Grafting of 18ß-Glycyrrhetinic Acid and Sialic Acid onto Chitosan to Produce a New Amphipathic Chitosan Derivative: Synthesis, Characterization, and Cytotoxicity.

Chitosan is the only cationic polysaccharide found in nature. It has broad application prospects in biomaterials, but its application is limited due to its poor solubility in water. A novel chitosan derivative was synthesized by amidation of chitosan with 18ß-glycyrrhetinic acid and sialic acid. The chitosan derivatives were characterized by Fourier transform infrared spectroscopy, thermogravimetric analysis, and measurement of the zeta potential. We also investigated the solubility, cytotoxicity, and blood compatibility of chitosan derivatives. 18ß-glycyrrhetinic acid and sialic acid could be grafted onto chitosan molecular chains. The thermal stability of the synthesized chitosan derivatives was decreased and the surface was positively charged in water and phosphate-buffered saline. After chitosan had been modified by 18 ß-glycyrrhetinic acid and sialic acid, the solubility of chitosan was improved greatly in water and phosphate-buffered saline, and percent hemolysis was <5%. Novel amphiphilic chitosan derivatives could be suitable polymers for biomedical purposes.

An enzyme free fluorescence resonance transfer strategy based on hybrid chain reaction and triplex DNA for Vibrio parahaemolyticus.

In this work, an enzyme-free fluorescence resonance energy transfer (FRET) strategy was established for rapid and specific detection of the DNA sequence from Vibrio parahaemolyticus (VP) using hybridization chain reaction (HCR) amplification and triplex DNA. The triplex forming oligonucleotide (TFO) was labelled with carboxyfluorescein (FAM) as fluorescence donor, and hairpin sequence H1 was labelled by tetramethylrhodamine (TAMRA) as fluorescence receptor. In the present target VP DNA, the hairpin structure of molecular beacon (MB) was opened, the free end was released and hybridized with H1-TAMRA, and the HCR reaction was triggered by the alternate supplementation of H1-TAMRA and H2 to produce the notch double helix analogue. After the addition of TFO-FAM, a triplex structure was formed between HCR products (H1-TAMRA/H2) and TFO-FAM. A close contact between the donor and the receptor resulted in FRET. Under the optimal conditions, the fluorescence quenching value was inversely proportional to the concentration of target VP DNA in the range of 0.1-50 nmol L-1, and the detection limit was 35 pmol L-1.

Mussel-Inspired Catechol-Functionalized Hydrogels and Their Medical Applications.

Mussel adhesive proteins (MAPs) have a unique ability to firmly adhere to different surfaces in aqueous environments via the special amino acid, 3,4-dihydroxyphenylalanine (DOPA). The catechol groups in DOPA are a key group for adhesive proteins, which is highly informative for the biomedical domain. By simulating MAPs, medical products can be developed for tissue adhesion, drug delivery, and wound healing. Hydrogel is a common formulation that is highly adaptable to numerous medical applications. Based on a discussion of the adhesion mechanism of MAPs, this paper reviews the formation and adhesion mechanism of catechol-functionalized hydrogels, types of hydrogels and main factors affecting adhesion, and medical applications of hydrogels, and future the development of catechol-functionalized hydrogels.

The effect of melanin-free extract from Sepia esculenta ink on lipid peroxidation, protein oxidation and water-holding capacity of tilapia fillet during cold storage.

BACKGROUND: Preservative effect of melanin-free extract of Sepia esculenta ink (MFESI) on Sparus latus fillet has been verified in our previous work. This study aims to further approach the mechanism of MFESI for extending the shelf-life of fish fillet during cold storage. Tilapia fillets were treated with different dosage of MFESI (0, 15, 25 and 35 mg/ml) and packed with preservative film for succedent cold-storage at 4 °C for scheduled time. Contents of total volatile basic nitrogen and sulfydryl and carbanyl groups were measured for evaluating protein oxidation. Malondialdehyde contents were measured for estimating lipid peroxidation and loss of water was used to determine water-holding capacity of fillet. RESULTS: The data indicated that MFESI not only possessed certain degree of antioxidant capacity in vitro, also lengthened shelf-life of tilapia fillet in cold-storage condition. Apart from 15 mg/ml, both 25 and 35 mg/ml of MFESI obviously prevented lipid and protein from oxidation and reduced loss of water from tilapia fillets, and the latter was more effective than the former. CONCLUSION: MFESI can repress lipid peroxidation and protein oxidation and reduce water loss, maintain the tilapia fillets quality and, thus, it could be an effective and natural preservative for extending the shelf-life of tilapia fillets during cold storage.

Squid ink polysaccharide prevents chemotherapy induced injury in the testes of reproducing mice.

