Baker’s yeast glucan (BYG) has been reported to be an anti-diabetic agent. In the work described herein, further study on the effect of orally administered BYG on glucose and lipid homeostasis in the livers of ob/ob mice was performed. It was found that BYG decreased the blood glucose and the hepatic glucose and lipid disorders. Western blotting analysis revealed that BYG up-regulated p-AKT and p-AMPK, and down-regulated p-Acc in the liver. Furthermore, RNA-Seq analysis indicated that BYG down-regulated genes responsible for gluconeogenesis (G6pase and Got1), fatty acid biosynthesis (Acly, Acc, Fas, etc.), glycerolipid synthesis (Gpam and Lipin1/2), and cholesterol synthesis (Hmgcr, Fdps, etc.). Additionally, BYG decreased glucose transporters SGLT1 and GLUT2, fat emulsification, and adipogenic genes/proteins in the intestine to decrease glucose and lipid absorption. All these findings demonstrated that BYG is beneficial for regulating glucose and lipid homeostasis in diabetic mice, and thus has potential applications in anti-diabetic foods or drugs.Keywords: blood glucose; lipid metabolism; liver; type 2 diabetes; yeast β-glucan;

•t-LNT self-assembled into dendrite-like aggregates via parallel alignment in water.•t-LNT enhanced the blue fluorescence of TPE in water.•t-LNT/TPE showed high water-dispersibility, good stability and biocompatibility.We report a unique self-assembly of lentinan, a triple helical β-(1→3)-glucan (t-LNT), in water. By molecular dynamics simulation, it was found that t-LNT aggregated preferentially along the chain direction to form long chains, accompanied by side-direction linkage to form branches. Transmission electron microscopy images demonstrated that t-LNT formed dendrite-like fibers, which further formed fishnet-like porous/mesoporous aggregates with increasing concentration. The meshes in the fishnet were ascribed to the intersection of branches. The major driving force for aggregation was expected to be hydrogen bonding between hydroxyl groups in t-LNT chains. Based on this self-assembly behavior, a novel composite was prepared from t-LNT and tetraphenylethylene (TPE) by entrapping TPE aggregates into the meshes of t-LNT fishnets. The as-prepared t-LNT/TPE composite largely enhanced the blue fluorescence of TPE in water, exhibiting stable optical property and good biocompatibility, and t-LNT is expected to show great potential as a carrier of hydrophobic molecules for biomedical application.Download high-res image (166KB)Download full-size image

According to the principles of green chemistry and co-efficiency, natural polysaccharides with biological activities, particularly immunoregulation and antitumor activities, have attracted increasing attention. However, the lack of information concerning the pharmacokinetics of polysaccharides is one of the major obstacles limiting their rational clinical use. In this study, triple helical β-glucan (THG), a β-1,6-branched β-1,3-glucan isolated from Lentinus edodes, was studied to clarify its cellular uptake after parenteral (e.g. intraperitoneal) administration and to determine its effect on immune cells in murine tumor models. The results obtained from size exclusion chromatography with static light scattering, differential refractometry and viscometry (SEC-LLS–RI–Vis) and atomic force microscopy (AFM) confirmed that all three THG samples displayed an extended stiff chain conformation with apparent average contour lengths of 598 nm, 510 nm and 117 nm, respectively. The ex vivo results indicated that THG directly promoted cell proliferation of peritoneal macrophages, whole spleen cells and lymphocytes, and activated peritoneal macrophages with TNF-α production. In our findings, the intraperitoneally (i.p.) administrated THG was initially ingested by peritoneal resident macrophages, which transported it to lymph nodes, thymus and even tumor. Simultaneously, THG in macrophages was biodegraded into fragments with granular shapes and small size, which were sufficiently active to be easily ingested by neutrophils. Furthermore, THG fragments could promote the infiltration of macrophages, neutrophils and DCs into tumors, and also activate these cells to enhance their cytotoxicity toward tumor cells, leading to their apoptosis. This study provides important information concerning the ingestion and processing in vivo of triple helical β-glucan from natural products, leading to a better understanding of its antitumor mechanism through activating immune cells in murine tumor models.

