Monocyte Chemotactic Protein-1 Production Research Articles

CYLD Deubiquitinates Nicotinamide Adenine Dinucleotide Phosphate Oxidase 4 Contributing to Adventitial Remodeling.

Transdifferentiation of adventitial fibroblasts (AFs) into myofibroblasts plays a critical role during the vascular remodeling that occurs during atherosclerosis, restenosis, and aortic aneurysm. The ubiquitination/deubiquitination regulatory system is essential for the quality control of proteins. The involvement of ubiquitination/deubiquitination during AF transdifferentiation remains largely unknown. In this study, we determined the role of cylindromatosis (CYLD), a deubiquitinase, in the process of AF differentiation and activation in vitro and in vivo. Transforming growth factor-β1 and homocysteine, 2 known inducers of AF transdifferentiation, greatly upregulated CYLD expression in a time- and dose-dependent manner. The silencing of CYLD significantly inhibited AF transdifferentiation and activation as evidenced by the expression of contractile proteins, the production of the proinflammatory cytokines MCP-1 (monocyte chemotactic protein 1) and IL-6 (interleukin-6), the deposition of extracellular matrix, and cell migration. We further asked whether CYLD mediates AF activation via the regulation of nicotinamide adenine dinucleotide phosphate oxidase 4 (Nox4) as it is an essential factor during AF transdifferentiation. Indeed, the silencing of CYLD repressed transforming growth factor-β1-induced and homocysteine-induced Nox4 upregulation and reactive oxygen species production, whereas Nox4 overexpression greatly rescued the inhibitory effect on AF activation by CYLD silencing. Most interestingly, transforming growth factor-β1 and homocysteine repressed Nox4 ubiquitination and prolonged the half-life of Nox4. Moreover, Nox4 was deubiquitinated via a direct interaction with the ubiquitin-specific protease domain of CYLD. In accordance, hyperhomocysteinemia significantly increased adventitial CYLD and Nox4 expression, promoted AF transdifferentiation, and aggravated CaPO4-induced abdominal aortic aneurysm in mice. These effects were abolished in CYLD-/- mice. CYLD contributes to the transdifferentiation of AFs via deubiquitinating Nox4 and may play a role in vascular remodeling.

Beneficial Effects of 6-Month Supplementation with Omega-3 Acids on Selected Inflammatory Markers in Patients with Chronic Kidney Disease Stages 1-3.

Introduction Chronic kidney disease (CKD) is accompanied by inflammation. The aim of this study was to evaluate the effect of 6-month supplementation with omega-3 acids on selected markers of inflammation in patients with CKD stages 1–3. Methods Six-month supplementation with omega-3 acids (2 g/day) was administered to 87 CKD patients and to 27 healthy individuals. At baseline and after follow-up, blood was taken for C-reactive protein (CRP) and monocyte chemotactic protein-1 (MCP-1) concentration and white blood cell (WBC) count. Serum concentration of omega-3 acids—eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and alpha-linolenic acid (ALA)—was determined using gas chromatography. And 24-hour urinary collection was performed to measure MCP-1 excretion. Results After six-month omega-3 supplementation, ALA concentration increased in CKD patients and in the reference group, while EPA and DHA did not change. At follow-up, a significant decrease in urinary MCP-1 excretion in CKD (p = 0.0012) and in the reference group (p = 0.001) was found. CRP, serum MCP-1, and WBC did not change significantly. The estimated glomerular filtration rate (eGFR) did not change significantly in the CKD group. Conclusions The reduction of urinary MCP-1 excretion in the absence of MCP-1 serum concentration may suggest a beneficial effect of omega-3 supplementation on tubular MCP-1 production. Trial Registration This study was registered in ClinicalTrials.gov (identifier: NCT02147002).

Anti-inflammatory Effects of Nortrachelogenin in Murine J774 Macrophages and in Carrageenan-Induced Paw Edema Model in the Mouse.