The present study was conducted to investigate the preventive effects of squid ink polysaccharides (SIP) on the damage of sperm and reproduction induced by cyclophosphamide that is most commonly used for treating clinically cancers. Male Kunming mice exposed to cyclophosphamide were administered with SIP and were sacrificed to determine sperm parameters, testicular antioxidant ability and reproductive capacity. Data indicated that cyclophosphamide caused obvious changes in mice such as significant reduction (P<0.01) of glutathione reductase activity (GR), vitamin C (Vc) content and total antioxidant capacity (T-AOC) in the testes, as well as elevation (P<0.01) of abnormal rates of sperm and fetus, and a decrease in the total fetal count and average fetal count (P<0.01), were totally alleviated by SIP. From these findings it can be concluded that SIP decreases chemotherapeutic damage to sperm and reproduction in mice induced by cyclophosphamide.

Inhibition of chemotherapy-induced apoptosis of testicular cells by squid ink polysaccharide.

The aim of this study was to determine the mechanisms driving the protective effects of squid ink polysaccharide (SIP) against cyclophosphamide (CP)-induced testicular damage, focusing on germ cells. In the testes of mice exposed to CP and/or SIP, the present study examined the levels of reactive oxygen species (ROS) and malondialdehyde, activity of superoxide dismutase levels, protein expression levels of B-cell lymphoma 2 (Bcl2), Bcl2-associated X protein (Bax), and total Caspase 3, activation of p-p38 and p-Akt proteins, and tissue morphology. The findings indicated that CP induced ROS production and oxidative stress, resulting in testicular damage. However, under administration of SIP, oxidative stress was impaired and the testicular toxicity induced by CP was weakened, which implied that SIP may have an important role in preventing chemotherapeutic damage to the male reproductive system via promoting antioxidant ability. Furthermore, the altered expression levels, including the upregulation of Bax and Caspase 3, downregulation of Bcl-2 and the increased Bax/Bcl-2 ratio, indicated that apoptosis occurred in CP exposed testes of mice; however, the alterations were reversed in mice treated with SIP. Moreover, in CP-exposed testes, p38 and Akt proteins were significantly phosphorylated (P<0.05), whereas in the testes of mice co-treated with SIP and CP, phosphorylation of the two proteins was inhibited, demonstrating that the two signalling pathways participated in the regulative processes of the deleterious effects caused by CP, and the preventive effects SIP mediated.

Inhibition of acrolein-induced autophagy and apoptosis by a glycosaminoglycan from Sepia esculenta ink in mouse Leydig cells.

In our recent reports, a squid ink polysaccharide (SIP) was found having preventive activity against cyclophosphamide induced damage in mouse testis and ovary. Here we further reveal the regulative mechanism of SIP against chemical toxicity on testis. Leydig cells exposed to acrolein (ACR) underwent apoptosis at 12h and 24h. Before apoptosis, cells occurred autophagy that was confirmed by high autophagic rate and Beclin-1 protein content at 3h. PI3K/Akt and p38 MAPK signal pathways involved in the regulatory mechanisms. These outcomes of ACR were recovered completely by SIP, which was demonstrated by attenuated disruption of redox equilibrium and increased testosterone production, through suppressing ACR-caused autophagy and apoptosis regulated by PI3K/Akt and p38 MAPK signal pathways in Leydig cells. Summarily, autophagy occurred before apoptosis caused by ACR-activated p38 MAPK and PI3K/Akt pathways were blocked by SIP, resulting in survival and functional maintenance of Leydig cells.

Squid ink polysaccharide prevents autophagy and oxidative stress affected by cyclophosphamide in Leydig cells of mice: a pilot study.

OBJECTIVES: The aim of this study was to explore the effects of Squid ink polysaccharide (SIP) on prevention of autophagy and oxidative stress induced by cyclophosphamide (CP) in Leydig cells of mice. MATERIALS AND METHODS: Examination of reproductive organ exponents, abnormal sperm rate, activities of superoxide dismutase (SOD), catalase (CAT), contents of malondialdehyde (MDA), and histological structure were performed to detect the optimal dose of SIP against oxidative stress damage in vivo, and autophagy-associated protein LC3 and Beclin-1 were examined by immunofluorescence, and their expression was detected by Western blot analysis. Leydig cells ultrastructural changes were observed by transmission fluorescent microscope. RESULTS: SIP significantly inhibited sperm aberration, histological structure and injury of seminiferous tubules caused by CP, as well as the antioxidant activity of SOD and CAT were increased; contents of MDA were decreased. The optimal dose of SIP for prevention of oxidative stress injury by CP was 80 mg/kg. In addition, LC3 and Beclin-1 fluorescent granules were much less in the Leydig cell layer after treatment via SIP compared with the CP-treated group, and the expression levels of LC3 and Beclin-1 were also decreased. Furthermore, characteristics of cell autophagy such as mitochondrial swelling, autophagic vacuoles, and chromatin pyknosis were observed in CP-treated Leydig cells, but SIP could effectively weaken injury of Leydig cell ultrastructure by CP. CONCLUSION: SIP, as an antioxidant, prevents the cytoskeleton damage through up-regulation antioxidant capacity and inhibition autophagy caused by CP.

Preventive Effects of a Novel Polysaccharide from Sepia esculenta Ink on Ovarian Failure and Its Action Mechanisms in Cyclophosphamide-Treated Mice.

