Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/97741
標題: 豬β2整合素基因轉殖小鼠對胸膜肺炎放線桿菌之感受性探討
Sensitivity of porcine β2 integrin transgenic mice to Actinobacillus pleuropneumoniae
作者: 洪榮昭
Jung-Chao Hung
關鍵字: 胸膜肺炎放線桿菌;β2整合素;基因轉殖小鼠;Actinobacillus pleuropneumoniae;β2 integrin;transgenic mice
引用: 林俊宏,2009。豬胸膜肺炎放線桿菌及豬巴氏桿菌疫苗市場現況與發展趨勢。農業生技產業季刊。20:18-23。 張兆緯,2016。β2整合素在胸膜肺炎放線桿菌致病性中所扮演的角色。國立中興大學獸醫病理生物學研究所碩士論文。 Abbas, A. K., Lichtman, A. H., Pillai, S., 2014. Basic immunology: functions and disorders of the immune system. Elsevier Health Sciences. Atapattu, D. N., Albrecht, R. M., McClenahan, D. J., Czuprynski, C. J., 2008. Dynamin-2-dependent targeting of mannheimia haemolytica leukotoxin to mitochondrial cyclophilin D in bovine lymphoblastoid cells. Infect. Immun. 76, 5357-5365. Atapattu, D. N., Czuprynski, C. J., 2005. Mannheimia haemolytica leukotoxin induces apoptosis of bovine lymphoblastoid cells (BL-3) via a caspase-9-dependent mitochondrial pathway. Infect. Immun. 73, 5504-5513. Atapattu, D. N., Czuprynski, C. J., 2007. Mannheimia haemolytica leukotoxin binds to lipid rafts in bovine lymphoblastoid cells and is internalized in a dynamin-2- and clathrin-dependent manner. Infect. Immun. 75, 4719-4727. Baarsch, M. J., Scamurra, R. W., Burger, K., Foss, D. L., Maheswaran, S. K., Murtaugh, M. P., 1995. Inflammatory cytokine expression in swine experimentally infected with Actinobacillus pleuropneumoniae. Infect. Immun. 63, 3587-3594. Bandara, A. B., Lawrence, M. L., Veit, H. P., Inzana, T. J., 2003. Association of Actinobacillus pleuropneumoniae capsular polysaccharide with virulence in pigs. Infect. Immun. 71, 3320-3328. Bertram, T. A., 1985. Quantitative morphology of peracute pulmonary lesions in swine induced by Haemophilus pleuropneumoniae. Vet. Pathol. 22, 598-609. Bertram, T. A., 1988. Pathobiology of acute pulmonary lesions in swine infected with Haemophilus (Actinobacillus) pleuropneumoniae. Can. Vet. J. 29, 574-577. Blackall, P. J., Klaasen, H. L., van den Bosch, H., Kuhnert, P., Frey, J., 2002. Proposal of a new serovar of Actinobacillus pleuropneumoniae: serovar 15. Vet. Microbiol. 84, 47-52. Bosse, J. T., Li, Y., Sarkozi, R., Fodor, L., Lacouture, S., Gottschalk, M., Casas Amoribieta, M., Angen, O., Nedbalcova, K., Holden, M. T. G., Maskell, D. J., Tucker, A. W., Wren, B. W., Rycroft, A. N., Langford, P. R., BRaDP1T consortium, 2018. Proposal of serovars 17 and 18 of Actinobacillus pleuropneumoniae based on serological and genotypic analysis. Vet. Microbiol. 217, 1-6. Bosse, J. T., Li, Y., Sarkozi, R., Gottschalk, M., Angen, O., Nedbalcova, K., Rycroft, A. N., Fodor, L., Langford, P. R., 2017. A unique capsule locus in the newly designated Actinobacillus pleuropneumoniae serovar 16 and development of a diagnostic PCR assay. J. Clin. Microbiol. 55, 902-907. Chang, N. Y., Chen, Z. W., Chen, T. H., Liao, J. W., Lin, C. C., Chien, M. S., Lee, W. C., Lin, J. H., Hsuan, S. L., 2014. Elucidating the role of ApxI in hemolysis and cellular damage by using a novel apxIA mutant of Actinobacillus pleuropneumoniae serotype 10. J. Vet. Sci. 15, 81-89. Charlesworth, B., 1996. The evolution of chromosomal sex determination and dosage compensation. Curr. Biol. 6, 149-162. Chatellier, S., Harel, J., Dugourd, D., Chevallier, B., Kobisch, M., Gottschalk, M., 1999. Genomic relatedness among Actinobacillus pleuropneumoniae