Regulatory Effects of Alhagi Honey Small-Molecule Sugars on Growth Performance and Intestinal Microbiota of Lambs.

The present study was designed to assess the impact of Alhagi honey small-molecule sugars (AHAS) on Hu lambs. Therefore, in this study, AHAS low-dose (AHAS-L, 200 mg/ kg per day), AHAS medium-dose (AHAS-M, 400 mg/kg per day), and AHAS high-dose (AHAS-H, 800 mg/kg per day) were administered to Hu lambs to investigate the regulatory effects of AHAS on growth performance, oxidation index, immune system enhancement, and intestinal microbiota. The results showed that lambs in the AHAS-H group exhibited significantly increased in average daily weight gain, and growth performance compared to those in the control group (p < 0.05). Moreover, AHAS-H supplementation resulted in increased levels of serum antioxidant enzymes (SOD, GSH-Px, and T-AOC), serum antibodies (IgA, IgG, and IgM), and cytokines (IL-4, 10,17, IFN-γ, and TNF-α) compared with the control group (p < 0.05). Additionally, it increased the quantity and richness of beneficial bacteria at such as Sphingomonas, Ralstonia, and Flavobacterium, activating various metabolic pathways and promoting the production of various short-chain fatty acids. In summary, our findings highlight the potential of AHAS-H treatment in enhancing intestinal health of lambs by improving intestinal function, immunity, and related metabolic pathways. Consequently, these results suggest that AHAS holds promising potential as a valuable intervention for optimizing growth performance and intestinal health in lambs.

The first molecular detection of benzimidazole resistance in Haemonchus contortus from sheep in Hejing and Minfeng counties of Southern Xinjiang.

To understand the benzimidazole (BZ) resistance of Haemonchus contortus in Southern Xinjiang, three single nucleotide polymorphisms (SNPs) designated as F167Y, E198A, and F200Y, in the isotype-1 ß-tubulin gene which are associated with BZ resistance, were investigated for H. contortus populations from sheep in Hejing and Minfeng counties of Southern Xinjiang. In brief, a total of 190 H. contortus adults were collected from 52 out of 70 slaughtered sheep in city abattoirs across two regions in Southern Xinjiang. The species identity of each adult worm was confirmed by PCR amplification of ITS-2 using H. contortus-specific primers targeting the ITS-2. The samples were then investigated for BZ-related SNPs at locus 167, 198, and 200, by PCR-sequencing of the isotype-1 ß-tubulin gene. The results showed that only E198A and F200Y mutations were detected in the investigated H. contortus populations. The E198A mutation (homozygous and heterozygote resistant: found in 40% and 30% of sequenced samples from Minfeng and Hejing counties, respectively) was predominant compared with the F200Y mutation (homozygous and heterozygote resistant: found in 14% and 13.3% of sequenced samples from Minfeng and Hejing counties, respectively). The results indicate a high prevalence of BZ resistance in H. contortus populations from certain areas of Southern Xinjiang. Our findings provide valuable information for the prevention and control of H. contortus in areas with similar conditions.

Pomegranate flower polysaccharide improves mastitis in mice by regulating intestinal flora and restoring the blood-milk barrier.

This study explored the inhibitory effect of pomegranate flower polysaccharide (PFPS) on mastitis through in vitro and in vivo models. PFPS is a new type of polysaccharide isolated and extracted from pomegranate flowers. The result revealed that PFPS consists of GalA, Ara, and Gal, and the residues consist of 1,4-GalpA, 1,4-Galp, and 1,3,6-Galp, which contain HG-type and RG-I-type pectin structural domains. In vitro studies showed that PFPS could inhibit LPS-enhanced phagocytosis of RAW 264.7 cells and the release of IL-1ß, IL-10, and TNF-α. In vivo, studies showed that PFPS improved xylene-induced mouse ear swelling and carrageenan-induced mouse paw edema by inhibiting inflammatory factors. In the mouse mastitis model, PFPS significantly improved LPS-induced inflammation and oxidative stress in mammary tissue. Intestinal flora sequencing results showed that PFPS could effectively regulate the intestinal flora of mice, reduce the relative abundance of pathogenic bacteria Oscillospira and AF12, and increase the probiotics Blautia, Parabacteroides, Allobaculum, and Clostridiaceae_Clostridium. Therefore, PFPS ultimately played a role in preventing mastitis by regulating the intestinal flora and further improving the blood-milk barrier. This study provides a scientific basis for PFPS as a potential candidate drug for the treatment of mastitis.

