Targeting fibroblast activation protein inhibits endothelial-mesenchymal transition by affecting cancer-associated fibroblasts derived exosomes / 中国药理学通报

Aim To investigate whether targeted inhibition of fibroblast activation protein (FAP) can inhibit the endothelial-to-mesenchymal transition (EndMT) of vascular endothelial cells by affecting exosomes (Exo) of cancer-associated fibroblasts (CAFs) and explore the underlying mechanisms. Methods Primary CAFs and peri-tumor fibroblasts (PTFs) were obtained from lung cancer and peri-cancer tissues, and CAFs-exo and PTFs-exo were collected from culture medium, respectively. Exosomes from CAFs treated with specific FAP inhibitor (3.3 nmol • L-

Inducible co-stimulatory molecules participate in mesenteric vascular endothelial-mesenchymal transition and sclerosis of mesenteric vessels in spontaneously hypertensive rats / 南方医科大学学报

OBJECTIVE@#To investigate the correlation of inducible co-stimulatory molecules (ICOS) with mesenteric vascular endothelial- mesenchymal transition (EndMT) and sclerosis in spontaneously hypertensive rats (SHR).@*METHODS@#Twenty 4-week-old WKY rats and 20 SHRs of the same strain were both randomly divided into 4 groups for observation at 4, 6, 10 and 30 weeks of age. ICOS expression frequency in rat spleen CD4+T cells was analyzed using flow cytometry, and the expressions of ICOS, VE-cad, α-SMA and Col3 mRNA in rat mesentery were detected by RT-PCR. The distributions of ICOS, IL-17A and TGF-β in rat mesentery were detected by immunohistochemistry. The levels of IL-17A and TGF-β in rat plasma were measured using ELISA. The morphological changes of rat mesenteric vessels were observed with Masson staining. Spearman or Pearson correlation analyses were used to evaluate the correlation between ICOS expression and the expressions of the markers of vascular EndMT and sclerosis.@*RESULTS@#Compared with the control WKY rats, the SHRs began to show significantly increased systolic blood pressure and ICOS expression frequency on CD4+T cells at 6 weeks of age (P < 0.05). In the SHRs, the mRNA and protein expressions of ICOS, α-SMA, Col3, IL-17A and TGF-β in the mesentery were significantly higher than those in control group (P < 0.05), while the mRNA and protein expressions of VE-cad started to reduce significantly at 10 weeks of age (P < 0.05). The plasma levels of IL-17A and TGF-β were significantly increased in SHRs since 6 weeks of age (P < 0.05) with progressive worsening of mesenteric vascular sclerosis (P < 0.05). ICOS mRNA and protein expression levels in the mesenteric tissues of SHRs began to show positive correlations with α-SMA and Col3 expression levels and the severity of vascular sclerosis at 6 weeks of age (P < 0.05) and a negative correlation with VE-cad expression level at 10 weeks (P < 0.05).@*CONCLUSION@#ICOS play an important pathogenic role in EndMT and sclerosis of mesenteric vessels in essential hypertension by mediating related immune responses.

Impact of the transforming growth factor-β pathway on vascular restenosis and its mechanism / 中南大学学报(医学版)

As a crucial regulatory molecule in the context of vascular stenosis, transforming growth factor-β (TGF-β), plays a pivotal role in its initiation and progression. TGF-β, a member of the TGF-β superfamily, can bind to the TGF-β receptor and transduce extracellular to intracellular signals through canonical Smad dependent or noncanonical signaling pathways to regulate cell growth, proliferation, differentiation, and apoptosis. Restenosis remains one of the most challenging problems in cardiac, cerebral, and peripheral vascular disease worldwide. The mechanisms for occurrence and development of restenosis are diverse and complex. The TGF-β pathway exhibits diversity across various cell types. Hence, clarifying the specific roles of TGF-β within different cell types and its precise impact on vascular stenosis provides strategies for future research in the field of stenosis.

