Immunomodulatory effects of Radix isatidis polysaccharides <em>in vitro</em> and <em>in vivo</em>

Tao,Wei; Fu,Ting; He,Zhuo-Jing; Zhou,Han-Peng; Hong,Yan

doi:10.3892/etm.2021.10841

Immunomodulatory effects of Radix isatidis polysaccharides in vitro and in vivo

Authors:
- Wei Tao
- Ting Fu
- Zhuo-Jing He
- Han-Peng Zhou
- Yan Hong
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Affiliations: School of Bioengineering, Hangzhou Medical College, Hangzhou, Zhejiang 310013, P.R. China
Published online on: October 4, 2021 https://doi.org/10.3892/etm.2021.10841
Article Number: 1405

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Abstract

Radix isatidis (R. isatidis) is a commonly used traditional Chinese herbal medicine, which has been used for thousands of years in China and is believed to have the pharmacological properties of heat‑clearing and detoxification. Heat‑clearing and detoxification are theories of traditional Chinese medicine meaning that R. isatidis could treat febrile disease by clearing heat and reducing swelling. Polysaccharides isolated from R. isatidis by water extraction and alcohol precipitation have exhibited numerous biological activities, including antiviral and immunomodulatory effects. The present study was performed to investigate the immunomodulatory effects of water‑soluble R. isatidis polysaccharides (RIPs) on RAW264.7 macrophages and murine splenocytes, and attempt to preliminarily identify the mechanism of immunomodulation. In vitro, RIPs had a low cytotoxicity, as shown by CellTiter 96^® AQueous One Solution Cell Proliferation Assay. RAW264.7 cells treated with different concentrations of RIP displayed different morphological changes, from a round shape and aggregation to polygonal shape and dispersion in a dose‑dependent manner. In the 5 mg/ml RIP‑treated group, the changes of morphology were as same as the lipopolysaccharide‑treated group. RIP also significantly enhanced the release of nitric oxide as shown by Griess method, and the secretion of TNF‑α and IL‑6 in RAW264.7 cells was confirmed by ELISA assay. Western blotting revealed a significant increase of toll‑like receptor‑4 (TLR‑4) in RIP‑treated RAW264.7, suggesting that TLR‑4 may be associated with the immunomodulatory mechanism of RIP. Animal experiments also demonstrated through ELISA assays a significant increase in IFN‑γ and IL‑10 levels after the splenocytes of RIP‑immunized mice were stimulated by inactivated herpes simplex virus type 2. The immune function of RIP‑immunized mice was improved. The present study suggested that RIP could be potentially used as a novel immunomodulator.