Extraction and immunomodulatory effects of acid Lagenaria siceraria (Molina) Standl. Polysaccharide on chickens.

Herbal polysaccharides are extensively studied as vaccine adjuvants due to their safety and potent immunoenhancing activity. This study aimed to analyze the structure of Lagenaria siceraria (Molina) Standl polysaccharide (LSP50) and investigate its adjuvant activity for the H9N2 vaccine in broiler chickens. Structural analysis revealed that LSP50 primarily consisted of rhamnose, arabinose, xylose, mannose, glucose, and galactose with molar ratios of 23.12: 12.28: 10.87: 8.26: 2.64: 22.82 respectively. The adjuvant activity of LSP50 was evaluated, which showing significant enhancements compared to the H9N2 group. Parameters including the immune organ index, H9N2 specific IgG level, cytokines contents (IFN-γ, IL-2, IL-4, and IL-5), and the proportion of CD3e+CD8aT+cells were significantly increased in the LSP50 group (P < 0.05). Additionally, sequencing results showed that LSP50 modulates the immune response by regulating PLA2G12B and PTGDS genes involved in the arachidonic acid pathway. These findings were further validated through qPCR analysis to affirm the reliability of the sequencing data. In conclusion, our results demonstrate that LSP50 exhibits potent adjuvant activity, enhancing both cellular and humoral immunity.

Effects of Alhagi maurorum Medik polysaccharide derived from different regions on the intestinal immune functions of lambs.

Introduction: Plant polysaccharide are widely studied as potential prebiotics because of their potential to protect and enhance the immunity of lambs. Methods: In this study, the polysaccharide content of Alhagi maurorum Medik from Aksu (AK) and Shanshan (SS) at different cutting periods was determined, and the functions of Alhagi maurorum Medik polysaccharide were investigated to useas an immunomodulator. Results: Our results indicated that the content of Alhagi maurorum Medik polysaccharide is the highest at the maturity stage, and the polysaccharide content of Alhagi maurorum Medik produced in Shanshan area is higher as compared to the Aksu area. The serum IgG, duodenum IgA, TNF-α, IL-4, IL-10 contents, jejunum IgA, TNF-α, IL-4, IL-17 contents, ileum IgA, IL-17 contents, duodenum villus height, crypt depth and jejunum crypt depth of lambs were significantly adjusted in the SS group as compared to CK control group and AK groups (p < 0.05). Furthemore, the sequencing results showed that SS polysaccharide promoted the release of large amounts of IgA and enhanced the immunal function of intestine by regulating the IgA production pathway and B-cell receptor signaling to activate B cells in the T-dependent pathway. Discussion: Altogether, Alhagi maurorum Medik polysaccharide from SS group holds a promising potential to be used as a valuable immunopotentiator for optimizing the immune system of intestine in lambs.

Structure characterization and intestinal immune promotion effect of polysaccharide purified from Alhagi camelorum Fisch.

