Effect of cladribine on growth inhibition and autocrining cytokines in human umbilical vein endothelial cell line EA.hy926

AIM: To study the effects of cladribine on growth and secretion activity of human umbilical vein endothelial cell line EA.hy926, and to investigate the mechanism of its anti-tumor effect by inhibiting endothelial cells. METHODS: The effects of cladribine at different concentrations on the cell viability were detected by CCK-8 assay. Apoptosis and cell cycle distribution were examined by flow cytometry. The protein expression levels were determined by Western blot. The levels of tumor necrosis factor-α (TNF-α), transforming growth factor-β1 (TGF-β1) and vascular endothelial growth factor (VEGF) secreted by EA.hy926 cells with cladribine treatment for 48 h were analyzed by ELISA. The nitric oxide (NO) production was measured by Gries method. RESULTS: Cladribine at 0.4~1 μmol/L inhibited the viability of EA.hy926 cells in time-and dose-dependent manners. The IC₅₀ was about 3.644 μmol/L. The results showed 43.74% cells in S phase when the concentration of cladribine was 0.4 μmol/L, and 77.23% cells in S phase when the concentration of cladribine was 1 μmol/L. The apoptosis was not induced by cladribine at 0.4~10 μmol/L. The protein expression of Bax and caspase-3 did not change. The expression of p21 increased and the p53 decreased (P<0.05). The levels of TNF-α and TGF-β1 secreted by EA.hy926 cells increased after cladribine treatment for 48 h. The levels of VEGF and NO decreased. CONCLUSION: Cladribine obviously inhibits the viability of EA.hy926 cells. The mechanism is related to the cell cycle arrest. Cladribine promotes the secretion of TNF-α and TGF-β1 by EA.hy926 cells and inhibits the secretion of VEGF and NO.     

Expression of Smoothened and its role in endothelial cells in synovial tissues of rheumatoid arthritis

AIM: To investigate the expression of Sonic Hedgehog signaling pathway-associated factor Smoothened (Smo) and its role in endothelial cells in synovial tissue of active rheumatoid arthritis (RA). METHODS: Smo expression in synovial tissue from 4 RA patients and 4 patients with trauma or meniscal injury (without arthritis, used for control) was detected by the method of immunohistochemistry. Human umbilical vein endothelial cell line EA.hy926 was used as the model of synovial vascular endothelial cells. The expression of Smo was detected by Western blotting after TNF-α treatment. The small interfering RNA (siRNA) specifically targeting Smo gene was synthesized and transfected into EA.hy926 cells. The interference efficiency of the siRNA on the production of Smo protein was determined by Western blotting. The cells were treated with TNF-α and actinomycin D (ActD) 24 h after siRNA transfection. The cell survival rate was determined by CCK-8 assay and the apoptotic rate was examined by flow cytometry. RESULTS: Smo was highly expressed in synovial tissue from active RA patients, especially in endothelial cells as compared with control group. TNF-α significantly increased the protein expression of Smo in EA.hy926 cells. EA.hy926 cells transfected with Smo-siRNA showed a significant decrease in the cell viability with the cell survival rate of (24.30±0.45)% and the apoptotic rate of (48.00±1.96)%, as compared with those in negative control group ［(36.86±0.62）% and （31.70±0.82）%, respectively］. CONCLUSION: Smo may play a role in the regulation of apoptosis in endothelial cells in RA synovium.     

Lycium barbarum polysaccharides attenuate oxidative stress injury of human endothelium-like EA.hy926 cells induced by hydrogen peroxide

