Isoleucine, an Essential Amino Acid, Induces the Expression of Human <i>β</i>Defensin 2 through the Activation of the G-Protein Coupled Receptor-ERK Pathway in the Intestinal Epithelia

Anti-microbial peptides are essential for the intestinal innate immunity that protects the intestinal epithelia from attacks by foreign pathogens. Human β-defensin (HBD) is one of the pivotal anti-microbial peptides that are expressed in the colonic epithelia. This study investigated the effect and the signaling mechanism of inducible β-defensin HBD2 by an essential amino acid, isoleucine (Ile) in colonic epithelial cells. Here we examined the expression level of HBD2 on induction of Ile in epithelial cells, and checked this pathway. HBD2 mRNA was induced by co-incubation with IL-1α and Ile in Caco2 cells, but not by Ile alone. An inhibitor of either ERK or Gi, a subunit of G-proteins, reduced the induction of HBD2 mRNA by Ile. The treatment with Ile also increased the intracellular calcium ion concentration, thus suggesting that the GPCR and ERK signaling pathway mediate the effects of Ile. These results indicate that an essential amino acid, Ile, enhances the expression of an inducible β-defensin, namely HBD2, by IL-1α through the activation of GPCRs and ERK signaling pathway. The administration of Ile may therefore represent a possible option to safely treat intestinal inflammation.

Synthesis and cytotoxicity of 28-carboxymethoxy lupane triterpenoids.Preference of 28-O-acylation over 28-O-alkylation of betulm by ethyl bromoacetate

28-Carboxymethoxy lupane tritepenoids 3 and 4 were synthesized by alkylation of betulin with the THP protected 2- hydroxyethyl iodide followed by oxidation and reduction. Direct reaction of betulin （5） or betulone （10） with ethyl bromoacetate led to 28-O-acylation, instead of 28-O-alkylation. The targeted compounds 3 and 4 were not cytotoxic at the highest concentration tested （75 μmol/L）, suggesting that elongation of the chain length at the 28-position in both betulinic acid （1） and betulonic acid （2） was detrimental to the cytotoxicity. The acylation products 28-O-bromoacetates （Sa, 8b and 11） and 28-O-methoxyacetate 13 exhibited cytotoxicity against several cancer cell lines tested.

The Number of Cyclic Stretch Regulates Cellular Elasticity in C2C12 Myoblasts

Mechanical stimulations have been shown to regulate cellular mechanical properties. However, the stimulation patterns for effective regulation are as yet unclear. We investigated the effects of application of differing numbers of mechanical stimulation sets, each set consisting of 8% extension and compression to cells via deformation of cell culture elastic chamber, on cellular elasticity. Elasticity increased with only a single step-like stretch and with a single step-like stretch after 1 set of mechanical stimulation, whereas elasticity did not change with a single step-like stretch after 10 sets of mechanical stimulation. These results indicate that the increase in cellular elasticity with the single step-like stretch depends on the number of applied mechanical stimulations. Immunofluorescence staining showed that phosphorylation and dephosphorylation of myosin regulatory light chain (MRLC), which regulates intracellular contractile force and cellular elasticity, accompanied cellular elasticity changes. These findings suggest that cellular elasticity changes under cyclic and step-like stretches are mediated by MRLC.

Chlorella Residue Functions as a Bio-Stimulant to Promote Plant Growth and Improve Soil Fertility

Chlorella residues are currently underutilized. Therefore, in this study, we analyzed the nutritional components of Chlorella residue, and investigated its potential use as an organic fertilizer/bio-stimulant. Composition analyses revealed that the Chlorella residue contained a substantial amount of nitrogen (97,910 mg/kg), and significant quantities of secondary macronutrients, such as calcium (4300 mg/kg) and magnesium (9700 mg/kg), and micronutrients, such as iron (1850 mg/L) and manganese (359 mg/kg). The application of Chlorella residue to soil resulted in increased soil bacterial biomass. When Chlorella residue was added to the soil at a rate of 0.5% or 1.0% (w/w), the fresh weights of Brassica rapa and Spinacia oleracea were significantly increased. Furthermore, the application of Chlorella residue to the soil of B. rapa suppressed the reduction of the microbiome caused by clubroot disease and decreased the clubroot disease index. Therefore, Chlorella residue can be included in organic fertilizers that effectively improve soil nutrient contents, promote plant growth, and reduce the incidence of disease.