Polysaccharides are naturally occurring biological macromolecules that are envisaged as promising substitutes of non-degradable polymers due to their outstanding inherent properties of biodegradability, biocompatibility, low-cost, and availability. Their utilization in the development of nanostructured hybrid materials has numerous advantages in theranostics, the integrated approach of therapeutics and diagnostics. In particular, some rigid polysaccharides occur in nature, which can self-assemble into ordered hierarchical structures, facilitating the controllable fabrication of various nanocomposites. These rigid polysaccharides, including Lentinan, Curdlan, Schizophyllan, Scleroglucan, Auricularian, and yeast glucan, possess the linear β-(1→3)-D-glucan as the backbone with or without β-(1→6)-linked glucose as the branch, having diverse biological activities. This review focuses on the incorporation of nanoparticles, such as gold (AuNPs), silver (AgNPs), selenium (SeNPs), silica, carbon nanotubes, homo-polynucleotides, bio-imaging probes, and drugs, into different rigid polysaccharide matrices through self-assembly and summarizes their biological functions as well as the correlation to their conformations. Additionally, we addressed the use of the as-engineered polysaccharide nanocomposites as effective therapeutic agents, and the challenges or ambiguity issues concerning further practical clinic therapeutic applications.

Dendritic nanotubes (DNTs) with hydrophobic cavities were constructed directly from rigid branched β-1,3-D-glucan (AF1) in aqueous solution, and the AF1 sample was isolated from the fruiting bodies of Auricularia auricula-judae, a household nutritional food. The structure of AF1 dendritic nanotubes was demonstrated with a transmission electron microscope (TEM) and a scanning electron microscope (SEM), and a schematic diagram was proposed to describe the formation process, which was supported by the results of static/dynamic light scattering (SLS/DLS) and atomic force microscopy (AFM). In solution, a sequential self-assembly of the AF1 chains in a parallel manner occurred to form lamellas followed by self-curling into nanotubes with the mean diameters from 20 to 80 nm, depending on the concentration and molecular weight of AF1, through hydrogen bonding and hydrophilic/hydrophobic interaction. As a result of the dendritic structure, the AF1 aggregates exhibited highly condensed hydrophobic regions, which could be used as carriers to achieve a high concentration of the target molecules. In our findings, the anticancer drug DOX and the fluorescent probe TPA-BMO could be loaded into the hydrophobic region of DNTs. Interestingly, DOX-loaded DNTs of AF1 exhibited high drug loading capacity and pH-triggered sustained release behaviors (>23 days) with reduced cytotoxicity in vitro. Moreover, the bioimaging experiment demonstrated that TPA-BMO-loaded DNTs of AF1 induced stronger fluorescence intensity than TPA-BMO alone, and maintained a longer duration time (18 days) in vivo. Therefore, the DNTs of AF1 have promising applications as bioactive carriers, especially in the fields of drug delivery and bioimaging.

Mushrooms are known as a delicacy due to their delicious taste and rich nutrition, and their β-glucans have antitumor activity. Here, a triple helical β-glucan (THG) isolated from Lentinus edodes was successfully fractionated into nine fractions with different weight-average molecular weights (Mw) through ultrasonic irradiation. The Mw, radius of gyration (〈Rg〉z), hydrodynamic radius (〈Rh〉z), structure-sensitive parameters (ρ), contour length (L), persistence length (q) and molar mass per contour length (ML) were characterized by static light scattering (SLS), dynamic light scattering (DLS), and atomic force microscopy (AFM). The results indicated that THG displayed an extended chain conformation with ρ values of 2.1 ± 0.1 for the nine fractions, as well as ML and q values of 2160 nm−1 and 110 nm in water, which were consistent with the data for triple helical polysaccharides. In combination with the apparent length (Lap) values visualized with AFM and Mw, the molar mass per apparent contour length (MLap) was calculated to be 2242 nm−1, which was similar to that obtained from SLS according to the wormlike cylinder model. Thus, we established a novel method using AFM for characterizing the chain stiffness of polysaccharides. The results from an animal assay demonstrated that THG significantly inhibited H22 tumor growth without damage to organisms, and the THG fractions with relatively low molecular weight and/or higher stiffness showed stronger antitumor activity, revealing the significant molecular weight- and chain conformation-dependences of antitumor activity. Moreover, a schematic model to describe the interaction between THG and receptors on the immune cell membrane was proposed to illustrate these results. This work provides important information for characterizing the chain conformation of polysaccharides and understanding the relationship between structure and antitumor activity, which is relevant for the treatment of hepatocellular carcinoma in clinic.

