3 . 2022

Phenotypic and functional characteristic analysis of THP-1 cell line as a model of infl ammation

Abstract

Introduction. THP-1 cell line represents one of the common research models for monocyte/macrophage-mediated inflammation.

Aim – to study the properties of THP-1 monocytes activated with bacterial lipopolysaccharides (LPS), influenza virus strain A/Puerto Rico/8/34 (H1N1), and X-ray irradiation.

Material and methods. The cell phenotypic and functional profiles were studied with light microscopy, RT-PCR with reverse transcription, and flow cytometry.

Results. We determined that HSP90, HPRT, and GAPDH were suitable reference genes for mRNA expression normalization upon LPS, viral, or irradiation exposure, respectively. The gene metabolic markers of THP-1 cell responses to LPS (GAPDH), virus (HSP90, GAPDH), and irradiation (B2M) were determined suggesting the new targets for monocyte profiling in pathological inflammation. Finally, the comparison of pro-inflammatory cytokines and antigen-presenting activities of THP-1 cells showed the differential cytokine expression upon various stimuli.

Conclusion. The profiles of metabolic and functional activities can be further used for THP-1 and monocyte comparative analysis and confirm the utility of THP-1 cell line as a model of inflammation.

Keywords:THP-1; monocytes; inflammation; virus; radiotherapy; infection; housekeeping genes

For citation: Kokinos Е.K., Kuzmina D.О., Kuchur О.А., Tsymbal S.А., Vasilichin V.A., Galochkina A.V., Zavirskyi A.V., Basharin V.A., Shtro А.А., Shtil А.А., Dukhinova М.S. Phenotypic and functional characteristic analysis of THP-1 cell line as a model of inflammation. Immunologiya. 2022; 43 (3): 277–87. DOI: https://doi.org/10.33029/0206-4952-2022-43-3-277-287 (in Russian)

Funding. This work was supported by the Russian Science Foundation (Grant No. 20-75-10112), Russian Foundation of Basic Research (Grant No. 19-315-60012).

Conflict of interests. Authors declare no conflict of interests.

Authors’ contribution. Study concept and design – Dukhinova M.S.; data generation and analysis – Kokinos Е.К., Kuzmina D.О., Kuchur О.А., Tsymbal S.A., Vasilichin V.A., Galochkina A.V., Zavirskyi A.V.; statistical analysis – Kokinos Е.К., Kuzmina D.О., Kuchur О.А., Vasilichin V.A., Tsymbal S.A.; text writing – Kokinos Е.К., Kuchur О.А., Tsymbal S.A.; text editing – Basharin V.A., Shtro А.А., Shtil А.А., Dukhinova М.S.

References

1. Budikhina A.S., Murugina N.E., Maksimchik P.V., Dagil’ Yu.A., Nikolaeva A.M., Balyasova L.S., Murugin V.V., Chkadua G.Z., Pinegin B.V., Pashchenkov M.V. On the role of glycolysis in pro-inflammatory cytokine production by macrophages. Immunologiya. 2019; 40 (5): 11–22. DOI: https://doi.org/10.24411/0206-4952-2019-15002 (in Russian)

2. Gan X., Wang H., Yu Y., Yi W., Zhu S., Li E., et al. Epigenetically repressing human cytomegalovirus lytic infection and reactivation from latency in THP-1 model by targeting H3K9 and H3K27 histone demethylases. PLoS One. 2017; 12 (4): e0175390. DOI: https://doi.org/10.1371 /journal.pone.0175390

3. Barcia-Macay M., Seral C., Mingeot-Leclercq M.P., Tulkens P.M., Van Bambeke F. Pharmacodynamic evaluation of the intracellular activities of antibiotics against Staphylococcus aureus in a model of THP-1 macrophages. Antimicrob. Agents Chemother. 2006; 50 (3): 841–51. DOI: https://doi.org/10.1128/AAC.50.3.841-851.2006

4. Chanput W., Mes J.J., Wichers H.J. THP-1 cell line: an in vitro cell model for immune modulation approach. Int. Immunopharmacol. 2014; 23 (1): 37–45. DOI: https://doi.org/10.1016/j.intimp.2014.08.002