AIM: To study the effect of Lycium barbarum polysaccharides (LBP) on oxidative stress injury of human endothelium-like EA.hy926 cells induced by hydrogen peroxide (H₂O₂). METHODS: The EA.hy926 cell model of oxidative stress injury was established by H₂O₂ treatment. The EA.hy926 cells were divided into 5 groups:control group, damage (H₂O₂ at 50 mmol/L) group, LBP (100 mg/L) group, anti-damage groups (LBP at 50 mg/L, 100 mg/L or 200 mg/L+50 mol/L H₂O₂), and LY294002 (20 μmol/L) group. The effect of LBP at different concentrations on the cell viability of EA.hy926 cells was measured by CCK-8 assay, and the optimum concentration of LBP was screened out. The apoptotic of EA.hy926 cells was analyzed by flow cytometry. Acridine orange/ethidium bromide (AO/EB) staining was used to observe the morphological characteristics of the apoptotic cells. The cell migration ability was detected by scratch method. The levels of nitric oxide (NO) and vascular endothelial growth factor (VEGF) in the cell culture medium were examined. The protein levels of cleaved caspase-3, Bax, Bcl-2, endothelial NO synthase (eNOS), p-eNOS and p-Akt were determined by Western blot. RESULTS: LBP at concentration of 100 mg/L significantly attenuated the injury of EA.hy926 cells induced by H₂O₂, as indicated by improved cell viability (P<0.05) and decreased apoptosis (P<0.05). Pretreatment with LBP elevated the levels of NO and VEGF (P<0.05), and promoted the migration ability of EA.hy926 cells. LBP also increased the Bcl-2/Bax ratio, down-regulated the protein level of cleaved caspase-3, and up-regulated the protein levels of eNOS and p-eNOS. The protective effect of LBP were abolished by pretreatment of the EA.hy926 cells with the inhibitor of PI3K (P<0.05). As a result, the protein level of p-Akt was down-regulated, and the level of NO was also significantly reduced. CONCLUSION: LBP has protective effect on H₂O₂ -induced EA.hy926 cells by attenuating apoptosis of the cells. The mechanism is closely related to the activation of PI3K/Akt/eNOS signaling pathway.     

Salvianolate attenuates oxidative stress injury of human endothelial EA.hy926 cells induced by hydrogen peroxide

AIM: To investigate the effect of salvianolate on oxidative damage induced by hydrogen peroxide in human endothelial EA.hy926 cells.METHODS: EA.hy926 cells were cultured in vitro and divided into the following groups:control group, damage group, and anti-damage groups (salvianolate+damage groups). The cell viability was measured by CCK-8 assay. The migration ability of the EA.hy926 cells was detected by Transwell assay. The content of nitric oxide (NO) in the culture supernatant of the EA.hy926 cells was examined. The levels of vascular endothelial growth factor (VEGF) were detected by ELISA. The apoptosis,mitochondrial membrane potential and intracellular superoxide anion content of the EA.hy926 cells were analyzed by flow cytometry. The protein levels of caspase-3, cleaved caspase-3, Bcl-2, Bax, NF-κB and p53 were determined by Western blot. RESULTS: Compared with damage group, the viability of EA.hy926 cells pretreated with salvianolate at different concentrations was significantly increased (P<0.05). The apoptotic rate was significantly decreased (P<0.05). Savianolate enhanced the migration ability of the cells. The levels of VEGF, NO and mitochondrial transmembrane potential were increased (P<0.05), and the intracellular ROS level was significantly decreased (P<0.05). The protein levels of NF-κB, p53, Bax and cleaved caspase-3 were significantly decreased, and the protein level of Bcl-2 was markedly increased(P<0.05). CONCLUSION: Savianolate reduces the damage of EA.hy926 cells by hydrogen peroxide exposure, and its mechanism may be related to the blocking of NF-κB signaling pathway.     

AngiotensinⅡ activates autophagy pathway through elevating ROS level in vascular endothelial cells

AIM: To evaluate the effects of angiotensinⅡ (AngⅡ) on autophagy induction in vascular endothelial cells. METHODS: Human vascular endothelial EA.hy926 cells were used in the study. Intracellular reactive oxygen species (ROS) levels were detected by a microplate reader after the cells were treated with AngⅡ (10^-7 mol/L) or AngⅡ combined with antioxidant N-acetyl-L-cysteine (NAC,50 μmol/L) for 24 h. The protein levels of LC3-Ⅱ was detected by Western blotting after the cells were stimulated by different concentrations (10^-8, 10^-7, 10^-6 mol/L) of AngⅡ for 24 h or by AngⅡ (10^-7mol/L) for different time (0 h, 6 h, 12 h, 24 h, 36 h). The number of autophagosomes was evaluated by fluorescence microscopy after stained with acridine orange. Similarly, the protein level of LC3-Ⅱ and the number of autophagosomes were detected after treated with AngⅡ(10^-7mol/L), AngⅡ combined with autophagy inhibitor 3-methyladenine (3-MA) at concentration of 2 mmol/L or AngⅡ combined with NAC at concentration of 50 μmol/L. RESULTS: Intracellular ROS level and LC3-Ⅱprotein level were significantly increased (P<0.05) after the cells were treated with AngⅡ, accompanied by the significant increase in the number of autophagosomes. AngⅡ-induced autophagy (as showed both in LC3-Ⅱprotein level and autophagosomes) was dramatically down-regulated by the treatment with 3-MA or NAC in EA.hy926 cells (P<0.05). CONCLUSION: AngⅡ induces autophagy through elevating ROS levels in EA.hy926 cells.     

