An NADPH-oxidase complex containing at least two protein components (gp91-phox and p22-phox) and a unique low redox potential (-245 mV) cytochrome b-245 is the source of superoxide generated for bacterial killing in neutrophils and has been suggested as the oxygen sensor in the carotid body. In pure cultures of smooth muscle cells from calf small pulmonary arteries (300 microns diameter) the presence of the 91 kD protein specific to this cytochrome was demonstrated by Western blot analysis with monoclonal antibody 48. Low-temperature-difference spectrophotometry of homogenates of these cells demonstrated the characteristic cytochrome b-245 spectrum when titrated between redox potentials of -152 and -345 mV, consistent with the low redox potential form. When these same cells were exposed to hypoxia (approximately 40 mmHg), superoxide production increased significantly from 1.4 +/- 0.2 to 73 +/- 12 nmoles.min-1 mg-1 protein. Hypoxic generation of superoxide was inhibited by the NADPH-oxidase inhibitor diphenyleneiodonium (DPI: 10 microM) but not by the mitochondrial inhibitor myxathiazole (10 microM). The hypoxic superoxide increase was significantly greater than that observed from smooth muscle cells from large pulmonary arteries or from large or small systemic arteries. Fluorescence immunocytochemistry revealed the presence of the NADPH-oxidase protein in the walls of pulmonary arteries in rat lung slices, and confocal microscopy showed the complex to be widely distributed in the vicinity of the arterial smooth muscle walls. In hypoxia or norepinephrine (NP)-induced vasoconstriction of pulmonary artery rings from cats, the sensitivity to inhibition by DPI was observed to be significantly greater for hypoxia (ED50 = 0.8 microM) than for NP-induced (ED50 = 13.4 microM) constriction. Together these observations demonstrate that the unique cytochrome b-245 containing NADPH-oxidase is present in pulmonary artery smooth muscle and that an NADPH-oxidase or NADH-oxidoreductase complex is activated to release superoxide by hypoxic conditions. It is concluded that a trans-membrane NADPH-oxidase is the most likely and that activation of this system may be involved in the initiation of hypoxic pulmonary vasoconstriction.

In this review, the second of a two part series, the analytic techniques introduced in the first part are applied to a broad range of pulmonary pathophysiologic conditions. The contributions of hypoxic pulmonary vasoconstriction to both homeostasis and pathophysiology are quantitated for atelectasis, pneumonia, sepsis, pulmonary embolism, chronic obstructive pulmonary disease and adult respiratory distress syndrome. For each disease state the influence of principle variables, including inspired oxygen concentration, cardiac output and severity of pathology are explored and the actions of selected drugs including inhaled nitric oxide and infused vasodilators are illustrated. It is concluded that hypoxic pulmonary vasoconstriction is often a critical determinant of hypoxemia and/or pulmonary hypertension. Furthermore this analysis demonstrates the value of computer simulation to reveal which of the many variables are most responsible for pathophysiologic results.

These studies document striking pulmonary vasoconstrictor response to nitric oxide synthase (NOS) inhibition in monocrotaline (MCT) pulmonary hypertension in rats. This constriction is caused by elevated endothelin (ET)-1 production acting on ETA receptors. Isolated, red blood cell plus buffer-perfused lungs from rats were studied 3 wk after MCT (60 mg/kg) or saline injection. MCT-injected rats developed pulmonary hypertension, right ventricular hypertrophy, and heightened pulmonary vasoconstriction to ANG II and the NOS inhibitor N G-monomethyl-l-arginine (l-NMMA). In MCT-injected lungs, the magnitude of the pulmonary pressor response to NOS inhibition correlated strongly with the extent of pulmonary hypertension. Pretreatment of isolated MCT-injected lungs with combined ETA (BQ-123) plus ETB (BQ-788) antagonists or ETA antagonist alone prevented thel-NMMA-induced constriction. Addition of ETA antagonist reversed establishedl-NMMA-induced constriction; ETB antagonist did not. ET-1 concentrations were elevated in MCT-injected lung perfusate compared with sham-injected lung perfusate, but ET-1 levels did not differ before and after NOS inhibition. NOS inhibition enhanced hypoxic pulmonary vasoconstriction in both sham- and MCT-injected lungs, but the enhancement was greater in MCT-injected lungs. Results suggest that in MCT pulmonary hypertension, elevated endogenous ET-1 production acting through ETA receptors causes pulmonary vasoconstriction that is normally masked by endogenous NO production.