Research progress of endothelial-to-mesenchymal transition in ocular diseases / 国际眼科杂志(Guoji Yanke Zazhi)

@#Endothelial-to-mesenchymal transition(EndoMT)is a change in the transformation or differentiation of endothelial cells into mesenchymal cells under physiological or pathological conditions, accompanied by changes in phenotype and function, and is an important part of fiber repair. It is widely involved in the pathophysiological process of embryonic development, tumor invasion and a variety of fibrotic diseases. Research on the role of EndoMT in ocular diseases has also made some progress. This article will review the basic biological characteristics, mechanism and research results of EndoMT in ophthalmological diseases, intending to theoretically reveal its possibility as a treatment target and a key point of regenerative medicine technology in related diseases, provide a reference for clinical practice and scientific research.

Research advances on pathogenesis of diabetic pulmonary fibrosis / 中国药理学通报

As a pulmonary complication of diabetes, diabetic pulmonary fibrosis has gradually entered people's sight, but its mechanism is still poorly understood.This is the first systematic review of the mechanisms of autonomic neuropathy, pulmonary microangiopathy, accumulation of advanced glycosylation end products, oxidative stress, inflammation, epithelial-mesenchy- mal transition and endothelial-mesenchymal transition, cell se¬nescence and I)NA damage, etc.in diabetic pulmonary fibrosis.which aims to provide inquiring ideas for exploring the specific molecule mechanism and a reference for the development of ther¬apeutic drugs for diabetic pulmonary fibrosis.,,,,.

Study on the effect and mechanism of Pirfenidone on endothelial-mesenchymal transition in endothelial cells / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM:To establish the hypoxia induced endothelial-mesenchymal transition(EndoMT)model of endothelial cells, and to investigate the effect and mechanism of Pirfenidone(PFD)on inhibiting the subretinal fibrosis progression.<p>METHODS: Primary cultured human umbilical vein endothelial cells(HUVEC), 4-7 passages were used for experiments after cell identification. CoCl₂ induced hypoxia to establish the transformation model of endothelial cells into fibroblasts. CCK-8 was performed to detect cell proliferation rate and chose the optimal drug concentration. All cells were divided into 4 groups: control group(FBS-free), CoCl₂(200μmol/L)group, CoCl₂+0.3mg/mL PFD group, CoCl₂+0.6mg/mL PFD group. The protein expression of CD31, VE-cadherin, α-SMA, FSP1, p-p38 and p38 were detected by Western blot. Double immunofluorescence labeling method was used to observe the CD31/α-SMA expression. Wound healing assay detected the cell migration. The q-PCR was applied to detect the mRNA levels of TGF-β1 and SNAI1.<p>RESULTS: Compared with CoCl₂ group, PFD increased cell proliferation rate and inhibited cell migration significantly under hypoxia(<i>P</i><0.05). PFD decreased the protein expression of the mesenchymal markers α-SMA and FSP1, and increased the protein level of the endothelial markers CD31 and VE-cadherin(<i>P</i><0.05). Double immunofluorescence results showed that PFD could reduce the expression of α-SMA and increase the level of CD31(<i>P</i><0.05). In the process of EndoMT, the p38 protein expression level was stable(<i>P</i>>0.05). PFD down-regulated significantly the high protein expression of p-p38, and high mRNA expression of TGF-β1 and SNAI1 compared with control group(<i>P</i><0.05). There was no significant difference between the 0.3 and 0.6mg/mL PFD groups in all results above.<p>CONCLUSION: PFD can inhibit the formation of fibrosis in endothelial cells. TGF-β/p38MAPK signaling pathway might be one of the mechanisms that PFD regulates EndoMT progression. PFD will be expected to become a potential new sight on the treatment of subretinal fibrosis.

