Endothelin-1 does not mediate the endothelium-dependent hypoxic contractions of small pulmonary arteries in rats.

Various pulmonary artery preparations in vitro demonstrate sustained endothelium-dependent contractions upon hypoxia. To determine whether endothelin-1 could mediate this phenomenon, we examined the effect of bosentan, a new antagonist of both the ETA and ETB subtypes of the endothelin receptor. Small (300 pm) pulmonary arteries from rats were mounted on a myograph, precontracted with prostaglandin F2 alpha and exposed to hypoxia (PO2, 10 to 15 mm Hg, measured on-line) for 45 min. Endothelium-intact control rings exhibited a biphasic response, with a transient initial vasoconstriction (phase 1) followed by a second slowly developing sustained contraction (phase 2). Expressed in percent of the maximal response to 80 mmol/L KCl, the amplitudes of phase 1 (peak tension) and 2 (tension after 45 min of hypoxia) averaged 37 +/- 12% and 17 +/- 14%, respectively (n = 11). In endothelium-denuded rings, phase 1 persisted while the amplitude of phase 2 was reduced to 2 +/- 12% (p < 0.05, n = 8), showing the endothelium dependence of this contraction. Neither phase was significantly decreased in rings treated with 10(-5) mmol/L bosentan (38 +/- 15% and 17 +/- 12%, respectively, n = 6). The PO2 threshold for onset of hypoxic contraction was not significantly different among these three groups and averaged 32 +/- 24 mm Hg. In a separate experiment, we assessed the inhibitory effect of 10(-5) mol/L bosentan on the response to 10(-8) mol/L endothelin-I. Rings treated for 45 min with 10(-8) mol/L endothelin-1 alone exhibited a maximal contraction of 75 +/- 27% (n = 6). This was reduced to 4 +/- 17% (p < 0.01, n = 6) in rings treated with both 10(-8) mol/L endothelin-1 and 10(-5) mol/L bosentan. We conclude that complete blockade of all endothelin receptor subtypes has no effect on either endothelium-dependent or -independent hypoxic contractions in this preparation. This suggests that endothelial factors other than endothelin-I mediate the acute hypoxic contractions of small pulmonary arteries in the rat.

CD1d-antibody fusion proteins target iNKT cells to the tumor and trigger long-term therapeutic responses.

Despite the well-established antitumor activity of CD1d-restricted invariant natural killer T lymphocytes (iNKT), their use for cancer therapy has remained challenging. This appears to be due to their strong but short-lived activation followed by long-term anergy after a single administration of the CD1d agonist ligand alpha-galactosylceramide (αGC). As a promising alternative, we obtained sustained mouse iNKT cell responses associated with prolonged antitumor effects through repeated administrations of tumor-targeted recombinant sCD1d-antitumor scFv fusion proteins loaded with αGC. Here, we demonstrate that CD1d fusion proteins bound to tumor cells via the antibody fragment specific for a tumor-associated antigen, efficiently activate human iNKT cell lines leading to potent tumor cell lysis. The importance of CD1d tumor targeting was confirmed in tumor-bearing mice in which only the specific tumor-targeted CD1d fusion protein resulted in tumor inhibition of well-established aggressive tumor grafts. The therapeutic efficacy correlated with the repeated activation of iNKT and natural killer cells marked by their release of TH1 cytokines, despite the up-regulation of the co-inhibitory receptor PD-1. Our results demonstrate the superiority of providing the superagonist αGC loaded on recombinant CD1d proteins and support the use of αGC/sCD1d-antitumor fusion proteins to secure a sustained human and mouse iNKT cell activation, while targeting their cytotoxic activity and cytokine release to the tumor site.

Tissue oxygenation and hemodynamic response to NO synthase inhibition in septic shock.

