تاثیر آنتی‌ژن ویروس عامل بیماری IPN بارگذاری شده در ذرات کیتوزان و در ماتریس آلژینات بر بیان ژن های 12-IL، IL-17و ترکیب گلبول های سفید خون در ماهی قزل‌آلای رنگین کمان (Oncorhynchus mykiss)

نوع مقاله : مقاله پژوهشی

نویسندگان
بهنام بختیاری مقدم 1
شفیق شفیعی 2
صادق شیریان 2
اسماعیل میرزایی 3
اعظم مختاری 2
زهره خورشیدوند 4
1 دانشجوی دکتری تخصصی بهداشت وبیماری های آبزیان دانشگاه شهرکرد
2 دانشگاه دولتی شهرکرد
3 علوم پزشکی شیراز
4 دانشگاه علومپزشکی همدان
10.22034/ijvcs.2026.14960.1096
چکیده
بیماری نکروز عفونی پانکراس (Infectious pancreatic necrosis) یکی از چالش‌های اصلی صنعت آبزی‌پروری، به‌ویژه در پرورش ماهی قزل‌آلا است. این مطالعه با هدف بررسی تأثیر واکسن غیرفعال خوراکی حاوی آنتی‌ژن ویروس IPN بارگذاری‌شده در نانوذرات کیتوزان و ماتریس آلژینات بر بیان ژن‌هایIL-12، IL-17 و ترکیب گلبول‌های سفید در ماهی قزل‌آلای رنگین‌کمان (Oncorhynchus mykiss) انجام شد. تعداد ۵۴۰ قطعه ماهی با میانگین وزن 5/0± 3 گرم به شش گروه شامل کنترل، کنترل مثبت (نانوذرات بدون آنتی‌ژن) و چهار گروه دریافت‌کننده آنتی‌ژن آزاد (06/0 یا 12/0 سی‌سی به ازای هر ماهی) و نانوذرات حاوی آنتی‌ژن (06/0 یا 12/0سی‌سی) تقسیم شدند. واکسیناسیون خوراکی در روزهای صفر و ۱۵ انجام و نمونه‌برداری در روز ۳۰ صورت گرفت. نتایج نشان داد که دریافت آنتی‌ژن، به‌ویژه در دوز بالاتر و همراه با نانوذرات، باعث افزایش معنی‌دار درصد لنفوسیت‌ها و کاهش معنی‌دار درصد نوتروفیل‌ها شد (05/0P<). بیان ژن IL-12 در گروه‌های دریافت‌کننده آنتی‌ژن، به‌خصوص در دوز 12/0 سی‌سی و گروه نانوذرات حاوی آنتی‌ژن با دوز 12/0 سی‌سی، افزایش معنی‌داری یافت(05/0P<)، در حالی که بیان IL-17 تغییر معنی‌داری نشان نداد. نتیجه‌گیری می‌شود که بارگذاری آنتی‌ژن IPNV در نانوذرات کیتوزان-آلژینات به‌عنوان یک سامانه رهش کنترل‌شده، می‌تواند پاسخ‌های ایمنی تطبیقی از نوع Th1 را از طریق افزایش IL-12 تقویت کرده و ترکیب گلبول‌های سفید را به سمت ایمنی اختصاصی‌تر سوق دهد. با این حال، سنجش تیتر آنتی‌بادی و آزمون چالش برای نتیجه‌گیری قطعی ضروری است.
