scholarly journals Acid sphingomyelinase promotes SGK1-dependent vascular calcification

2021 ◽  
Vol 135 (3) ◽  
pp. 515-534
Author(s):  
Trang Thi Doan Luong ◽  
Rashad Tuffaha ◽  
Mirjam Schuchardt ◽  
Barbara Moser ◽  
Nadeshda Schelski ◽  
...  
Keyword(s):  
Vascular Calcification ◽  
Ex Vivo ◽  
Acid Sphingomyelinase ◽  
Upstream Regulator ◽  
Key Factor ◽  
Complex Signaling ◽  
High Phosphate

Abstract In chronic kidney disease (CKD), hyperphosphatemia is a key factor promoting medial vascular calcification, a common complication associated with cardiovascular events and high mortality. Vascular calcification involves osteo-/chondrogenic transdifferentiation of vascular smooth muscle cells (VSMCs), but the complex signaling events inducing pro-calcific pathways are incompletely understood. The present study investigated the role of acid sphingomyelinase (ASM)/ceramide as regulator of VSMC calcification. In vitro, both, bacterial sphingomyelinase and phosphate increased ceramide levels in VSMCs. Bacterial sphingomyelinase as well as ceramide supplementation stimulated osteo-/chondrogenic transdifferentiation during control and high phosphate conditions and augmented phosphate-induced calcification of VSMCs. Silencing of serum- and glucocorticoid-inducible kinase 1 (SGK1) blunted the pro-calcific effects of bacterial sphingomyelinase or ceramide. Asm deficiency blunted vascular calcification in a cholecalciferol-overload mouse model and ex vivo isolated-perfused arteries. In addition, Asm deficiency suppressed phosphate-induced osteo-/chondrogenic signaling and calcification of cultured VSMCs. Treatment with the functional ASM inhibitors amitriptyline or fendiline strongly blunted pro-calcific signaling pathways in vitro and in vivo. In conclusion, ASM/ceramide is a critical upstream regulator of vascular calcification, at least partly, through SGK1-dependent signaling. Thus, ASM inhibition by repurposing functional ASM inhibitors to reduce the progression of vascular calcification during CKD warrants further study.

Circulating uromodulin inhibits vascular calcification by interfering with pro-inflammatory cytokine signalling

2020 ◽  
Author(s):  
Ioana Alesutan ◽  
Trang T D Luong ◽  
Nadeshda Schelski ◽  
Jaber Masyout ◽  
Susanne Hille ◽  
...  
Keyword(s):  
Vascular Calcification ◽  
Serum Levels ◽  
Vascular Pathology ◽  
Adeno Associated Virus ◽  
Aortic Smooth Muscle ◽  
Key Factor ◽  
Factor Α

Abstract Aims Uromodulin is produced exclusively in the kidney and secreted into both urine and blood. Serum levels of uromodulin are correlated with kidney function and reduced in chronic kidney disease (CKD) patients, but physiological functions of serum uromodulin are still elusive. This study investigated the role of uromodulin in medial vascular calcification, a key factor associated with cardiovascular events and mortality in CKD patients. Methods and results Experiments were performed in primary human (HAoSMCs) and mouse (MOVAS) aortic smooth muscle cells, cholecalciferol overload and subtotal nephrectomy mouse models and serum from CKD patients. In three independent cohorts of CKD patients, serum uromodulin concentrations were inversely correlated with serum calcification propensity. Uromodulin supplementation reduced phosphate-induced osteo-/chondrogenic transdifferentiation and calcification of HAoSMCs. In human serum, pro-inflammatory cytokines tumour necrosis factor α (TNFα) and interleukin-1β (IL-1β) co-immunoprecipitated with uromodulin. Uromodulin inhibited TNFα and IL-1β-induced osteo-/chondrogenic signalling and activation of the transcription factor nuclear factor kappa-light-chain-enhancer of activated β cells (NF-kB) as well as phosphate-induced NF-kB-dependent transcriptional activity in HAoSMCs. In vivo, adeno-associated virus (AAV)-mediated overexpression of uromodulin ameliorated vascular calcification in mice with cholecalciferol overload. Conversely, cholecalciferol overload-induced vascular calcification was aggravated in uromodulin-deficient mice. In contrast, uromodulin overexpression failed to reduce vascular calcification during renal failure in mice. Carbamylated uromodulin was detected in serum of CKD patients and uromodulin carbamylation inhibited its anti-calcific properties in vitro. Conclusions Uromodulin counteracts vascular osteo-/chondrogenic transdifferentiation and calcification, at least in part, through interference with cytokine-dependent pro-calcific signalling. In CKD, reduction and carbamylation of uromodulin may contribute to vascular pathology.