5. Caras I., Tucureanu C., Lerescu L., Pitica R., Melinceanu L., Neagu S., et al. Influence of tumor cell culture supernatants on macrophage functional polarization: In vitro models of macrophage-tumor environment interaction. Tumori. 2011; 97 (5): 647–54. DOI: https://doi.org/10.1177/030089161109700518

6. Gudima G.O., Khaitov R.M., Kudlay D.A., Khaitov M.R. Molecular immunological aspects of diagnostics, prevention and treatment of coronavirus infection. Immunologiya. 2021; 42 (3): 198–210. DOI: https://doi.org/10.33029/0206-4952-2021-42-3-198-210 (in Russian)

7. Dukhinova M., Kokinos E., Kuchur P., Komissarov A., Shtro A. Macrophage-derived cytokines in pneumonia: linking cellular immunology and genetics. Cytokine Growth Factor Rev. 2021; 59: 46–61. DOI: https://doi.org/10.1016/j.cytogfr.2020.11.003

8. Sun L., Li X., Xu M., Yang F., Wang W., Niu X. In vitro immunomodulation of magnesium on monocytic cell toward anti-inflammatory macrophages. Regen. Biomater. 2020; 7 (4): 391–401. DOI: https://doi.org/10.1093/rb/rbaa010

9. Medvedeva G.F., Kuzmina D.O., Nuzhina J., Shtil A.A., Dukhinova M.S. How macrophages become transcriptionally dysregulated: a hidden impact of antitumor therapy. Int. J. Mol. Sci. 2021; 22 (5): 2662. DOI: https://doi.org/10.3390/ijms22052662

10. Frey B., Hehlgans S., Rödel F., Gaipl U.S. Modulation of inflammation by low and high doses of ionizing radiation: implications for benign and malign diseases. Cancer Lett. 2015; 368 (2): 230–7. DOI: https://doi.org/10.1016/j.canlet.2015.04.010

11. Di Maggio F.M., Minafra L., Forte G.I., Cammarata F.P., Lio D., Messa C., et al. Portrait of inflammatory response to ionizing radiation treatment. J. Inflamm. (United Kingdom). 2015; 12 (1): 1–11. DOI: https://doi.org/10.1186/s12950-015-0058-3

12. Wan J., Shan Y., Fan Y., Fan C., Chen S., Sun J., et al. NFB inhibition attenuates LPS-induced TLR4 activation in monocyte cells. Mol. Med. Rep. 2016; 14 (5): 4505–10. DOI: https://doi.org/10.3892/mmr. 2016.5825

13. TFBS – Home. URL: http://tfbsdb.systemsbiology.net/ (date of access May 21, 2021)

14. Moniruzzaman M., Ghosal I., Das D., Chakraborty S.B. Melatonin ameliorates H2O2-induced oxidative stress through modulation of Erk/Akt/NFkB pathway. Biol. Res. 2018; 51 (1): 17. DOI: https://doi.org/10.1186/s40659-018-0168-5

15. Prodromou C. Regulatory mechanisms of Hsp90. Biochem. Mol. Biol. J. 2017; 3 (1): 2. DOI: https://doi.org/10.21767/2471-8084.100030

16. Yan M., Hou M., Liu J., Zhang S., Liu B., Wu X., et al. Regulation of iNOS-derived ROS generation by HSP90 and Cav-1 in porcine reproductive and respiratory syndrome virus-infected swine lung injury. Inflammation. 2017; 40 (4): 1236–44. DOI: https://doi.org/10.1007/s10753-017-0566-9

17. Zhang C., Yang Y., Zhou X., Yang Z., Liu X., Cao Z., et al. The NS1 protein of influenza a virus interacts with heat shock protein Hsp90 in human alveolar basal epithelial cells: Implication for virus-induced apoptosis. Virol. J. 2011; 8 (1): 1–9. DOI: https://doi.org/10.1186/1743-422X-8-181

18. Cline T.D., Beck D., Bianchini E. Influenza virus replication in macrophages: Balancing protection and pathogenesis. J. Gen. Virol. 2017; 98 (10): 2401–12. DOI: https://doi.org/10.1099/jgv.0.000922

