scholarly journals Downregulation of miR-31 in Diabetic Nephropathy and its Relationship with Inflammation

2018 ◽  
Vol 50 (3) ◽  
pp. 1005-1014 ◽  
Author(s):  
Susana Rovira-Llopis ◽  
Irene Escribano-Lopez ◽  
Noelia Diaz-Morales ◽  
Francesca Iannantuoni ◽  
Sandra Lopez-Domenech ◽  
...  
Keyword(s):  
Diabetic Nephropathy ◽  
Pilot Study ◽  
Leukocyte Adhesion ◽  
Flow Chamber ◽  
Leukocyte Rolling ◽  
Rolling Velocity ◽  
Vascular Walls ◽  
Biochemical Evaluation ◽  
Vitro Model

Background/Aims: There is a lack of reliable biological markers for the early diagnosis of diabetic nephropathy (DN) during type 2 diabetes. In this pilot study we aim to assess whether miR-31 levels are modulated by the presence of DN and whether the expression of this miRNA is related to leukocyte-endothelial interactions and inflammation. Methods: Thirty-one T2D patients were enrolled in this pilot study; 18 with no diabetic complications and 13 with diabetic nephropathy. 24 non-diabetic subjects and 13 T2D patients with retinopathy (absent of other complications) were included to test the specificity of miR-31. Following anthropometric and biochemical evaluation, serum miR-31 levels were assessed by Real Time-PCR. Leukocyte-endothelial interactions were evaluated by a parallel flow chamber in vitro model. Serum TNFα, IL-6 and ICAM-1 levels were determined by XMAP-technology in a flow cytometry-based Luminex 200 instrument. Results: Serum miR-31 levels were similar between control and T2D subjects. However, T2D patients with DN displayed reduced levels of miR-31 with respect to patients without complications. This decrease in miR-31 was more pronounced in patients with macroalbuminuria than in those with microalbuminuria and was specific for DN, since patients with retinopathy displayed unaltered miR-31 levels. The presence of DN involved a lower leukocyte rolling velocity and an increased rolling flux and adhesion. miR-31 levels were positively correlated with leukocyte rolling velocity and negatively associated to leukocyte adhesion, TNFα, IL-6 and ICAM-1 levels. Conclusion: Serum miR-31 may be a biomarker for DN in T2D patients. The regulation of this miRNA seems to be related to the recruitment of leukocytes to vascular walls induced by pro-inflammatory and adhesion molecules.

scholarly journals L-Selectin–Dependent Leukocyte Adhesion to Microvascular But Not to Macrovascular Endothelial Cells of the Human Coronary System

Blood ◽  
1997 ◽  
Vol 89 (9) ◽  
pp. 3228-3235 ◽  
Author(s):  
A. Zakrzewicz ◽  
M. Gräfe ◽  
D. Terbeek ◽  
M. Bongrazio ◽  
W. Auch-Schwelk ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Time Course ◽  
Leukocyte Adhesion ◽  
Flow Chamber ◽  
Shear Stresses ◽  
Selectin Ligands ◽  
Tnf Α ◽  
Factor Α

Abstract To characterize L-selectin–dependent cell adhesion to human vascular endothelium, human cardiac microvascular endothelial cells (HCMEC) and human coronary endothelial cells (HCEC) were isolated from explanted human hearts. The adhesion behavior of human (NALM-6) and mouse (300.19) pre-B cells transfected with cDNA encoding for human L-selectin was compared with that of the respective nontransfected cells in a flow chamber in vitro. More than 80% of the adhesion to tumor necrosis factor-α (TNF-α)–stimulated HCMEC at shear stresses <2 dyne/cm2 was L-selectin dependent and could be equally well blocked by an anti–L-selectin antibody or a L-selectin-IgG-chimera. No L-selectin dependent adhesion to HCEC could be shown. The L-selectin dependent adhesion to HCMEC was insensitive to neuraminidase, but greatly inhibited by addition of NaClO3 , which inhibits posttranslational sulfation and remained elevated for at least 24 hours of stimulation. E-selectin dependent adhesion of HL60 cells to HCMEC was blocked by neuraminidase, but not by NaClO3 and returned to control levels within 18 hours of HCMEC stimulation. It is concluded that microvascular, but not macrovascular endothelial cells express TNF-α–inducible sulfated ligand(s) for L-selectin, which differ from known L-selectin ligands, because sialylation is not required. The prolonged time course of L-selectin dependent adhesion suggests a role in sustained leukocyte recruitment into inflammatory sites in vivo.

