Takeshi Tsuka

Effects of oral administration of fucoidan extracted from Cladosiphon okamuranus on tumor growth and survival time in a tumor-bearing mouse model

We evaluated the anti-tumor activities of the oral administration of fucoidan extracted from Cladosiphon okamuranus using a tumor (colon 26)-bearing mouse model. The materials used included low-molecular-weight fucoidan (LMWF: 6.5–40 kDa), intermediate-molecular-weight fucoidan (IMWF: 110–138 kDa) and high-molecular-weight fucoidan (HMWF: 300–330 kDa). The IMWF group showed significantly suppressed tumor growth. The LMWF and HMWF groups showed significantly increased survival times compared with that observed in the control group (mice fed a fucoidan-free diet). The median survival times in the control, LMWF, IMWF and HMWF groups were 23, 46, 40 and 43 days, respectively. It was also found that oral administration of fucoidan increased the population of natural killer cells in the spleen. Furthermore, from the results of the experiment using Myd-88 knockout mice, it was found that these effects are related to gut immunity. These results suggest that fucoidan is a candidate anti-tumor functional food.

α-Chitin nanofibrils improve inflammatory and fibrosis responses in inflammatory bowel disease mice model

We evaluated the anti-inflammatory and anti-fibrosis effects of α-chitin nanofibrils in a mouse model of dextran sulfate sodium (DSS)-induced acute ulcerative colitis (UC). α-Chitin nanofibrils decreased positive areas of nuclear factor-κB staining in the colon tissue (7.2±0.5%/fields in the α-chitin nanofibrils group vs. 10.7±0.9%/fields in the control group; p<0.05). α-Chitin nanofibrils also decreased serum monocyte chemotactic protein-1 concentration in DSS-induced acute UC (24.1±7.8 pg/ml in the α-chitin nanofibrils group vs. 53.5±3.1 pg/ml in the control group; p<0.05). Moreover, α-chitin nanofibrils suppressed the increased positive areas of Massons trichrome staining in colon tissue (6.8±0.6%/fields in the α-chitin nanofibrils group vs. 10.1±0.7%/fields in the control group; p<0.05). On the other hand, α-chitin powder suspension did not show these effects in DSS-induced acute UC mice model. Our results indicated that α-chitin nanofibrils have the anti-inflammatory effect via suppressing NF-κB activation and the anti-fibrosis effects in DSS-induced acute UC mice model.

Evaluation of the effects of chitin nanofibrils on skin function using skin models.

Chitins are highly crystalline structures that are predominantly found in crustacean shells. Alpha-chitin is composed of microfibers, which are made up of nanofibrils that are 2-5 nm in diameter and 30 nm in length and embedded in a protein matrix. Crystalline nanofibrils can also be prepared by acid treatment. We verified the effect of chitin nanofibrils (NF) and nanocrystals (NC) on skin using a three-dimensional skin culture model and Franz cells. The application of NF and NC to skin improved the epithelial granular layer and increased granular density. Furthermore, NF and NC application to the skin resulted in a lower production of TGF-β compared to that of the control group. NF and NC might have protective effects to skin. Therefore, their potential use as components of skin-protective formulations merits consideration.

Biological adhesive based on carboxymethyl chitin derivatives and chitin nanofibers

Novel biological adhesives made from chitin derivatives were prepared and evaluated for their adhesive properties and biocompatibility. Chitin derivatives with acrylic groups, such as 2-hydroxy-3-methacryloyloxypropylated carboxymethyl chitin (HMA-CM-chitin), were synthesized and cured by the addition of an aqueous hydrogen peroxide solution as a radical initiator. The adhesive strength of HMA-CM-chitin increased when it was blended with chitin nanofibers (CNFs) or surface-deacetylated chitin nanofibers (S-DACNFs). HMA-CM-chitin/CNFs or HMA-CM-chitin/S-DACNFs have almost equal adhesive strength compared to that of a commercial cyanoacrylate adhesive. Moreover, quick adhesion and induction of inflammatory cells migration were observed in HMA-CM-chitin/CNF and HMA-CM-chitin/S-DACNF. These findings indicate that the composites prepared in this study are promising materials as new biological adhesives.

Favorable effects of superficially deacetylated chitin nanofibrils on the wound healing process.

Previous reports indicate that the beneficial effect of chitin nanofibrils (CNFs), and chitosan nanofibrils (CSNFs) for wound healing. In this study, the wound healing effects of superficially deacetylated chitin nanofibrils (SDACNFs) were evaluated using an experimental model. In the experiments using circular excision wound model, SDACNFs induced re-epithelium and proliferation of the fibroblasts and collagen tissue. In the chitin, CNFs, and CSNFs, on the other hand, the e-epithelium and proliferation of the fibroblasts and collagen tissue were not induced perfectly compared with the SDACNFs group. In particular, re-epithelization was observed on day 4 in the only SDACNF group. Moreover, SDACNFs did not induce severe systemic inflammation in the linear incision wound model. The data indicated that SDACNFs effectively induced the proliferation and re-modeling phases compared with chitin, CNFs, and CSNFs in the wound. These data indicate that SDACNFs can be beneficial as a new biomaterial for wound healing.

Anti-inflammatory effects of orally administered glucosamine oligomer in an experimental model of inflammatory bowel disease.

