Biomedical subjects

J M Nicolás

Publications and source records attributed to J M Nicolás.

At least 19 recordsLinked to original sources

PubMed2002

Differential effects of moderate or heavy alcohol consumption on circulating adhesion molecule levels.

Epidemiological studies suggest that moderate but not heavy alcohol consumption provides protection against coronary heart disease. We assessed the relationship between alcohol consumption and serum levels of adhesion molecules involved in the pathogenesis of early atherosclerosis. One-hundred apparently healthy men with similar cardiovascular risk factors were divided into four groups according to ethanol intake. Moderate drinkers (20-40 g/day) showed lower serum intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) levels than abstainers (p < 0.05; both), as well as lower serum ICAM-1, VCAM-1 and E-selectin levels than heavy drinkers (p = 0.01; all). The latter also showed higher serum ICAM-1 and E-selectin levels than abstainers (p < 0.001; both). We conclude that moderate drinkers show a significant reduction of soluble endothelial adhesion molecule levels compared to abstainers and heavy drinkers, that may contribute to the protective effect of moderate alcohol consumption against atherosclerosis.

Adult↗

PubMed2001

Human hepatic stellate cells secrete adrenomedullin: potential autocrine factor in the regulation of cell contractility.

BACKGROUND/AIMS: Hepatic stellate cells (HSCs) are perisinusoidal pericytes which have receptors for vasoactive factors, such as endothelin-1, which can regulate cell contractility in an autocrine manner. It is unknown whether human HSCs have receptors for and are able to synthesize the vasodilator peptide adrenomedullin (ADM), a peptide produced by most contractile cells. METHODS AND RESULTS: Stimulation of HSCs with ADM resulted in a dose-dependent raise in cAMP concentration (radioimmunoassay) and markedly blunted the endothelin-induced increase in [Ca2+]i and cell contraction, as assessed in cells loaded with fura-2 using a morphometric method. The existence of the receptor CRLR for ADM and their associated proteins RAMP-1 and RAMP-2 was demonstrated by reverse transcriptase-polymerase chain reaction (RT-PCR). Moreover, activated human HSCs spontaneously secreted ADM in the culture medium in a time-dependent manner. ADM secretion was markedly enhanced by tumour necrosis factor-alpha and interleukin-1beta. Specific mRNA for ADM (RT-PCR and Northern blot) was detected in HSCs and increased after incubation of cells with cytokines. CONCLUSIONS: Human HSCs have functional receptors for ADM, the stimulation of which blunts the contractile effect of endothelin-1. Cultured human HSCs secrete ADM in baseline conditions. This secretion is markedly increased by cytokines. These results suggest that ADM can regulate HSCs' contractility in an autocrine manner.

Adrenomedullin↗

PubMed2001

Human myofibroblastic hepatic stellate cells express Ca(2+)-activated K(+) channels that modulate the effects of endothelin-1 and nitric oxide.

BACKGROUND/AIMS: High-conductance Ca(2+)-activated K(+) (BK(Ca)) channels modulate the effects of vasoactive factors in contractile cells. It is unknown whether hepatic stellate cells (HSCs) contain BK(Ca) channels and what their role in the regulation of HSCs contractility is. METHODS: The presence of BK(Ca) channels in HSCs was assessed by the patch-clamp technique. The functional role of BK(Ca) channels was investigated by measuring intracellular calcium concentration ([Ca(2+)](i)) and cell contraction in individual cells after stimulation with endothelin-1 in the presence or absence of specific modulators of BK(Ca) channels. RESULTS: BK(Ca) channels were detected by patch-clamp in most of the activated HSCs studied. Incubation of cells with iberiotoxin, a BK(Ca) channel blocker, increased both the sustained phase of [Ca(2+)](i) elicited by endothelin-1 and the number of cells undergoing contraction, while the use of NS1619, a BK(Ca) channel opener, induced opposite effects. Stimulation of HSCs with S-nitroso-N-acetyl-penicillamine (SNAP), a nitric oxide (NO)-donor, increased the opening of BK(Ca) channels and reduced the effects of endothelin-1. Conversely, iberiotoxin abolished the inhibitory effect of SNAP on endothelin-induced [Ca(2+)](i) increase and cell contraction. CONCLUSIONS: Activated human HSCs contain BK(Ca) channels that modulate the contractile effect of endothelin-1 and mediate the inhibitory action of NO.

