Open Access
Research (Published online: 28-01-2018)
14. In vitro immunomodulatory potential of Artemisia indica Willd. in chicken lymphocytes
Pushpa Ruwali, Tanuj Kumar Ambwani and Pankaj Gautam
Veterinary World, 11(1): 80-87

Pushpa Ruwali: Department of Biotechnology, Graphic Era University, Dehradun - 248 002, Uttarakhand, India.
Tanuj Kumar Ambwani: Department of Veterinary Physiology and Biochemistry, College of Veterinary & Animal Sciences, G. B. Pant University of Agriculture & Technology, Pantnagar - 263 145; Uttarakhand, India.
Pankaj Gautam: Department of Biotechnology, Graphic Era University, Dehradun - 248 002, Uttarakhand, India.

doi: 10.14202/vetworld.2018.80-87

Share this article on [Facebook] [LinkedIn]

Article history: Received: 11-08-2017, Accepted: 12-12-2017, Published online: 28-01-2018

Corresponding author: Tanuj Kumar Ambwani

E-mail: tambwani@yahoo.com

Citation: Ruwali P, Ambwani TK, Gautam P (2018) In vitro immunomodulatory potential of Artemisia indica Willd. in chicken lymphocytes, Veterinary World, 11(1): 80-87.
Abstract

Aim: Evaluation of the in vitro immunomodulatory potential of Artemisia indica Willd. methanolic extract in chicken lymphocyte culture system through lymphocyte (B and T cells) proliferation assay, after standardizing the maximum non-cytotoxic dose (MNCD) in chicken lymphocytes.

Materials and Methods: Fresh aerial parts of A. indica Willd. (family: Asteraceae) specimens were collected (altitude 1560 m), gotten authenticated, processed, dried, and Soxhlet extracted to yield methanolic extract (AME). Chicken splenocytes were isolated from spleens collected from healthy birds; lymphocytes were separated by density gradient centrifugation, percentage cell viability determined and final cell count adjusted to 107 cells/ml in RPMI-1640 medium. MNCD of AME in chicken lymphocytes was determined through 3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyltetrazolium bromide dye reduction assay. Immunomodulatory potential of AME was evaluated through lymphocytes proliferation or B and T cells blastogenesis assay in the presence of appropriate mitogens, namely, lipopolysaccharide (LPS) and concanavalin A (Con A), respectively.

Results: Maximum concentration of AME exhibiting 100% cell viability (MNCD) was 200 μg/ml and was selected for further in vitro analysis. The in vitro exposure of chicken lymphocytes to 200 μg/ml dose of AME, resulted in significant (p<0.05) upregulation of 11.76% in B cell proliferation in the presence of B cell mitogen (LPS) and a significant (p<0.05) increase of 12.018% T cells proliferation in the presence of the mitogen (Con A), as compared to the control.

Conclusion: The significant upregulation in the proliferation of two major cell types modulating the immune system is an indication of the immunostimulatory potential of the plant. It would be worthwhile to further evaluate A. indica on relevant immunomodulatory aspects, especially the in vivo studies in a poultry system.

Keywords: Artemisia indica Willd., immunomodulation, in vitro chicken lymphocytes, lymphocyte proliferation assay, plant extract.

References

1. Ishizuka, M., Kawatsu, M., Yamashita, T., Ueno, M. and Takeuchi, T. (1995) Low molecular weight immunomodulators produced by microorganisms. Int. J. Immunopharmacol., 17: 133-139. [Crossref]

2. Rang, H.P., Dale, M.M., Ritter, J.M. and Moore, P.K. (2003) Pharmacology. 5th ed. Churchill Livingstone, Edinburgh.

3. Ruwali, P., Gautam, P., Ambwani, S., Dadhich, A. and Ambwani, T.K. (2013) Immunomodulatory potential of genus Artemisia. J. Immunol. Immunopathol., 15(1): 99-101.

4. Ruwali, P., Ambwani, T.K., Gautam, P. and Thapliyal, A. (2015) Qualitative and Quantitative phytochemical analysis of Artemisia indica Willd. J. Chem. Pharm. Res., 7: 942-949.

5. Ruwali, P., Ambwani, T.K., Gautam, P. and Palni, L.M.S. (2017) Artemisia indica Willd. plant as a potential source of fodder and ethnoveterinary medicine in sheep and goat. In: National Seminar on 'Improvement of Small Ruminant Production System for Livelihood Security', March 9-10, ICAR-CSWRI, Avikanagar, Rajasthan, SS4-33: 208-209.