Rat lungs were perfused in an in vitro circuit with separate control of alveolar and pulmonary arterial O2 tension. With perfusion flow constant, the hypoxic pulmonary vasoconstrictor (HPV) response was measured as changes of perfusion pressure. HPV was a function of both alveolar O2 tension (PvO2) and was described by a double sigmoid response surface. Where RA-v is this pressure response expressed as a percent of the maximum, the linearized form of the response surface is given by log [RA-v/(100-RA-v)] = 3.93 - 1.029 (log PvO2) - 1.623 (log PAO2). From this relationship it was concluded that 1) HPV is determined by PAO2 and PvO2; 2) the fundamental stimulus-response relationship is a sigmoid with a 50% response when both PAO2 and PvO2 are 30.3 Torr; 3) PAO2 has a greater effect than PvO2 due in part to the geometry of the vascular wall but principally due to O2 exchange between alveolar gas and blood in small pulmonary arteries; 4) there is not a localized sensor for HPV (the response is accounted for by each smooth muscle cell in the pulmonary arterial wall responding to the O2 tension in its vicinity); and 5) the characteristics of the response suggest that the cell sensor resembles a cytochrome.

Six pentobarbital-anesthetized dogs were prepared with endobronchial tubes and electromagnetic flow probes. The effects of changing inspired oxygen concentrations (FIO2 = 1, 0.21, 0.15, 0.1, 0.075, 0.05, and 0) were tested on test segments of different size corresponding to left lower lobe, left upper lobe-lingula, left lung, right lung, right lung plus left lower lobe, right lung plus left upper lobe-lingula, and whole lung. In each test the rest of the lung received oxygen. Hypoxic pulmonary vasoconstriction is demonstrated by both diversion of blood flow away from hypoxic test segments and by increased perfusion pressure. Flow diversion (FD%) decreases with the size of the hypoxic test segment (%QSN) from a maximum of 75% for very small segments to zero when the whole lung is hypoxic. FD% increases linearly as alveolar oxygen tension (PAO2) of the test segment is decreased in the range of 130--28 Torr. When mixed venous oxygen tension (PVO2) is less than 45 Torr FD% is reduced. These relationships are described by FD% = [74.99 - 0.0778 (%QSN) - 0.00661 (%QSN)2] [1.268 - 0.0096 (PAO2)] [0.47 + 0.012 (PVO2)], with r = 0.92 and standard error for prediction of 8.4%. Pulmonary perfusion pressure changes (PPH/PPN) increase with the size of the hypoxic test segments from 0 with very small segments to approximately 2.2 for the hypoxic whole lung. For all test segments PPH/PPN increases linearly with PAO2. These relationships are described by PPH/PPN = 1 + [0.0043 (%QSN) + 0.000072 (%QSN)2] [1.234 - 0.0096 (PAO2)], with r = 0.91 and standard error for prediction of 0.3 units. Responses to hypoxic pulmonary vasoconstriction in dogs are therefore shown to be predictable and continuous, and the physiological basis for action of each of the variables is discussed.

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The regulation of the distribution of ventilation/perfusion ratios by hypoxic pulmonary vasoconstriction contributes to both the efficiency of gas exchange and to pulmonary hemodynamics. In this review, the first of two part series, are summarized the physiologic principles on which the analysis of ventilation/perfusion ratios and of pressure-flow relationships are based. A new combined analysis is introduced that permits the important contributions of hypoxic pulmonary vasoconstriction to overall gas exchange to be demonstrated in the circumstances of clinical complexity.