Establishment and Characterization of an Immortalized Red River Hog Blood-derived Macrophage Cell Line

Overview

Journal Front Immunol

Specialty Allergy & Immunology

Date 2024 Sep 26

PMID 39324137

Authors

Takato Takenouchi

Kentaro Masujin

Rina Ikeda

Seiki Haraguchi

Shunichi Suzuki

Hirohide Uenishi

Eiji Onda

Takehiro Kokuho

Affiliations

Soon will be listed here.

Abstract

Red river hogs (RRHs) (), a wild species of living in Africa with a major distribution in the Guinean and Congolian forests, are natural reservoirs of African swine fever virus (ASFV) and typically are asymptomatic. Since blood and tissue macrophages of suid animals are target cell lineages of ASFV, RRH-derived macrophages are expected to play an important role in suppressing disease development in infected individuals. In the present study, we successfully isolated RRH-derived blood macrophages using co-culture techniques of RRH blood cells with porcine kidney-derived feeder cells and immortalized them by transferring SV40 large T antigen and porcine telomerase reverse transcriptase genes. The newly established macrophage cell line of the RRH-derived blood cell origin (RZJ/IBM) exhibited an Iba1-, CD172a-, and CD203a-positive typical macrophage-like phenotype and up-regulated the phosphorylation of nuclear factor-κB p65 subunit and p38 mitogen-activated protein kinase in response to the bacterial cell wall components, lipopolysaccharide (LPS) and muramyl dipeptide. In addition, RZJ/IBM cells produced the precursor form of interleukin (IL)-1β and IL-18 upon a stimulation with LPS, leading to the conversion of IL-18, but not IL-1β, into the mature form. Time-lapse live cell imaging with pHrodo dye-conjugated BioParticles demonstrated the phagocytotic activity of RZJ/IBM cells. It is important to note that RZJ/IBM cells are clearly susceptible to ASFV infection and support viral replication . Therefore, the RZJ/IBM cell line provides a unique model for investigating the pathogenesis of ASFV.

References

Takenouchi T, Masujin K, Miyazaki A, Suzuki S, Takagi M, Kokuho T . Isolation and immortalization of macrophages derived from fetal porcine small intestine and their susceptibility to porcine viral pathogen infections. Front Vet Sci. 2022; 9:919077. PMC: 9339801. DOI: 10.3389/fvets.2022.919077. View

Kaplanski G . Interleukin-18: Biological properties and role in disease pathogenesis. Immunol Rev. 2017; 281(1):138-153. PMC: 7165732. DOI: 10.1111/imr.12616. View

Masujin K, Kitamura T, Kameyama K, Okadera K, Nishi T, Takenouchi T . An immortalized porcine macrophage cell line competent for the isolation of African swine fever virus. Sci Rep. 2021; 11(1):4759. PMC: 7910288. DOI: 10.1038/s41598-021-84237-2. View

Takenouchi T, Suzuki S, Shinkai H, Tsukimoto M, Sato M, Uenishi H . Extracellular ATP does not induce P2X7 receptor-dependent responses in cultured renal- and liver-derived swine macrophages. Results Immunol. 2014; 4:62-7. PMC: 4213840. DOI: 10.1016/j.rinim.2014.07.002. View

Alvarez B, Revilla C, Poderoso T, Ezquerra A, Dominguez J . Porcine Macrophage Markers and Populations: An Update. Cells. 2023; 12(16). PMC: 10453229. DOI: 10.3390/cells12162103. View

Gaudreault N, Madden D, Wilson W, Trujillo J, Richt J . African Swine Fever Virus: An Emerging DNA Arbovirus. Front Vet Sci. 2020; 7:215. PMC: 7237725. DOI: 10.3389/fvets.2020.00215. View

Tschopp J, Schroder K . NLRP3 inflammasome activation: The convergence of multiple signalling pathways on ROS production?. Nat Rev Immunol. 2010; 10(3):210-5. DOI: 10.1038/nri2725. View

Hu J, Liu K, Liu J, Jiang X, Wang X, Chen Y . CD163 as a marker of M2 macrophage, contribute to predicte aggressiveness and prognosis of Kazakh esophageal squamous cell carcinoma. Oncotarget. 2017; 8(13):21526-21538. PMC: 5400603. DOI: 10.18632/oncotarget.15630. View

Gordon S, Pluddemann A . Tissue macrophages: heterogeneity and functions. BMC Biol. 2017; 15(1):53. PMC: 5492929. DOI: 10.1186/s12915-017-0392-4. View

10.

