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  • Research article
  • Open Access
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Ginkgolide B inhibits platelet and monocyte adhesion in TNFα-treated HUVECs under laminar shear stress

Contributed equally
BMC Complementary and Alternative Medicine201818:220

https://doi.org/10.1186/s12906-018-2284-8

  • Received: 21 March 2018
  • Accepted: 11 July 2018
  • Published:
Open Peer Review reports

Abstract

Background

Endothelial cells are sensitive to changes in both blood components and mechanical stimuli. Endothelial cells may undergo phenotypic changes, such as changes in adhesion protein expression, under different shear stress conditions. Such changes may impact platelet and monocyte adhesion to endothelial cells. This phenomenon is linked to chronic vascular inflammation and the development of atherosclerosis. In the present study, we investigated the effects of ginkgolide B on platelet and monocyte adhesion to human umbilical vein endothelial cells (HUVECs) under different conditions of laminar shear stress.

Methods

Platelet and monocyte adhesion to endothelial cells was determined by the Bioflux 1000. HUVECs were incubated with ginkgolide B or aspirin for 12 h, and then TNFα was added for 2 h to induce the inflammatory response under conditions of 1 and 9 dyn/cm2 laminar shear stress. The protein expression was analyzed by Western blot.

Results

The number of platelets that adhered was greater under conditions of 1 dyn/cm2 than under conditions of 9 dyn/cm2 of laminar shear stress (74.8 ± 19.2 and 59.5 ± 15.1, respectively). Ginkgolide B reduced the tumor necrosis factor α (TNFα)-induced increase in platelet and monocyte adhesion to HUVECs at 1 and 9 dyn/cm2 of laminar shear stress. In TNFα-treated HUVECs, the number of monocytes that adhered was greater under conditions of 1 dyn/cm2 of laminar shear stress compared with 9 dyn/cm2 (29.1 ± 4.9 and 22.7 ± 3.7, respectively). Ginkgolide B inhibited the TNFα-induced expression of vascular cell adhesion molecule-1(VCAM-1), VE-cadherin, and Cx43 in HUVECs at 1 and 9 dyn/cm2. The expression of these proteins was not different between 1 and 9 dyn/cm2.

Conclusions

Ginkgolide B suppressed platelet and monocyte adhesion under different conditions of laminar shear stress. Moreover, ginkgolide B reduced VCAM-1, VE-cadherin and Cx43 expression in TNFα-treated HUVECs under laminar shear stress. This suggested that ginkgolide B might shed light on the treatment of inflammation in atherosclerosis.

Keywords

  • Ginkgolide B
  • Endothelial cells
  • Platelets
  • VCAM-1
  • VE-cadherin
  • Shear stress

Background

Endothelial cells are the initial barriers on the vessel wall. Endothelial cell dysfunction is a primary cause of cardiovascular disease. Endothelial cells are sensitive to changes in both blood components and mechanical stimuli [13]. In humans, normal physiological flow ranges from 10 to 50 dyn/cm2 and is highly pulsatile in arteries and ~ 10-fold less with minimal pulsation in veins [4]. High shear stress that results from laminar flow promotes endothelial cell survival and quiescence, alignment in the direction of the flow, and the secretion of substances that promote vasodilation and anticoagulation [5, 6]. Low shear stress or changes in the direction of shear stress, as found in turbulent flow, promotes vasoconstriction, coagulation, and platelet aggregation [7, 8]. The precise mechanisms by which endothelial cells sense shear stress to affect cell-cell interactions are still not completely understood. Endothelial cells may undergo phenotypic changes, such as changes in adhesion protein expression, under different shear stress conditions. Such changes may impact platelet and monocyte adhesion to endothelial cells. This phenomenon is linked to chronic vascular inflammation and the development of atherosclerosis [911].

In the physiological state, endothelial cells maintain vessel integrity through junction proteins. VE-cadherin is a calcium-dependent cell-cell adhesion protein that is composed of five extracellular cadherin repeats and a transmembrane region [12]. VE-cadherin antibodies were shown to increase monolayer permeability in cultured cells [13]. We recently reported that VE-cadherin was involved in monocyte translocation in oxidized low-density lipoprotein (ox-LDL)-treated endothelial cells. Treatment with VE-cadherin siRNA reduced monocyte translocation in ox-LDL-treated endothelial cells [14]. Cx43 is also a junction protein that belongs to the gap junction protein family. Gap junctions play a role in intercellular communication between cells to regulate cell death, proliferation, and differentiation [15]. Gap junctions are involved in connecting adjacent cells to permit the exchange of low-molecular-weight molecules, such as ions and secondary messengers, to maintain homeostasis [16, 17].

