Study on Echinacea Extract for Animal Immune System

Echinacea, also known as “golden chrysanthemum”, belongs to the Asteraceae family and is a perennial plant that originated in eastern North America and has been used as a medicinal plant in North America for hundreds of years, and was first used for the treatment of colds and upper respiratory tract infections [1-2]. There are nine species of Echinacea, three of which have been widely used in medicine, namely, Echinacea purpurea (purple-flowered Echinacea), Echinacea angustifolia (narrow-leaved Echinacea) and Echinacea pallida (white-flowered Echinacea) [3]. According to statistics, Echinacea purpurea is the best-selling herbal plant in the U.S. and European markets, accounting for 10% of the total herbal market [1, 4]. In recent years, the application of Echinacea and its extracts has gradually shifted from clinical treatment to animal production, and the results of a large number of in vivo and in vitro experiments have shown that Echinacea and its extracts mainly exert anti-inflammatory, anti-bacterial, and anti-viral effects by enhancing the immunity of the body.

In recent years, Echinacea and its extracts have been used as a kind of feed additives in livestock and poultry production, and their main effects are reflected in the improvement of animal feed intake, so that the relative increase in body weight, enhancement of the survival rate of the born piglets and reduction of the occurrence of deformities, enhancement of the resistance to disease and reduction of unnecessary deaths of the animals, anti-bacterial and de-worming, protection of livestock and poultry health, and improvement of the quality of animal products and so on［5-6］. -However, Echinacea purpurea and its extracts have been shown to be effective in improving the quality of animal products. However, these effects of Echinacea purpurea and its extracts are reflected to different degrees in the regulation of immune function. In this paper, we present a brief overview of the studies on Echinacea and its extracts in regulating the immune function in recent years, in order to provide a reference for the study of the mechanism of Echinacea and its extracts and to provide a scientific basis for the popularization and application of Echinacea and its extracts in animal husbandry production.

1 Effects of Echinacea and its extracts on the immune function of animals

1.1 In vivo study

Under normal feeding conditions, the addition of Echinacea or its extracts to animal diets has the effect of regulating their immune function. Aly et al. [7] added 0.25 mg/kg of Echinacea extract to the feed of tilapia, which significantly increased daily weight gain and blood lysozyme activity. Oskoii et al. [8] added 0.25% or 0.50% Echinacea extract to the feed of rainbow trout. Oskoii et al. [8] added 0.25% or 0.50% Echinacea to rainbow trout diets and increased the number of leukocytes and lymphocytes in the blood of rainbow trout, and Arafa et al. [9] showed that feeding Echinacea-supplemented diets to weaned rabbits reduced their mortality rate, which was related to the enhancement of the immune function and the antioxidant defense system of the rabbits, and Ebrahimi et al. [10] used broilers fed diets supplemented with organic acids, probiotics, or Echinacea. Ebrahimi et al. [10] studied broiler chickens fed diets supplemented with organic acids, probiotics or Echinacea extracts. The results showed that the relative weights of spleen and bursa of the supplemented groups increased compared with those of the control group, and the Echinacea group had the best effect. Hao Zhi et al. [11] showed that Echinacea extract could improve the growth performance of broiler chickens, and at the same time increase the index of immune organs.

Niu Xiaofei et al. [12] showed that Echinacea extract could significantly increase the antibody titer of bursa virus and cytokine (e.g., interleukin-2 (IL-2), tumor necrosis factor-α (TNF-α)) content in the peripheral blood of chickens, and improve the production performance of broiler chickens. The results of the study showed that the Ni Yaodi et al. [13] found that Echinacea can increase the activity of non-specific T cells, the content of CD4+ T lymphocytes and the ratio of CD4+/CD8+ T lymphocyte subpopulations in peripheral blood of chicks, which is a kind of immunomodulatory agent with obvious effects. Song Hongwei et al. [14] showed that Echinacea extract is an effective immunomodulator, which can promote the development of immune organs in broiler chicks and increase the production of Newcastle Disease antibodies in the peripheral blood, and Kim et al. [15] found that feeding Echinacea-added diets to BALB/c mice could enhance the function of antigen-presenting cells by decreasing the number and function of regulatory T-cells in the spleen. Wu et al. [16] used Kunming mice in a carbon profile test and showed that 0.025 μg/g of Echinacea purpurea chloroform extract alleviated the inhibitory effect of hydrocortisone immunization on the function of monocyte-macrophage system in Kunming mice. Li Wan et al. [17] showed that Echinacea treatment could significantly increase the spleen index of immunosuppressed mice.

