Introduction of N-cadherin-binding motif to alginate hydrogels for controlled stem cell differentiation

Lee, Jae Won,An, Hyoseok,Lee, Kuen Yong Elsevier 2017 Colloids and surfaces. B, Biointerfaces Vol.155 No.-

<P><B>Abstract</B></P> <P>Control of stem cell fate and phenotype using biomimetic synthetic extracellular matrices (ECMs) is an important tissue engineering approach. Many studies have focused on improving cell-matrix interactions. However, proper control of cell-cell interactions using synthetic ECMs could be critical for tissue engineering, especially with undifferentiated stem cells. In this study, alginate hydrogels were modified with a peptide derived from the low-density lipoprotein receptor-related protein 5 (LRP5), which is known to bind to N-cadherin, as a cell-cell interaction motif. <I>In vitro</I> changes in the morphology and differentiation of mouse bone marrow stromal cells (D1 stem cells) cultured in LRP5-alginate hydrogels were investigated. LRP5-alginate gels successfully induced stem cell aggregation and enhanced chondrogenic differentiation of D1 stem cells, compared to RGD-alginate gels, at low cell density. This approach to tailoring synthetic biomimetic ECMs using cell-cell interaction motifs may be critical in tissue engineering approaches using stem cells.</P> <P><B>Highlights</B></P> <P> <UL> <LI> LRP5 peptide can be successfully conjugated to alginate hydrogel. </LI> <LI> LRP5-alginate gel induces N-cadherin-mediated cell-cell interaction. </LI> <LI> LRP5-alginate gel enhances stem cell aggregation and chondrogenic differentiation. </LI> <LI> Controlling cell-cell interaction is critical in tissue engineering applications. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

Monitoring of cell-cell interaction in a programmable static droplet array with a confined microenvironment

김동영,진시형,이병진,정성근,이성식,( Matthias Peter ),이창수 한국공업화학회 2019 한국공업화학회 연구논문 초록집 Vol.2019 No.0

Cell-cell interaction can occur through biochemical signaling molecules, regulating a variety of biological responses such as quorum sensing, biofilm formation, and inflammatory response. However, traditional cell culture systems are difficult to accurately reproduce the effects of cell-cell interactions and mixed cell populations, have no single cell resolution, and often have limited sensitivity and accuracy. Thus, there is a growing need for a novel platform for isolating, manipulating, tracking, and imaging cells to better study the various cell-cell interaction scenarios. Here we present a programmable static droplet array (SDA) for monitoring of the effect of population on natural cell-cell contact and signaling processes in a confined microenvironment. The SDA provides a valuable research tool to study cell-cell communication and signaling in a controlled microenvironment with the sensitivity and accuracy required for screening and long-term phenotypic analysis.

Development of a 2-Chamber Culture System for Impedimetric Monitoring of Cell-cell Interaction

Kin Fong Lei,Meng-Tsan Tsai,Ming-Hong Zhong,Chia-Hao Huang,Ngan-Ming Tsang,Ming-Yih Lee 한국바이오칩학회 2017 BioChip Journal Vol.11 No.2

In cancer research, study of cell-cell interaction is important to understand tumor initiation, progression, metastasis, and therapeutic resistance. Conventionally, transwell system was adopted and cell proliferation was quantified by end-point bio-assays. The operations are labor-intensive and time-consuming while studying of the dynamic cellular responses of cell-cell interaction. Although impedance measurement was suggested to be a promising technique to monitor cellular responses, electrodes cannot be integrated into the transwell for the measurement purpose. In this work, a 2-chamber culture system incorporated with impedance measurement technique was developed to quantitatively study cell-cell interaction. The chamber was composed of 2 sub-chambers separated with a barrier. By this design, two types of cells could be independently cultured and concurrently monitored under common medium supplied. Cell-cell interaction was demonstrated by aberrant cell proliferation induced by the EGF secreted from the transfected cells cultured on another sub-chamber. Real-time and non-invasive monitoring of cell-cell interaction was successfully demonstrated. This work provides a practical solution for monitoring the dynamic cellular responses of cellcell interaction during the culture course. It is a reliable and convenient platform and facilitate more quantitative assessments in cancer research.