Effects of metformin on epithelial-mesenchymal transition of rat alveolar epithelial type II cells induced by TGF-β / 中国临床药理学与治疗学

AIM: To observe the effect of metformin (Met) on the endothelial-mesenchymal transition (EMT) of rat alveolar epithelial type II cells and its mechanism. METHODS: The RLE-6TN cells were divided into 6 groups as follows: Control group; transforming growth factor-β

Salidroside inhibits endothelial-mesenchymal transition by regulating KLF4/eNOS signaling pathway / 中国药理学通报

Aim To investigate the effect and mechanism of salidroside (SAL) on homocysteine (Hcy)-induced endothelial-mesenchymal transition (EndMT) based on the KLF4/eNOS signaling pathway. Methods (1) Salidroside inhibited Hcy-induced EndMT. HUVECs were pretreated with different concentrations of SAL for 2 h, then followed by Hcy (1 mmol · L-l) co-incubation for 48 hours to induce EndMT. The expression levels of VE-cadherin, α-SMA, KLF4 and eNOS were detected by Western blot, scratch repair experiment was used to determin cell migration ability, the level of NO in cells was determined by nitrate reductase method, and the expression and location of KLF4 were observed by immunofluorescence technology. (2) The signal mechanism of SAL inhibiting End-MT through KLF4/eNOS signal was studied. siRNA mediated knockdown of KLF4 in HUVECs, and the protein expression levels of VE-cadherin, α-SMA, KLF4 and eNOS in each group were determined by Western blot. Results Western blotting demonstrated that SAL significantly reversed the Hcy-induced increase in α-SMA and KLF4 expression and decrease in VE-cadherin and eNOS expression. The immunofluorescence analysis suggested that SAL inhibited the translocation of KLF4 from cytoplasm to nucleus. (2) Silencing KLF4 down-regulated the expression of α-SMA and KLF4, and up-regulated the expression of VE-cadherin and eNOS. Compared with the group of SAL + siKLF4, there was no significant difference in the effect of SAL group and siKLF4 group on phenotypic markers. Conclusions Salidroside inhibits EndMT induced by Hcy, which may be related to the regulation of KLF4/eNOS signaling pathway.

The endothelial-mesenchymal transition in glioma and strategies for targeted therapy: research progress / 国际药学研究杂志

Endothelial-mesenchymal transition(EndoMT)is a pathophysiological progress that endothelial cells lose their specific phenotype,function and morphology and gradually transform to mesenchymal cells under the effect of certain stimulating factors. More and more researches have shown that EndoMT plays a key role in the occurrence and development as well as the invasion and metastasis of glioma. This paper reviews the EndoMT-related signaling pathways and their important effect on the angiogenesis,invasion,metastasis,immunosuppression and the drug resistance of glioma. Meanwhile, strategies for the EndoMT-targeted therapy are also summarized, so as to provide a reference for related studies and clinical treatment of glioma.

Research progress of endothelial mesenchymal transition in myocardial fibrosis / 中国药理学通报

Endothelial mesenchymal transition ( EndMT) , a biological process in which endothelial cells under specific environmental stimuli, gradually lose close connection and expression of specific markers, and obtain stromal cells or muscle fiber cell plienolypic expression and characteristics of movement and shrinkage. EndMT not only plays a crucial role in the development of heart and vascular, but lias been widely studied in the process of organ fibrosis such as heart, lung and kidney. It may be the key step in the prevention and treatment of myocardial fibrosis. We review the research progress of EndMT in myocardial fibrosis.

Role of EndoMT in HHPH based on TGF-β/Smads signaling pathway and regulated by Yiqi-Wenyang-Huoxue-Huatan formula / 中国病理生理杂志