field strains of sterotypes 1 and 5 isolated from healthy and diseased pigs. Can. J. Vet. Res. 63, 170-176. Chen, Z. W., Chien, M. S., Chang, N. Y., Chen, T. H., Wu, C. M., Huang, C., Lee, W. C., Hsuan, S. L., 2011. Mechanisms underlying Actinobacillus pleuropneumoniae exotoxin ApxI induced expression of IL-1beta, IL-8 and TNF-alpha in porcine alveolar macrophages. Vet. Res. 42, 25. Cheng, M. K., Disteche, C. M., 2004. Silence of the fathers: early X inactivation. Bioessays 26, 821-824. Chi, D. S., Harris, N. S., 1978. A simple method for the isolation of murine peripheral blood lymphocytes. J. Immunol. Methods 19, 169-172. Chien, M. S., Chan, Y. Y., Chen, Z. W., Wu, C. M., Liao, J. W., Chen, T. H., Lee, W. C., Yeh, K. S., Hsuan, S. L., 2009. Actinobacillus pleuropneumoniae serotype 10 derived ApxI induces apoptosis in porcine alveolar macrophages. Vet. Microbiol. 135, 327-333. Chiers, K., De Waele, T., Pasmans, F., Ducatelle, R., Haesebrouck, F., 2010. Virulence factors of Actinobacillus pleuropneumoniae involved in colonization, persistence and induction of lesions in its porcine host. Vet. Res. 41, 65. Chiers, K., Donne, E., Van Overbeke, I., Ducatelle, R., Haesebrouck, F., 2002. Actinobacillus pleuropneumoniae infections in closed swine herds: infection patterns and serological profiles. Vet. Microbiol. 85, 343-352. Choi, C., Kwon, D., Min, K., Chae, C., 1999. In-situ hybridization for the detection of inflammatory cytokines (IL-1, TNF-alpha and IL-6) in pigs naturally infected with Actinobacillus pleuropneumoniae. J. Comp. Pathol. 121, 349-356. Chung, W. B., Backstrom, L. R., McDonald, J., Collins, M. T., 1993. The (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium) colorimetric assay for the quantitation of Actinobacillus pleuropneumoniae cytotoxin. Can. J. Vet. Res. 57, 159-165. Clinkenbeard, K. D., Mosier, D. A., Confer, A. W., 1989. Transmembrane pore size and role of cell swelling in cytotoxicity caused by Pasteurella haemolytica leukotoxin. Infect. Immun. 57, 420-425. Costantini, F., Lacy, E., 1981. Introduction of a rabbit beta-globin gene into the mouse germ line. Nature 294, 92-94. Das, P. J., Mishra, D. K., Ghosh, S., Avila, F., Johnson, G. A., Chowdhary, B. P., Raudsepp, T., 2013. Comparative organization and gene expression profiles of the porcine pseudoautosomal region. Cytogenet. Genome Res. 141, 26-36. Dileepan, T., Thumbikat, P., Walcheck, B., Kannan, M. S., Maheswaran, S. K., 2005. Recombinant expression of bovine LFA-1 and characterization of its role as a receptor for Mannheimia haemolytica leukotoxin. Microb. Pathog. 38, 249-257. Dubreuil, J. D., Jacques, M., Mittal, K. R., Gottschalk, M., 2000. Actinobacillus pleuropneumoniae surface polysaccharides: their role in diagnosis and immunogenicity. Anim. Health Res. Rev. 1, 73-93. Eckmann, C. R., Rammelt, C., Wahle, E., 2011. Control of poly(A) tail length. Wiley Interdiscip Rev RNA 2, 348-361. Enriquez-Verdugo, I., Guerrero, A. L., Serrano, J. J., Godinez, D., Rosales, J. L., Tenorio, V., de la Garza, M., 2004. Adherence of Actinobacillus pleuropneumoniae to swine-lung collagen. Microbiology 150, 2391-2400. Evans, R., Patzak, I., Svensson, L., De Filippo, K., Jones, K., McDowall, A., Hogg, N., 2009. Integrins in immunity. J. Cell Sci. 122, 215-225. Fenwick, B., Henry, S., 1994. Porcine pleuropneumonia. J. Am. Vet. Med. Assoc. 