Analysis of the neutrophils role in the development of inflammation in monocrotaline-induced pulmonary arterial hypertension

Original article

УДК 577.3’32/.’36; 577.34
DOI: 10.57034/2618723X-2023-04-06

G.N. Semenkova1*,
PhD (Biology), associate professor, leading researcher,
I.E. Adzerikho1,
PhD, MD (Medicine), professor,
N.V. Amaegberi2,
PhD (Biology), researcher,
T.E. Vladimirskaja1,
PhD (Biology), associate professor, leading researcher,
O.N. Yatsevich1,

1 Belarusian Medical Academy of Postgraduate Education,
Brovki st., 3, 220013, Minsk, Belarus.
2 Belarusian State University,
Nezavisimosti Ave., 4, 220030, Minsk, Belarus.

* E-mail: [email protected]

Keywords: pulmonary arterial hypertension; neutrophils; monocrotaline; reactive oxygen and chlorine species; myeloperoxidase


This work was supported by Belarusian Republican Foundation for Fundamental Research (grant No. М17-113).

For citation:

Semenkova G.N., Adzerikho I.E., Amaegberi N.V., Vladimirskaja T.E., Yatsevich O.N. Analysis of the neutrophils role in the development of inflammation in monocrotaline-induced pulmonary arterial hypertension. Laboratory Animals for Science. 2023; 4.


Pulmonary arterial hypertension (PAH) is a clinical syndrome with dismal outcome. This disease is associated with endothelial dysfunction or damage, pulmonary vascular remodeling, and increased vascular resistance and pulmonary artery blood pressure. All this provokes right ventricular failure and pulmonary edema, which leads to premature death of patients. PAH is accompanied by the inflammatory process deve­lopment, one of the main causes of which is the increased production of reactive oxygen and chlorine species by stimulated neutrophils. The purpose of our study was to study the role of these cells in the oxidative stress formation in experimental model of PAH.

The studies were conducted on outbred white rats. Modeling of PAH was carried out by subcutaneous administration of a monocrotaline solution at a dose of 60 mg/kg. The control group consisted of 10 healthy animals. After monocrotaline injection, PAH animals were divided into 4 groups: 10 rats in the 2-week group, 10 rats in the 4-week group, 14 rats in the 6-week group, and 20 rats in the 8-week group. After collecting peripheral blood and isolating neutrophils, reactive oxygen and chlorine species production, secretory degranulation and the unbound calcium ions concentration in the cytosol were determined using spectral analysis methods. The results were compared with corresponding data obtained for 10 healthy animals.

It has been established that the development of PAH in rats over 8 weeks is accompanied by neutrophil priming and oxidative stress formation as a result of “respiratory burst” mechanisms modification. Changes in reactive oxygen and chlorine species production, disruption of Ca2+-dependent intracellular signaling involving cyclooxygenase 1/2 and phosphatidylinositol 3-kinase, and increased secretion of myeloperoxidase from neutrophils were revealed.

Conflict of interest

The authors declare no conflict of interest.

Authors contribution

G.N. Semenkova — concept and design of the study, writing the text of the article.
I.E. Adzerikho — concept and design of the study.
N.V. Amaegberi — obtaining and processing experimental data.
T.E. Vladimirskaja — obtaining, processing, analysis of experimental data.
O.N. Yatsevich — obtaining and processing experimental data.
All authors read and agreed to the final version of the manuscript.


