Folic acid, either solely or combined with L‐citrulline, improves NO signaling and ameliorates chronic hypoxia‐induced pulmonary hypertension in newborn pigs

Abstract

Concomitant with developing pulmonary hypertension (PH), newborn piglets exposed to chronic hypoxia develop pulmonary vascular NO signaling impairments. PH is reduced and NO signaling is improved in chronically hypoxic piglets treated with the NO‐arginine precursor, L‐citrulline. Folic acid positively impacts NO signaling. We evaluated whether the effect on NO signaling and PH is greater using co‐treatment with folic acid and L‐citrulline than either alone. From day 3 to day 10 of hypoxia, piglets were treated solely with folic acid, solely with L‐citrulline, or co‐treated with both. Catheters were placed to measure in vivo hemodynamics. NO production was measured in vitro in dissected pulmonary arteries. Compared to normoxic piglets, pulmonary vascular resistance (PVR) was elevated and NO production was reduced in untreated hypoxic piglets. Regardless of treatment strategy, PVR was less in all three treated groups of hypoxic piglets when compared to the untreated hypoxic group. In addition, for all three groups of treated hypoxic piglets, NO production was higher than the untreated group. Improvements in PVR and NO production did not differ between piglets co‐treated with folic acid and L‐citrulline and those treated solely with either. Thus, the impact on NO production and PVR was not augmented by combining folic acid and L‐citrulline treatments. Nonetheless, treatment with folic acid, either singly or when combined with L‐citrulline, increases NO production and inhibits PH in chronically hypoxic newborn piglets. Folic acid merits consideration as a therapy for PH in human infants with chronic heart and lung conditions that are associated with chronic hypoxia.

Impact of l‐citrulline on nitric oxide signaling and arginase activity in hypoxic human pulmonary artery endothelial cells

“…It is important to emphasize that we are aware that our findings in this study might have differed had we used other concentrations of folic acid and/or l ‐citrulline. In fact, in a previous study using our chronically hypoxic piglet model, we did not find a greater impact on NO signaling from co‐treatment with folic acid and l ‐citrulline than either alone 38 . Of note, plasma folic acid levels averaged 0.8 μM in the folic acid‐treated piglets, both when treated solely with folic acid or co‐treated with l ‐citrulline 38 .…”

“…In fact, in a previous study using our chronically hypoxic piglet model, we did not find a greater impact on NO signaling from co‐treatment with folic acid and l ‐citrulline than either alone. 38 Of note, plasma folic acid levels averaged 0.8 μM in the folic acid‐treated piglets, both when treated solely with folic acid or co‐treated with l ‐citrulline. 38 It is quite possible that this plasma level of folic acid led to an increase in intracellular oxidative stress, as has been reported to occur in Caenorhabditis elegans .…”

“… 38 Of note, plasma folic acid levels averaged 0.8 μM in the folic acid‐treated piglets, both when treated solely with folic acid or co‐treated with l ‐citrulline. 38 It is quite possible that this plasma level of folic acid led to an increase in intracellular oxidative stress, as has been reported to occur in Caenorhabditis elegans . 58 Supportive of this possibility, folic acid treatment failed to reduce superoxide generation in pulmonary arteries from the chronically hypoxic piglets.…”

“…Another reason underlying our interest in evaluating folic acid is that previous studies by us with chronically hypoxic piglets and by another group of investigators with chronically hypoxic adult mice have shown that folic acid treatment improves PH in chronically hypoxic animal models. 38 Moreover, although the impact on human pulmonary vascular disease has not been studied, there are a number of publications providing evidence that folic acid can be an effective treatment for a variety of cardiovascular diseases in humans. [39][40][41][42] The choice of concentration of folic acid used in this study with hypoxic human PAECs merits discussion.…”

“…58 Supportive of this possibility, folic acid treatment failed to reduce superoxide generation in pulmonary arteries from the chronically hypoxic piglets. 38 Indeed, although folic acid has been considered to be safe, there is growing awareness of the need for more research to assess the health effects of using folic acid in amounts exceeding current recommendations for upper limits. 59,60 Limitations of our study should be mentioned.…”