Immunosuppressive mesenchymal stem cells aggregates incorporating hydrogel microspheres promote an in vitro invasion of cancer cells

[Introduction] The objective of this study is to design a co-culture system of cancer cells and three-dimensional (3D) mesenchymal stem cells (MSC) aggregates for the in vitro evaluation of cancer invasion. [Methods] First, the MSC of an immunosuppressive phenotype (MSC2) were prepared by the MSC stimulation of polyriboinosinic polyribocytidylic acid. By simple mixing MSC2 and gelatin hydrogel microspheres (GM) in a U-bottomed well of 96 well plates which had been pre-coated with poly (vinyl alcohol), 3D MSC2 aggregates incorporating GM were obtained. The amount of chemokine (C–C motif) ligand 5 (CCL5) secreted from the MSC2 aggregates incorporating GM. Finally, an invasion assay was performed to evaluate the cancer invasion rate by co-cultured cancer cells and the 3D MSC2 incorporating GM. [Results] The amount of CCL5 secreted for the 3D MSC2 aggregates incorporating GM was significantly higher than that of two-dimensional (2D) MSC, 2D MSC2, and 3D MSC aggregates incorporating GM. When MDA-MB-231 human breast cancer cells were co-cultured with the 3D MSC2 aggregates incorporating GM, the invasion rate of cancer cells was significantly high compared with that of 2D MSC or 2D MSC2 and 3D MSC aggregates incorporating GM. In addition, high secretion of matrix metalloproteinase-2 was observed for the 3D MSC2 aggregates/cancer cells system. [Conclusions] It is concluded that the co-culture system of 3D MSC2 aggregates incorporating GM and cancer cells is promising to evaluate the invasion of cancer cells in vitro

Effect of ProNectin F derivatives on cell attachment and proliferation.

ProNectin F (PnF) was chemically modified by introducing some functional groups to prepare various derivatives of primary amino (PnF-N(1)), tertiary amino (PnF-N(3)), quaternary ammonium (PnF-N(4)), carboxyl (PnF-COOH) and sulfonyl groups (PnF-SO(3)H). When C3H10T1/2 cells were cultured on non-treated dishes coated with the derivatives, the number of mesenchymal cells attached to the culture dishes increased for the coating with PnF-COOH and PnF-SO(3)H, even at their low adsorption amount. The cytotoxicity was high for the coating of PnF-N(1) and PnF-N(4) compared with that of the PnF-N(3), PnF-COOH and PnF-SO(3)H. The treatment with integrin α5 and αV antibodies suppressed the cell attachment to the dishes coated with PnF-COOH and PnF-SO(3)H. The phosphorylation of extracellular signal-regulated kinase (ERK) was upregulated for cells attached to the dishes coated with PnF-COOH and PnF-SO(3)H, indicating their enhanced proliferation. It is concluded that the chemical derivatization of PnF enhanced the ability of cell attachment and proliferation

高分子微粒子のマクロファージによる貪食とマクロファージの抗腫瘍活性化

京都大学0048新制・課程博士工学博士甲第3891号工博第991号新制||工||709(附属図書館)UT51-63-C109京都大学大学院工学研究科高分子化学専攻(主査)教授 筏 義人, 教授 東村 敏延, 教授 今西 幸男学位規則第5条第1項該当Kyoto UniversityDFA

Enhancement of ectopic osteoid formation following the dual release of bone morphogenetic protein 2 and Wnt1 inducible signaling pathway protein 1 from gelatin sponges.

Bone morphogenetic protein (BMP) 2-incorporated gelatin sponge is effective for in vivo osteoinduction. However, the modeling capacity of bone decreases with age. As atrial to stimulate effective bone formation for animals with decreased osteogenic potential, Wnt1 inducible signaling pathway protein (WISP) 1, an osteoblastic regulator, was combined with gelatin sponge incorporating BMP2. Osteopontin (Opn) geneexpression was increased in vitro for mouse bone marrow stromal cells (BMSC) cultured in gelatin sponges incorporating BMP2 and WISP1 compared with those incorporating BMP2 or WISP1 alone. In vivo synergistic effect of BMP2 and WISP1 on the ectopic osteoid formation was observed when gelatin sponges incorporating BMP2 and WISP1 were implanted subcutaneously into middle-aged mice with decreased bone formation potential. It is concluded that the scaffold incorporating multiple osteoinductive agents could be effective in inducing bone formation in those with age-related decreased potential of bone formation