AIM:To investigate the relationship between transforming growth factor-β(TGF-β)/Smads signa-ling pathway and pulmonary arterial endothelial-mesenchymal transition(EndoMT)in hypoxia-hypercapnia pulmonary hy-pertension(HHPH)process and the regulatory effect of Yiqi-Wenyang-Huoxue-Huatan formula(YWHHF).METHODS:Healthy male SD rats were randomly divided into 5 groups:normal control(N)group,hypoxia-hypercapnia(HH)group, high-dose YWHHF(YH)group,middle-dose YWHHF(YM)group and low-dose YWHHF(YL)group.The rats in N group was housed in normoxic environment,and the rats in the other 4 groups were housed in hypoxia-hypercapnia environ-ment(9%~11%O2and 5%~6%CO2)for 4 weeks,8 h/d,6 d/week.The excess water vapor was absorbed by anhy-drous CaCl2,and CO2was absorbed by sodium hydroxide.The rats in YWHHF groups were put into the oxygen chamber before the same volume of YWHHF at different concentrations were given(200 g/L for YH group,100 g/L for YM group and 50 g/L for YL group).The average pulmonary artery pressure and the average carotid artery pressure were measured during the operation.After operation,the right ventricular free wall and left ventricle plus interventricular septum were col -lected for determining the right ventricular hypertrophy index.Moreover,the morphological changes of the lung tissues were observed under light microscope.The mRNA and protein levels of α-smooth muscle actin(α-SMA),CD31,TGF-β1 and Smad2/3,and the protein level of p-Smad2/3 were detected by RT-PCR and Western blot.RESULTS:Compared with N group,the pulmonary artery mean pressure,the mRNA and protein expression of α-SMA,TGF-β1 and Smad2/3,and the protein level of p-Smad2/3 were increased, the levels of CD31 were decreased(P<0.05), and the lung tissue damage was observed in the other 4 groups.Compared with HH group,the pulmonary artery mean pressure,the mRNA and protein expression of α-SMA,TGF-β1 and Smad2/3,and the protein level of p-Smad2/3 were decreased, while the mRNA and protein levels of CD31 were increased.Moreover,the lung tissue damage was reduced in YH,YM and YL groups.CON-CLUSION:TGF-β/Smads pathway may be involved in the process of EndoMT under hypoxia and hypercapnia condition, and YWHHF may reduce EndoMT by inhibiting the expression of TGF-β/Smads pathway-related molecules.

The effect of TGF-β/ILK pathway in endothelial-mesenchymal transition induced by parathyroid hormone in human vascular endothelial cells / 重庆医学

Objective To investigate the molecular mechanism of endothelial-mesenchymal transition (EndMT) induced by uremic toxin,parathyroid hormone (PTH),in vascular endothelial cells.Methods PTH (1 × 10-8 mol/L) was used to induce EndMT in human aortic endothelial cells (HAECs).TGF-β signaling inhibitor,including SB431542 and pirfenidone (PFD),and integrin-linked kinase (ILK) inhibitor Cpd22 were used to investigate the potential mechanism of EndMT induced by PTH in HAECs.Then the vascular endothelial cell markers VE-cadherin and CD31,and the mesenchymal marker α-SMA were detected by western blot.Results PTH reduced the expression levels of vascular endothelial cell marker CD31 and VE-cadherin (P＜0.05),while significantly increased the expression level of fiber cell marker α-SMA(P＜0.05).Furthermore,the TGF-βsignaling inhibitors (SB431542 and PFD) and ILK inhibitor (Cpd22) were able to partially reverse the EndMT induced by PTH in HAECs,which reversed the effect of PTH on reducing vascular endothelial cell marker expression and increasing fiber cell marker expression.Conclusion PTH could induce EndMT in HAECs via TGF-β/ILK pathway.

Regulation and its signaling mechanism of miR-328on endothelial-mesenchymal transition induced by high glucose of humanumbilical vein endothelial cells / 基础医学与临床

