RESUMEN
Here, we tested the hypothesis that TNAP (tissue nonspecific alkaline phosphatase) modulates vascular responsiveness to norepinephrine. In the isolated, Tyrode's-perfused rat mesentery, 50 µmol/L of L-p-bromotetramisole (L-p-BT; selective TNAP inhibitor, Ki=56 µmol/L) significantly reduced TNAP activity and caused a significant 9.0-fold rightward-shift in the norepinephrine concentration versus vasoconstriction relationship. At 100 µmol/L, L-p-BT further reduced mesenteric TNAP activity and caused an additional significant right-shift of the norepinephrine concentration versus vasoconstriction relationship. A higher concentration (200 µmol/L) of L-p-BT had no further effect on either mesenteric TNAP activity or norepinephrine-induced vasoconstriction. L-p-BT did not alter vascular responses to vasopressin, thus ruling-out nonspecific suppression of vascular reactivity. Since in the rat mesenteric vasculature α1-adrenoceptors mediate norepinephrine-induced vasoconstriction, these finding indicate that TNAP inhibition selectively interferes with α1-adrenoceptor signaling. Additional experiments showed that the effects of TNAP inhibition on norepinephrine-induced vasoconstriction were not mediated by accumulation of pyrophosphate or ATP (TNAP substrates) nor by reduced adenosine levels (TNAP product). TNAP inhibition significantly reduced the Hillslope of the norepinephrine concentration versus vasoconstriction relationship from 1.8±0.2 (consistent with positive cooperativity of α1-adrenoceptor signaling) to 1.0±0.1 (no cooperativity). Selective activation of A1-adenosine receptors, which are known to participate in coincident signaling with α1-adrenoceptors, reversed the suppressive effects of L-p-BT on norepinephrine-induced vasoconstriction. In vivo, L-p-BT administration achieved plasma levels of ≈60 µmol/L and inhibited mesenteric vascular responses to exogenous norepinephrine and sympathetic nerve stimulation. TNAP modulates vascular responses to norepinephrine likely by affecting positive cooperativity of α1-adrenoceptor signaling via a mechanism involving A1 receptor signaling.
Asunto(s)
Fosfatasa Alcalina/metabolismo , Proteínas de la Membrana/metabolismo , Mesenterio/efectos de los fármacos , Norepinefrina/farmacología , Tetramisol/análogos & derivados , Vasoconstricción/efectos de los fármacos , Vasoconstrictores/farmacología , Antagonistas del Receptor de Adenosina A1/farmacología , Fosfatasa Alcalina/antagonistas & inhibidores , Fosfatasa Alcalina/genética , Animales , Masculino , Proteínas de la Membrana/antagonistas & inhibidores , Proteínas de la Membrana/genética , Mesenterio/metabolismo , Ratas , Tetramisol/farmacología , Xantinas/farmacologíaRESUMEN
OBJECTIVES: To determine the production of 9-hydroxyoctadecadienoic acid and 13-hydroxyoctadecadienoic acid during cardiopulmonary bypass in infants and children undergoing cardiac surgery, evaluate their relationship with increase in cell-free plasma hemoglobin, provide evidence of bioactivity through markers of inflammation and vasoactivity (WBC count, milrinone use, vasoactive-inotropic score), and examine their association with overall clinical burden (ICU/hospital length of stay and mechanical ventilation duration). DESIGN: Prospective observational study. SETTING: Twelve-bed cardiac ICU in a university-affiliated children's hospital. PATIENTS: Children were prospectively enrolled during their preoperative clinic appointments with the following criteria: greater than 1 month to less than 18 years old, procedures requiring cardiopulmonary bypass INTERVENTIONS:: None. MEASUREMENTS AND MAIN RESULTS: Plasma was collected at the start and end of cardiopulmonary bypass in 34 patients. 9-hydroxyoctadecadienoic acid, 13-hydroxyoctadecadienoic acid, plasma hemoglobin, and WBC increased. 9:13-hydroxyoctadecadienoic acid at the start of cardiopulmonary bypass was associated with vasoactive-inotropic score at 2-24 hours postcardiopulmonary bypass (R = 0.25; p < 0.01), milrinone use (R = 0.17; p < 0.05), and WBC (R = 0.12; p < 0.05). 9:13-hydroxyoctadecadienoic acid at the end of cardiopulmonary bypass was associated with vasoactive-inotropic score at 2-24 hours (R = 0.17; p < 0.05), 24-48 hours postcardiopulmonary bypass (R = 0.12; p < 0.05), and milrinone use (R = 0.19; p < 0.05). 