Nortrachelogenin is a pharmacologically active lignan found in knot extracts of Pinus sylvestris. In previous studies, some lignans have been shown to have anti-inflammatory properties, which made nortrachelogenin an interesting candidate for our study. In inflammation, bacterial products and cytokines induce the expression of inducible nitric oxide synthase, cyclooxygenase-2, and microsomal prostaglandin E synthase-1. These enzymes synthesize factors, which, together with proinflammatory cytokines, are important mediators and drug targets in inflammatory diseases.The effects of nortrachelogenin on the expression of inducible nitric oxide synthase, cyclooxygenase-2, and microsomal prostaglandin E synthase-1 as well as on the production of nitric oxide, prostaglandin E2, and cytokines interleukin-6 and monocyte chemotactic protein-1 were investigated in the murine J774 macrophage cell line. In addition, we examined the effect of nortrachelogenin on carrageenan-induced paw inflammation in mice.Interestingly, nortrachelogenin reduced carrageenan-induced paw inflammation in mice and inhibited the production of inflammatory factors nitric oxide, prostaglandin E2, interleukin-6, and monocyte chemotactic protein-1 in J774 macrophages in vitro. Nortrachelogenin inhibited microsomal prostaglandin E synthase-1 protein expression but had no effect on cyclooxygenase-2 protein levels. Nortrachelogenin also had a clear inhibitory effect on inducible nitric oxide synthase protein expression but none on its mRNA levels, and the proteasome inhibitor lactacystin reversed the effect of nortrachelogenin on inducible nitric oxide synthase expression, suggesting a post-transcriptional mechanism of action.The results revealed hitherto unknown anti-inflammatory properties of nortrachelogenin, which could be utilized in the development of anti-inflammatory treatments.

High fat diet exacerbates Alzheimer's disease-related pathology in APPswe/PS1 mice.

Alzheimer's disease (AD) is mainly characterized by the accumulation and aggregation of amyloid-β (Aβ) peptides in brain parenchyma and cerebral microvasculature. Unfortunately, the exact causes of the disease are still unclear. However, blood-brain barrier (BBB) dysfunction and activation of inflammatory pathways are implicated in AD pathogenesis. Importantly, advanced age and high fat diet, two major risk factors associated with AD, were shown to deeply affect BBB function and modulate the immune response. As such, this study evaluated the impact of age and high fat diet on AD progression. For this purpose, 3 (i.e. young) and 12 (i.e. aged) months old APPswe/PS1 mice were fed for 4 months with a high fat diet (i.e. Western diet (WD)) or normal diet. Interestingly, neurobehavioral tests revealed that WD accelerates age-associated cognitive decline without affecting parenchymal Aβ. Nonetheless, WD decreases matrix metalloproteinase-9 enzymatic activity and brain-derived neurotrophic factor mRNA and protein levels in brain, suggesting loss of synaptic plasticity. In the periphery, WD promotes systemic inflammation by increasing the levels of blood-circulating monocytes and monocyte chemotactic protein-1 production, which is accompanied by an augmentation of oxidized-low density lipoprotein levels in blood circulation. At the BBB, WD potentiates the age-induced increase of Aβ 1-40 accumulation and exacerbates the oxidative stress, specifically in cerebral microvasculature. These effects were accompanied by the dysfunction of pericytes, thus altering BBB functionality without compromising its integrity. Our study provides new insights into the implication of high fat diet in accelerating the cognitive decline in AD.

SIRT1 inhibits inflammatory response partly through regulation of NLRP3 inflammasome in vascular endothelial cells.

Emerging evidence has indicated that vascular endothelial cells (ECs) not only form the barrier between blood and the vessel wall but also serve as conditional innate immune cells. Our previous study found that SIRT1, a class III histone deacetylase, inhibits the inflammatory response in ECs. Recent studies revealed that SIRT1 also participates in the modulation of immune responses. Although the NLRP3 inflammasome is known to be a crucial component of the innate immune system, there is no direct evidence demonstrating the anti-inflammatory effect of SIRT1 on ECs through the NLRP3 inflammasome. In this study, we observed that lipopolysaccharide (LPS) and adenosine triphosphate (ATP) triggered the activation of NLRP3 inflammasome in human umbilical vein ECs (HUVECs). Moreover, SIRT1 expression was reduced in HUVECs stimulated with LPS and ATP. SIRT1 activator inhibited the expression of monocyte chemotactic protein-1 (MCP-1) and C-reactive protein (CRP), whereas SIRT1 knockdown resulted in significant increases in MCP-1 and CRP levels in HUVECs stimulated with LPS and ATP. Importantly, the lack of SIRT1 enhanced NLRP3 inflammasome activation and subsequent caspase-1 cleavage. On the other hand, NLRP3 siRNA blocked the activation of the NLRP3 inflammasome in HUVECs stimulated with LPS plus ATP. Further study revealed that NLRP3 inflammasome blockade significantly reduced MCP-1 and CRP production in HUVECs. In vivo studies indicated that implantation of the periarterial carotid collar inhibited arterial SIRT1 expression in rabbits. Meanwhile, treatment with a SIRT1 activator decreased the expression levels of MCP-1 and CRP in collared arteries and the interleukin (IL)-1β level in serum. Taken together, these findings indicate that NLRP3 inflammasome activation promoted endothelial inflammation and that SIRT1 inhibits the inflammatory response partly through regulation of the NLRP3 inflammasome in ECs.