The objective of the study was to evaluate the tissue oxygenation and hemodynamic effects of NOS inhibition in clinical severe septic shock. Eight patients with septic shock refractory to volume loading and high level of adrenergic support were prospectively enrolled in the study. Increasing doses of NOS inhibitors [N(G)-nitro-L-arginine-methyl ester (L-NAME) or N(G)-monomethyl-L-arginine (L-NMMA)] were administered as i.v. bolus until a peak effect = 10 mmHg on mean blood pressure was obtained or until side effects occurred. If deemed clinically appropriate, a continuous infusion of L-NAME was instituted and adrenergic support weaning attempted. The bolus administration of NOS inhibitors transiently increased mean blood pressure by 10 mm Hg in all patients. Seven out of eight patients received an L-NAME infusion, associated over 24 h with a progressive decline in cardiac index (P < 0.001) and an increase in systemic vascular resistance (P < 0.01). Partial or total adrenergic support weaning was rapidly possible in 6/8 patients. Oxygen transport decreased (P < 0.001), but oxygen consumption remained unchanged in those patients in whom it could be measured by indirect calorimetry (5/8). Blood lactate and the difference between tonometric gastric and arterial PCO2 remained unchanged. There were 4/8 ICU survivors. We conclude that nitric oxide synthase inhibition in severe septic shock was followed with a progressive correction of the vasoplegic hemodynamic disturbances with finally normalization of cardiac output and systemic vascular resistances without any demonstrable deterioration in tissue oxygenation.

Effect of repetitive arm cycling following botulinum toxin injection for poststroke spasticity: evidence from FMRI.

BACKGROUND AND OBJECTIVE: Investigations were performed to establish if repetitive arm cycling training enhances the antispastic effect of intramuscular botulinum toxin (BTX) injections in postischemic spastic hemiparesis. Effects on cerebral activation were evaluated by functional magnetic resonance imaging (fMRI). METHODS: Eight chronic spastic hemisyndrome patients (49 ± 10 years) after middle cerebral artery infarction (5.5 ± 2.7 years) were investigated. BTX was injected into the affected arm twice, 6 months apart. Spasticity was assessed using the Ashworth Scale and range of motion before and 3 months after BTX injections. Images were analyzed using Brain Voyager QX 1.8, and fMRI signal changes were corrected for multiple comparisons. RESULTS: During passive movements of affected and nonaffected hands, fMRI activity was increased bilaterally in the sensorimotor cortex (MISI), secondary somatosensory areas (SII), and supplementary motor area predominantly in the contralesional hemisphere, compared with the rest. Following repetitive arm cycling, fMRI activity increased further in MISI of the lesioned hemisphere and SII of the contralesional hemisphere. For patients with residual motor activity, treatment-related fMRI activity increases were associated with reduced spasticity; in completely plegic patients, there was no fMRI activity change in SII but increased spasticity after training. CONCLUSION: Increased activity in SII of the contralesional hemisphere and in MISI of the lesioned hemisphere reflect a treatment-induced effect in the paretic arm. It is hypothesized that the increased BOLD activity results from increased afferent information related to the antispastic BTX effect reinforced by training.

Diabetic neuropathy is a more important determinant of baroreflex sensitivity than carotid elasticity in type 2 diabetes.

The object of this study was to evaluate the contribution of carotid distensibilty on baroreflex sensitivity in patients with type 2 diabetes mellitus with at least 2 additional cardiovascular risk factors. Carotid distensibility was measured bilaterally at the common carotid artery in 79 consecutive diabetic patients and 60 matched subjects without diabetes. Spontaneous baroreflex sensitivity assessment was obtained using time and frequency methods. Baroreflex sensitivity was lower in diabetic subjects as compared with nondiabetic control subjects (5.25+/-2.80 ms/mm Hg versus 7.55+/-3.79 ms/mm Hg; P<0.01, respectively). Contrary to nondiabetic subjects, diabetic subjects showed no significant correlation between carotid distensibility and baroreflex sensitivity (r2=0.08, P=0.04 and r2=0.04, P=0.13, respectively). In diabetic subjects, baroreflex sensitivity was significantly lower in subjects with peripheral neuropathy than in those with preserved vibration sensation (4.1+/-0.5 versus 6.1+/-0.4 ms/mm Hg, respectively; P=0.005). Age in nondiabetic subjects, diabetes duration, systolic blood pressure, peripheral or sensitive neuropathy, and carotid distensibility were introduced in a stepwise multivariate analysis to identify the determinants of baroreflex sensitivity. In diabetic patients, neuropathy is a more sensitive determinant of baroreflex sensitivity than the reduced carotid distensibility (stepwise analysis; F ratio=5.1, P=0.028 versus F ratio=1.9, P=0.16, respectively). In diabetic subjects with 2 additional cardiovascular risk factors, spontaneous baroreflex sensitivity is not related to carotid distensibility. Diabetic subjects represent a particular population within the spectrum of cardiovascular risk situations because of the marked neuropathy associated with their metabolic disorder. Therefore, neuropathy is a more significant determinant of baroreflex sensitivity than carotid artery elasticity in patients with type 2 diabetes.