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Abstract Helicobacter pylori (H. pylori) causes infection in the stomach and is a major factor for gastric carcinogenesis. The application of antimicrobial peptides (AMPs) as an alternative treatment to traditional antibiotics is limited by their facile degradation in the stomach, their poor penetration of the gastric mucosa, and the cost of peptide production. Here, the design and characterization of a genetically encoded H. pylori‐responsive microbicidal protein crystal Cry3Aa‐MIIA‐AMP‐P17 is described. This designed crystal exhibits preferential binding to H. pylori, and when activated, promotes the targeted release of the AMP at the H. pylori infection site. Significantly, when the activated Cry3Aa‐MIIA‐AMP‐P17 crystals are orally delivered to infected mice, the Cry3Aa crystal framework protects its cargo AMP against degradation, resulting in enhanced in vivo efficacy against H. pylori infection. Notably, in contrast to antibiotics, treatment with the activated crystals results in minimal perturbation of the mouse gut microbiota. These results demonstrate that engineered Cry3Aa crystals can serve as an effective platform for the oral delivery of therapeutic peptides to treat gastrointestinal diseases.

IntroductionLead is a ubiquitous environmental and industrial pollutant. Its nonbiodegradable toxicity induces a plethora of human diseases. A novel bioactive glycoprotein containing 1.15% carbohydrate, with the ability of adsorbing lead and effecting detoxification, has been purified from Auricularia polytricha and designated as APL. Besides, its mechanisms related to regulation of hepatic metabolic derangements at the proteome level were analyzed in this study.MethodsChromatographic techniques were utilized to purify APL in the current study. For investigating the protective effects of APL, Sprague-Dawley rats were given daily intraperitoneal injections of lead acetate for establishment of an animal model, and different dosages of APL were gastrically irrigated for study of protection from lead detoxification. Liver samples were prepared for proteomic analyses to explore the detoxification mechanisms.Results and discussionThe detoxifying glycoprotein APL displayed unique molecular properties with molecular weight of 252-kDa, was isolated from fruiting bodies of the edible fungus A. polytricha. The serum concentrations of lead and the liver function biomarkers aspartate and alanine aminotransferases were significantly (p

Abstract Background Previous studies have shown a major green tea polyphenol (−)-epigallocatechin-3-gallate ((−)-EGCG) as a powerful anti-cancer agent. However, its poor bioavailability and requirement of a high dosage to manifest activity have restricted its clinical application. Recently, our team synthesized a peracetate-protected derivative of EGCG, which can act as a prodrug of (−)-EGCG (ProEGCG) with enhanced stability and improved bioavailability in vitro and in vivo. Herein, we tested the therapeutic efficacy of this novel ProEGCG, in comparison to EGCG, toward human endometrial cancer (EC). Methods In this study, the effects of ProEGCG and EGCG treatments on cell growth, cell survival and modulation of intracellular signaling pathways in RL95–2 and AN3 CA EC cells were compared. The antiproliferative effect was evaluated by cell viability assay. Apoptosis was measured by annexin/propidium iodide staining. Expression of mitogen-activated protein kinases, markers of proliferation and apoptosis were measured by immunoblot analysis. In addition, the effects of ProEGCG and EGCG on tumor growth, vessel formation and gene expression profiles on xenograft models of the EC cells were investigated. Results We found that treatment with ProEGCG, but not EGCG, inhibited, in a time- and dose-dependent manner, the proliferation and increased apoptosis of EC cells. Treatment with low-dose ProEGCG significantly enhanced phosphorylation of JNK and p38 MAPK and inhibited phosphorylation of Akt and ERK which are critical mediators of apoptosis. ProEGCG, but not EGCG, elicited a significant decrease in the growth of the EC xenografts, promoted apoptotic activity of tumour cells in the EC xenografts, and decreased microvessel formation, by differentially suppressing anti-apoptotic molecules, NOD1 and NAIP. Notably, no obvious adverse effects were detected. Conclusions Taken together, ProEGCG at a low dose exhibited anticancer activity in EC cells through its anti-proliferative, pro-apoptotic and anti-tumor actions on endometrial cancer in vitro and in vivo. In contrast, a low dose of EGCG did not bring about similar effects. Importantly, our data demonstrated the efficacy and safety of ProEGCG which manifests the potential of a novel anticancer agent for the management of endometrial cancer.

