Mir-199a-3p Mediates Fluid Shear Stress-Induced Osteoblast Proliferation by Targeting CABLES-1 / 医用生物力学

Objective To explore the role of miR-199a-3p in osteoblast proliferation induced by fluid shear stress (FSS) and the potential molecular mechanism. Methods Osteoblast MC3T3-E1 was treated with 1. 2 Pa FSS with time gradients of 0, 15, 30, 45, 60, 75 and 90 min, respectively. MC3T3-E1 cells were transfected with miR-199a-3p mimic or miR-199a-3p inhibitor. MC3T3-E1 cells were transfected with miR-199a-3p mimic and itsnegative control and then treated with 1. 2 Pa FSS for 45 min. The pc DNA NC, pc DNA-CABLES -1, si RNA NC and si RNA CABLES-1 were transfected into MC3T3-E1 cells. The pc DNA-CABLES-1 and mir-199a-3p mimic and SI NA-cables-1 and miR-199a-3p inhibitor were co-transfected, respectively. Cell activity was detected by CCK-8 assay. Real-time quantitative PCR (RT-qPCR) was used to detect expression levels of CABLES-1, miR-199a-3p, CDK 6, Cyclin D1 and PCNA. Luciferase reporting assay was used to detect targeting relationship between CABLES-1 and miR-199a-3p. Immunofluorescence was used to detect protein expression of CABLES-1.Western blot was used to detect protein expression of CABLES-1, CDK 6, PCNA and Cyclin D1. Results Mir- 199a-3p in MC3T3-E1 cells was significantly down-regulated by FSS. Over-expressed miR-199a-3p inhibitedosteoblast proliferation, and down-regulated miR-199a-3p expression promoted osteoblast proliferation. miR-199a- 3p could reverse the FSS-induced proliferation in osteoblasts. Dual luciferase assay showed that miR-199a-3p targeted to CABLES-1 and over-expressed miR-199a-3p inhibited expression of CBALES-1 protein. CABLES-1 could promote proliferation of osteoblasts. miR-199a-3p inhibited osteoblast proliferation induced by FSS through CABLES-1. Conclusions FSS-induced osteoblast proliferation can be realized by down-regulated miR-199a-3p expression via targeting CABLES-1. The findings in this study provide new direction for researches on mechanism of FSS-induced osteoblast proliferation, as well as new ideas for future research on clinical application of mechanical loading in the treatment of bone and joint diseases.

CD44-HA Mediate Adhesion of MDA-MB-231 Cells and HL60 Cells under Fluid Shear Flow / 医用生物力学

Objective To explore how hyaluronic acid ( HA) in extracellular matrix regulates the adhesion ofCD44+tumor cells. Methods MDA-MB-231 cells or HL60 cells were perfused in a parallel plate chamber. Themovement of cells over immobilized HA was observed and analyzed to obtain the characteristics of cell adhesionand rolling. Results The adhesion number of MDA-MB-231 cells on HA substrate was positively regulated by HAconcentration, but not by HA molecular weight. Compared with physically adsorbed HA, immobilized HA byavidin-biotin could significantly improve the cell adhesion ratio. With the increase of shear stress in the range of30-50 mPa, the rolling velocity of cells increased and the adhesion ratio decreased, but the tether lifetime of cellswas not affected. In the same flow field, compared with MDA-MB-231 cells, HL60 cells with low expression ofCD44 rolled more quickly on immobilized HA, with shorter tether lifetime and much lower adhesion ratio(<1. 5% ). Conclusions Fluid shear stress might mediate the rolling velocity of MDA-MB-231 cells by regulatingthe CD44-HA association rate rather than their dissociation rate. The interaction between CD44 and HA is involved in the initial adhesion of HL60 cells, but it does not play a major role. This study will provide references for the design of anti-tumor drugs.