In addition, the addition of Echinacea or its extracts to diets can also improve the immune function of animals infected or attacked by diseases, and Pourgholam et al [18] showed that the addition of Echinacea extracts to diets could enhance the immune function of rainbow trout infected with streptococci, and El-Asely et al [19] found that the addition of Echinacea extracts to diets could enhance the immune function of tilapia fed in water containing Aeromonas hydrophilus. El-Asely et al. Qin Zhihua et al. [20] showed that Echinacea extract could reduce the mortality of turbot after the attack of Edwardsiella tarda, which was related to its ability to enhance the non-specific immune function. Ren Yonglin [21] showed that Echinacea purpurea extract could effectively enhance the immune function of carp and produce strong resistance to Aeromonas hydrophila. Xiao Zhengzhong et al. [22] showed in a study on mice injected intraperitoneally with Escherichia coli liquid that Echinacea purpurea significantly enhanced the immune function of mice, thus improving their resistance to E. coli infection.

1.2 In vitro studies

The regulation of immune function by Echinacea and its extracts has not only been demonstrated in in vivo tests, but also confirmed in in vitro tests. Sasagawa et al. [23] found that Echinacea extracts could inhibit the production of IL-2 when treating the blood T cells of leukemia patients and the inhibition had a dose-dependent effect. In a study on murine peritoneal macrophages, Zhai et al. [24] and Sullivan et al. [25] showed that treatment with Echinacea extract activated the innate immune response and regulated the production of interleukin-6, TNF-α, interleukin-12 and nitric oxide (niO) in the blood T cells of rats. Nitric oxide (NO) production; similar results were obtained by Yu et al. in the study of chicken peripheral blood mononuclear cells and murine macrophage cell line RAW264.7.

Classen et al. [27] showed that Echinacea extract had an effect on immunoglobulin M (IgM) production in mouse lymphocytes and stimulated the production of NO and IL-6 in mouse alveolar macrophages in a dose-dependent manner, and Groom et al. [28] found that Echinacea extract increased phagocytosis and interferon-γ synthesis in macrophages. Fast et al. [29] showed that Echinacea extract inhibited Pam3csk4-induced TNF-α secretion in the human leukemia cell line THP-1, and Sharma et al. [30-31] showed that Echinacea extract mitigated the attack-induced TNF-α secretion in human bronchial epithelial cells and skin tissues in an in vitro study of Propionibacterium acnes and rhinovirus attack. Echinacea extract attenuated the production of inflammatory cytokines in human bronchial epithelial cells and skin fibroblasts induced by viral attack and inhibited the proliferation of pathogenic bacteria.

The results of in vivo and in vitro studies show that Echinacea purpurea and its extracts have the ability to regulate the production of inflammatory factors and gonadotropins and enhance the production of antibodies, which helps to improve the immune function of the animal body, thereby increasing the survival rate, reducing the mortality rate and increasing the daily weight gain of the animal. In addition, Echinacea and its extracts can also produce a certain degree of immunoprotection for poisonous and immunosuppressed animals, that is to say, they can produce resistance to pathogenic bacteria and have an immune-enhancing effect.