  1. Galiè N., Hoeper M.M., Humbert M. et al. Guidelines for the diagnosis and treatment of pulmonary hypertension // Eur. Respir. J. 2009. Vol. 34. N. 6. P. 1219–1263. DOI: 10.1183/09031936.00139009.
  2. El Chami H., Hassoun P.M. Immune and inflammatory mechanisms in pulmonary arterial hypertension // Prog. Cardiovasc. Dis. 2012. Vol. 55. N. 2. P. 218–228. DOI: 10.1016/j.pcad.2012.07.006.
  3. Tang C., Luo Y., Li S. et al. Characteristics of inflammation process in monocrotaline-induced pulmonary arterial hypertension in rats // Biomed Pharmaco­ther. 2021. Vol. 133. P. 111081. DOI: 10.1016/j.biopha.2020.111081.
  4. Touyz R.M. Reactive oxygen species, vascular oxidative stress, and redox signaling in hypertension: what is the clinical significance? // Hypertension. 2004. Vol. 44. P. 248–252. DOI: 10.1161/01.HYP.0000138070.47616.9d.
  5. Rafikova O., Al Ghouleh I., Rafikov R. Focus on early events: pathogenesis of pulmonary arterial hypertension development // Antioxid. Redox. Signal. 2019. Vol. 31. N. 13. P. 933–953. DOI: 10.1089/ars.2018.7673.
  6. Arnhold J. The dual role of myeloperoxidase in immune response // Int. J. Mol. Sci. 2020. Vol. 21. P. 8057. DOI: 10.3390/ijms21218057.
  7. Hawkins C.L., Pattison D.I., Davies M.J. Hypochlorite-induced oxidation of amino acids, peptides and proteins // Amino Acids. 2003. Vol. 25. N. 3–4. P. 259–274. DOI: 10.1007/s00726-003-0016-x.
  8. Padrez Y., Golubewa L., Kulahava T. et al. Quantitative and qualitative analysis of pulmonary arterial hypertension fibrosis using wide-field second harmonic generation microscopy // Scientific reports. 2022. Vol. 12. N. 1. P. 7330. DOI: 10.1038/s41598-022-11473-5.
  9. Wilson D.W., Segall H.J., Pan L.C. et al. Mechanisms and pathology of monocrotaline pulmonary toxi­city // Crit. Rev. Toxicol. 1992. Vol. 22. P. 307–325. DOI: 10.3109/10408449209146311.
  10. Bøyum A. Isolation of lymphocytes, granulocytes and macrophages // Scand. J. Immunol. 1976. Suppl. 5. P. 9–15.
  11. Kavalenka A.I., Semenkova G.N., Cherenkevich S.N. Effects of hydrogen peroxide on neutrophil ability to ge­nerate reactive oxygen and chlorine species and to secrete myeloperoxidase in vitro // Cell Tissue Biol. 2007. Vol. 1. N. 6. P. 551–559.
  12. Patel A., Hirst R.A., Harrison C. et al. Measurement of [Ca2+] i in whole cell suspensions using fura-2 // Methods Mol. Biol. 2013. Vol. 937. P. 37–47. DOI: 10.1007/978-1-62703-086-1_2.
  13. Рощупкин Д.И., Белакина Н.С., Мурина М.А. Усиленная люминолом хемилюминесценция полиморфноядерных лейкоцитов кролика: природа оксидантов, непосредственно вызывающих окисление люминола // Биофизика. 2006. Т. 51. № 1. С. 99–107. [Roshchupkin D.I., Belakina N. S., Murina M.A. Luminol-enhanced chemiluminescence of rabbit polymorphonuc­lear leukocytes: the nature of oxidants directly responsible for luminol oxidation // Biophys. 2006. Vol. 51. N. 1. P. 99–107. (In Russ.)].
  14. Li Y., Zhu H., Kuppusamy P. Roubaud V., Zweier J.L., Trush M.A. Validation of lucigenin (bis-N-methylacri­dinium) as a chemilumigenic probe for detecting superoxide anion radical production by enzymatic and cellular systems // J. Biol. Chem. 1998. Vol. 273. N. 4. P. 2015–2023. DOI: 10.1074/jbc.273.4.2015.
  15. Immler R., Simon S.I., Sperandio M. Calcium signalling and related ion channels in neutrophil recruitment and function // Eur. J. Clin. Invest. 2018. Vol. 48. P. e12964. DOI: 10.1111/eci.12964.

Received: 2023-10-16
Reviewed: 2023-11-01
Accepted for publication: 2023-11-09

You may be interested