S100G expression and function in fibroblasts on colitis induction

Supplementation with interleukin (IL)-10, an important anti-inflammatory cytokine, has shown disappointing efficacy for inflammatory bowel diseases (IBD). IL-10 may down-regulate the expression of other anti-inflammatory mediators following colitis induction. We used a colitis model characterized by hapten-protein visualization, which indicates the site of hapten-protein formation after colitis induction for histological and gene expression analyses. Under IL-10 deficiency, following colitis induction inflammatory changes were reduced, and S100G expression was elevated. S100G was expressed in fibroblasts, and S100G expression was down-regulated by IL-10. S100G suppressed the production of monocyte chemotactic protein-1 (MCP-1) through the inhibition of NF-κB activation. Therefore, S100G, also known as Calbindin-D9k, may be an important anti-inflammatory mediator in fibroblasts following colitis induction, and down-regulation of S100G expression might be one reason for the insufficient performance of IL-10 supplementation.

IL-6 stimulates a concentration-dependent increase in MCP-1 in immortalised human brain endothelial cells.

Systemic inflammation is associated with neurodegeneration, with elevated interleukin-6 (IL-6) in particular being correlated with an increased risk of dementia. The brain endothelial cells of the blood brain barrier (BBB) serve as the interface between the systemic circulation and the brain microenvironment and are therefore likely to be a key player in the development of neuropathology associated with systemic inflammation. Endothelial cells are known to require soluble IL-6 receptor (sIL-6R) in order to respond to IL-6, but studies in rat models have shown that this is not the case for brain endothelial cells and studies conducted in human cells are limited. Here we report for the first time that the human cerebral microvascular cell line, hCMVEC, uses the classical mIL-6R signalling pathway in response to IL-6 in a concentration-dependent manner as measured by the production of monocyte chemotactic protein (MCP-1). This novel finding highlights a unique characteristic of human brain endothelial cells and that further investigation into the phenotype of this cell type is needed to elucidate the mechanisms of BBB pathology in inflammatory conditions.

Inhibitory Effects of Tranilast on Cytokine, Chemokine, Adhesion Molecule, and Matrix Metalloproteinase Expression in Human Corneal Fibroblasts Exposed to Poly(I:C)

ABSTRACTPurpose/Aim: Viral infection of the cornea can result in inflammation and scarring and eventually cause blindness. Polyinosinic–polycytidylic acid [poly(I:C)], an analog of viral double-stranded RNA, induces the synthesis of various cytokines, chemokines, adhesion molecules, and matrix metalloproteinases (MMPs) in corneal fibroblasts. The effects of tranilast on the expression of these molecules in human corneal fibroblasts were examined.Materials and methods: Human corneal fibroblasts were cultured with or without poly(I:C) or tranilast. The release of the proinflammatory cytokine interleukin (IL)-6 and of the chemokines IL-8 and monocyte chemotactic protein-1 (MCP-1) was measured with enzyme-linked immunosorbent assays. The expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), MMP-1, and MMP-3 was evaluated by immunoblot or immunofluorescence analysis. The phosphorylation of mitogen-activated protein kinases (MAPKs), c-Jun (a component of the transcription factor AP-1), and IκB-α (an endogenous inhibitor of the transcription factor NF-κB) was examined by immunoblot analysis.Results: Tranilast inhibited in a concentration- and time-dependent manner the production of IL-6, IL-8, MCP-1, ICAM-1, VCAM-1, MMP-1, and MMP-3 by corneal fibroblasts exposed to poly(I:C). It also inhibited the poly(I:C)-induced phosphorylation of c-Jun and the MAPK JNK without affecting that of IκB-α or the MAPKs ERK and p38.Conclusions: Tranilast inhibited proinflammatory cytokine, chemokine, adhesion molecule, and MMP expression in human corneal fibroblasts exposed to poly(I:C), with these effects likely being mediated by attenuation of JNK-AP-1 signaling. Tranilast might therefore be expected to limit immune cell infiltration and stromal degradation associated with viral infection of the cornea.