9:13-hydroxyoctadecadienoic acid at the start and end of cardiopulmonary bypass were associated with the changes in plasma hemoglobin (R = 0.21 and R = 0.23; p < 0.01). The changes in plasma hemoglobin was associated with milrinone use (R = 0.36; p < 0.001) and vasoactive-inotropic score less than 2 hours (R = 0.22; p < 0.01), 2-24 hours (R = 0.24; p < 0.01), and 24-48 hours (R = 0.48; p < 0.001) postcardiopulmonary bypass. Cardiopulmonary bypass duration, 9:13-hydroxyoctadecadienoic acid at start of cardiopulmonary bypass, and plasma hemoglobin may be risk factors for high vasoactive-inotropic score. Cardiopulmonary bypass duration, changes in plasma hemoglobin, 9:13-hydroxyoctadecadienoic acid, and vasoactive-inotropic score correlate with ICU and hospital length of stay and/mechanical ventilation days. CONCLUSIONS: In low-risk pediatric patients undergoing cardiopulmonary bypass, 9:13-hydroxyoctadecadienoic acid was associated with changes in plasma hemoglobin, vasoactive-inotropic score, and WBC count, and may be a risk factor for high vasoactive-inotropic score, indicating possible inflammatory and vasoactive effects. Further studies are warranted to delineate the role of hydroxyoctadecadienoic acids and plasma hemoglobin in cardiopulmonary bypass-related dysfunction and to explore hydroxyoctadecadienoic acid production as a potential therapeutic target.
Asunto(s)
Puente Cardiopulmonar/métodos , Ácidos Grasos Insaturados/sangre , Cardiopatías Congénitas/cirugía , Ácidos Linoleicos/sangre , Oxilipinas/sangre , Biomarcadores/sangre , Procedimientos Quirúrgicos Cardíacos/efectos adversos , Procedimientos Quirúrgicos Cardíacos/métodos , Puente Cardiopulmonar/efectos adversos , Niño , Preescolar , Ácidos Grasos Insaturados/metabolismo , Femenino , Cardiopatías Congénitas/tratamiento farmacológico , Hemoglobinas/análisis , Humanos , Lactante , Unidades de Cuidados Intensivos , Tiempo de Internación , Recuento de Leucocitos , Ácidos Linoleicos/metabolismo , Masculino , Milrinona/uso terapéutico , Oxilipinas/metabolismo , Estudios Prospectivos , Respiración Artificial , Factores de Riesgo , Vasodilatadores/uso terapéuticoRESUMEN
SDF-1α (stromal cell-derived factor-1α) is a CXCR4-receptor agonist and DPP4 (dipeptidyl peptidase 4) substrate. SDF-1α, particularly when combined with sitagliptin to block the metabolism of SDF-1α by DPP4, stimulates proliferation of cardiac fibroblasts via the CXCR4 receptor; this effect is greater in cells from spontaneously hypertensive rats versus Wistar-Kyoto normotensive rats. Emerging evidence indicates that ubiquitin(1-76) exists in plasma and is a potent CXCR4-receptor agonist. Therefore, we hypothesized that ubiquitin(1-76), similar to SDF-1α, should increase proliferation of cardiac fibroblasts. Contrary to our working hypothesis, ubiquitin(1-76) did not stimulate cardiac fibroblast proliferation, yet unexpectedly antagonized the proproliferative effects of SDF-1α combined with sitagliptin. In this regard, ubiquitin(1-76) was more potent in spontaneously hypertensive versus Wistar-Kyoto cells. In the presence of 6bk (selective inhibitor of insulin-degrading enzyme [IDE]; an enzyme known to convert ubiquitin(1-76) to ubiquitin(1-74)), ubiquitin(1-76) no longer antagonized the proproliferative effects of SDF-1α/sitagliptin. Ubiquitin(1-74) also antagonized the proproliferative effects of SDF-1α/sitagliptin, and this effect of ubiquitin(1-74) was not blocked by 6bk and was >10-fold more potent compared with ubiquitin(1-76). Neither ubiquitin(1-76) nor ubiquitin(1-74) inhibited the proproliferative effects of the non-CXCR4 receptor agonist neuropeptide Y (activates Y1 receptors). Cardiac fibroblasts expressed IDE mRNA, protein, and activity and converted ubiquitin(1-76) to ubiquitin(1-74). Spontaneously hypertensive fibroblasts expressed greater IDE activity. Extracellular ubiquitin(1-76) blocks the proproliferative effects of SDF-1α/sitagliptin via its conversion by IDE to ubiquitin(1-74), a potent CXCR4 antagonist. Thus, IDE inhibitors, particularly when combined with DPP4 inhibitors or hypertension, could increase the risk of cardiac fibrosis.