Antiviral Activity of Recombinant Porcine Interferon-α Against Porcine Transmissible Gastroenteritis Virus in PK-15 Cells

Bir rekombinant domuz interferon alfa (rPoIFN-?) geliştirilmiş ve patenti daha öncesinde alınmıştı (Çin patent numarası ZL200810020180.4). Bu çalışmada, rPoIFN-?'nın farlı dozlarının domuz transmissible gastroenteritis virüs (TGEv)'ün üremesi üzerindeki baskılayıcı etkisi domuz böbrek hücre kültüründe (PK-15) araştırıldı. rPoIFN-? ile viral büyümenin baskılanmasını kantitatif olarak belirlemek için TCID50 testi, plak oluşum testi, gerçek zamanlı qRT-PCR, western blot ve immunfloresan teknikleri viral enfeksiyöz partiküllerin değişimlerini, viral genom kopya sayılarını ve viral protein ekspresyon seviyelerini belirlemek amacıyla uygulandı. Araştırma sonuçları test edilen üç rPoIFN-?'nın da PK-15 hücrelerinde TGEv ile oluşturulmuş sitopatik etkisinin aynı derecede olduğunu göstermiştir. rPoIFN-?, TGEv proliferasyonunu insan IFN-? ürününden daha güçlü olarak inhibe etti. Kültürde TGEv büyümesine rPoIFN-?'nın baskılayıcı aktivitesi doza bağımlı olup aktivite rPoIFN-?'nın azalan dozu ile göreceli olarak azalma gösterdi

PK-15 Hücrelerinde Domuz Transmissible Gastroenteritis Virüsüne Karşı Rekombinant Domuz İnterferon-α’nın Antiviral Aktivitesi

A recombinant porcine interferon alpha (rPoIFN-α) has been developed and patented previously (Chinese patent number ZL200810020180.4). In the current study, we investigated the inhibitory effects of the rPoIFN-α on the propagation of porcine transmissible gastroenteritis virus (TGEv) at different doses in porcine kidney cell line (PK-15). To quantitatively determine the inhibition of viral growth by rPoIFN-α, TCID50 assay, plaque formation assay, real-time qRT-PCR, western blot and immunofluorescence assay were adopted to evaluate the changes of viral infectious particles, viral genome copy numbers and viral protein expression levels respectively. The results demonstrated that all the three batches of the rPoIFN-α tested inhibited TGEvinduced cytopathic effect in PK-15 cells with very similar potency. rPoIFN-α inhibited TGEv proliferation more strongly than human IFN-α product. The inhibitory activity of rPoIFN-α on TGEv growth in culture was dose dependent, and the activity was gradually reduced with the decreasing of the concentration of rPoIFN-α

PDF

___

1. Jansson AM: Structure of alphacoronavirus transmissible gastroenteritis virus nsp1 has implications for coronavirus nsp1 function and evolution. J Virol, 87, 2949-2955, 2013. DOI: 10.1128/JVI.03163-12
2. Lee BM, Han YW, Kim SB, Rahman MM, Uyangaa E, Kim JH, Roh YS, Kim B, Han SB, Hong JT, Kim K, Eo SK: Enhanced protection against infection with transmissible gastroenteritis virus in piglets by oral co-administration of live attenuated Salmonella enterica serovar Typhimurium expressing swine interferon-α and interleukin-18. Comp Immunol Microbiol Infect Dis, 34, 369-380, 2011. DOI: 10.1016/j. cimid.2011.05.001
3. Kowdley KV: Hematologic side effects of interferon and ribavirin therapy. J Clin Gastroenterol, 39, S3-S8, 2005. DOI: 10.1097/01.mcg. 0000145494.76305.11
4. Amsler L, Verweij MC, DeFilippis VR: The tiers and dimensions of evasion of the type I interferon response by human cytomegalovirus. J Mol Biol, 425, 4857-4871, 2013. DOI: 10.1016/j.jmb.2013.08.023
