Anti-dengue activity of synthetic peptides increases antiviral interferon-beta genes in LLC-MK2 cells

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Jundee Rabablert
Promsin Masrinoul
Sutee Yoksan
Supoth Rajakam
Supanyika Sengsai
Korakot Changirakul
Prayad Sangngam
Sunee Techaarpornkul
Sittiruk Roytrakul


Dengue virus (DV) causes dengue diseases in human via the Aedes mosquitoes. Fatal cases of dengue hemorrhagic fevers show a complication of acute kidney injury. Previous reports revealed that synthetic DV2(413-440), DV2(413-447), and DV2(419-447) peptides inhibit the dengue envelope proteins in Aedes albopictus C6/36 cells, but nothing is known about the effect of these peptides on mammalian cells in relation to adaptive and innate immunities. In this study, the anti-dengue activity of these synthetic peptides in Rhesus monkey kidney LLC-MK2 cells was investigated at the cellular and molecular levels. Moreover, the effects of these synthetic peptides on apoptotic caspase-10, pro-inflammatory interleukin-1beta, tumor necrosis factor-alpha, and antiviral interferon-beta genes in LLC-MK2 cells were also sought. The results revealed the maximum non-toxic doses of DV2 (413-440), DV2 (413-447), and DV2 (419-447) in LLC-MK2 cells, with values of 77.88± 0.52, 47.01±1.32, and 63.82±1.97 µM, respectively. At 25 µM concentration, synthetic DV2 (413-447) and DV2 (419-447) peptides showed 100% plaque inhibition in simultaneous treatment. By contrast, these peptides showed ≤58% plaque inhibition in pre- and post-treatment at 7 days post-incubation. These synthetic peptides also inhibited Dengue 2 virus, apoptotic caspase-10, pro-inflammatory tumor necrosis factor-alpha, and interleukin-1beta genes. On the other hand, these peptides upregulated the antiviral interferon-beta gene in innate immunity. This study is the first report to reveal the anti-dengue activity of synthetic DV2 (413-440), DV2 (413-447), and DV2 (419-447) peptides in adaptive and innate immunity.


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Akash, M. S. H., Rehman, K., and Liaqat, A. (2018). Tumor necrosis factor-alpha: role in development of insulin resistance and pathogenesis of type 2 diabetes mellitus. Journal of Cellular Biochemistry, 119(1), 105-110.

Akey, D. L., Brown, W. C., Dutta, S., Konwerski, J., Jose, J., Jurkiw, T. J., DelProposto, J., Ogata, C. M., Skiniotis, G., Kuhn, R. J., and Smith, J. L. (2014). Flavivirus NS1 structures reveal surfaces for associations with membranes and the immune system. Science, 343(6173), 881-885.

Bethell, D. B., Flobbe, K., Cao, X. T., Day, N. P., Pham, T. P., Buurman, W. A., Cardosa, M. J., White, N. J., and Kwiatkowski, D. (1998). Pathophysiologic and prognostic role of cytokines in dengue hemorrhagic fever. Journal of Infectious Diseases, 177(3), 778-782.

Chen, H. C., Hofman, F. M., Kung, J. T., Lin, Y. D., and Wu-Hsieh, B. A. (2007). Both virus and tumor necrosis factor alpha are critical for endothelium damage in a mouse model of dengue virus induced hemorrhage. Journal of Virology, 81(11), 5518-5526.

Chew, M. F., Poh, K. S., and Poh, C. L. (2017). Peptides as therapeutic agents for dengue virus. International Journal of Medical Sciences, 14(13), 1342-1359.

Clark, K. B., Onlamoon, N., Hsiao, H. M., Perng, G. C., and Villinger, F. (2013). Can non-human primates serve as models for investigating dengue disease pathogenesis? Frontiers in Microbiology, 4, 305.

Conceição, T. M., El-Bacha, T., Villas-Bôas, C. S. A., Coello, G., Ramirez, J., Montero-Lomeli, M., and Poian, A. T. D. (2010). Gene expression analysis during dengue virus infection in HepG2 cells reveals virus control of innate immune response. Journal of Infection, 60(1), 65-75.

Diamond, M. S., Roberts, T. G., Edgil, D., Lu, B., Ernst J., and Harris, E. (2000). Modulation of dengue virus Infection in human cells by alpha, beta, and gamma interferons. Journal of Virology, 74(11), 4957-4966.

Dinarello, C. A., Gatti, S., and Bartfai, T. (1999). Fever: Links with an ancient receptor. Current Biology, 9(4), R147- R150.