This study investigated the structure of acid Alhagi camelorum Fischa polysaccharide (aAP) and its impact on intestinal activity in mice. The results showed that aAP comprised of the fucose, arabinose, rhamnose, galactose, glucose, xylose, mannose, galacturonic acid, glucuronic acid with the molar ratio of 0.81:14.97:10.84:11.14:3.26:0.80:0.80:54.92:2.47 with the molecular weight (Mw) of 22.734 kDa. Additionally, the composition of aAP was assessed via FT-IR, methylation, and NMR analyses, indicating that the backbone of the aAP was consisted of →4)-α-D-GalpA-6-OMe-(1 â†’ 4)-α-GalpA-(1 â†’ and →4)-α-D-GalpA-6-OMe-(1 â†’ 2)-α-L-Rhap-(1→, as well as →4)-ß-D-Galp- and →5)-α-L-Araf- for the branched chain. Furthermore, ICR mice underwent intragastric administration of different concentrations of aAP for 7 consecutive days. The results showed that aAP enhanced the murine spleen and thymus indices, promoted the secretion of serum lgG antibody, intestinal lgA antibody and intestinal cytokines, improved the morphology of intestinal villi and crypts, enhanced quantity of intestinal IELs and IgA+ cells, and activated T lymphocytes and DC cells in MLNs. In summary, these findings suggest that the utilization of aAP could enhance the immune response of the murine intestinal mucosa.

Preparation and activity study of Ruoqiang jujube polysaccharide copper chelate.

Background: Polysaccharide metal chelate exhibit both immunoregulatory activity and metal element supplementation effects. Methods: In this study, Ruoqiang jujube polysaccharide copper chelate (RJP-Cu) was prepared and the preparation conditions were optimized using the response surface method. Subsequently, RJP-Cu was administered to lambs to evaluate its impact on growth performance, copper ion (Cu2+) supplementation, immune enhancement, and intestinal flora was evaluated. Results: The results indicated that optimal RJP-Cu chelation conditions included a sodium citrate content of 0.5 g, a reaction temperature of 50°C, and a solution pH of 8.0, resulting in a Cu2+ concentration of 583°mg/kg in RJP-Cu. Scanning electron microscopy (SEM) revealed significant structural changes in RJP before and after chelation. RJP-Cu displaying characteristic peaks of both polysaccharides and Cu2+ chelates. Blood routine indexes showed no significant differences among the RJP-Cu-High dose group (RJP-Cu-H), RJP-Cu-Medium dose group (RJP-Cu-M), RJP-Cu-low dose group (RJP-Cu-L) and the control group (p > 0.05). However, compared with the control group, the RJP-Cu-H, M, and L dose groups significantly enhanced lamb production performance (p < 0.05). Furthermore, RJP-Cu-H, M, and L dose groups significantly increased serum Cu2+ concentration, total antioxidant capacity (T-AOC), catalase (CAT), and total superoxide dismutase (T-SOD) contents compared with control group (p < 0.05). The RJP-Cu-H group exhibited significant increases in serum IgA and IgG antibodies, as well as the secretion of cytokines IL-2, IL-4, and TNF-α compared to the control group (p < 0.05). Furthermore, RJP-Cu-H group increased the species abundance of lamb intestinal microbiota, abundance and quantity of beneficial bacteria, and decrease the abundance and quantity of harmful bacteria. The RJP-Cu-H led to the promotion of the synthesis of various Short Chain Fatty Acids (SCFAs), improvements in atrazine degradation and clavulanic acid biosynthesis in lambs, while reducing cell apoptosis and lipopolysaccharide biosynthesis. Conclusion: Thus, these findings demonstrate that RJP-Cu, as a metal chelate, could effectively promote lamb growth performance, increase Cu2+ content, and potentially induce positive immunomodulatory effects by regulating antioxidant enzymes, antibodies, cytokines, intestinal flora, and related metabolic pathways.

Preparation and sulfate modified of Lagenaria siceraria (Molina) Standl polysaccharide and its immune-enhancing adjuvant activity.