2 Mechanisms of Echinacea and its extracts in regulating immune function

Analyzing the results of relevant studies in recent years, Echinacea and its extracts may regulate the immune function of the organism through the following three pathways: 1) activating the amplification of signaling mechanism and up-regulating the function of macrophages through the dependence mechanism of Toll-like receptor 4 (TLR4); 2) activating the amplification of signaling mechanism and up-regulating the function of macrophages through the dependence mechanism of TLR4; 3) activating the amplification of signaling mechanism and up-regulation of macrophages through the dependence mechanism of TLR4. Sullivan et al. [25] conducted a study on murine peritoneal macrophages, and the results showed that Echinacea polysaccharides have the ability to activate macrophage effector functions, and in this process, Echinacea polysaccharides mainly act as a ligand to bind to TLR4 on the surface of macrophages, which leads to the upregulation of the signaling pathway related proteins (e.g., extracellular regulated protein kinases (MAPK)) in the cell. The activation of proteins related to the signaling pathway (e.g., extracellular regulated protein kinase (ERK), p38, and amino-terminal kinase (c-Jun N-terminal kinase (JNK)), which ultimately activates nuclear factor-κB, was also found to be a TLR4-independent process, as well as a TLR4-independent process, which was found to be a TLR4-independent process. TLR4-independent mechanism was also found in this process.

Xu Xin [32] obtained similar results on mouse macrophages and further found that this process also involves the articulation of Toll-like receptors (TLRs), myeloid differentiation factor 88 (MyD88) and Toll/interleukin-1 receptor structure. The molecular myeloid differentiation factor 88 (MyD88) and the Toll/interleukin-1 receptor domain-containing adaptor inducing interferon-β (TRIF). 2) can act as ligands with cannabinoid receptors (cannabinoid receptor-2, TRIF). TRIF (2) binds to cannabinoid receptor-2 (CB2) as a ligand and activates related signaling pathways, thus exerting its immunomodulatory effects.

A study by Gertsch et al. [33] using Echinacea extracts to treat peripheral blood mononuclear cells showed that Echinacea extracts regulate immunity by binding to CB2, activating cyclic adenosine monophosphate (cAMP), p38MAPK and JNK pathways, increasing NF-κB and activating the transcription factor (transcriptional factor) -2/cyclic adenosine monophosphate response (transcriptional factor). The activation of cyclic adenosine monophosphate (cAMP), p38MAPK and JNK pathways, and the increase of NF-κB and acti- vating transcription factor-2 / cAMP-response element binding protein-1 (ATF-2 / CREB-1) activities, were achieved by the activation of NF-κB. The activity of ATF-2 / CREB-1 is realized by the activity of ATF-2 / CREB-1. Similar results were obtained by Chicca et al. [34] using the human embryonic kidney cell line HEK. 3) Influence immune function by regulating the expression of cellular immune-related genes. Wang et al. [35] used a targeted functional DNA microsequencing system to test the effects of Echinacea extracts from different parts of Echinacea purpurea on dendritic cells, and the results showed that Echinacea purpurea extracts from different parts of Echinacea purpurea affected the expression of CD83, CD34, and chemokines (CCL2, CCL3, CCL8, CDIA, CCR1, CCR9, etc). Brush et al. [36] also showed that Echinacea purpurea extracts had a similar effect. Brush et al. [36] also showed that Echinacea extract could increase the expression of CD69 in T cells.

In conclusion, innate immunity is the first line of defense of the host against foreign pathogens, in which TLRs are one of the keys to immunoregulation, and TLR4 is one of the most representative members of the TLR family, which is capable of recognizing exogenous pathogen molecules, especially Gram-negative molecules, and plays an important role in the natural immunity of the organism [37]. CB2 is mainly distributed in immune tissues and cells, with only a small amount of expression in the central nervous system, which is mainly involved in immunomodulation and related to neurodegenerative diseases [38]. Echinacea and its extracts can act as ligands to bind to receptors (TLR4 and CB2), activating their downstream pathways and causing a series of immune responses. In addition, Echinacea and its extracts can also regulate the expression of immunity-related genes and enhance the body's defense function, thus preventing the invasion of harmful pathogens in a rapid and timely manner.

