Cystathionine-β-synthase gene transfer and 3-deazaadenosine ameliorate inflammatory response in endothelial cells

2007 ◽  
Vol 293 (6) ◽  
pp. C1779-C1787 ◽  
Author(s):  
Utpal Sen ◽  
Neetu Tyagi ◽  
Munish Kumar ◽  
Karni S. Moshal ◽  
Walter E. Rodriguez ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Inflammatory Response ◽  
Protein Expression ◽  
Adhesion Molecules ◽  
Adhesion Molecule ◽  
Vascular Remodeling ◽  
Collagen Type ◽  
Gene Transfection ◽  
Collagen Type 1

Although elevated levels of homocysteine (Hcy) known as hyperhomocysteinemia (HHcy) are associated with increased inflammation and vascular remodeling, the mechanism of Hcy-mediated inflammation and vascular remodeling is unclear. The matrix metalloproteinases (MMPs) and adhesion molecules play an important role in vascular remodeling. We hypothesized that HHcy induces inflammation by increasing adhesion molecules and matrix protein expression. Endothelial cells were supplemented with high methionine, and Hcy accumulation was measured by HPLC. Nitric oxide (NO) bioavailability was detected by a NO probe. The protein expression was measured by Western blot analysis. MMP-9 activity was detected by gelatin-gel zymography. We demonstrated that methionine supplement promoted upregulation of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) through increased Hcy accumulation. In addition, increased synthesis of collagen type-1 was also observed. MMP-9 gene expression and protein activity were increased in methionine supplement groups. 3-Deazaadenosine (DZA), an adenosine analogue, prevented high methionine-induced ICAM-1 and VCAM-1 expression and collagen type-1 synthesis. Transfection of endothelial cells with cystathionine-β-synthase (CBS) gene construct, which converts Hcy to cystathionine, reduced Hcy accumulation in high methionine-fed cells. CBS gene transfection reduced the inflammatory response, as evident by attenuated ICAM-1 and VCAM-1 expression. Furthermore, collagen type-1 expression and MMP-9 activity were dramatically attenuated with CBS gene transfection. These results suggested that methionine supplement increased Hcy accumulation, which was associated with inflammatory response and matrix remodeling such as collagen type-1 synthesis and MMP-9 activity. However, in vitro DZA and CBS gene therapy successfully treated the HHcy-induced inflammatory reaction in the methionine metabolism pathway.

scholarly journals 1,25-Dihydroxyvitamin D3 Reduces TGF-β3-Induced Fibrosis-Related Gene Expression in Human Uterine Leiomyoma Cells

2011 ◽  
Vol 96 (4) ◽  
pp. E754-E762 ◽  
Author(s):  
Sunil K. Halder ◽  
J. Shawn Goodwin ◽  
Ayman Al-Hendy
Keyword(s):  
Protein Expression ◽  
Vitamin D3 ◽  
Uterine Leiomyoma ◽  
Collagen Type ◽  
Plasminogen Activator Inhibitor ◽  
Plasminogen Activator Inhibitor 1 ◽  
Dihydroxyvitamin D3 ◽  
Collagen Type 1 ◽  
Activator Inhibitor

Abstract Background: Uterine leiomyomas (fibroids) are the most common benign estrogen-dependent tumors of premenopausal women. TGF-β3 up-regulates the synthesis of many of extracellular matrix proteins that are associated with tissue fibrosis. Objective: To examine the effect of 1,25-dihydroxyvitamin D3 (vitamin D3) on TGF-β3-induced fibrosis-related protein expression in immortalized human uterine leiomyoma (HuLM) cells. Methods: HuLM cells were treated with TGF-β3 with or without vitamin D3. Western blot analyses were employed to test the effect of vitamin D3 on TGF-β3-induced protein expression of collagen type 1, fibronectin, and plasminogen activator inhibitor-1 proteins. Western blots as well as immunofluorescence analyses were used to verify the effect of vitamin D3 on TGF-β3-induced Smad activation involved in extracellular matrix protein synthesis and deposition, which ultimately lead to tissue fibrosis. Results: We observed that TGF-β3 induced fibronectin and collagen type 1 protein expression in HuLM cells, and that effect was suppressed by vitamin D3. TGF-β3 also induced protein expression of plasminogen activator inhibitor-1, an important TGF-β target, in HuLM cells, which was also inhibited by vitamin D3. Additionally, TGF-β3 induced phosphorylation of Smad2 as well as nuclear translocation of Smad2 and Smad3 in HuLM cells, whereas vitamin D significantly reduced all these TGF-β3-mediated effects. Therefore, our results suggest that vitamin D3 has consistently reduced TGF-β3 effects that are involved in the process of fibrosis in human leiomyoma cells. Conclusion: Vitamin D3 is an antifibrotic factor that might be potentially useful as a novel therapeutic for nonsurgical treatment of benign uterine fibroids.