Tumor necrosis factor α (TNFα) is a member of the cytokine family and involved in various inflammatory processes. TNFα is implicated in several human diseases, including atherosclerosis and cardiovascular disease. It is a potent inducer of nuclear factor κB (NF-кB) signaling, which is involved in the transcription of many inflammatory proteins. TNFα is also involved in endothelial cell injury under pathological conditions, such as atherosclerosis [18].

Ginkgolide B is a Ginkgo biloba leaf extract that can completely bind platelet-activating factor receptor (PAFR) and inhibit platelet activation [19]. Our recent studies showed that ginkgolide B inhibited inflammatory protein expression that was induced by ox-LDL in human umbilical vein endothelial cells (HUVECs) [20, 21]. However, remaining unknown is whether ginkgolide B inhibits platelet and monocyte adhesion to endothelial cells under different shear stress conditions. In the present study, we investigated the effects of ginkgolide B on platelet and monocytes adhesion to endothelial cells and phenotypic changes under different laminar shear stress conditions.

Methods

Materials

Ginkgolide B (95% purity) was purchased from Daguanyuan Company (Xuzhou, Jiangsu, China). Mouse tail type I collagen were purchased from Sigma-Aldrich (St. Louis, MO, USA). Monoclonal anti-connexin 43 antibody, polyclonal anti-VCAM-1 antibody and monoclonal anti-actin antibody were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Monoclonal anti-VE-cadherin antibody was purchased from Abcam (Boston, MA, USA). Bioflux 48-well plates (1–20 dyn/cm− 2; 910–0047) were purchased from Fluxion Biosciences (South San Francisco, CA, USA).

Preparation of platelets

Fresh citrate anti-coagulated venous blood was obtained from human donors who had not taken any medication for a minimum of 2 weeks before blood collection. The blood was centrifuged at 400 × g for 15 min to obtain platelet-rich plasma (PRP). The PRP was washed twice in Tyrode’s/HEPES buffer with 2 mM ethylene glycol tetraacetic acid (EGTA). Platelets were suspended in Tyrode’s/HEPES buffer at a concentration of 2 × 108 cells/ml [22].

Preparation of monocytes

The THP-1 human monocytic cell line was obtained from the American Type Culture Collection (Manassas, VA, USA). The cells were maintained at 37 °C in RPMI 1640 medium supplemented with 10% fetal calf serum (FCS), 100 IU penicillin, 100 μg/ml streptomycin, and 2 mM L-glutamine in a humidified 5% carbon dioxide atmosphere [23].

Preparation and culture of human umbilical vein endothelial cells

HUVECs were purchased from ScienCell Research Laboratories (Carlsbad, CA, USA). HUVECs were redissolved in a 37 °C constant temperature water bath. The cells were cultured in M199 medium that contained 10% fetal bovine serum (Gibco, NY, USA), 2 mM glutamine, 100 U/ml penicillin, 100 μg/ml streptomycin, and 20 ng/ml endothelial growth factor (R&D, Minneapolis, MN, USA) in an incubator at 37 °C and 5% CO2. Cells up to passage 4 were used in the experiments [24].

Platelet and monocyte adhesion to HUVECs assay

The Bioflux 1000 system (Fluxion Biosciences Inc., CA, USA) was used in the present study. The flow experiments were performed as previously described [25]. Briefly, the microfluidic channel was coated with type I collagen (20 μg/ml) that was dissolved in 0.02 M acetic acid. HUVECs (3 × 107) were seeded in microfluidic well plates until > 90% of the cells covered the well plate that was used for the experiments. HUVECs were treated with ginkgolide B (0.6 mg/ml) or aspirin (1 mM) for 12 h, and then TNFα was added for another 2 h under conditions of 1 and 9 dyn/cm2 laminar shear stress. The platelet suspension (300 μl, 2 × 108) or monocyte suspension (300 μl, 5 × 106) was added to the input channel for 45 min for cell adhesion under condition of 0.2 dyn/cm2. Images were captured by a Nikon Ti100 CCD camera in five locations in the flow chamber and analyzed using Bioflux Montage software.

