Optical control of cell differentiation on synthetic collagen-like scaffolds.

We have developed biocompatible scaffolds that enable cell fate control with visible light. The scaffolds are based on synthetic collagen-like polypeptide, poly(prolyl-hydroxyprolyl-glycyl) {poly(Pro-Hyp-Gly)} which has been used for cosmetics and other healthcare applications. Bioactive peptides were conjugated to the scaffolds via photoactivation reaction utilizing 488 nm visible light.

Calcium deposition in photocrosslinked poly(Pro-Hyp-Gly) hydrogels encapsulated rat bone marrow stromal cells.

Reproducing the features of the extracellular matrix is important for fabricating three-dimensional (3D) scaffolds for tissue regeneration. A collagen-like polypeptide, poly(Pro-Hyp-Gly), is a promising material for 3D scaffolds because of its excellent physical properties, biocompatibility, and biodegradability. In this paper, we present a novel photocrosslinked poly(Pro-Hyp-Gly) hydrogel as a 3D scaffold for simultaneous rat bone marrow stromal cell (rBMSC) encapsulation.

Cytocompatible polyion complex gel of poly(Pro-Hyp-Gly) for simultaneous rat bone marrow stromal cell encapsulation.

Polyion complex (PIC) gel of poly(Pro-Hyp-Gly) was successfully fabricated by simply mixing polyanion and polycation derivatives of poly(Pro-Hyp-Gly), a collagen-like polypeptide. The polyanion, succinylated poly(Pro-Hyp-Gly), and the polycation, arginylated poly(Pro-Hyp-Gly), contain carboxy (pK = 5.2) and guanidinium (pK = 12.

Nontubulation Repair of Peripheral Nerve Gap Using Heparin/Alginate Gel Combined with b-FGF.

All artificial nerve grafts have a tubular structure, and they guide axonal regrowth through the tube from the proximal side toward the peripheral side. Based on the results of our experimental study using animals, we used alginate gel without a tubular structure as an artificial nerve graft for digital nerve repair and evaluated peripheral nerve regeneration. In 2 patients, a gap due to digital nerve injury was bridged with controlled-release heparin/alginate gel combined with basic fibroblast growth factor, and restoration of the sensory function was serially evaluated.

Synthesis, Photophysical Properties, and Biological Evaluation of trans-Bisthioglycosylated Tetrakis(fluorophenyl)chlorin for Photodynamic Therapy.

trans-Bisthioglycosylated tetrakis(fluorophenyl)chlorin (7) was designed as a powerful photodynamic therapy (PDT) photosensitizer based on the findings of our systematic studies. We show here that the trans-bisthioglycosylated structure of 7 enhanced its uptake by HeLa cells and that the chlorin ring of 7 increased the efficiency of reactive oxygen species generation under the standard condition of our photocytotoxicity test. The versatility of 7 in PDT treatment was established using weakly metastatic B16F1 melanoma cells, metastatic 4T1 breast cancer cells, the RGK-1 gastric carcinoma mucosal cell line, and three human glioblastoma cell lines (U87, U251, and T98G).

Encapsulation of rat bone marrow stromal cells using a poly-ion complex gel of chitosan and succinylated poly(Pro-Hyp-Gly).

Encapsulation of stem cells into a three-dimensional (3D) scaffold is necessary to achieve tissue regeneration. Prefabricated 3D scaffolds, such as fibres or porous sponges, have limitations regarding homogeneous cell distribution. Hydrogels that can encapsulate cells such as animal-derived collagen gels need adjustment of the pH and/or temperature upon cell mixing.

Utilization of star-shaped polymer architecture in the creation of high-density polymer brush coatings for the prevention of platelet and bacteria adhesion.

We demonstrate utilization of star-shaped polymers as high-density polymer brush coatings and their effectiveness to inhibit the adhesion of platelets and bacteria. Star polymers consisting of poly(2-hydroxyethyl methacrylate) (PHEMA) and/or poly(methyl methacrylate) (PMMA), were synthesized using living radical polymerization with a ruthenium catalyst. The polymer coatings were prepared by simple drop casting of the polymer solution onto poly(ethylene terephthalate) (PET) surfaces and then dried.