Herbal polysaccharides and their modifiers used as vaccine adjuvants have been widely investigated due to their safety and good immunoenhancing activity. In this study, the 50% ethanol concentration precipitated Lagenaria siceraria(Molina) standl polysaccharide (LSP50) and sulfated modified LSP50 (sLSP50) was prepared, and their characterization was investigated. LSP50 and sLSP50-1.5 were used as vaccine adjuvants to immunize chickens, and the strength and type of immune responses induced by different adjuvants were detected. Our results showed that LSP50 was homogeneous polysaccharides, and the carbohydrate content was 98.6%. The sLSP50-1.5 with the DS value of 1.5 was optimized by response surface methodology. The sLSP50-1.5 has both characteristics of polysaccharide functional groups and sulfate functional groups. Adjuvant activity of LSP50 and sLSP50-1.5 showed that LSP50 and sLSP50-1.5 could induce long-lasting and high hemagglutination (HI) titers, antigen-specific lgG-NDV antibody, splenic lymphocyte proliferation, high immune organ index. Moreover, chicken immunized with sLSP50-1.5 showed a strong mixed Th1-type (IFN-γ and TNF-α) and Th2-type (IL-4 and IL-6) cytokines expression. Thus, these findings demonstrated that sLSP50-1.5 as a vaccine adjuvant can induce a mixed cellular and humoral immune response and can potentially serve as an effective vaccine adjuvant for NDV antigen.

Polyethyleneimine modified Pickering emulsion as a novel adjuvant to induce strong and long-lasting immune responses.

Poly (lactic-co-glycolic acid) (PLGA) nanoparticles are widely-investigated vaccine adjuvants owing to their safety, antigen slow-release ability, and good adjuvants activity. This study involved the preparation of the polyethyleneimine-modified immunopotentiator Alhagi honey polysaccharide encapsulated PLGA nanoparticles (PEI-AHPP) and the assembly of the Pickering emulsion with PEI-AHPP as shell and squalene as core (PEI-PPAS). Furthermore, PEI-AHPP and PEI-PPAS were characterized. To assess the strength and type of immune responses induced by different adjuvants, the chickens were immunized with H9N2-absorbed nanoparticle formulations. Our results showed that since the PEI-PPAS possess rough strawberry-like surfaces, a large number of antigens can be absorbed on their surfaces through simple mixing. Compared to PEI-AHPP, PEI-PPAS was found to exhibit better stability and antigen-loading efficiency. The adjuvant activity of the nanoparticles showed PEI-PPAS/H9N2 to induce long-lasting and high Hemagglutination inhibition (HI) titers, high thymus, spleen, and organ index of the bursa of Fabricius. Moreover, the chickens immunized with PEI-PPAS/H9N2 showed a mixture of high CD4+ and CD8a+ T-cells in the spleen and strong Th1 and Th2-type cytokines secretion. Thus, these findings demonstrated PEI-PPAS to be a vaccine adjuvant inducing a mixed cellular and humoral immune response, which can potentially serve as an effective vaccine delivery adjuvant system for the H9N2 antigen.

Poly (lactic-co-glycolic acid) nanoparticle-based vaccines delivery systems as a novel adjuvant for H9N2 antigen enhance immune responses.

Poly (lactic-co-glycolic acid) (PLGA) nanoparticle used as vaccine adjuvants have been widely investigated due to their safety, antigen slow-release ability, and good adjuvants activity. In this study, immunopotentiator Alhagi honey polysaccharide encapsulated PLGA nanoparticles (AHPP) and assembled pickering emulsion with AHPP as shell and squalene as core (PPAS) were prepared. Characterization of AHPP and PPAS were investigated. H9N2 absorbed nanoparticles formulations were immunized to chicken, then the magnitude and kinetics of antibody and cellular immune responses were assessed. Our results showed that PPAS had rough strawberry-like surfaces, a large number of antigens could be absorbed on their surfaces through simple mixing. Adjuvant activity of PPAS showed that, PPAS/H9N2 can induce long-lasting and high HI titers, high thymus, spleen, and bursa of fabricius organ index. Moreover, chicken immunized with PPAS/H9N2 showed a mixed high differentiation of CD4+ and CD8a+ T cell, and strong Th1 and Th2-type cytokines mRNA expression. Thus, these findings demonstrated that PPAS could induce a strong and long-term cellular and humoral immune response, and has the potential to serve as an effective vaccine delivery adjuvant system for H9N2 antigen.

Alhagi honey polysaccharides encapsulated into PLGA nanoparticle-based pickering emulsion as a novel adjuvant to induce strong and long-lasting immune responses.