5. García-Sastre A, Biron CA: Type 1 interferons and the virus-host relationship: A lesson in détente. Science, 312, 879-882, 2006. DOI: 10.1126/science.1125676
6. Liu SY, Sanchez DJ, Aliyari R, Lu S, Cheng G: Systematic identification of type I and type II interferon-induced antiviral factors. Proc Natl Acad Sci USA, 109, 4239-4244, 2012. DOI: 10.1073/pnas.1114981109
7. Diaz-San Segundo F, Moraes MP, de Los Santos T, Dias CC, Grubman MJ: Interferon-induced protection against foot-and-mouth disease virus infection correlates with enhanced tissue-specific innate immune cell infiltration and interferon-stimulated gene expression. J Virol, 84, 2063- 2077, 2010. DOI: 10.1128/JVI.01874-09
8. Nfon CK, Ferman GS, Toka FN, Gregg DA, Golde WT: Interferon-alpha production by swine dendritic cells is inhibited during acute infection with foot-and-mouth disease virus. Viral Immunol, 21, 68-77, 2008. DOI: 10.1089/vim.2007.0097
9. Xiong Y, Lin M, Yuan B, Yuan T, Zheng C: Expression of exogenous IFN-alpha by bypassing the translation block protects cells against FMDV infection. Antiviral Res, 84, 60-66, 2009. DOI: 10.1016/j.antiviral.2009.07.004
10. Buddaert W, Van Reeth K, Pensaert M: In vivo and in vitro interferon (IFN) studies with the porcine reproductive and respiratory syndrome virus (PRRSV). Adv Exp Med Biol, 440, 461-467, 1998.
11. Seya T, Kasamatsu J, Azuma M, Shime H, Matsumoto M: Natural killer cell activation secondary to innate pattern sensing. J Innate Immun, 3, 264-273, 2011. DOI: 10.1159/000326891
12. Pol JM, Broekhuysen-Davies JM, Wagenaar F, La Bonnardière C: The influence of porcine recombinant interferon-alpha 1 on pseudorabies virus infection of porcine nasal mucosa in vitro. J Gen Virol, 72, 933-938, 1991. DOI: 10.1099/0022-1317-72-4-933
13. Horisberger MA: Virus-specific effects of recombinant porcine interferon-gamma and the induction of Mx proteins in pig cells. J Interferon Res, 12, 439-444,1992. DOI: 10.1089/jir.1992.12.439
14. Xia C, Dan W, Wen-Xue W, Jian-Qing W, Li W, Tian-Yao Y, Qin W, Yi-Bao N: Cloning and expression of interferon-alpha/gamma from a domestic porcine breed and its effect on classical swine fever virus. Vet Immunol Immunopathol, 104, 81-89, 2005. DOI: 10.1016/j.vetimm. 2004.10.005
15. Barbé F, Saelens X, Braeckmans D, Lefèvre F, Reeth KV: Role of IFN-alpha during the acute stage of a swine influenza virus infection. Res Vet Sci, 88, 172-178, 2010. DOI: 10.1016/j.rvsc.2009.07.001
16. Osterlund P, Pirhonen J, Ikonen N, Rönkkö E, Strengell M, Mäkelä SM, Broman M, Hamming OJ, Hartmann R, Ziegler T, Julkunen I: Pandemic H1N1 2009 influenza A virus induces weak cytokine responses in human macrophages and dendritic cells and is highly sensitive to the antiviral actions of interferons. J Virol, 84, 1414-1422, 2010. DOI: 10.1128/ JVI.01619-09
17. Woo PC, Tung ET, Chan KH, Lau CC, Lau SK, Yuen KY: Cytokine profiles induced by the novel swine-origin influenza A/H1N1 virus: Implications for treatment strategies. J Infect Dis, 201, 346-353, 2010. DOI: 10.1086/649785
18. Du Y, Dai J, Li Y, Li C, Qi J, Duan S, Jiang P: Immune responses of recombinant adenovirus co-expressing VP1 of foot-and-mouth disease virus and porcine interferon alpha in mice and guinea pigs. Vet Immunol Immunopathol, 124, 274-283, 2008. DOI: 10.1016/j.vetimm.2008.04.011