The water soluble β-(1 → 3)-D-glucan with short branches (AF1) isolated from Auricularia auricula-judae was successfully fractionated by ultrasonication into three fractions with different weight-average molecular weights (Mws). The results of static and dynamic laser light scattering, viscometry and atomic force microscopy confirmed that the AF1 samples adopted a stiff chain conformation in water, and the coexistence of individuals and aggregates occurred gradually with increasing concentration. The AF1 sample with the highest Mw easily self-entangled, and exhibited a strong shear rate-dependence of viscosity in water. The glucans displayed anti-hepatoma activity and significantly inhibited H22 tumour growth without cytotoxicity towards normal tissues. They displayed both molecular weight- and dosage-dependencies of anti-tumour activity, and the sample with an Mw of 7.7 × 105 at the dosage of 5 mg kg−1 exhibited the highest inhibition ratio of ∼77% against H22 tumour, even significantly higher than the positive control of cytoxan. The immunohistochemical and western blot analyses revealed that the AF1 glucans triggered cell apoptosis, indicated by the activation of caspase 3/9 and down-regulated tumour angiogenesis factors of VEGF and CD31. The underlying antitumor mechanism was suggested to induce tumour cell apoptosis and to inhibit angiogenesis in tumour tissues via enhancement of the immune-response. Taken together, the AF1 β-glucan was a potent natural drug candidate with high anti-cancer activities and less cytotoxicity, and the AF1 sample with a moderate molecular weight existed in aqueous solution as a more extended chain conformation, which plays an important role in activating immune responses.

•PR-CA was characterized to be a β-(1→3)-d-glucan with multiple branches.•The chain stiffness of PR-CA in water was directly visualized by AFM and TEM.•The aggregation behavior of PR-CA was studied in water/DMSO solutions.•The branched PR-CA chains took parallel aggregation to form hollow nanofibers.A polysaccharide coded as PR-CA was extracted from Polyporus rhinoceros and determined to be a β-(1→3)-d-glucan with multiple branches. The weight-average molecular weights (Mw) of PR-CA in dimethylsulfoxide (DMSO) and in water were determined with static light scattering (SLS) to be 3.57 × 105 and 1.79 × 107, indicating existence of the single chains in DMSO and co-existence of single chains and aggregates in water. Moreover, the stiffness of single chains of PR-CA in water was directly visualized by atomic force microscopy (AFM) and transmission electron microscopy (TEM). The hollow structure of PR-CA nanofibers with width of 30–40 nm and length of ∼350 nm formed in the water/DMSO (9:1, v:v) was demonstrated by a fluorescent probe tetraphenylethylene (TPE) via aggregation-induced emission (AIE). The formation of PR-CA nanofibers was ascribed to the parallel aggregation of the extended PR-CA chains due to the hydrogen bonding and hydrophobic interaction. This work offered valuable results for promising applications of natural branched β-glucans in the biological fields of drug inclusion, delivery and disease diagnosis.

•Stiff branched β-glucan in water transformed into flexible chain in DMSO.•The viscosity and chain size of AF1 drop abruptly for vDMSO >0.8.•Breaking of multiple hydrogen bonds led to the conformation transition of AF1.The conformation transition of the short branched β-glucan AF1 isolated from Auricularia auricula-judae in DMSO/water solutions was investigated with viscometry and static/dynamic light scattering (SLS/DLS). AF1 in the mixed solution exhibited a sharp decrease in viscosity and molecular size in a narrow DMSO volume fraction (vDMSO) range of 0.80–1.0. It indicated that the stiff AF1 transformed into flexible chains probably, resulting from the destruction of intra- and inter-molecular hydrogen bonds of the polysaccharides. AF1 solutions with dialyzed treatment gave a more narrow transition range of 0.90–1.0. The conformation transition of AF1 was further conformed by transition electron microscopy (TEM) and atomic force microscopy (AFM). The multiple hydrogen bonds of polysaccharides affect the chain conformation, which in turn affect the application in food and pharmaceutical fields.