Lectins are a unique group of nonimmune carbohydrate-binding proteins or glycoproteins that exhibit specific and reversible carbohydrate-binding activity in a non-catalytic manner. Lectins have diverse sources and are classified according to their origins, such as plant lectins, animal lectins, and fish lectins. Marine organisms including fish, crustaceans, and mollusks produce a myriad of lectins, including rhamnose binding lectins (RBL), fucose-binding lectins (FTL), mannose-binding lectin, galectins, galactose binding lectins, and C-type lectins. The widely used method of extracting lectins from marine samples is a simple two-step process employing a polar salt solution and purification by column chromatography. Lectins exert several immunomodulatory functions, including pathogen recognition, inflammatory reactions, participating in various hemocyte functions (e.g., agglutination), phagocytic reactions, among others. Lectins can also control cell proliferation, protein folding, RNA splicing, and trafficking of molecules. Due to their reported biological and pharmaceutical activities, lectins have attracted the attention of scientists and industries (i.e., food, biomedical, and pharmaceutical industries). Therefore, this review aims to update current information on lectins from marine organisms, their characterization, extraction, and biofunctionalities.

Chitosan is a natural polycationic linear polysaccharide derived from chitin. The low solubility of chitosan in neutral and alkaline solution limits its application. Nevertheless, chemical modification into composites or hydrogels brings to it new functional properties for different applications. Chitosans are recognized as versatile biomaterials because of their non-toxicity, low allergenicity, biocompatibility and biodegradability. This review presents the recent research, trends and prospects in chitosan. Some special pharmaceutical and biomedical applications are also highlighted.

Peptides are important bioactive natural products which are present in many marine species. These marine peptides have high potential nutraceutical and medicinal values because of their broad spectra of bioactivities. Their antimicrobial, antiviral, antitumor, antioxidative, cardioprotective (antihypertensive, antiatherosclerotic and anticoagulant), immunomodulatory, analgesic, anxiolytic anti-diabetic, appetite suppressing and neuroprotective activities have attracted the attention of the pharmaceutical industry, which attempts to design them for use in the treatment or prevention of various diseases. Some marine peptides or their derivatives have high commercial values and had reached the pharmaceutical and nutraceutical markets. A large number of them are already in different phases of the clinical and preclinical pipeline. This review highlights the recent research in marine peptides and the trends and prospects for the future, with special emphasis on nutraceutical and pharmaceutical development into marketed products.

Abnormalities in the melatonin signaling pathway and the involvement of melatonin receptor MT2 have been reported in patients with adolescent idiopathic scoliosis (AIS). Whether these abnormalities were involved in the systemic abnormal skeletal growth in AIS during the peripubertal period remain unknown. In this cross-sectional case-control study, growth plate chondrocytes (GPCs) were cultured from twenty AIS and ten normal control subjects. Although the MT2 receptor was identified in GPCs from both AIS and controls, its mRNA expression was significantly lower in AIS patients than the controls. GPCs were cultured in the presence of either the vehicle or various concentrations of melatonin, with or without the selective MT2 melatonin receptor antagonist 4-P-PDOT (10 µM). Then the cell viability and the mRNA expression of collagen type X (COLX) and alkaline phosphatase (ALP) were assessed by MTT and qPCR, respectively. In the control GPCs, melatonin at the concentrations of 1, 100 nM and 10 µM significantly reduced the population of viable cells, and the mRNA level of COLX and ALP compared to the vehicle. Similar changes were not observed in the presence of 4-P-PDOT. Further, neither proliferation nor differentiation of GPCs from AIS patients was affected by the melatonin treatment. These findings support the presence of a functional abnormality of the melatonin signaling pathway in AIS GPCs, which might be associated with the abnormal endochondral ossification in AIS patients.