Benzimidazoles↗

PubMed2001

In vitro and in vivo activation of rat hepatic stellate cells results in de novo expression of L-type voltage-operated calcium channels.

Following chronic liver injury, hepatic stellate cells (HSCs) transdifferentiate into myofibroblast-like cells, which develop contractile properties and contribute to increased resistance to blood flow. We investigated whether this phenotypic activation includes changes in the expression of L-type voltage-operated Ca2+ channels (VOCC), which mediate Ca2+ influx and regulate cell contraction in vascular cell types. Rat HSCs were studied in the quiescent phenotype and after their activation in vitro (cultured on plastic for 14 days) and in vivo (isolated from rats with CCl(4)-induced cirrhosis). Patch-clamp studies showed Ca2+ currents through L-type VOCC in HSCs activated both in vitro and in vivo, whereas no currents were detected in quiescent HSCs. Moreover, binding studies with (3)H-isradipine, a specific L-type VOCC antagonist, showed a large number of binding sites in activated HSCs, while no specific binding was found in quiescent HSCs. Finally, messenger RNA (mRNA) encoding L-type VOCC was not detected in quiescent HSCs as assessed by reverse transcription-polymerase chain reaction (RT-PCR) and Northern blot analysis, whereas it was present in activated HSCs. Stimulation of L-type VOCC with KCl resulted in a marked increase in [Ca2+](i) followed by cell contraction in HSCs activated both in vitro and in vivo, whereas no effects were observed in quiescent HSCs. We conclude that the activation of HSCs is associated with up-regulation of L-type VOCC that mediate Ca2+ influx and cell contraction. These results may be relevant to the pathogenesis of portal hypertension.

Animals↗

PubMed2001

Sudden death in Staphylococcus aureus-associated infective endocarditis due to perforation of a free-wall myocardial abscess.

Free-wall myocardial abscess perforation with hemopericardium and sudden death is an extremely infrequent complication of infective endocarditis (IE). We describe a case of Staphylococcus aureus-associated native aortic and tricuspid valve endocarditis complicated by a septic myocardial infarction and abscess formation of embolic origin, with fatal rupture into the pericardium. To our knowledge, only 2 cases of myocardial abscess rupture have previously been reported in relation to IE.

Abscess↗

PubMed2001

Reappraisal of the aetiology and prognostic factors of severe acute respiratory failure in HIV patients.

The introduction of highly active antiretroviral therapy with protease inhibitors in 1996 has changed the morbidity and mortality of acquired immune deficiency syndrome patients. Therefore, the aetiologies and prognostic factors of human immunodeficiency virus (HIV)-infected patients with life-threatening respiratory failure requiring intensive care unit (ICU) admission need to be reassessed. From 1993 to 1998, we prospectively evaluated 57 HIV patients (mean+/-SEM age 36.5+/-1.3 yrs) admitted to the ICU showing pulmonary infiltrates and acute respiratory failure. A total of 21 and 30 patients were diagnosed as having Pneumocystis carinii and bacterial pneumonia, respectively, of whom 13 and eight died during their ICU stay (p=0.01). Both groups of patients had similar age, Acute Physiology and Chronic Health Evaluation (APACHE) II score, and severity in respiratory failure. The number of cases with bacterial pneumonia admitted to ICU decreased after 1996 (p=0.05). Logistic regression analysis showed that (APACHE) II score >17, serum albumin level <25 g.(-1), and diagnosis of P. carinii pneumonia were the only factors at entry associated with ICU mortality (p=0.02). Patients with bacterial pneumonia are less frequently admitted to the intensive care unit after the introduction of highly active antiretroviral therapy with protease inhibitors in 1996. Compared to the previous series, it was observed that the few Pneumocystis carinii pneumonia patients that need intensive care still have a bad prognosis.

AIDS-Related Opportunistic Infections↗

PubMed2001

Increased circulating leptin levels in chronic alcoholism.

