Research On The Mechanism Of Polysaccharide From The Roots Of Radix Cyathulae Officinalis Kuan (RC) On Immune Responses In Mice

Infectious diseases, especially virus diseases, are worldwide concern as they usually cause great loss in domestic animal and poultry industry. Vaccination remains the most effective biomedical approach for the control of infectious diseases. Although the strict immune program had been taken in the farms, some infectious diseases are still hard to be controlled due to the lack of effective immune responses to vaccination. The majority of vaccines require association with adjuvants capable of increasing the potency or stimulating the appropriate immune response. The application of adjuvant, one of the methods to improve the efficacy of vaccine, has been considered as an important strategy. There are many growing evidences that medicinal herbs and the ingredients enhance the immune response to vaccines direct against infectious agents. Several herbal extracts as adjuvant of vaccines has been use in the human and animal clinical trials."Sichuan herbs" are well known because the medicinal quality of the herbals Produced in Si Chuan Province believed to be better than in other Province of China. We choose ten well known herbs include Cyathula officinalis Kuan, Eucommia ulmoides Oliv, Chuanminshen violaceum, Ophiopogon japonicus,Alisma orientalis,Salvia miltiorrhiza, Dipsacaceae, Rhizoma Ligustici Chuanxiong, Radix Angelicae Dahuricae, bergamol. screening the immunoenhancement herbs from those ten "Sichuan herbs"and following to study the Effects of oral administration of herbals extract on immuno response to the vaccination in mice. The relating studies carried out and results obtained are as follows:1. Effects of ten Sichuan herbals Extract on the Immune Responses to Ovalbumin in MiceObjective This study was investigated the effects of oral administration the water extract from the ten Sichuan herbs on the immune responses in mice immunized with ovalbumin (OVA). Method One hundred fifty five Institute of Cancer Research (ICR) mice were randomly divided into Thirty one groups with five animals in each group, and daily orally administered daily for4days with a high, medium or low dose of herbal exract respectively. After that, the animals were subcutaneously immunized twice with OVA (10μg) at2-week intervals. Blood samples were collected on day14after,boosting for, measurement of OVA-specific IgG titers and the IgG subclasses. Results Our data indicated that the OVA-specific IgG titers and the level of IgG subclass was significantly enhanced in mice orally administered exact of Radix Cyathulae officinalis Kuan (RC) when compared to the control group (P<0.05). OVA-specific IgG titers were significantly improved in mice orally administered the exact from Eucommia ulmoides Oliv and the Chuanminshen violaceum when compared to the control group (P<0-05).2. Effects of the Extract from Radix Cyathulae (RC) on the Immune Responses to Ovalbumin in MiceObjective This study was designed to evaluate the effects of oral administration of the water extract from the Radix Cyathula (RC) on the immune responses in mice immunized with ovalbumin (OVA). Method Forty two Institute of Cancer Research (ICR) mice were randomly divided into six groups with seven animals in each group, and orally administered daily for4days at a dose equivalent to0,0.05,0.1,0.2,0.4or0.8g of the RC exract, respectively. After that, the animals were subcutaneously immunized twice with OVA at2-week intervals. Blood samples and spleen were collected on day14after boosting for measurement of OVA-specific IgG titers and the IgG subclasses, lymphocyte proliferation as well as the production of IL-4and IFN-y. Results Our data indicated that the OVA-specific IgG titers and the level of IgG subclass was significantly enhanced in mice orally administered RC at the dose of0.2,0.4or0.8g when compared to the control group (P<0.05). The splenocyte proliferation response and the production of IL-4and IFN-y were also significantly improved (P<0.05). Besides, the body weight of the mice administered with the RC extract (0,0.1,0.2,0.4or0.8g) was no significant difference compared to the mice administered saline.Considering the immunomodulatory effect and safety of RC presented in this study, it deserves further investigation to evaluate the potential improvement of vaccination in other animals such as pigs, goats and cattle. Conclution Our data show that RC significantly enhanced OVA-specific IgG and IgG subclasses titers, lymphocyte proliferation, the production of IL-4and IFN-y.3. Effects of Polysaccharide from the Radix Cyathulae officinalis Kuan (RCPS) on the Immune Activity of Lymphocyte in vitroObjective This study was to extract Polysaccharide from the Radix Cyathulae officinalis Kuan (RCPS) and to explore the effects on the immune activity in vitro. Method firstly, the highest non-toxic concentrations of RCPS on splenocytes were measurement. Effect of RCPS on lymphocyte proliferation, the production of IL-4and IFN-y from