Biomechanical Environment Regulates Biomimetic Mineralization of Bone Matrix / 医用生物力学

Bone defects have always been an important cause of threat to human health, and artificial biomimetic bone repair replacement materials are currently one of the most effective and feasible solution approaches to treat bone damage. To develop artificial bone biomimetic materials, an in vitro biomimetic mineralization system must be constructed first to study in vitro biomimetic mineralization mechanism of natural bone matrix. Collagen is a template for mineralization, and its properties such as crosslinking degree, diameter, osmotic pressure, and surface charge can all directly affect mineralization progress. The biochemical and mechanical environments in which mineralization occurs are also quite distinct in their effects on mineralization process, particularly noncollagenous proteins and fluid shear stress (FSS). FSS is considered to be the main mechanical stimulation of bone tissues in micro-environment, which is of great significance to bone growth, repair and health maintenance. FSS at different levels and loading regimes has significant effects on transformation of amorphous calcium phosphate to bone apatite, self-assembly and directional alignment of collagen fibrils, and formation of hierarchical intrafibrillar mineralization. In this paper, the factors affecting collagen mineralization and their mechanism were summarized, with focus on regulation of FSS on collagen mineralization, and development direction in future was also prospected.

Chemokine CXCL12-Induced Calcium Response of Jurkat T Cells under Fluid Shear Stress / 医用生物力学

Objective To investigate the mechanism of mechano-chemical coregulation in chemokine-induced calcium response of Jurkat T cells under fluid shear stress (FSS). Methods By using parallel-plate flow chamber combined with fluorescence microscope, the calcium response of Jurkat T cells on CXCL12 was observed to extract the corresponding characteristic parameters under static or flow state, with or without extracellular Ca2+, respectively. Results Immobilized CXCL12 could induce firm adhesion of the circulating Jurkat T cells, and the arrested cells increased with the increase of CXCL12 concentration. Force could trigger the calcium response of Jurkat T cells and sharply raised the activation ratio from 4% up to 75% when the FSS increased from 0 to 20 mPa. Under 20 mPa FSS, extracellular Ca2+ could stimulate quickly the calcium response by shortening the delay time (about 23 s), and enhance calcium intensity by prolonging the climbing time (about 7 s) and half time (about 20 s). Conclusions The cooperation between FSS and extracellular Ca2+ would accelerate and enhance CXCL12-mediated-calcium response of Jurkat T cells, which indicated a fast mechanosensitive pathway through ‘extracellular calcium influx-intracellular calcium store release’. The research results would contribute to understanding the process of T cells activation and providing the clue for relevant pathological and drug research.

Fluid-solid coupling numerical simulation on ideal porous structure of rat alveolar bone / 生物医学工程学杂志

Fluid shear stress (FSS) caused by interstitial fluid flow within trabecular bone cavities under mechanical loading is the key factor of stimulating biological response of bone cells. Therefore, to investigate the FSS distribution within cancellous bone is important for understanding the transduction process of mechanical forces within alveolar bone and the regulatory mechanism at cell level during tooth development and orthodontics. In the present study, the orthodontic tooth movement experiment on rats was first performed. Finite element model of tooth-periodontal ligament-alveolar bone based on micro computed tomography (micro-CT) images was established and the strain field in alveolar bone was analyzed. An ideal model was constructed mimicking the porous structure of actual rat alveolar bone. Fluid flow in bone was predicted by using fluid-solid coupling numerical simulation. Dynamic occlusal loading with orthodontic tension loading or compression loading was applied on the ideal model. The results showed that FSS on the surface of the trabeculae along occlusal direction was higher than that along perpendicular to occlusal direction, and orthodontic force has little effect on FSS within alveolar bone. This study suggests that the orientation of occlusal loading can be changed clinically by adjusting the shape of occlusal surface, then FSS with different level could be produced on trabecular surface, which further activates the biological response of bone cells and finally regulates the remodeling of alveolar bone.