19. La Bonnardière C, Lefèvre F, Charley B: Interferon response in pigs: Molecular and biological aspects. Vet Immunol Immunopathol, 43, 29- 36, 1994. DOI: 10.1016/0165-2427(94)90117-1
20. Lefèvre F, Mège D, L’Haridon R, Bernard S, De Vaureix C, La Bonnardière C: Contribution of molecular biology to the study of the porcine interferon system. Vet Microbiol, 23, 245-257, 1990. DOI: 10.1016/0378-1135(90)90155-O
21. Lefevre F, La Bonnardiere C: Molecular cloning and sequencing of a gene encoding biologically active porcine alpha-interferon. J Interferon Res, 6, 349-360, 1986. DOI: 10.1089/jir.1986.6.349
22. Lefèvre F, L’Haridon R, Borras-Cuesta F, La Bonnardière C: Production, purification and biological properties of an Escherichia coliderived recombinant porcine alpha interferon. J Gen Virol, 71, 1057-1063, 1990. DOI: 10.1099/0022-1317-71-5-1057
23. Jordan LT, Derbyshire JB: Antiviral action of interferon-alpha against porcine transmissible gastroenteritis virus. Vet Microbiol, 45, 59- 70, 1995. DOI: 10.1016/0378-1135(94)00118-G
24. Weingartl HM, Derbyshire JB: Antiviral activity against transmissible gastroenteritis virus, and cytotoxicity, of natural porcine interferons alpha and beta. Can J Vet Res, 55,143-149, 1991.
25. Reed LI, Muench H: A simple method of estimating fifty percent end points. Am J Hyg, 27, 493-497, 1938.
26. Sachs LA, Schnurr D, Yagi S, Lachowicz-Scroggins ME, Widdicombe JH: Quantitative real-time PCR for rhinovirus, and its use in determining the relationship between TCID50 and the number of viral particles. J Virol Methods, 171, 212-218, 2011. DOI: 10.1016/j.jviromet.2010.10.027
27. Liu K, Liao X, Zhou B, Yao H, Fan S, Chen P, Miao D: Porcine alpha interferon inhibit Japanese encephalitis virus replication by different ISGs in vitro. Res Vet Sci, 95, 950-956, 2013. DOI: 10.1016/j.rvsc.2013.08.008
28. Dash P, Barnett PV, Denyer MS, Jackson T, Stirling CM, Hawes PC, Simpson JL, Monaghan P, Takamatsu HH: Foot-and-mouth disease virus replicates only transiently in well-differentiated porcine nasal epithelial cells. J Virol, 84, 9149-9160, 2010. DOI: 10.1128/JVI.00642-10
29. An K, Fang L, Luo R, Wang D, Xie L, Yang J, Chen H, Xiao S: Quantitative proteomic analysis reveals that transmissible gastroenteritis virus activates the JAK-STAT1 signaling pathway. J Proteome Res, 13, 5376- 5390, 2014. DOI: 10.1021/pr500173p
30. Jordan LT, Derbyshire JB: Antiviral action of interferon-alpha against porcine transmissible gastroenteritis virus. Vet Microbio, 45, 59- 70, 1995. DOI: 10.1016/0378-1135(94)00118-G
31. Lee BM, Han YW, Kim SB, Rahman MM, Uyangaa E, Kim JH, Roh YS, Kim B, Han SB, Hong JT, Kim K, Eo SK: Enhanced protection against infection with transmissible gastroenteritis virus in piglets by oral co-administration of live attenuated Salmonella enterica serovar Typhimurium expressing swine interferon-α and interleukin-18. Comp Immunol Microbiol Infect Dis, 34, 369-380, 2011. DOI: 10.1016/j. cimid.2011.05.001
32. Zhu YP, Xu H, Chen YH, Wang Y, Cen L, Guo XF: Modifications of the porcine interferon-α gene and its antiviral activity in vitro and in vivo. Revue Méd Vét, 162, 143-149, 2011.