•A highly purified β-glucan (BBG) was acquired by hot water and alkali extraction.•BBG activates macrophages to produce cytokines TNF-α and MCP-1.•BBG activates macrophages through targeting CR3/TLR2 signaling pathway.Yeast β-glucan has many formulations with different chemical structures, water solubility and purity. In particular, the purity of β-glucan in these formulations is variable and relatively low, contributing to different data on its biological activity. In this study, the major polysaccharide component in the crude Baker’s yeast polysaccharides coded as BBG with high purity of 99% was obtained, and its chemical structure was determined to be a linear β-(1,3)-glucan. It was found that BBG interacted with complement receptor 3 (CR3) and toll-like receptor 2 (TLR2) on the surface of macrophage-like RAW264.7 cells, and initiated activation of RAW264.7 cells characterized by significant production of tumor necrosis factor-α (TNF-α) and monocyte chemoattractant protein 1 (MCP-1). Additionally, activation of the nuclear factor kappaB p65 (NF-κB p65), c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) induced by BBG, were also observed, further confirming the stimulation of RAW264.7 cells by BBG. All these findings provided important scientific evidences for better understanding the molecular mechanism of action for the linear β-(1,3)-glucan in cells.

A novel gene carrier was prepared from a triple-helical β-glucan LNT (t-LNT) on the basis of the special interaction between the single chain of t-LNT (s-LNT) and polydeoxyadenylic acid (poly(dA)). The effects of the chain length of poly(dA) in the target DNA sequence on the stability and gene transfection efficiency of the gene carrier of the poly(dA)–s-LNT complex were evaluated by agarose gel retardation assay, circular dichroism, flow cytometry, and ELISA kits. All the results demonstrated that the chain length of poly(dA) that produced the strongest interaction with s-LNT was favorable for the stability and gene transfection efficiency. Moreover, the gene transfection efficiency was strikingly dependent on the transfection time and temperature, suggesting the internalization of the gene delivery system via endocytosis. The high expressions of receptors including TLR2 and CR3 (CD11b/CD18) on the surfaces of RAW264.7 cells were confirmed by confocal microscopy and flow cytometry. By blocking TLR2, CD11b, and CD18 with the corresponding specific antibodies, the transfection efficiency was hardly changed at 4 °C or 37 °C. This suggested that the internalization pattern of such a delivery system was not via the usual β-glucan receptor-mediated endocytosis (such as dectin-1, TLR2, and CR3), which was different from the reported uptake mechanism. Overall, this work facilitates the understanding of the pathways of delivery systems involving β-glucans, which has significant implications for potential applications in disease treatment.

•We report on a green procedure for the stabilization of selenium nanoparticles by a β-glucan.•Selenium nanoparticles/β-glucan composites with different size were constructed.•The antitumor activity of selenium nanoparticles was enhanced by complexation with β-glucan.•The effect of size of selenium nanoparticles on antitumor activity was investigated.We report on a green procedure for the stabilization of selenium nanoparticles (SeNPs) by a naturally occurring β-glucan with triple helical conformation known as Lentinan (t-LNT) in water after denaturing into single chains (s-LNT) at 140 °C. The results demonstrated that the s-LNT can interact with SeNPs through SeOH interaction. Transmission electron microscopy (TEM), energy dispersive X-ray (EDX) spectra, UV/vis, X-ray diffraction (XRD) and dynamic light scattering (DLS) showed that s-LNT coated SeNPs to form a stable nano-composite Se/s-LNT, leading to good dispersion of SeNPs. Especially, the as-prepared Se/s-LNT composite in the solution could remain homogeneous and translucent for 30 days without any precipitates. Different size distribution of SeNPs was prepared by simply controlling the concentrations of selenite sodium and the corresponding reducing agent ascorbic acid. The size effect of SeNPs on anti-tumor activity was revealed that the SeNPs with more evenly particle size distribution show the higher anticancer activity.

The intra- and intermolecular interactions of chitin in NaOH/urea aqueous system were studied by a combination of NMR measurements (including 13C NMR, 23Na NMR, and 15N NMR) and differential scanning calorimetry. The results revealed that the NaOH and chitin formed a hydrogen-bonded complex that was surrounded by the urea hydrates to form a sheath-like structure, leading to the good dissolution. The optimal concentration range, in which chitin was molecularly dispersed in NaOH/urea aqueous, was found to investigate the chain conformation in the dilute solution with a combination of static and dynamic light scattering. The weight-average molecular weight (Mw), radii of gyration (⟨Rg⟩z), and hydrodynamic radii (⟨Rh⟩z) values of chitin were determined, and the structure-sensitive parameter (ρ) and persistent length (Lp) were calculated to be >2.0 and ∼30 nm, respectively, suggesting an extended wormlike chain conformation. The visualized images from TEM, cryo-TEM, and AFM indicated that, chitin nanofibers were fabricated from the parallel aggregation of chitin chains in NaOH/urea system. This work would provide a theoretical guidance for constructing novel chitin-based nanomaterials via “bottom-up” method at the molecular level.