BACKGROUND: Malnutrition seen in chronic alcoholics is partly due to reduced energy intake. Leptin is a peptide hormone implicated in the regulation of appetite and expenditure of energy. The prevalence and significance of abnormal circulating leptin levels in alcoholics, as well as the relationship of these levels with nutritional status, liver disease, and ethanol consumption, remain uncertain. METHODS: Serum leptin levels were measured in 60 active asymptomatic alcoholics, 20 active alcoholics with cirrhosis of the liver, 20 abstinent alcoholics, and 60 controls. Nutritional status and ethanol consumption also were assessed. RESULTS: In the control group, circulating leptin levels (mean 4.7+/-0.3 microg/liter) correlated with body fat stores. Despite showing a lower fat area of the arm, active alcoholics had significantly higher leptin levels than the controls (p < 0.001), regardless of the presence of cirrhosis. By contrast, none of the abstinent alcoholics showed hyperleptinemia. In the multivariate regression analysis, the fat area of the arm (p < 0.001), the lifetime ethanol consumption (p = 0.007), and the number of cigarettes smoked per day (p = 0.02) were found to be independent factors that influenced leptin levels in active alcoholics. After we adjusted for age, fat area of the arm, and tobacco consumption, a significant correlation was observed between lifetime consumption of ethanol and serum leptin concentrations (r = 0.36, p < 0.001). CONCLUSIONS: Circulating leptin levels are increased in a dose-dependent manner in chronic alcoholism, regardless of nutritional status or the presence of compensated liver disease.

Adult↗

PubMed2000

Angiotensin II induces contraction and proliferation of human hepatic stellate cells.

BACKGROUND & AIMS: Circulating levels of angiotensin II (ANGII), a powerful vasoconstrictor factor, are frequently increased in chronic liver diseases. In these conditions, hepatic stellate cells (HSCs) proliferate and acquire contractile properties. This study investigated the presence of receptors for ANGII and the effects of ANGII in human HSCs activated in culture. METHODS: The presence of ANGII receptors was assessed by binding studies. The effects of ANGII on intracellular calcium concentration ([Ca(2+)](i)), cell contraction, and cell proliferation were also assessed. RESULTS: Binding studies showed the presence of ANGII receptors of the AT1 subtype. ANGII elicited a marked dose-dependent increase in [Ca(2+)](i) and cell contraction. Moreover, ANGII stimulated DNA synthesis and increased cell number. All these effects were totally blocked by losartan and reduced by nitric oxide donors or prostaglandin E(2). The effects of ANGII were barely detectable in quiescent cells (2 days in culture), suggesting that phenotypic transformation of HSCs is associated with a marked increase in the effects of ANGII. CONCLUSIONS: ANGII induces contraction and is mitogenic for human-activated HSCs by acting through AT1 receptors. These results suggest that activated HSCs are targets of the vasoconstrictor action of ANGII in the intrahepatic circulation.

Angiotensin II↗

PubMed2000

Low-dose ethanol consumption allows strength recovery in chronic alcoholic myopathy.

Chronic skeletal myopathy may affect one third of chronic alcohol misusers. It is generally accepted that abstinence allows partial recovery, and that continued high-dose ethanol consumption progressively deteriorates muscle function. However, the effect of low-dose ethanol consumption in alcoholic myopathy has not been studied. We studied 58 chronic alcoholic male patients with biopsy-proven chronic alcoholic myopathy over 5 years. We evaluated ethanol intake, biochemical and nutritional parameters, and assessed muscle strength. Eighteen patients who remained abstinent showed marked improvement in muscle strength. As expected, the 19 patients who persisted in high-dose ethanol consumption further diminished in their muscle strength. In the 11 patients who maintained low-dose (</=60 g ethanol/day) 'controlled' drinking, muscle strength improved (p=0.003), despite no change in nutritional and exercise status. There is a dose-dependent recovery in muscle strength according to the degree of ethanol consumption, and moderate controlled drinking of up to 60 g ethanol/day still allows improvement in muscle strength.

Adult↗

PubMed2000

High ethanol intake and malnutrition in alcoholic cerebellar shrinkage.