Growth and Migration of BALB/3T3 Fibroblast Cells on Nano-engineered Silica Beads Surface

Jihee Kim,Prakash Chandra,Jiyoon Yang,Seog Woo Rhee 대한화학회 2013 Bulletin of the Korean Chemical Society Vol.34 No.12

In this study, the behavior of cells on the modified surface, and the correlation between the modified substrates and the response of cells is described. A close-packed layer of nano-sized silica beads was prepared on a coverslip, and the adhesion, proliferation, and migration of BALB/3T3 fibroblast cells on the silica layer was monitered. The 550 nm silica beads were synthesized by the hydrolysis and condensation reaction of tetraethylorthosilicate in basic solution. The amine groups were introduced onto the surfaces of silica particles by treatment with 3-aminopropyltrimethoxysilane. The close-packed layer of silica beads on the coverslip was obtained by the reaction of the amine-functionalized silica beads and the (3-triethoxysilyl)propylsuccinic anhydride treated coverslip. BALB/3T3 fibroblast cells were loaded on bare glass, APTMS coated glass, and silica bead coated glass with the same initial cell density, and the migration and proliferation of cells on the substrates was investigated. The cells were fixed and stained with antibodies in order to analyze the changes in the actin filaments and nuclei after culture on the different surfaces. The motility of cells on the silica bead coated glass was greater than that of the cells cultured on the control substrate. The growth rate of cells on the silica bead coated glass was slower than that of the control. Because the close-packed layer of silica beads gave an embossed surface, the adhesion of cells was very weak compared to the smooth surfaces. These results indicate that the adhesion of cells on the substrates is very important, and the actin filaments might play key roles in the migration and proliferation of cells. The nuclei of the cells were shrunk on the weakly adhered surfaces, and the S1 stage in which DNA is duplicated in the cell dividing processes might be retarded. As a result, the rate of proliferation of cells was decreased compared to the smooth surface of the control. In conclusion, the results described here are very important in the understanding of the interaction between implanted materials and biosystems.

The Ring-H2 Finger Motif of CKBBP1/SAG Is Necessary for Interaction with Protein Kinase CK2 and Optimal Cell Proliferation

( Yun Sook Kim ),( Kwon Soo Ha ),( Young Ho Kim ),( Young Seuk Bae ) 생화학분자생물학회 2002 BMB Reports Vol.35 No.6

Protein kinase CKII (CKII) is required for progression through the cell division. We recently reported that the β subunit of protein kinase CKII (CKIIβ) associates with CKBBP1 that contains the Ring-H2 finger motif in the yeast two-hybrid system. We demonstrate here that the Ring-H2 finger-disrupted mutant of CKBBP1 does not interact with purified CKIIβ in vitro, which shows that the Ring-H2 finger motif is critical for direct interaction with CKIIβ. The CKII holoenzyme is efficiently co-precipitated with the wild-type CKBBP1, but not with the Ring-H2 finger-disrupted CKBBP1, from whole cell extracts when epitope-tagged CKBBP1 is transiently expressed in HeLa cells. Disruption of the Ring-H2 finger motif does not affect the cellular localization of CKBBP1 in HeLa cells. The increased expression of either the wild-type CKBBP1 or Ring-H2 finger-disrupted CKBBP1 does not modulate the protein or the activity levels of CKII in HeLa cells. However, the stable expression of Ring-H2 finger-disrupted CKBBP1 in HeLa cells suppresses cell proliferation and causes the accumulation of the G1/G0 peak of the cell cycle. The Ring-H2 finger motif is required for maximal CKBBP1 phosphorylatlon by CKII, suggesting that the stable binding of CKBBP1 to CKII is necessary for its efficient phosphorylation. Taken together, these results suggest that the complex formation of CKIIβ with CKBBP1 and/or CKII-mediated CKBBP1 phosphorylation is important for the G1/S phase transition of the cell cycle.