scholarly journals Long Non-Coding RNA NEAT1 Regulates Pyroptosis in Diabetic Nephropathy via Mediating the miR-34c/NLRP3 Axis

2020 ◽  
Vol 45 (4) ◽  
pp. 589-602 ◽  
Author(s):  
Jin-Feng Zhan ◽  
Hong-Wei Huang ◽  
Chong Huang ◽  
Li-Li Hu ◽  
Wen-Wei Xu
Keyword(s):  
Diabetic Nephropathy ◽  
Target Gene ◽  
Molecular Mechanisms ◽  
Mesangial Cells ◽  
Receptor Protein ◽  
Luciferase Reporter ◽  
Glomerular Mesangial Cells ◽  
Non Coding Rna ◽  
Vitro Model

Introduction: Diabetic nephropathy (DN) is a serious complication of diabetes mellitus and is considered to be a sterile inflammatory disease. Increasing evidence suggest that pyroptosis and subsequent inflammatory response play a key role in the pathogenesis of DN. However, the underlying cellular and molecular mechanisms responsible for pyroptosis in DN are largely unknown. Methods: The rat models of DN were successfully established by single 65 mg/kg streptozotocin treatment. Glomerular mesangial cells were exposed to 30 mmol/L high glucose media for 48 h to mimic the DN environment in vitro. Gene and protein expressions were determined by quantitative real-time PCR and Western blot. Cell viability and pyroptosis were measured by MTT assay and flow cytometry analysis, respectively. The relationship between lncRNA NEAT1, miR-34c, and Nod-like receptor protein-3 (NLRP3) was confirmed by luciferase reporter assay. Results: We found that upregulation of NEAT1 was associated with the increase of pyroptosis in DN models. miR-34c, as a target gene of NEAT1, mediated the effect of NEAT1 on pyroptosis in DN by regulating the expression of NLRP3 as well as the expressions of caspase-1 and interleukin-1β. Either miR-34c inhibition or NLRP3 overexpression could reverse the accentuation of pyroptosis and inflammation by sh-NEAT1 transfection in the in vitro model of DN. Conclusions: Our findings suggested NEAT1 and its target gene miR-34c regulated cell pyroptosis via mediating NLRP3 in DN, providing new insights into understanding the molecular mechanisms of pyroptosis in the pathogenesis of DN.

Preconditioning and adenosine in I/R-induced leukocyte-endothelial cell interactions

1998 ◽  
Vol 274 (4) ◽  
pp. H1230-H1238 ◽  
Author(s):  
Paul Kubes ◽  
Derrice Payne ◽  
Lena Ostrovsky
Keyword(s):  
Endothelial Cell ◽  
Leukocyte Adhesion ◽  
Vascular Dysfunction ◽  
Ischemia Reperfusion ◽  
Microvascular Dysfunction ◽  
Cell Interactions ◽  
Minor Role ◽  
Neutrophil Adhesion ◽  
Rolling Velocity

Recently, it was reported that preconditioning reduced leukocyte adhesion following ischemia-reperfusion (I/R). We further examined the effects of preconditioning and adenosine not only on neutrophil adhesion but also on neutrophil rolling and vascular dysfunction. Intravital microscopy revealed a decrease in neutrophil rolling velocity; a profound increase in neutrophil rolling, adhesion, and microvascular dysfunction; and a reduction in venular shear rates associated with 60 min ischemia and 60 min reperfusion in the feline mesentery. Preconditioning (5 min ischemia/10 min reperfusion) prevented subsequent I/R-induced slow neutrophil rolling, neutrophil adhesion, and microvascular dysfunction but did not affect the flux of rolling neutrophils. Adenosine deaminase A1and A2adenosine-receptor antagonists had only minor effects on the preconditioning responses. Pretreatment of vessels with exogenous adenosine reduced neutrophil adhesion and microvascular permeability and improved neutrophil rolling velocity and shear forces associated with I/R, but the flux of rolling neutrophils was not affected. Finally, in vitro experiments revealed that adenosine had absolutely no direct effect on neutrophil-endothelial cell interactions. In conclusion, our data suggest that adenosine plays only a minor role in preconditioned vessels and that adenosine per se may not directly affect neutrophil-endothelial cell interactions.

