Arginine modulates endothelial phenotypes in experimental bronchopulmonary dysplasia-associated pulmonary hypertension: potential involvement of RARS1–HES1 signaling

Abstract

Background: Bronchopulmonary dysplasia (BPD) is a complex chronic lung disease caused by the interaction between lung development and prenatal and postnatal injury events. Up to one-third of infants with BPD further develop pulmonary vascular dysfunction, leading to chronic pulmonary hypertension (PH). Infants with BPD-PH face increased mortality, a higher risk of adverse neurodevelopmental outcomes, and a lack of effective treatments.
Methods: A total of 30 study participants were recruited from the Children’s Hospital Affiliated to Shandong University, with the data collection period spanning from January 2023 to May 2025. Through metabolite analysis, we observed serum arginine concentrations were lower in the BPD-PH group than in the comparison group, but the between-group difference did not reach statistical significance (P = 0.0581). Subsequently, in vitro experiments were conducted to investigate the effects of arginine on the proliferation and migration abilities of vascular endothelial cells and to elucidate the underlying molecular mechanisms. Finally, in vivo experiments were performed to validate the protective role of arginine in BPD-PH.
Results: The results of the in vitro experiments indicate that arginine inhibits the proliferation and migration of vascular endothelial cells. Molecular mechanism studies revealed that arginine reduces the expression of α-smooth muscle actin, hypoxia-inducible factor-1α, and related proteins through the arginyl-tRNA synthetase-1 (RARS1)-hairy and enhancer of split homologue-1 (HES1) pathway, RARS1 and HES1 were detected in the same immunoprecipitated protein complex, supporting a possible involvement of RARS1–HES1 signaling. In neonatal rats exposed to hyperoxia, arginine supplementation was associated with changes in selected lung-tissue mRNA markers. Pulmonary hemodynamics, right-ventricular remodeling, vascular morphometry, and alveolar structure were not assessed.
Conclusion: Our results indicate that arginine alleviates BPD-PH by promoting vascular endothelial cell remodeling through the RARS1-HES1 pathway. Arginine supplementation may represent a novel therapeutic alternative for preventing BPD.

Keywords

Arginine / Arginyl-tRNA synthetase-1 / Bronchopulmonary dysplasia / Hairy and enhancer of split homologue-1 / Pulmonary hypertension

References

[1] Course CW, Kotecha EA, Course K, Kotecha S. The respiratory consequences of preterm birth: from infancy to adulthood. Br J Hosp Med (Lond). 2024; 85(8): 1-11. doi: 10.12968/hmed.2024.0141

[2] Lu W, Li M, Ji F, et al. Potential impact of specific therapy on pregnant women with pulmonary arterial hypertension without cardiac shunt: a descriptive study in northern China. Emerg Crit Care Med. 2024; 4(2): 60-66. doi: 10.1097/ec9.0000000000000103

[3] Lu W, Li M, Ji F, et al. General anesthesia is an independent predictor for worse maternal outcome in pregnant pulmonary arterial hypertension patients without cardiac shunt but not for those with shunt. Emerg Crit Care Med. 2025; 5(1): 3-8. doi: 10.1097/ec9.0000000000000104

[4] Mascarenhas D, Al-Balushi M, Al-Sabahi A, Weisz DE, Jain A, Jasani B. Pulmonary hypertension in preterm neonates with bronchopulmonary dysplasia: a meta-analysis. Arch Dis Child Fetal Neonatal Ed. 2025; 110(4): 344-352. doi: 10.1136/archdischild-2024-327547

[5] El-Saie A, Varghese NP, Webb MK, et al. Bronchopulmonary dysplasia-associated pulmonary hypertension: an updated review. Semin Perinatol. 2023; 47(6): 151817. doi: 10.1016/j.semperi.2023.151817

[6] Gentle SJ, Travers CP, Nakhmani A, Indic P, Carlo WA, Ambalavanan N. Intermittent hypoxemia and bronchopulmonary dysplasia with pulmonary hypertension in preterm infants. Am J Respir Crit Care Med. 2023; 207(7): 899-907. doi: 10.1164/rccm.202203-0580OC

[7] Evans CE, Cober ND, Dai Z, Stewart DJ, Zhao YY. Endothelial cells in the pathogenesis of pulmonary arterial hypertension. Eur Respir J. 2021; 58(3): 2003957. doi: 10.1183/13993003.03957-2020

[8] Fung TS, Ryu KW, Thompson CB. Arginine: at the crossroads of nitrogen metabolism. EMBO J. 2025; 44(5): 1275-1293. doi: 10.1038/s44318-025-00379-3

[9] Anand SK, Governale TA, Zhang X, et al. Amino acid metabolism and atherosclerotic cardiovascular disease. Am J Pathol. 2024; 194(4): 510-524. doi: 10.1016/j.ajpath.2023.12.006

[10] Shiraseb F, Asbaghi O, Bagheri R, Wong A, Figueroa A, Mirzaei K. Effect of l-arginine supplementation on blood pressure in adults: a systematic review and dose-response meta-analysis of randomized clinical trials. Adv Nutr. 2022; 13(4): 1226-1242. doi: 10.1093/advances/nmab155

[11] Tratsiakovich Y, Yang J, Gonon AT, Sjöquist PO, Pernow J. Arginase as a target for treatment of myocardial ischemia-reperfusion injury. Eur J Pharmacol. 2013; 720(1-3): 121-123. doi: 10.1016/j.ejphar.2013.10.040

[12] Heffernan KS, Fahs CA, Ranadive SM, Patvardhan EA. L-arginine as a nutritional prophylaxis against vascular endothelial dysfunction with aging. J Cardiovasc Pharmacol Ther. 2010; 15(1): 17-23. doi: 10.1177/1074248409354599