A naturally occurring β-glucan from Lentinus edodes (t-LNT) has been proved to adopt a unique triple helix structure due to strong hydrogen bonding. The single chain of t-LNT (s-LNT) can interact with polydeoxyadenylic acid (poly(dA)) through hydrogen bonding between s-LNT and poly(dA). Based on this, a novel strategy for an efficient gene transfection was developed as follows. The target DNA was attached to poly(dA)50 through a disulfide bond to form DNA–SS–poly(dA)50, and s-LNT was then combined with the poly(dA)50 segment to form a DNA–SS–poly(dA)50/s-LNT complex through hydrogen bonding, which could be taken up by cells followed by the release of target DNA in the cytoplasm via the breaking of the disulfide bond under the treatment of reducing agents such as GSH. The experimental results from the agarose gel retardation assay and circular dichroism demonstrated the formation of the complex DNA–SS–poly(dA)50/s-LNT. Confocal microscopy and flow cytometry indicated that the target DNA was delivered into the cells with high transfection efficiency, which was strongly dependent on molecular weight and the concentration of s-LNT. The uptake mechanism of the complex was ascribed to endocytosis. The significant enhancement of IL-12p40 production in macrophage RAW 264.7 cells stimulated by the transfected CpG DNA with immunomodulating activity further confirmed the feasibility of this system. Additionally, gene transfection with high efficiency could be performed in tumor cells (HeLa). The cytotoxicity of s-LNT against RAW 264.7 and HeLa cells by the MTT assay showed good safety. Overall, s-LNT could be used as an excellent gene carrier, indicating its promising application in gene delivery.

The development of biological high-performance materials fabricated from natural polysaccharides has attracted great attention for a sustainable world. In this work, hollow fibers with high strength were spun from a polysaccharide aqueous solution at a concentration of 0.02 g mL−1. The polysaccharide was a comb-like β-glucan with short branches isolated from Auricularia auricula-judae, coded as AF1. Atomic force microscopy (AFM) and transmission electron microscopy (TEM) confirmed directly that AF1 existed as a stiff chain conformation in water, and displayed parallel self-orientation behavior. AF1 could self-assemble into well defined hollow nanofibers with diameters less than 100 nm and lengths of tens of micrometers in dilute solution, supported by scanning electron microscopy (SEM). Moreover, AF1 in the disulfonated tetraphenylethene (TPE-SO3Na) aqueous solution exhibited strong luminescence, indicating that the TPE-SO3Na molecules without luminescence in water were trapped in the cavities of the hollow nanofibers through hydrophobic interactions, leading to the aggregation-induced emission (AIE). The nanofibers were composed of relatively hydrophobic inner-walls and hydrophilic shells in water. Interestingly, SEM and polarized light microscopy verified that the nanofibers fused to form an ordered architecture of lamella and then tended to curl into hollow fibers in relatively concentrated solution. The hollow fibers exhibited excellent tensile strength, biocompatibility, organic solvent resistance and birefringence. A schematic model was proposed to describe the construction of the hollow fibers via the hierarchical self-assembly process. The new materials would have potential applications such as drug release as a new class of fibrous carrier, indicators with fluorescence to detect cell growth in cell transplantation, and biomolecular recognition (e.g., DNA).