19. Holm C.K., Rahbek S.H., Gad H.H., Bak R.O., Jakobsen M.R., Jiang Z., et al. Influenza A virus targets a cGAS-independent STING pathway that controls enveloped RNA viruses. Nat. Commun. 2016; 7 (1): 1–9. DOI: https://doi.org/10.1038/ncomms10680

20. Menachery V.D., Schäfer A., Burnum-Johnson K.E., Mitchell H.D., Eisfeld A.J., Walters K.B., et al. MERS-CoV and H5N1 influenza virus antagonize antigen presentation by altering the epigenetic landscape. Proc. Natl Acad. Sci. USA. 2018; 115 (5): E1012–21. DOI: https://doi.org/10.1073/pnas.1706928115

21. Tanaka A., To J., O’Brien B., Donnelly S., Lund M. Selection of reliable reference genes for the normalisation of gene expression levels following time course LPS stimulation of murine bone marrow derived macrophages. BMC Immunol. 2017; 18 (1): 43. DOI: https://doi.org/10.1186/s12865-017-0223-y

22. Liu L., Zhou J., Wang Y., Mason R.J., Funk C.J., Du Y. Proteome alterations in primary human alveolar macrophages in response to influenza A virus infection. J. Proteome Res. 2012; 11 (8): 4091–101. DOI: https://doi.org/10.1021/pr3001332

23. Qin Y., Yu M., Zhou L., Jiang L., Huang M. Durable response to combination radiotherapy and immunotherapy in EP-resistant lung large-cell neuroendocrine carcinoma with B2M and STK11 mutations: a case report. Immunotherapy. 2020; 12 (4): 223–7. DOI: https://doi.org/10.2217/imt-2019-0166

24. Nomura T., Huang W.C., Zhau H., Josson S., Mimata H., Chung L. B2-microglobulin-mediated signaling as a target for cancer therapy. Anticancer Agents Med. Chem. 2014; 14 (3): 343–52. DOI: https://doi.org/10.2174/18715206113139990092

25. Reits E.A., Hodge J.W., Herberts C.A., Groothuis T.A., Chakraborty M., Wansley E.K., et al. Radiation modulates the peptide repertoire, enhances MHC class I expression, and induces successful antitumor immunotherapy. J. Exp. Med. 2006; 203 (5): 1259–71. DOI: https://doi.org/10.1084/jem.20052494

26. Biondillo D.E., Konicek S.A., Iwamoto G.K. Interferon-γ regulation of interleukin 6 in monocytic cells. Am. J. Physiol. Lung Cell. Mol. Physiol. 1994; 267 (5): 564–8. DOI: https://doi.org/10.1152/ajplung.1994.267.5.l564

27. Groves A.M., Johnston C.J., Williams J.P., Finkelstein J.N. Role of infiltrating monocytes in the development of radiation-induced pulmonary fibrosis. Radiat. Res. 2018; 189 (3): 300. DOI: https://doi.org/10.1667/RR14874.1

28. Tu M.M., Abdel-Hafiz H.A., Jones R.T., Jean A., Hoff K.J., Duex J.E., et al. Inhibition of the CCL2 receptor, CCR2, enhances tumor response to immune checkpoint therapy. Commun. Biol. 2020; 3 (1): 1–12. DOI: https://doi.org/10.1038/s42003-020-01441-y

29. Mikhalkevich N., O’Carroll I.P., Tkavc R., Lund K., Sukumar G., Dalgard C.L., et al. Response of human macrophages to gamma radiation is mediated via expression of endogenous retroviruses. PLoS Pathogens. 2021; 17 (2): e1009305. DOI: https://doi.org/10.1371/JOURNAL.PPAT.1009305

30. Osman A.M., Sun Y., Burns T.C., He L., Kee N., Oliva-Vilarnau N., et al. Radiation triggers a dynamic sequence of transient microglial alterations in juvenile brain. Cell Rep. 2020; 31 (9): 107699. DOI: https://doi.org/10.1016/j.celrep.2020.107699

All articles in our journal are distributed under the Creative Commons Attribution 4.0 International License (CC BY 4.0 license)

JOURNALS of «GEOTAR-Media»