemia, both subgroups of relatively favourable prognosis; 9 patients with refractory anaemia with blastic excess and 3 with refractory anaemia with blastic excess in transformation, subgroups of unfavourable prognosis. We determined factor B, alternate and classical hemolytic activity, as well as C3 and C4 quantification of the complement system. Circulating immune complexes detection was performed by the C1q deviation test and polyethylene glycol 6000 precipitation method at 3.75% final concentration. RESULTS: A significant decrease in C3 level, alternate haemolytic activity and factor B was demonstrated in the unfavourable prognosis subgroups in comparison with patients of favourable prognosis and normal controls. CONCLUSIONS: Our results suggest the possibility of subclinical infections in patients with unfavourable prognosis.

Adult↗
PubMed1997

[Subisthmic aortic coarctation. Apropos a rare case of arterial hypertension].

An uncommon case of high blood pressure secondary to subisthmic aortic coarctation and cured surgically by arterial revascularization is reported. It is probably a congenital coarctation, because the extensive clinical evaluation and condition of the patient ruled out an acquired disease of the aorta, confirmed by the morphologic operative findings and pathological examinations of the aortic wall. In this case, the clinical diagnosis was initially suggested due to an abdominal systolic-diastolic bruit, associated to hypertension and weak femoral pulses. Although doppler ultrasonography and nuclear magnetic resonance were useful diagnostic tools, a complete angiography should be performed because it is currently the most informative technique to evaluate the aorta and its collaterals, frequently involved in association with subisthmic aortic coarctation. Surgical exploration revealed a hypoplastic descending thoracic aorta and histological data confirmed the nonexistence of acquired pathology of the aortic wall. Surgical management was accomplished successfully and the patient was asymptomatic 41 months after the operation with normal blood pressure, no treatment, and a normal pattern of life. Based on this presentation, the authors describe the etiology, morphology and anatomic classification, clinical manifestations, diagnosis and management of subisthmic aortic coarctation.

Adolescent↗