To determine the influence of chronic ethanol intake and nutritional status on cerebellar shrinkage in alcoholism, we studied 12 undernourished patients with acute Wernicke's encephalopathy (WE), 12 undernourished and 24 well-nourished asymptomatic chronic alcoholics, and 24 age-matched well-nourished controls, using morphometric analysis of MRI scans with volumetry of the cerebellum. Alcoholics reported a mean daily intake of ethanol of 177+/-8 g over a period of 27+/-1 years. Most undernourished alcoholics and half of the well-nourished alcoholics, compared to one-tenth of the controls, showed a significant reduction in cerebellar volume (p< or =0.01, both). Alcoholics with cerebellar shrinkage (n=33) were older (p=0.05) and tended to report greater daily ethanol intake than alcoholics without cerebellar shrinkage (n=15), although not significantly so (p=0.09). Cerebellar volume correlated negatively with age in controls and asymptomatic alcoholics (r> or =0.52, p< or =0.01, both), with a significantly greater shrinkage for age in the latter (p=0.003). Logistic regression analysis showed that malnutrition (OR 6.6 [95%CI 1.7-25.6], p=0.005) and a daily ethanol intake of more than 140 g over ten years (OR 6.1 [95%CI 1.8-20.5], p=0.003) were independently associated with the development of cerebellar shrinkage.

Adult↗

PubMed2000

Comparison of systemic cytokine levels in patients with acute respiratory distress syndrome, severe pneumonia, and controls.

BACKGROUND: The inflammatory response has been widely investigated in patients with acute respiratory distress syndrome (ARDS) and pneumonia. Studies investigating the diagnostic values of serum cytokine levels have yielded conflicting results and only little information is available for the differential diagnosis between ARDS and pneumonia. METHODS: Clinical and physiological data, serum concentrations of tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta and IL-6, and quantitative cultures of lower respiratory tract specimens were obtained from 46 patients with ARDS and 20 with severe pneumonia within 24 hours of the onset of the disease and from 10 control subjects with no inflammatory lung disease. Cytokine concentrations were compared between groups and determinants in addition to the diagnosis were tested. RESULTS: Serum TNF-alpha levels were significantly higher in ARDS patients (67 (57) pg/ml) than in patients with severe pneumonia (35 (20) pg/ml; p = 0.031) or controls (17 (8) pg/ml; p = 0.007). For IL-1beta and IL-6 the observed differences were not statistically significant between patients with ARDS (IL-1beta: 34 (65) pg/ml; IL-6: 712 (1058) pg/ml), those with severe pneumonia (IL-1beta: 3 (4) pg/ml, p = 0.071; IL-6: 834 (1165) pg/ml, p = 1.0), and controls (IL-1beta: 6 (11) pg/ml, p = 0.359; IL-6: 94 (110) pg/ml, p = 0.262). TNF-alpha (standardised coefficient beta = 0.410, p<0.001) and IL-1beta (standardised coefficient beta = 0.311, p = 0.006) were most strongly associated with the degree of lung injury, even when the diagnostic group was included in the statistical model. CONCLUSIONS: Serum TNF-alpha levels were higher in patients with ARDS than in those with severe pneumonia or in control subjects. Multivariate results suggest that the levels of systemic TNF-alpha and IL-1beta reflect the severity of the lung injury rather than the diagnosis.

Biomarkers↗

PubMed2000

Heart mitochondrial respiratory chain complexes are functionally unaffected in heavy ethanol drinkers without cardiomyopathy.