Western blot

HUVECs were incubated with ginkgolide B (0.6 mg/ml) or aspirin (1 mM) for 12 h, and then TNFα (20 ng/ml) was added for 2 h under conditions of 1 and 9 dyn/cm2 laminar shear stress. The platelet or monocyte suspension was perfused for 45 min under condition of 0.2 dyn/cm2 for adhesion. After that the microfluidic channel was washed with PBS to remove unadhered platelets or monocytes for 5 min under condition of 2 dyn/cm2, and then lysis buffer (1% Triton X-100, 100 mM Tris/HCl [pH 7.2], 50 mM NaCl, 5 mM ethylenediaminetetraacetic acid [EDTA], 5 mM EGTA, 1 μM phenylmethylsulfonyl fluoride [PMSF], and 100 μg/ml leupeptin) was added. Lysates were centrifuged at 12000 × g at 4 °C for 5 min. To obtain sufficient protein, three parallel microfluidic flow channels were used in each group. The cell lysates were separated by 10% sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE), and transferred to a polyvinylidene difluoride membrane (Millipore, Billerica, MA, USA. Primary antibody incubations were performed overnight at 4 °C. Horseradish peroxidase-conjugated secondary antibody was applied for 1 h at room temperature and developed using Super Signal developing reagent (Pierce, Thermo Scientific). Blot densitometry was then performed, and the bands were analyzed using the Gene Genius Bio Imaging System.

Statistical analysis

Quantitative data are presented as mean ± SEM. Significant differences between two groups were analyzed by two-tail unpaired Student’s t-test. All of the calculations were performed using SPSS 18.0 software (Armonk, NY, USA). Values of p < 0.05 were considered statistically significant.

Results

Ginkgolide B decreases platelet adhesion to TNFα-treated HUVECs under conditions of laminar shear stress

To investigate the interaction between platelets and HUVECs, platelet adhesion was determined in TNFα-treated HUVECs under conditions of 1 and 9 dyn/cm2 of shear stress. Our previous studies showed that 0.6 mg/ml ginkgolide B and 1 mM aspirin significantly inhibited the inflammatory response of endothelial cells. Therefore, we applied the same doses of ginkgolide B and aspirin in the present study. As shown in Fig. 1, at 1 dyn/cm2, the number of platelets that adhered was 74.8 ± 19.2 and 36.1 ± 4.4 in TNFα-treated and -untreated HUVECs, respectively. The number of platelets that adhered was 39.5 ± 12.3 in ginkgolide B-treated HUVECs. We used aspirin (1 mM) as a control. Aspirin treatment also reduced the number of platelets that adhered (35.8 ± 10.1) in TNFα-treated HUVECs. At 9 dyn/cm2, the number of platelets that adhered was 59.5 ± 15.1 and 28.8 ± 3.7 in TNFα-treated and -untreated HUVECs. The number of platelets that adhered was 31.5 ± 4.9 and 32.1 ± 4.3 in the ginkgolide B- and aspirin-treated groups, respectively.
Fig. 1
Fig. 1

Ginkgolide B reduced platelet adhesion in TNFα-treated HUVECs under different conditions of laminar shear stress. HUVECs were incubated with ginkgolide B (0.6 mg/ml) or aspirin (1 mM) for 12 h, and then TNFα (20 ng/ml) was added for another 2 h. The platelet suspension was then added to the input well under conditions of 1 and 9 dyn/cm2 of laminar shear stress for 45 min. The data were obtained from three independent experiments. a Ginkgolide B decreased the number of platelets that adhered to TNFα-treated HUVECs at 1 and 9 dyn/cm2. b Platelet adhesion to HUVECs. ##p < 0.01, significant difference between TNFα-treated and -untreated HUVECs. **p < 0.01, significant difference between TNFα-treated cells and ginkgolide B- and aspirin-treated cells

Ginkgolide B suppresses VCAM-1, VE-cadherin, and Cx43 expression in TNFα-treated HUVECs that present platelet adhesion under conditions of laminar shear stress

We first investigated the effects of ginkgolide B on VCAM-1 expression under laminar shear stress. As shown in Fig. 2a, at 1 dyn/cm2, VCAM-1 expression increased by 31.7% ± 2.9% in TNFα-treated HUVECs compared with the control. Ginkgolide B and aspirin completely inhibited TNFα-induced VCAM-1 expression. VCAM-1 expression increased by 46.9% ± 10.1% in TNFα-treated HUVECs under conditions of 9 dyn/cm2 of laminar shear stress. Ginkgolide B (0.6 mg/ml) almost completely attenuated TNFα-induced VCAM-1 expression. Similar results were found in the aspirin-treated group.
Fig. 2
Fig. 2