204, 1334-1340. Fiser, R., Konopasek, I., 2009. Different modes of membrane permeabilization by two RTX toxins: HlyA from Escherichia coli and CyaA from Bordetella pertussis. Biochim. Biophys. Acta 1788, 1249-1254. Fong, K. P., Pacheco, C. M., Otis, L. L., Baranwal, S., Kieba, I. R., Harrison, G., Hersh, E. V., Boesze-Battaglia, K., Lally, E. T., 2006. Actinobacillus actinomycetemcomitans leukotoxin requires lipid microdomains for target cell cytotoxicity. Cell. Microbiol. 8, 1753-1767. Frey, J., 2011. The role of RTX toxins in host specificity of animal pathogenic Pasteurellaceae. Vet. Microbiol. 153, 51-58. Frey, J., Bosse, J. T., Chang, Y. F., Cullen, J. M., Fenwick, B., Gerlach, G. F., Gygi, D., Haesebrouck, F., Inzana, T. J., Jansen, R., et al., 1993. Actinobacillus pleuropneumoniae RTX-toxins: uniform designation of haemolysins, cytolysins, pleurotoxin and their genes. J. Gen. Microbiol. 139, 1723-1728. Frey, J., Kuhnert, P., 2002. RTX toxins in Pasteurellaceae. Int. J. Med. Microbiol. 292, 149-158. Frey, J., Nicolet, J., 1990. Hemolysin patterns of Actinobacillus pleuropneumoniae. J. Clin. Microbiol. 28, 232-236. Fuller, C. A., Yu, R., Irwin, S. W., Schryvers, A. B., 1998. Biochemical evidence for a conserved interaction between bacterial transferrin binding protein A and transferrin binding protein B. Microb. Pathog. 24, 75-87. Goel, M. K., Khanna, P., Kishore, J., 2010. Understanding survival analysis: Kaplan-Meier estimate. Int J Ayurveda Res 1, 274-278. Gonzalez, G. C., Yu, R. H., Rosteck, P. R., Jr., Schryvers, A. B., 1995. Sequence, genetic analysis, and expression of Actinobacillus pleuropneumoniae transferrin receptor genes. Microbiology 141 ( Pt 10), 2405-2416. Gordon, J. W., Ruddle, F. H., 1981. Integration and stable germ line transmission of genes injected into mouse pronuclei. Science 214, 1244-1246. Gordon, J. W., Scangos, G. A., Plotkin, D. J., Barbosa, J. A., Ruddle, F. H., 1980. Genetic transformation of mouse embryos by microinjection of purified DNA. Proc. Natl. Acad. Sci. U. S. A. 77, 7380-7384. Gottschalk, M., Broes, A., Mittal, K. R., Kobisch, M., Kuhnert, P., Lebrun, A., Frey, J., 2003. Non-pathogenic Actinobacillus isolates antigenically and biochemically similar to Actinobacillus pleuropneumoniae: a novel species? Vet. Microbiol. Gram, T., Ahrens, P., Andreasen, M., Nielsen, J. P., 2000. An Actinobacillus pleuropneumoniae PCR typing system based on the apx and omlA genes--evaluation of isolates from lungs and tonsils of pigs. Vet. Microbiol. 75, 43-57. Haesebrouck, F., Chiers, K., Van Overbeke, I., Ducatelle, R., 1997. Actinobacillus pleuropneumoniae infections in pigs: the role of virulence factors in pathogenesis and protection. Vet. Microbiol. 58, 239-249. Harlan, J. M., Vedder, N. B., Winn, R. K., Rice, C. L., 1991. Mechanisms and consequences of leukocyte-endothelial interaction. West. J. Med. 155, 365-369. Hsu, C. W., Li, S. C., Chang, N. Y., Chen, Z. W., Liao, J. W., Chen, T. H., Wang, J. P., Lin, J. H., Hsuan, S. L., 2016. Involvement of NF-kappaB in regulation of Actinobacillus pleuropneumoniae exotoxin ApxI-induced proinflammatory cytokine production in porcine alveolar macrophages. Vet. Microbiol. 195, 128-135. Inzana, T. J., Ma, J., Workman, T., Gogolewski, R. P., Anderson, P., 1988. Virulence properties and protective efficacy of the capsular polymer of Haemophilus (Actinobacillus) pleuropneumoniae serotype 5. Infect. Immun. 