Diptyanusa, A., Phumratanaprapin, W., Phonrat, B., Poovorawan, K., Hanboonkunupakarn, B., Sriboonvorakul, N., and Thisyakorn, U. (2019). Characteristics and associated factors of acute kidney injury among adult dengue patients: A retrospective single-center study. PloS One, 14(1), e0210360.

Fernandez-Mestre, M. T., Gendzekhadze, K., Rivas-Vetencourt, P., and Layrisse, Z. (2004). TNF-α-308A allele, a possible severity risk factor of hemorrhagic manifestation in dengue fever patients. Tissue Antigens, 64(4), 469-472.

Huang, Y. W., Lee, C. T., Wang, T. C., Kao, Y. C., Yang, C. H., Lin, Y. M., and Huang, K. S. (2018). The development of peptide-based antimicrobial agents against dengue virus. Current Protein and Peptide Science, 19(10), 998-1010.

Iglesias, N. G., Filomatori, C. V., and Gamarnik, A. V. (2011). The F1 motif of dengue virus polymerase NS5 is involved in promoter-dependent RNA synthesis. Journal of Virology, 85(12), 5745-5756.

Jeewandara, C., Gomes, L., Wickramasinghe, N., Gutowska-Owsiak, D., Waithe, D., Paranavitane, S. A., Shyamali, N. L. A., Ogg, G. S., and Malavige, G. N. (2015). Platelet activating factor contributes to vascular leak in acute dengue infection. PLoS Neglected Tropical Diseases, 9(2), e0003459.

Kittigul, L., Temprom, W., Sujirarat, D., and Kittigul, C. (2000). Determination of tumor necrosis factor-alpha levels in dengue virus infected patients by sensitive biotin-streptavidin enzyme-linked immunosorbent assay. Journal of Virological Methods, 90(1), 51-57.

Lee, I. K., Liu, J. W., and Yang, K. D. (2009). Clinical characteristics, risk factors, and outcomes in adults experiencing dengue hemorrhagic fever complicated with acute renal failure. American Journal of Tropical Medicine and Hygiene, 80(4), 651-655.

Liu, T., Tang, L., Tang, H., Pu, J., Gong, S., Fang, D., Zhang, H., Li, Y-P, Zhu, X., Wang, W., Wu, M., Liao, Y., Li, C., Zhou, H., and Huang, X. (2019). Zika virus infection induces acute kidney injury through activating NLRP3 inflammasome via suppressing Bcl-2. Frontiers in Immunology, 10, 1925.

Lizarraga, K. J., and Nayer, A. (2014). Dengue-associated kidney disease. Journal of Nephropathology, 3(2), 57-62.

Man, S. M., and Kanneganti, T. D. (2016). Converging roles of caspases in inflammasome activation, cell death and innate immunity. Nature Reviews Immunology, 16(1), 7-21.

Martínez-Betancur, V., and Martínez-Gutierrez, M. (2016). Proteomic profile of human monocytic cells infected with dengue virus. Asian Pacific Journal of Tropical Biomedicine, 6(11), 914-923.

Masood, K. I., Jamil, B., Rahim, M., Islam, M., Farhan, M., and Hasen, Z. (2018). Role of TNF α, IL-6 and CXCL10 in Dengue disease severity. Iranian Journal of Microbiology, 10(3), 202-207.

Mehaffey, E., and Majid, D. S. A. (2017). Tumor necrosis factor-α, kidney function, and hypertension. American Journal of Physiology-Renal Physiology, 313(4), F1005-F1008.

Mladinich, K. M., Piaskowski, S. M., Rudersdorf, R., Eernisse, C. M., Weisgrau, K. L., Martins, M. A., Furlott, J. R., Partidos, C. D., Brewoo, J. N., Osorio, J. E., Wilson, N. A., Rakasz, E. G., and Watkins, D. I. (2012). Dengue virus-specific CD4+ and CD8+ T lymphocytes target NS1, NS3 and NS5 in infected Indian rhesus macaques. Immunogenetics, 64(2), 111-121.

Mohr, A., Deedigan L., Jencz, S., Mehrabadi, Y., Houlden, L., Albarenque, S. M., and Zwacka, R. M. (2018). Caspase-10: a molecular switch from cell-autonomous apoptosis to communal cell death in response to chemotherapeutic drug treatment. Cell Death and Differentiation, 25(2), 340-352.