Alhagi honey polysaccharides, extracted from a perennial plant Alhagi pseudalhagi syn, possessed many biological activities such as immune enhancement, anti-tumor effect, and antioxygenation. In this study, we used Alhagi honey polysaccharide encapsulated (poly lactic-co-glycolic acid) (PLGA) nanoparticles to prepare an assembled particles-oil pickering emulsion: PPAS and PEI-PPAS. We investigated the characterization of two pickering emulsions, and the possible mechanism to enhance immune responses. The results showed that PPAS and PEI-PPAS both could load high adsorption of OVA and had ability to sustained controlled release OVA. In vivo experiment, PEI-PPAS/OVA enhanced the levels of IgG and cytokines. Meanwhile, it could effectively target dendritic cells (DCs), promoted the cellular uptake of OVA then activated DCs in lymph nodes. And this effect of PEI-PPAS might be induced through the MHC II and MHC I pathway in DCs. Thus, these findings demonstrated that PEI-PPAS could induce a strong and long-term cellular and humoral immune response, and have potential to applied to vaccine adjuvant delivery system.

Supplementation of Alhagi honey polysaccharides contributes to the improvement of the intestinal immunity regulating the structure of intestinal flora in mice.

Alhagi honey polysaccharides (AH), a main active component of Alhagi honey, are known to possess excellent pharmacological activities and have been widely used as dietary supplements in traditional Chinese medicine for thousands of years. This study is aimed to investigate the heath effect of AH on murine intestinal mucosal immune function and composition of the gut microbiome. ICR mice received daily intragastric administration of AH (three dosages, 200 mg kg-1, 400 mg kg-1, and 800 mg kg-1) or saline for 7 consecutive days. Results indicated an improvement in the intestinal barrier function through increases in secretory immunoglobulin A (sIgA) and ß-defensins. Simultaneously, AH also significantly stimulated IL-2, IL-4, IL-6, IL-10, IL-17, IFN-γ, and TNF-α cytokine secretion as compared to the control samples. Moreover, hematoxylin and eosin staining showed that AH enhanced the number of intraepithelial lymphocytes (IELs) in the small intestine. An obvious increase in the ratio of IgA+ cells of AH-treatment samples in the lamina propria was also detected by immunohistochemical staining. In addition, the CD3+, CD4+ and CD8+ T-cell ratio in mesenteric lymph nodes and Peyer's patches in the AH-treatment was significantly higher than that in the control group. Furthermore, 16S rDNA gene sequencing was used to monitor the dynamic changes in the gut microbiota. The result revealed that AH significantly increased the indexes of Shannon and obviously decreased the indexes of Simpson, suggesting the enhancement of the diversity and richness of the intestinal microbiome. Moreover, AH modulated the gut microbiome via increasing the abundance of probiotics and decreasing the levels of pathogenic bacteria. In summary, these results indicated that AH could be used as a prebiotic to enhance murine intestinal mucosal immunity and to modulate the gut microbiome.

Alhagi honey polysaccharides attenuate intestinal injury and immune suppression in cyclophosphamide-induced mice.

Cyclophosphamide (CY), extensively used as an anti-cancer agent, could cause diverse side effects, such as immunosuppression and intestinal barrier damage. Alhagi honey polysaccharides (AH), polysaccharides isolated from Alhagi honey, are widely known for their anti-tumor and immunomodulatory activities. Herein, AH are evaluated for their ability to protect mice from CY-induced toxicity. The results demonstrated that treatment with AH could prevent the reduction in spleen and thymus indices as well as body weight, and significantly increase the Peyer's patch count in CY-induced mice and the levels of IL-2, IL-6, and TNF-α in serum, suggesting the role of Alhagi honey polysaccharides in alleviating the immunosuppression induced by CY. Moreover, administration of AH significantly increased the SOD activity and the expression level of ß-defensin while decreasing the MDA content and DAO activity in CY-treated mice, which suggested a protective effect of AH on the intestinal barrier. Simultaneously, a CY-induced decrease in the ratio of villi length/crypt depth and the number of intraepithelial lymphocytes and goblet cells was reversed by AH treatment, as were the alterations in the expression of ZO-1, mucin-2, E-cadherin and occludin in the intestine and the concentrations of SCFAs in the colon. Furthermore, AH have the ability to regulate the MAPK pathway in CY-mice models to reduce CY-induced toxicity, evidenced by the increased expression of p-ERK and inhibited production of both p-JNK and p-p38. Overall, these results showed that AH could be used as protective agents to mitigate intestinal injury and immune suppression in mice induced by CY.