33. Weissmann C, Weber H:The interferon genes. Prog Nucleic Acid Res Mol Biol, 33, 251-300, 1986. DOI: 10.1016/S0079-6603(08)60026-4
34. Familletti PC, McCandliss R, Pestka S: Production of high levels of human leukocyte interferon from a continuous human myeloblast cell culture. Antimicrob Agents Chemother, 20, 5-9, 1981. DOI: 10.1128/ AAC.20.1.5
35. Yerle M, Gellin J, Echard G, Lefevre F, Gillois M: Chromosomal localization of leukocyte interferon gene in the pig (Sus scrofa domestica L.) by in situ hybridization.Cytogenet Cell Genet, 42, 129-132, 1986. DOI: 10.1159/000132265
36. Zanotti C, Razzuoli E, Crooke H, Soule O, Pezzoni G, Ferraris M, Ferrari A, Amadori M: Differential biological activities of swine interferon-α subtypes. J Interferon Cytokine Res, 35, 990-1002, 2015. DOI: 10.1089/jir.2015.0076
37. Cheng G, Zhao X, Chen W, Yan W, Liu M, Chen J, Zheng Z: Detection of differential expression of porcine IFN-alpha subtypes by reverse transcription polymerase chain reaction. J Interferon Cytokine Res, 27, 579-587, 2007. DOI: 10.1089/jir.2006.0126
38. Razzuoli E, Villa R, Sossi E, Amadori M: Reverse transcription real-time PCR for detection of porcine interferon α and β genes.Scand J Immunol, 74, 412-418, 2011. DOI: 10.1111/j.1365-3083.2011.02586.x
39. Sang Y, Rowland RR, Hesse RA, Blecha F: Differential expression and activity of the porcine type I interferon family. Physiol Genomics, 42, 248-258, 2010. DOI: 10.1152/physiolgenomics.00198.2009
40. Sosan O, Graham S, Everett H, Crudgington B, Bodman-Smith K, Crooke H: Differential antiviral effect of porcine interferon alpha subtypes on classical swine fever virus infection of porcine monocytes. Cytokine, 59, 552-552, 2012. DOI: 10.1016/j.cyto.2012.06.192
41. Wang YB, Wang ZY, Chen HY, Cui BA, Wang YB, Zhang HY, Wang R: Secretory expression of porcine interferon-gamma in baculovirus using HBM signal peptide and its inhibition activity on the replication of porcine reproductive and respiratory syndrome virus. Vet Immunol Immunopathol, 132, 314-317, 2009. DOI: 10.1016/j.vetimm.2009.05.017
42. Yu R, Dong S, Zhu Y, Jin H, Gao M, Duan Z, Zheng Z, Shi Z, Li Z: Effective and stable porcine interferon-alpha production by Pichia pastoris fed-batch cultivation with multi-variables clustering and analysis. Bioprocess Biosyst Eng, 33, 473-483, 2010. DOI: 10.1007/s00449- 009-0356-3
43. Kim SM, Kim SK, Park JH, Lee KN, Ko YJ, Lee HS, Seo MG, Shin YK, Kim B: A recombinant adenovirus bicistronically expressing porcine interferon-α and interferon-γ enhancesantiviral effects against footand-mouth disease virus. Antiviral Res, 104, 52-58, 2014. DOI: 10.1016/j. antiviral.2014.01.014
44. Huang L, Cao RB, Wang N, Liu K, Wei JC, Isahg H, Song LJ, Zuo WY, Zhou B, Wang WW, Mao X, Chen PY: The design and recombinant protein expression of a consensus porcine interferon: CoPoIFN-α. Cytokine, 57, 37-45, 2012. DOI: 10.1016/j.cyto.2011.10.011
45. Ma SJ, Li K, Li XS, Guo XQ, Fu PF, Yang MF, Chen HY: Expression of bioactive porcine interferon-alpha in Lactobacillus casei. World J Microbiol Biotechnol, 30, 2379-2386, 2014. DOI: 10.1007/s11274-014-1663-7
46. Mahmoudi S, Abtahi H, Bahador A, Mosayebi G, Salmanian AH, Teymuri M: Optimizing of nutrients for high level expression of recombinant streptokinase using pET32a expression system. Maedica (Buchar), 7, 241-246, 2012.
47. Wang Q, Zhu F, Xin Y, Liu J, Luo L, Yin Z: Expression and purification of antimicrobial peptide buforin IIb in Escherichia coli. Biotechnol Lett, 33, 2121-2126, 2011. DOI: 10.1007/s10529-011-0687-4