کلیدواژه‌ها
ویروس نکروز عفونی پانکراس
نانوذرات کیتوزان
آلژینات
IL-12
IL-17
قزل‌آلای رنگین‌کمان
IPN
موضوعات

عنوان مقاله English

Effect of IPN virus antigen loaded in chitosan particles and in alginate matrix on IL-12, IL-17 gene expression and white blood cell composition in rainbow trout (Oncorhynchus mykiss)

نویسندگان English

Behnam Bakhtiarimoghadam 1
Shafigh shafiei 2
Sadegh Shirian 2
Esmail Mirzaei 3
Azam Mokhtari 2
Zohre Khorshidvand 4
1 shahre-kord university
2 Shahre kord university
3 Shiraz Medical Sciences
4 Hamedan Medical Sciences
چکیده English

Infectious pancreatic necrosis (IPN) is one of the major challenges in the aquaculture industry, particularly in in rainbow trout farming. This study aimed to evaluate the effect of an inactivated oral vaccine containing IPNV antigen loaded into chitosan nanoparticles and alginate matrix on the gene expression of IL-12, IL-17, and white blood cell composition in rainbow trout (Oncorhynchus mykiss). A total of 540 fish with an average weight of 3 ± 0.5 g were divided into six groups: control, positive control (nanoparticles without antigen), and four groups receiving free antigen (0.06 or 0.12 cc per fish) or antigen-loaded nanoparticles (0.06 or 0.12 cc per fish). Oral vaccination was performed on days 0 and 15, and sampling was conducted on day 30. The results showed that antigen administration, particularly at the higher dose and when associated with nanoparticles, significantly increased the percentage of lymphocytes and significantly decreased the percentage of neutrophils (P < 0.05). The expression of IL-12 significantly increased in antigen-receiving groups, especially at the dose of 0.12 cc and in the group receiving antigen-loaded nanoparticles at 0.12 cc (P < 0.05), whereas IL-17 expression showed no significant change. It is concluded that encapsulation of IPNV antigen in chitosan-alginate nanoparticles, as a controlled-release system, can enhance Th1-type adaptive immune responses through upregulation of IL-12 and shift white blood cell composition toward more specific immunity. However, antibody titer measurement and challenge tests are necessary for definitive conclusions.

کلیدواژه‌ها English

Infectious Pancreatic Necrosis virus
Chitosan nanoparticles
Alginate
IL-12
IL-17
Rainbow trout
IPN
Abourehab MA, Rajendran RR, Singh A, Pramanik S, Shrivastav P, Ansari MJ, Manne R, Amaral LS, Deepak A. Alginate as a promising biopolymer in drug delivery and wound healing: A review of the state-of-the-art. International Journal of Molecular Sciences. 2022 Aug 12;23(16):9035.
2.  Ahmadivand S, Soltani M, Behdani M, Evensen Ø, Alirahimi E, Hassanzadeh R, Soltani E. Oral DNA vaccines based on CS-TPP nanoparticles and alginate microparticles confer high protection against infectious pancreatic necrosis virus (IPNV) infection in trout. Developmental & Comparative Immunology. 2017 Sep 1;74:178-189.
3.  Akhter S, Tasnin FM, Islam MN, Rauf A, Mitra S, Emran TB, Alhumaydhi FA, Ahmed Khalil A, Aljohani AS, Al Abdulmonem W, Thiruvengadam M. Role of Th17 and IL-17 Cytokines on Inflammatory and Auto-immune Diseases. Current Pharmaceutical Design. 2023 Jul 1;29(26):2078-2090.
4. Ballesteros NA, Alonso M, Saint-Jean SR, Perez-Prieto SI. An oral DNA vaccine against infectious haematopoietic necrosis virus (IHNV) encapsulated in alginate microspheres induces dose-dependent immune responses and significant protection in rainbow trout (Oncorrhynchus mykiss). Fish & shellfish immunology. 2015 Aug 1;45(2):877-888.
5. Boroumand H, Badie F, Mazaheri S, Seyedi ZS, Nahand JS, Nejati M, Baghi HB, AbbasiKolli M, Badehnoosh B, Ghandali M, Hamblin MR. Chitosan-based nanoparticles against viral infections. Frontiers in Cellular and Infection Microbiology. 2021 Mar 17;11:643953.
6. Cabillon NA, Lazado CC. Mucosal barrier functions of fish under changing environmental conditions. Fishes. 2019 Jan 10;4(1):2.