Selective accumulation of [⁶²Zn]-labeled glycoconjugated porphyrins as multi-functional positron emission tomography tracers in cancer cells.

Positron-emission tomography (PET) can be used to visualize active stage cancer. Fluorine-18 ([(18)F])-labeled 2-([(18)F])2-deoxy-2-fluoroglucose (([(18)F])-FDG), which accumulates in glucose-dependent tissues, is a good cancer-targeting tracer. However, ([(18)F])-FDG is obscured in glucose-dependent normal tissues.

Sugar and heavy atom effects of glycoconjugated chlorin palladium complex on photocytotoxicity.

Palladium(II) complexes of glycoconjugated porphyrin and pyrrolidine-fused chlorin were prepared to examine sugar and heavy atom effects on in vitro photocytotoxicity. Cellular uptake into HeLa cells was enhanced by introducing sugar units regardless of other features, such as the central ion (free base or palladium(II) ion) and the ring structure (porphyrin or chlorin). The palladium(II) complex of glycoconjugated pyrrolidine-fused chlorin (PdPC2) exerted an excellent degree of photocytotoxicity not only on HeLa cells, but also on metastatic B16-BL6 cells, weakly metastatic B16F1 cells, and metastatic 4T1 cells.

Collagen-like polypeptide poly(Pro-Hyp-Gly) conjugated with Gly-Arg-Gly-Asp-Ser and Pro-His-Ser-Arg-Asn peptides enchances cell adhesion, migration, and stratification.

Collagens are widely used in medical applications, including as a scaffold for tissue regeneration. However, animal-derived collagens have several drawbacks, such as low thermal stability, nonspecific cell adhesion, antigenicity, and contamination with pathogenic substances. To overcome these problems, we chemically synthesized the collagen-like polypeptide, poly(prolyl-hydroxyprolyl-glycyl) (poly(Pro-Hyp-Gly)), which forms a collagen-like triple-helical structure and shows biodegradability and biocompatibility.

Delivery of small interfering RNA with a synthetic collagen poly(Pro-Hyp-Gly) for gene silencing in vitro and in vivo.

Silencing gene expression by small interfering RNAs (siRNAs) has become a powerful tool for the genetic analysis of many animals. However, the rapid degradation of siRNA and the limited duration of its action in vivo have called for an efficient delivery technology. Here, we describe that siRNA complexed with a synthetic collagen poly(Pro-Hyp-Gly) (SYCOL) is resistant to nucleases and is efficiently transferred into cells in vitro and in vivo, thereby allowing long-term gene silencing in vivo.

Anticoagulant activity of enzymatically synthesized amylose derivatives containing carboxy or sulfonate groups.

Heparin is an extracellular matrix polysaccharide. It is widely employed as an anticoagulant and can be used to form an anticoagulant surface on various medical devices such as renal dialysis devices to prevent thrombosis. However, heparin may cause hemorrhage and thrombocytopenia.

Synthesis, photophysical properties and photocytotoxicity of mono-, di-, tri- and tetra-glucosylated fluorophenylporphyrins.

In order to explore the effect of substitution patterns on the photocytotoxicity of glycoconjugated porphyrins, we synthesized and characterized a 'complete set' of tetrakis(perfluorophenyl)porphyrins having beta-d-glucopyranosylthio groups on the phenyl ring. Among five possible derivatives, trans-substituted S-glucosylated porphyrin trans-2(OH) exerted outstanding photocytotoxicity (EC(50) value was < 5 nM) in HeLa cells. The excellent photocytotoxicity of trans-2(OH) was found to be attributable to several factors, namely high optical transition probability in aqueous media, efficient type I photoreactions and enhanced cellular uptake.

The aromatic amino acid residues of tumor necrosis factor receptor-1-derived peptide are important for promoting differentiation of neural stem cells.