6. Obistioiu, D., Cristina, R.T., Schmerold, I., Chizzola, R., Stolze, K., Nichita, I. and Chiurciu, V. (2014) Chemical characterization by GC-MS and in vitro activity against Candida albicans of volatile fractions prepared from Artemisia dracunculus, Artemisia abrotanum, Artemisia absinthium and Artemisia vulgaris. Chem. Cent. J., 8(6): 1-11. [Crossref]

7. Shah, N.C. (2014) The economic and medicinal Artemisia species in India. Scitech. J., 1(1): 29-38.

8. Rather, M.A., Dar, B.A., Shah, W.A., Prabhakar, A., Bindu, K., Banday, J.A. and Qurishi, M.A. (2017) Comprehensive GC-FID, GC-MS and FT-IR spectroscopic analysis of the volatile aroma constituents of Artemisia indica and Artemisia vestita essential oils. Arabian J. Chem., 10: 3798-3803. [Crossref]

9. Choi, E., Park, H., Lee, J. and Kim, G. (2013) Anticancer, antiobesity, and anti-inflammatory activity of Artemisia species in vitro. J. Tradit. Chin. Med., 33(1): 92-97. [Crossref]

10. Afsar, S.K., Kumar, R., Gopal, J. and Raveesha, P. (2013) Assessment of anti-inflammatory activity of Artemisia vulgaris leaves by cotton pellet granuloma method in wistar albino rats. J. Pharm. Res., 7: 463-467. [Crossref]

11. Kim, W., Choi, W.J., Lee, S., Kim, W.J., Lee, D.C., Sohn, U.D., Shin, H. and Kim, W. (2015) Anti-inflammatory, antioxidant and antimicrobial effects of artemisinin extracts from Artemisia annua L. Korean J. Physiol. Pharmacol., 19: 21-27. [Crossref] [PubMed] [PMC]

12. WHO. (2013) The WHO Traditional Medicine (TM) Strategy 2014-2023. World Health Organization, WHO Press, Geneva, Switzerland. p16.

13. Janossy, G. and Greaves, M.F. (1971) Lymphocyte activation: I.-Response of T and B lymphocytes to phytomitogens. Clin. Exp. Immunol., 9(4): 483-498. [PubMed] [PMC]

14. Rose, N.R. and Friedman, H. (1976) Use of cell mediated lympholysis test in transplantation immunity. Manual of Clinical Immunology. American Society of Microbiology, Washington, USA. p851-857.

15. Boyse, E.A., Old, L.J. and Chouroulinkov, I. (1964) Cytotoxic test for demonstration of mouse antibody. Methods Med. Res., 10: 39-47. [PubMed]

16. Mosmann, T. (1983) Rapid colorimetric assay for cellular growth and survival: Application to proliferation and cytotoxicity assays. J. Immunol. Methods, 65: 55-63. [Crossref]

17. Lee, C.H., Kim, J.K., Kim, H.Y., Park, S.M. and Lee, S.M. (2009) Immunomodulating effects of Korean mistletoe lectin in vitro and in vivo. Int. Immunopharmacol., 9: 1555-1561. [Crossref] [PubMed]

18. Snedecor, G.W. and Cochran, W.G. (1989) Statistical Methods. 8th ed. Iowa State University Press, Iowa, USA.

19. Boardman, P.E., Sanz-Ezquerro, J., Overton, I.M., Burt, D.W., Bosch, E., Fong, W.T., Tickle, C., Brown, W.R.A., Wilson, S.A. and Hubbard, S.J. (2002) A comprehensive collection of chicken cDNAs. Curr. Biol., 12: 1965-1969. [Crossref]

20. Ali, M.M. (2015) Emerging prospective of Indian livestock: A study on poultry industry. Asian J. Manag. Sci., 4(1): 33-39.

21. Arreola, R., Quintero-Fabian, S., Lopez-Roa, R.I., Flores-Gutierrez, E.O., Reyes-Grajeda, J.P., Carrera Quintanar, L. and Ortuno-Sahagun, D. (2015) Immunomodulation and anti-inflammatory effects of garlic compounds. J. Immunol. Res., 2015: 401630. [Crossref] [PubMed] [PMC]

22. Croteau, R., Kutchan, T.M. and Lewis, N.G. (2000) 'Natural products (secondary metabolites). In: Buchanan, B.B., Gruissem, W. and Jones, R.L., editors. 'Biochemistry and Molecular Biology of Plants', Chapter: 24, American Society of Plant Physiologists, Rockville, Maryland, USA. p1250-1318.