7. Campbell JH, Dixon B, Whitehouse LM. The intersection of stress, sex and immunity in fishes. Immunogenetics. 2021 Feb;73(1):111-129.
8. Chang J, Zhu W, Huo X, Qiao M, Yang C, Zhang Y, Su J. Oral Lactobacillus casei expressing VP56310–500 and adjuvant flagellin C delivered by alginate-chitosan microcapsules remarkably enhances the immune protection against GCRV infection in grass carp. Aquaculture. 2023 Mar 30;567:739301.
9. Chimal-Ramírez GK, Espinoza-Sánchez NA, Fuentes-Pananá EM. Protumor activities of the immune response: insights in the mechanisms of immunological shift, oncotraining, and oncopromotion. Journal of oncology. 2013;2013(1):835956.
10. Collet B. Innate immune responses of salmonid fish to viral infections. Developmental & Comparative Immunology. 2014 Apr 1;43(2):160-173.
11. Dmour I, Islam N. Recent advances on chitosan as an adjuvant for vaccine delivery. International Journal of Biological Macromolecules. 2022 Mar 1;200:498-519.
12. Dopazo CP. The infectious pancreatic necrosis virus (IPNV) and its virulence determinants: What is known and what should be known. Pathogens. 2020 Feb 4;9(2):94.
13. Duan K, Tang X, Zhao J, Ren G, Shao Y, Lu T, He B, Xu L. An inactivated vaccine against infectious pancreatic necrosis virus in rainbow trout (Oncorhynchus mykiss). Fish & Shellfish Immunology. 2022 Aug 1;127:48-55.
14. Duan T, Du Y, Xing C, Wang HY, Wang RF. Toll-like receptor signaling and its role in cell-mediated immunity. Frontiers in immunology. 2022 Mar 3;13:812774.
15. Elveborg S, Monteil VM, Mirazimi A. Methods of inactivation of highly pathogenic viruses for molecular, serology or vaccine development purposes. Pathogens. 2022 Feb 19;11(2):271.
16. Farshidfar N, Iravani S, Varma RS. Alginate-based biomaterials in tissue engineering and regenerative medicine. Marine Drugs. 2023 Mar 18;21(3):189.
17. Fayaz I, Bhat RA, Tandel RS, Dash P, Chandra S, Dubey MK, Ganie PA. Comprehensive review on infectious pancreatic necrosis virus. Aquaculture. 2023 Sep 15;574:739737.
18. Gaglio SC, Perduca M, Zipeto D, Bardi G. Efficiency of chitosan nanocarriers in vaccinology for mucosal immunization. Vaccines. 2023 Aug 6;11(8):1333.
19. Gao H, Li K, Ai K, Geng M, Cao Y, Wang D, Yang J, Wei X. Interleukin-12 induces IFN-γ secretion and STAT signaling implying its potential regulation of Th1 cell response in Nile tilapia. Fish & Shellfish Immunology. 2023 Sep 1;140:108974.
20. Giri SS, Kim SG, Kang JW, Kim SW, Kwon J, Lee SB, Jung WJ, Park SC. Applications of carbon nanotubes and polymeric micro‐/nanoparticles in fish vaccine delivery: progress and future perspectives. Reviews in Aquaculture. 2021 Sep;13(4):1844-1863.
21. Gote V, Bolla PK, Kommineni N, Butreddy A, Nukala PK, Palakurthi SS, Khan W. A comprehensive review of mRNA vaccines. International journal of molecular sciences. 2023 Jan 31;24(3):2700.
22. Gregory AE, Titball R, Williamson D. Vaccine delivery using nanoparticles. Frontiers in cellular and infection microbiology. 2013 Mar 25;3:13.