The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

2012 ◽  
Vol 82 (3) ◽  
pp. 228-232 ◽  
Author(s):  
Mauro Serafini ◽  
Giuseppa Morabito
Keyword(s):  
Antioxidant Capacity ◽  
Dietary Intervention ◽  
Ex Vivo ◽  
The Body ◽  
Dietary Polyphenols ◽  
Enzymatic Antioxidant ◽  
Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

scholarly journals Heparan sulfate and heparin interactions with proteins

2015 ◽  
Vol 12 (110) ◽  
pp. 20150589 ◽  
Author(s):  
Maria C. Z. Meneghetti ◽  
Ashley J. Hughes ◽  
Timothy R. Rudd ◽  
Helena B. Nader ◽  
Andrew K. Powell ◽  
...  
Keyword(s):  
Heparan Sulfate ◽  
Ex Vivo ◽  
Sequence Specificity ◽  
Structure Activity ◽  
Multiple Binding Sites ◽  
Multiple Binding ◽  
Heparin Activity

Heparan sulfate (HS) polysaccharides are ubiquitous components of the cell surface and extracellular matrix of all multicellular animals, whereas heparin is present within mast cells and can be viewed as a more sulfated, tissue-specific, HS variant. HS and heparin regulate biological processes through interactions with a large repertoire of proteins. Owing to these interactions and diverse effects observed during in vitro , ex vivo and in vivo experiments, manifold biological/pharmacological activities have been attributed to them. The properties that have been thought to bestow protein binding and biological activity upon HS and heparin vary from high levels of sequence specificity to a dependence on charge. In contrast to these opposing opinions, we will argue that the evidence supports both a level of redundancy and a degree of selectivity in the structure–activity relationship. The relationship between this apparent redundancy, the multi-dentate nature of heparin and HS polysaccharide chains, their involvement in protein networks and the multiple binding sites on proteins, each possessing different properties, will also be considered. Finally, the role of cations in modulating HS/heparin activity will be reviewed and some of the implications for structure–activity relationships and regulation will be discussed.

scholarly journals The pathophysiologic role of the protein kinase Cδ pathway in the intervertebral discs of rabbits and mice: In vitro, ex vivo, and in vivo studies

2012 ◽  
Vol 64 (6) ◽  
pp. 1950-1959 ◽  
Author(s):  
Michael B. Ellman ◽  
Jae-Sung Kim ◽  
Howard S. An ◽  
Jeffrey S. Kroin ◽  
Xin Li ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Ex Vivo ◽  
Intervertebral Discs ◽  
In Vivo Studies ◽  
Protein Kinase Cδ

Abstract P283: Parkin Mediates Mitophagy in Cardioprotection

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Allen M Andres ◽  
Chengqun Huang ◽  
Eric P Ratliff ◽  
Genaro Hernandez ◽  
Pamela Lee ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Wild Type ◽  
Simulated Ischemia ◽  
Selective Targeting ◽  
Cardioprotective Effects ◽  
Mitochondrial Turnover ◽  
First Time

Autophagy-dependent mitochondrial turnover in response to cellular stress is necessary for maintaining cellular homeostasis. However, the mechanisms that govern the selective targeting of damaged mitochondria are poorly understood. Parkin, an E3 ubiquitin ligase, has been shown to be essential for the selective clearance of damaged mitochondria. Parkin is expressed in the heart, yet its function has not been investigated in the context of cardioprotection. We previously reported that autophagy is required for cardioprotection by ischemic preconditioning (IPC). In the present study, we used simulated ischemia in vitro and IPC in hearts (in vivo and ex vivo) to investigate the role of Parkin in mediating cardioprotection. In HL-1 cells, simulated ischemia induced Parkin translocation to mitochondria and mitochondrial elimination. Mitochondrial loss was blunted in Atg5-deficient cells, revealing the requirement for autophagy in mitochondrial elimination. Consistent with previous reports implicating p62/SQSTM1 in mitophagy, we found that downregulation of p62 attenuated mitophagy and exacerbated cell death in HL-1 cardiomyocytes subjected to simulated ischemia. While wild type mice showed p62 translocation to mitochondria after IPC, Parkin knockout mice exhibited attenuated translocation of p62 to mitochondria. Importantly, ablation of Parkin in mice abolished the cardioprotective effects of IPC. These results reveal for the first time the crucial role of Parkin and mitophagy in cardioprotection.

scholarly journals ROLE OF NITRIC OXIDE (NO) IN CAPSAICIN MEDIATED ANTI-PLATELET ACTIVITY IN IN VITRO, IN VIVO, EX-VIVO MODEL OF PLATELET AGGREGATION ASSAY AND ARTERIAL THROMBOSIS IN RAT: POTENTIAL THERAPEUTIC TARGET?