Endothelial E- and P-selectin expression in iNOS- deficient mice exposed to polymicrobial sepsis

2001 ◽  
Vol 280 (2) ◽  
pp. G291-G297 ◽  
Author(s):  
Cameron W. Lush ◽  
Gediminas Cepinskas ◽  
William J. Sibbald ◽  
Peter R. Kvietys
Keyword(s):  
Leukocyte Adhesion ◽  
Cecal Ligation ◽  
Leukocyte Rolling ◽  
Wild Type ◽  
Endothelial Adhesion Molecule ◽  
Acute Peritonitis ◽  
Leukocyte Accumulation ◽  
Deficient Mice

In vitro, nitric oxide (NO) decreases leukocyte adhesion to endothelium by attenuating endothelial adhesion molecule expression. In vivo, lipopolysaccharide-induced leukocyte rolling and adhesion was greater in inducible NO synthase (iNOS)−/− mice than in wild-type mice. The objective of this study was to assess E- and P-selectin expression in the microvasculature of iNOS−/− and wild-type mice subjected to acute peritonitis by cecal ligation and perforation (CLP). E- and P-selectin expression were increased in various organs within the peritoneum of wild-type animals after CLP. This CLP-induced upregulation of E- and P-selectin was substantially reduced in iNOS−/− mice. Tissue myeloperoxidase (MPO) activity was increased to a greater extent in the gut of wild-type than in iNOS−/− mice subjected to CLP. In the lung, the reduced expression of E-selectin in iNOS−/− mice was not associated with a decrease in MPO. Our findings indicate that NO derived from iNOS plays an important role in sepsis-induced increase in selectin expression in the systemic and pulmonary circulation. However, in iNOS−/− mice, sepsis-induced leukocyte accumulation is affected in the gut but not in the lungs.

Leukocyte adhesion in local versus hemorrhage-induced ischemia

1992 ◽  
Vol 263 (3) ◽  
pp. H810-H815 ◽  
Author(s):  
M. A. Perry ◽  
D. N. Granger
Keyword(s):  
Endothelial Cell ◽  
Leukocyte Adhesion ◽  
Ischemia Reperfusion ◽  
Leukocyte Rolling ◽  
Leukocyte Adherence ◽  
Rolling Velocity ◽  
Local Ischemia ◽  
Cell Adhesive ◽  
Local Restriction ◽  
Adhesive Interactions

The objective of this study was to compare the leukocyte-endothelial cell adhesive interactions elicited in postcapillary venules by either local ischemia-reperfusion or hemorrhage-reperfusion. Leukocyte rolling, adherence, and emigration were monitored in cat mesenteric venules exposed to an 85% reduction in blood flow (induced by either hemorrhage or local restriction of arterial inflow) for 1 h, followed by 1 h reperfusion. Leukocyte-endothelial cell interactions, venular diameter, and red blood cell velocity were measured during baseline, ischemia, and reperfusion periods. Both local and hemorrhage-induced ischemia reperfusion caused a reduction in leukocyte rolling velocity and increases in leukocyte adherence and emigration. Quantitatively, the adherence and emigration responses in both ischemia models were nearly identical. However, the two models differed in their response to immunoneutralization of the leukocyte adhesion glycoprotein CD11/CD18 with monoclonal antibody (MAb) IB4. The MAb had a more profound effect in attenuating leukocyte adherence and emigration in the local ischemia model. These results indicate that different factors may contribute to leukocyte-endothelial cell adhesive interactions observed in local vs. systemic models of ischemia-reperfusion.

scholarly journals Sialyl Lewisx (sLex) and an sLexMimetic, CGP69669A, Disrupt E-Selectin–Dependent Leukocyte Rolling In Vivo