Ginkgolide B suppressed VCAM-1, VE-cadherin, and Cx43 expression in TNFα-treated HUVECs that presented platelet adhesion under different conditions of laminar shear stress. HUVECs were incubated with ginkgolide B (0.6 mg/ml) or aspirin (1 mM) for 12 h, and then TNFα (20 ng/ml) was added for another 2 h. The platelet suspension was then added to the input well at 1 and 9 dyn/cm2 of laminar shear stress for 45 min. Unadhered platelets were removed by the addition of PBS buffer for 2 min. Lysates were then collected by the addition of lysis buffer. Protein expression was analyzed by Western blot. The data were obtained from three independent experiments. a Ginkgolide B inhibited TNFα-induced VCAM-1 expression at 1 and 9 dyn/cm2. b Ginkgolide B decreased TNFα-induced VE-cadherin expression at 1 and 9 dyn/cm2. c Ginkgolide B reduced TNFα-induced Cx43 expression at 1 and 9 dyn/cm2. #p < 0.05, significant difference between TNFα-treated and -untreated HUVECs. **p < 0.01, significant difference between TNFα-treated cells and ginkgolide B- and aspirin-treated cells

We next evaluated the effects of ginkgolide B on the expression of the tight junction proteins VE-cadherin and Cx43 under different conditions of laminar shear stress. As shown in Fig. 2b and c, TNFα treatment increased VE-cadherin expression by 31.8% ± 2.2% and 38.9 ± 9.1% under conditions of 1 and 9 dyn/cm2 of laminar shear stress, respectively. Ginkgolide B (0.6 mg/ml) and aspirin (1 mM) completely abolished TNFα-induced VE-cadherin expression at 1 and 9 dyn/cm2. Cx43 expression increased by 37.7% ± 9.8 and 30.5% ± 6.1% in TNFα-treated HUVECs that presented platelet adhesion at 1 and 9 dyn/cm2, respectively. Ginkgolide B (0.6 mg/ml) and aspirin (1 mM) significantly inhibited TNFα-induced Cx43 expression at both 1 and 9 dyn/cm2.

Ginkgolide B decreases monocyte adhesion to TNFα-treated HUVECs under conditions of laminar shear stress

Furthermore, monocyte adhesion to endothelial cells was evaluated under conditions of laminar shear stress. As shown in Fig. 3a, at 1 dyn/cm2, the number of monocytes that adhered was 29.1 ± 4.9 in TNFα-treated HUVECs. The number of monocytes that adhered was 11.3 ± 1.9 in the control group. At 9 dyn/cm2, the number of monocytes that adhered was 22.7 ± 3.7 in TNFα-treated HUVECs. The number of monocytes that adhered was 14.3 ± 4.5 in the control group. Ginkgolide B (0.6 mg/ml) inhibited monocyte adhesion to TNFα-treated HUVECs. The number of monocytes that adhered was 17.0 ± 2.8 at 1 dyn/cm2 and 15.7 ± 4.8 at 9 dyn/cm2. In the aspirin-treated group, the number of monocytes that adhered was 20.7 ± 1.8 and 14.7 ± 5.4 at 1 and 9 dyn/cm2, respectively. Significant differences were found between TNFα-treated cells and ginkgolide B- and aspirin-treated cells.
Fig. 3
Fig. 3

Ginkgolide B reduced monocyte adhesion to TNFα-treated HUVECs under different conditions of laminar shear stress. HUVECs were incubated with ginkgolide B (0.6 mg/ml) or aspirin (1 mM) for 12 h, and then TNFα (20 ng/ml) was added for another 2 h. The monocyte suspension was then added to the input well at 1 and 9 dyn/cm2 of laminar shear stress for 45 min. The data were obtained from three independent experiments. a Ginkgolide B reduced TNFα-induced monocyte adhesion to HUVECs at 1 and 9 dyn/cm2. b Monocyte adhesion to HUVECs. #p < 0.05, significant difference between TNFα-treated and -untreated HUVECs. *p < 0.01, significant difference between TNFα-treated cells and ginkgolide B- and aspirin-treated cells

Ginkgolide B suppresses VCAM-1, VE-cadherin, and Cx43 expression in TNFα-treated HUVECs that present monocyte adhesion under conditions of laminar shear stress