Anti-inflammatory effects of oral administration of the glucosamine oligomers (chito-oligosaccharides: COS) were evaluated in an experimental model of inflammatory bowel disease (IBD). Oral administration of COS improved shortening of colon length and tissue injury (as assessed by histology) in mice. Oral administration of COS inhibited inflammation in the colonic mucosa by suppression of myeloperoxidase activation in inflammatory cells, as well as activation of nuclear factor-kappa B, cyclooxygenase-2, and inducible nitric oxide synthase. Oral administration of COS also reduced serum levels of pro-inflammatory cytokines (tumor necrosis factor-α and interleukin-6). Moreover, it prolonged survival time in mice. These data suggest that COS have anti-inflammatory effects in an experimental model of IBD, and could be new functional foods for IBD patients.

Anti-tumor properties of orally administered glucosamine and N-acetyl-D-glucosamine oligomers in a mouse model.

The current study evaluated the anti-tumor activities of N-acetyl-d-glucosamine oligomer (NACOS) and glucosamine oligomer (COS) after their oral administration in a tumor (colon 26)-bearing mouse model. Compared to the control group, NACOS and COS groups showed significantly suppressed tumor growth, and apparent, marked apoptosis in tumor tissues. Furthermore, serum interleukin-12p70 and interferon-γ levels significantly increased in the NACOS and COS groups compared to the corresponding levels in the control group. Collectively, the results indicate the oral administration of NACOS and COS could enhance innate immunity. Results of experiments in Myd-88 knockout mice revealed that the apparent effects were related to both Myd-88-dependent and Myd-88-independent pathways. The data indicated that oral administration of NACOS and COS produced anti-tumor effects through the induction of apoptosis and stimulation of the immune system, which suggests that NACOS and COS are candidate anti-tumor functional foods.

Antitumor effects of high‑temperature hyperthermia on a glioma rat model

High-temperature hyperthermia (HTH) is an established treatment option for cancer. The aim of the present study was to reveal the exact correlation between HTH at temperatures of 50–70°C and the resulting antitumor effects, using a glioma rat model. In the 60°C (T-60) and 70°C (T-70) HTH groups, tumor growth rates were significantly suppressed compared with those in the nontreatment (NT) group. In the 50°C (T-50) HTH group, tumor growth rates were not suppressed compared with those in the NT group. The numbers of terminal dUTP nick-end labeling-positive cells in tumor tissue were significantly higher in the T-50, T-60 and T-70 groups than those in the NT group. The Ki-67-positive areas were significantly decreased in the T-70 group compared with those in the NT and T-60 groups. Our data indicate that HTH at 60 and 70°C suppresses tumor growth in a glioma rat model. In particular, cell proliferation was significantly suppressed by HTH at 70°C. However, differences in the mechanism of action of HTH at 60 and 70°C were observed.

Quantitative evaluation of bone development of the distal phalanx of the cow hind limb using computed tomography

Computed tomography (CT) was performed on 400 claws (200 inner and 200 outer claws) of 100 pairs of bovine hind limbs to investigate the etiological theory that an exacerbating factor for ulceration is exostosis of the tuberculum flexorium within the distal phalanx. A variety of morphological changes of the tuberculum flexorium of bovine hind limb claws was visualized by 3-dimensional CT, and the geometry of these claws suggested a growth pattern of bone development with respect to the assumed daily loading patterns. This growth occurs initially at the abaxial caudal aspect of the distal phalanx and is followed by horizontal progression toward the axial aspect. The length of downward bone development on the solar face of the distal phalanx was 2.73±1.32 mm in the outer claws, significantly greater than in the inner claws (2.38±0.96 mm). Ratios of downward (vertical) bone development to the thickness of the subcutis and the corium (VerBD ratios) did not differ between the outer and inner claws (36.7 vs. 38.3%, respectively). Ratios of horizontal bone development to the axial-to-abaxial line of the tuberculum flexorium (HorBD ratios) were approximately 60% for both outer and inner claws. These quantitative measures regarding horizontal and vertical bone development within the distal phalanx were positively correlated with age and VerBD ratios (r=0.53 and r=0.36 for the inner and outer claws, respectively). Correlations between claw width of the outer claw and length of vertical bone development (r=0.43), the HorBD ratio (r=0.51), and the VerBD ratio (r=0.42) suggested that the relative size difference between the inner and outer claws enhances bone development in the outer claw. Correlation coefficients between VerBD and HorBD ratios (r=0.52 and 0.63 for the inner and outer claws, respectively) suggested that horizontal and vertical bone development occurs as a synchronized process within the tuberculum flexorium. This age-related progress of bone development within the tuberculum flexorium is associated with increased exposure to several exacerbating factors and the laminitic process.

Effects of Surface-Deacetylated Chitin Nanofibers in an Experimental Model of Hypercholesterolemia

This study evaluated the effects of oral administration of surface-deacetylated chitin nanofibers (SDACNFs) on hypercholesterolemia using an experimental model. All rats were fed a high cholesterol diet with 1% w/w cholesterol and 0.5% w/w cholic acid for 28 days. Rats were divided equally into four groups: the control group was administered 0.05% acetic acid dissolved in tap water, and the SDACNF, chitosan (CS), and cellulose nanofiber (CLNF) groups were administered 0.1% CNF, CS, or CLNF dissolved in the tap water, respectively, during the experimental period. Changes in body weight, intake of food and water, and organ weight were measured. Serum blood chemistry and histopathological examination of the liver were performed. Administration of SDACNF did not affect body weight change, food and water intake, or organ weights. Administration of SDACNF and CS decreased the diet-induced increase in serum total cholesterol, chylomicron, very-low-density lipoprotein, and phospholipid levels on day 14. Moreover, oral administration of SDACNFs suppressed the increase of alanine transaminase levels on day 29 and suppressed vacuolar degeneration and accumulation of lipid droplets in liver tissue. These data indicate that SDACNF has potential as a functional food for patients with hypercholesterolemia.