IL-6 stimulates a concentration-dependent increase in MCP-1 in immortalised human brain endothelial cells

Systemic inflammation is associated with neurodegeneration, with elevated interleukin-6 (IL-6) in particular being correlated with an increased risk of dementia. The brain endothelial cells of the blood brain barrier (BBB) serve as the interface between the systemic circulation and the brain microenvironment and are therefore likely to be a key player in the development of neuropathology associated with systemic inflammation. Endothelial cells are known to require soluble IL-6 receptor (sIL-6R) in order to respond to IL-6, but studies in rat models have shown that this is not the case for brain endothelial cells and studies conducted in human cells are limited. Here we report for the first time that the human cerebral microvascular cell line, hCMVEC, uses the classical mIL-6R signalling pathway in response to IL-6 in a concentration-dependent manner as measured by the production of monocyte chemotactic protein (MCP-1). This novel finding highlights a unique characteristic of human brain endothelial cells and that further investigation into the phenotype of this cell type is needed to elucidate the mechanisms of BBB pathology in inflammatory conditions.

Geniposide attenuates inflammatory response by suppressing P2Y14 receptor and downstream ERK1/2 signaling pathway in oxygen and glucose deprivation-induced brain microvascular endothelial cells

Ethnopharmacological relevanceFructus gardenia is widely used for treatment of stroke and infectious diseases in Chinese medicine. Geniposide is the key bioactive compound related to the pharmacodynamic actions of gardenia on ischemic stroke. The molecular mechanism by which geniposide improves the ischemic brain injury was observed in the study. Aim of the studyRecent studies showed that geniposide had protective activities against the inflammatory response in ischemic stroke. However, the molecular mechanism of geniposide anti-inflammatory role has not yet been fully elucidated. In this study, we investigated the effect of geniposide on the expression of P2Y14 receptor and downstream signaling pathway in brain microvascular endothelial cells (BMECs). Materials and methodsAn in vitro model of cerebral ischemia in BMECs was established by oxygen-glucose-deprivation (OGD). To further confirm the specific effect of geniposide on P2Y14 receptor and downstream signaling pathways, we set up a UDP-glucose (an agonist of the P2Y14 receptor) stimulated model. After administration of geniposide, the expression of P2Y14 receptor, phosphorylation of RAF-1, mitogen activated protein kinase kinase1/2 (MEK1/2), extracellular signal-regulated kinase 1/2 (ERK1/2), level of interleukin-8 (IL-8), interleukin-1β (IL-1β), monocyte chemotactic protein 1 (MCP-1) in BMECs were determined. ResultsThe mRNA and protein expression of P2Y14 in the rat BMECs were up-regulated in OGD-induced injury. After administration of Geniposide, the expression of P2Y14 receptor was significantly down-regulated, the phosphorylation of RAF-1, MEK1/2, ERK1/2 were suppressed. Similar data were obtained in UDP-glc stimulated model. We also observed that geniposide markedly declined the production of IL-8, IL-1β and MCP-1 in OGD-induced BMECs. ConclusionGeniposide exerted anti-inflammatory effects by interfering with the expression of P2Y14 receptor, which subsequently inhibits the downstream ERK1/2 signaling pathways and the release of the pro-inflammatory cytokines IL-8, MCP-1, IL-1β. Therefore, this study provides the evidence for gardenia's clinical application in cerebral ischemia.

IL-6 stimulates a concentration-dependent increase in MCP-1 in immortalised human brain endothelial cells

Systemic inflammation is associated with neurodegeneration, with elevated interleukin-6 (IL-6) in particular being correlated with an increased risk of dementia. The brain endothelial cells of the blood brain barrier (BBB) serve as the interface between the systemic circulation and the brain microenvironment and are therefore likely to be a key player in the development of neuropathology associated with systemic inflammation. Endothelial cells are known to require soluble IL-6 receptor (sIL-6R) in order to respond to IL-6, but studies in rat models have shown that this is not the case for brain endothelial cells and studies conducted in human cells are limited. Here we report for the first time that the human cerebral microvascular cell line, hCMVEC, uses the classical mIL-6R signalling pathway in response to IL-6 in a concentration-dependent manner as measured by the production of monocyte chemotactic protein (MCP-1). This novel finding highlights a unique characteristic of human brain endothelial cells and that further investigation into the phenotype of this cell type is needed to elucidate the mechanisms of BBB pathology in inflammatory conditions.