Abstract 452: Serine Carboxypeptidase 1 Mediates Vascular Remodeling by Increasing Inflammatory Cell Infiltration

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Ting-Hein Lee ◽  
Joseph Miano
Keyword(s):  
Endothelial Cells ◽  
Cell Adhesion ◽  
Adhesion Molecules ◽  
Adhesion Molecule ◽  
Vascular Remodeling ◽  
Cell Adhesion Molecule ◽  
Inflammatory Cell ◽  
Catalytic Triad ◽  
Serine Carboxypeptidase ◽  
Cell Adhesion Molecule 1

In pathological vascular remodeling, contractile vascular smooth muscle cells (VSMCs) switch their phenotype to highly proliferative and migratory states leading to neointimal formation. Inflammatory cell recruitment and infiltration, which is dependent on the increased expression of adhesion molecules on the endothelial cells, is a key event to initiate SMC phenotypic modulation in vascular remodeling. Serine carboxypeptidase 1 (scpep1), a novel protease containing the putative catalytic triad (Ser-Asp-His) common to all members of the serine protease family, has been proved to be involved in vascular remodeling by promoting SMC proliferation and migration in a catalytic triad-dependent manner. To determine whether Scpep1 modulates leukocyte adhesion and infiltration, a flow-induced model of vascular remodeling was conducted in wild-type (WT) or Scpep1 knockout (KO) mice. Scpep1-null mice show a decreased number of infiltrated leukocytes into the intima and media compared to WT mice. Further, mice devoid of Scpep1 show a dramatic reduction of vascular cell adhesion molecule-1 (VCAM-1) and intercellular cell adhesion molecule-1 (ICAM-1) expression in vessels in comparison with that of WT mice. Consistent with our in vivo data, the expression levels of ICAM-1 and VCAM-1 on human umbilical vein endothelial cells (HUVECs) transfected with SiRNA against Scpep1 were significantly decreased after TNF-α treatment. Taken together, these data suggest that Scpep1 may increase leukocyte extravasation by increasing the expression of VCAM-1 and ICAM-1 adhesion molecules.

scholarly journals Direct comparison of 3D and 2D cultivation reveals higher osteogenic capacity of elderly osteoblasts in 3D

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Stephan Payr ◽  
Elizabeth Rosado-Balmayor ◽  
Thomas Tiefenboeck ◽  
Tim Schuseil ◽  
Marina Unger ◽  
...  
Keyword(s):  
Gene Expression ◽  
Collagen Type ◽  
3D Culture ◽  
Osteogenic Potential ◽  
Donor Age ◽  
Collagen Type 1 ◽  
2D And 3D ◽  
Human Primary Osteoblasts ◽  
Gene Expression Levels

Abstract Background The aim of this study was the investigation of the osteogenic potential of human osteoblasts of advanced donor age in 2D and 3D culture. Methods Osteoblasts were induced to osteogenic differentiation and cultivated, using the same polystyrene material in 2D and 3D culture for 2 weeks. Samples were taken to evaluate alkaline phosphatase (ALP) activity, mineralization and gene expression. Results Osteoprotegerin (OPG) levels were significantly increased (8.2-fold) on day 7 in 3D compared to day 0 (p < 0.0001) and 11.6-fold higher in 3D than in 2D (p < 0.0001). Both culture systems showed reduced osteocalcin (OC) levels (2D 85% and 3D 50% of basic value). Collagen type 1 (Col1) expression was elevated in 3D on day 7 (1.4-fold; p = 0.009). Osteopontin (OP) expression showed 6.5-fold higher levels on day 7 (p = 0.002) in 3D than in 2D. Mineralization was significantly higher in 3D on day 14 (p = 0.0002). Conclusion Advanced donor age human primary osteoblasts reveal significantly higher gene expression levels of OPG, Col1 and OP in 3D than in monolayer. Therefore, it seems that a relatively high potential of bone formation in a natural 3D arrangement is presumably still present in osteoblasts of elderly people. Trial registration 5217/11 on the 22nd of Dec. 2011.

Adenosine inhibits cytokine release and expression of adhesion molecules by activated human endothelial cells

1996 ◽  
Vol 270 (2) ◽  
pp. C522-C529 ◽  
Author(s):  
M. G. Bouma ◽  
F. A. van den Wildenberg ◽  
W. A. Buurman
Keyword(s):  
Endothelial Cells ◽  
Adhesion Molecules ◽  
Adhesion Molecule ◽  
Vascular Endothelium ◽  
Cell Activation ◽  
Umbilical Vein ◽  
Intercellular Adhesion Molecule ◽  
Cytokine Release ◽  
Adenosine Deaminase Activity ◽  
Anti Inflammatory