BACKGROUND: Harmful effects of chronic ethanol intake on liver mitochondria have been clearly demonstrated; however, mitochondria from skeletal muscle are preserved, and the effect of ethanol on heart mitochondria remains controversial. We assessed individual enzyme activity of mitochondrial respiratory chain (MRC) complexes and membrane oxidative damage of heart mitochondria in active ethanol drinkers before the development of dilated cardiomyopathy. PATIENTS AND METHODS: Heart samples were obtained from otherwise healthy organ donor individuals with a sudden death of traumatic or neurological cause in whom hearts could not be used because of absence of matched receptors or size inadequacy. Detailed history of alcohol intake was achieved from the relatives. Citrate synthase activity was spectrophotometrically assayed, as well as absolute (nmol x min(-1) x mg protein(-1)) and relative (corrected by citrate synthase) activities of complex I, II, III, and IV of the MRC. Oxidative damage of myocardium membranes was assessed measuring the degree of lipid peroxidation by fluorescence using cis-parinaric acid as probe. RESULTS: We included 10 ethanol drinkers (age 53 +/- 13 years, 100% males, mean lifetime intake of 15.6 +/- 7.9 kg ethanol kg body weight(-1)) and 12 controls (age 60 +/- 10 years, 75% males). Mitochondrial content did not differ between the two groups. Absolute enzyme activities for ethanol drinkers and controls were, respectively, 145 +/- 75 and 130 +/- 50 for complex I (p = NS); 399 +/- 193 and 376 +/- 100 for complex II (p = NS); 719 +/- 288 and 714 +/- 308 for complex III (p = NS); and 475 +/- 139 and 570 +/- 160 for complex IV (p = NS). After correcting such activities by citrate synthase activity, we failed again to demonstrate differences between ethanol drinkers and controls. Lipid peroxidation of myocardium membranes was similar in both groups. CONCLUSIONS: Chronic ethanol drinkers without cardiomyopathy exhibit normal MRC activity in the heart and do not show increased oxidative damage in myocardial membranes.

Adult↗

PubMed2000

Diastolic function impairment in alcoholics.

BACKGROUND: Chronic excessive ethanol consumption exerts a deleterious effect on the myocardium. Although the effects of chronic alcoholism on systolic cardiac function are well known, diastolic involvement has been evaluated only partially. Therefore, we determined the presence of left ventricular diastolic impairment in chronic alcoholics and its relation with simultaneous systolic dysfunction. We also assessed the influence of ethanol consumption in diastolic impairment. METHODS: Thirty-five alcoholics with cardiomyopathy (ejection fraction < or = 50%) and 77 alcoholics with normal systolic function (ejection fraction > 50%) were evaluated. Assessment of New York Heart Association functional class, history of ethanol intake, technetium-99m radionuclide angiocardiography, and bidimensional Doppler echocardiography with evaluation of systolic and diastolic left ventricular function were performed. RESULTS: Diastolic function impairment was present in one third of the alcoholics without cardiomyopathy, compared with two thirds of the patients with cardiomyopathy (p < 0.01). A pseudonormalization phenomenon of diastolic function was observed in patients with more advanced systolic dysfunction (ejection fraction < 32%). The deterioration of the diastolic parameters correlated with ethanol consumption, regardless of age (r = 0.44, p < 0.001 for ratio of peak velocity of the transmitral flow in early diastole and peak velocity of atrial contraction flow, with lifetime dose of ethanol). CONCLUSIONS: There seems to be a dose-dependent effect of ethanol on systolic and diastolic heart function. Diastolic function impairment is present in one third of alcoholics with normal systolic function and is even more frequent when systolic dysfunction coexists.

Adult↗

PubMed1999

Atrial natriuretic peptide antagonizes endothelin-induced calcium increase and cell contraction in cultured human hepatic stellate cells.

Hepatic stellate cells (HSCs) participate in the regulation of hepatic microcirculation and have receptors for many vasoconstrictor factors. It is unknown whether HSCs have receptors for circulating vasodilators such as atrial natriuretic peptide (ANP). This study investigated the presence of ANP receptors in human HSCs and whether ANP antagonizes the effects of endothelin-1 in these cells. ANP receptors were assessed by binding and cross-linking studies, reverse-transcriptase polymerase chain reaction (PCR), and measuring intracellular cyclic guanosine monophosphate concentration. Intracellular calcium concentration ([Ca(2+)](i)) and cell contraction were measured in individual cells loaded with fura-2 using a morphometric method. Binding and cross-linking affinity experiments showed the existence of ANP receptors in human HSCs. PCR products with the expected length were obtained for guanylate cyclase A receptor, the physiological receptor of ANP, both in quiescent and activated human cells. ANP induced a dose-dependent increase in intracellular cyclic guanosine monophosphate concentration and blunted the increase in [Ca(2+)](i) elicited by endothelin-1. Most importantly, ANP markedly reduced cell contraction induced by endothelin-1. HSCs isolated from rats with carbon tetrachloride-induced cirrhosis showed a higher number of ANP receptors compared with HSCs isolated from normal rats, indicating that in vivo activation of HSCs is associated with an up-regulation of ANP receptors. These results indicate that human HSCs have receptors for ANP, the activation of which reduces the effects of endothelin-1 on [Ca(2+)](i) and cell contraction. ANP could participate in regulating the contractility of HSCs by antagonizing the effect of vasoconstrictors.