Blood ◽  
1998 ◽  
Vol 91 (2) ◽  
pp. 475-483 ◽  
Author(s):  
Keith E. Norman ◽  
Gary P. Anderson ◽  
Hartmut C. Kolb ◽  
Klaus Ley ◽  
Beat Ernst
Keyword(s):  
Leukocyte Rolling ◽  
Sialyl Lewisx ◽  
Necrosis Factor Alpha ◽  
Rolling Velocity ◽  
Beneficial Effects ◽  
Selectin Family ◽  
Factor Alpha ◽  
Observable Event

Abstract Leukocyte rolling is the earliest observable event in their recruitment from the circulation to inflamed tissue. This rolling is mediated largely by interaction between the selectin family of adhesion molecules and their glycosylated ligands. Although the nature of these ligands and their interaction with the selectins is not fully understood, it is accepted that expression of fucosylated sialylated glycans such as sialyl Lewisx (sLex) is required for function. Despite findings that sLex inhibits binding of leukocytes to E-selectin in vitro, and has beneficial effects in inflammatory disease models, inhibition of E-selectin–dependent leukocyte rolling in vivo has not been described. Functional overlap between the selectins has been noted and reduction of rolling by E-selectin antibodies only occurs if P-selectin is absent or blocked. We demonstrate that leukocyte rolling velocity in tumor necrosis factor alpha (TNFα)-stimulated mouse cremaster is increased following treatment with either sLex or the sLex-mimetic CGP69669A and that rolling is dramatically reduced if CGP69669A is applied in the presence of anti–P-selectin antibody. These effects are characteristic of E-selectin antagonism. In contrast, surgically stimulated (L- or P-selectin–dependent) rolling is unaffected by either sLex or CGP69669A. Our data demonstrate that CGP69669A is an effective and selective antagonist of E-selectin in vivo.

scholarly journals Mechanical Shedding of L-selectin from the Neutrophil Surface during Rolling on Sialyl Lewis x under Flow

2006 ◽  
Vol 282 (7) ◽  
pp. 4812-4820 ◽  
Author(s):  
Dooyoung Lee ◽  
Joanne B. Schultz ◽  
Philip A. Knauf ◽  
Michael R. King
Keyword(s):  
Flow Chamber ◽  
Mechanical Force ◽  
Mitogen Activated Protein Kinase ◽  
Sialyl Lewis X ◽  
Lewis X ◽  
Rolling Velocity ◽  
Sialyl Lewis ◽  
Mitogen Activated Protein ◽  
Neutrophil Surface

The interaction of L-selectin expressed on leukocytes with endothelial cells leads to capture and rolling and is critical for the recruitment of leukocytes into sites of inflammation. It is known that leukocyte activation by chemoattractants, the change of osmotic pressure in cell media, or cross-linking of L-selectin all result in rapid shedding of L-selectin. Here we present a novel mechanism for surface cleavage of L-selectin on neutrophils during rolling on a sialyl Lewis x-coated surface that involves mechanical force. Flow cytometry and rolling of neutrophils labeled with Qdot®-L-selectin antibodies in an in vitro flow chamber showed that the mechanical shedding of L-selectin occurs during rolling and depends on the amount of shear applied. In addition, the mechanical L-selectin shedding causes an increase in cell rolling velocity with rolling duration, suggesting a gradual loss of L-selectin and is mediated by p38 mitogen-activated protein kinase activation. Thus, these data show that mechanical force induces the cleavage of L-selectin from the neutrophil surface during rolling and therefore decreases the adhesion of cells to a ligand-presenting surface in flow.

scholarly journals TNF-α activation of arterioles and venules alters distribution and levels of ICAM-1 and affects leukocyte-endothelial cell interactions

2006 ◽  
Vol 291 (5) ◽  
pp. H2116-H2125 ◽  
Author(s):  
Ronen Sumagin ◽  
Ingrid H. Sarelius
Keyword(s):  
Spatial Distribution ◽  
Endothelial Cell ◽  
Inflammatory Responses ◽  
Leukocyte Adhesion ◽  
Leukocyte Rolling ◽  
Cremaster Muscle ◽  
Rolling Velocity ◽  
Tnf Α ◽  
Arteriolar Wall ◽  
Microvessel Wall