Immunoenhancement effects of chitosan-modified ginseng stem-leaf saponins-encapsulated cubosomes as an ajuvant.

Nanoparticle delivery of functional molecules and vaccine is a promising method for enhancing the immune response. The objective of this study was to design chitosan (CS)-modified ginseng stem-leaf saponins (GSLS)-encapsulated cubosomes (Cub-GSLSCS) as a vaccine delivery system and explore its immunologic activity and adjuvanticity. In this study, CS-modified GSLS-encapsulated cubosomes (Cub-GSLSCS) were prepared. The storage stability of GSLS and that of ovalbumin (OVA) were measured. Additionally, the immunopotentiation of Cub-GSLSCS were assessed on potentiating macrophage in vitro, and the adjuvant activity was evaluated through immune response triggered by OVA model antigen. The encapsulation efficiency of optimized Cub-GSLSCS was about 65 % with Im3m nanostructure. The Cub-GSLSCS showed excellent stability and sustained release for up to 28 days. In vitro, Cub-GSLSCS nanoparticles improved cellular uptake, stimulated cytokines secretion of IL-6, IL-12, TNF-α, and generated more inducible nitric oxide synthase (iNOS) to produce higher levels of nitric oxide (NO) compared with other groups. Furthermore, the immunoadjuvant effects of OVA encapsulated Cub-GSLSCS nanoparticles (Cub-GSLSCS-OVA) were observed through immunized mice. Results showed that the ratio of CD4+/CD8 + T lymphocytes was increased in Cub-GSLSCS-OVA group. In addition, Cub-GSLSCS-OVA nanoparticles induced dramatically high OVA-specific IgG, IgG1, and IgG2a levels and stimulated the secretion of cytokines. Cub-GSLSCS may be a potential vaccine delivery system and induce a long-term sustained immunogenicity.

Cationic polymer-modified Alhagi honey polysaccharide PLGA nanoparticles as an adjuvant to induce strong and long-lasting immune responses.

Alhagi honey polysaccharide (AHP) exhibit an excellent immune adjuvant effect, but low bioavailability in the body limits its application. Cationic polymer-modified poly (lactic-co-glycolic acid) (PLGA) nanoparticles have been widely investigated as vaccine delivery systems owing to their excellent antigen-loading efficiency. In this study, three kinds of cationic polymer were used to coat AHP-encapsulated PLGA nanoparticles (AHPP) to build positively charged antigen carriers. Among them, H5N1-loaded PEI-AHPP formulation could induce highest hemagglutination inhibition (HI) titer, IgG-subtype, and cytokines, activated dendritic cells (DCs) in lymph nodes, and CD3e+CD4+ and CD3e+CD8a+ T cells in the spleen of immunized mice. PEI-AHPP could stimulate DCs to highly express MHCI and MHCII molecules and had good antigen slow-release effect at the injected site along with lymph node targeting. These findings demonstrate that PEI-AHPP has the potential to be an effective adjuvant to induce strong and long-lasting Th1 and Th2 mixed immune responses.

Surface-Engineered Cubosomes Serve as a Novel Vaccine Adjuvant to Modulate Innate Immunity and Improve Adaptive Immunity in vivo.