Numerical Simulation on Fluid Flow within Rat Alveolar Bone under Orthodontic and Occlusal Loading / 医用生物力学

Objective To study fluid flow within alveolar bone under orthodontic and occlusal loading, so as to provide references for understanding the regulatory mechanism of bone remodeling during orthodontics. Methods An animal model for orthodontic tooth movement on rats was first constructed. The finite element model of tooth-periodontal ligament-alveolar bone was established based on micro-CT images and the strain field in alveolar bone under orthodontic or constant occlusal loading was analyzed. Then finite element model of alveolar bone was constructed from the bone near the cervical margin or apical root of mesial root. The fluid flow in this model under orthodontic and cyclic occlusal loading was further predicted by using fluid-solid coupling numerical simulation. Results The fluid velocity within alveolar bone cavity mainly distributed at 0-10 μm/s, and the fluid shear stress (FSS) was mainly distributed at 0-10 Pa. FSS on the surface of alveolar bone near the apical root was higher than that close to the cervical margin. Conclusions FSS at different levels could be produced at different location within alveolar bone cavity under orthodontic and cyclic occlusal loading, which might further activate biological response of bone cells on the surface of trabeculae and finally regulate the remodeling of alveolar bone and orthodontic movement of tooth. The results provide theoretical guidance for the clinical treatment of orthodontics.

Effects of Fluid Shear Stress on PCP Signaling Pathway and Ciliogenesis in Vascular Endothelial Cells / 医用生物力学

Objective To investigate the effect of different fluid shear stress (FSS) on the regulation of planar cell polarity (PCP) signaling, and further to explore the relationship among FSS, PCP signaling pathway and ciliogenesis. Methods The hydrodynamic cell model of adjustable FSS was established. qPCR and immunofluorescence were used to detect the mRNA expression of PCP signaling pathway core protein Dvl2 and cilia assembly protein IFT88, cell targeting and co-localization under different FSS. Western blot (WB) was used to detect the protein expression of Dvl2 at 18 h under different FSS. Results The qPCR result showed that compared with 1.5 Pa FSS, under 0.1 Pa FSS, the mRNA expression of Dvl2 was higher at 6 h and 18 h (P＜0.05), significantly higher at 12 h (P＜0.01); the mRNA expression of IFT88 was significantly higher at 18 h (P＜0.01). The WB result showed that compared with 0 h, under 0.1 Pa FSS, the protein expression of Dvl2 was higher at 18 h (P＜0.05), significantly lower under 1.5 Pa FSS (P＜0.01); compared with 1.5 Pa FSS, the protein expression of Dvl2 was higher at 18 h under 0.1 Pa FSS (P＜0.05). The immunofluorescence result showed that the positive expression of Dvl2 increased with the loading time on FSS increasing, and gradually aggregated at a point around the nucleus; the positive expression of IFT88 was gradually transferred from the nucleus to the cytoplasm and aggregated at a point under 0.1 Pa FSS, and gradually decreased and depolymerized under 1.5 Pa FSS. Protein Dvl2 and IFT88 were located in the same position in cells under 0.1 Pa FSS and before 18 h under 1.5 Pa FSS, and colocalization of proteins Dvl2 and IFT88 was not observed after 18 h under 1.5 Pa FSS due to IFT88 depolymerization. Conclusions Laminar FSS played an inhibition on the transduction of PCP signaling pathway and a hindrance on the process of ciliogenesis, while low FSS played a promotion on the transduction. PCP signaling pathway might regulate FSS-induced ciliogenesis by Dvl2.