Monterio, M. J. C., Oliveira, M. D., Dias, R. S., Nacif-Marçal, L., Feio, R. N., Ferreira, S. O., Oliveira, L. L., Silva, C. C., and Paula, S. O. (2018). The antimicrobial peptide HS-1 inhibits dengue virus infection. Virology, 514, 79-87.

Morrison, J., and García-Sastre, A. (2014). STAT2 signaling and dengue virus infection. JAK-STAT, 3(1), e27715.

Munoz-Jordan, J. L. (2010). Subversion of interferon by dengue virus. Current Topics in Microbiology and Immunology, 338, 35-44.

Nitsche, C., Holloway, S., Schirmeister, T., and Klein, C. D. (2014). Biochemistry and medicinal chemistry of the dengue virus protease. Chemical Reviews, 114(22), 11348-11381.

Onlamoon, N., Noisakran, S., Hsiao, H. M., Duncan, A., Villinger, F., Ansari, A. A., and Perng, G. C. (2010). Dengue virus-induced hemorrhage in a nonhuman primate model. Blood, 115(9), 1823-1834.

Pan, P., Zhang, Q., Liu, W., Wang, W., Yu, Z., Lao, Z., and Zhang, W., Shen, M., Wan, P., Xiao, F., Shereen, M. A., Zhang, W., Tan, Q., Liu, Y., Liu, X., Wu, K., Liu, Y., Li, G., and Wu, J. (2019). Dengue virus infection activates Interleukin-1β to induce tissue Injury and vascular leakage. Frontiers in Microbiology, 10, 2637.

Pang, X., Zhang, R., and Cheng, G. (2017). Progress towards understanding the pathogenesis of dengue hemorrhagic fever. Virologica Sinica, 32(1), 16-22.

Pichyangkul, S., Endy, T. P., Kalayanarooj, S., Nisalak, A., Yongvanitchit, K., Green, S., Rothman, A. L., Ennis, F. A., and Libraty, D. H. (2003). A blunted blood plasmacytoid dendritic cell response to an acute systemic viral infection is associated with increased disease severity. Journal of Immunology, 171(10), 5571-5578.

Póvoa, T. F., Oliveira, E. R. A., Basilio-de-Oliveira, C. A., Nuovo, G. J., Chagas, V. L. A., Salomão, N. G., Mota, E. M., and Paes, M. V. (2016). Peripheral organs of dengue fatal cases present strong pro-inflammatory response with participation of IFN-gamma-, TNF-alpha- and RANTES-producing cells. PLoS One, 11(12), e0168973.

Rabablert, J., Rajakam, S., Chaiyo, K., and Yoksan, S. (2020). Attenuation biomarkers of dengue 4 virus strain 1036 PDK40 infectious clone. Southeast Asian Journal of Tropical Medicine and Public Health, 51(1), 103-114.

Rodriguez-Madoz, J. R., Bernal-Rubio, D., Kaminski, D., Boyd, K., and Fernandez-Sesma, A. (2010). Dengue virus inhibits the production of type I interferon in primary human dendritic cells. Journal of Virology, 84(9), 4845-4850.

Schleich, K., Nurnberger, C., Sobanski, A., and Efferth, T. (2011). Vaccination and antiviral treatment of neglected diseases caused by flaviviral infections. Current Medicinal Chemistry, 18(4), 604-614.

Schmidt, A. G., Yang, P. L., and Harrison, S. C. (2010a). Peptide inhibitors of dengue-virus entry target a late-stage fusion intermediate. PLoS Pathogens, 6(4), e1000851.

Schmidt, A. G., Yang, P. L., and Harrison, S. C. (2010b). Peptide inhibitors of flavivirus entry derived from the E protein stem. Journal of Virology, 84(24), 12549-12554.

Seo, Y. J., and Hahm, B. (2010). Type I interferon modulates the battle of host immune system against viruses. Advances in Applied Microbiology, 73, 83-101.

Shresta, S., Kyle, J. L., Snider H. M., Basavapatna, M., Beatty, P. R., and Harris, E. (2004). Interferon-dependent immunity is essential for resistance to primary dengue virus infection in mice, whereas T- and B-cell-dependent immunity are less critical. Journal of Virology, 78(6), 2701-2710.

Shresta, S., Sharar, K. L., Prigozhin, D. M., Snider, H. M., Beatty, P. R., and Harris, E. (2005). Critical roles for both STAT1-dependent and STAT1-independent pathways in the control of primary dengue virus infection in mice. Journal of Immunology, 175(6), 3946-3954.