56, 1880-1889. Jacques, M., 2004. Surface polysaccharides and iron-uptake systems of Actinobacillus pleuropneumoniae. Can. J. Vet. Res. 68, 81-85. Jaenisch, R., Mintz, B., 1974. Simian virus 40 DNA sequences in DNA of healthy adult mice derived from preimplantation blastocysts injected with viral DNA. Proc. Natl. Acad. Sci. U. S. A. 71, 1250-1254. Jurisic, V., Konjevic, G., Jancic-Nedeljkov, R., Sretenovic, M., Banicevic, B., Colovic, M., Spuzic, I., 2004. The comparison of spontaneous LDH release activity from cultured PBMC with sera LDH activity in non-Hodgkin's lymphoma patients. Med. Oncol. 21, 179-185. Lally, E. T., Hill, R. B., Kieba, I. R., Korostoff, J., 1999. The interaction between RTX toxins and target cells. Trends Microbiol. 7, 356-361. Lally, E. T., Kieba, I. R., Sato, A., Green, C. L., Rosenbloom, J., Korostoff, J., Wang, J. F., Shenker, B. J., Ortlepp, S., Robinson, M. K., Billings, P. C., 1997. RTX toxins recognize a beta2 integrin on the surface of human target cells. J. Biol. Chem. 272, 30463-30469. Lessard, P. A., Kulaveerasingam, H., York, G. M., Strong, A., Sinskey, A. J., 2002. Manipulating gene expression for the metabolic engineering of plants. Metab Eng 4, 67-79. Meshulam, T., Levitz, S. M., Christin, L., Diamond, R. D., 1995. A simplified new assay for assessment of fungal cell damage with the tetrazolium dye, (2,3)-bis-(2-methoxy-4-nitro-5-sulphenyl)-(2H)-tetrazolium-5-carboxanil ide (XTT). J. Infect. Dis. 172, 1153-1156. Morova, J., Osicka, R., Masin, J., Sebo, P., 2008. RTX cytotoxins recognize beta2 integrin receptors through N-linked oligosaccharides. Proc. Natl. Acad. Sci. U. S. A. 105, 5355-5360. Park, C., Ha, Y., Kim, S., Chae, C., Ryu, D. Y., 2009. Construction and characterization of an Actinobacillus pleuropneumoniae serotype 2 mutant lacking the Apx toxin secretion protein genes apxIIIB and apxIIID. J. Vet. Med. Sci. 71, 1317-1323. Pohl, S., Bertschinger, H. U., Frederiksen, W., Mannheim, W., 1983. Transfer of Haemophilus pleuropneumoniae and the Pasteurella haemolytica-like organism causing porcine necrotic pleuropneumonia to the genus Actinobacillus (Actinobacillus pleuropneumoniae comb. nov.) on the basis of phenotypic and deoxyribonucleic acid relatedness. Int. J. Syst. Evol. Microbiol. 33, 510-514. Preiss, T., Muckenthaler, M., Hentze, M. W., 1998. Poly(A)-tail-promoted translation in yeast: implications for translational control. RNA 4, 1321-1331. Ramjeet, M., Deslandes, V., St Michael, F., Cox, A. D., Kobisch, M., Gottschalk, M., Jacques, M., 2005. Truncation of the lipopolysaccharide outer core affects susceptibility to antimicrobial peptides and virulence of Actinobacillus pleuropneumoniae serotype 1. J. Biol. Chem. 280, 39104-39114. Rayamajhi, N., Shin, S. J., Kang, S. G., Lee, D. Y., Ahn, J. M., Yoo, H. S., 2005. Development and use of a multiplex polymerase chain reaction assay based on Apx toxin genes for genotyping of Actinobacillus pleuropneumoniae isolates. J. Vet. Diagn. Invest. 17, 359-362. Reed, L. J., Muench, H., 1938. A simple method of estimating fifty per cent endpoints. Am. J. Epidemiol. 