The observation that leukocyte-endothelial cell (EC) interactions are localized to specific regions on the microvessel wall suggests that adhesion molecule distribution is not uniform. We investigated ICAM-1 distribution and leukocyte-EC interactions in blood-perfused microvessels (<80 μm) in cremaster muscle of anesthetized mice, using intravital confocal microscopy and immunofluorescent labeling. Variability of ICAM-1 expression directly determines leukocyte adhesion distribution within the venular microcirculation and contributes to leukocyte rolling in arterioles during inflammation. The number of rolling interactions increased with ICAM-1 intensity ( r2 = 0.69, P < 0.05), and rolling velocity was lower in regions of higher ICAM-1 intensity. In controls, venular ICAM-1 expression was approximately twofold higher than in arterioles. After TNF-α treatment, ICAM-1 expression was significantly increased, 2.8 ± 0.2-fold in arterioles and 1.7 ± 0.2-fold in venules ( P < 0.05). ICAM-1 expression on activated arteriolar ECs only reached the level of control venular ICAM-1. Arteriolar but not venular ECs underwent redistribution of ICAM-1 among cells; some cells increased and some decreased ICAM-1 expression, magnifying the variability of ICAM-1. TNF-α treatment increased the length of bright fluorescent regions per unit vessel length (42%, control; 70%, TNF-α) along the arteriolar wall, whereas no significant change was observed in venules (60%, control; 63%, TNF-α). The spatial distribution and expression levels of adhesion molecules in the microcirculation determine the timing and placement of leukocyte interactions and hence significantly impact the inflammatory response. That arteriolar ECs respond to TNF-α by upregulation of ICAM-1, although in a different way compared with venules, suggests an explicit role for arterioles in inflammatory responses.

scholarly journals Endothelial PI 3-kinase activity regulates lymphocyte diapedesis

2007 ◽  
Vol 293 (6) ◽  
pp. H3608-H3616 ◽  
Author(s):  
Maryam Nakhaei-Nejad ◽  
Amer M. Hussain ◽  
Qiu-Xia Zhang ◽  
Allan G. Murray
Keyword(s):  
Kinase Activity ◽  
Leukocyte Adhesion ◽  
Flow Chamber ◽  
Regulatory Subunit ◽  
Intercellular Adhesion Molecule ◽  
Intercellular Adhesion Molecule 1 ◽  
Lymphocyte Migration ◽  
Actin Remodeling ◽  
Lymphocyte Adhesion

Lymphocyte recruitment to sites of inflammation involves a bidirectional series of cues between the endothelial cell (EC) and the leukocyte that culminate in lymphocyte migration into the tissue. Remodeling of the EC F-actin cytoskeleton has been observed after leukocyte adhesion, but the signals to the EC remain poorly defined. We studied the dependence of peripheral blood lymphocyte transendothelial migration (TEM) through an EC monolayer in vitro on EC phosphatidylinositol 3-kinase (PI 3-kinase) activity. Lymphocytes were perfused over cytokine-activated EC using a parallel-plate laminar flow chamber. Inhibition of EC PI 3-kinase activity using LY-294002 or wortmannin decreased lymphocyte TEM (48 ± 6 or 34 ± 7%, respectively, vs. control; mean ± SE; P < 0.05). Similarly, EC knockdown of the p85α regulatory subunit of PI 3-kinase decreased lymphocyte transmigration. Treatment of EC with jasplakinolide to inhibit EC F-actin remodeling also decreased lymphocyte TEM to 24 ± 10% vs. control ( P < 0.05). EC PI 3-kinase inhibition did not change the strength of lymphocyte adhesion to the EC or formation of the EC “docking structure” after intercellular adhesion molecule-1 ligation, whereas this was inhibited by jasplakinolide treatment. A similar fraction of lymphocytes migrated on control or LY-294002-treated EC and localized to interendothelial junctions. However, lymphocytes failed to extend processes below the level of vascular endothelial (VE)-cadherin on LY-294002-treated EC. Together these observations indicate that EC PI 3-kinase activity and F-actin remodeling are required during lymphocyte diapedesis and identify a PI 3-kinase-dependent step following initial separation of the VE-cadherin barrier.

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