3 Conclusion

In summary, Echinacea and its extracts can regulate the immune function both in vitro and in vivo, and they can be used as feed additives to improve the immune function of animals, enhance their resistance to adversity and promote their growth. The mechanisms by which Echinacea and its extracts regulate the immune function mainly involve the binding of Echinacea to cell surface receptors (e.g., TLR4 and CB2) and the activation of related signaling pathways, as well as the expression of immune-related factors. At the same time, it can be seen that there are many shortcomings in the research on Echinacea and its extracts, i.e., there are fewer studies on its application in livestock; the mechanisms of regulating the immune function are limited to in vitro cellular assays and have not been verified in vivo; and the safety of Echinacea and its extracts as feed additives has not been reported. Future research on Echinacea and its extracts should focus on these aspects, which will provide theoretical basis for its safe and efficient use.

Reference:

［1 ］ MACCHIA  M ，ANGELIN I  L   G，CECCARIN I  L .Methods to overcome seed dormancy in Echinacea an- gustifolia  DC ［J］．Scientia   Horticulturae，2001，89  (4) :317 －324 .

［2 ］  HUDSON  J   B．Applications  of  the  phytomedicine  Echinacea purpurea  (purple coneflower) in infectious  diseases［J］．Journal of Biomedicine and Biotechnolo- gy，2012，doi :10．1155 /2012 /769896 .

［3 ］ CHICCA A，ADINOLFI B，MARTINOTTI E，et al.   Cytotoxic effects  of  Echinacea  root  hexanic  extracts  on human cancer cell lines［J］．Journal of Ethnophar- macology，2007，110(1) :148 －153 .

［4 ］ SKOPI N' SKA -R6ZEWSKA   E ，STRZELECKA   H， WASIUTYN' SKI A，et al．Aqueous and hydro-alcohol- ic extracts of Echinacea purpurea  (L) Moench as tra- ditional  herbal  remedies   with  immunotropic  activity  ［J］．Central-European Journal of Immunology，2008， 33 (2) :78 －82 .

［5 ］ BARNES J，ANDERSON  L  A，GIBBONS  S，et  al.   Echinacea  species  ( Echinacea   angustifolia   ( DC．) Hell．，Echinacea pallida   ( Nutt．) Nutt．，Echinacea  purpurea  ( L．) Moench) :a  review  of  their  chemistry，pharmacology  and  clinical  properties［J］．Journal of Pharmacy  and Pharmacology，2005，57 (8 ) :929 -  954 .

［6 ］  GILROY C M ，STEINER J F，BYERS T，et al．Echi- nacea and  truth  in  labeling［J］．Archives  of  Internal Medicine，2003，163 (6) :699 －704 .

［7 ］  ALY  S  M ，MOHAMMED  M  F，JOHN  G．Echinacea  as  immunostimulatory   agent  in   Nile  tilapia  ( Oreo- chromis niloticus) via earthen pond experiment［C］/ /  Proceedings of the 8th International Symposium on Ti- lapia in Aquaculture．Cairo:［s．n．］，2008 .

［8 ］ OSKOII S  B，KOHYANI  A  T，PARSEH  A，et  al.   Effects  of  dietary   administration  of  Echinacea pur- purea on growth indices and biochemical and hemato- logical   indices    in    rainbow    trout    ( Oncorhynchus  mykiss ) fingerling s ［J］．Fish  Physiology  and  Bio- chemistry，2012，38 (4) :1029 －1034 .

［9 ］  ARAFA N M  S，SALEM  S  M  A，FARID  O A  H A .   Influence of Echinacea  extract pre- or  postnatal  sup- plementation on immune and oxidative status of grow - ing  rabbits［J］．Italian  Journal  of  Animal  Science， 2010，9 (3) :e63 .