Protective effect of SIRT1 on rat mesangial cells by decreasing high glucose-induced acetylation of NF-κB p65

AIM: To investigate the effects of silent information regulator 1 (SIRT1) on high glucose-induced acetylation of NF-κB p65 subunit and its protective role in rat mesangial cells. METHODS: Rat mesangial cells were cultured in DMEM supplemented with 10% FBS and were divided into control group, mannitol group, high glucose group, resveratrol group and SIRT1 RNAi group. The cell viability was determined by MTT assay. The mRNA expression of SIRT1, monocyte chemoattratant protein 1 (MCP-1), vascular cell adhesion molecule 1 (VCAM-1-1), tumor necrosis factor α (TNF-α), transforming growth factor β1 (TGF-β1) was analyzed by real-time quantitative PCR. The protein expression of SIRT1 and the acetylation of NF-κB p65 subunit were determined by Western blotting. The protein concentrations of MCP-1, VCAM-1, TNF-α, TGF-β1 and malondialdehyde (MDA) were detected by ELISA. RESULTS: The cell viability, superoxide dismutase (SOD) activity, and the expression of SIRT1 at mRNA and protein levels were decreased by high glucose treatment as compared with control group. The acetylation of NF-κB p65 subunit was significantly increased after interfered with high glucose, resulting in the increase in the secretion of MCP-1, VCAM-1, TNF-α and TGF-β1. Resveratrol decreased high glucose-induced acetylation of NF-κB p65 subunit. However, silencing SIRT1 significantly enhanced the acetylation of NF-κB p65 subunit and the expression of MCP-1, VCAM-1, TNF-α and TGF-β1. CONCLUSION: SIRT1 remarkably inhibits the inflammatory reactions by deacetylating NF-κB p65, suggesting that SIRT1 is a possible target for preventing diabetic nephropathy.     

Effect of N,N-dimethylsphingosine on the expression of adhesion molecules in human umbilical vein endothelial cell line EA.hy926

AIM: To investigate whether and how N, N-dimethylsphingosine (DMS) plays a role in modulating the adhesion of monocytes to vascular endothelial cells, and identify whether human umbilical vein endothelial cell line EA.hy926 take place of the vascular endothelial cells.METHODS: Adhesion ratio was measured by flow cytometry, and immunohistochemistry was used to detect the expression of ICAM-1 and P-selectin in HUVEC: EA.hy926 cells after the effect of DMS. RESULTS: DMS inhibited the adhesion of monocytes to HUVEC: EA.hy926 cells in a time-dependent and concentration-dependent manner by reducing the expression of ICAM-1 and P-selectin. CONCLUSIONS: DMS reduced adhesion molecule expression in vascular endothelial cells. DMS may be an important contributor to reduce adhesion ratio, suggesting that DMS plays a negative role in proinflammatory and immune functions of the modified vascular endothelial cells during atherosclerosis and restenosis.     

ZNF580 is up-regulated in S1P-induced migration and proliferation of endothelial cells