27, 493-497. Shope, R. E., 1964. Porcine contagious pleuropneumonia. I. Experimental transmission, etiology, and pathology. J. Exp. Med. 119, 357-368. Sibille, Y., Reynolds, H. Y., 1990. Macrophages and polymorphonuclear neutrophils in lung defense and injury. Am. Rev. Respir. Dis. 141, 471-501. Steffens, W. L., Byrd, W., Kadis, S., 1990. Identification and localization of surface sialylated glycoconjugates in Actinobacillus pleuropneumoniae by direct enzyme-colloidal gold cytochemistry. Vet. Microbiol. 25, 217-227. Strathdee, C. A., Lo, R. Y., 1989. Cloning, nucleotide sequence, and characterization of genes encoding the secretion function of the Pasteurella haemolytica leukotoxin determinant. J. Bacteriol. 171, 916-928. van den Bosch, H., Frey, J., 2003. Interference of outer membrane protein PalA with protective immunity against Actinobacillus pleuropneumoniae infections in vaccinated pigs. Vaccine 21, 3601-3607. Van Overbeke, I., Chiers, K., Charlier, G., Vandenberghe, I., Van Beeumen, J., Ducatelle, R., Haesebrouck, F., 2002. Characterization of the in vitro adhesion of Actinobacillus pleuropneumoniae to swine alveolar epithelial cells. Vet. Microbiol. 88, 59-74. Vanden Bergh, P. G., Fett, T., Zecchinon, L. L., Thomas, A. V., Desmecht, D. J., 2005. The CD11a partner in Sus scrofa lymphocyte function-associated antigen-1 (LFA-1): mRNA cloning, structure analysis and comparison with mammalian homologues. BMC Vet. Res. 1, 5. Vanden Bergh, P. G., Zecchinon, L. L., Fett, T., Desmecht, D., 2008. Probing of Actinobacillus pleuropneumoniae ApxIIIA toxin-dependent cytotoxicity towards mammalian peripheral blood mononucleated cells. BMC Res. Notes 1, 121. Vanden Bergh, P. G., Zecchinon, L. L., Fett, T., Desmecht, D., 2009. Porcine CD18 mediates Actinobacillus pleuropneumoniae ApxIII species-specific toxicity. Vet. Res. 40, 33. Wang, C., Yu, X., Cao, Q., Wang, Y., Zheng, G., Tan, T. K., Zhao, H., Zhao, Y., Wang, Y., Harris, DCh, 2013. Characterization of murine macrophages from bone marrow, spleen and peritoneum. BMC Immunol. 14, 6. Wang, Y. C., Chan, J. P., Yeh, K. S., Chang, C. C., Hsuan, S. L., Hsieh, Y. M., Chang, Y. C., Lai, T. C., Lin, W. H., Chen, T. H., 2010. Molecular characterization of enrofloxacin resistant Actinobacillus pleuropneumoniae isolates. Vet. Microbiol. 142, 309-312. White, M. A., Ikeda, A., Payseur, B. A., 2012. A pronounced evolutionary shift of the pseudoautosomal region boundary in house mice. Mamm. Genome 23, 454-466. Wu, C. M., Chen, Z. W., Chen, T. H., Liao, J. W., Lin, C. C., Chien, M. S., Lee, W. C., Hsuan, S. L., 2011. Mitogen-activated protein kinases p38 and JNK mediate Actinobacillus pleuropneumoniae exotoxin ApxI-induced apoptosis in porcine alveolar macrophages. Vet. Microbiol. 151, 372-378. Xu, Z., Zhou, Y., Li, L., Zhou, R., Xiao, S., Wan, Y., Zhang, S., Wang, K., Li, W., Li, L., Jin, H., Kang, M., Dalai, B., Li, T., Liu, L., Cheng, Y., Zhang, L., Xu, T., Zheng, H., Pu, S., Wang, B., Gu, W., Zhang, X. L., Zhu, G. F., Wang, S., Zhao, G. P., Chen, H., 2008. Genome biology of Actinobacillus pleuropneumoniae JL03, an isolate of serotype 3 prevalent in China. PLoS One 3, e1450. Yang, C. Y., Lin, C. N., Lin, C. F., Chang, T. C., Chiou, M. T., 2011. Serotypes, antimicrobial susceptibility, and minimal inhibitory concentrations of Actinobacillus pleuropneumoniae isolated from slaughter pigs in Taiwan (2002-2007). J. Vet. Med. Sci. 73, 205-208. Zimmerman, J. J., Karriker, L. A., Ramirez, A., Schwartz, K. J., Stevenson, G. W., 2012. Diseases of Swine. 10th edition.
