Rho kinase inhibitor Y-27632 promotes the differentiation of human bone marrow mesenchymal stem cells into keratinocyte-like cells in xeno-free conditioned medium

INTRODUCTION: Bone marrow mesenchymal stem cells (BMSCs), which have the ability to self-renew and to differentiate into multiple cell types, have recently become a novel strategy for cell-based therapies. The differentiation of BMSCs into keratinocytes may be beneficial for patients with burns, disease, or trauma. However, the currently available cells are exposed to animal materials during their cultivation and induction. These xeno-contaminations severely limit their clinical outcomes. Previous studies have shown that the Rho kinase (ROCK) inhibitor Y-27632 can promote induction efficiency and regulate the self-renewal and differentiation of stem cells. In the present study, we attempted to establish a xeno-free system for the differentiation of BMSCs into keratinocytes and to investigate whether Y-27632 can facilitate this differentiation. METHODS: BMSCs isolated from patients were cultured by using a xeno-free system and characterised by using flow cytometric analysis and adipogenic and osteogenic differentiation assays. Human primary keratinocytes were also isolated from patients. Then, the morphology, population doubling time, and β-galactosidase staining level of these cells were evaluated in the presence or absence of Y-27632 to determine the effects of Y-27632 on the state of the keratinocytes. Keratinocyte-like cells (KLCs) were detected at different time points by immunocytofluorescence analysis. Moreover, the efficiency of BMSC differentiation under different conditions was measured by quantitative real-time-polymerase chain reaction (RT-PCR) and Western blot analyses. RESULTS: The ROCK inhibitor Y-27632 promoted the proliferation and lifespan of human primary keratinocytes. In addition, we showed that keratinocyte-specific markers could be detected in BMSCs cultured in a xeno-free system using keratinocyte-conditioned medium (KCM) independent of the presence of Y-27632. However, the efficiency of the differentiation of BMSCs into KLCs was significantly higher in the presence of Y-27632 using immunofluorescence, quantitative RT-PCR, and Western blot analyses. CONCLUSIONS: This study demonstrated that Y-27632 could promote the proliferation and survival of human primary keratinocytes in a xeno-free culture system. In addition, we found that BMSCs have the ability to differentiate into KLCs in KCM and that Y-27632 can facilitate this differentiation. Our results suggest that BMSCs are capable of differentiating into KLCs in vitro and that the ROCK pathway may play a critical role in this process

Leukadherin-1-Mediated Activation of CD11b Inhibits LPS-Induced Pro-inflammatory Response in Macrophages and Protects Mice Against Endotoxic Shock by Blocking LPS-TLR4 Interaction

Dysregulation of macrophage has been demonstrated to contribute to aberrant immune responses and inflammatory diseases. CD11b, expressed on macrophages, plays a critical role in regulating pathogen recognition, phagocytosis, and cell survival. In the present study, we explored the effect of leukadherin-1 (LA1), an agonist of CD11b, on regulating LPS-induced pro-inflammatory response in macrophages and endotoxic shock. Intriguingly, we found that LA1 could significantly reduce mortalities of mice and alleviated pathological injury of liver and lung in endotoxic shock. In vivo studies showed that LA1-induced activation of CD11b significantly inhibited the LPS-induced pro-inflammatory response in macrophages of mice. Moreover, LA1-induced activation of CD11b significantly inhibited LPS/IFN-γ-induced pro-inflammatory response in macrophages by inhibiting MAPKs and NF-κB signaling pathways in vitro. Furthermore, the mice injected with LA1-treated BMDMs showed fewer pathological lesions than those injected with vehicle-treated BMDMs in endotoxic shock. In addition, we found that activation of TLR4 by LPS could endocytose CD11b and activation of CD11b by LA1 could endocytose TLR4 in vitro and in vivo, subsequently blocking the binding of LPS with TLR4. Based on these findings, we concluded that LA1-induced activation of CD11b negatively regulates LPS-induced pro-inflammatory response in macrophages and subsequently protects mice from endotoxin shock by partially blocking LPS-TLR4 interaction. Our study provides a new insight into the role of CD11b in the pathogenesis of inflammatory diseases