AIM: To investigate whether a novel human C2H2-type zinc finger protein ZNF580 is involved in the proliferation and migration of endothelial cells induced by sphingosine 1-phosphate (S1P). METHODS: The cDNA of EA.hy926 cells was analyzed by RT-PCR to determine the S1P receptor expression profile. The cells were incubated with S1P at different concentrations and for different time intervals. Total RNA and protein in treated EA.hy926 cells were analyzed by RT-PCR and Western blotting. SB203580, a chemical inhibitor of p38 MAPK, was used to determine whether p38 MAPK pathway had any effect on the up-regulation of ZNF580 expression by S1P. The plasmid pEGFP-ZNF580 or the synthetic ZNF580-siRNA was transfected into EA.hy926 cells with Lipofectamine 2000 for 48 h. Cell migration assay and MTT colorimetric assay were used to investigate the effects of ZNF580 on the motility and growth of endothelial cells. RESULTS: EA.hy926 endothelial cells expressed S1P1, S1P3 and S1P5 receptors. Furthermore, S1P up-regulated ZNF580 at mRNA and protein levels in a concentration- and time-dependent manner. The p38 MAPK pathway specific inhibitor SB203580 blocked the S1P-induced up-regulation of ZNF580 expression. Moreover, overexpression/silencing of ZNF580 in EA.hy926 cells led to enhancement/decrease of the migration and proliferation of the cells. CONCLUSION: S1P-induced migration and proliferation of endothelial cells are critical for angiogenesis. ZNF proteins usually play an essential role in altering gene expression and regulating the angiogenesis.     

Effects of resveratrol on TNF-α-induced injury and expression of MCP-1 in isolated rat pulmonary artery endothelial cells

AIM: To investigate the possible mechanism of resveratrol (Res) on tumor necrosis factor-α (TNF-α)-induced monocyte chemoattractant protein-1 (MCP-1) expression in primary rat pulmonary artery endothelial cells (RPAECs).METHODS: RPAECs were randomly divided into 4 groups:control group, solvent (1% DMSO) group, TNF-α group and Res group. Each group was divided into 1 h, 4 h and 8 h subgroups (n=6 per time point). The TNF-α+C1142 (a rodent chimeric mAb that neutralizes rat MCP-1) group was set up at the 8 h time point. At each time point, the protein and mRNA expression of MCP-1 was measured by Western blot and real-time PCR.RESULTS: Pretreatment of the RPAECs with C1142 significantly down-regulated the expression of MCP-1 (P<0.05). The protein and mRNA expression of MCP-1 was markedly increased in TNF-α group (P<0.05). Notably, incubation with Res down-re-gulated the protein and mRNA expression of MCP-1, which was significantly lower than that in TNF-α group (P<0.05).CONCLUSION: MCP-1 was involved in the process of TNF-α-induced injury of RPAECs. Res down-regulates the expression of MCP-1 in RPAECs, thus attenuating cell injury.     

Curcumin reduces neuroinflammation stimulated by Aβ25-35 in primary rat microglial cells

AIM: To investigate the effects of curcumin (Cur) on the expression of High mobility group box 1 protein (HMGB1), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) in amyloid-β (Aβ)-induced primary rat microglial cells. METHODS: Microglia were derived from the cerebral cortices of postnatal rat brains. The cells were identified by immunocytochemistry using mouse anti rat Iba-1 monoclonal antibody. A cell model using primary rat microglial cells incubated with Aβ_25-35 as an inflammation model of Alzheimer's disease (AD) was set up. The morphological characters of primary rat microglial cells were observed. The concentration of Aβ_25-35 and the treatment concentration of curcumin were selected by CCK-8 assay. Cultured primary rat microglial cells were divided into 5 groups:normal cell group, Aβ_25-35 group, Cur group, Aβ_25-35+Cur group and Aβ_25-35+DMSO group. The expression of HMGB1, NF-κB, and receptor for advanced glycation end products (RAGE) was detected by Western blot. The levels of HMGB1, IL-1β, and TNF-α in the culture supernatant were measured by ELISA. RESULTS: The purity of primary microglias determined by Iba-1 immunofluorescence was more than 95%. The protein levels of HMGB1, RAGE and NF-κB were significantly increased after Aβ_25-35 stimulation. After treatment with Cur, the protein levels of HMGB1, RAGE and NF-κB were significantly decreased (P<0.05). The levels of HMGB1, IL-1β and TNF-α in the supernatant were significantly increased after Aβ_25-35 stimulation. Cur significantly decreased the level of HMGB1, IL-1β and TNF-α in the supernatant. CONCLUSION: Curcumin significantly inhibits neuroinflammation stimulated by Aβ_25-35 in primary rat microglial cells.     

