DFMG reverses proliferation and migration of vascular smooth muscle cells induced by co-culture with injured vascular endothelial cells via suppression of the TLR4-mediated signaling pathway.

7-Difluoromethoxy-5,4'-dimethoxy-genistein (DFMG) is a novel chemical compound synthesized using genistein. Previous studies have indicated that DFMG can reverse the apoptosis of vascular endothelial cells (VECs) by regulating the mitochondrial apoptosis pathway. The present study aimed to investigate the activity and molecular mechanism underlying DFMG­mediated protection of vascular smooth muscle cell (VSMCs) using a non­contact co­culture model established by using Transwell insert. Secretion of interleukin­6 (IL­6) and tumor necrosis factor­α (TNF­α) were measured by ELISA. Proliferation and migration of VSMCs were assessed using a Cell Counting kit­8 and wound healing assays, respectively. Toll­like receptor 4 (TLR4) mRNA and protein levels were detected by reverse transcription-quantitative polymerase chain reaction and western blotting analyses, respectively. In the present study, lysophosphatidylcholine (LPC) significantly increased the secretion of IL­6 and TNF­α in VECs. VECs treated with LPC markedly increased proliferation and migration of VSMCs, which were inhibited by DFMG. Transfection of either TLR4 short hairpin RNA (shRNA) or TLR4 cDNA in VECs inhibited and increased proliferation and migration of VSMCs, respectively. Furthermore, transfection of VECs with TLR4 shRNA suppressed the proliferation and migration of VSMCs induced by co­culture with injured VECs, which was further enhanced by treatment with DFMG. By contrast, transfection of VECs with TLR4 cDNA enhanced proliferation and migration of VSMCs and this effect was inhibited by treatment with DFMG. Taken together, the results of the present study demonstrated that DFMG can reverse proliferation and migration of VSMCs induced by co­culture with injured VECs via suppression of the TLR4­mediated signaling pathway.

Application of cell co-culture techniques in central nervous system diseases / 药学学报

The study of central nervous system disease is dependent on in vitro culture of neuronal cells. However, it is hard to determine the interaction between cells in culture of single type of neuronal cells. The co-culture system is able to mimic the cell-cell interaction in the brain and to facilitate investigation into the interaction between different types of cells, as well as cell-environment interaction. The co-culture of neurocytes is more and more popular in the disease study of central nervous system in vitro, such as stroke, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, etc. Neurovascular unit (NVU), which is composed of neurons, brain microvascular endothelial cells and astrocytes, can reflect the structure and function of brain in state of the art. Establishment of NVU in vitro is important in the study of the brain diseases. In this paper, several co-culture models of the central nervous system are reviewed in techniques for two-dimensional and three-dimensional culturing. Cell contact and non-contact methods are compared. Moreover, their application in the relevant research and the future direction are explored.