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Abstract Background We have recently found a phenomenon that spontaneous regeneration of a hyaline cartilage-like tissue can be induced in a large osteochondral defect by implanting a double-network (DN) hydrogel plug, which was composed of poly-(2-Acrylamido-2-methylpropanesulfonic acid) and poly-(N, N'-Dimetyl acrylamide), at the bottom of the defect. The purpose of this study was to clarify gene expression profile of the regenerated tissue in comparison with that of the normal articular cartilage. Methods We created a cylindrical osteochondral defect in the rabbit femoral grooves. Then, we implanted the DN gel plug at the bottom of the defect. At 2 and 4 weeks after surgery, the regenerated tissue was analyzed using DNA microarray and immunohistochemical examinations. Results The gene expression profiles of the regenerated tissues were macroscopically similar to the normal cartilage, but showed some minor differences. The expression degree of COL2A1, COL1A2, COL10A1, DCN, FMOD, SPARC, FLOD2, CHAD, CTGF, and COMP genes was greater in the regenerated tissue than in the normal cartilage. The top 30 genes that expressed 5 times or more in the regenerated tissue as compared with the normal cartilage included type-2 collagen, type-10 collagen, FN, vimentin, COMP, EF1alpha, TFCP2, and GAPDH genes. Conclusions The tissue regenerated by using the DN gel was genetically similar but not completely identical to articular cartilage. The genetic data shown in this study are useful for future studies to identify specific genes involved in spontaneous cartilage regeneration. more...

The purposes of this study were to identify signaling pathways that were specifically activated in ATDC5 cells cultured on poly (2-acrylamido-2-methylpropanesulfonic acid) (PAMPS) gel in insulin-free maintenance medium and to evaluate the significance of the determined signaling pathways in the chondrogenic differentiation induced by this gel. In this study, ATDC5 cells cultured on PAMPS gel using the maintenance medium without insulin (PAMPS Culture) were compared with cells cultured on polystyrene using the differentiation medium containing insulin (PS-I Culture). The microarray analysis, Western blot analysis, and real-time PCR analysis demonstrated that the TGF-β/BMP signaling pathway was significantly enhanced at Days 1, 2, and 3 in the PAMPS Culture when compared with the PS-I Culture. Inhibition of the BMP type-I receptor reduced the phosphorylation level of Smad1/5 and expression of type-2 collagen and aggrecan mRNA in the cells accompanied by a reduction in cell aggregation at Day 13 in the PAMPS Culture. The inhibition of the TGF-β/BMP signaling pathway significantly inhibited the chondrogenic differentiation induced by the PAMPS gel. The present study demonstrated that synthetic PAMPS gel activates the TGF-β/BMP/Smad signaling pathway in the ATDC5 cells in the absence of insulin, and that this activation plays a significant role in the chondrogenic differentiation induced by PAMPS gel. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 734-746, 2016., (© 2015 Wiley Periodicals, Inc.) more...

A double-network (DN) gel, which was composed of poly(2-acrylamido-2-methylpropanesulfonic acid) and poly(N,N'-dimethyl acrylamide) (PAMPS/PDMAAm), has the potential to induce chondrogenesis both in vitro and in vivo. The present study investigated the biomechanical and biological responses of chondrogenic progenitor ATDC5 cells cultured on the DN gel. ATDC5 cells were cultured on a polystyrene surface without insulin (Culture 1) and with insulin (Culture 2), and on the DN gel without insulin (Culture 3). The cultured cells were evaluated using micropipette aspiration for cell Young's modulus and qPCR for gene expression of chondrogenic and actin organization markers on days 3, 7 and 14. On day 3, the cells in Culture 3 formed nodules, in which the cells exhibited an actin cortical layer inside them, and gene expression of type-II collagen, aggrecan, and SOX9 was significantly higher in Culture 3 than Cultures 1 and 2 (p<0.05). Young's modulus in Culture 3 was significantly higher than that in Culture 1 throughout the testing period (p<0.05) and that in Culture 2 on day 14 (p<0.01). There was continuous expression of actin organization markers in Culture 3. This study highlights that the cells on the DN gel increased the modulus and mRNA expression of chondrogenic markers at an earlier time point with a greater magnitude compared to those on the polystyrene surface with insulin. This study also demonstrates a possible strong interrelation among alteration of cell mechanical properties, changes in actin organization and the induction of chondrogenic differentiation., (Copyright © 2014 Elsevier Ltd. All rights reserved.) more...