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1	Tzianabos AO: Polysaccharide immunomodulators as therapeutic agents: structural aspects and biologic function. Clin Microbiol Rev. 13:523–533. 2000.PubMed/NCBI View Article : Google Scholar
2	Murray HW: Current and future clinical applications of interferon-gamma in host antimicrobial defense. Intensive Care Med. 22 (Suppl 4):S456–S461. 1996.PubMed/NCBI View Article : Google Scholar
3	Nemunaitis J: A comparative review of colony-stimulating factors. Drugs. 54:709–729. 1997.PubMed/NCBI View Article : Google Scholar
4	Hamilton JA and Anderson GP: GM-CSF biology. Growth Factors. 22:225–231. 2004.PubMed/NCBI View Article : Google Scholar
5	Singh N and Bhattacharyya D: Collagenases in an ether extract of bacterial metabolites used as an immunostimulator induces TNF-α and IFN-γ. Int Immunopharmacol. 23:211–221. 2014.PubMed/NCBI View Article : Google Scholar
6	Wasser SP: Medicinal mushrooms as a source of antitumor and immunomodulating polysaccharides. Appl Microbiol Biotechnol. 60:258–274. 2002.PubMed/NCBI View Article : Google Scholar
7	Li XY: Immunomodulating Chinese herbal medicines. Mem Inst Oswaldo Cruz. 86 (Suppl 2):S159–S164. 1991.PubMed/NCBI View Article : Google Scholar
8	Wei W, Xiao HT, Bao WR, Ma DL, Leung CH, Han XQ, Ko CH, Lau CB, Wong CK, Fung KP, et al: TLR-4 may mediate signaling pathways of Astragalus polysaccharide RAP induced cytokine expression of RAW264.7 cells. J Ethnopharmacol. 179:243–252. 2016.PubMed/NCBI View Article : Google Scholar
9	Hou C, Chen L, Yang L and Ji X: An insight into anti-inflammatory effects of natural polysaccharides. Int J Biol Macromol. 153:248–255. 2020.PubMed/NCBI View Article : Google Scholar
10	Chen X, Han W, Wang G and Zhao X: Application prospect of polysaccharides in the development of anti-novel coronavirus drugs and vaccines. Int J Biol Macromol. 164:331–343. 2020.PubMed/NCBI View Article : Google Scholar
11	Cai Z, Li W, Wang H, Yan W, Zhou Y, Wang G, Cui J and Wang F: Anti-tumor and immunomodulating activities of a polysaccharide from the root of Sanguisorba officinalis L. Int J Biol Macromol. 51:484–488. 2012.PubMed/NCBI View Article : Google Scholar
12	Yu Y, Shen M, Song Q and Xie J: Biological activities and pharmaceutical applications of polysaccharide from natural resources: A review. Carbohydr Polym. 183:91–101. 2018.PubMed/NCBI View Article : Google Scholar
13	Zhou W and Zhang XY: Research progress of Chinese herbal medicine Radix isatidis (banlangen). Am J Chin Med. 41:743–764. 2013.PubMed/NCBI View Article : Google Scholar
14	Sun B, Yu S, Zhao D, Guo S, Wang X and Zhao K: Polysaccharides as vaccine adjuvants. Vaccine. 36:5226–5234. 2018.PubMed/NCBI View Article : Google Scholar
15	Li Z, Li L, Zhou H, Zeng L, Chen T, Chen Q, Zhou B, Wang Y, Chen Q, Hu P and Yang Z: Radix isatidis polysaccharides inhibit influenza a virus and influenza A virus-induced inflammation via suppression of host TLR3 signaling in vitro. Molecules. 22(116)2017.PubMed/NCBI View Article : Google Scholar
16	Wei ZY, Wang XB, Zhang HY, Yang CH, Wang YB, Xu DH, Chen HY and Cui BA: Inhibitory effects of indigowoad root polysaccharides on porcine reproductive and respiratory syndrome virus replication in vitro. Antivir Ther. 16:357–363. 2011.PubMed/NCBI View Article : Google Scholar
17	Wang T, Wang X, Zhuo Y, Si C, Yang L, Meng L and Zhu B: Antiviral activity of a polysaccharide from Radix isatidis (Isatis indigotica Fortune) against hepatitis B virus (HBV) in vitro via activation of JAK/STAT signal pathway. J Ethnopharmacol. 257(112782)2020.PubMed/NCBI View Article : Google Scholar
18	Liao HF, Lu MC, Chang HC, Wei CC, Kao CH, Chen ZH, Huang CC and Li C: Effects of herbal medicinal formulas on suppressing viral replication and modulating immune responses. Am J Chin Med. 38:173–190. 2010.PubMed/NCBI View Article : Google Scholar
