Role of Vimentin in the Inhibitory Effects of Low-Molecular-Weight Heparin on PC-3M Cell Adhesion to, and Migration through, Endothelium

Abstract

Low-molecular-weight heparin (LMWH) has been used in cancer patients with venous thromboembolic complications, resulting in a higher survival rate and an inhibitory action on experimental metastasis. In the present study, human umbilical vein endothelial cells (HUVECs) were treated with LMWH for 24 h. We found that the resulting HUVECs could significantly inhibit the highly metastatic human prostate cancer cell line (PC-3M) in terms of its adhesion to the endothelium and migration across the endothelium, according to scanning electron microscopy. We also determined the elevated levels of endothelial intercellular Ca 2ϩ concentration after the adhesion of PC-3M cells to HUVECs was greatly reduced by incubation with LMWH. Using proteomics, we surveyed the global protein changes in HUVECs after LMWH treatment and identified four down-regulated proteins that were possible isoforms of cytoskeletal vimentin intermediate filaments, cartilage-derived Ctype lectin, and serine/threonine protein phosphatase 1␤ (PP-1B). LMWH affected the morphology of vimentin and the expression levels of ␣ v integrin and PP-1B in HUVECs bound to PC-3M cells. Vimentin assists in the adhesion of PC-3M cells, which was confirmed by short interfering RNA experiments. Furthermore, the direct binding of purified vimentin protein with LMWH was detected with surface plasmon resonance methods. However, when we used fluorescence-labeled heparin for 24 h to identify whether this binding occurred within cells, heparin was distributed principally around endothelial cells. Taken together, these findings suggest that the monoincubation of LMWH with HUVECs could inhibit PC-3M cell adhesion to, and migration through, endothelium. LMWH's regulation of vimentin plays a role in the antimetastatic action.

Enoxaparin Sensitizes Human Non–Small-Cell Lung Carcinomas to Gefitinib by Inhibiting DOCK1 Expression, Vimentin Phosphorylation, and Akt Activation

“…found that vimentin could be regulated by LMWH and played a critical role in the antiangiogenesis and antimetastasis effects of LMWH (Pan et al, 2011). In the present study, we found that vimentin activity was influenced in the group administered enoxaparin with gefitinib in A549 cells (Fig.…”

“…Our previous study showed that unfractionated heparin does not easily cross the cell membrane (Pan et al, 2011). Therefore, it is possible for enoxaparin to exert its function through interaction with proteins present on the cell surface, including DOCK1, which could be translocated across the plasma membrane (Kobayashi et al, 2001).…”

“…Vimentin, a major constituent of the cytoskeleton IF family of proteins, plays an integral role in the progression of lung cancers, including cell migration and invasion, EMT, regulation of major signal transduction pathways, the positioning and anchorage of organelles, such as mitochondria, and cell-cell and cell-substrate adhesion (Tzivion et al 2000;Mendez et al, 2010;Kidd et al, 2014). Our previous studies (Pan et al, 2011) have shown that vimentin intermediate filaments are an important factor in the antiadhesion and migration effects of LMWH in tumor cells that travel to the endothelium. In the present study, enoxaparin 1 gefitinib produce a better effective inhibitory effect on p-vimentin expression, while the expression of the vimentin protein showed no significant decrease after enoxaparin 1 gefitinib treatment.…”

“…Combinations of low doses of anticancer agents that have different mechanisms of action may have significantly improved efficacy in inhibiting tumor formation and tumor growth when compared with individual anticancer substances alone (Chen et al, 2012). Although low molecular weight heparins (LMWHs) (e.g., enoxaparin, dalteparin, reviparin, and tinzaparin) have been widely studied as antitumor drugs and it has been shown that LMWH has an antimetastatic effect on multiple neoplasms with improved animal survival rates (Mousa and Petersen, 2009;Pan et al, 2011;Walenga and Lyman, 2013), the anticancer effects of LMWH in combination with gefitinib in NSCLC A549 and H1975 cells as well as xenograft lung tumors in immune-deficient mice are not known. Gefitinib targets and inhibits EGFRtyrosine kinase, leading to constitutive inhibition of downstream signaling transduction pathways, including RAF-extracellular signal-related kinase 1/2 (Erk1/2) and phosphoinositide 3-kinase (PI3K)-Akt.…”

Roles of vimentin and 14-3-3 zeta/delta in the inhibitory effects of heparin on PC-3M cell proliferation and B16-F10-luc-G5 cells metastasis

“…Previously, based on the hypothesis that heparin regulates vimentin in the tumor-endothelium interaction and the finding of endothelial 2-DE experiment that vimentin expression altered greatly by low molecular weight heparin, we demonstrated that vimentin partly plays a role in the anti-metastatic effects of low molecular weight heparin [16] . In the present study, we further analyzed and confirmed the down-regulated expression of vimentin, which was also one of the proteins influenced by heparin and identified in both the PC-3M cells and lungs of mice burdened with B16-F10-luc-G5 cells, along with 14-3-3 zeta/delta.…”