摘要: 
胸膜肺炎放線桿菌(Actinobacillus pleuropneumoniae, AP)感染豬隻呼吸道所引起的纖維素性、出血性及壞死性胸膜肺炎會造成豬隻死亡。AP所分泌外毒素,包含ApxI~ApxIV,Apx依結構特色被歸類為Repeat in Toxin(RTX)家族,RTX經由與白血球上的β2整合素(CD11/CD18)結合,造成細胞毒殺作用。過去研究指出,ApxIII對豬隻CD18(porcine CD18, pCD18)具物種專一性。本研究為探討pCD18與AP致病機轉之相關性,以C57BL/6JNarl小鼠構築表現pCD18基因轉殖(Transgenic, TG)小鼠,並分析TG小鼠對AP之感受性。首先,分離小鼠腹腔灌洗細胞與脾臟細胞,並以流式細胞儀分析其中巨噬細胞、B細胞及T細胞族群比例,發現TG與Wild type(WT)小鼠相似,但TG小鼠表現pCD18比例明顯高於WT小鼠,且母鼠表現量顯著高於公鼠。此外,以AP血清型第1型(AP1)和第2型(AP2)菌株進行腹腔內攻毒實驗。結果顯示,攻毒後TG母鼠較快出現死亡,TG與WT母鼠於AP1的半致死劑量並無差異,而TG母鼠對AP2的半致死劑量比WT母鼠低7.5倍,說明TG母鼠對AP菌株具較高敏感性。另為探討AP所分泌Apx對小鼠免疫細胞所造成的影響,本研究自WT與TG母鼠分離脾臟單核細胞(SMCs),分別與AP10所分泌之ApxI或AP2所分泌之ApxII/III進行感作,結果發現無論ApxI或ApxII/III對TG母鼠SMCs皆造成較嚴重的細胞毒性。為進一步探討pCD18表現比例與Apx致病性關係,自TG母鼠分離腹腔灌洗細胞與SMCs,利用磁珠抗體分選高純度pCD18陽性細胞與無標記的陰性細胞,分別與1 ~ 10 CU ApxII/III進行感作。結果顯示,ApxII/III對pCD18陽性細胞具劑量效應,毒素濃度越高造成更高比例之細胞膜損傷,其中對腹腔灌洗細胞造成更嚴重的細胞損傷情形。綜合上述結果,本研究已建立pCD18基因轉殖小鼠對AP疾病動物模式,證實pCD18於AP致病性和Apx外毒素細胞毒殺作用中扮演重要的角色,此基因轉殖小鼠具有應用於AP致病機轉探討或作為效力檢定工具之潛力。

Actinobacillus pleuropneumoniae (AP) causes fibrinous, hemorrhagic, and necrotizing pleuropneumonia in pigs. The exotoxins secreted by AP, including ApxI to ApxIV, are classified as members of the Repeats-in-Toxin (RTX) family based on their structural characteristics. It has been demonstrated that RTX toxins target specifically to β2 integrin (CD11/CD18) and lead to cell death. Previous studies indicate that the porcine CD18 (pCD18) mediates ApxIII-induced species-specific toxicity on pig leukocytes. The aim of this study was to investigate the role of pCD18 in AP pathogenicity. A line of transgenic (TG) C57BL/6JNarl mouse expressing pCD18 was generated and studied. To characterize the expression of pCD18, we isolated the peritoneal lavage cells (PLCs) and splenocytes, and analyzed the percent of macrophages, B cells, and T cells among them. A similar percentage of cell population was found in both wild type (WT) and TG mice. PLCs and splenocytes derived from TG mice, especially from TG female mice, had a higher expression level of pCD18 as compared to WT mice or TG male mice. Therefore, female TG mice were used in the subsequent experiments. To investigate the susceptibility of TG and WT mice to AP, mice were intraperitoneally inoculated with AP serotype 1 (AP1) or 2 (AP2), and the mortality assessed and compared. It was found that the death of TG mice occurred at time points earlier than that of WT mice after challenge. The LD50 of AP1 in TG mice was equivalent to that in WT mice, while the LD50 of AP2 in TG mice was 7.5 times lower than that in WT mice. We inferred that the higher susceptibility of TG mice to AP may result from the expression of pCD18. To test this hypothesis, PLCs and splenic mononuclear cells (SMCs) of TG mice were collected and purified using magnetic beads coupled with antibodies recognizing porcine CD18. The susceptibility of pCD18+-cells towards Apx toxins were assessed and compared to pCD18--cells using the lactate dehydrogenase assay. The results showed that pCD18+-cells were more susceptible toward ApxI and ApxII/III comparing to pCD18--cells. Apx toxins induced significant levels of cytotoxicity in a concentration-dependent manner in pCD18+-cells, which signifies an indispensable role of pCD18 in Apx cytotoxic effect. In conclusion, we have successfully generated the TG mice expressing pCD18 and demonstrate the role of pCD18 in AP pathogenicity and in Apx cytotoxicity. The TG mice might be a valuable tool for the study of AP pathogenicity and for the evaluation of vaccine efficacy in the future.
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