Viscosity Behavior of P(DAC-AM) with Serial Cationicity and Intrinsic Viscosity in Inorganic Salt Solutions

The poly(acryloyloxyethyl trimethyl ammonium chloride–co–acrylamide), P(DAC-AM), is a kind of cationic polyelectrolyte usually applied in a solution form, and its performance is affected by its structure and the environment where it is used. In particular, its viscosity properties in salt solutions are directly related to its efficacy in various applications, and the performance is one of the most important solution properties. Therefore, in this paper, the effects of the salt concentration and valence of seven kinds of inorganic salts, NaCl, LiCl, KCl, MgCl2, AlCl3, Na2SO4, and Na3PO4, on the values of apparent viscosity (ηa) of P(DAC-AM) samples with cationicity of 10%, 50%, and 90%, and intrinsic viscosity ([η]) of 5, 10, and 15 dL/g were investigated. The ηa was determined using a rotational viscometer. The interaction mechanism between the polymers and salt ions was also investigated. The results showed that depending on the salt concentration, the ηa firstly decreased sharply to the inflection point which indicated the minimum volume of the molecule shrinking, and then either maintained the value unchanged or increased. The salt concentration corresponding to the inflection point decreased with the increase of the salt ion valence but with the reduction of the cationicity of the polymer. The ηa at the inflection point increased as the [η] of the polymer grew. This indicated that the salt concentration and the salt ion valence had a notable impact on the stretch of the cationic polymer molecule in the salt solutions. It was discovered that the phenomenon of the increase of the ηa of P(DAC-AM) samples in the multivalent salt solutions after the inflection point was caused by not only the increase of the ηa of the complexes formed from the pure salts, but also the viscosity resistance of the charge and volume between the polymer molecules and salt ions, as well as the complexes themselves. The linear relationship between the increased ηa and the salt concentration, representing the interaction both among the complexes themselves and between the polymer and complexes, was obtained. Furthermore, the interaction model between the salt ions and P(DAC-AM) molecules in a wide range of salt concentrations was illustrated.</jats:p

Kinetics of acryloyloxyethyl trimethyl ammonium chloride polymerization in aqueous solutions

KINETICS OF ACRYLOYLOXYETHYL TRIMETHYL AMMONIUM CHLORIDE POLYMERIZATION IN AQUEOUS SOLUTIONS e-Polymers (Rights reserved) (-) Kinetics of acryloyloxyethyl trimethyl ammonium chloride polymerization in aqueous solutions / Fu, Xingqin (CC BY) (-

Research Progress on Typical Quaternary Ammonium Salt Polymers

Quaternary ammonium salt polymers, a kind of polyelectrolyte with a quaternary ammonium group, are widely used in traditional and emerging industries due to their good water-solubility, adjustable cationicity and molecular weight, high efficiency and nontoxicity. In this paper, firstly, the properties and several synthesis methods of typical quaternary ammonium salt monomers were introduced. Secondly, the research progress on the synthesis of polymers was summarized from the perspective of obtaining products with high molecular weight, narrow molecular weight distribution and high monomer conversion, and special functional polymers. Thirdly, the relationships between the structures and properties of the polymer were analyzed from the perspectives of molecular weight, charge density, structural stability, and microstructural regulation of the polymer chain unit. Fourthly, typical examples of quaternary ammonium salt polymers in the application fields of water treatment, daily chemicals, petroleum exploitation, papermaking, and textile printing and dyeing were listed. Finally, constructive suggestions were put forward on developing quaternary ammonium salt polymers with high molecular weights, strengthening the research on the relationships between the structures and their properties and pinpointing relevant application fields