Adjuvants enhance immunogenicity and sustain long-term immune responses. As vital components of vaccines, efficient adjuvants are highly desirable. Recent evidence regarding the potential of carbon nanotubes (CNTs) to act as a support material has suggested that certain properties, such as their unique hollow structure, high specific surface area, and chemical stability, make CNTs desirable for a variety of antigen-delivery applications. Lentinan, a β-1,3-glucohexaose with β-1,6-branches that is extracted from the mushroom Lentinus edodes, is an effective immunostimulatory drug that has been clinically used in Japan and China, and recent studies have proved that specific beta-glucans can bind to various immune receptors. In this research, we covalently attached lentinan to multiwalled carbon nanotubes (MWCNTs) and tested their ability to enhance immune responses as a vaccine delivery system. In vitro study results showed that the nanotube constructs could rapidly enter dendritic cells and carry large amounts of antigen. Moreover, maturation markers were significantly upregulated versus the control. Thus, lentinan-modified multiwalled carbon nanotubes (L-MWCNTs) were regarded as an effective intracellular antigen depot and a catalyzer that could induce phenotypic and functional maturation of dendritic cells. Furthermore, compared with L-MWCNTs (35 μg/mL), a corresponding concentration of carboxylic carbon nanotubes (C-MWCNTs, 31.8 μg/mL) and an equivalent concentration of lentinan (3.2 μg/mL) did not remarkably influence the immune reaction in vitro or in vivo. Hence, we can hypothesize that the capability of L-MWCNTs was a consequence of the increased intracellular quantity of lentinan grafted onto the nanotubes. Overall, our studies demonstrated that L-MWCNTs significantly increased antigen accumulation in the cells and potentiated cellular and humoral immunity. In conclusion, L-MWCNTs constitute a potential vaccine delivery system to enhance immunogenicity for therapeutic purposes.Keywords: adjuvant; carbon nanotube; immune response; lentinan; OVA

Highlights•GLPL/OVA were able to maintain good colloidal stability and higher OVA-EE when stored at 4 °C.•GLPL/OVA could induce more powerful antigen-specific immune responses.•GLPL/OVA generate a stronger Th1-biased immune response.•The strong immune response of GLPL/OVA might be attributed to efficient activation and mature of DC in draining lymph nodes.Liposome-based vaccine delivery systems are known to enhance immune responses. Ganoderma lucidum polysaccharides (GLP) have been widely studied as immunomodulator and it could be as inducers of strong immune responses. In the research, GLP and ovalbumin (OVA) were encapsulated into liposome as vaccine and inoculated to mice. The magnitude and kinetics of the humoral and cellular immune responses were investigated. The results showed that GLP–OVA-loaded liposomes (GLPL/OVA) could induce more powerful antigen-specific immune responses than each single-component formulation. Mice immunized with GLPL/OVA displayed higher antigen-specific IgG antibodies, better splenocytes proliferation, higher cytokine secretion by splenocytes and significant activation of CD3 + CD4+ and CD3 + CD8+ T cells. Thus the GLPL/OVA formulation produced a heightened humoral and cellular immune response, with an overall Th1 bias. Enhanced immune responses elicited by the GLPL/OVA formulation might be attributed to effective activation and mature of DC in draining lymph nodes. Overall, these findings indicate that GLPL have the potential to enhance immune responses as vaccine delivery systems.

•RSM was chosen as the optimal method for preparation conditions of LBPL.•LBPL with high entrapping efficiency and small particle size could be prepared by the reverse-phase evaporation method.•LBPL could effectively enhance peritoneal macrophages phagocytosis and promote NO secretion of the peritoneal macrophages.The purpose of this study was to optimize the preparation conditions of Lycium barbarum polysaccharides liposome (LBPL) by response surface methodology (RSM) and to investigate the effect of LBPL activating function of peritoneal macrophages. LBPL was prepared using the reverse-phase evaporation method. The optimal preparation conditions of LBPL by RSM were as follows: the ratio of lipid to drug (w/w) of 25:1, the ultrasound time of 14 min and the ratio of soybean phospholipids to cholesterol (w/w) of 2.4:1. Under these conditions, the experimental encapsulation efficiency of LBPL was 86.37 ± 0.63%, which was close to the predicted value. These indicated that LBPL with high entrapping efficiency and small particle size could be prepared by the reverse-phase evaporation method, which is applied easily. Furthermore, macrophages are the key players in the innate immune system. LBPL could effectively enhance peritoneal macrophages phagocytosis and resulted in inducing NO (nitric oxide) production in mouse peritoneal macrophages.