Protective Effect of Danggui Buxue Tang on Impaired Functional Activity of Endothelial Cells Exposed to Low Flow Shear Stress / 中国实验方剂学杂志

Objective: To investigate the protective effect of Danggui Buxue Tang(DGBX)with Angelicae Sinensis Radix(AS) and Astragali Radix(AR)at different radios on impaired functional activity of endothelial cells(ECs)exposed to low-fluid shear stress (FSS). Method: Low FSS was loaded by a parallel plate flow chamber,and ECs were divided into normal FSS group,low FSS group(each preincubated with M199 medium for 2 h),simvastatin group(preincubated with 0.1 μmol·L-1 simvastatin for 2 h),and 3 DGBX groups(preincubated with 3 g·L-1 AS and AR at 1:1,1:3,1:5 for 2 h,respectively). Then,the normal group was exposed to 1.2 Pa FSS,while the rest groups were all exposed to low FSS. At time points of 30,60, 360 min,the proliferation was detected by methyl thiazoly tetrazolium(MTT),the secretion of nitric oxide(NO) was detected by nitrase reduction test,and the mRNA and protein expressions of endothelial nitric oxide synthase(eNOS) were detected by Real-time fluorescence guantitative polymerase chain reaction(Real-time PCR) and Western blot,respectively. Result: Compared with the normal group, the secretion of NO and the expression of eNOS in ECs were both increased significantly at 60 min (PConclusion: DGBX could protect the functional activity of ECs exposed to low FSS.

Effects of shear stress on morphology, adhesion and proliferation of human umbilical cord blood mesenchymal stem cells / 中华实验眼科杂志

Objective@#To explore the effects of shear stress on morphology, adhesion and proliferation of human umbilical cord blood mesenchymal stem cells(hUC-MSCs).@*Methods@#The hUC-MSCs were cultured in vitro until confluence and then placed in a flow system.The cells were subjected to different shear stress (1, 2, 3, 4 dye/cm2) for 2 hours and 6 hours, and no shear stress treatment cells served as a static control.The morphological changes of hUC-MSCs in different groups were analyzed by phase contrast microscopy and immunofluorescence.The mRNA expression levels of intercellular adhesion molecule-1 (ICAM-1) and Ki67 were detected by real-time PCR.@*Results@#Compared with the static control group, the hUC-MSCs cells were arranged in the direction of fluid after treated with shear stress.The immunofluorescence results showed that the cytoskeletal protein F-actin filaments was prolonged after shear stress.The cytoskeleton was further elongated in the 2 dye/cm2 for 6 hours group when compared with 2 dye/cm2 for 2 hours group, and the cytoskeleton was loosened when time extended to 6 hours.Real-time PCR results showed that the relative expressions of ICAM-1 mRNA and Ki67 mRNA in the static control group and different gradient shear stress groups were significantly different, with significant differences among them (F=17.141, P=0.000; F=11.336, P=0.001). The relative expression of ICAM-1 mRNA in the 1, 2, 3, 4 dye/cm2 shear stress group was 2.74±0.32, 9.77±1.19, 6.70±0.92 and 5.69±0.72, respectively, which was significantly higher than 1.00±0.28 in the static control group, with significant differences between them (all at P<0.05). The relative expression of Ki67 mRNA in the 3 dye/cm2 and 4 dye/cm2 shear stress group was 0.39±0.09 and 0.04±0.02, respectively, which was significantly lower than 1.00±0.24 in the static control group, with statistically significant differences between them (both at P<0.05).@*Conclusions@#After treated with fluid shear stress, hUC-MSCs are arranged in the direction of fluid.Shear stress can promote the adhesion of hUC-MSCs.As the increase of shear stress intensity, cell proliferation is inhibited.

Optimization of the theoretical model for growth rate of mesenchymal stem cells on three-dimensional scaffold under fluid shear stress / 生物医学工程学杂志

Bone tissue engineering is considered as one of the most promising way to treat large segmental bone defect. When constructing bone tissue engineering graft , suitable bioreactor is usually used to incubate cell-scaffold complex under perfusion to obtain bone tissue engineering graft with good repair efficiency. However, the theoretical model for growth rate of single cell (especially for stem cell) during this process still has many defects. The difference between stem cells and terminally differentiated cells is always ignored. Based on our previous studies, this study used self-made perfusion apparatus to apply different modes and strengths of fluid shear stress (FSS) to the cells seeded on scaffolds. The effects of FSS on the proliferation and osteogenic differentiation of mesenchymal stem cells (MSCs) were investigated. The regression analysis model of the effect of FSS on the single-cell growth rate of MSCs was further established. The results showed that 0.022 5 Pa oscillatory shear stress had stronger ability to promote proliferation and osteogenic differentiation of MSCs, and the growth rate of a single MSC cell under FSS was modified. This study is expected to provide theoretical guidance for optimizing the perfusion culture condition of bone tissue engineering grafts .

