Background-Alveolar hypoxia acutely elicits pulmonary vasoconstriction (HPV). Chronic hypoxia (CH), despiteattenuating HPV, causes pulmonary hypertension (CH-PHT). HPV results, in part, from inhibition of O 2 -sensitive, voltage-gated potassium channels (Kv) in pulmonary artery smooth muscle cells (PASMCs). CH decreases Kv channel current/expression and depolarizes and causes Ca 2ϩ overload in PASMCs. We hypothesize that Kv gene transfer would normalize the pulmonary circulation (restore HPV and reduce CH-PHT), despite ongoing hypoxia. Methods and Results-Adult male Sprague-Dawley rats were exposed to normoxia or CH for 3 to 4 weeks and then nebulized orotracheally with saline or adenovirus (Ad5) carrying genes for the reporter, green fluorescent protein reporterϮhuman Kv1.5 (cloned from normal PA). HPV was assessed in isolated lungs. Hemodynamics, including Fick and thermodilution cardiac output, were measured in vivo 3 and 14 days after gene therapy by use of micromanometer-tipped catheters. Transgene expression, measured by quantitative RT-PCR, was confined to the lung, persisted for 2 to 3 weeks, and did not alter endogenous Kv1.5 levels. Ad5-Kv1.5 caused no mortality or morbidity, except for sporadic, mild elevation of liver transaminases. Ad5-Kv1.5 restored the O 2 -sensitive K ϩ current of PASMCs, normalized HPV, and reduced pulmonary vascular resistance. Pulmonary vascular resistance decreased at day 2 because of increased cardiac output, and remained reduced at day 14, at which time there was concomitant regression of right ventricular hypertrophy and PA medial hypertrophy. Conclusions-Kv1.5 is an important O 2 -sensitive channel and potential therapeutic target in PHT. Kv1.5 gene therapy restores HPV and improves PHT. This is, to the best of our knowledge, the first example of K ϩ channel gene therapy for a vascular disease.
The between-day repeatability of simultaneous measures of brachial artery diameter (D) (echo Doppler) and mean blood velocity (MBV) (pulsed Doppler) was tested during rest and exercise. On 3 separate days, six volunteers performed one trial of 1-min rest followed by a step increase in dynamic handgrip exercise for 4 min which required the lifting and lowering of a 4.4-kg weight (approximately 8-12% MVC) in a 1s/2s (work/rest) cadence. Measures for MBV and D were collected continuously on a beat-by-beat basis during the transition from rest to end exercise. The mean rest values over one min, and single data points at 30, 60, 120, and 240 s of exercise were extracted from the time series data. At all exercise time points, MBV was greater than rest (P < 0.05), but these levels were not different across test days. Arterial D at all exercise time points ranged from 3.8 +/- 0.1 mm to 4.1 +/- 0.1 mm (mean +/- SEM) and did not differ from rest (3.9 +/- 0.1 mm) (P > 0.05), nor did D differ between days. The mean between-day coefficient of variation for D was 4.08 +/- 0.7% at rest and ranged from 2.90 +/- 0.4% to 3.96 +/- 0.5% during exercise. The coefficient of variation for MBV was 13.2 +/- 2.6% at rest and reached 20.2 +/- 3.1% during the final min of exercise; the exercise variability was reduced to 14.9 +/- 2.4% by averaging MBV over 3 s (the duration of a contraction/relaxation duty cycle) (P < 0.05) with no further advantage of averaging over ten 60-s sample periods. The data indicate that, for the six subjects tested, Doppler ultrasound measures of arterial MBV and diameter during both rest and exercise were reproducible across different test days and can be used as a reliable, noninvasive means of testing hypotheses pertaining to blood flow control.
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