2P-329 Co-culture system using reversible cell layering mediated by ionic crosslinking of chitosan and functionalized cell surface membrane

유승미,김현범,강미경,황석연,김병수 한국공업화학회 2017 한국공업화학회 연구논문 초록집 Vol.2017 No.1

Cells in living tissues are engaged in complex cell-cell interaction with heterogeneous cells in the face of a dynamically changing in vivo environment. In order to recapitulate the complex microenvironment, various cellular assembly approaches, mostly rely on irreversible cell layering, have been proposed. Inefficient interactions among cells in the techniques prevent characterization and therapeutic applications of the cells following co-culture. Here, we develop a reversible cell layering system for heterogeneous cell assembly mediated by ionic cross-linking of chitosan and a functionalized cell surface membrane. Anionic maleimide-chondroitin-sulfate is grafted onto the surface membrane of myogenic cells and human mesenchymal stem cells (hMSCs) via ionic cross-linking forming assembled double-layered cell constructs. hMSCs differentiated through this system showed higher levels of myogenic marker expression during muscle regeneration.

Growth and Migration of BALB/3T3 Fibroblast Cells on Nano-engineered Silica Beads Surface

Kim, Jihee,Chandra, Prakash,Yang, Jiyoon,Rhee, Seog Woo Korean Chemical Society 2013 Bulletin of the Korean Chemical Society Vol.34 No.12

In this study, the behavior of cells on the modified surface, and the correlation between the modified substrates and the response of cells is described. A close-packed layer of nano-sized silica beads was prepared on a coverslip, and the adhesion, proliferation, and migration of BALB/3T3 fibroblast cells on the silica layer was monitered. The 550 nm silica beads were synthesized by the hydrolysis and condensation reaction of tetraethylorthosilicate in basic solution. The amine groups were introduced onto the surfaces of silica particles by treatment with 3-aminopropyltrimethoxysilane. The close-packed layer of silica beads on the coverslip was obtained by the reaction of the amine-functionalized silica beads and the (3-triethoxysilyl)propylsuccinic anhydride treated coverslip. BALB/3T3 fibroblast cells were loaded on bare glass, APTMS coated glass, and silica bead coated glass with the same initial cell density, and the migration and proliferation of cells on the substrates was investigated. The cells were fixed and stained with antibodies in order to analyze the changes in the actin filaments and nuclei after culture on the different surfaces. The motility of cells on the silica bead coated glass was greater than that of the cells cultured on the control substrate. The growth rate of cells on the silica bead coated glass was slower than that of the control. Because the close-packed layer of silica beads gave an embossed surface, the adhesion of cells was very weak compared to the smooth surfaces. These results indicate that the adhesion of cells on the substrates is very important, and the actin filaments might play key roles in the migration and proliferation of cells. The nuclei of the cells were shrunk on the weakly adhered surfaces, and the S1 stage in which DNA is duplicated in the cell dividing processes might be retarded. As a result, the rate of proliferation of cells was decreased compared to the smooth surface of the control. In conclusion, the results described here are very important in the understanding of the interaction between implanted materials and biosystems.

Cellular behavior in micropatterned hydrogels by bioprinting system depended on the cell types and cellular interaction

Hong, S.,Song, S.J.,Lee, J.Y.,Jang, H.,Choi, J.,Sun, K.,Park, Y. Society for Bioscience and Bioengineering, Japan ; 2013 Journal of bioscience and bioengineering Vol.116 No.2