The absence of fluid shear stress reduces AMPK activity in the renal I/R process / 中华器官移植杂志

Objective To investigate the mechanism of renal injury induced by changes in flow shear stress (FSS) during renal ischemia/reperfusion (I/R).Methods 1.In vitro,HUVECs were divided into 4 groups:(1) HUVECs were loaded with 12 dyn/cm2 force for 30,45,and 90 min by using plate fluid chamber system.(2) Cells were loaded with FSS for 2 h,and then cultured for 1,3,8 and 12 h respectively;(3) HUVECs were pretreated with 0,1,2,4 and 8 mmol metformin and cultured for 24 h.(4) HUVECs in control group were cultured normally.The expression of p-AMPK/AMPK protein was detected by Western blotting in each group.2.In vivo,16 SD rats with successful establishment of IR model were randomly divided into 4 groups (n =4 in each group):(1) static cold storage (CS) group:isolated kidneys were stored for 4 h;(2) hypothermia machine perfusion (HMP) group:isolated kidneys were continuously perfused with 0 ℃ lactated Ringer's solution for 4 h;(3)metformin treatment group (Met-CS):metformin was intraperitoneally injected 3 days before surgery,and the isolated kidneys were obtained after cold preservation for 4 h;(4)rat kidneys of control group were just subjected to thermal ischemia for 30 min.The injury of renal tissue in each group was observed by TUNEL and HE staining.The expression and distribution of p-AMPK protein in renal tissues were detected by immunohistochemistry.The correlation between FSS loss and AMPK expression in kidney tissue was analyzed.Results The expression of p-AMPK in HUVECs could be up-regulated by FSS,and the expression of p-AMPK protein increased with the prolongation of time.After stopping FSS,the expression of p-AMPK protein in HUVECs gradually decreased with time (P＜0.05).Metformin could activate AMPK activity in a concentration-dependent manner (P＜0.05).The content of p-AMPK in renal tissue of HMP group was significantly higher than that of CS group (P＜0.05).The expression of p-AMPK in renal tissue of HMP group mainly distributed in the renal tubules,and few in glomerular endothelial cells and blood vessels.The apoptosis rate of renal tissue in HMP group was significantly lower than that in CS group (P＜0.05).In the HMP group,the damage of the renal tissue was mild,there was no swelling,and the renal tubules were slightly expanded.In the CS group,the renal tissue was severely damaged and the renal tubules were markedly swollen.Conclusion During the course of renal IR in rats,changes in FSS may affect renal tissue damage through the AMPK pathway.

Vascular Endothelial Glycocalyx and Lipoproteins / 医用生物力学

Vascular endothelial cell glycocalyx is a layer of glycoprotein complex located on the surface of endothelial cells, forming a selective permeation barrier on the surface of endothelial cells. In the present review, after a brief introduction of glycocalyx, the relationship between glycocalyx and mass transport under fluid sheer stress (FSS), especially the relationship between glycocalyx and macromolecules such as low density lipoprotein (LDL) has been discussed. This relationship was reflected as following: on the one hand, the thickness and integrity of the glycocalyx affects the concentration polarization of LDL and its transendothelial transport and heparan sulfate proteoglycan (HSPG) participates in the whole process of residual lipoproteins metabolism. On the other hand, ox-LDL, an oxidized product of LDL, destroys heparan sulfate (HS) which is a major component of the endothelial cell glycocalyx. The study on relationship between vascular endothelial glycocalyx and lipoproteins will provide a new clue to elucidate the pathogenesis of atherosclerosis and provide more evidence to view the glycocalyx as a new control target.