•RSM was chosen as the optimal method for preparation conditions of GLPL.•The GLPL particles showed nearly spherical shape with uniform size.•GLPL significantly stimulated proliferation of murine splenic lymphocyte singly or synergistically with PHA and LPS.The aim of this study was to investigate the optimizing preparation conditions of Ganoderma lucidum polysaccharide liposome (GLPL) with response surface methodology (RSM) and the immunological enhancement activity of GLPL. The immunological enhancement activity of GLPL on splenocyte proliferation was measured. The optimum formulation of GLPL, in which the ratio of soybean phospholipid to cholesterol(w/w) of 11:1, the ratio of soybean phospholipid to tween-80 (w/w) of 10.5:1 and ultrasonic time(min) of 11, had higher entrapment efficiency (EE) of 71.43 ± 0.49% with spherical shape and uniform sizes. In addition, GLPL could significantly promote splenocyte proliferation singly or synergistically with PHA and LPS. That indicated that the immunological enhancement of Ganoderma lucidum polysaccharide (GLP) was significantly enhanced after encapsulation with the liposome.

•We studied the isolation of Cynanchum paniculatum (Bunge) Kitag and obtained twenty-nine compounds.•The structures of these compounds were elucidated by spectroscopic methods.•Five compounds were firstly reported from Asclepiadaceae family.•Compounds 7, 15–17, 19, 20 and 27 were obtained from C. paniculatum firstly for the first time.•The isolated constituents exhibited the relationships between C. paniculatum and some other plants of Cynanchum genus.Twenty-nine compounds, including five acetophenone derivatives (1–5), three phenanthroindolizidine alkaloids (12–14), seven pentacyclic triterpenoids (15–21) and five C21 steroidal sapogenins (22–26), were isolated from the root of Cynanchum paniculatum (Bunge) Kitag. Their structures were determined by spectroscopic methods and comparison with reported data. Moreover, the chemotaxonomic relationships were also discussed. As a result, acetophenone derivatives, pentacyclic triterpenoids and C21 pregnane sapogenins can be recognized as chemotaxonomic markers for Cynanchum genus, and C. paniculatum has close relationships with some species of genus Cynanchum.

•Reverse-phase evaporation method was chosen as the optimal method for preparation of RGPL.•Based on single factor investigation, RSM was employed to determine the optimum preparation conditions of RGPL.•RGPL significantly stimulated proliferation of murine splenic lymphocyte in vitro.The aim of this study was to investigate the optimizing preparation conditions and immunological enhancement activity of Rehmannia glutinosa polysaccharide liposome (RGPL). RGPL was prepared using the reverse-phase evaporation method and optimized using the response surface methodology. The immunological enhancement activity of RGPL on splenocyte proliferation was measured. These results showed that the optimum preparation conditions were: a soybean phosphatide to cholesterol ratio of 8:1, a chloroform to methanol ratio of 3:5, a soybean phosphatide to tween 80 ratio of 10:1, a temperature (°C) of 66 °C and an entrapment efficiency of 72.753 ± 0.318%. In a single stimulation of drugs, RGPL could significantly promote splenocyte proliferation, specifically at 200 μg mL−1. Moreover, in a synergistic stimulation of drugs with LPS or PHA, a significant difference was obtained between the RGPL and RGP at 100–400 μg mL−1, which indicated that the immunological enhancement of RGP was significantly improved after encapsulation with the liposome.