2018 ◽  
Vol 10 (4) ◽  
pp. 44
Author(s):  
Mihir K Patel ◽  
Kiranj K. Chaudagar ◽  
Anita A. Mehta
Keyword(s):  
Platelet Aggregation ◽  
Arterial Thrombosis ◽  
Ex Vivo ◽  
Platelet Activity ◽  
Aggregation Assay ◽  
Thrombosis Model ◽  
Platelet Aggregation Assay

Objective: Although recent advances in the treatment of congestive heart disease, mortality among patients’ remains a questionable remark. Therefore, we evaluated the role of capsaicin on in vitro and ex vivo platelet aggregation induced by Adenosine Di-Phosphate (ADP) as well as in in vivo thrombosis models and role of NO, KATP was also identified in the capsaicin-induced anti-platelet animal model as well as in vivo model of arterial thrombosis.Methods: According to body weight wistar rats were divided into five groups. Group I and Group II was treated with saline and capsaicin (3 mg/kg, i. v), while animals from Group III were treated with N(ω)-nitro-L-arginine methyl ester (L-NAME) (30 mg/kg, i. v) 30 min before administration of capsaicin (3 mg/kg, i. v). Group IV animals were treated with glibenclamide (10 mg/kg,i. v) 30 min before administration of capsaicin (3 mg/kg, i. v). Group V was considered as a positive control and administered clopidogrel (30 mg/kg, p. o). Animals were subjected for in vitro, ex-vivo platelet aggregation assay. ADP (30µM) was utilized as an aggregating agent in these experiments. After these assays; animals of each group were subjected for subaqueous tail bleeding time in a rat model and FeCl3-induced arterial thrombosis model in rats.Results: In ADP-induced in vitro platelet aggregation, a significant reduction in % platelet aggregation was observed at 50µM (64.35±4.641) and 100µM (52.72±4.192) concentration of capsaicin as compared to vehicle control (85.82±3.716). Capsaicin (3 mg/kg, i. v) also showed a significant reduction (49.53±4.075) in ex-vivo ADP-induced platelet aggregation as compared to vehicle control (89.38±2.057). In FeCl3 induced arterial thrombosis model, Capsaicin (3 mg/kg, i. v) exhibited an increase in time to occlusion in this rodent model and presence of the L-NAME and glibenclamide had inhibited the activity of capsaicin.Conclusion: In our study, capsaicin (50 µM, 100µM) exhibited potent anti-platelet activity in ADP-induced platelet aggregation, similarly capsaicin exhibited significant anti-platelet action in the ex-vivo study. Moreover, the presence of L-NAME and glibenclamide inhibited the anti-thrombotic and anti-platelet action of capsaicin. Therefore, it was concluded that NO and KATP may be involved in the anti-thrombotic action of capsaicin.

scholarly journals In Vivo Role of Dendritic Cells in a Murine Model of Pulmonary Cryptococcosis

2006 ◽  
Vol 74 (7) ◽  
pp. 3817-3824 ◽  
Author(s):  
Karen L. Wozniak ◽  
Jatin M. Vyas ◽  
Stuart M. Levitz
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Cell Activation ◽  
Cell Activation ◽  
Ex Vivo ◽  
Interleukin 2 ◽  
Mouse Lung

ABSTRACT Dendritic cells (DC) have been shown to phagocytose and kill Cryptococcus neoformans in vitro and are believed to be important for inducing protective immunity against this organism. Exposure to C. neoformans occurs mainly by inhalation, and in this study we examined the in vivo interactions of C. neoformans with DC in the lung. Fluorescently labeled live C. neoformans and heat-killed C. neoformans were administered intranasally to C57BL/6 mice. At specific times postinoculation, mice were sacrificed, and lungs were removed. Single-cell suspensions of lung cells were prepared, stained, and analyzed by microscopy and flow cytometry. Within 2 h postinoculation, fluorescently labeled C. neoformans had been internalized by DC, macrophages, and neutrophils in the mouse lung. Additionally, lung DC from mice infected for 7 days showed increased expression of the maturation markers CD80, CD86, and major histocompatibility complex class II. Finally, ex vivo incubation of lung DC from infected mice with Cryptococcus-specific T cells resulted in increased interleukin-2 production compared to the production by DC from naïve mice, suggesting that there was antigen-specific T-cell activation. This study demonstrated that DC in the lung are capable of phagocytosing Cryptococcus in vivo and presenting antigen to C. neoformans-specific T cells ex vivo, suggesting that these cells have roles in innate and adaptive pulmonary defenses against cryptococcosis.