Shresta, S., Sharar, K. L., Prigozhin, D. M., Beatty, P. R., and Harris, E. (2006). Murine model for dengue virus-induced lethal disease with increased vascular permeability. Journal of Virology, 80(20), 10208-10217.

Shrivastava, G., Visoso-Carvajal, G., Garcia-Corder, J., Leon-Juarez, M., Chavez-Munguia, B., Lopez, T., Nava, P., Villegas-Sepulveda, N., and Cedillo-Barron, L. (2020). Dengue virus serotype 2 and its non-structural proteins 2A and 2B activate NLRP3 inflammasome. Frontiers in Immunology, 11, 352.

Smit, J. M., Moesker B., Rodenhuis-Zybert, I., and Wilschut, J. (2011). Flavivirus cell entry and membrane fusion. Viruses, 3(2), 160-171.

Sydow, F. F., Santiago, M. A., Neves-Souza, P. C., Cerqueira, D. I., Gouvea, A. S., Lavatori, M. F., Bertho, A. L., and Kubelka, C. F. (2000). Comparison of dengue infection in human mononuclear leukocytes with mosquito C6/36 and mammalian Vero cells using flow cytometry to detect virus antigen. Memórias do Instituto Oswaldo Cruz, 95(4), 483-489.

Tan, T. Y., and Chu, J. J. H. (2013). Dengue virus-infected human monocytes trigger late activation of caspase-1, which mediates pro-inflammatory IL-1β secretion and pyroptosis. Journal of General Virology, 94(10), 2215-2220.

Thepparit, C. Khakpoor, A., Khongwichit, S., Wikan, N., Fongsaran, C., Chingsuwanrote, P., Panraksa, P., and Smith, D. R. (2013). Dengue 2 infection of HepG2 liver cells results in endoplasmic reticulum stress and induction of multiple pathways of cell death. BMC Research Notes. 6, 372.

Torrentes-Carvalho, A., Azeredo, E. L., Reis, S. R., Miranda, A. S., Gandini, M., Barbosa, L. S., and Kubelka, C. F. (2009). Dengue-2 infection and the induction of apoptosis in human primary monocytes. Memórias do Instituto Oswaldo Cruz, 104(8), 1091-1099.

Uno, N., and Ross, T. M. (2018). Dengue virus and the host innate immune response. Emerging Microbes and Infections, 7(1), 167.

Wang, J., Chun, H. J., Wong, W., Spencer, D. M., and Lenardo, M. J. (2001). Caspase-10 is an initiator caspase in death receptor signaling. Proceedings of the National Academy of Sciences of the United States of America, 98(24), 13884-13888.

Wang, W., Li, G., Wu, D., Luo, Z., Pan, P., Tian, M., Wang, Y., Xiao, F., Li, A., Wu, K., Liu, X., Rao, L., Liu, F., Liu, Y., and Wu, J. (2018). Zika virus infection induces host inflammatory responses by facilitating NLRP3 inflammasome assembly and interleukin-1β secretion. Nature Communications, 2018(9), 106.

Yen, Y. T., Chen, H. C., Lin, Y. D., Shieh, C. C., and Wu-Hsieh, B. A. (2008). Enhancement by tumor necrosis factor alpha of dengue virus-induced endothelial cell production of reactive nitrogen and oxygen species is key to hemorrhage development. Journal of Virology, 82(24), 12312-12324.

Yoksan, S., Bhamarapravati, N., and Halstead, S. B. (1986). Dengue virus vaccine development: study on biological markers of uncloned dengue 1-4 viruses serially passaged in primary kidney cells. In Arbovirus Research in Australia: Proceedings 4th Symposium (George, T. D., Kay, B. H., Block, J., eds.), pp. 35-38. Brisbane: CSIRO Division of Tropical Animal Science.

Yoksan, S., Rabablert, J., Chaiyo, K., Rajchakam, S., Tiewcharoen, S., Rabablert, N., Kerdkriangkrai, S., Samngamnim, N., Phurttikul, W., and Luangboribun, T. (2013). Cytokine gene expression in human hepatocytes infected with dengue virus serotype 3 (strain-16562). Health, 5(9), 1516-1525.

Yoksan, S., Chaiyo, K., Rajakam, S., Kerdkriangkrai, S., Khawsithiwong, P., and Rabablert, J. (2018). Molecular epidemiology of dengue viruses isolated from patients with suspected dengue fever in Bangkok, Thailand during 2006-2015. Southeast Asian Journal of Tropical Medicine and Public Health, 49(4), 604-616.