Objective The study is to investigate the role of miR-328 in endothelial mesenchymal transition (EMT)induced by high glucose in human umbilical vein endothelial cells (HUVECs)and its signaling mechanism.Methods HUVECs were cultured in high glucose environment to induce EMT;The recombinant lentiviruses were created by miR-328 and antagomiR- 328 transfection of HUVECs.The experiment was divided into seven groups: normal glucose;mannitol group;high glucose;miR-328;miR-328 virus negative control;high glucose + U0126;miR-328 + U0126.Double immunofluorescent staining was used to determine expression of EMT markers;Changes in miR-328 expression is examined by RT-qPCR;The expressions of type Ⅰ/Ⅲ collagen,p-MEK1/2 and p-ERK1/2 are examined by Western blot.Results 1)HUVECs showed positive staining for CD31 and α-SMA in high glucose group.2)The expression of miR-328 was up-regulated(P<0.05)in HUVECs treated by high glucose or miR-328.Compared with high glucose group or miR-328 group,miR-328 expression was less pronounced aftertreatment with U0126.3)The expressions of type Ⅰ/Ⅲ collagen increased in HUVECs treated by high glucose or miR-328(P<0.05) Compared with high glucose group or miR-328 group,typeⅠ/Ⅲ collagen expressions were less pronounced after treatment with U0126.4)The expressions of p-MEK1/2 and p-ERK1/2 were increased in HUVECs treated by high glucose or miR-328 in comparison to the control group (P<0.05);a lower expression of p-MEK1/2 and p-ERK1/2 were observed in U0126 group than in high glucose group or miR-328 group.Conclusions The phenomenon of EMT in HUVECs is induced by high glucose with increased expression of miR-328;overexpression of miR-328 induced EMT in HUVECs;miR-328 induced EMT is related with MEK1/2-ERK1/2 signaling pathway.

Celastrol Protects TGF-β1-induced Endothelial-mesenchymal Transition / 华中科技大学学报（医学）（英德文版）

The endothelial-to-mesenchymal transition (EndMT) in endothelial cells contributes to the development of cardiac fibrosis,ultimately leading to cardiac remodeling.In this study,the effects and molecular mechanisms of celastrol (CEL) on transforming growth factor-β1 (TGF-β1)-induced EndMT in human umbilical vein endothelial (HUVEC-12) cells were investigated.The presented data demonstrated.that CEL significantly blocked the morphology change of HUVEC-12 cells induced by TGF-β1 without cell cytotoxicity.In accordance with these findings,CEL blocked TGF-β1-induced EndMT as evidenced by the inhibition of the mesenchymal markers,including collagen Ⅰ,Ⅲ,α-SMA,fibronectin mRNA expression,and the increase in the mRNA expression of endothelial cell marker CD31.These changes were also confirmed by double immunofluorescence staining of CD31 and vimentin.The in vitro scratch assay showed that CEL inhibited the migration capacity of the transitioned endothelial cells induced by TGF-β1.Further experiments showed that the beneficial effect of CEL on blocking the EndMT in HUVEC-12 cells was associated with the suppression of the TGF-β1/Smads signalling pathway,which was also confirmed by the inhibition of its downstream transcription factor snail1,twistl,twist2,ZEB1 and ZEB2.These results indicate that CEL blocks TGF-β1-induced EndMT through TGF-β1/Smads signalling pathway and suggest that it may be a feasible therapy for cardiac fibrosis diseases.

Aspirin-Triggered Resolvin D1 Inhibits TGF-β1-Induced EndMT through Increasing the Expression of Smad7 and Is Closely Related to Oxidative Stress

The endothelial-mesenchymal transition (EndMT) is known to be involved in the transformation of vascular endothelial cells to mesenchymal cells. EndMT has been confirmed that occur in various pathologic conditions. Transforming growth factor β1 (TGF-β1) is a potent stimulator of the vascular endothelial to mesenchymal transition (EMT). Aspirin-triggered resolvin D1 (ATRvD1) has been known to be involved in the resolution of inflammation, but whether it has effects on TGF-β1-induced EndMT is not yet clear. Therefore, we investigated the effects of AT-RvD1 on the EndMT of human umbilical vein vascular endothelial cells line (HUVECs). Treatment with TGF-β1 reduced the expression of Nrf2 and enhanced the level of F-actin, which is associated with paracellular permeability. The expression of endothelial marker VE-cadherin in HUVEC cells was reduced, and the expression of mesenchymal marker vimentin was enhanced. AT-RvD1 restored the expression of Nrf2 and vimentin and enhanced the expression of VE-cadherin. AT-RvD1 did also affect the migration of HUVEC cells. Inhibitory κB kinase 16 (IKK 16), which is known to inhibit the NF-κB pathway, had an ability to increase the expression of Nrf2 and was associated with the inhibition effect of AT-RvD1 on TGF-β1-induced EndMT, but it had no effect on TGF-β1-induced EndMT alone. Smad7, which is a key regulator of TGF-β/Smads signaling by negative feedback loops, was significantly increased with the treatment of AT-RvD1. These results suggest the possibility that AT-RvD1 suppresses the TGF-β1-induced EndMT through increasing the expression of Smad7 and is closely related to oxidative stress.

