Induction of CXC chemokines in human mesenchymal stemcells by stimulation with secreted frizzled-related proteinsthrough non-canonical Wnt signaling

• 作者: David S Bischoff, Jian-Hua Zhu, Nalini S Makhijani Dean T Yamaguchi
• 来源期刊: 世界干细胞杂志：英文版（电子版）
• 学科: 医学
• 关键词: CXC CHEMOKINES MESENCHYMAL stem cell Osteogenesis Differentiation Wnt signaling pathway Frizzled-related protein FRIZZLED receptors
• 年，卷（期）: 2015,（11）
• 页码范围: 1262-1273
• 页数: 12页
• 分类号: R

摘要

AIM To investigate the effect of secreted frizzledrelatedproteins （sFRPs） on CXC chemokine expressionin human mesenchymal stem cells （hMSCs）.METHODS： CXC chemokines such as CXCL5 andCXCL8 are induced in hMSCs during differentiation withosteogenic differentiation medium （OGM） and may beinvolved in angiogenic stimulation during bone repair.hMSCs were treated with conditioned medium （CM） fromL-cells expressing non-canonical Wnt5a protein, or withcontrol CM from wild type L-cells, or directly with sFRPsfor up to 10 d in culture. mRNA expression levels of bothCXCL5 and CXCL8 were quantitated by real-time reversetranscriptase-polymerase chain reaction and secretedprotein levels of these proteins determined by ELISA.Dose- （0-500 ng/mL） and time-response curves weregenerated for treatment with sFRP1. Signal transductionpathways were explored by western blot analysis withpan- or phosphorylation-specific antibodies, through useof specific pathway inhibitors, and through use of siRNAstargeting specific frizzled receptors （Fzd）-2 and 5 or the receptor tyrosine kinase-like orphan receptor-2 （RoR2）prior to treatment with sFRPs.RESULTS： CM from L-cells expressing Wnt5a, a noncanonicalWnt, stimulated an increase in CXCL5 mRNAexpression and protein secretion in comparison to controlL-cell CM. sFRP1, which should inhibit both canonicaland non-canonical Wnt signaling, surprisingly enhancedthe expression of CXCL5 at 7 and 10 d. Dickkopf1, aninhibitor of canonical Wnt signaling prevented the sFRPstimulatedinduction of CXCL5 and actually inhibitedbasal levels of CXCL5 expression at 7 but not at 10 d posttreatment. In addition, all four sFRPs isoforms inducedCXCL8 expression in a dose- and time-dependent mannerwith maximum expression at 7 d with treatment at 150ng/mL. The largest increases in CXCL5 expression wereseen from stimulation with sFRP1 or sFRP2. Analysis ofmitogen-activated protein kinase signaling pathwaysin the presence of OGM showed sFRP1-induced phosphorylationof extracellular signal-regula