Based on the fluorescence resonance energy transfer (FRET) and fluorescence anisotropy (FA), the present study reported proof-of-principle for a highly sensitive and rapid detection technique that can be precisely utilized for investigating the self-assembly of polydeoxyadenylic acid (poly(dA)) and β-glucan, and the interactions of the poly(dA)–β-glucan complex on the surface of graphene oxide (GO). Due to the noncovalent assembly of fluorescein amidite (FAM)-labeled poly(dA) and GO via π–π stacking, the fluorescence of (FAM)-labeled poly(dA) as a molecular aptamer beacon (MAB) was completely quenched by GO. Conversely, the addition of single-stranded lentinan (s-LNT) resulted in the significant restoration of fluorescence due to the formation of poly(dA)–s-LNT complexes with a stiff rod-like structure, which had a weak affinity to GO and kept the dyes away from GO. However, relatively weak fluorescence restoration was observed by adding another single-stranded curdlan (s-CUR) for positive control, indicative of complex formation with higher binding ability to GO. The fluorescence anisotropy (FA) was also combined to confirm the occurrence with different increments of anisotropy relative to the free poly(dA), which could be conveniently extended for detecting the assembly of other biomolecules with higher sensitivity.

We report on a green procedure for the synthesis and stabilization of gold nanoparticles (AuNPs) from chlorauric acid (HAuCl4) with the use of a β-glucan known as Lentinan (LNT) without external reducing or stabilizing agents in aqueous medium. LNT adopted triple helical conformation in water, which was first denatured into single chains (s-LNT) at a high temperature of 140 °C before mixing with HAuCl4. Results from UV–vis absorption spectroscopy, transmission electron microscopy (TEM), and energy dispersive X-ray (EDX) spectra suggested that AuCl4– was rapidly reduced to AuNPs by s-LNT. Moreover, the as-prepared AuNPs could be converted into nanobelt, spherical nanoparticles, and nanowire morphology simply by controlling the s-LNT concentration, reaction time, and temperature. In particular, the AuNPs nanowire was confirmed as the most stable shape in water, which was predominately ascribed to the hydrophobic cavity in the helical center of the renatured triple helical LNT (r-LNT) from s-LNT. Namely, AuNPs were entrapped in the hydrophobic cavity of r-LNT to form nanowire with an outer layer of water-soluble r-LNT, leading to stable dispersion of AuNPs. All the data demonstrated that the β-glucan of s-LNT can be used as a reducing and stabilizing agent to synthesize and disperse AuNPs in water. The whole process of reduction and stabilization was free of organic solvent and thus very safe, which is important for the potential application of AuNPs in biotechnology and biomedicine.

Dendritic nanotubes (DNTs) with hydrophobic cavities were constructed directly from rigid branched β-1,3-D-glucan (AF1) in aqueous solution, and the AF1 sample was isolated from the fruiting bodies of Auricularia auricula-judae, a household nutritional food. The structure of AF1 dendritic nanotubes was demonstrated with a transmission electron microscope (TEM) and a scanning electron microscope (SEM), and a schematic diagram was proposed to describe the formation process, which was supported by the results of static/dynamic light scattering (SLS/DLS) and atomic force microscopy (AFM). In solution, a sequential self-assembly of the AF1 chains in a parallel manner occurred to form lamellas followed by self-curling into nanotubes with the mean diameters from 20 to 80 nm, depending on the concentration and molecular weight of AF1, through hydrogen bonding and hydrophilic/hydrophobic interaction. As a result of the dendritic structure, the AF1 aggregates exhibited highly condensed hydrophobic regions, which could be used as carriers to achieve a high concentration of the target molecules. In our findings, the anticancer drug DOX and the fluorescent probe TPA-BMO could be loaded into the hydrophobic region of DNTs. Interestingly, DOX-loaded DNTs of AF1 exhibited high drug loading capacity and pH-triggered sustained release behaviors (>23 days) with reduced cytotoxicity in vitro. Moreover, the bioimaging experiment demonstrated that TPA-BMO-loaded DNTs of AF1 induced stronger fluorescence intensity than TPA-BMO alone, and maintained a longer duration time (18 days) in vivo. Therefore, the DNTs of AF1 have promising applications as bioactive carriers, especially in the fields of drug delivery and bioimaging.