•RGP significantly stimulated proliferation of murine splenic lymphocyte.•RGP significantly upregulated IL-2 and IFN-γ production of T lymphocyte.•RGP could improve DCs stimulating on proliferation of T cells.•RGP could enhance the ability of antigen presenting of DCs.The aim of this study is to investigate immunomodulatory effect of rehmannia glutinosa polysaccharide (RGP) on murine splenic lymphocyte and bone marrow derived dendritic cells (DCs). Splenic lymphocytes obtained from mice were co-cultured with RGP for 48 h and then harvested for analyzing with MTT method. The cytokine production of T lymphocytes was measured by ELISA. Effects of RGP treatment on DCs were investigated and assessed by MTT method. The results showed RGP significantly stimulated lymphocyte proliferation and the growth rate of T cell was more significant. The IL-2 and IFN-γ production of T lymphocyte were significantly upregulated after being stimulated with RGP. DCs stimulating on proliferation of T cells and the ability of antigen presenting of DCs have been enhanced under the stimulation of RGP. In conclusion, these findings provided valuable information that RGP possessed strong immunoenhancement activity, which provided the theoretical basis for the further experiment.

Two experiments were carried out. In vitro, the effects of Astragalus polysaccharide liposome (APSL) on chicken's T and B lymphocytes proliferation were determined. The results showed that APSL could significantly promote T and B lymphocytes proliferation singly or synergistically with PHA and LPS and the efficacy were superior to those of Astragalus polysaccharide (APS) and blank liposome (BL) at most concentrations. In immune response experiment, the adjuvanticity of APSL at three doses, APS and BL were compared on chickens vaccinated with ND vaccine. The results showed that APSL could significantly promote lymphocyte proliferation, enhance antibody titer, promote IFN-γ, IL-2, IL-4 and IL-10 secretion, and its medium dose possessed the best efficacy. These results indicated that APSL could significantly improve the adjuvanticity and drug action of APS, and its medium dose possessed the best efficacy, the liposome would be expected to exploit into a new-type preparation of APS.Highlights► APSL could significantly promote T and B lymphocytes proliferation in vitro. ► APSL could significantly improve the adjuvanticity of APS in vivo. ► The medium dose of APSL possessed the best efficacy. ► The liposome would be expected to exploit into a new-type preparation of APS.

•Salidroside liposome could promote mature, stimulating proliferation of MLR and antigen presentation of DC cells in vitro.•Salidroside liposome enhanced the immune response in vivo.•Immunological adjuvant of salidroside liposome is better than those of salidroside and liposome.Salidroside, the important composition, of Rhodiola rosea L. has been reported to have various pharmacological properties. Liposome is known to be effective as drug carriers and immune adjuvant. Therefore, the aim of this study is to investigate immunological adjuvant activity of salidroside liposome. Here we reported the preparation, the effect on DCs in vitro and the immune response in vivo. The immunological adjuvant activity of salidroside liposome formulation was compared with that of salidroside and liposome. The result showed that salidroside liposome formulation not only could promote the maturation of DCs, the stimulation of DCs on MLR proliferation and the antigen presenting ability, but also induced the sustained cellular immune and humoral immune response. Overall, the results showed that salidroside liposome formulation had the potential to act as effective sustained release vaccine delivery systems.

•The average diameters of liposomes (LBPL, BL) and liposome-formulated vaccines (LBPL-PCV2, BL-PCV2) were less than 200 nm and stable.•LBPL could significantly promote splenocyte proliferation and the cytokine secretion of macrophages in vitro.•LBPL as a vaccine adjuvant has good improvement effect on immune responses against PCV2 in vivo.In previous researches, the results showed that Lycium barbarum polysaccharides (LBP) encapsulated with liposome could enhance the immune activity of LBP. Therefore, the present study was designed to investigate the effects of LBPL on spleen lymphocytes and macrophages of mice in vitro and evaluate the immunological adjuvant activity of PCV2 vaccine in vivo. The results showed that LBPL could significantly promote splenocyte proliferation synergistically with PHA or LPS, increase the ratio of CD4+ to CD8+ T cells and promote the cytokine secretion of macrophages; enhance PCV2-specific IgG antibody responses, promote Th1 cytokines (IFN-γ and TNF-a) and Th2 cytokine (IL-4) secretion. The histomorphological observation of spleen demonstrated that LBPL as a vaccine adjuvant also has good improvement and stimulating effect on the immune organ.