Polyzwitterionic materials, which have both cationic and anionic groups in the polymeric repeat unit, show excellent anti-biofouling properties and are drawing more attention in the biomedical field. In this study, we have successfully synthesized novel single network hydrogels and double network (DN) hydrogels from the zwitterionic monomer, N-(carboxymethyl)-N,N-dimethyl-2-(methacryloyloxy) ethanaminium, inner salt (CDME). The polyCDME (PCDME) single network hydrogel behaves like a hydrophilic neutral hydrogel and its properties are not sensitive to temperature, pH, or ionic strength over a wide range. DN hydrogels using the poly(2-acrylamido-2-methylpropanesulfonic) (PAMPS) as the first network and PCDME as the second network, having a Young's modulus of 0.2-0.9 MPa, possess excellent mechanical strength (fracture stress: 1.2-1.4 MPa, fracture strain: 2.2-6.0 mm/mm) and toughness (work of extension at fracture: 0.9-2.4 MJ m -3 ) depending on the composition ratio of PCDME to PAMPS. The strength and toughness of the optimized PAMPS/PCDME DN is comparable to the normal PAMPS/PAAm DN hydrogels that use poly(acrylamide) (PAAm) as the second network. By macrophage adhesion test, both the PCDME hydrogels and the PAMPS/PCDME DN hydrogels have shown excellent anti-biofouling properties. These results demonstrate that the PCDME-based DN hydrogels have high potential as a novel soft and wet biomaterial. more...

Marine sessile organisms easily adhere to submerged solids such as rocks, metals and plastics, but not to seaweeds and fishes, which are covered with soft and wet 'hydrogel'. Inspired by this fact, we have studied long-term antifouling properties of hydrogels against marine sessile organisms. Hydrogels, especially those containing hydroxy group and sulfonic group, show excellent antifouling activity against barnacles both in laboratory assays and in the marine environment. The extreme low settlement on hydrogels in vitro and in vivo is mainly caused by antifouling properties against the barnacle cypris. more...

This study evaluated the wear property of four novel double-network (DN) hydrogels, which was composed of two kinds of hydrophilic polymers, using pin-on-flat wear testing. The gels involve PAMPS-PAAm gel which consists of poly(2-acrylamide-2-metyl-propane sulfonic acid) and polyacrylamide, PAMPS-PDAAAm gel which consists of poly(2-acrylamide-2-metyl-propane sulfonic acid) and poly(N,N'-dimetyl acrylamide), Cellulose/PDMAAm gel which consists of bacterial Cellulose and poly dimetyl-acrylamide, and Cellulose-Gelatin gel which consists of bacterial Cellulose and Gelatin. Ultra-high molecular weight polyethylene (UHMWPE) was used as a control of a clinically available material. Using a reciprocating apparatus, 10(6) cycles of friction between a flat specimen and ceramic pin were repeated in water under a contact pressure of 0.1 MPa. To determine the depth and the roughness of the concave lesion created by wear, a confocal laser microscope was used. As a result, the maximum wear depth of the PAMPS-PDMAAm gel (3.20 microm) was minimal in the five materials, while there was no significant difference compared to UHMWPE. There were significant differences between UHMWPE and one of the other three gels. The PAMPS-PAAm gel (9.50 microm), the Cellulose-PDMAAm gel (7.80 microm), and the Cellulose-Gelatin gel (1302.40 microm). This study demonstrated that the PAMPS-PDMAAm DN gel has an amazing wear property as a hydrogel that is comparable to the UHMWPE. In addition, the PAMPS-PAAm and Cellulose-PDMAAm DN gels are also resistant to wear to greater degrees than conventionally reported hydrogels. On the other hand, this study showed that the Cellulose-Gelatin DN gel was not resistant to wear. more...

It was found that when an aqueous solution of vinyl monomers is polymerized on a hydrophobic substrate, obvious heterogeneity occurs in the region of the interface. This substrate effect was observed on polytetrafluroethylene (Teflon), polypropylene (PP), polyethylene (PE), polystyrene (PS), and polyvinylchloride (PVC), but not on hydrophilic substrates. Compared with synthesis on hydrophilic surfaces, the surfaces of hydrogels synthesized on a hydrophobic substrate exhibit a larger degree of swelling, a lower surface coefficient of friction and elastic modulus, weaker interfacial adhesion, and reduced interaction with biological cells. This substrate effect has been observed for many types of aqueous monomer solutions. It was found that the above properties are related to the loosely cross-linked architecture, containing some graft-like polymer chains, that is formed on the gel surface when the gel is prepared on a hydrophobic substrate. To understand the mechanism of the substrate effect, two novel optical methods, electric speckle pattern interferometry (ESPI) and real-time laser sheet refraction (RT-LSR), were developed. It was found that oxygen trapped in the composite interface between the monomer solution and rough hydrophobic substrates played an important role in the substrate effect., (Copyright 2003 The Japan Chemical Journal Forum and Wiley Periodicals, Inc., Chem Rec 3: 40-50; 2003: Published online in Wiley InterScience (www.interscience.wiley.com) DOI 10.1002/tcr.10048) more...