19	Zhao YL, Wang JB, Shan LM, Jin C, Ma L and Xiao XH: Effect of Radix isatidis polysaccharides on immunological function and expression of immune related cytokines in mice. Chin J Integr Med. 14:207–211. 2008.PubMed/NCBI View Article : Google Scholar
20	Ma L, Feng SH, Tang JY, Zhao YL and Xiao XH: Influence of Radix isatidis polysaccharides produced by different preparation technologies on proliferation function of mouse splenic lymphocyte. China Pharm (Chin). 18:169–171. 2007.
21	Wang X, Chen Z, Chen T, Li X, Huang S, Wang H, Tong C and Liu F: Isatis root polysaccharide promotes maturation and secretory function of monocyte-derived dendritic cells. BMC Complement Med Ther. 20(301)2020.PubMed/NCBI View Article : Google Scholar
22	Du Z, Liu H, Zhang Z and Li P: Antioxidant and anti-inflammatory activities of Radix isatidis polysaccharide in murine alveolar macrophages. Int J Biol Macromol. 58:329–335. 2013.PubMed/NCBI View Article : Google Scholar
23	Lian ZY, Hou JS, Sun GR, Zhao BS and Xin GH: Radix isatidis polysaccharide enhances killing effect of NK cells on esophageal cancer cells by promoting expression of NKG2D ligand and its mechanism. Chin J Immunol. 36:965–970. 2020.
24	Li JP, Zhu GH, Yuan Y and Liu MX: Anti-tumor and immune function regulation effects of Radix isatidis polysaccharides in vivo. Nat Prod Res Dev. 29:2010–2016. 2017.
25	Shan J, Sun G, Ren J, Zhu T, Jia P, Qu W, Li Q, Wu J, Ma H, Wen S and Wang Y: An α-glucan isolated from root of Isatis indigotica, its structure and adjuvant activity. Glycoconj J. 31:317–326. 2014.PubMed/NCBI View Article : Google Scholar
26	Zhang W, Zheng X, Cheng N, Gai W, Xue X, Wang Y, Gao Y, Shan J, Yang S and Xia X: Isatis indigotica root polysaccharides as adjuvants for an inactivated rabies virus vaccine. Int J Biol Macromol. 87:7–15. 2016.PubMed/NCBI View Article : Google Scholar
27	Zhou HP, Tao W, Fu T, He ZJ and Hong Y: Influence of isatis root polysaccharide on immune effect of herpes simplex virus type-2 DNA vaccine. Chin J Biol. 33:609–613. 2020.(In Chinese).
28	Lim KH and Staudt LM: Toll-like receptor signaling. Cold Spring Harb Perspect Biol. 5(a011247)2013.PubMed/NCBI View Article : Google Scholar
29	Yang X, Zhao Y, Wang H and Mei Q: Macrophage activation by an acidic polysaccharide isolated from Angelica sinensis (Oliv.) diels. J Biochem Mol Biol. 40:636–643. 2007.PubMed/NCBI View Article : Google Scholar
30	Masuko T, Minami A, Iwasaki N, Majima T, Nishimura S and Lee YC: Carbohydrate analysis by a phenol-sulfuric acid method in microplate format. Anal Biochem. 339:69–72. 2005.PubMed/NCBI View Article : Google Scholar
31	Zhu H, Ding X, Hou Y, Li Y and Wang M: Structure elucidation and bioactivities of a new polysaccharide from Xiaojin boletus speciosus frost. Int J Biol Macromol. 126:697–716. 2019.PubMed/NCBI View Article : Google Scholar
32	Lilley E, Andrews MR, Bradbury EJ, Elliott H, Hawkins P, Ichiyama RM, Keeley J, Michael-Titus AT, Moon LDF, Pluchino S, et al: Refining rodent models of spinal cord injury. Exp Neurol. 328(113273)2020.PubMed/NCBI View Article : Google Scholar
33	Gao B, Li L, Zhu P, Zhang M, Hou L, Sun Y, Liu X, Peng X and Gu Y: Chronic administration of methamphetamine promotes atherosclerosis formation in ApoE-/- knockout mice fed normal diet. Atherosclerosis. 243:268–277. 2015.PubMed/NCBI View Article : Google Scholar
34	Wynn TA and Vannella KM: Macrophages in tissue repair, regeneration, and fibrosis. Immunity. 44:450–462. 2016.PubMed/NCBI View Article : Google Scholar
35	Lee JS, Kwon DS, Lee KR, Park JM, Ha SJ and Hong EK: Mechanism of macrophage activation induced by polysaccharide from Cordyceps militaris culture broth. Carbohydr Polym. 120:29–37. 2015.PubMed/NCBI View Article : Google Scholar
36	Yang F, Li X, Yang Y, Ayivi-Tosuh SM, Wang F, Li H and Wang G: A polysaccharide isolated from the fruits of physalis alkekengi L. induces RAW264.7 macrophages activation via TLR2 and TLR4-mediated MAPK and NF-κB signaling pathways. Int J Biol Macromol. 140:895–906. 2019.PubMed/NCBI View Article : Google Scholar