Murine macrophage response from peritoneal cavity requires signals mediated by chemokine receptor CCR-2 during Staphylococcus aureus infection.

C-C chemokine receptor-2 (CCR-2) is a cognate receptor for monocyte chemotactic protein-1 (MCP-1), and recent studies revealed that MCP-1-CCR-2 signaling is involved in several inflammatory diseases characterized by macrophage infiltration. Currently, there is no study on the involvement of CCR-2 in the killing of S. aureus by macrophages of Swiss albino mice, and its substantial role in host defense against S. aureus infection in murine macrophages is still unclear. Therefore, the present study was aimed to investigate the functional and interactive role of CCR-2 and MCP-1 in regulating peritoneal macrophage responses with respect to acute S. aureus infection. We found that phagocytosis of S. aureus can serve as an important stimulus for MCP-1 production by peritoneal macrophages, which is dependent directly or indirectly on cytokines, reactive oxygen species and nitric oxide. Neutralization of CCR-2 in macrophages leads to increased production of IL-10 and decreased production of IFN-γ and IL-6. In CCR-2 blocked macrophages, pretreatment with specific blocker of NF-κB or p38-MAPK causes elevation in MCP-1 level and subsequent downregulation of CCR-2 itself. We speculate that CCR-2 is involved in S. aureus-induced MCP-1 production via NF-κB or p38-MAPK signaling. We also hypothesized that unnaturally high level of MCP-1 that build up upon CCR-2 neutralization might allow promiscuous binding to one or more other chemokine receptors, a situation that would not occur in CCR-2 non-neutralized condition. This may be the plausible explanation for such observed Th-2 response in CCR-2 blocked macrophages infected with S. aureus in the present study.

IL-22 capacitates dermal fibroblast responses to TNF in scleroderma

ObjectivesInterleukin (IL) 22 mRNA in systemic sclerosis (SSc) skin and Th22 cells in SSc peripheral blood are increased, but the role of IL-22 in fibrosis development remains poorly understood.MethodsBiopsies were...

Attenuation of the macrophage inflammatory activity by TiO₂ nanotubes via inhibition of MAPK and NF-κB pathways.

Biomaterial implantation in a living tissue triggers the activation of macrophages in inflammatory events, promoting the transcription of pro-inflammatory mediator genes. The initiation of macrophage inflammatory processes is mainly regulated by signaling proteins of mitogen-activated protein kinase (MAPK) and by nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathways. We have previously shown that titania nanotubes modified Ti surfaces (Ti/TiO2) mitigate the immune response, compared with flat Ti surfaces; however, little is known regarding the underlying mechanism. Therefore, the aim of this study is to investigate the mechanism(s) by which this nanotopography attenuates the inflammatory activity of macrophages. Thus, we analyzed the effects of TiO2 nanotubes on the activation of MAPK and NF-κB signaling pathways in standard and lipopolysaccharide-evoked conditions. Results showed that the Ti/TiO2 significantly reduce the expression levels of the phosphorylated forms of p38, ERK1/2, c-Jun NH2-terminal kinase (JNK), IKKβ, and IkB-α. Furthermore, a significant reduction in the p65 nuclear accumulation on the nanotubular surface was remarked. Following, by using specific MAPK inhibitors, we observed that lipopolysaccharide-induced production of monocyte chemotactic protein-1 and nitric oxide was significantly inhibited on the Ti/TiO2 surface via p38 and ERK1/2, but not via JNK. However, the selective inhibitor for JNK signaling pathway (SP600125) was effective in reducing tumor necrosis factor alpha release as well as monocyte chemotactic protein-1 and nitric oxide production. Altogether, these data suggest that titania nanotubes can attenuate the macrophage inflammatory response via suppression of MAPK and NF-κB pathways providing a potential mechanism for their anti-inflammatory activity.

Baicalein Inhibits Amadori-Glycated Albumin-Induced MCP-1 Expression in Retinal Ganglion Cells via a MicroRNA-124-Dependent Mechanism.