We also evaluated the effects of ginkgolide B on adhesion proteins and tight junction proteins under conditions of laminar shear stress and monocyte adhesion in HUVECs. As shown in Fig. 4a-c, VCAM-1 expression increased by 29.8% ± 2.9% at 1 dyn/cm2 and by 48.6% ± 10.1% at 9 dyn/cm2 in TNFα-treated HUVECs. Ginkgolide B inhibited TNFα-induced VCAM-1 expression at both 1 and 9 dyn/cm2. Similar results were found in aspirin-treated HUVECs.
Fig. 4
Fig. 4

Ginkgolide B suppressed VCAM-1, VE-cadherin, and Cx43 expression in TNFα-treated HUVECs that presented monocyte adhesion under different conditions of laminar shear stress. HUVECs were incubated with ginkgolide B (0.6 mg/ml) or aspirin (1 mM) for 12 h, and then TNFα (20 ng/ml) was added for another 2 h. The monocyte suspension was then added to the input well at 1 and 9 dyn/cm2 of laminar shear stress for 45 min. Unadhered monocytes were removed by the addition of PBS buffer for 2 min. Lysates were then collected by the addition of lysis buffer. Protein expression was analyzed by Western blot. a Ginkgolide B inhibited TNFα-induced VCAM-1 expression at 1 and 9 dyn/cm2. b Ginkgolide B decreased TNFα-induced VE-cadherin expression at 1 and 9 dyn/cm2. c Ginkgolide B reduced TNFα-induced Cx43 expression at 1 and 9 dyn/cm2. #p < 0.05, significant difference between TNFα-treated and -untreated HUVECs. *p < 0.05, **p < 0.01, significant difference between TNFα-treated cells and ginkgolide B- and aspirin-treated cells

TNFα treatment increased VE-cadherin expression by 36.9% ± 6.6 and 28.2% ± 6.0% at 1 and 9 dyn/cm2, respectively. Ginkgolide B and aspirin reduced TNFα-induced VE-cadherin expression at 1 and 9 dyn/cm2. Cx43 expression increased by 20.9% ± 7.1 and 21.8% ± 5.4% at 1 and 9 dyn/cm2 in TNFα-treated HUVECs, respectively. Both ginkgolide B and aspirin completely suppressed TNFα-induced Cx43 expression at 1 and 9 dyn/cm2.

Discussion

Platelets, monocytes, and endothelial cells interact in pathological states. Platelets release several inflammatory mediators, such as platelet factor 4 (PF4), regulated on activation, normal T-cell expressed and secreted chemokine (RANTES), P-selectin, and CD40 ligand, to recruit monocytes to the site of damaged endothelial cells [2628]. The adhesion of platelets through the expression of adhesion proteins can also elicit an inflammatory response in endothelial cells [29]. Monocyte adhesion to endothelial cells and enter intima where phagocytosis lipids and transform into macrophage and foam cells [3032]. This process is involved in plaque formation in atherosclerosis. Therefore, the inhibition of platelet and monocyte adhesion to endothelial cells might be a strategy for preventing atherosclerosis.

In the present study, we used the Bioflux 1000 microfluidic device to investigate platelet and monocyte adhesion to endothelial cells under different conditions of laminar shear stress. High shear stress in the laminar flow promotes endothelial cell survival and anticoagulation. Low shear stress in the laminar flow promotes endothelial cell proliferation, apoptosis, and coagulation [33]. However, the precise mechanisms by which these processes occur are still unknown. To determine the effects of different conditions of shear stress on the interaction between platelets/monocytes and endothelial cells, two conditions of shear stress were used (1 and 9 dyn/cm2). Laminar shear stress at 1 dyn/cm2 reflects a pathological state, and 9 dyn/cm2 reflects a physiological state [34]. Areas of low shear stress in vessels may be linked to endothelial cell dysfunction, reflected by lower nitric oxide and prostacyclin production. The present results showed that the number of platelets that adhered to HUVECs at 1 dyn/cm2 was greater than the number that adhered at 9 dyn/cm2. Monocyte adhesion presented a similar trend. The number of monocytes that adhered to HUVECs at 1 dyn/cm2 was greater than the number that adhered at 9 dyn/cm2. These results support the hypothesis that platelets and monocytes under conditions of low shear stress easily adhere to endothelial cells. We also observed phenotypic changes in endothelial cells under conditions of low and high shear stress. The expression of VCAM-1, VE-cadherin, and Cx43 was not different between 1 and 9 dyn/cm2. This implies that 9 dyn/cm2 of laminar shear stress does not impact TNFα-induced VCAM-1, VE-cadherin, or Cx43 expression. Both ginkgolide B and aspirin exerted protective actions against the expression of these proteins that was induced by TNFα under conditions of laminar shear stress. This is consistent with our previous study, in which increases in VE-cadherin and Cx43 expression were linked to monocyte migration that was induced by ox-LDL. Furthermore, the knockdown of VE-cadherin and Cx43 gene expression by siRNA decreased the number of monocytes that migrated in ox-LDL-treated HUVECs [14]. This suggests VE-cadherin and Cx43 mediate monocyte migration, and this phenomenon might occur independently of their function at cell junctions. In the present study, platelet and monocyte adhesion decreased under conditions of high shear stress, but the underlying mechanism needs clarification.