Effect of TGF-β/Smad signal pathway mediated by integrin linked kinase in the progress of renal fibrosis / 重庆医学

Objective To observe the effect of TGF‐β1 in the endothelial‐mesenchymal transition EndM T of glomeruli ,and to explore the effect of TGF‐β/Smad signaling pathway mediated by integrin linked kinase(ILK ) in the progress of renal fibrosis . Methods Human glomerular endothelial cells (HGEnC) incubated in vitro were divided into blank control group ,TGF‐β1 (12 .5 , 25 .0 ,50 .0 ng/mL) group .TGF‐β1 50 .0 ng/mL receptor type one antagonist (LY364749)5 .0 μmol/L group ,ILK (QLT‐0267)5 .0μmol/L antagonist group .The mRNA level of P‐Smad 2/3 and ILK was determined by RT‐PCR ,and the protein level of P‐Smad 2/3 ,ILK ,E‐cadherin ,CD31 ,α‐SMA and FSP‐1 was determined by Western blot after 48 h and 72 h after incubation in each group un‐der the above‐mentioned condition .Results (1)TGF‐β1 could significantly increased the mRNA level of P‐Smad2/3 and ILK (P0 .05) .Conclusion TGF‐β1 as the effector molecule in downstream can promote endothelial‐mesenchymal transition of HGEnC ,and TGF‐β/Smad signaling pathway mediated integrin linked kinase participate in this process ,which probably play important role in the progress of renal fibrosis .

Effect of high glucose on glomerular endothelial-mesenchymal transition and its possible mechanism / 中华肾脏病杂志

Objective To investigate whether high glucose can induce endothelial-mesenchymal transition (EndMT) in glomerular endothelial cells and the role of TGF-β in the process.Methods Rat glomerular endothelial cells were divided into five groups:normal glucose (NG,5.5mmol/L),high glucose (HG,15,30 mmol/L),TGF-β inhibition (HG+ LY36,30 mmol/L glucose + 10 μmol/L LY364947),hyperosmotic control (M,5.5 mmol/L glucose+25.5 mmol/L mannitol) and solvent control (D,5.5 mmol/L glucose + 1 ml/L DMSO).Western blotting was performed to detect relative protein quantities of endothelial marker claudin 5 and mesenchymal marker α-smooth muscle actin (α-SMA).TGF-β1 and TGF-β2 mRNA levels were measured by real-time PCR.Vascular endothelial marker VE-cadherin and mesenchymal marker α-SMA were detected by immunofluorescent stain and observed by confocal microscopy.Results Compared with NG,the expression of claudin5 protein in HG (15 or 30 mmol/L) was up-regulated while expression of α-SMA protein was down-regulated (P ＜0.05).Both TGF-β1 and TGF-β2 mRNA levels increased as well (P ＜ 0.05).However,when compared with HG,the claudin 5 levels increased while α-SMA decreased in TGF-β inhibition group.No significant changes were observed in hyperosmotic or solvent control group.Confocal microscopy showed the transformation of cells from a cobblestone-liked shape to a spindle one,and a decreasing expression of VE-cadherin while an increasing α-SMA in HG group (P ＜ 0.05),whereas TGF-β inhibition partly attenuated those changes in both morphological and protein levels.Conclusions High glucose treatment of glomerular endothelial cells results in an increase in the level of TGF-β1 and TGF-β2 mRNA and leads to endothelial-mesenchymal transiton.Inhibition of TGF-β partly prevents this process,indicating that TGF-β plays a crucial role in high-glucose-induced glomerular endothelial-mesenchymal transiton.