The purpose of this study was to characterize the inflammatory effect of amadori-glycated albumin (AGA) in cultured rat retinal ganglion cells (RGCs) and to further explore the potential mechanism of the anti-inflammatory effects of baicalein. Primary rat retinal neurons were separated and cultured. The levels of monocyte chemotactic protein-1 (MCP-1) mRNA and soluble MCP-1 produced by RGCs in response to AGA were measured with quantitative reverse transcription-PCR (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA). In addition, the expression of microRNA-124 (miR-124) and histone deacetylases (HDACs) were detected in cultured rat RGCs by qRT-PCR in the presence or absence of baicalein. Luciferase reporter assays were used to validate the regulation of a putative target of miR-124. Amadori-glycated albumin stimulation increased the expression of MCP-1 and inhibited the expression of miR-124 in cultured rat RGCs. In addition, miR-124 directly controlled MCP-1 expression by binding directly to 3'-untranslated region (3'-UTR) of MCP-1. Next, we demonstrated that miR-124 expression was suppressed by HDACs and that treatment of RGCs with HDACs inhibitors increased miR-124 expression and decreased MCP-1 production. Furthermore, application of baicalein in RGCs attenuated AGA-induced MCP-1 expression and upregulated expression of miR-124 by controlling HDAC4 and HDAC5. Collectively, these data suggest that baicalein inhibits AGA-induced MCP-1 expression in retinal ganglion cells via a microRNA-124-dependent mechanism.

Nicotinic acid inhibits vascular inflammation via the SIRT1-dependent signaling pathway

Nicotinic acid (NA) has recently been shown to inhibit inflammatory response in cardiovascular disease. Sirtuin1 (SIRT1), a NAD+-dependent class III histone deacetylase, participates in the regulation of cellular inflammation. We hypothesized that dietary supplementation of NA could attenuate vascular inflammation via modulation of SIRT1 pathway. New Zealand White rabbits received chow or chow supplemented with 0.6% (wt/wt) NA for 2 weeks. Acute vascular inflammation was induced in the animals by placing a non-occlusive silastic collar around the left common carotid artery. At 24 h after collar implantation, the collar-induced production of C-reactive protein and monocyte chemotactic protein-1 was significantly suppressed in the NA-supplemented animals. Meanwhile, NA also decreased the expression of cluster of differentiation 40 (CD40) and CD40 ligand, but up-regulated SIRT1 expression, both in rabbits and in lipopolysaccharide-stimulated endothelial cells. Moreover, knockdown of SIRT1 reversed the inhibitory effect of NA on CD40 expression. Further study revealed that NA also decreased the expression of CD40 partly through mammalian target of rapamycin. These results indicate that NA protects against vascular inflammation via the SIRT1/CD40-dependent signaling pathway.

High Concentrations of TNF-α Induce Cell Death during Interactions between Human Umbilical Cord Mesenchymal Stem Cells and Peripheral Blood Mononuclear Cells

Human umbilical cord mesenchymal stromal cells (hUC-MSCs) are currently being used as novel therapeutic agents in numerous clinical trials. Previous works have shown that hUC-MSCs possess profound immunomodulatory capacities through IL-1 stimulation produced by peripheral blood mononuclear cells (PBMCs), their main cellular partner in most pathophysiological and therapeutic situations. The present study was designed to explore the role of TNF-α in these interactions. In these experiments, we demonstrated that TNF-α originated from PBMCs under the influence of IL-1. We also showed that TNF-α acted differently depending upon the concentrations reached. At low concentrations it clearly contributed to IL-6 and monocyte chemotactic protein 1 (MCP-1) production. At high concentrations, used alone or in association with the TNF-related apoptosis-inducing ligand, TNF-α also stimulated hUC-MSC IL-6 but, more intensely, MCP-1 production. This stimulation was associated but independent of apoptosis induction in a process involving Inhibitor of Apoptosis Proteins. Interferon gamma (IFN-γ), tested to stimulate PBMC and tissue activation, amplified IL-6 and MCP-1 production and cell death by, apparently, a different process involving necrosis. Our findings bring new insights into the complex interactions between hUC-MSCs and PBMCs, involving cytokines, chemokines and cell death, and are of fundamental importance for tissue homeostasis.