Growing evidence has shown the protective action of ginkgolide B on cardiovascular and nervous system diseases. Our previous studies showed that ginkgolide B inhibited platelet aggregation and reduced CD40L, RANTES and PF4 secretion induced by thrombin and collagen. Moreover, ginkgolide B treatment decreased platelet adhesion on aortic plaque in Apo E gene defective mice [20]. Recent a study reported that ginkgolide B promoted microglia/macrophage transferring from inflammatory M1 phenotype to anti-inflammatory phenotype M2 in vivo and in vitro [35]. In recent years Ginkgo biloba extracts have been widely used as a phytomedicine in Europe and in the United States. These studies demonstrated that ginkgolide B might be a promising drug in clinic application.

Conclusion

In conclusion, we found that platelet and monocyte adhesion was stronger under conditions of 1 dyn/cm2 of laminar shear stress compared with 9 dyn/cm2. Ginkgolide B inhibited TNFα-induced platelet and monocyte adhesion to endothelial cells and attenuated VCAM-1, VE-cadherin, and Cx43 expression under conditions of laminar shear stress. No differences in the expression of these proteins were found between 1 and 9 dyn/cm2. These findings suggest that ginkgolide B might shed light on the treatment of inflammation in atherosclerosis.

Notes

Abbreviations

NF-кB: 

Nuclear factor κB

TNFα: 

Tumor necrosis factor α

EDTA: 

Ethylenediaminetetraacetic acid

EGTA: 

Ethylene glycol tetraacetic acid

HUVECs: 

Human umbilical vein endothelial cells

ox-LDL: 

Oxidized low-density lipoprotein

PMSF: 

Phenylmethylsulfonyl fluoride

RANTES: 

Regulated on activation, normal T-cell expressed and secreted chemokine

VCAM-1: 

Vascular cell adhesion molecule-1

Declarations

Acknowledgements

We thank Dr. Yun You and Dr. Fulong Liao provides Bioflux 1000 facility for the present study. We would like to thank all the participants in the present study. We also thank the funding NSFC for the financial support.

Funding

This study was funded by National Natural Science Foundation of China (NSFC) (grant numbers 80471051, 81270379, 81070231, and 91649110). The funding agency had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Availability of data and materials

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request. All data generated or analyzed during this study are included in this published article.

Authors’ contributions

Conceived and designed the experiments: RMQ and YY. Performed the experiments: JS, MZ, YYZ, BDC, and HG. Analyzed the data: MZ and JS. Wrote the paper: RMQ. All author reviewed and approved the manuscript.

Ethics approval and consent to participate

Blood was collected from healthy donors who provided written informed consent. The experiments were conducted according to the principles of the Declaration of Helsinki. This study was approved by the Ethics Committee of the Beijing Institute of Geriatrics (no. 2015026).

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

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Authors’ Affiliations

(1)
MOH Key Laboratory of Geriatrics, Beijing Hospital, National Center of Gerontology, Beijing, China
(2)
Graduate School of Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China
(3)
Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, China