Mushrooms are known as a delicacy due to their delicious taste and rich nutrition, and their β-glucans have antitumor activity. Here, a triple helical β-glucan (THG) isolated from Lentinus edodes was successfully fractionated into nine fractions with different weight-average molecular weights (Mw) through ultrasonic irradiation. The Mw, radius of gyration (〈Rg〉z), hydrodynamic radius (〈Rh〉z), structure-sensitive parameters (ρ), contour length (L), persistence length (q) and molar mass per contour length (ML) were characterized by static light scattering (SLS), dynamic light scattering (DLS), and atomic force microscopy (AFM). The results indicated that THG displayed an extended chain conformation with ρ values of 2.1 ± 0.1 for the nine fractions, as well as ML and q values of 2160 nm−1 and 110 nm in water, which were consistent with the data for triple helical polysaccharides. In combination with the apparent length (Lap) values visualized with AFM and Mw, the molar mass per apparent contour length (MLap) was calculated to be 2242 nm−1, which was similar to that obtained from SLS according to the wormlike cylinder model. Thus, we established a novel method using AFM for characterizing the chain stiffness of polysaccharides. The results from an animal assay demonstrated that THG significantly inhibited H22 tumor growth without damage to organisms, and the THG fractions with relatively low molecular weight and/or higher stiffness showed stronger antitumor activity, revealing the significant molecular weight- and chain conformation-dependences of antitumor activity. Moreover, a schematic model to describe the interaction between THG and receptors on the immune cell membrane was proposed to illustrate these results. This work provides important information for characterizing the chain conformation of polysaccharides and understanding the relationship between structure and antitumor activity, which is relevant for the treatment of hepatocellular carcinoma in clinic.

Polysaccharides are naturally occurring biological macromolecules that are envisaged as promising substitutes of non-degradable polymers due to their outstanding inherent properties of biodegradability, biocompatibility, low-cost, and availability. Their utilization in the development of nanostructured hybrid materials has numerous advantages in theranostics, the integrated approach of therapeutics and diagnostics. In particular, some rigid polysaccharides occur in nature, which can self-assemble into ordered hierarchical structures, facilitating the controllable fabrication of various nanocomposites. These rigid polysaccharides, including Lentinan, Curdlan, Schizophyllan, Scleroglucan, Auricularian, and yeast glucan, possess the linear β-(1→3)-D-glucan as the backbone with or without β-(1→6)-linked glucose as the branch, having diverse biological activities. This review focuses on the incorporation of nanoparticles, such as gold (AuNPs), silver (AgNPs), selenium (SeNPs), silica, carbon nanotubes, homo-polynucleotides, bio-imaging probes, and drugs, into different rigid polysaccharide matrices through self-assembly and summarizes their biological functions as well as the correlation to their conformations. Additionally, we addressed the use of the as-engineered polysaccharide nanocomposites as effective therapeutic agents, and the challenges or ambiguity issues concerning further practical clinic therapeutic applications.

The development of biological high-performance materials fabricated from natural polysaccharides has attracted great attention for a sustainable world. In this work, hollow fibers with high strength were spun from a polysaccharide aqueous solution at a concentration of 0.02 g mL−1. The polysaccharide was a comb-like β-glucan with short branches isolated from Auricularia auricula-judae, coded as AF1. Atomic force microscopy (AFM) and transmission electron microscopy (TEM) confirmed directly that AF1 existed as a stiff chain conformation in water, and displayed parallel self-orientation behavior. AF1 could self-assemble into well defined hollow nanofibers with diameters less than 100 nm and lengths of tens of micrometers in dilute solution, supported by scanning electron microscopy (SEM). Moreover, AF1 in the disulfonated tetraphenylethene (TPE-SO3Na) aqueous solution exhibited strong luminescence, indicating that the TPE-SO3Na molecules without luminescence in water were trapped in the cavities of the hollow nanofibers through hydrophobic interactions, leading to the aggregation-induced emission (AIE). The nanofibers were composed of relatively hydrophobic inner-walls and hydrophilic shells in water. Interestingly, SEM and polarized light microscopy verified that the nanofibers fused to form an ordered architecture of lamella and then tended to curl into hollow fibers in relatively concentrated solution. The hollow fibers exhibited excellent tensile strength, biocompatibility, organic solvent resistance and birefringence. A schematic model was proposed to describe the construction of the hollow fibers via the hierarchical self-assembly process. The new materials would have potential applications such as drug release as a new class of fibrous carrier, indicators with fluorescence to detect cell growth in cell transplantation, and biomolecular recognition (e.g., DNA).