Lipopolysaccharide induces inducible nitric oxide synthase-dependent podocyte dysfunction via a hypoxia-inducible factor 1α and cell division control protein 42 and Ras-related C3 botulinum toxin substrate 1 pathway

Urine protein loss in immune complex-mediated diseases such as lupus nephritis is associated with podocyte foot process effacement (podocytopathy) but is not always dependent on glomerular immune complex deposition. Several murine and human studies have associated lupus nephritis with inducible nitric oxide synthase (iNOS) expression in what appear to be podocytes. This study was conducted to determine mechanisms of immune-complex-independent and iNOS-dependent podocyte dysfunction. Conditionally immortalized podocytes were cultured with lipopolysaccharide (LPS) and nitric oxide (NO), superoxide (SO), or peroxynitrite donors in the presence or absence of inhibitors of iNOS, reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase or monocyte chemotactic protein 1 (MCP-1), or with sepiapterin to increase coupling of iNOS homodimers. Podocyte NO, SO, and MCP-1 production and nitrotyrosine modifications were determined. The podocytopathy phenotype was determined by measuring cell motility and membrane permeability to albumin. This study determined that NO produced by iNOS is sufficient and necessary to induce podocytopathy. NO probably induces this phenotype via hypoxia-inducible factor 1α and cell division control protein 42 and Ras-related C3 botulinum toxin substrate 1 pathways. With LPS stimulation, neither SO nor peroxynitrite produced by uncoupled iNOS or NADPH oxidase nor MCP-1 was sufficient to induce the full phenotype. This study supports the notion that iNOS may induce autocrine podocyte dysfunction. Thus, targeting iNOS or the pathways of its induction may have therapeutic benefit.

Proinflammatory cytokines modulate the chemokine CCL2 (MCP-1) in human annulus cells in vitro: CCL2 expression and production

Chemokines are important secondary inflammatory mediators released in response to stimuli which act as second-order cytokines with specialized functions in inflammation. The role of many of these specialized mediators is as yet poorly understood in the human intervertebral disc. Here we investigated CCL2 (chemokine (C–C motif) ligand 2, also known as monocyte chemotactic protein-1 (MCP-1)) in a study of its immunolocalization in disc tissue, and then hypothesized that exposure of cultured human annulus cells to proinflammatory cytokines might alter CCL2 gene expression and CCL2 production. CLL2 was localized to many disc cells in both herniated and non-herniated tissue specimens. Molecular analyses showed that cells exposed to IL-1β showed a 5.5 fold upregulation in CCL2 gene expression vs. controls, p=0.017. Cells exposed to TNF-α showed a 7.7 fold upregulation vs. controls, p=0.005. Cultured cells (grades II–V) showed increased MCP-1 production in IL1-β-treated cells vs. controls (p=0.016), with no significant difference in production in TNF-α-treated cells. Local production of CCL2 in vivo and vitro suggests that annulus cells may be primary effector cells (as well as target cells), with the ability to mediate physiological immune-related processes during disc degeneration by both autocrine and paracrine signaling.

Poly(ADP-ribose) polymerase 1 activation links ischemic acute kidney injury to interstitial fibrosis.

Inactivation of poly(ADP-ribose) polymerase 1 (PARP1) has been found to be protective in several disease models; however, the role of PARP1 in acute kidney injury-induced interstitial fibrosis has not been studied. Herein, we tested whether PARP1 inactivation by treatment with PJ34 (a PARP1 inactivator; 10 mg/kg body weight/day, intraperitoneal implantation of a miniosmotic pump at 2 days after the onset) contributed to the decrease in interstitial fibrosis induced by ischemia-reperfusion injury (IRI) in mouse kidneys. IRI increased PARP1 activation represented by poly(ADP-ribose) expression from 4 to 16 days postinjury, whereas treatment with PJ34 at 2 days after the onset efficaciously abolished the increase in PARP1 activation at 4, 8 and 16 days after IRI. Pharmacological inactivation of PARP1 significantly reduced interstitial fibrosis as represented by the collagen deposition and transforming growth factor-β1 level at 8 and 16 days after IRI. Consistent with collagen deposition, myofibroblast activation represented by α-smooth muscle actin expression was also reduced by PARP1 inactivation at 8 and 16 days after IRI. Furthermore, IRI enhanced macrophage influx, but PARP1 inactivaton remarkably reduced macrophage influx for 4 through 16 days after the injury. Among the chemoattractants for monocytes/macrophages and neutrophils, monocyte chemotactic protein-1 (MCP-1) production in IRI kidneys was significantly reduced by PARP1 inactivation from 4 to 16 days postinjury. These data demonstrate that PARP1 activation contributes to IRI-induced MCP-1 production and in turn to macrophage influx, resulting in the promotion of interstitial fibrosis.