23. Halimi M, Alishahi M, Abbaspour MR, Ghorbanpoor M, Tabandeh MR. Valuable method for production of oral vaccine by using alginate and chitosan against Lactococcus garvieae/Streptococcus iniae in rainbow trout (Oncorhynchus mykiss). Fish & shellfish immunology. 2019 Jul 1;90:431-9.
24. Heidarieh M, Moodi S, Katuli KK, Unger H. Biochemical effects of encapsulated radiovaccine via alginate nanoparticles as useful strategy for booster in immunized rainbow trout against Ichthyophytirius multifiliis. (2015): 1330.
25. Hildenbrand K, Aschenbrenner I, Franke FC, Devergne O, Feige MJ. Biogenesis and engineering of interleukin 12 family cytokines. Trends in Biochemical Sciences. 2022 Nov 1;47(11):936-49.
26.  Huo X, Zhang Q, Chang J, Yang G, He S, Yang C, Liang X, Zhang Y, Su J. Nanopeptide C-I20 as a novel feed additive effectively alleviates detrimental impacts of soybean meal on mandarin fish by improving the intestinal mucosal barrier. Frontiers in Immunology. 2023 Jun 26;14:1197767.
27. Hwang J, Yadav D, Lee PC, Jin JO. Immunomodulatory effects of polysaccharides from marine algae for treating cancer, infectious disease, and inflammation. Phytotherapy Research. 2022 Feb;36(2):761-777.
28. Ji J, Torrealba D, Ruyra À, Roher N. Nanodelivery systems as new tools for immunostimulant or vaccine administration: targeting the fish immune system. Biology. 2015 Oct 19;4(4):664-696.
29.  Kang HJ, Li J, Razzak MA, Eom GD, Yoon KW, Mao J, Chu KB, Jin H, Choi SS, Quan FS. Chitosan-alginate polymeric nanocomposites as a potential oral vaccine carrier against influenza virus infection. ACS applied materials & interfaces. 2023 Oct 30;15(44):50889- 50897.
30.  Kole S, Qadiri SS, Shin SM, Kim WS, Lee J, Jung SJ. Nanoencapsulation of inactivatedviral vaccine using chitosan nanoparticles: evaluation of its protective efficacy and immune modulatory effects in olive flounder (Paralichthys olivaceus) against viral haemorrhagic septicaemia virus (VHSV) infection. Fish & Shellfish Immunology. 2019 Aug 1;91:136-147.
31.  Kole S, Kumari R, Anand D, Kumar S, Sharma R, Tripathi G, Makesh M, Rajendran KV, Bedekar MK. Nanoconjugation of bicistronic DNA vaccine against Edwardsiella tarda using chitosan nanoparticles: Evaluation of its protective efficacy and immune modulatory effects in Labeo rohita vaccinated by different delivery routes. Vaccine. 2018 Apr 12;36(16):2155- 165.
32. Kumar A, Middha SK, Menon SV, Paital B, Gokarn S, Nelli M, Rajanikanth RB, Chandra HM, Mugunthan SP, Kantwa SM, Usha T. Current challenges of vaccination in fish health management. Animals. 2024 Sep 16;14(18):2692.
33. Lakshmi B, Syed S, Buddolla V. Current advances in the protection of viral diseases in aquaculture with special reference to vaccination. Recent developments in applied microbiology and biochemistry. 2019 Jan 1:127-146.
34. Li L, Liu W, Zhang Z, Zhao J, Lu T, Shao Y, Xu L. IPNV Inactive Vaccine Supplemented with GEL 02 PR Adjuvant: Protective Efficacy, Cross-protection, and Stability. Fish & Shellfish Immunology. 2025 Jan 30:110167.
35. Lima BV, Oliveira MJ, Barbosa MA, Goncalves RM, Castro F. Immunomodulatory potential of chitosan-based materials for cancer therapy: a systematic review of in vitro, in vivo and clinical studies. Biomaterials Science. 2021;9(9):3209-3227.
36. Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method. methods. 2001 Dec 1;25(4):402-428.
