Abstract
Aim:
Recent evidence shows that cultured mycelium Cordyceps sinensis (CMCS) effectively protects against liver fibrosis in mice. Here, we investigated whether the anti-fibrotic action of CMCS was related to its regulation of the activity of hepatic natural killer (NK) cells in CCl4-treated mice.
Methods:
C57BL/6 mice were injected with 10% CCl4 (2 mL/kg, ip) 3 times per week for 4 weeks, and received CMCS (120 mg·kg−1·d−1, ig) during this period. In another part of experiments, the mice were also injected with an NK cell-deleting antibody ASGM-1 (20 μg, ip) 5 times in the first 3 weeks. After the mice were sacrificed, serum liver function, and liver inflammation, hydroxyproline content and collagen deposition were assessed. The numbers of hepatic NK cells and expression of NKG2D (activation receptor of NK cells) on isolated liver lymphocytes were analyzed using flow cytometry. Desmin expression and cell apoptosis in liver tissues were studied using desmin staining and TUNEL assay, respectively. The levels of α-SMA, TGF-β, RAE-1δ and RAE-1ε in liver tissues were determined by RT-qPCR.
Results:
In CCl4-treated mice, CMCS administration significantly improved liver function, attenuated liver inflammation and fibrosis, and increased the numbers of hepatic NK cells and expression level of NKG2D on hepatic NK cells. Furthermore, CMCS administration significantly decreased desmin expression in liver tissues, and increased TUNEL staining adjacent to hepatic stellate cells. Injection with NK cell-deleting ASGM-1 not only diminished the numbers of hepatic NK cells, but also greatly accelerated liver inflammation and fibrosis in CCl4-treated mice. In CCl4-treated mice with NK cell depletion, CMCS administration decelerated the rate of liver fibrosis development, and mildly upregulated the numbers of hepatic NK cells but without changing NKG2D expression.
Conclusion:
CMCS allevi¬ates CCl4-induced liver inflammation and fibrosis via promoting activation of hepatic NK cells. CMCS partially reverses ASGM-1-induced depletion of hepatic NK cells.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
References
Elsharkawy AM, Oakley F, Mann DA . The role and regulation of hepatic stellate cell apoptosis in reversal of liver fibrosis. Apoptosis 2005; 10: 927–39.
Pinzani M, Macias-Barragan J . Update on the pathophysiology of liver fibrosis. Expert Rev Gastroenterol Hepatol 2010; 4: 459–72.
Issa R, Zhou X, Constandinou CM, Fallowfield J, Millward-Sadler H, Gaca MD, et al. Spontaneous recovery from micronodular cirrhosis: evidence for incomplete resolution associated with matrix cross-linking. Gastroenterology 2004; 126: 1795–808.
Farag SS, Caligiuri MA . Human natural killer cell development and biology. Blood Rev 2006; 20: 123–37.
Tian Z, Chen Y, Gao B . Natural killer cells in liver disease. Hepatology 2013; 57: 1654–62.
Lanier LL . NK cell recognition. Annu Rev Immunol 2005; 23: 225–74.
Raulet DH . Roles of the NKG2D immunoreceptor and its ligands. Nat Rev Immunol 2003; 3: 781–90.
Ortaldo JR, Young HA . Mouse Ly49 NK receptors: balancing activation and inhibition. Mol Immunol 2005; 42: 445–50.
Ishiyama K, Ohdan H, Ohira M, Mitsuta H, Arihiro K, Asahara T . Difference in cytotoxicity against hepatocellular carcinoma between liver and periphery natural killer cells in humans. Hepatology 2006; 43: 362–72.
Gur C, Doron S, Kfir-Erenfeld S, Horwitz E, Abu-Tair L, Safadi R, et al. NKp46-mediated killing of human and mouse hepatic stellate cells attenuates liver fibrosis. Gut 2012; 61: 885–93.
Melhem A, Muhanna N, Bishara A, Alvarez CE, Ilan Y, Bishara T, et al. Anti-fibrotic activity of NK cells in experimental liver injury through killing of activated HSC. J Hepatol 2006; 45: 60–71.
Radaeva S, Sun R, Jaruga B, Nguyen VT, Tian Z, Gao B . Natural killer cells ameliorate liver fibrosis by killing activated stellate cells in NKG2D-dependent and tumor necrosis factor-related apoptosis-inducing ligand-dependent manners. Gastroenterology 2006; 130: 435–52.
Jeong WI, Park O, Radaeva S, Gao B . STAT1 inhibits liver fibrosis in mice by inhibiting stellate cell proliferation and stimulating NK cell cytotoxicity. Hepatology 2006; 44: 1441–51.
Yoshida K, Ohishi W, Nakashima E, Fujiwara S, Akahoshi M, Kasagi F, et al. Lymphocyte subset characterization associated with persistent hepatitis C virus infection and subsequent progression of liver fibrosis. Hum Immunol 2011; 72: 821–6.
Gao B, Radaeva S . Natural killer and natural killer T cells in liver fibrosis. Biochim Biophys Acta 2013; 1832: 1061–9.
Jeong WI, Park O, Suh YG, Byun JS, Park SY, Choi E, et al. Suppression of innate immunity (natural killer cell/interferon-gamma) in the advanced stages of liver fibrosis in mice. Hepatology 2011; 53: 1342–51.
Won SY, Park EH . Anti-inflammatory and related pharmacological activities of cultured mycelia and fruiting bodies of Cordyceps militaris. J Ethnopharmacol 2005; 96: 555–61.