Animals↗

PubMed1999

Bacterial colonization patterns in mechanically ventilated patients with traumatic and medical head injury. Incidence, risk factors, and association with ventilator-associated pneumonia.

We prospectively evaluated the relation of upper airway, lower airway, and gastric colonization patterns with the development of pneumonia and its etiology in 48 patients with surgical (n = 25) and medical (n = 23) head injury. Initial colonization was assessed by cultures of nasal and pharyngeal swabs, tracheobronchial aspirates, gastric juice, and bronchoscopically retrieved protected specimen brush. Follow-up colonization was determined until the end points extubation, suspected ventilator-associated pneumonia (VAP), or death. The initial colonization rate at any site at ICU admission was 39/47 (83%). It mainly accounted for Group I pathogens (Streptococcus pneumoniae, Staphylococcus aureus, Hemophilus influenzae) of the upper and lower airways. At follow-up, colonization rates with Group II pathogens (Gram-negative enteric bacilli and Pseudomonas spp.) increased significantly. The high initial bacterial load with Group I pathogens of the upper airways and trachea decreased during Days 2 to 4, whereas that of Group II pathogens increased. Upper airway colonization was an independent predictor of follow-up tracheobronchial colonization (odds ratio [OR], 9.9; 95% confidence interval [CI], 1.8 to 56.3 for initial colonization with Group I pathogens; OR, 23.9; 95% CI, 3.8 to 153.3 for follow-up colonization with Group II pathogens). Previous (short-term) antibiotics had a protective effect against colonization with Group I pathogens of the lower respiratory tract (OR, 0.2; 95% CI, 0.05 to 0.86), but they were a risk factor for colonization with Group II pathogens (OR, 6.1; 95% CI, 1.3 to 29). Initial tracheobronchial colonization with Group I pathogens was associated with a higher probability of early onset pneumonia (OR, 4. 1; 95% CI, 0.7 to 23.3), whereas prolonged antibiotic treatment (> 24 h) independently predicted late-onset pneumonia (OR, 9.2; 95% CI, 1.7 to 51.3). We conclude that patients with head injury are colonized in the airways mainly by Group I pathogens early in the evolution of illness. The upper airways represent the main reservoir for subsequent lower airway colonization with Group I pathogens. Previous (short-term) antibiotic treatment is protective against initial tracheobronchial colonization with Group I pathogens, but it represents a risk factor for subsequent lower airway colonization by Group II pathogens.

Adult↗

PubMed1999

Upregulated expression of VLA proteins and CD29 in peripheral blood lymphocytes of chronic alcoholics without ethanol-related diseases.

To analyze adhesion molecule expression on peripheral blood mononuclear cells (PBMCs) and on different lymphocyte subpopulations (CD2+, CD8+, CD19+, and CD56+ subsets) in chronic alcoholism, 30 well-nourished chronic alcoholics without ethanol-related diseases and 30 matched controls were included in the study. Adhesion molecules that mediate adhesion to other cells and to extracellular matrix proteins, and whose cellular expression is modified during lymphocyte activation, were selected for study. A detailed clinical evaluation, laboratory analysis, nutritional assessment, and study of adhesion molecule expression was performed. A significant higher expression of CD29 (beta1-integrin) (p = 0.001), VLA-3 (p = 0.002), VLA-4 (p = 0.03), and VLA-5 (p = 0.001) were observed on PBMCs of chronic alcoholics, compared with control subjects, whereas no changes were observed in CD18 (beta2-integrin) and CD50 (ICAM-3) expression. The upregulation of CD29 and VLA proteins only affected T lymphocytes (CD2+/CD8+/CD4+ cells). These data confirm that T cells of chronic alcoholics are basally activated and that changes in adhesion molecule expression on PBMCs may be responsible of disturbances of adhesion processes in chronic alcoholics without ethanol-related diseases.