Ischemia induces excessive ATP catabolism with subsequent local release of its metabolite adenosine, an autacoid with anti-inflammatory properties. Because activation of the vascular endothelium is critical to the inflammatory host response during ischemia and reperfusion, the effects of adenosine on two major determinants of endothelial cell activation (i.e., the release of proinflammatory cytokines and the expression of adhesion molecules) were studied. Adenosine dose dependently inhibited the release of interleukin (IL)-6 and IL-8 by stimulated human umbilical vein endothelial cells (HUVEC). Expression of E-selectin and vascular cell adhesion molecule 1 (VCAM-1), but not intercellular adhesion molecule 1 (ICAM-1), by activated HUVEC was also reduced by adenosine. Inhibition of endogenous adenosine deaminase activity by erythro-9-(2-hydroxy-3-nonyl)adenine or 2'-deoxycoformycin strongly enhanced the inhibitory effects of exogenous adenosine on cytokine release and expression of E-selectin and VCAM-1. However, a clear role for specific adenosine receptors in the described inhibitory events could not be established. Together, these data imply that the vascular endothelium constitutes an important target for the anti-inflammatory actions of adenosine.

scholarly journals Evaluation of Collagen Type-1 Production and Anti-Inflammatory Activities of Human Placental Extracts in Human Gingival Fibroblasts

2016 ◽  
Vol 25 (3) ◽  
pp. 277-281 ◽  
Author(s):  
Homare Akagi ◽  
Yasuhiro Imamura ◽  
Yoshimasa Makita ◽  
Hiroe Nakamura ◽  
Naomi Hasegawa ◽  
...  
Keyword(s):  
Collagen Type ◽  
Human Gingival Fibroblasts ◽  
Gingival Fibroblasts ◽  
Anti Inflammatory ◽  
Collagen Type 1

scholarly journals Evaluation of Secretome Tenogenic Potential from Adipose Stem Cells (ACS) in Hypoxic Condition with Fresh Frozen Tendon Scaffold Using Scleraxis (Scx), Insulin-Like Growth Factor 1 (IGF-1) and Collagen Type 1

2021 ◽  
Vol 49 ◽  
pp. 111-118
Author(s):  
Ramadhan Ananditia Putra ◽  
Heri Suroto
Keyword(s):  
Cell Culture ◽  
Collagen Type ◽  
Hypoxic Condition ◽  
Adipose Stem Cells ◽  
Immunohistochemical Examination ◽  
Hypoxic Conditions ◽  
Fresh Frozen ◽  
Collagen Type 1

Various studies have been conducted to see the scaffold that supports the regeneration of tendon. This study aims to analyze thein vitrosecretome tenogenic potential produced by ASCs culture with fresh frozen tendon scaffold in hypoxic conditions. ELISA tests for Scx and IGF-1 levels in secretome were obtained from ASC culture with fresh frozen tendon scaffold under normoxic (21%) and hypoxia (2%) conditions. The immunohistochemical examination of COL-1 was also carried out on the 2ndand 6thdays of cell culture. The secretion of Scx and IGF-1 was increased in secretome from ASC cultures using a fresh frozen tendon scaffold compared with those which did not (p <0.05). In the normoxia condition, Scx and IGF-1 in secretome with fresh frozen tendons had better results than hypoxic conditions (p <0.05). The highest Scx levels were obtained in culture on the 6thday (p <0.05), while the highest IGF-1 levels were obtained in the culture on the 2ndday (p <0.05). There was an increase in the secretion of Scx and IGF-1 from ASC cultures with fresh frozen tendon scaffold under the hypoxic condition of 2%.

Hypoxia-induced expression of complement receptor type 1 (CR1, CD35) in human vascular endothelial cells

1999 ◽  
Vol 276 (2) ◽  
pp. C450-C458 ◽  
Author(s):  
Charles D. Collard ◽  
Cuneyt Bukusoglu ◽  
Azin Agah ◽  
Sean P. Colgan ◽  
Wende R. Reenstra ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Protein Expression ◽  
Vascular Endothelial Cells ◽  
Umbilical Vein ◽  
Receptor Type ◽  
Complement Receptor ◽  
Complement Receptor Type ◽  
Umbilical Vein Endothelial Cells ◽  
Complement Receptor Type 1

Reoxygenation of hypoxic human umbilical vein endothelial cells (HUVECs) increases protein expression of the complement regulators CD46 and CD55. As the receptor for C3b is known to be present on injured bovine endothelial cells, we investigated whether hypoxia or inflammatory mediators induce complement receptor type 1 (CR1; CD35) expression on HUVECs. CR1 protein expression increased 3.7 ± 0.6-fold as measured by ELISA on HUVECs following hypoxia (48 h, 1% O2). Colocalization of CD35 and von Willebrand factor by confocal microscopy confirmed that CD35 was predominantly intracellular. Lipopolysaccharide or tumor necrosis factor-α also significantly increased HUVEC CR1 protein expression. Western blot analysis of neutrophil or hypoxic HUVEC lysates revealed a 221-kDa CR1 band under nonreducing conditions. RT-PCR of hypoxic HUVEC mRNA revealed a single band that, after sequencing, was identified as CD35. In situ hybridization of hypoxic HUVECs, but not normoxic HUVECs or fibroblasts, demonstrated increased CD35 mRNA. Hypoxic HUVECs bound immune complexes and acted as a cofactor for factor I-mediated cleavage of C3b. Thus hypoxia induces functional HUVEC CR1 expression.

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