OBJECTIVE: Recent studies have revealed the adjuvant activity of cubosomes and their potential utility as an antigen delivery system. In this study, to further enhance the adjuvant activity of cubosomes, two cationic polymers are modified on the surface of cubosomes. METHODS: Here, we exploit the effects of surface chemistry on the adjuvant activity of Ganoderma lucidum polysaccharide cubosomes (GLPC) by placing two kinds of molecules, that is, cetyltrimethylammonium bromide (CTAB) and poly(diallydimethyl ammonium chloride) (PDDAC), on their surface. RESULTS: CTAB- or PDDAC-modified GLPC were found to significantly promote humoral and cellular immune responses, as well as the proliferation of CD3+ CD4+ or CD3+ CD8+T cells through the powerful activation of dendritic cells (DCs). The enhanced immune responses of PDDAC-modified GLPC might be attributed to the maturation of DCs into draining lymph nodes and the activation of spleen and cytokines in serum. CONCLUSION: PDDAC modification is beneficial for enhancing humoral and cellular immune response, suggesting that PDDAC-GLPC-OVA has the ability to be a potential adjuvant for vaccine.

Polyethylenimine-coated PLGA nanoparticles-encapsulated Angelica sinensis polysaccharide as an adjuvant for H9N2 vaccine to improve immune responses in chickens compared to Alum and oil-based adjuvants.

Inactivated H9N2 influenza vaccines required adjuvants to induce strong immune responses to protect poultry from the infections of H9N2 influenza viruses. Recently, positively charged nanoparticles-based adjuvant delivery systems have been extensively investigated as the novel vaccine adjuvant due to the protection antigens and drugs from degradation, promoting antigens and drugs uptake by antigen presenting cells (APCs), and inducing strong humoral and cellular immune responses. In this study, the immunostimulant Angelica sinensis polysaccharide (ASP) was encapsulated into Poly (lactic-co-glycolic acid) PLGA nanoparticles, and the Polyethylenimine (PEI) was coated on the nanoparticles to develop a novel adjuvant (ASP-PLGA-PEI). To further investigate the adjuvant activities of ASP-PLGA-PEI nanoparticles for H9N2 vaccines in chickens and compare the adjuvant activities of nanoparticles adjuvant and conventional adjuvants (Alum and oil-based adjuvant), the H9N2 antigen was incubated with three different adjuvants and then immunized with chickens to evaluate the ability of inducing humoral and cellular immune responses. The results revealed that compared to Alum adjuvant, ASP-PLGA-PEI nanoparticles adjuvant stimulated higher antibody responses, promoted the activation of CD4+ T cells and CD8+ T cells, increased the expression of Th1 cytokines IFN-γ. Compared to oil-based adjuvant (ISA-206), ASP-PLGA-PEI nanoparticles adjuvant induced comparable antibody immune responses at later period after immunization, improved the activation of CD4+ T cells and CD8+ T cells. Therefore, compared to Alum and oil-based adjuvant, the ASP-PLGA-PEI nanoparticles serve as an efficient adjuvant for H9N2 vaccine and have the potential to induce vigorous humoral and cellular immune responses in chickens.

The Immunoenhancement Effects of Polyethylenimine-Modified Chinese Yam Polysaccharide-Encapsulated PLGA Nanoparticles as an Adjuvant.