Adult↗

PubMed1999

Bradykinin decreases outflow facility in perfused anterior segments and induces shape changes in passaged BTM cells in vitro.

PURPOSE: To investigate the effects of bradykinin (BK) on the outflow facility (C) of human and bovine perfused anterior segments, the [Ca2+]i of cultured bovine trabecular meshwork (BTM) cells, and the area and major axis of BTM cells. METHODS: Cellular studies were performed using first- through third-passage cultures of BTM cells. For [Ca2+]i and shape change assessment, BTM cells were loaded with fura-2 acetoxymethyl ester, and individual fluorescence images were analyzed after the different experimental manipulations. C studies were performed in vitro using human and bovine anterior segments perfused at constant pressure. RESULTS: Bradykinin at 10(-6) M elicited a [Ca2+]i increase of 8 to 10 times the basal levels in 90% of the studied cells. From the responder cells, 60% elicited a 15%+/-1% reduction of the initial cell area, and 37% showed a 13%+/-2% reduction of their major axis. Bradykinin failed to induce any effect in the presence of the BK-B2 receptor antagonist HOE-140. Zero [Ca2+]o the depletion of intracellular stores with thapsigargin, or the presence of the calmodulin antagonist W13, decreased the BK response significantly (P < 0.001; P < 0.001; and P < 0.05). A second application of BK elicited a significantly lower (P < 0.001) response than the previous one. Perfusion with 10(-6) M BK decreased CD, calculated as the area under the curve, by 13%+/-4% (P < 0.05) in human anterior segments and 12%+/-4% (P < 0.05) in bovine anterior segments. The presence of 10(-6) M HOE-140, a BK-B2 receptor antagonist, completely blocked the decrease in C after perfusion with BK. CONCLUSIONS: The C of human and bovine trabecular meshwork (perfused anterior segments) is decreased by BK, acting through BK-B2 receptors. Primary cultured BTM cells respond to BK stimulation by increasing their [Ca2+]i by mobilization of extracellular and intracellular Ca2+. Moreover, these cells are reduced in area and their major axis shortened after the [Ca2+]i peak elicited by BK through BK-B2 receptors. The [Ca2+]i mobilization and shape changes are calmodulin dependent. Taking into account the [Ca2+]i mobilization, the BTM shape changes, the decrease of C, and the temporal sequence of these parameters, a contraction of trabecular meshwork cells related to the functional role of trabecular meshwork is discussed.

Aged↗

PubMed1998

CD148 is a membrane protein tyrosine phosphatase present in all hematopoietic lineages and is involved in signal transduction on lymphocytes.

Evidence is presented showing that a protein tyrosine phosphatase different from CD45 is present on the membrane of human hematopoietic cells. The molecule recognized by the monoclonal antibody 143-41, which has been classified as CD148 in the VI International Workshop on Leukocyte Differentiation Antigens, was immunopurified and sequenced. The sequence obtained from N-terminus as well as from two different CNBr-digested peptides showed a close identity with a previously described tyrosine phosphatase named HPTP-eta/DEP-1. CD148 is present on all hematopoietic lineages, being expressed with higher intensity on granulocytes than on monocytes and lymphocytes. Interestingly, whereas it is clearly present on peripheral blood lymphocytes, it is poorly expressed on different lymphoid cell lines of T and B origin. When this protein tyrosine phosphatase was cocrosslinked with CD3, an inhibition of the normally observed calcium mobilization was observed. This inhibition correlates with a decrease in phospholipase C-gamma (PLC-gamma) phosphorylation and is similar to the one observed with CD45. In addition, it is shown that the crosslinking of the CD148 alone is also able to induce an increase in [Ca2+]i. This increase is abolished in the presence of genistein and by cocrosslinking with CD45. These data, together with the induction of tyrosine phosphorylation on several substrates, including PLC-gamma, after CD148 crosslinking, suggest the involvement of a tyrosine kinase-based signaling pathway in this process. In conclusion, the data presented show that CD148 corresponds to a previously described protein tyrosine phosphatase HPTP-eta/DEP-1 and that this molecule is involved in signal transduction in lymphocytes.

Amino Acid Sequence↗