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A dimeric 64-kDa glucosamine-specific lectin was purified from seeds of Phaseolus vulgaris cv. "brown kidney bean." The simple 2-step purification protocol involved affinity chromatography on Affi-gel blue gel and gel filtration by FPLC on Superdex 75. The lectin was absorbed on Affi-gel blue gel and desorbed using 1M NaCl in the starting buffer. Gel filtration on Superdex 75 yielded a major absorbance peak that gave a single 32-kDa band in SDS-PAGE. Hemagglutinating activity was completely preserved when the ambient temperature was in the range of 20 °C-60 °C. However, drastic reduction of the activity occurred at temperatures above 65 °C. Full hemagglutinating activity of the lectin was observed at an ambient pH of 3 to 12. About 50% activity remained at pH 0-2, and only residual activity was observed at pH 13-14. Hemagglutinating activity of the lectin was inhibited by glucosamine. The brown kidney bean lectin elicited maximum mitogenic activity toward murine splenocytes at 2.5 µM. The mitogenic activity was nearly completely eliminated in the presence of 250 mM glucosamine. The lectin also increased mRNA expression of the cytokines IL-2, TNF-α and IFN-γ. The lectin exhibited antiproliferative activity toward human breast cancer (MCF7) cells, hepatoma (HepG2) cells and nasopharyngeal carcinoma (CNE1 and CNE2) cells with IC(50) of 5.12 µM, 32.85 µM, 3.12 µM and 40.12 µM respectively after treatment for 24 hours. Flow cytometry with Annexin V and propidum iodide staining indicated apoptosis of MCF7 cells. Hoechst 33342 staining also indicated formation of apoptotic bodies in MCF7 cells after exposure to brown kidney bean lectin. Western blotting revealed that the lectin-induced apoptosis involved ER stress and unfolded protein response.

Breast cancer ranks as a common and severe neoplasia in women with increasing incidence as well as high risk of metastasis and relapse. Translational and laboratory-based clinical investigations of new/novel drugs are in progress. Medicinal plants are rich sources of biologically active natural products for drug development. The 27-kDa trichosanthin (TCS) is a ribosome inactivating protein purified from tubers of the Chinese herbal plant Trichosanthes kirilowii Maximowicz (common name Tian Hua Fen). In this study, we extended the potential medicinal applications of TCS from HIV, ferticide, hydatidiform moles, invasive moles, to breast cancer. We found that TCS manifested anti-proliferative and apoptosis-inducing activities in both estrogen-dependent human MCF-7 cells and estrogen-independent MDA-MB-231 cells. Flow cytometric analysis disclosed that TCS induced cell cycle arrest. Further studies revealed that TCS-induced tumor cell apoptosis was attributed to activation of both caspase-8 and caspase-9 regulated pathways. The subsequent events including caspase-3 activation, and increased PARP cleavage. With regard to cell morphology, stereotypical apoptotic features were observed. Moreover, in comparison with control, TCS- treated nude mice bearing MDA-MB-231 xenograft tumors exhibited significantly reduced tumor volume and tumor weight, due to the potent effect of TCS on tumor cell apoptosis as determined by the increase of caspase-3 activation, PARP cleavage, and DNA fragmentation using immunohistochemistry. Considering the clinical efficacy and relative safety of TCS on other human diseases, this work opens up new therapeutic avenues for patients with estrogen-dependent and/or estrogen-independent breast cancers.

Background: The fresh leaves of Eucalyptus globulus Labill. (Lan An) have been used in Chinese medicine for many years to treat dermatomycosis. Macrocarpal C was isolated from this herb and identiied as its major antifungal com- ponent by bioassay-guided puriication. This study aims to investigate the antifungal activity of macrocarpal C against Trichophyton mentagrophytes, which can cause tinea pedis. Methods: Fresh leaves of E. globulus were extracted with 95 % ethanol, and the resulting ethanolic extracts were dried before being partitioned with n-hexane. The n-hexane layer was then subjected to chromatographic puriication to give macrocarpal C. The antifungal minimum inhibitory concentration (MIC) of macrocarpal C was determined using the standard M38-A2 method described by the Clinical Laboratory Standards Institute (CLSI). The mode of action of macrocarpal C was elucidated using three in vitro assays, including (1) a fungal membrane permeability test using SYTOX® Green; (2) a reactive oxygen species (ROS) production test using 5-(and-6)-carboxy-2',7'-dihydrodichloroluorescein diacetate as a cell-permeable luorogenic probe; and (3) a DNA fragmentation test based on terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) detection. Terbinaine hydrochloride and nystatin were used as positive controls. Results: The suppression in the growth of T. mentagrophytes following its treatment with macrocarpal C was associated with an increase in the permeability of the fungal membrane (P = 0.0043 when compared to control); an increase in the production of intracellular ROS (P = 0.0063); and the induction of apoptosis as a consequence of DNA fragmentation (P = 0.0007). Conclusion: This study demonstrated that the antifungal action of macrocarpal C was associated with increases of membrane permeability, intracellular ROS and DNA fragmentation. [ABSTRACT FROM AUTHOR]