References

  1. Nakajima H, Yamamoto K, Agarwala S, Terai K, Fukui H, Fukuhara S, et al. Flow-dependent endothelial YAP regulation contributes to vessel maintenance. Dev Cell 2017;40:523–536.Google Scholar
  2. Urschel K, Garlichs CD, Daniel WG, Cicha I. VEGFR2 signalling contributes to increased endothelial susceptibility to TNF-alpha under chronic non-uniform shear stress. Atherosclerosis. 2011;219:499–509.View ArticlePubMedGoogle Scholar
  3. Johnson BD, Mather KJ, Wallace JP. Mechanotransduction of shear in the endothelium: basic studies and clinical implications. Vasc Med. 2011;16:365–77.View ArticlePubMedGoogle Scholar
  4. Paszkowiak JJ, Dardik A. Arterial wall shear stress: observations from the bench to the bedside. Vasc Endovasc Surg. 2003;37:47–57.View ArticleGoogle Scholar
  5. Schuler D, Sansone R, Freudenberger T, Rodriguez-Mateos A, Weber G, Momma TY, et al. Measurement of endothelium-dependent vasodilation in mice--brief report. Arterioscler Thromb Vasc Biol. 2014;34:2651–2657.Google Scholar
  6. Inaba H, Takeshita K, Uchida Y, Hayashi M, Okumura T, Hirashiki A, et al. Recovery of flow-mediated vasodilatation after repetitive measurements is involved in early vascular impairment: comparison with indices of vascular tone. PLoS One 2014;9:e83977.Google Scholar
  7. Verbeke FH, Pannier B, Guérin AP, Boutouyrie P, Laurent S, London GM. Flow-mediated vasodilation in end-stage renal disease. Clin J Am Soc Nephrol. 2011;6:2009–15.View ArticlePubMedPubMed CentralGoogle Scholar
  8. Okorie UM, Denney WS, Chatterjee MS, Neeves KB, Diamond SL. Determination of surface tissue factor thresholds that trigger coagulation at venous and arterial shear rates: amplification of 100 fM circulating tissue factor requires flow. Blood. 2008;111:3507–13.View ArticlePubMedPubMed CentralGoogle Scholar
  9. Ahmadsei M, Lievens D, Weber C, Von HP, Gerdes N. Immune-mediated and lipid-mediated platelet function in atherosclerosis. Curr Opin Lipidol. 2015;26:438–48.View ArticlePubMedGoogle Scholar
  10. Soehnlein O. Monocytes chat with atherosclerotic lesions. Arterioscler Thromb Vasc Biol. 2016;36:1720–1.View ArticlePubMedGoogle Scholar
  11. Badimon L, Suades R, Fuentes E, Palomo I, Padró T. Role of platelet-derived microvesicles as crosstalk mediators in Atherothrombosis and future pharmacology targets: a link between inflammation, atherosclerosis, and thrombosis. Front Pharmacol. 2016;7:293.View ArticlePubMedPubMed CentralGoogle Scholar
  12. Gavard J. Endothelial permeability and VE-cadherin: a wacky comradeship. Cell Adhes Migr. 2013;7:455–61.View ArticleGoogle Scholar
  13. Corada M, Liao F, Lindgren M, Lampugnani MG, Breviario F, Frank R, et al. Monoclonal antibodies directed to different regions of vascular endothelial cadherin extracellular domain affect adhesion and clustering of the protein and modulate endothelial permeability. Blood 2001;97:1679–1684.Google Scholar
  14. Liu X, Sun W, Zhao Y, Chen B, Wu W, Li B, et al. Ginkgolide B inhibits JAM-A, Cx43, and VE-cadherin expression and reduces monocyte transmigration in oxidized LDL-stimulated human umbilical vein endothelial cells. Oxidative Med Cell Longev 2015;2015:907926.Google Scholar
  15. Cheng JC, Chang HM, Fang L, Sun YP, Leung PC. TGF-β1 up-regulates connexin43 expression: a potential mechanism for human trophoblast cell differentiation. J Cell Physiol. 2015;230:1558–66.View ArticlePubMedGoogle Scholar
  16. Laird DW. Syndromic and non-syndromic disease-linked Cx43 mutations. FEBS Lett. 2014;588:1339–48.View ArticlePubMedGoogle Scholar
  17. Ghosh S, Kumar A, Chandna S. Connexin-43 downregulation in G2/M phase enriched tumour cells causes extensive low-dose hyper-radiosensitivity (HRS) associated with mitochondrial apoptotic events. Cancer Lett. 2015;363:46–59.View ArticlePubMedGoogle Scholar