On the basis of our findings about chemo-preventive roles of squid ink polysaccharide and the well-known toxicity of cyclophosphamide (CP) on female gonad, this research investigated the protective effects of a novel polysaccharide from Sepia esculenta ink (SEP) on the ovarian failure resulting from CP, as well as the action mechanisms underpinning this. The results indicated that CP destroyed the ovaries of mice which caused depletion of various follicles, and led to a reduction in estradiol content, increases in FSH and LH contents in sera, decreases in ovary and uterus masses and their relative mass ratios, disruption of the ultrastructure of granulosa cells, as well as induction of apoptosis and autophagy via p38 MAPK and PI3K/Akt signaling pathways. The phenomenon resulted in ovarian failure. However, SEP exposure altered the negative effects completely. The data indicated that SEP can effectively prevent ovarian failure CP caused in mice by inhibiting the p38 MAPK signaling pathway and activating the PI3K/Akt signaling pathway as regulated by CP. SEP was a novel polysaccharide from Sepia esculenta ink with a unique primary structure mainly composed of GalN and Ara that accounted for almost half of all monosaccharides: their ratio was nearly one-to-one. Besides, the polysaccharide contained a small number of Fuc and tiny amounts of Man, GlcN, GlcA, and GalA.

Polysaccharide from Sepia esculenta ink and cisplatin inhibit synergistically proliferation and metastasis of triple-negative breast cancer MDA-MB-231 cells.

OBJECTIVES: This paper aims to investigate synergistic inhibition of polysaccharide from Sepia esculenta ink (SIP), a newly isolated marine polysaccharide in our laboratory, on breast cancer MDA-MB-231 cells exposed to cisplatin. MATERIALS AND METHODS: Cell viability of MDA-MB-231 cells was determined by CCK 8 assay. Median-effect concentration was analyzed using Chou-Talalay method that was also subjected to determine cell inhibition ratio and combined index, as well as interaction between SIP and cisplatin. Proliferation and migration abilities were detected with plate colony formation assay and cell wound scratch assay, respectively. Expression of MMP-2 and MMP-9 proteins was measured with Western blot assay. RESULTS: Data showed that SIP not only suppressed proliferation and migration of MDA-MB-231 cells, and expression of MMP-2 and MMP-9 proteins, also promoted inhibition of cisplatin on proliferation, migration and MMPs expression of MDA-MB-231 cells, which indicates synergy inhibition of drug combination of SIP and cisplatin on breast cancer cells. The median-effect concentrations of cisplatin and SIP were 4.9 and 1659.6 µg/ml, respectively. Whereas the concentration of combination drug was 158.5 µg/ml. The data indicated that drug combination can decrease dosages of the two single agents, especially the usual dosage of cisplatin. CONCLUSION: This research demonstrated that SIP repressed proliferation and metastasis of MDA-MB-231 cells and promoted anticancer effect of cisplatin on the breast cancer cells. The data suggested that SIP is a potential natural drug that can be used as an auxiliary medicine alongside chemotherapy in treating breast cancer.

Protective effects of squid ink extract towards hemopoietic injuries induced by cyclophosphamine.

To investigate the protective effects of squid ink in chemotherapy, BALB/c mice were used as animal models of injuries induced by cyclophosphamine, a well known chemotherapeutic drug. The mice were randomly divided into five groups with the same number of males and females in each group. At the end of the experiment, animals were sacrificed to investigate organ indexes and antioxidant ability of the spleen, peripheral blood profile and quantities of bone marrow nucleated cells. Results showed that the hemopoietic function of mice was injured by cyclophosphamine, as indicated by decreases of contents of erythrocytes, leukocytes, hemoglobin and bone marrow nucleated cells (P<0.01), while platelets were not affected (P>0.05), as well as modification of organ indexes (P<0.05) and spleen antioxidant ability (P<0.05 or P<0.01), whereas sepia extract markedly increased the levels of erythrocytes, leukocytes, hemoglobin and bone marrow nucleated cells (P<0.01), but not platelets (P>0.05), and reversed the effects of cyclophosphamine on organ indexes and antioxidant ability of spleen (P<0.01 or P<0.05). In addition, squid ink extract did not change marrow hemopoiesis but improved the antioxidant ability of spleen in the animals. The data suggest that squid ink extract can protect the hemopoietic system from chemotherapeutic injury and could be employed to develop cell-protective drugs for use in clinical treatment of tumours.

Expression of basic fibroblast growth factor results in the decrease of myostatin mRNA in murine C2C12 myoblasts.

During the development and regeneration of skeletal muscle, many growth factors, such as basic fibroblast growth factor (bFGF, FGF-2) and myostatin, have been shown to play regulating roles. bFGF contributes to promote proliferation and to inhibit differentiation of skeletal muscle, whereas myostatin plays a series of contrasting roles. In order to elucidate whether the expression of bFGF has any relationship with the expression of myostatin in skeletal muscle cells, we constructed a eukaryotic expression vector for the expression of exogenous bFGF in murine C2C12 myoblasts. Quantitative RT-PCR assays indicated that with the increase of the expression of exogenous bFGF gene, the expression of endogenous myostatin gene was suppressed at mRNA level and protein level.