Pickles are popular in China and exhibits health-promoting effects. However, nitrite produced during fermentation adversely affects health due to formation of methemoglobin and conversion to carcinogenic nitrosamine. Fruiting bodies of the mushroom Boletus edulis were capable of inhibiting nitrite production during pickle fermentation. A 90-kDa nitrite reductase (NiR), demonstrating peptide sequence homology to fungal nitrite reductase, was isolated from B. edulis fruiting bodies. The optimum temperature and pH of the enzyme was 45 °C and 6.8, respectively. B. edulis NiR was capable of prolonging the lifespan of nitrite-intoxicated mice, indicating that it had the action of an antidote. The enzyme could also eliminate nitrite from blood after intragastric administration of sodium nitrite, and after packaging into capsule, this nitrite-eliminating activity could persist for at least 120 minutes thus avoiding immediate gastric degradation. B. edulis NiR represents the first nitrite reductase purified from mushrooms and may facilitate subsequent applications. [ABSTRACT FROM AUTHOR]

Chitosan is a natural polycationic linear polysaccharide derived from chitin. The low solubility of chitosan in neutral and alkaline solution limits its application. Nevertheless, chemical modification into composites or hydrogels brings to it new functional properties for different applications. Chitosans are recognized as versatile biomaterials because of their non-toxicity, low allergenicity, biocompatibility and biodegradability. This review presents the recent research, trends and prospects in chitosan. Some special pharmaceutical and biomedical applications are also highlighted. [ABSTRACT FROM AUTHOR]

A 5443 Da peptide with sequence homology to defensins was purified from purple pole beans (Phaseolus vulgaris cv. 'Extra-long Purple Pole bean'). This peptide was isolated by adsorption on an affinity chromatographic medium Affi- Gel Blue gel and ion-exchange chromatographic media SP-Sepharose (sulfopropyl-Sepharose) and Mono S and by gel filtration on Superdex peptide. The peptide inhibited mycelial growth in Mycosphaerella arachidicola, Helminthosporium maydis, Fusarium oxysporum, Verticillium dahliae, Rhizoctonia solani, Candida albicans and Setosphaeria turcica with an IC50 of 0.8, 0.9, 2.3, 3.2, 4.3, 4.8 and 9.8 μM respectively. Its antifungal potency was higher than that of the plant defensin coccinin (IC50 >50 μM). It induced membrane permeabilization in C. albicans as evidenced by SYTOX Green uptake, but did not affect erythrocyte membrane permeability. It inhibited growth in M. arachidicola by inducing chitin accumulation at hyphal tips as was shown by Congo Red staining. The antifungal activity was pH stable and thermostable. The peptide inhibited the proliferation of hepatoma (HepG2), breast cancer (MCF7), colon cancer (HT29) and cervical cancer (SiHa) cells but not that of human embryonic liver (WRL68) cells. Its anti- HepG2 activity (IC50 =4.1±0.8 μM, n=3) was higher than that of another plant defensin, gymnin (IC50 >50 μM). Its anti-MCF7 activity (IC50 =8.3±0.3 μM, n=3) was similar to that of other plant defensins. It reduced the activity of HIV-1 reverse transcriptase with an IC50 of 0.5±0.1 μM, n=3, much more potently than other plant defensins (IC50 >40 μM). There is the possibility of using the purple pole bean defensin for producing antifungal drugs and/or transgenic plants with fungal resistance. [ABSTRACT FROM AUTHOR]

A lectin has been isolated from seeds of the Phaseolus vulgaris cv. "Anasazi beans" using a procedure that involved affinity chromatography on Affi-gel blue gel, fast protein liquid chromatography (FPLC)-ion exchange chromatography on Mono S, and FPLC-gel filtration on Superdex 200. The lectin was comprised of two 30-kDa subunits with substantial N-terminal sequence similarity to other Phaseolus lectins. The hemagglutinating activity of the lectin was stable within the pH range of 1-14 and the temperature range of 0-80°C. The lectin potently suppressed proliferation of MCF-7 (breast cancer) cells with an IC50 of 1.3 μM, and inhibited the activity of HIV-1 reverse transcriptase with an IC50 of 7.6 μM. The lectin evoked a mitogenic response from murine splenocytes as evidenced by an increase in [3H-methyl]-thymidine incorporation. The lectin had no antifungal activity. It did not stimulate nitric oxide production by murine peritoneal macrophages. Chemical modification results indicated that tryptophan was crucial for the hemagglutinating activity of the lectin. [ABSTRACT FROM AUTHOR]