Lee HH, Park H, Sung GH, Lee K, Lee T, Lee I, et al. Anti-influenza effect of Cordyceps militaris through immunomodulation in a DBA/2 mouse model. J Microbiol 2014; 52: 696–701.
Yoon TJ, Yu KW, Shin KS, Suh HJ . Innate immune stimulation of exo-polymers prepared from Cordyceps sinensis by submerged culture. Appl Microbiol Biotechnol 2008; 80: 1087–93.
Liu C, Lu S, Ji MR . Effects of Cordyceps sinensis (CS) on in vitro natural killer cells. Zhongguo Zhong Xi Yi Jie He Za Zhi 1992; 12: 267–9, 59. Chinese.
Peng Y, Chen Q, Yang T, Tao Y, Lu X, Liu C . Cultured mycelium Cordyceps sinensis protects liver sinusoidal endothelial cells in acute liver injured mice. Mol Biol Rep 2014; 41: 1815–27.
Peng Y, Tao Y, Wang Q, Shen L, Yang T, Liu Z, et al. Ergosterol Is the Active Compound of Cultured Mycelium Cordyceps sinensis on Antiliver Fibrosis. Evid Based Complement Alternat Med 2014; 2014: 537234.
Habu S, Fukui H, Shimamura K, Kasai M, Nagai Y, Okumura K, et al. In vivo effects of anti-asialo GM1. I. Reduction of NK activity and enhancement of transplanted tumor growth in nude mice. J Immunol 1981; 127: 34–8.
Jamall IS, Finelli VN, Que Hee SS . A simple method to determine nanogram levels of 4-hydroxyproline in biological tissues. Anal Biochem 1981; 112: 70–5.
Safadi R, Ohta M, Alvarez CE, Fiel MI, Bansal M, Mehal WZ, et al. Immune stimulation of hepatic fibrogenesis by CD8 cells and attenuation by transgenic interleukin-10 from hepatocytes. Gastroenterology 2004; 127: 870–82.
Kataoka S, Konishi Y, Nishio Y, Fujikawa-Adachi K, Tominaga A . Antitumor activity of eosinophils activated by IL-5 and eotaxin against hepatocellular carcinoma. DNA Cell Biol 2004; 23: 549–60.
Nishikado H, Mukai K, Kawano Y, Minegishi Y, Karasuyama H . NK cell-depleting anti-asialo GM1 antibody exhibits a lethal off-target effect on basophils in vivo. J Immunol 2011; 186: 5766–71.
Slifka MK, Pagarigan RR, Whitton JL . NK markers are expressed on a high percentage of virus-specific CD8+ and CD4+ T cells. J Immunol 2000; 164: 2009–15.
Trambley J, Bingaman AW, Lin A, Elwood ET, Waitze SY, Ha J, et al. Asialo GM1(+) CD8(+) T cells play a critical role in costimulation blockade-resistant allograft rejection. J Clin Invest 1999; 104: 1715–22.
Wiltrout RH, Santoni A, Peterson ES, Knott DC, Overton WR, Herberman RB, et al. Reactivity of anti-asialo GM1 serum with tumoricidal and non-tumoricidal mouse macrophages. J Leukoc Biol 1985; 37: 597–614.
Kasai M, Iwamori M, Nagai Y, Okumura K, Tada T . A glycolipid on the surface of mouse natural killer cells. Eur J Immunol 1980; 10: 175–80.
Eckelhart E, Warsch W, Zebedin E, Simma O, Stoiber D, Kolbe T, et al. A novel Ncr1-Cre mouse reveals the essential role of STAT5 for NK-cell survival and development. Blood 2011; 117: 1565–73.
Jessen B, Maul-Pavicic A, Ufheil H, Vraetz T, Enders A, Lehmberg K, et al. Subtle differences in CTL cytotoxicity determine susceptibility to hemophagocytic lymphohistiocytosis in mice and humans with Chediak-Higashi syndrome. Blood 2011; 118: 4620–9.
Gao B, Radaeva S, Park O . Liver natural killer and natural killer T cells: immunobiology and emerging roles in liver diseases. J Leukoc Biol 2009; 86: 513–28.
Fowell AJ, Iredale JP . Emerging therapies for liver fibrosis. Dig Dis 2006; 24: 174–83.
Matheson DS, Green BJ, Minuk GY . Natural killer-cell activity and the response to interferons alpha, beta, and gamma in patients with primary biliary cirrhosis. J Allergy Clin Immunol 1989; 84: 214–8.
Friedman SL . Hepatic stellate cells: protean, multifunctional, and enigmatic cells of the liver. Physiol Rev 2008; 88: 125–72.
Acknowledgements
This work was supported by grants from the National Natural Science Foundation of China (No 81173405, 81102701 and 81503322), the National Science and Technology Major Project (2014ZX10005001), the International S&T Cooperation Program of China (2014DFA31440), the Program for Outstanding Medical Academic Leader of Shanghai Municipality (LJ10005), the E-Institute (E03008), “Three-year action plan” for Development of TCM in Shanghai (ZY3-CCCX-2-1003) and China Postdoctoral Science Foundation (No 2015M571595).
Author information
Authors and Affiliations
Corresponding authors
Rights and permissions
About this article
Cite this article
Peng, Y., Huang, K., Shen, L. et al. Cultured Mycelium Cordyceps sinensis allevi¬ates CCl4-induced liver inflammation and fibrosis in mice by activating hepatic natural killer cells. Acta Pharmacol Sin 37, 204–216 (2016). https://doi.org/10.1038/aps.2015.129
Received:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/aps.2015.129