E-Selectin Mediated-Calcium Response of Neutrophils under Fluid Shear Stresses / 医用生物力学

Objective To investigate the E-selectin mediated-calcium response of neutrophils under flow fields. Methods A parallel-plate flow chamber combined with a fluorescence microscope was used to observe the adhesion and subsequent calcium response of neutrophils on E-selectin at different concentrations under fluid shear stress (FSS) of 0-600 mPa. Results E-selectin could capture neutrophils from the flow to the chamber substrate and induce further intracellular calcium flux of firmly adhered cells. The arrested cell and activation ratio increased gradually as the concentration of E-selectin increased. Only immobilized E-selectin could conduct the external force-signal to trigger the calcium response of neutrophils effectively. By increasing FSS, not only was the activation ratio increased from 23% to 70%, but the calcium response intensity also increased from 0.92 to 1.45, while the delay time from cell adhesion to calcium response was greatly reduced from 70 s to 27 s. Conclusions FSS can modulate the calcium response of neutrophils in cooperation with E-selectin, and it positively regulates the activation rate and level of calcium response. This study may deepen the understanding of the immune response of leukocytes under a hemodynamic environment.

Effects of Fluid Shear Stress on Gene Expression of Piezo1 in the Cells During Osteogenic Differentiation / 医用生物力学

Objective To investigate the gene expression of Piezo1 in four types of bone cells at different stages of osteogenic differentiation under fluid shear stress (FSS). Methods The mouse-derived mesenchymal stem cells (MSC), osteoblast-like cells MC3T3-E1, post-osteoblasts MLO-A5 and osteocytes MLO-Y4 were exposed to FSS at different magnitude (0.1, 1.1 Pa) with a custom-made cone-plate flow chamber for 0.5, 1, 3, 6, 12 h, respectively. The expression of Piezo1 mRNA was assessed by quantitative real-time polymerase chain reaction. Results Both Piezo1 and Piezo2 were expressed in four types of bone cells. The expression of Piezo1 was significantly up-regulated in all cells under FSS stimulation, and the expression level under 1.1 Pa FSS was significantly higher than that under 0.1 Pa FSS. In addition, the expression of Piezo1 in MSC, MC3T3-E1 and MLO-A5 cells increased to the highest level at 1 h under FSS stimulation. The expression of Piezo1 in MC3T3-E1 cells was much higher than that in the other three types of cells. Conclusions The expression of Piezo1 was related to the process of osteogenic differentiation, FSS level and loading time, and this research finding is of great significance to reveal the mechanism of mechanotransduction in bone tissues and to establish clinical treatment for bone diseases.

Research progress on Mechanism of Fluid Shear Stress in Osteogenesis of Bone Mesenchymal Stem Cells / 医用生物力学

Cells are exposed to mechanical stress, such as fluid shear stress (FSS), mechanical strain, hydrostatic pressure in vivo. FSS is considered to be the most important stress during bone homeostasis and remodeling. At present, most studies are mainly about the FSS effect on osteocytes and osteoblasts. However, the effects of FSS on bone mesenchymal stem cell (BMSCs) are not fully understood. BMSCs are of great significance in bone reconstruction and clinical treatment, so researchers increasingly focus on the response of BMSCs to FSS. The response of BMSCs to FSS depends on the alteration of cytoskeleton, matrix stiffness and elasticity, osteogenic signaling pathways and so on. In this review paper, the recent researches about the mechanotransduction mechanism of FSS, and its effect on differentiation and function of BMSCs are summarized, so as to provide new insights for studying construction of tissue engineered bone and treatment of bone diseases.