Effect of probucol and simvastatin on reactive oxygen species production and HO-1 expression induced by advanced glycation end products in rat renal microvascular endothelial cells

AIM: To investigate the effect of probucol and simvastatin on the production of reactive oxygen species (ROS) and heme oxygenase-1 (HO-1) expression induced by advanced glycation end products (AGEs) in rat renal microvascular endothelial cells (RMECs). METHODS: RMECs isolated and cultured from rat kidney were divided into 4 groups: normal control group, AGEs group, probucol group and simvastatin group. The levels of ROS were determined by the molecular probes of DCFH-DA. The expression of HO-1 at mRNA and protein levels was detected by RT-PCR and Western blotting, respectively. RESULTS: (1) AGEs up-regulated ROS production and HO-1 expression in RMECs. (2) Probucol up-regulated HO-1 expression in RMECs, and inhibited the increasing level of ROS and expression of HO-1 in RMECs induced by AGEs. (3) Simvastatin also inhibited the increasing level of ROS in RMECs induced by AGEs, but it had no effect on HO-1 expression in RMECs with or without AGEs.CONCLUSION: Protective effect of probucol on the dysfunction of RMECs induced by AGEs may be related with its effect on the expression of HO-1 at mRNA and protein levels. Simvastatin also plays roles in antioxidation and renal protection, but is ineffective in the modulation of HO-1 expression.     

Advanced glycation end products-modified protein up-regulates expression of adhesion molecules in human umbilical vein endothelial cells

AIM: Advanced glycation end products (AGEs)-modified proteins are found in plasma and atherosclerosis lesion of diabetes mellitus patients. The study was conducted to elucidate the effect of AGE on expression of adhesion molecules in human endothelial cells. METHODS: Human endothelial cells derived from umbilical veins (HUVECs) were coincubated in vitro with native human serum albumin (HSA) or HSA modified with advanced glycation end products (AGE-HSA). The expression of adhesion molecule intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were determined by immunofluorescent staining and flow cytometry analysis. RESULTS: ICAM-1 and VCAM-1 were constitutively expressed on HUVECs. AGE-HAS enhanced the expression of ICAM-1 and VCAM-1 on HUVECs in a time- and dose-dependent manner. HSA had no effect on the expression of adhesion molecules. CONCLUSIONS: AGE-HSA up-regulates the expression of adhesion molecules in human endothelial cells. AGEs may therefore promote the infiltration of monocytes in the vascular endothelium and have an important effect in the generation and progress of atherosclerosis.     

DENV-2 induces apoptosis of EA.hy926 cells through mitochondrial pathway

AIM：To explore the effect of dengue virus type 2 (DENV-2) infection on the change of mitochondrial membrane potential (Δψm) in EA.hy926 cells. METHODS：The inhibitory effect of DENV-2 infection on EA.hy926 cell growth was examined by MTT assay. The changes of Δψm were analyzed by flow cytometry or observed under fluorescence microscope with JC-1 staining. The activity of caspase-9 was measured by a colorimetric kit. RESULTS：Infection of DENV-2 for 24 h, 36 h and 48 h inhibited the viability of EA.hy926 cells. After DENV-2 infection, the changes of Δψm in EA.hy926 cells were observed. Compared with the normal control cells, Δψm in DENV-2-infected EA.hy926 cells was notably decreased. The activity of caspase-9 increased at early stage after infection of DENV-2 and maintained at a high level at least to 48 h. CONCLUSION：DENV-2 infection decreases the mitochondrial membrane potential and increases the activity of caspase-9 in EA.hy926 cells in the early stage of proliferation, thus promoting the process of apoptosis.     

miR-126 regulates expression of vascular endothelial growth factor in EA.hy926 cells

AIM：To explore the effects of miR-126 on the expression of vascular endothelial growth factor (VEGF) in vascular endothelial cell line EA.hy926. METHODS：EA.hy926 cells were cultured in vitro and transfected with miR-126 mimics or miR-126 inhibitor by cation-mediated transfection method. The total RNA was extracted from the culture cells 36 h after transfection of miR-126 mimics or miR-126 inhibitor. The expression of VEGF at mRNA and protein levels was detected by real-time PCR and Western blotting. RESULTS：Thirty-six hours after transfection of miR-126 inhibitor at concentration of 50 nmol/L, the expression of VEGF at mRNA and protein levels increased significantly as compared with the negative control (P<001). However, transfection of miR-126 mimics at concentration of 50 nmol/L for 36 h significantly decreased the expression of VEGF at mRNA and protein levels as compared with the negative control. CONCLUSION：miR-126 inhibits the expression of VEGF. VEGF may be one of the target genes regulated by miR-126.     