Temperature-dependence of the DnaA–DNA interaction and its effect on the autoregulation of dnaA expression

2012 ◽  
Vol 449 (2) ◽  
pp. 333-341 ◽  
Author(s):  
Chiara Saggioro ◽  
Anne Olliver ◽  
Bianca Sclavi
Keyword(s):  
Temperature Dependence ◽  
Binding Sites ◽  
Dna Interaction ◽  
Bacterial Dna ◽  
Dnaa Protein ◽  
High Affinity ◽  
Key Factor

The DnaA protein is a key factor for the regulation of the timing and synchrony of initiation of bacterial DNA replication. The transcription of the dnaA gene in Escherichia coli is regulated by two promoters, dnaAP1 and dnaAP2. The region between these two promoters contains several DnaA-binding sites that have been shown to play an important role in the negative auto-regulation of dnaA expression. The results obtained in the present study using an in vitro and in vivo quantitative analysis of the effect of mutations to the high-affinity DnaA sites reveal an additional effect of positive autoregulation. We investigated the role of transcription autoregulation in the change of dnaA expression as a function of temperature. While negative auto-regulation is lost at dnaAP1, the effects of both positive and negative autoregulation are maintained at the dnaAP2 promoter upon lowering the growth temperature. These observations can be explained by the results obtained in vitro showing a difference in the temperature-dependence of DnaA–ATP binding to its high- and low-affinity sites, resulting in a decrease in DnaA–ATP oligomerization at lower temperatures. The results of the present study underline the importance of the role for autoregulation of gene expression in the cellular adaptation to different growth temperatures.

scholarly journals Metformin-stimulated AMPK-α1 promotes microvascular repair in acute lung injury

2013 ◽  
Vol 305 (11) ◽  
pp. L844-L855 ◽  
Author(s):  
Ming-Yuan Jian ◽  
Mikhail F. Alexeyev ◽  
Paul E. Wolkowicz ◽  
Jaroslaw W. Zmijewski ◽  
Judy R. Creighton
Keyword(s):  
Endothelial Cells ◽  
Acute Lung Injury ◽  
Lung Injury ◽  
Ex Vivo ◽  
Endothelial Permeability ◽  
Beneficial Effects ◽  
Isolated Lung

Acute lung injury secondary to sepsis is a leading cause of mortality in sepsis-related death. Present therapies are not effective in reversing endothelial cell dysfunction, which plays a key role in increased vascular permeability and compromised lung function. AMP-activated protein kinase (AMPK) is a molecular sensor important for detection and mediation of cellular adaptations to vascular disruptive stimuli. In this study, we sought to determine the role of AMPK in resolving increased endothelial permeability in the sepsis-injured lung. AMPK function was determined in vivo using a rat model of endotoxin-induced lung injury, ex vivo using the isolated lung, and in vitro using cultured rat pulmonary microvascular endothelial cells (PMVECs). AMPK stimulation using N1-(α-d-ribofuranosyl)-5-aminoimidizole-4-carboxamide or metformin decreased the LPS-induced increase in permeability, as determined by filtration coefficient ( Kf) measurements, and resolved edema as indicated by decreased wet-to-dry ratios. The role of AMPK in the endothelial response to LPS was determined by shRNA designed to decrease expression of the AMPK-α1 isoform in capillary endothelial cells. Permeability, wounding, and barrier resistance assays using PMVECs identified AMPK-α1 as the molecule responsible for the beneficial effects of AMPK in the lung. Our findings provide novel evidence for AMPK-α1 as a vascular repair mechanism important in the pulmonary response to sepsis and identify a role for metformin treatment in the management of capillary injury.

scholarly journals Inhibition of Gastrin-Releasing Peptide Attenuates Phosphate-Induced Vascular Calcification

Cells ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 737 ◽  
Author(s):  
Hyun-Joo Park ◽  
Yeon Kim ◽  
Mi-Kyoung Kim ◽  
Jae Joon Hwang ◽  
Hyung Joon Kim ◽  
...  
Keyword(s):  
Vascular Calcification ◽  
Vascular System ◽  
Ex Vivo ◽  
Calcium Deposition ◽  
Gastrin Releasing Peptide ◽  
Cardiovascular Morbidity And Mortality ◽  
Potential Strategy ◽  
Grp Receptor

Vascular calcification is the pathological deposition of calcium/phosphate in the vascular system and is closely associated with cardiovascular morbidity and mortality. Here, we investigated the role of gastrin-releasing peptide (GRP) in phosphate-induced vascular calcification and its potential regulatory mechanism. We found that the silencing of GRP gene and treatment with the GRP receptor antagonist, RC-3095, attenuated the inorganic phosphate-induced calcification of vascular smooth muscle cells (VSMCs). This attenuation was caused by inhibiting phenotype change, apoptosis and matrix vesicle release in VSMCs. Moreover, the treatment with RC-3095 effectively ameliorated phosphate-induced calcium deposition in rat aortas ex vivo and aortas of chronic kidney disease in mice in vivo. Therefore, the regulation of the GRP-GRP receptor axis may be a potential strategy for treatment of diseases associated with excessive vascular calcification.

Sign in / Sign up

Export Citation Format

Share Document