The water soluble β-(1 → 3)-D-glucan with short branches (AF1) isolated from Auricularia auricula-judae was successfully fractionated by ultrasonication into three fractions with different weight-average molecular weights (Mws). The results of static and dynamic laser light scattering, viscometry and atomic force microscopy confirmed that the AF1 samples adopted a stiff chain conformation in water, and the coexistence of individuals and aggregates occurred gradually with increasing concentration. The AF1 sample with the highest Mw easily self-entangled, and exhibited a strong shear rate-dependence of viscosity in water. The glucans displayed anti-hepatoma activity and significantly inhibited H22 tumour growth without cytotoxicity towards normal tissues. They displayed both molecular weight- and dosage-dependencies of anti-tumour activity, and the sample with an Mw of 7.7 × 105 at the dosage of 5 mg kg−1 exhibited the highest inhibition ratio of ∼77% against H22 tumour, even significantly higher than the positive control of cytoxan. The immunohistochemical and western blot analyses revealed that the AF1 glucans triggered cell apoptosis, indicated by the activation of caspase 3/9 and down-regulated tumour angiogenesis factors of VEGF and CD31. The underlying antitumor mechanism was suggested to induce tumour cell apoptosis and to inhibit angiogenesis in tumour tissues via enhancement of the immune-response. Taken together, the AF1 β-glucan was a potent natural drug candidate with high anti-cancer activities and less cytotoxicity, and the AF1 sample with a moderate molecular weight existed in aqueous solution as a more extended chain conformation, which plays an important role in activating immune responses.

A novel gene carrier was prepared from a triple-helical β-glucan LNT (t-LNT) on the basis of the special interaction between the single chain of t-LNT (s-LNT) and polydeoxyadenylic acid (poly(dA)). The effects of the chain length of poly(dA) in the target DNA sequence on the stability and gene transfection efficiency of the gene carrier of the poly(dA)–s-LNT complex were evaluated by agarose gel retardation assay, circular dichroism, flow cytometry, and ELISA kits. All the results demonstrated that the chain length of poly(dA) that produced the strongest interaction with s-LNT was favorable for the stability and gene transfection efficiency. Moreover, the gene transfection efficiency was strikingly dependent on the transfection time and temperature, suggesting the internalization of the gene delivery system via endocytosis. The high expressions of receptors including TLR2 and CR3 (CD11b/CD18) on the surfaces of RAW264.7 cells were confirmed by confocal microscopy and flow cytometry. By blocking TLR2, CD11b, and CD18 with the corresponding specific antibodies, the transfection efficiency was hardly changed at 4 °C or 37 °C. This suggested that the internalization pattern of such a delivery system was not via the usual β-glucan receptor-mediated endocytosis (such as dectin-1, TLR2, and CR3), which was different from the reported uptake mechanism. Overall, this work facilitates the understanding of the pathways of delivery systems involving β-glucans, which has significant implications for potential applications in disease treatment.

A naturally occurring β-glucan from Lentinus edodes (t-LNT) has been proved to adopt a unique triple helix structure due to strong hydrogen bonding. The single chain of t-LNT (s-LNT) can interact with polydeoxyadenylic acid (poly(dA)) through hydrogen bonding between s-LNT and poly(dA). Based on this, a novel strategy for an efficient gene transfection was developed as follows. The target DNA was attached to poly(dA)50 through a disulfide bond to form DNA–SS–poly(dA)50, and s-LNT was then combined with the poly(dA)50 segment to form a DNA–SS–poly(dA)50/s-LNT complex through hydrogen bonding, which could be taken up by cells followed by the release of target DNA in the cytoplasm via the breaking of the disulfide bond under the treatment of reducing agents such as GSH. The experimental results from the agarose gel retardation assay and circular dichroism demonstrated the formation of the complex DNA–SS–poly(dA)50/s-LNT. Confocal microscopy and flow cytometry indicated that the target DNA was delivered into the cells with high transfection efficiency, which was strongly dependent on molecular weight and the concentration of s-LNT. The uptake mechanism of the complex was ascribed to endocytosis. The significant enhancement of IL-12p40 production in macrophage RAW 264.7 cells stimulated by the transfected CpG DNA with immunomodulating activity further confirmed the feasibility of this system. Additionally, gene transfection with high efficiency could be performed in tumor cells (HeLa). The cytotoxicity of s-LNT against RAW 264.7 and HeLa cells by the MTT assay showed good safety. Overall, s-LNT could be used as an excellent gene carrier, indicating its promising application in gene delivery.