Protective effect of procyanidin single active ingredient B2 on human endothelial progenitor cells under high glucose stimulation

AIM: To study the protective effect of procyanidin single active ingredient B2(PC-B2) on human endothelial progenitor cells(EPCs) stimulated with high glucose. METHODS: The human EPCs were isolated from peripheral blood of healthy people and identified. The EPCs were divided into control group(PBS treatment), hypertonic control group(25 mmol/L mannitol treatment), high glucose(30 mmol/L) group, and different concentrations(2, 10 and 50 mg/L) of PC-B2+30 mmol/L glucose groups. The viability of EPCs was detected by CCK-8 assay. The levels of LDH, MDA, SOD and GSH in the EPCs were detected. The changes of NO, ET-1, ICAM-1 and VCAM-1 in the EPCs cultured medium were measured by ELISA. The cell apoptotic rate and reactive oxygen species(ROS) in the EPCs were analyzed by flow cytometry. The expression of VEGF and VEGFR-2 in the EPCs were determined by Western blot. RESULTS: Compared with control group, the viability of human EPCs was decreased significantly in 30 mmol/L glucose group(P<0.05). The LDH leakage, MDA content and the releases of ET-1, ICAM-1 and VCAM-1 were induced significantly(P<0.05), but SOD and GSH activity and NO production were decreased significantly(P<0.05). The ROS and cell apoptotic rate were increased significantly(P<0.05). The expression of VEGF and VEGFR-2 in the EPCs were decreased(P<0.05). When human EPCs were treated with different concentrations of PC-B2 and 30 mmol/L glucose, the viability was obviously rebounded(P<0.05), the LDH leakage, MDA content and the releases of ET-1, ICAM-1 and VCAM-1 were decreased gradually(P<0.05), the SOD, GSH activity and NO production were increased significantly(P<0.05), the ROS and cell apoptotic rate were decreased significantly(P<0.05), and the expression of VEGF and VEGFR-2 in the EPCs was increased gradually(P<0.05).CONCLUSION: PC-B2 enhances the viability of human EPCs under high glucose condition, reduces high glucose-induced oxidative damage, restores the EPCs normal function, and reduces the releases of inflammatory cytokines and apoptosis, thus playing a protective effect on human EPCs through inducing the expression of VEGF and VEGFR-2.     

Effects of Xijiao Dihuang and Yinqiao San decoction on ICAM-1 and VCAM-1 expression in mouse lung tissues and rat pulmonary microvascular endothelial cells infected with influenza virus

AIM：To investigate the effects of Xijiao Dihuang and Yinqiao San decoction (XDY) on the expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in mouse lung tissues and rat pulmonary microvascular endothelial cells (RPMVECs) infected with influenza virus, and to explore its mechanism for treatment of viral pneumonia. METHODS：Fifty-four male BALB/c mice were randomly divided into normal group, model group and XDY group (n=18 in each group). The viral pneumonia model was established by intranasally dripping influenza A (H1N1) virus into the mice. The mice in XDY group were treated with XDY 1 h after dripping the virus. The expression of ICAM-1 and VCAM-1 in lung tissues was examined by immunohistochemical staining 2, 4 and 6 d after infection. On the other hand, RPMVECs were obtained from male Wistar rats and primarily cultured. The cells were randomly divided into control group, virus group, virus+XDY group, tumor necrosis factor α (TNF-α) group and TNF-α+XDY group. The mRNA and protein expression of ICAM-1 and VCAM-1 was evaluated by real-time PCR and flow cytometry 24 h after infection. RESULTS：Virus exposure increased ICAM-1 and VCAM-1 expression in mouse lung tissues (P<0.01), and XDY treatment attenuated this effect (P<0.01). Virus and TNF-α both led to the increases in mRNA and protein expression of ICAM-1 and VCAM-1 in RPMVECs (P<0.01), which were also reduced by treatment with XDY (P<0.01). CONCLUSION：Treatment with XDY decreases virus-induced ICAM-1 and VCAM-1 expression, suggesting an important role of XDY in treatment of viral pneumonia.     