[13] Scott JA, Maarsingh H, Holguin F, Grasemann H. Arginine therapy for lung diseases. Front Pharmacol. 2021; 12: 627503. doi: 10.3389/fphar.2021.627503

[14] He XD, Gong W, Zhang JN, et al. Sensing and transmitting intracellular amino acid signals through reversible lysine aminoacylations. Cell Metab. 2018; 27(1): 151-166.e6. doi: 10.1016/j.cmet.2017.10.015

[15] Nangle LA, Xu Z, Siefker D, et al. A human histidyl-tRNA synthetase splice variant therapeutic targets NRP2 to resolve lung inflammation and fibrosis. Sci Transl Med. 2025; 17(789): eadp4754. doi: 10.1126/scitranslmed.adp4754

[16] Zhang L, Zhang Q, Guo C, et al. Hes1 in malignant tumors: from molecular mechanism to therapeutic potential. Front Immunol. 2025; 16: 1585624. doi: 10.3389/fimmu.2025.1585624

[17] Yang L, Wang X, Sun J, et al. Neuritin promotes angiogenesis through inhibition of DLL4/Notch signaling pathway. Acta Biochim Biophys Sin (Shanghai). 2021; 53(6): 663-672. doi: 10.1093/abbs/gmab039

[18] Takeshita K, Satoh M, Ii M, et al. Critical role of endothelial Notch1 signaling in postnatal angiogenesis. Circ Res. 2007; 100(1): 70-78. doi: 10.1161/01.RES.0000254788.47304.6e

[19] Yang XY, Jiang D, Wang YZ, et al. Chlorogenic acid alleviates renal fibrosis by reducing lipid accumulation in diabetic kidney disease through suppressing the Notch1 and Stat3 signaling pathway. Ren Fail. 2024; 46(2): 2371988. doi: 10.1080/0886022X.2024.2371988

[20] Sajadimajd S, Mohammadi B, Bahrami G, Madani SH, Hatami R, Miraghaee SS. Modulation of Notch signaling and angiogenesis via an isolated polysaccharide from Momordica charantia in diabetic rats. J Food Biochem. 2022; 46(2): e14033. doi: 10.1111/jfbc.14033

[21] Shin S, Gombedza FC, Awuah Boadi E, Yiu AJ, Roy SK, Bandyopadhyay BC. Reduction of TRPC1/TRPC3 mediated Ca(2+)-signaling protects oxidative stress-induced COPD. Cell Signal. 2023; 107: 110681. doi: 10.1016/j.cellsig.2023.110681

[22] Zhu Y, Chu Y, Lan Y, et al. Loss of endothelial TRPC1 induces aortic hypercontractility and hypertension. Circ Res. 2025; 136(5): 508-523. doi: 10.1161/CIRCRESAHA.124.325574

[23] Chen S, Zhang J, Li M, Zhou J, Zhang Y. Danhong injection combined with tPA protects the BBB through Notch-VEGF signaling pathway on long-term outcomes of thrombolytic therapy. Biomed Pharmacother. 2022; 153: 113288. doi: 10.1016/j.biopha.2022.113288

[24] Song Y, Zhang Y, Jiang H, et al. Activation of Notch3 promotes pulmonary arterial smooth muscle cells proliferation via Hes1/p27Kip1 signaling pathway. FEBS Open Bio. 2015; 5: 656-660. doi: 10.1016/j.fob.2015.08.007

[25] Liu C, Fu C, Sun Y, et al. Itaconic acid regulation of TFEB-mediated autophagy flux alleviates hyperoxia-induced bronchopulmonary dysplasia. Redox Biol. 2024; 72: 103115. doi: 10.1016/j.redox.2024.103115

[26] Xiong Z, Zhou X, Yue SJ. Methods for establishing animal model of bronchopulmonary dysplasia and their evaluation. Zhongguo Dang Dai Er Ke Za Zhi. 2017; 19(1): 121-125. doi: 10.7499/j.issn.1008-8830.2017.01.020

[27] Schreckenberg R, Schulz R, Itani N, et al. Inhibition of MMP2 activity mitigates N-omega-nitro-l-arginine-methyl ester (l-NAME)-induced right heart failure. Redox Biol. 2024; 76: 103308. doi: 10.1016/j.redox.2024.103308

[28] Vadivel A, Aschner JL, Rey-Parra GJ, et al. L-citrulline attenuates arrested alveolar growth and pulmonary hypertension in oxygen-induced lung injury in newborn rats. Pediatr Res. 2010; 68(6): 519-525. doi: 10.1203/PDR.0b013e3181f90278

[29] Gentle SJ, Travers CP, Clark M, Carlo WA, Ambalavanan N. Patent ductus arteriosus and development of bronchopulmonary dysplasia-associated pulmonary hypertension. Am J Respir Crit Care Med. 2023; 207(7): 921-928. doi: 10.1164/rccm.202203-0570OC

[30] Xu W, Janocha AJ, Erzurum SC. Metabolism in pulmonary hypertension. Annu Rev Physiol. 2021; 83: 551-576. doi: 10.1146/annurev-physiol-031620-123956

[31] Zhao J, Pan B, Fina M, et al. α-Synuclein arginylation in the human brain. Transl Neurodegener. 2022; 11(1): 20. doi: 10.1186/s40035-022-00295-0

[32] Kwon YT, Kashina AS, Davydov IV, et al. An essential role of N-terminal arginylation in cardiovascular development. Science. 2002; 297(5578): 96-99. doi: 10.1126/science.1069531