37	Liu X, Xie J, Jia S, Huang L, Wang Z, Li C and Xie M: Immunomodulatory effects of an acetylated cyclocarya paliurus polysaccharide on murine macrophages RAW264.7. Int J Biol Macromol. 98:576–581. 2017.PubMed/NCBI View Article : Google Scholar
38	Zheng P, Fan W, Wang S, Hao P, Wang Y, Wan H, Hao Z, Liu J and Zhao X: Characterization of polysaccharides extracted from platycodon grandiflorus (Jacq.) A.DC. affecting activation of chicken peritoneal macrophages. Int J Biol Macromol. 96:775–785. 2017.PubMed/NCBI View Article : Google Scholar
39	Zhang M, Tian X, Wang Y, Wang D, Li W, Chen L, Pan W, Mehmood S and Chen Y: Immunomodulating activity of the polysaccharide TLH-3 from Tricholomalobayense in RAW264.7 macrophages. Int J Biol Macromol. 107:2679–2685. 2018.PubMed/NCBI View Article : Google Scholar
40	Škrnjug I, Guzmán CA and Rueckert C: Cyclic GMP-AMP displays mucosal adjuvant activity in mice. PLoS One. 9(e110150)2014.PubMed/NCBI View Article : Google Scholar
41	Ning H, Zhang W, Kang J, Ding T, Liang X, Lu Y, Guo C, Sun W, Wang H, Bai Y and Shen L: Subunit vaccine ESAT-6:c-di-AMP delivered by intranasal route elicits immune responses and protects against mycobacterium tuberculosis infection. Front Cell Infect Microbiol. 11(647220)2021.PubMed/NCBI View Article : Google Scholar
42	Schroder K, Hertzog PJ, Ravasi T and Hume DA: Interferon-gamma: An overview of signals, mechanisms and functions. J Leukoc Biol. 75:163–189. 2004.PubMed/NCBI View Article : Google Scholar
43	Levy Y and Brouet JC: Interleukin-10 prevents spontaneous death of germinal center B cells by induction of the bcl-2 protein. J Clin Invest. 93:424–428. 1994.PubMed/NCBI View Article : Google Scholar
44	Rousset F, Garcia E, Defrance T, Péronne C, Vezzio N, Hsu DH, Kastelein R, Moore KW and Banchereau J: Interleukin 10 is a potent growth and differentiation factor for activated human B lymphocytes. Proc Natl Acad Sci USA. 89:1890–1893. 1992.PubMed/NCBI View Article : Google Scholar
45	Rousset F, Peyrol S, Garcia E, Vezzio N, Andujar M, Grimaud JA and Banchereau J: Long-term cultured CD40-activated B lymphocytes differentiate into plasma cells in response to IL-10 but not IL-4. Int Immunol. 7:1243–1253. 1995.PubMed/NCBI View Article : Google Scholar
46	Wang X, Wong K, Ouyang W and Rutz S: Targeting IL-10 family cytokines for the treatment of human diseases. Cold Spring Harb Perspect Biol. 11(a028548)2019.PubMed/NCBI View Article : Google Scholar
47	Mazer M, Unsinger J, Drewry A, Walton A, Osborne D, Blood T, Hotchkiss R and Remy KE: IL-10 has differential effects on the innate and adaptive immune systems of septic patients. J Immunol. 203:2088–2099. 2019.PubMed/NCBI View Article : Google Scholar
48	Davinelli S, Melvang HM, Andersen LP, Scapagnini G and Nielsen ME: Astaxanthin from shrimp cephalothorax stimulates the immune response by enhancing IFN-γ, IL-10, and IL-2 secretion in splenocytes of helicobacter pylori-infected mice. Mar Drugs. 17(382)2019.PubMed/NCBI View Article : Google Scholar
49	Lin HY, Juan SH, Shen SC, Hsu FL and Chen YC: Inhibition of lipopolysaccharide-induced nitric oxide production by flavonoids in RAW264.7 macrophages involves heme oxygenase-1. Biochem Pharmacol. 66:1821–32. 2003.PubMed/NCBI View Article : Google Scholar
50	Means TK, Lien E, Yoshimura A, Wang S, Golenbock DT and Fenton MJ: The CD14 ligands lipoarabinomannan and lipopolysaccharide differ in their requirement for toll-like receptors. J Immunol. 163:6748–6755. 1999.PubMed/NCBI
51	Pi J, Li T, Liu J, Su X, Wang R, Yang F, Bai H, Jin H and Cai J: Detection of lipopolysaccharide induced inflammatory responses in RAW264.7 macrophages using atomic force microscope. Micron. 65:1–9. 2014.PubMed/NCBI View Article : Google Scholar
52	Yin M, Zhang Y and Li H: Advances in research on immunoregulation of macrophages by plant polysaccharides. Front Immunol. 10(145)2019.PubMed/NCBI View Article : Google Scholar