23. Leung, K.N., Leung, P.Y., Kong, L.P. and Leung, P.K. (2005) Immunomodulatory effects of esculetin (6,7-dihydroxycoumarin) on murine lymphocytes and peritoneal macrophages. Cell. Mol. Immunol., 2: 181-188. [PubMed]

24. Shi, C., Li, H., Yang, Y. and Hou, L. (2015) Anti-inflammatory and immunoregulatory functions of artemisinin and its derivatives. Mediators Inflamm., 2015: 435713. [Crossref] [PubMed] [PMC]

25. Ashok, P.K. and Upadhyaya, K. (2013) Evaluation of analgesic and anti-inflammatory activities of Aerial parts of Artemisia vulgaris L. in experimental animal models. J. Biol. Act. Prod. Nat., 3(1): 101-105.

26. Sagar, M.K., Ashok, P.K., Chopra, H. and Upadhyaya, K. (2010) Phytochemical and pharmacological potential of Artemisia indica in experimental models. Pharmacologyonline, 2: 1-4.

27. Li, C.R., Santoso, S. and Lo, D.D. (2007) Quantitative analysis of T cell homeostatic proliferation. Cell. Immunol., 250(1-2): 40-54. [Crossref] [PubMed] [PMC]

28. Gholamrezaie, S.L., Mohammadi, M., Jalali-Sendi, J., Abolghasemi, S.A. and Roostaie, A.M.M. (2013) Extract and leaf powder effect of Artemisia annua on performance, cellular and humoral immunity in broilers. Iran. J. Vet. Res., 14: 15-20.

29. Koo, K., Kwak, J., Lee, K., Zee, O., Woo, E., Park, H.K. and Youn, H.J. (1994) Antitumor and immunomodulating activities of the polysaccharide fractions from Artemisia selengensis and Artemisia iwayomogi. Arch. Pharm. Res., 17(5): 371-374. [Crossref]

30. Lee, K.R., Zee, O.P., Kwak, J.H., Kim, Y.S., Park, H.K., Koo, K.A. and Youn, H.J. (1993) The polysaccharide fractions of Artemisia species. Kor. J. Pharmacogn., 24: 289-295.

31. Lan, M.B., Zhang, Y.H., Zheng, Y., Yuan, H.H., Zhao, H.L. and Gao, F. (2010) Antioxidant and immunomodulatory activities of polysaccharides from Moxa (Artemisia argyi) leaf. Food Sci. Biotechnol., 19(6): 1463-1469. [Crossref]

32. Ambwani, T.K. (2013) In vitro evaluation of antioxidative, immunomodulatory and antiviral potential of Bacopa monnieri (Linn.) in Cell Culture System. PhD. Thesis, G.B. Pant University of Agriculture and Technology, Pantnagar, Uttarakhand, India.

33. Cundell, D.R. (2014) Herbal phytochemicals as immunomodulators. Curr. Immunol. Rev., 10(2): 1-18. [Crossref]

34. Fan, Y.M., Xu, L.Z., Gao, J., Wang, Y., Tang, X.H., Zhao, X.N. and Zhang, Z.X. (2004) Phytochemical and anti-inflammatory studies on Terminalia catappa. Fitoterapia, 75: 253-260. [Crossref] [PubMed]

35. Alcaraz, M.J. and Jimenez, M.I. (1988) Flavonoids as anti-inflammatory agents. Fitoterapia, 59: 25-38.

36. Bhargava, K.P., Gupta, M.B., Gupta, G.P. and Mitra, C.R. (1970) Anti-inflammatory activity of saponin and other natural products. Indian J. Med. Res., 58(6): 724-730. [PubMed]

37. Anam, E.M. (2001) Anti-inflammatory activity of compounds isolated from the aerial parts of Abrus precatorius (Fabaceae). Phytomedicine, 8: 24-27. [Crossref] [PubMed]

38. Russo, A., Izzo, A.A., Borrelli, F., Renis, M. and Vanella, A. (2003) Free radical scavenging capacity and protective effect of Bacopa monniera L. on DNA damage. Phytother. Res.,17: 870-875. [Crossref] [PubMed]

39. Bukharia, V., Charde, R. and Charde, M. (2010) Evaluation of ethanolic extract of Moringa oleifera for wound healing, anti-inflammatory and antioxidant activities on rats. In: International Conference on Folk and Herbal Medicine, November 5-27 (2010), M. S. University, Udaipur, PB-40. p180.

40. Bhati, P. and Charde, R. (2010) Antioxidant and anti-inflammatory activities of bark extract of Cassia fistula Linn. In: International Conference on Folk and Herbal Medicine, November 5-27 (2010), M. S. University, Udaipur, PB-38. p179.