Local Regeneration of Dentin-Pulp Complex Using Controlled Release of FGF-2 and Naturally Derived Sponge-Like Scaffolds

Restorative and endodontic procedures have been recently developed in an attempt to preserve the vitality of dental pulp after exposure to external stimuli, such as caries infection or traumatic injury. When damage to dental pulp is reversible, pulp wound healing can proceed, whereas irreversible damage induces pathological changes in dental pulp, eventually requiring its removal. Nonvital teeth lose their defensive abilities and become severely damaged, resulting in extraction. Development of regeneration therapy for the dentin-pulp complex is important to overcome limitations with presently available therapies. Three strategies to regenerate the dentin-pulp complex have been proposed; regeneration of the entire tooth, local regeneration of the dentin-pulp complex from amputated dental pulp, and regeneration of dental pulp from apical dental pulp or periapical tissues. In this paper, we focus on the local regeneration of the dentin-pulp complex by application of exogenous growth factors and scaffolds to amputated dental pulp

Efficacy and duration of analgesia from a sustained-release lidocaine sheet in humans

BACKGROUNDWe have synthesized a sustained-releaselidocaine sheet (SRLS) and injectable sustained-release lidocaine particles(SRLP) using biodegradable polymers. In the present study, we performed anexploratory first clinical trial of the SRLS in healthy volunteers as a preludeto patient administration. This trial is meant as an initial intervention inultimately developing and refining the SRLP. METHODSWe evaluated the intensity and duration ofanalgesia of the SRLS compared with 8% lidocaine spray. In Protocol 1, weapplied the SRLS piece to the mucous membrane of the nasal vestibule. Weexamined the local pain threshold over 72 h after administration, and removedthe SRLS after 72 h. Individuals that finished Protocol 1 underwent Protocol 2,in which we applied 8% lidocaine spray. RESULTSTwelve volunteers were enrolled and seven ofthese volunteers finished Protocol 1. All seven individuals who completedProtocol 1 also completed Protocol 2. The mean pain thresholds were 32 g, 78 g,90 g, 90 g, 87 g, and 87 g at pre-administration and 4 h, 10 h, 24 h, 48 h, and72 h after administration, respectively, in Protocol 1, and 36 g, 85 g, 49 g,and 33 g at pre-administration and 15 min, 2 h, and 4 h, respectively, inProtocol 2. CONCLUSIONA sustained-release lidocaine usingbiodegradable polymers was applied as a sheet in humans for the first time inthe world. It maintained significant analgesia for 72 h without majortoxicities. Furthermore, degree of analgesia provided by the SRLS throughoutthe entire study was similar to that provided by the 8% lidocaine spray. It may suitable for management ofpostoperative pain especially in outpatients

Gelatin hydrogel nonwoven fabrics of a cell culture scaffold to formulate 3-dimensional cell constructs

The objective of this study is to evaluate the possibility of gelatin hydrogel nonwoven fabrics (GHNF) of a cell culture scaffold to formulate 3-dimensional (3D) cell construct. The thickness of cell construct is about 1 mm and the cells inside are live and bio-active, irrespective of their internal distribution. The GHNF were prepared by the solution blow method of gelatin, following by dehydrothermal crosslinking. The GHNF showed a mechanical strength strong enough not to allow the shape to deform even in a wet state. The wet GHNF also showed resistance against repeated compression. After human mesenchymal stromal cells (hMSC) were seeded and cultured, the inner distribution in GHNF, the apoptosis, hypoxia inducible factor (HIF)-1α, Ki67, collagen or sulfated glycosaminoglycan (sGAG) secretion of cells were evaluated. The hMSC proliferated inside the GHNF with time while a homogeneous distribution in the number of cells proliferated from the surface to the 1000 μm depth of GHNF was observed. The number of apoptosis and HIF-1α positive cells was significantly low compared with that of polypropylene nonwoven fabrics with the similar fiber diameters and intra-structure. The GHNF were degraded during cell culture, and completely replaced by collagen and sGAG secreted. It is concluded that the GHNF is a promising cell culture scaffold for 3D cell constructs