Bmi-1 expression of bone marrow mesenchymal stem cells induced by fluid shear stress mediated with Akt signal molecule / 医用生物力学

Objective To investigate the effect of fluid shear stress (FSS) on the expression of B lymphoma MoMLV insertion region 1 (Bmi-1) in bone mesenchymal stem cells (BMSCs) and possible signal transduction mechanism.Methods BMSCs were isolated from SD rats and FSS at different magnitude (0.5,1.5,3.0 Pa)and under different time phase (1,2,6,24 h) were loaded by parallel-plate flow chamber system.The expression of Bmi-1 was measured by real-time RT-PCR at mRNA level and the levels of phosphorylated Akt (p-Akt)and extracellular signalregulated kinase 1/2 (p-ERK1/2) were detected by Western blotting.The signaling inhibitors,wortmannin (PI3K specific inhabitor) and PD98059 (ERK1/2 specific inhabitor),were used to investigate possible mechanical signal transduction pathway.Results Bmi-1mRNA expression increased when BMSCs were exposed to 1.5 Pa FSS for 1 h and reached the peak at 24 h.All FSS with different magnitude could increase Bmi-1 expression,especial at high FSS (3.0 Pa).Meanwhile,FSS resulted in a significant activation of p-Akt and p-ERK1/2 in BMSCs.After treated with wortmannin,the expression of Bmi-1 was inhibited prominently,however,PD98059,the expression of Bmi-1 did not change.Conclusions FSS can activate the expression of Bmi-1,the amount of Bmi-1 expression was closely related to the stimulating time and the magnitude of FSS,and Akt signal molecule plays an important role during the process.These findings provide significant references for studying the mechanical biological mechanisms of stem cell differentiation.

Bmi-1 expression of bone marrow mesenchymal stem cells induced by fluid shear stress mediated with Akt signal molecule / 医用生物力学

Objective To investigate the effect of fluid shear stress (FSS) on the expression of B lymphoma MoMLV insertion region 1 (Bmi-1) in bone mesenchymal stem cells (BMSCs) and possible signal transduction mechanism.Methods BMSCs were isolated from SD rats and FSS at different magnitude (0.5,1.5,3.0 Pa)and under different time phase (1,2,6,24 h) were loaded by parallel-plate flow chamber system.The expression of Bmi-1 was measured by real-time RT-PCR at mRNA level and the levels of phosphorylated Akt (p-Akt)and extracellular signalregulated kinase 1/2 (p-ERK1/2) were detected by Western blotting.The signaling inhibitors,wortmannin (PI3K specific inhabitor) and PD98059 (ERK1/2 specific inhabitor),were used to investigate possible mechanical signal transduction pathway.Results Bmi-1mRNA expression increased when BMSCs were exposed to 1.5 Pa FSS for 1 h and reached the peak at 24 h.All FSS with different magnitude could increase Bmi-1 expression,especial at high FSS (3.0 Pa).Meanwhile,FSS resulted in a significant activation of p-Akt and p-ERK1/2 in BMSCs.After treated with wortmannin,the expression of Bmi-1 was inhibited prominently,however,PD98059,the expression of Bmi-1 did not change.Conclusions FSS can activate the expression of Bmi-1,the amount of Bmi-1 expression was closely related to the stimulating time and the magnitude of FSS,and Akt signal molecule plays an important role during the process.These findings provide significant references for studying the mechanical biological mechanisms of stem cell differentiation.

Bmi-1 expression of bone marrow mesenchymal stem cells induced by fluid shear stress mediated with Akt signal molecule / 医用生物力学