37. Lv B, Shen N, Cheng Z, Chen Y, Ding H, Yuan J, Zhao K, Zhang Y. Strategies for biomaterial-based spinal cord injury repair via the TLR4-NF-κB signaling pathway. Frontiers in Bioengineering and Biotechnology. 2022 Apr 29;9:813169.
38. Malek-Khatabi A, Tabandeh Z, Nouri A, Mozayan E, Sartorius R, Rahimi S, Jamaledin R. Long-term vaccine delivery and immunological responses using biodegradable polymerbased carriers. ACS Applied Bio Materials. 2022 Oct 10;5(11):5015-5040.
39. Ma Y, Zhang Y, Zhu L. Role of neutrophils in acute viral infection. Immunity, inflammation and disease. 2021 Dec;9(4):1186-1196.
40. Mills KH. IL-17 and IL-17-producing cells in protection versus pathology. Nature Reviews Immunology. 2023 Jan;23(1):38-54.
41. Mondal H, Thomas J. A review on the recent advances and application of vaccines against fish pathogens in aquaculture. Aquaculture international. 2022 Aug;30(4):1971- 2000.
42. Mu R, Dong L, Wang C. Carbohydrates as putative pattern recognition receptor agonists in vaccine development. Trends in immunology. 2023 Oct 1;44(10):845-57. 
43. Najafi A, Ghazvini K, Sankian M, Gholami L, Amini Y, Zare S, Khademi F, Tafaghodi M. T helper type 1 biased immune responses by PPE17 loaded core-shell alginate-chitosan nanoparticles after subcutaneous and intranasal administration. Life Sciences. 2021 Oct 1;282:119806.
44. Najafi A, Ghazvini K, Sankian M, Gholami L, Zare S, Arvand AY, Tafaghodi M. Mucosal and systemic immunization against tuberculosis by ISCOMATRIX nano adjuvant coadministered with alginate coated chitosan nanoparticles. Iranian Journal of Basic Medical Sciences. 2023;26(10):1162.
45. Nandanpawar P, Badhe M, Rather MA, Sharma R. Chitosan nanoparticles for gene delivery in Macrobrachium rosenbergii (DE MAN 1879). Journal of Cell and Tissue Research. 2013 Aug 1;13(2):3619.
46. Oroojalian F, Beygi M, Baradaran B, Mokhtarzadeh A, Shahbazi MA. Immune cell membrane‐coated biomimetic nanoparticles for targeted cancer therapy. Small. 2021 Mar;17(12):2006484.
47. Patra P, Upadhyay TK, Alshammari N, Saeed M, Kesari KK. Alginate-Chitosan biodegradable and biocompatible based hydrogel for breast cancer immunotherapy and diagnosis: a comprehensive review. ACS Applied Bio Materials. 2024 May 24;7(6):3515- 3534.
48. Rane BR, Patil VL, Mhatre NR, Padave AP, Mane NP, Gavit MR, Mutkule DS, Gawade SS, Udmale AV, Chaure PP, Jain AS. Polymer-Based Vaccines. Polymers in Modern Medicine-Part 2. 2024 Dec 13:135.
49. Radhakrishnan A, Vaseeharan B, Ramasamy P, Jeyachandran S. Oral vaccination for sustainable disease prevention in aquaculture—an encapsulation approach. Aquaculture International. 2023 Apr;31(2):867-891.
50. Reed LJ, Muench H. A simple method of estimating fifty per cent endpoints. 1938: 493- 497.
51. Reyes-Cerpa S, Maisey K, Reyes-López F, Toro-Ascuy D, Sandino AM, Imarai M. Fish cytokines and immune response. New advances and contributions to fish biology. 2012 Nov 21;1.
52. Rivas-Aravena A, Sandino AM, Spencer E. Nanoparticles and microparticles of polymers and polysaccharides to administer fish vaccines. Biological research. 2013;46(4):407-419.