  18. Chin-Feng H, Hsia-Fen H, Wei-Kung T, Thung-Lip L, Yu-Feng W, Kwan-Lih H, et al. Glossogyne tenuifolia extract inhibits TNF-α-induced expression of adhesion molecules in human umbilical vein endothelial cells via blocking the NF-kB signaling pathway. Molecules 2015;20:16908–16923.Google Scholar
  19. Hu L, Chen Z, Xie Y, Jiang Y, Zhen H. New products from alkali fusion of Ginkgolides a and B. J Asian Nat Prod Res. 2000;2:103–10.View ArticlePubMedGoogle Scholar
  20. Liu X, Zhao G, Yan Y, Bao L, Chen B, Qi R. Ginkgolide B reduces atherogenesis and vascular inflammation in ApoE(−/−) mice. PLoS One. 2012;7:e36237.View ArticlePubMedPubMed CentralGoogle Scholar
  21. Zhang S, Chen B, Wu W, Bao L, Qi R. Ginkgolide B reduces inflammatory protein expression in oxidized low-density lipoprotein-stimulated human vascular endothelial cells. J Cardiovasc Pharmacol. 2011;57:721–7.View ArticlePubMedGoogle Scholar
  22. Reiss C, Mindukshev I, Bischoff V, Subramanian H, Kehrer L, Friebe A, et al. The sGC stimulator riociguat inhibits platelet function in washed platelets but not in whole blood. Br J Pharmacol 2015; 172:5199–5210.Google Scholar
  23. Scipione CA, Sayegh SE, Romagnuolo R, Tsimikas S, Marcovina SM, Boffa MB, et al. Mechanistic insights into Lp(a)-induced IL-8 expression: a role for oxidized phospholipid modification of apo(a). J Lipid Res 2015;56:2273–2285.Google Scholar
  24. Qin WD, Mi SH, Li C, Wang GX, Zhang JN, Wang H, et al. Low shear stress induced HMGB1 translocation and release via PECAM-1/PARP-1 pathway to induce inflammation response. PLoS One 2015;10:e0120586.Google Scholar
  25. Sato M, Levesque MJ, Nerem RM. Micropipette aspiration of cultured bovine aortic endothelial cells exposed to shear stress. Arteriosclerosis. 1987;7:276–86.View ArticlePubMedGoogle Scholar
  26. Woller G, Brandt E, Mittelstädt J, Rybakowski C, Petersen F. Platelet factor 4/CXCL4-stimulated human monocytes induce apoptosis in endothelial cells by the release of oxygen radicals. J Leukoc Biol. 2008;83:936–45.View ArticlePubMedGoogle Scholar
  27. Von HP, Weber KS, Huo Y, Proudfoot AE, Nelson PJ, Ley K, et al. RANTES deposition by platelets triggers monocyte arrest on inflamed and atherosclerotic endothelium. Circulation 2001;103:1772–1777.Google Scholar
  28. Lievens D, Zernecke A, Seijkens T, Soehnlein O, Beckers L, Munnix IC, et al. Platelet CD40L mediates thrombotic and inflammatory processes in atherosclerosis. Blood 2010;116:4317–4327.Google Scholar
  29. Blann AD, Nadar SK, Lip GY. The adhesion molecule P-selectin and cardiovascular disease. Eur Heart J. 2003;24:2166–79.View ArticlePubMedGoogle Scholar
  30. Hilgendorf I, Swirski FK, Robbins CS. Monocyte fate in atherosclerosis. Arterioscler Thromb Vasc Biol. 2015;35:272–9.View ArticlePubMedGoogle Scholar
  31. Plotkin JD, Elias MG, Dellinger AL, Kepley CL. NF-κB inhibitors that prevent foam cell formation and atherosclerotic plaque accumulation. Nanomedicine. 2017;13:2037–48.View ArticlePubMedGoogle Scholar
  32. Fong GH. Potential contributions of intimal and plaque hypoxia to atherosclerosis. Curr Atheroscler Rep. 2015;17:1–10.View ArticleGoogle Scholar
  33. Cunningham KS, Gotlieb AI. The role of shear stress in the pathogenesis of atherosclerosis. Lab Investig. 2005;85:9–23.View ArticlePubMedGoogle Scholar
  34. Stepp DW, Nishikawa Y, Chilian WM. Regulation of shear stress in the canine coronary microcirculation. Circulation. 1999;100:1555–61.View ArticlePubMedGoogle Scholar
  35. Shu ZM, Shu XD, Li HQ, Sun Y, Shan H, Sun XY, et al: Ginkgolide B protects against ischemic stroke via modulating microglia polarization in mice. CNS Neurosci Ther 2016;22:729–739.Google Scholar

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