Transcriptomic Changes in Osteoblasts Following Endothelial Cell‐Cocultivation Suggest a Role of Extracellular Matrix in Cellular Interaction

Abstract

Vascularization is important for bone development, fracture healing and engineering of artificial bone tissue. In the context of bone tissue engineering, it was shown that coimplantation of human primary umbilical vein endothelial cells (HUVECs) and human osteoblasts (hOBs) results in the formation of functional blood vessels and enhanced bone regeneration. Implanted endothelial cells do not only contribute to blood vessel formation, but also support proliferation, cell survival and osteogenic differentiation of coimplanted hOBs. These effects are partially mediated by direct heterotypic cell contacts. In a previous report we could show that cocultivated hOBs strongly increase the expression of genes involved in extracellular matrix (ECM) formation in HUVECs, suggesting that ECM may be involved in the intercellular communication between hOBs and HUVECs. The present study aimed at investigating whether comparable changes occur in hOBs. We therefore performed a microarray analysis of hOBs cultivated in direct contact with HUVECs, revealing 1,004 differentially expressed genes. The differentially expressed genes could be assigned to the functional clusters ECM, proliferation, apoptosis and osteogenic differentiation. The microarray data could be confirmed by performing quantitative real time RT-PCR on selected genes. Furthermore, we could show that the ECM produced by HUVECs increased the expression of the osteogenic differentiation marker alkaline phosphatase (ALP) in hOBs. In summary, our data demonstrate that HUVECs provoke complex changes in gene expression patterns in cocultivated hOBs and that ECM plays and important role in this interaction. J. Cell. Biochem. 117: 1869-1879, 2016. © 2016 Wiley Periodicals, Inc.

KLF2+ stemness maintains human mesenchymal stem cells in bone regeneration

“…Recently, coculture of MSCs with ECs has attracted a great deal of interest in the development of strategies to achieve vascularized bone. A number of coculture models of ECs with osteoblasts or human osteoprogenitors derived from MSCs have been reported, demonstrating enhanced osteogenesis or angiogenesis . There have been few studies regarding the roles and timing of angiogenesis and osteogenesis in coculture systems.…”

A novel method to improve the osteogenesis capacity of hUCMSCs with dual‐directional pre‐induction under screened co‐culture conditions

“…One is soluble cytokines secreted by endothelial cells, such as bone morphogenetic protein‐2 (BMP‐2), VEGF, PDGF acting on osteoblasts in the form of paracrine to promote their differentiation . Secondly, the extracellular matrix plays a pivotal role in the communication between two kinds of cells . Finally, multiple scholars believe that the direct contact between cells is the premise of endothelial cells to promote the functions of osteoblasts, and gap junctions are decisive .…”

Calmodulin Regulated Spectrin Associated Protein 1 mRNA is Directly Regulated by miR‐126 in Primary Human Osteoblasts

“…Genes involved in extracellular matrix formation and turnover were the above‐mentioned TNFAIP6 and tetranectin, as well as PLAUR [Furlan et al, ]. The work from several independent groups [Kang et al, ; Gandavarapu et al, ] defined the role of ECM in osteoblast differentiation; and work from our laboratory [Simunovic et al, ; Lampert et al, ] accentuated its importance in hOB/HUVEC interaction. Besides its role as mechanical support for the investing cells, it is known that ECM regulates cellular differentiation and behavior, mainly through storage and release of signaling molecules and growth factors [Hynes, ], and these properties make it an interesting field of research in hOB/HUVEC interaction.…”

“…The isolation of primary human osteoblasts (hOBs) was established in our lab by Wenger et al []. Since then, hOBs have been characterized by histology (such as von Kossa staining and Alizarin Red staining), osteogenic differentiation (such as alkaline phosphatase expression), and an osteogenic phenotype by microarray data [Stahl et al, ; Schmidt et al, ; Lampert et al, ]. Given that we utilized hOBs which were isolated and cultivated by a validated procedure, for the actual study, no further characterization of hOBs was performed.…”

“…The nature of this interaction is important for tissue engineering, as its manipulation might improve the survival of implanted bone and blood vessel cells. Direct cellular interaction through gap junctions, indirect communication via soluble factors, and communication via the extracellular matrix has been considered [Stahl et al, ; Sun et al, ; Guillotin et al, ; Grellier et al, ; Hager et al, ; Steiner et al, ; Simunovic et al, ; Lampert et al, ]. The role of miRNAs has not been extensively studied in this system.…”