Neural stem cells have the self-renewal capacity and the ability to differentiate into all types of nerve cells. We previously reported that the tumor necrosis factor receptor-1-derived peptide promotes neural differentiation of fetal rat hippocampal neural stem cells. The tumor necrosis factor receptor-1-derived peptide contains six aromatic amino acid residues among its 14 amino acid residues.

Synthesis, photophysical properties and sugar-dependent in vitro photocytotoxicity of pyrrolidine-fused chlorins bearing S-glycosides.

Eight S-glycosylated 5,10,15,20-tetrakis(tetrafluorophenyl)porphyrins (1a', 1b', 1a and 1b (a: S-glucosylated, b: S-galactosylated)) and their 1,3-dipolar cycloadducts, i.e. chlorins 2a', 2b', 2a and 2b were prepared by nucleophilic substitution of the pentafluorophenyl groups with S-glycoside.

Effects of surface carboxylic acid groups of cerasomes, morphologically stable vesicles having a silica surface, on biomimetic deposition of hydroxyapatite in body fluid conditions.

Biomimetic mineralization of supramolecular scaffolds consisting of biomolecules or their analogues has received much attention recently from the viewpoint of creation of novel biomaterials. This study investigated biomimetic deposition of hydroxyapatite (HAp) on cerasomes, morphologically stable organic-inorganic hybrid vesicles. Scanning electron microscopy, energy-dispersive X-ray spectroscopy, and X-ray diffraction studies revealed that the pristine cerasomes induced heterogeneous nucleation of HAp when they were immersed in 1.

Synthesis and photocytotoxicity of S-glucosylated 5,10,15,20-Tetrakis(tetrafluorophenyl)porphyrin metal complexes as efficient (1)O(2)-generating glycoconjugates.

5,10,15,20-Tetrakis(4-(2,3,4,6-tetra-O-acetyl-beta-d-glucopyranosylthio)-2,3,5,6-tetrafluorophenyl)porphyrin 2a and its Zn(II), Pd(II), and Pt(II) complexes 2b, 2c, and 2d were prepared in excellent yields by nucleophilic substitution of the corresponding free-base porphyrin and metalloporphyrins with acetyl 2,3,4,6-tetra-O-acetyl-1-thio-beta-d-glucopyranoside. Deprotection of 2a, 2b, 2c, and 2d by alkaline hydrolysis afforded the corresponding S-glucosylated porphyrin 3a and its metal complexes 3b, 3c, and 3d. The structures and purity of all new photosensitizers were confirmed by elemental analysis and (1)H, (13)C, and (19)F NMR, UV-vis, and steady-state luminescence spectroscopy.

In vitro heavy-atom effect of palladium(II) and platinum(II) complexes of pyrrolidine-fused chlorin in photodynamic therapy.

Introduction of a heavy atom into photosensitizers generally facilitates intersystem crossing and improves the quantum yield (Phi(Delta)) of singlet oxygen ((1)O(2)), which is a key species in photodynamic therapy (PDT). However, little information is available about the physiological importance of this heavy-atom effect. The aim of this study is to examine the heavy-atom effect in simple metallochlorins in vitro at the cellular level.

Structure-photodynamic effect relationships of 24 glycoconjugated photosensitizers in HeLa cells.

The photodynamic effect of the glycoconjugated photosensitizer library containing 16 glycoconjugated 5,10,15,20-tetraphenylporphyrins and 8 glycoconjugated 5,10,15,20-tetraphenylchlorins were examined in HeLa cells, and analyzed by two approaches, namely, physiological properties (cellular uptake and reactive oxygen species (ROS)) and structural features of glycoconjugated photosensitizers. All glycoconjugated photosensitizers showed no cytotoxicity in the dark at a concentration of 5 muM. The photocytotoxicity profiles poorly related to the amount of cellular uptake of the photosensitizers.

A synthetic peptide corresponding to residues 301-320 of human Wnt-1 promotes PC12 cell adhesion and hippocampal neural stem cell differentiation.