53. Robledo D, Taggart JB, Ireland JH, McAndrew BJ, Starkey WG, Haley CS, Hamilton A, Guy DR, Mota-Velasco JC, Gheyas AA, Tinch AE. Gene expression comparison of resistant and susceptible Atlantic salmon fry challenged with Infectious Pancreatic Necrosis virus reveals a marked contrast in immune response. BMC genomics. 2016 Dec;17:1-16.
54. Sell S. How vaccines work: immune effector mechanisms and designer vaccines. Expert review of vaccines. 2019 Oct 3;18(10):993-1015.
55. Silva BC, Martins ML, Jatobá A, Buglione Neto CC, Vieira FN, Pereira GV, Jerônimo GT, Seiffert WQ, Mouriño JL. Hematological and immunological responses of Nile tilapia after polyvalent vaccine administration by different routes. Pesquisa Veterinária Brasileira. 2009 Nov;29(11):874-80.
56. Su L, Feng Y, Wei K, Xu X, Liu R, Chen G. Carbohydrate-based macromolecular biomaterials. Chemical reviews. 2021 Aug 2;121(18):10950-1029.
57. Tapia D, Eissler Y, Reyes‐Lopez FE, Kuznar J, Yáñez JM. Infectious pancreatic necrosis virus in salmonids: Molecular epidemiology and host response to infection. Reviews in Aquaculture. 2022 Mar;14(2):751-769.
58.Ullrich KA, Schulze LL, Paap EM, Müller TM, Neurath MF, Zundler S. Immunology of IL-12: An update on functional activities and implications for disease. EXCLI journal. 2020 Dec 11;19:1563.
59. Vasquez-Martínez N, Guillen D, Moreno-Mendieta SA, Sanchez S, Rodríguez-Sanoja R. The role of mucoadhesion and mucopenetration in the immune response induced by polymer-based mucosal adjuvants. Polymers. 2023 Mar 24;15(7):1615.
60.Vimal S, Majeed SA, Nambi KS, Madan N, Farook MA, Venkatesan C, Taju G, Venu S, Subburaj R, Thirunavukkarasu AR, Hameed AS. Delivery of DNA vaccine using chitosan– tripolyphosphate (CS/TPP) nanoparticles in Asian sea bass, Lates calcarifer (Bloch, 1790) for protection against nodavirus infection. Aquaculture. 2014 Jan 15;420:240-246.
61. Vimal S, Taju G, Nambi KN, Majeed SA, Babu VS, Ravi M, Hameed AS. Synthesis and characterization of CS/TPP nanoparticles for oral delivery of gene in fish. Aquaculture. 2012 Aug 15;358:14-22.
62. Vinay TN, Bhat S, Gon Choudhury T, Paria A, Jung MH, Shivani Kallappa G, Jung SJ. Recent advances in application of nanoparticles in fish vaccine delivery. Reviews in Fisheries Science & Aquaculture. 2018 Jan 2;26(1):29-41.
63. Zhang Z, Guan C, Zhao J, Lin J, Shao Y, Li L, Lu T, Chen P, Zhang YA, Xu L. Production and evaluation of a bivalent adjuvant inactivated vaccine against infectious hematopoietic necrosis virus and infectious pancreatic necrosis virus. Aquaculture. 2025 Feb 15;597:741914.
64. Zhao Z, Peng Y, Shi X, Zhao K. Chitosan derivative composite nanoparticles as adjuvants enhance the cellular immune response via activation of the cGAS-STING pathway. International Journal of Pharmaceutics. 2023 Apr 5;636:122847.
65. Zhao T, Cai Y, Jiang Y, He X, Wei Y, Yu Y, Tian X. Vaccine adjuvants: mechanisms and platforms. Signal transduction and targeted therapy. 2023 Jul 19;8(1):283.
علوم درمانگاهی دامپزشکی ایران
دوره 19، شماره 2 - شماره پیاپی 33
پاییز و زمستان 1404
اسفند 1404
صفحه 71-84