Objective To investigate the effect of fluid shear stress (FSS) on the expression of B lymphoma MoMLV insertion region 1 (Bmi-1) in bone mesenchymal stem cells (BMSCs) and possible signal transduction mechanism.Methods BMSCs were isolated from SD rats and FSS at different magnitude (0.5,1.5,3.0 Pa)and under different time phase (1,2,6,24 h) were loaded by parallel-plate flow chamber system.The expression of Bmi-1 was measured by real-time RT-PCR at mRNA level and the levels of phosphorylated Akt (p-Akt)and extracellular signalregulated kinase 1/2 (p-ERK1/2) were detected by Western blotting.The signaling inhibitors,wortmannin (PI3K specific inhabitor) and PD98059 (ERK1/2 specific inhabitor),were used to investigate possible mechanical signal transduction pathway.Results Bmi-1mRNA expression increased when BMSCs were exposed to 1.5 Pa FSS for 1 h and reached the peak at 24 h.All FSS with different magnitude could increase Bmi-1 expression,especial at high FSS (3.0 Pa).Meanwhile,FSS resulted in a significant activation of p-Akt and p-ERK1/2 in BMSCs.After treated with wortmannin,the expression of Bmi-1 was inhibited prominently,however,PD98059,the expression of Bmi-1 did not change.Conclusions FSS can activate the expression of Bmi-1,the amount of Bmi-1 expression was closely related to the stimulating time and the magnitude of FSS,and Akt signal molecule plays an important role during the process.These findings provide significant references for studying the mechanical biological mechanisms of stem cell differentiation.

Bmi-1 expression of bone marrow mesenchymal stem cells induced by fluid shear stress mediated with Akt signal molecule / 医用生物力学

Objective To investigate the effect of fluid shear stress (FSS) on the expression of B lymphoma Mo-MLV insertion region 1 (Bmi-1) in bone mesenchymal stem cells (BMSCs) and possible signal transduction mechanism. Methods BMSCs were isolated from SD rats and FSS at different magnitude (0.5, 1.5, 3.0 Pa) and under different time phase (1, 2, 6, 24 h) were loaded by parallel-plate flow chamber system. The expression of Bmi-1 was measured by real-time RT-PCR at mRNA level and the levels of phosphorylated Akt (p-Akt) and extracellular signalregulated kinase 1/2 (p-ERK1/2) were detected by Western blotting. The signaling inhibitors, wortmannin (PI3K specific inhabitor) and PD98059 (ERK1/2 specific inhabitor), were used to investigate possible mechanical signal transduction pathway. Results Bmi-1mRNA expression increased when BMSCs were exposed to 1.5 Pa FSS for 1 h and reached the peak at 24 h. All FSS with different magnitude could increase Bmi-1 expression, especial at high FSS (3.0 Pa). Meanwhile, FSS resulted in a significant activation of p-Akt and p-ERK1/2 in BMSCs. After treated with wortmannin, the expression of Bmi-1 was inhibited prominently, however, PD98059, the expression of Bmi-1 did not change. Conclusions FSS can activate the expression of Bmi-1, the amount of Bmi-1 expression was closely related to the stimulating time and the magnitude of FSS, and Akt signal molecule plays an important role during the process. These findings provide significant references for studying the mechanical biological mechanisms of stem cell differentiation.

Fluid shear stress regulates epithelial-mesenchymal transition (EMT) in Hep2 cells / 医用生物力学

Objective To examine the effects of fluid shear stress (FSS) on epithelial-mesenchymal transition (EMT) in Hep2 cells. Methods Hep2 cells were exposed to 140 mPa FSS. The morphologic changes of Hep2 cells exposed to FSS at different durations were observed using inverted microscope. The migration ability of Hep2 cells after FSS loading was investigated using scratch wound assay. The distribution and expression of cytoskeleton protein F-actin were examined by confocal microscope. The expression of the EMT marker proteins were detected by Western blotting. Results Most of Hep2 cells changed their morphology from polygon to elongated spindle with well-organized F-actin under FSS. After removing FSS, Hep2 cells recovered their initial morphology with flat polygon. FSS regulated Hep2 cells to enhance their migration capacity in a time-dependent manner. FSS promoted the rearrangement of cytoskeletal protein F-actin，which enhanced the migration behavior of Hep2 cells. In addition, FSS induced a time regularity of expression of the EMT marker proteins in Hep2 cells. Conclusions FSS as an important physical factor can induce EMT in Hep2 cells.