［10］ EBRAHIM I H，RAHIM I S，KHAKI P．The effect of  organic acid，probiotic and Echinacea purpurea usage  on gastrointestinal  microflora  and  immune  system   of  broiler chickens［J］．Journal of Veterinary  Research， 2015，70(3) :293 －299 .

［11］ Hao Zhiwei, Chen Zhuanduo, Qiu Mei, et al. Effects of different Echinacea extracts on the immune function of broiler chickens［J］. Journal of Chinese Veterinary Medicine, 2010, 29 (2) :7 -11 .

［12］ Niu Xiaofei, Shi Wanyu, Ni Yaodi, et al. Effect of Echinacea purpurea on the immunization effect of infectious bursa vaccine［J］. Animal husbandry and veterinary medicine, 2008, 40 (9) :5 -8 .

［13］ Ni Yaodi, Xu Li, Du Jian. Effects of Echinacea polysaccharide on peripheral blood T cell subpopulations in IBDV-vaccinated chicks［J］. Chinese Journal of Veterinary Medicine, 2014, 50(5) :57 -59 .

［14］ Song Hongwei, Li Xiuju, Yue Li. Effect of Echinacea extract on immune function of broiler chicks［J］. Feed Expo, 2014(6) :7 -10 .

［15］  KIM  H  R，OH S  K，LIM  W，et al．Immune enhancing effects of Echinacea purpurea root extract by reducing regulatory  T  cell  number  and  function［J］．Natural Product Communications，2014，9 (4) :511 －514 .

［16］ Wu Jiangtao, Liu Ke. Effect of Echinacea extract on the systemic function of mouse monocyte macrophage［J］. Journal of Yantai University: Natural Science and Engineering Edition, 2001, 14(2) :131 -134.

［17］ Li Wan, Liu Ping, Shi Ruonan, et al. Effects of Echinacea extract on immune organ index and serum hemolysin level in mice[J]. Journal of Chinese Veterinary Medicine, 2015, 34(6) :40-42 .［18］ POURGHOLAM  R，SHARIF  ROHANI  M ，SAFARI  R，et al．Effect of Echinacea purpurea  extract on the  immune   system   of    rainbow   trout    ( Oncorhynchus  mykiss) and its resistance to Streptococcusis［J］．Iranian Scientific Fisheries Journal，2013，22(3) :1 －12 .

［19］ EL-ASELY A  M ，AMIN  R A，EL-HABASHI N  M .Effect of dietary administration of Echinacea purpurea  on immune responses，histopathological alteration and  microbial safety  in Nile tilapia  ( Oreochromis  nilotic- us) infected  with  Aeromonas  hydrophila［C］/ /Pro- ceedings of the 5th  Global Fisheries  and Aquaculture  Research  Conference．Giza : Faculty   of  Agriculture， Cairo University，2012 :100 －114 .

［20］ Qin Zhi-Hua, Dong Wen-Bin, Jiang Ling-Xu, et al. Effects of Echinacea extract on nonspecific immune function of turbot (Scophthalmus maximus) [J]. Ocean and Lake, 2015, 46(3) :665 -669 .

［21］ Ren Yonglin. Effects of Echinacea purpurea extract on production performance and immune function of common carp [D]. D]. Master's degree thesis. Ya'an: Sichuan Agricultural University, 2008.

[22] Xiao Zhengzhong, Wu Suxiao. Effects of Echinacea purpurea extract on immune function in mice［J］. Jiangsu Agricultural Science, 2011, 39 (3) :275 -276 .

［23］  SASAGAWA M ，CECH N B，GRAY D E，et al．Echi- nacea alkylamides  inhibit interleukin-2  production  by  Jurkat T  cells［J］．International  Immunopharmacolo- gy，2006，6(7) :1214 －1221 .

［24］ ZHAI Z  L，HANEY  D，WU  L  K，et  al．Alcohol  ex- tracts of Echinacea  inhibit  production  of  nitric  oxide  and  tumor  necrosis  factor-alpha   by  macrophages  in  vitro［J］.Food  and  Agricultural  Immunology，2007， 18 (3 /4) :221 －236 .