The fabrication of patterned microstructures within three-dimensional (3D) matrices is a challenging subject in tissue engineering and regenerative medicine. A 3D, free-moving bioprinting system was developed and hydrogels were patterned by varying the process parameters of z-axis moving velocity and ejection velocity. The patterning of hydrogel based microfibers in a 3D matrigel was achieved with dimensions of 4.5 mm length and widths from 79 to 200 μm. Hyaluronan-based hydrogels mixed with fibroblasts (L929), mouse endothelial cells (MS1), or human mesenchymal stem cells (hMSCs) were patterned using a 3D moving axis bioprinter and cell behavior was monitored in culture for up to 16 days. L929 and MS1 cells and hMSCs in patterned hydrogel revealed cell-cell interactions and a morphological dependency on cell types. HMSCs formed spheres through cell aggregation, while L929 cells increased in cellular mass without cell aggregation and MS1 dispersed into the matrix instead of aggregating. The aggregation of hMSCs was attenuated by treatment with Rho kinase (ROCK) inhibitor and cadherin antibody. This reflected the close relationship between cell aggregation and migration with RhoA and cell-cell adhesion molecules. Angiogenic-specific gene expression profiles showed that expression of CD105 decreased to 22% in the ROCK inhibitor group compared to control group. These results showed that cell-based patterns in a 3D matrix are highly dependent on both cell aggregation and migration over time.

Th17 Cell and Inflammatory Infiltrate Interactions in Cutaneous Leishmaniasis: Unraveling Immunopathogenic Mechanisms

Abraham U. Morales-Primo,Ingeborg Becker,Claudia Patricia Pedraza-Zamora,Jaime Zamora-Chimal The Korean Association of Immunobiologists 2024 Immune Network Vol.24 No.2

The inflammatory response during cutaneous leishmaniasis (CL) involves immune and non-immune cell cooperation to contain and eliminate Leishmania parasites. The orchestration of these responses is coordinated primarily by CD4⁺ T cells; however, the disease outcome depends on the Th cell predominant phenotype. Although Th1 and Th2 phenotypes are the most addressed as steers for the resolution or perpetuation of the disease, Th17 cell activities, especially IL-17 release, are recognized to be vital during CL development. Th17 cells perform vital functions during both acute and chronic phases of CL. Overall, Th17 cells induce the migration of phagocytes (neutrophils, macrophages) to the infection site and CD8⁺ T cells and NK cell activation. They also provoke granzyme and perforin secretion from CD8⁺ T cells, macrophage differentiation towards an M2 phenotype, and expansion of B and Treg cells. Likewise, immune cells from the inflammatory infiltrate have modulatory activities over Th17 cells involving their differentiation from naive CD4⁺ T cells and further expansion by generating a microenvironment rich in optimal cytokines such as IL-1β, TGF-β, IL-6, and IL-21. Th17 cell activities and synergies are crucial for the resistance of the infection during the early and acute stages; however, if unchecked, Th17 cells might lead to a chronic stage. This review discusses the synergies between Th17 cells and the inflammatory infiltrate and how these interactions might destine the course of CL.

High throughput approaches for controlled stem cell differentiation

Kim, H.D.,Lee, E.A.,Choi, Y.H.,An, Y.H.,Koh, R.H.,Kim, S.L.,Hwang, N.S. Elsevier BV 2016 ACTA BIOMATERIALIA Vol.34 No.-

Stem cells have unique ability to undergo self-renewal indefinitely in culture and potential to differentiate into almost all cell types in the human body. However, the developing a method for efficiently differentiating or manipulating these stem cells for therapeutic purposes remains a challenging problem. Pluripotent stem cells, as well as adult stem cells, require biological cues for their proliferation and differentiation. These cues are largely controlled by cell-cell, cell-insoluble factors (such as extracellular matrix), and cell-soluble factors (such as cytokine or growth factors) interactions. In this review, we describe a state of research on various stem cell-based tissue engineering applications and high throughput strategies for developing synthetic or biosynthetic microenvironments to allow efficient commitments in stem cells. Statement of Significance: Nowadays, pluripotency of stem cells have received much attention to use therapeutic purpose. However, a major difficulty with stem cell therapy is to control its differentiation through desired cells or tissues. In other words, various microenvironment factors are involved during stem cell differentiation, including dimensionality, growth factors, cell junctions, nutritional status, matrix stiffness, matrix composition, mechanical stress, and cell-matrix adhesion. Therefore, researchers have engineered a variety of platforms to enable controlling and monitoring bioactive factors to induce stem cell commitment. In this review, we report on recent advancements in a novel technology based on high-throughput strategies for stem cell-based tissue engineering applications.