Self-assembled structures of a semi-rigid polyanion in aqueous solutions and hydrogels

Poly(2,2'-disulfonyl-4,4'-benzidine terephthalamide) (PBDT), a kind of liquid-crystalline (LC) molecule, has high molecular weight, negative charge and a semi-rigid structure. The aqueous solution of PBDT shows nematic liquid crystalline state above a critical PBDT concentration, CLC*, of 2 wt%-3wt%. Different from the flexible polyelectrolyte, PBDT shows a variety of self-assembling structures in aqueous solution with and without salt due to the semi-rigid nature and highly charged property. In addition, the hydrogels with ordered structure are developed by polymerizing a cationic monomer N-[3-(N,N-dimethylamino) propyl] acrylamide methyl chloride quarternary (DMAPAA-Q) in the presence of a small amount of PBDT below the CLC*. During the polymerization of cationic monomer, the polycations form a complex with semi-rigid PBDT through electrostatic interaction; these complexes self-assemble into ordered structures that are frozen in the hydrogel. Several different structures, including the anisotropic, dual network-like structure, and cylindrically symmetric structure, with various length scales from micrometer to millimeter, are observed. The hydrogels with ordered liquid crystalline assemblies and particular optical properties should promise applications in many fields, such as in bionics, tissue engineering, and mechano-optical sensors.

SYNTHETIC HYDROGELS AS SCAFFOLDS FOR MANIPULATING ENDOTHELIUM CELL BEHAVIORS

Synthetic hydrogels can be used as scaffolds that not only favor endothelial cells （ECs） proliferation but also manipulate the behaviors and functions of the ECs. In this review paper, the effect of chemical structure, Young＇s modulus （E） and zeta potential （ζ） of synthetic hydrogel scaffolds on static cell behaviors, including cell morphology, proliferation, cytoskeleton structure and focal adhesion, and on dynamic cell behaviors, including migration velocity and morphology oscillation, as well as on EC function such as anti-platelet adhesion, are reported. It was found that negatively charged hydrogels, poly（2-acrylamido-2-methylpropanesulfonic sodium） （PNaAMPS） and poly（sodium p-styrene sulphonate） （PNaSS）, can directly promote cell proliferation, with no need of surface modifcation by any cell-adhesive proteins or peptides at the environment of serum-containing medium. In addition, the Young＇s modulus （E） and zeta potential （ζ） of hydrogel scaffolds are quantitatively tuned by copolymer hydrogels, poly（NaAMPS-co-DMAAm） and poly（NaSS-co-DMAAm）, in which the two kinds of negatively charged monomers NaAMPS and NaSS are copolymerized with neutral monomer, N,N-dimethylacrylamide （DMAAm）. It was found that the critical zeta potential of hydrogels manipulating EC morphology, proliferation, and motility is ζcritical= -20.83 mV and ζcritical = -14.0 mV for poly（NaAMPS-co-DMAAm） and poly（NaSS-co-DMAAm）, respectively. The above mentioned EC behaviors well correlate with the adsorption of fibronectin, a kind of cell-adhesive protein, on the hydrogel surfaces. Furthermore, adhered platelets on the EC monolayers cultured on the hydrogel scaffolds obviously decreases with an increase of the Young＇s modulus （E） of the hydrogels, especially when E 〉 60 kPa. Glycocalyx assay and gene expression of ECs demonstrate that the anti-platelet adhesion well correlates with the EC-specific glycocalyx. The above investigation suggests that understanding the relationship between physic-chemical properties of synthetic hydrogels and cell responses is essential to design optimal soft and wet scaffolds for tissue engineering. Keywords： Synthetic hydrogel; Scaffold; Endothelial cell; Cell behavior

Facile tuning of hydrogel properties by manipulating cationic-aromatic monomer sequences

Copolymer hydrogels formed from cationic and aromatic monomers with identical monomer compositions but different average sequences were synthesized by free-radical copolymerization in various solvents.We found that hydrogels with one-componentrich segments are mechanically stronger than those with adjacent-rich monomer sequences in water,while hydrogels with a rich cation-πadjacent sequence showed excellent mechanical strength and underwater adhesion in saltwater(0.7 M Na Cl).The molecular mechanisms for these behaviors are discussed in terms of polymer structures.This work reveals the importance of monomeric sequences in determining hydrogel properties and provides a facile approach to develop hydrogels with different properties but the same monomer composition.

Aggregated structures and their functionalities in hydrogels

Biological soft tissues and hydrogels belong to the same category of soft and wet matter.Both of them are composed of polymer network and a certain amount of water,and permeable to small molecules.Biological tissues possess elaborated structures and exhibit outstanding functionalities.On the other hand,hydrogels are usually amorphous with poor functionality.In recent years,various hydrogels with robust functionalities have been developed by introducing aggregated structures into the gel networks,widely extend their applications in diverse fields,such as soft actuators,biological sensors,and structural biomaterials.Four strategies are usually used to fabricate aggregated structure into hydrogels,including molecular self-assembling,microphase separation,crystallization,and inorganic additives.Different aggregated structures entail the gel very different functionalities.A simple aggregated structure is able to bring multiple functionalities and a combination of mechanical performances of the hydrogels.In this review,we describe the strategies used to construct aggerated structure in hydrogels and discuss about the close relation between the aggregated structure and functionality.We also highlight the role of nonequilibrium aggregated structure in fabricating hydrogels with dynamic memorizing-forgetting behavior and point out the remaining challenges.