41. Leong, A.C.N., Kinjo, Y., Tako, M., Iwasaki, H., Oku, H. and Tamaki, H. (2010) Flavonoid glycosides in the shoot system of okinawa taumu (Colocasia esculenta S). Food Chem., 119: 630-635. [Crossref]

42. Aruoma, O.I. (2003) Methodological considerations for characterizing potential antioxidant actions of bioactive components in food plants. Mutat. Res., 523-524: 9-20. [Crossref]

43. Valentao, P., Fernandes, E., Carvalho, F., Andrade, P.B., Seabra, R.M. and Bastos, M.L. (2002) Antioxidant activity of Hypericum androsaemum Infusion: Scavenging activity against superoxide radical, hydroxyl radical and hypochlorous acid. Biol. Pharm. Bull., 25(10): 1320-1323. [Crossref]

44. Bendich, A. (1993) Physiological role of antioxidant in the immune system. J. Dairy Sci., 76: 2789-2794. [Crossref]

45. Kong, X., Hu, Y., Rui, R., Wang, D. and Li, X. (2004) Effects of Chinese herbal medicinal ingredients on peripheral lymphocyte proliferation and serum antibody titer after vaccination in chicken. Int. Immunopharmacol., 4: 975-982. [Crossref] [PubMed]

46. Taheri, H.R., Rahmani, H.R. and Pourreza, J. (2005) Humoral immunity of broilers is affected by oil extracted propolis (OEP) in the diet. Int. J. Poult. Sci., 4: 414-417. [Crossref]

47. Ruwali, P., Ambwani, T.K. and Gautam, P. (2017) In vitro antioxidative potential of Artemisia indica willd. Indian J. Anim. Sci., 87(11): 1326-1331.

48. Oblakova, M.G., Sotirov, L.K., Lalev, M.T., Hristakieva, P., Mincheva, N., Ivanova, I., Bozakova, N.A. and Koynarski, T.S. (2015) Growth performance and natural humoral immune Status in broiler chickens treated with the immunomodulator natstim@. Int. J. Curr. Microbiol. App. Sci., 4(11): 1-7.

49. Webel, D.M., Finck, B.N., Baker, D.H. and Jonson, R.W. (1997) Time course of increased plasma cytokines, cortisone and urea nitrogen in pigs following intraperitoneal injection of lipopolisaccharide. J. Anim. Sci., 75: 1514-1520. [Crossref] [PubMed]

50. Schepetkin, I.A. and Quinn, M.T. (2006) Botanical polysaccharides: Macrophage immunomodulation and therapeutic potential. Int. Immunopharmacol., 6: 317-333. [Crossref] [PubMed]

51. Choque-Delgado, G.T., Tamashiro, W.M.S.C. and Pastore, G.M. (2010) Immunomodulatory effects of fructans. Food Res. Int., 43(5): 1231-1236. [Crossref]

52. Pereira, L., De, P., Da-Silva, R.O., Bringel, P.H., De, S.F., Da-Silva, K.E.S., Assreuy, A.M.S. and Pereira, M.G. (2012) Polysaccharide fractions of Caesalpinia ferrea pods: Potential anti-inflammatory usage. J. Ethnopharmacol., 139: 642-648. [Crossref]

53. Correa-Ferreira, M.L., Noleto, G.R. and Petkowicz, C.L.O. (2014) Artemisia absinthium and Artemisia vulgaris: A comparative study of infusion polysaccharides. Carbohydr. Polym., 102: 738-745. [Crossref] [PubMed]

54. Lee, J.A., Sung, H.N., Jeon, C.H., Gill, B.C., Oh, G.S., Youn, H.J. and Park, J.H. (2008) Carbohydrate fraction from Artemisia iwayomogi, modulate the functional differentiation of bone marrow-derived dendritic cells. Int. Immunopharmacol., 8(4): 534-541. [Crossref] [PubMed]

55. Bao, X., Yuan, H., Wang, C., Liu, J. and Lan, M. (2013) Antitumor and immunomodulatory activities of a polysaccharide from Artemisia argyi. Carbohydr. Polym., 98: 1236-1243. [Crossref] [PubMed]

56. Xie, G., Schepetkin, I.A., Siemsen, D.W., Kirpotina, L.N., Wiley, J.A. and Quinn, M.T. (2008) Fractionation and characterization of biologically-active polysaccharides from Artemisia tripartita. Phytochemistry, 69(6): 1359-1371. [Crossref] [PubMed] [PMC]

57. Hamberger, M. and Hastettman, K. (1991) Bioactivity in plants: The link between phytochemistry and medicine. Phytochemistry, 30: 3864-3874. [Crossref]

58. Bouayed, J. and Bohn, T. (2010) Exogenous antioxidants-double-edged swords in cellular redox state. Health beneficial effects at physiologic doses versus deleterious effects at high doses. Oxid. Med. Cell. Longev., 3(4): 228-237. [Crossref] [PubMed] [PMC]