BACKGROUND: Poly(lactic-co-glycolic acid) (PLGA) has been extensively applied for sustained drug delivery and vaccine delivery system. However, vaccines delivered by PLGA nanoparticles alone could not effectively activate antigen-presenting cells (APCs) to induce strong immune responses. PURPOSE: The aim of the present study was to design polyethylenimine (PEI)-modified Chinese yam polysaccharide (CYP)-encapsulated PLGA nanoparticles (CYPP-PEI) as a vaccine delivery system and evaluate the adjuvant activities in vitro and in vivo. MATERIALS AND METHODS: Cationic-modified nanoparticles exhibited high antigen absorption and could be efficiently taken by APCs to enhance the immune responses. Therefore, PEI-modified CYP-encapsulated PLGA nanoparticles (CYPP-PEI) were prepared. The storage stability and effective adsorption capacity for porcine circovirus-2 (PCV-2) antigen of these antigen-absorbed nanoparticles were measured for one month. Furthermore, the adjuvant activity of CYPP-PEI nanoparticles was evaluated on macrophages in vitro and through immune responses triggered by PCV-2 antigen in vivo. RESULTS: The PCV-2 absorbed CYPP-PEI nanoparticles showed excellent storage stability and high absorption efficiency of PCV-2 antigen. In vitro, CYPP-PEI nanoparticles promoted antigen uptake, enhanced surface molecular expressions of CD80 and CD86, and improved cytokine secretion of TNF-α, IFN-γ, and IL-12p70 in macrophages. After immunization with CYPP-PEI/PCV-2 formulation in mice, the expressions of surface activation markers on dendritic cells which located in draining lymph nodes were increased, such as MHCI, MHCII, and CD80. In addition, CYPP-PEI nanoparticles induced dramatically high PCV-2-specific IgG levels which could last for a long time and stimulated the secretion of subtype antibodies and cytokines. The results showed that CYPP-PEI could induce Th1/Th2 mixed but Th1-biased type immune responses. CONCLUSION: Polyethylenimine-modified Chinese yam polysaccharide-encapsulated PLGA nanoparticle was a potential vaccine delivery system to trigger strong and persistent immune responses.

Immune-adjuvant activity of lentinan-modified calcium carbonate microparticles on a H5N1 vaccine.

In recent years, the high prevalence of avian influenza viruses especially H5N1 subtype isolated from poultry and human has become a major public health concern. Vaccination is still a major strategy for preventing H5N1 infections. Lentinan (LNT), a ß-1,3-glucohexaose with ß-1,6-branches, is extracted from Lentinus edodes and has been extensively studied for its immunoenhancement effects. In this study, we synthesized and characterized calcium carbonate (CaCO3) microparticles which modified with LNT as an adjuvant for H5N1 vaccine and investigated their ability to enhance immune responses. We prepared spherical and uniform CaCO3-LNT microparticles with a mean hydrodynamic size was around 2 µm. The H5N1 antigen-loaded CaCO3-LNT particles were injected into mice to evaluate their effectiveness as an adjuvant for H5N1 vaccines. The results demonstrated that CaCO3-LNT/H5N1 significantly enhanced the expression of MHC-II and CD86 in lymph node dendritic cells, and increased the ratio of CD4+ to CD8+ T cells in lymphocytes. Moreover, CaCO3-LNT/H5N1 surprisingly increased the HI titers and induced the secretion of IgG subtypes (IgG1 and IgG2b) and Th-associated cytokines (TNF-α, IFN-γ and IL-4) in immunized mice. Therefore, by combining with the immunostimulatory activity of LNT and the drug/antigen delivery capabilities of CaCO3, the CaCO3-LNT/H5N1 could induce a stronger cellular and humoral immune response and could be a potential adjuvant for the H5N1 vaccine.

Lentinan-Functionalized Graphene Oxide Is an Effective Antigen Delivery System That Modulates Innate Immunity and Improves Adaptive Immunity.

Graphene oxide (GO) and lentinan have received great attention because of their utility in biomedical applications. Graphene oxide is utilized in drug- and vaccine-delivery systems due to its biocompatibility, large surface area, and outstanding adsorption capability, while lentinan has immunity-enhancing effects. In this study, we synthesized and characterized GO grafted with lentinan (LNT) as an adjuvant and investigated how to impact the immune responses. Lentinan-modified GO (GO-LNT) facilitated antigen uptake in macrophages and improved the efficiency of antigen application in vitro. Furthermore, in vivo, compared with GO/OVA, GO-LNT/OVA decreased the release rate of ovalbumin (OVA) to sustain long-term immune responses and boost the levels of IgG and IgG subtypes. Hence, we can infer that the effects of GO-LNT were a result of the increased amounts of antigen uptake by cells. Overall, our studies demonstrated that GO-LNT could suffice for a safe and effective vaccine-delivery system as well as an excellent adjuvant that both elicits a long-term immune memory response and potentiates cellular and humoral immunity.