lymphocyte, NK cell Killing activity were investigated. Results The results showed that the highest non-toxic concentrations of RCPS at500μg/mL, The splenocyte proliferation response and the production of IL-4and IFN-γ were also significantly improved (P<0.05) after the splenocytes were stimulated by RCPS. Besides, RCPS promoted the NK cell activity. Conclution Our findings show that RCPS significantly enhanced lymphocyte proliferation, the production of IL-4and IFN-y and NK activity in vitro.4. Effects of the Polysaccharide from the Radix Cyathulae officinalis Kuan (RCPS) on the Immune Response to Ovalbumin in MiceObjective This study was designed to investigate the effects of oral administration of the polysaccharide from the Radix Cyathulae officinalis Kuan (RCPS) on the specific cellular and humoral immune responses in mice. Methods Forty two ICR mice were randomly divided into six groups (n=7), and orally administered at a dose equivalent to0,25mg,50mg,100mg of the RCPS for continuous4days respectively. After that, the animal were subcutaneously immunized twice with OVA at2weeks intervals. Blood samples and spleen collected on day14after boosting for measurement including serum OVA-specific antibody level, lymphocyte proliferation, expression of surface and intracellular molecules, phagocytic capacity of peritoneal macrophage, NK (Nature killer) and CTL (cytotoxic lymphocyte) Killing activity, The frequency of costimulatory molecule (CD40, CD80, CD86) and CD4+CD25+Foxp3+Treg cells. Results In this study, our findings demonstrated that oral administration of RCPS significantly enhanced OVA-specific IgG, IgG1, IgG2a, and IgG2b antibody titers, splenocyte proliferation, the phagocytic capacity of peritoneal macrophage, and the activity of natural killer (NK) cells and cytotoxic T lymphocytes (CTL). Furthermore, RCPS promoted the level of interleukin-2(IL-2), IFN-γ and IL-4in CD4+T cells and level of IFN-y in CD8+T cells. In addition, RCPS enhanced the expression of CD40, CD80and CD86on the dendritic cells (DCs). Importantly, RCPS down-regulated the frequency of CD4+CD25+Foxp3+Treg cells. C Taken together, these results suggested that RCPS could increase both cellular and humoral immune responses via up-regulating-DCs maturation, and suppressing Treg frequency.5. Effects of the Polysaccharide from the Radix Cyathulae officinalis Kuan (RC)on the immune Response to Foot-and-Mouth Disease Vaccine in MiceObjective This study was designed to investigate the effects of oral administration of the polysaccharide from the Radix Cyathulae officinalis Kuan (RCPS) on the specific cellular and humoral immune responses in mice to explore its mechanisms. Methods Sixty ICR mice were randomly divided into six groups (n=10), and orally administered at a dose equivalent to0,25,50,100mg of the RCPS for continuous4days respectively. After that, the animal were subcutaneously immunized twice with FMDV vaccine at2weeks intervals. Blood samples and spleen collected on day14after boosting for measurement of were determined, including serum antibody level, lymphocyte proliferation, the phagocytic capacity of peritoneal macrophage, Natural killer cell (NK) and cytotoxic T lymphocyte(CTL) activity, expression of surface and intracellular molecules, the frequency of CD4+CD25+Foxp3+regulatory T cells (Treg cells), the mRNA level of interleukin-2(IL-2), interleukin-4(IL-4), interleukin-10(IL-10), interferon-y (IFN-y), major histocompatibility complex Ⅰ (MHC Ⅰ), major histocompatibility complex Ⅱ (MHCⅡ), CXC chemokine receptor4(CXCR4), CC chemokine receptor7(CCR7), Toll like receptor2(TLR-2), Toll like receptor4(TLR-4), transcription factor T-bet, GATA-3,transforming growth factor-β (TGF-β) were detected by Real-Time PCR. Results In this study, our findings show that addition of the RCPS increased FMDV-specific antibody production, lymphocyte proliferation, phagocytic capacity of peritoneal macrophage, NK and CTL activity. Production of IL-2, IFN-y and IL-4in CD4+T cells and IFN-y in CD8+T cells were dramatically:increased. Expression-of CD40,CD80, CD80,CD86were was significantly increased. In addition, the frequency of CD4+CD25+Foxp3+regulatory T cells were down-regulated. The mRNA expression of the MHC I, MHC II, CXCR4, TLR-2TLR-4, T-bet, GATA-3, IL-2, IL-4, IL-10, IFN-y were up-regulated. The mRNA level of TGF-β were down-regulated. Conclution Taken together, these results demonstrate that RCPS can enhance both cellular and humoral immune responses via up-regulating DCs maturation through TLR2, TLR4signaling pathway, and inhibit the expression of TGF-β and frequency of Treg cells.6. Conclution of this studyTaken together, these results demonstrate that RCPS can enhance both cellular and humoral immune responses in mice immunized with FMD vaccine via up-regulating DCs maturation through TLR2, TLR4signaling pathway, and inhibit the expression of TGF-β and frequency of Treg cells. RCPS promoted a balanced Thl/Th2immune response via the develop of ThO cells to Thl/Th2cells through up-regulated T-bet, GATA-3mRNA level.