Regulation of Smooth Muscle Cell Proliferation by NADPH Oxidases in Pulmonary Hypertension

Hyperproliferation of pulmonary arterial smooth muscle cells is a key component of vascular remodeling in the setting of pulmonary hypertension (PH). Numerous studies have explored factors governing the changes in smooth muscle cell phenotype that lead to the increased wall thickness, and have identified various potential candidates. A role for reactive oxygen species (ROS) has been well documented in PH. ROS can be generated from a variety of sources, including mitochondria, uncoupled nitric oxide synthase, xanthine oxidase, and reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. In this article, we will review recent data supporting a role for ROS generated from NADPH oxidases in promoting pulmonary arterial smooth muscle cell proliferation during PH

Blockade of Endothelin-1 Receptor Type B Ameliorates Glucose Intolerance and Insulin Resistance in a Mouse Model of Obstructive Sleep Apnea

Obstructive sleep apnea (OSA) is associated with insulin resistance (IR) and glucose intolerance. Elevated endothelin-1 (ET-1) levels have been observed in OSA patients and in mice exposed to intermittent hypoxia (IH). We examined whether pharmacological blockade of type A and type B ET-1 receptors (ETA and ETB) would ameliorate glucose intolerance and IR in mice exposed to IH. Subcutaneously implanted pumps delivered BQ-123 (ETA antagonist; 200 nmol/kg/day), BQ-788 (ETB antagonist; 200 nmol/kg/day) or vehicle (saline or propyleneglycol [PG]) for 14 days in C57BL6/J mice (10/group). During treatment, mice were exposed to IH (decreasing the FiO2 from 20.9% to 6%, 60/h) or intermittent air (IA). After IH or IA exposure, insulin (0.5 IU/kg) or glucose (1 mg/kg) was injected intraperitoneally and plasma glucose determined after injection and area under glucose curve (AUC) was calculated. Fourteen-day IH increased fasting glucose levels (122 ± 7 vs. 157 ± 8 mg/dL, PG: 118 ± 6 vs. 139 ± 8; both p < 0.05) and impaired glucose tolerance (AUCglucose: 19,249 ± 1105 vs. 29,124 ± 1444, PG AUCglucose: 18,066 ± 947 vs. 25,135 ± 797; both p < 0.05) in vehicle-treated animals. IH-induced impairments in glucose tolerance were partially ameliorated with BQ-788 treatment (AUCglucose: 21,969 ± 662; p < 0.05). Fourteen-day IH also induced IR (AUCglucose: 7185 ± 401 vs. 8699 ± 401; p < 0.05). Treatment with BQ-788 decreased IR under IA (AUCglucose: 5281 ± 401, p < 0.05) and reduced worsening of IR with IH (AUCglucose: 7302 ± 401, p < 0.05). There was no effect of BQ-123 on IH-induced impairments in glucose tolerance or IR. Our results suggest that ET-1 plays a role in IH-induced impairments in glucose homeostasis

SUN-LB121 Nifedipine Worsens Glucose Tolerance in C57BL/6J Mice Exposed to Intermittent Hypoxia

Background: Intermittent hypoxemia (IH), a pathognomonic component of obstructive sleep apnea (OSA), has been independently associated with development of glucose intolerance, insulin resistance, and type 2 diabetes. L-type calcium channel blockers (CCB) influence glucose homeostasis including insulin sensitivity and secretion. To date, the potential impact of the combined effects of L-type CCB and IH on fasting glycemia and glucose tolerance have not been examined. The objective of this study was to determine whether CCB alters glucose metabolism in a murine model of IH. Methods: Adult male C57BL6/J mice (age 19-week-old) were exposed to IH using an automated system with specially-modified cages that oscillated FiO 2 from 21% to 5.5% at a target rate of 60 events/h during a 12 h (7am – 7pm) light cycle to simulate severe OSA for 5 days. The L-type CCB, nifedipine, or vehicle (polyethleneglycol-400) were administered at a dose of 20mg/kg/day via subcutaneous osmotic pumps (Alzet model 2001). Mice were exposed to IH or intermittent air (IA) with four resulting groups: IA-vehicle (n=12), IH-vehicle (n=16), IA–nifedipine (n=10), and IH–nifedipine (n=13). Fasting glucose, intraperitoneal glucose tolerance test, and insulin levels were obtained after exposures. Results: In the absence of a L-type CCB, IH increased fasting (105.1 vs. 71.2 mg/dL; p<0.001) and 2-hour glucose levels (104.8 vs. 82.0 mg/dL; p=0.003). The area under the glucose tolerance curve (AUC) was also higher with IH than IA in mice treated with vehicle (17896.3 vs.13965.8 mg-min/dL; p<0.001). Although the effects of IH on fasting glucose levels were comparable with and without L-type CCB treatment, the 2-hour glucose levels and the AUCs were substantially different. A statistically significant interaction was noted for the 2-hr glucose levels between IH and treatment with a L-type CCB (IH-CCB: 193.7; IH-V: 122.6; IA-CCB: 103.5; and IA-V: 82.0 mg/dL; p<0.05 for interaction between IH and CCB). Finally, the AUC for IH-CCB treated mice was significantly higher than the AUC for IH-V treated mice (IH-CCB: 30223.1; IH-V: 17896.3. mg-min/dL; p=0.0001) Conclusions: In a murine model of IH, treatment with an L-type CCB exacerbates the deleterious effects of IH on glucose tolerance. Thus, use of CCB in patients with OSA should take into consideration these unfavorable effects particularly in those who are metabolically compromised

Endothelin-1 augments Na⁺/H⁺ exchange activity in murine pulmonary arterial smooth muscle cells via Rho kinase.

Excessive production of endothelin-1 (ET-1), a potent vasoconstrictor, occurs with several forms of pulmonary hypertension. In addition to modulating vasomotor tone, ET-1 can potentiate pulmonary arterial smooth muscle cell (PASMC) growth and migration, both of which contribute to the vascular remodeling that occurs during the development of pulmonary hypertension. It is well established that changes in cell proliferation and migration in PASMCs are associated with alkalinization of intracellular pH (pH(i)), typically due to activation of Na(+)/H(+) exchange (NHE). In the systemic vasculature, ET-1 increases pH(i), Na(+)/H(+) exchange activity and stimulates cell growth via a mechanism dependent on protein kinase C (PKC). These results, coupled with data describing elevated levels of ET-1 in hypertensive animals/humans, suggest that ET-1 may play an important role in modulating pH(i) and smooth muscle growth in the lung; however, the effect of ET-1 on basal pH(i) and NHE activity has yet to be examined in PASMCs. Thus, we used fluorescent microscopy in transiently (3-5 days) cultured rat PASMCs and the pH-sensitive dye, BCECF-AM, to measure changes in basal pH(i) and NHE activity induced by increasing concentrations of ET-1 (10(-10) to 10(-8) M). We found that application of exogenous ET-1 increased pH(i) and NHE activity in PASMCs and that the ET-1-induced augmentation of NHE was prevented in PASMCs pretreated with an inhibitor of Rho kinase, but not inhibitors of PKC. Moreover, direct activation of PKC had no effect on pH(i) or NHE activity in PASMCs. Our results indicate that ET-1 can modulate pH homeostasis in PASMCs via a signaling pathway that includes Rho kinase and that, in contrast to systemic vascular smooth muscle, activation of PKC does not appear to be an important regulator of PASMC pH(i)