Kitani H, Yoshioka M, Takenouchi T, Sato M, Yamanaka N . Characterization of the liver-macrophages isolated from a mixed primary culture of neonatal swine hepatocytes. Results Immunol. 2014; 4:1-7. PMC: 3973824. DOI: 10.1016/j.rinim.2014.01.001. View

11.

Carrascosa A, Bustos M, de Leon P . Methods for growing and titrating African swine fever virus: field and laboratory samples. Curr Protoc Cell Biol. 2011; Chapter 26:26.14.1-26.14.25. DOI: 10.1002/0471143030.cb2614s53. View

12.

Takenouchi T, Masujin K, Suzuki S, Haraguchi S, Hiramatsu K, Kokuho T . Establishment and characterization of the immortalized porcine lung-derived mononuclear phagocyte cell line. Front Vet Sci. 2022; 9:1058124. PMC: 9715978. DOI: 10.3389/fvets.2022.1058124. View

13.

Finlay B, McFadden G . Anti-immunology: evasion of the host immune system by bacterial and viral pathogens. Cell. 2006; 124(4):767-82. DOI: 10.1016/j.cell.2006.01.034. View

14.

Borca M, Rai A, Espinoza N, Ramirez-Medina E, Spinard E, Velazquez-Salinas L . African Swine Fever Vaccine Candidate ASFV-G-ΔI177L Produced in the Swine Macrophage-Derived Cell Line IPKM Remains Genetically Stable and Protective against Homologous Virulent Challenge. Viruses. 2023; 15(10). PMC: 10612016. DOI: 10.3390/v15102064. View

15.

Takenouchi T, Kitani H, Suzuki S, Nakai M, Fuchimoto D, Tsukimoto M . Immortalization and Characterization of Porcine Macrophages That Had Been Transduced with Lentiviral Vectors Encoding the SV40 Large T Antigen and Porcine Telomerase Reverse Transcriptase. Front Vet Sci. 2017; 4:132. PMC: 5566601. DOI: 10.3389/fvets.2017.00132. View

16.

Keller M, Ruegg A, Werner S, Beer H . Active caspase-1 is a regulator of unconventional protein secretion. Cell. 2008; 132(5):818-31. DOI: 10.1016/j.cell.2007.12.040. View

17.

Duan T, Du Y, Xing C, Wang H, Wang R . Toll-Like Receptor Signaling and Its Role in Cell-Mediated Immunity. Front Immunol. 2022; 13:812774. PMC: 8927970. DOI: 10.3389/fimmu.2022.812774. View

18.

Takenouchi T, Morozumi T, Wada E, Suzuki S, Nishiyama Y, Sukegawa S . Dexamethasone enhances CD163 expression in porcine IPKM immortalized macrophages. In Vitro Cell Dev Biol Anim. 2021; 57(1):10-16. PMC: 7862206. DOI: 10.1007/s11626-020-00535-5. View

19.

Gomez-Villamandos J, Bautista M, Sanchez-Cordon P, Carrasco L . Pathology of African swine fever: the role of monocyte-macrophage. Virus Res. 2013; 173(1):140-9. DOI: 10.1016/j.virusres.2013.01.017. View

20.

Stefanovic S, Windsor M, Nagata K, Inagaki M, Wileman T . Vimentin rearrangement during African swine fever virus infection involves retrograde transport along microtubules and phosphorylation of vimentin by calcium calmodulin kinase II. J Virol. 2005; 79(18):11766-75. PMC: 1212593. DOI: 10.1128/JVI.79.18.11766-11775.2005. View