Vaccine 29 (2011) 3888–3894 Contents lists available at ScienceDirect Vaccine journal homepage: www.elsevier.com/locate/vaccine Improvement of the immunity of pig to Hog cholera vaccine by recombinant plasmid with porcine interleukin-6 gene and CpG motifs Dong Li 1 , Jian-Lin Chen 1 , Huan Zhang 1,2 , Xiao Yang, Xiao-Ping Wan, Chi Cheng 3 , Yue Li, Ze-Zhou Wang, Xue-Bin Lv 4 , Hong-Ning Wang, Hai-Yan Wang ∗ , Jiang-Lin Li ∗,4 , Rong Gao ∗ Key Laboratory for Bio-Resource and Eco-Environment of Ministry Education, Key Laboratory for Animal Disease Prevention and Food Safety, Life Science College, Sichuan University, Wangjiang Road 29th, Chengdu 610064, Sichuan, PR China article info Article history: Received 2 September 2008 Received in revised form 8 March 2011 Accepted 12 March 2011 Available online 26 March 2011 Keywords: Pig Hog cholera vaccine Immunity Pig interleuin-6 gene CpG motifs abstract In order to observe the dosage-effect of recombinant pig interleukin-6 gene and CpG motifs on the immune responses of swine to vaccine, a novel recombinant eukaryotic VPIL6C plasmid was packed with chitosan nanoparticles (CNP) prepared by ionic cross linkage, which contains pig interleukin-6 gene and immunostimulatory sequence consisted of 11 CpG motifs. CNP-VRIL6C was then utilized to inoc- ulate 30-day-old piglets intramuscularly at the dosage of 0.5, 1.0 and 1.5 mg/per capita, respectively. Meanwhile, the piglets were injected with attenuated classical Hog cholera vaccine and designated as A1, A2 and A3 group. The blood was weekly collected from the piglets after vaccination to detect the changes of immunoglobulins, specific antibody, interleukins, IFN- and immune cells. The results were found that compared to those of the control piglets injected with VR1020-CNP, the content of IgG, IgA and IgM, specific antibodies, IL-2, IL-6 and IFN- significantly increased in the sera from the treated three groups from 14 to 70 days after vaccination (P < 0.05); the number of T H ,T C and CD3 + positive T cells raised obviously in the blood of VPIL6C treated piglets (P < 0.05). Also the above immune indexes of A1 group were significantly lower to different extent in comparison with those of A2 and A3 group from 14 to 56 days post inoculation (P > 0.05). Moreover, the lymphocytes also remarkably elevated in the treated groups (P < 0.05). These indicate that VPIL6C entrapped with CNP is a novel effective adjuvant to boost the humoral and cellular immunity of pig to Hog cholera, implying it’s potentiality to enhance the resistance of pig against infectious diseases. © 2011 Elsevier Ltd. All rights reserved. 1. Introduction Classical swine fever (CSF) or hog cholera is one of the most devastating porcine haemorrhagic viral diseases. Outbreaks of CSF usually lead to significant losses in many countries worldwide [1]. CSF virus (CSFV) mainly infects endothelial cells and macrophages and at the same time promotes bystander apoptosis of the sur- rounding T cells, causing strong immune suppression and high mortality rates [2–5]. Presently, novel vaccine and adjuvant is crit- Abbreviations: CNP, chitosan nanoparticle; VPIL6C, recombinant eukaryotic VR1020 plasmid inserted with pig IL-6 and CpG; VPIL6C- CNP (CNPs), VPIL6C enwrapped into CNP; PIL-6, pig interleukin-6 gene. ∗ Corresponding authors. Tel.: +86 28 85411033; fax: +86 28 85471599. E-mail address: lake96@qq.com (R. Gao). 1 These authors contribute equally to the research. 2 Biology group of North Sichuan Medical College, Nan Chong 637007, Sichuan, China. 3 Bioengineering Department of Sichuan University of Science & Engineering, Zigong 643000, China. 4 Animal Academy of Sichuan province, Chengdu 610035, China. ical for enhancing immunity and resistance of pig against various infections. Vaccinations are frequently compromised due to rapid mutation of virus and bacteria. Nowadays adjuvant is still important for boosting immunity and improving resistance in animals. Traditional adjuvants are mainly consisting of chemical molecule and microbial components, such as Al (OH) 3 , BCG and incomplete Freund’s adjuvant, derived bacteria and plant components [6]. Recently cytokines had been utilized as new effective adjuvants [7,8]. Moreover, pig Interleukin (IL-6) gene was ever proved to be able to enhance the immune responses of animal [9], which was one of key cytokines balanc- ing between inflammation and the immune responses during the infection or injury process [10]. CpG motifs were also reported to represent another emerging adjuvant to strengthen the innate immunity of animal. The unique unmethylated CpG motifs could activate B cells, dendritic cells (DCs), NK T cells and monocytes and play an important role in infection and vaccination [11,12]. Clinical studies demonstrate that CpG could be applied in therapy against infectious diseases, cancer, asthma and allergy [13]. The adjuvant activity of CpG ODN was demonstrated in humans and numerous animal species [14–16]. But they are usually not safe or effective 0264-410X/$ – see front matter © 2011 Elsevier Ltd. All rights reserved. doi:10.1016/j.vaccine.2011.03.036