［25］ SULLIVAN  A  M ，LABA  J  G，MOORE  J  A，et  al.Echinacea-induced macrophage activation［J］．Immu- nopharmacology   and    Immunotoxicology ，2008 ，30  (3) :553 －574 .

［26］  YU B，LEE T T．Immunostimulant activity of Echina- cea extracts in vitro［C］/ /Proceedings of 2010 Inter- national Conference  on  Biotechnology  and  Food  Sci- ence．［S．l］:［s．n．］，2010 :37－41 .

［27］ CLASSEN B，THUDE S，BLASCHEK  W，et al．Im - munomodulatory   effects   of  arabinogalactan-proteins  from  Baptisia  and  Echinacea ［J］．Phytomedicine， 2006，13 (9 /10) :688 －694 .

［28］  GROOM  S N，JOHNS T，OLDFIELD P R．The poten- cy of immunomodulatory  herbs  may  be  primarily  de- pendent  upon  macrophage  activation［J］．Journal  of  Medicinal Food，2007，10(1) :73 －79 .

［29］ FAST D  J，BALLES  J  A，SCHOLTEN  J  D，et  al.   Echinacea purpurea  root extract inhibits TNF release  in response to  Pam3Csk4  in  a  phosphatidylinositol-3 - kinase  dependent  manner［J］．Cellular  Immunology， 2015，297 (2) :94 －99 .

［30］  SHARMA M ，SCHOOP R，SUTER A，et al．The po- tential  use  of  Echinacea  in  acne : control  of  Propi- onibacterium  acnes  growth  and  inflammation ［J］.   Phytotherapy Research，2011，25 (4) :517 －521 .

［31］  SHARMA M ，SCHOOP R，HUDSON J B．Echinacea  as an antiinflammatory agent :the influence of phy sio- logically  relevant  parameters ［J］．Phytotherapy   Re- search，2009，23 (6) :863 －867 .

［32］ Xu Xin. Effects of Echinacea extract on immunity and resistance to Salmonella infection in mice and its mechanism [D]. Doctoral dissertation. Hangzhou: Zhejiang University, 2014.

［33］  GERTSCH J，SCHOOP R，KUENZLE U，et al．Echinacea  alkylamides  modulate  TNF-α  gene expression via  cannabinoid  receptor   CB2   and   multiple   signal transduction pathways［J］．FEBS  Letters，2004，577 (3) :563 －569 .

［34］  CHICCA A，RADUNER S，PELLATI F，et al．Syner- gistic  immunomopharmacological  effects  of  N-alkyl- amides in Echinacea purpurea herbal extracts［J］．In-ternational   Immunopharmacology，2009，9  ( 7 /8 ) : 850 －858 .

［35］  WANG  C Y，CHIAO M T，YEN P J，et al．Modulato- ry  effects  of  Echinacea purpurea  extracts  on  human  dendritic cells :a  cell- and gene-based  study［J］．Ge- nomics，2006，88 (6) :801 －808 .

［36］ BRUSH J，MENDENHALL E，GUGGENHEIM  A，et  al．The effect of Echinacea purpurea，Astragalus mem- branaceus  and  Glycyrrhiza  glabra  on  CD69  expres- sion and immune cell activation in humans［J］．Phyto- therapy Research，2006，20(8) :687 －695 .

［37］  MOLTENI M ，GEMMA S，ROSSETTI C．The role of  toll-like receptor 4  in infectious and noninfectious in- flammation ［J］．Mediators  of  Inflammation，2016， 2016(7) :1 －9 .

［38］ Zhang Shujuan, Tang Wangxian. Role of cannabinoids and their receptors in immunomodulation［J］. Journal of Huazhong University of Science and Technology :Medical Edition, 2014, 43 (2) : 236 -238, 241 .