Wnt signaling cascades play a crucial role in the maintenance of stem cell niches in many tissues as well as in embryonic patterning and cell-fate determination. Wnt signaling pathways have been well studied; however, the precise binding mechanism of Wnt protein to its receptor has not yet been clarified. Here we show the design and synthesis of seven novel peptide candidates for a receptor-binding site of human Wnt-1 based on its hydrophilicity and beta-turn profiles.

The biodegradability of poly(Pro-Hyp-Gly) synthetic polypeptide and the promotion of a dermal wound epithelialization using a poly(Pro-Hyp-Gly) sponge.

Collagens are widely used in medical applications, but animal-derived collagens have several drawbacks, such as low thermal stability, nonspecific cell attachment, and susceptibility to contamination by infectious pathogens, such as prions, which may transfect humans. We have previously reported the chemical synthesis of polypeptides consisting of a Pro-Hyp-Gly sequence and the high thermostability of their triple-helical structure. To clarify the biomaterial characteristics of the poly(Pro-Hyp-Gly) polypeptide, we assessed its biodegradability and its capability for skin regeneration.

Biomimetic deposition of hydroxyapatite on a synthetic polypeptide with beta sheet structure in a solution mimicking body fluid.

Deposition of a hydroxyapatite layer with similar structure to bone mineral is an attractive approach to the fabrication of bioactive coating layers to achieve direct bonding to living bone. To get successful coating of a hydroxyapatite layer on an organic polymer using a biomimetic solution, it is essential to find organic substrates that can effectively induce heterogeneous nucleation of hydroxyapatite after exposure to the body environment. Our previous study showed that sericin, a type of silk protein, has the ability to induce hydroxyapatite nucleation in a biomimetic solution when the sericin has a beta sheet structure.

Sugar-dependent photodynamic effect of glycoconjugated porphyrins: a study on photocytotoxicity, photophysical properties and binding behavior to bovine serum albumin (BSA).

The photocytotoxicity of four glycoconjugated porphyrins, namely 5,10,15,20-tetrakis[4-(beta-D-glucopyranosyloxy)phenyl]porphyrin (p-1a), 5,10,15,20-tetrakis[4-(beta-D-galactopyranosyloxy)phenyl]porphyrin (p-1b), 5,10,15,20-tetrakis[4-(beta-D-xylopyranosyloxy)phenyl]porphyrin (p-1c) and 5,10,15,20-tetrakis[4-(beta-D-arabinopyranosyloxy)phenyl]porphyrin (p-1d), was evaluated in HeLa cells in the concentration range from 1 to 7 microM using a light dose of 16 J x cm(-2) with a wavelength greater than 500 nm. The photocytotoxicity depends on the sugar moieties, and increases in the order of p-1d View Article and Find Full Text PDF

In vitro apatite formation on polyamide containing carboxyl groups modified with silanol groups.

Modification of organic polymer with silanol groups in combination with calcium salts enables the polymer to show bioactivity, that is, the polymer forms apatite on its surface after exposure to body environment. However, how modification with silanol groups influences ability of apatite formation on the polymer substrate and adhesive strength between polymer and apatite is not yet known. In the present study, polyamide containing carboxyl groups was modified with different amounts of silanol groups, and its apatite-forming ability in 1.

A novel role of L-serine (L-Ser) for the expression of nuclear factor of activated T cells (NFAT)2 in receptor activator of nuclear factor kappa B ligand (RANKL)-induced osteoclastogenesis in vitro.

Multinucleated cell formation is crucial for osteoclastogenesis, and the expression of nuclear factor of activated T cells (NFAT)2 (NFATc1) is essential for this process. We previously found, using mouse RAW264 cells, that culture at high cell density blocked progression to the multinucleated cell stage induced by stimulation with receptor activator of nuclear factor kappaB ligand (RANKL). Here, we have confirmed this finding in a bone marrow cell system and extended the analysis further.