Effects of Huaiyu decoctum on serum concentrations of TNF-α, IL-6 and IL-10 in rats after anorectal operation

ATM: To investigate the effects of Huaiyu decoctum on the serum concentrations of TNF-α, IL-6 and IL-10 in rats after anorectal operation. METHODS: Sprague-Dawley rats (n=40) were randomly divided into 4 groups:normal group, model group, low-dose Huaiyu decoctum group and high-dose Huaiyu decoctum group. The concentrations of TNF-α, IL-6 and IL-10 in the rat serum were measured by ELISA. The pathologic changes of the anorectal tissues were observed under microscope with HE staining. The protein expression of ICAM-1, VCAM-1 and NF-κB was determined by Western blotting. RESULTS: After Huaiyu decoctum administration, TNF-α and IL-6 concentrations in the serum were significantly decreased, and IL-10 concentration was increased as compared with model group. Moreover, Huaiyu decoctum markedly attenuated edema and hyperemia in the rats after anorectal operation. The protein expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and NF-κB in the anorectal tissues was obviously inhibited by Huaiyu decoctum treatment. CONCLUSION: Huaiyu decoctum improves the recovery of anorectal tissues after operation by decreasing the serum concentrations of TNF-α, IL-6 and IL-10, and reducing the protein expression of ICAM-1, VCAM-1 and NF-κB in the anorectal tissues.     

Effect of water extract propolis on expression of vascular endothelial cell adhesion molecules

AIM: To explore the mechanism of propolis on the inhibition of atherosclerosis and thrombosis in injured human umbilical vascular endothelial cells (HUVECs) induced by tumor necrosis factor alpha (TNF-α)in vitro.METHODS: TNF-α at the concentration of 50 μg/L was used to induce the injury of HUVECs. The injured HUVECs were treated with water extract propolis (WEP) at the concentrations of 50, 100 and 200 mg/L for 6 h, 12 h and 24 h. The expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) was examined by flow cytometry.RESULTS: The expression of ICAM-1 and VCAM-1 was significantly higher in injured HUVECs (P<0.01) than that in the control cells. The expression of ICAM-1 and VCAM-1 was downregulated by WEP treatment in a dose-dependent manner. Between the groups of 100 and 200 mg/L WEP, the difference was significant. In the injured HUVECs treated with 50 mg/L WEP, the inhibitory effect on the expression of ICAM-1 and VCAM-1 was presented in a time-dependent manner. Compared to the single administration, the use of WEP combined with fluvastatin showed better inhibitory effect on the expression of ICAM-1 and VCAM-1 in the injured HUVECs induced by TNF-α (P<0.01).CONCLUSION: WEP may be helpful for the protection of vascular endothelial cells by inhibiting the expression of ICAM-1 and VCAM-1 in a time-and dose-dependent manner. The protective effect of WEP on endothelial cells may be synergic with the inhibitor of HMG-CoA reductase such as fluvastatin sodium.     

Injuring effect of DMSO-soluble particles from cigarette smoke on human umbilical vein endothelial cell line EA. hy 926 in vitro

AIM: To investigate the injuring effect of DMSO-soluble particles from cigarette smoke(DSP) on human umbilical vein endothelial cells. METHODS: Human umbilical vein endothelial cell line EA. hy 926 was used as target cells in the study. The growth and viability of the cells treated with various dosages (1, 2, 4 or 4 mL/L) of DSP and low dose (2 mL/L) of DSP at different time points were evaluated by MTT colorimetric assay and celllular protein assay in 96-well plates. Transmission electron microscopy study was carried out to observe the ultrastructure of human umbilical vein endothelial cells under DSP treatment.RESULTS: DSP inhibited the proliferation of human umbilical vein endothelial cell line EA. hy 926. Under DSP treatment, the reducing cellular protein and increasing cell death(mainly necrosis) were observed in time-dependent and dosage-dependent manners.CONCLUSIONS: These results indicated that the toxic effect of DSP caused functional disturbance and structural damage of human endothelial cells.