Toxicology
Volume 336,
2 October 2015
, Pages 70-78
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https://doi.org/10.1016/j.tox.2015.08.003Get rights and content
Abstract
Placental angiogenesis is dramatically increased during pregnancy in association with the elevated placental blood flows to support the rapidly growing fetus. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is an environmental toxicant and a ligand of aryl hydrocarbon receptor (AhR). Herein, we investigated the effects of TCDD on proliferation, migration, and viability of fetoplacental endothelial cells in response to a complete growth medium which contained serum and growth supplement using human umbilical cord vein (HUVECs) and artery (HUAECs) cells as models. We found that TCDD dose- and time-dependently inhibited (p<0.05) proliferation of HUVECs and HUAECs. Treatment with TCDD at 10nM for 6 days inhibited (p<0.05) migration (by ∼30%) of HUAECs, but not HUVECs. TCDD at 10nM also decreased (p<0.05) viability of HUVECs and HUAECs. Interestingly, specific AhR siRNA blocked (p<0.05) the TCDD-inhibited cellular responses in HUAECs, but not HUVECs. Nonetheless, TCDD at 10nM neither affected the cell cycle progression, nor did it induce cell apoptosis in HUVECs and HUAECs. In addition, TCDD at 10nM also did not alter activation of ERK1/2 and AKT1 in HUVECs and HUAECs. Collectively, TCDD suppresses proliferation and/or migration (two key steps of angiogenesis) of HUVECs and HUAECs independent and dependent of AhR, respectively. These data suggest that TCDD inhibited growth of HUVECs and HUAECs via decreasing cell viability. Thus, TCDD may inhibit fetoplacental angiogenesis, leading to negative pregnancy outcomes.
Introduction
During pregnancy, fetoplacental angiogenesis is dramatically increased in association with the elevated placental blood flows to support the rapidly growing fetus (Magness and Zheng, 1996, Reynolds et al., 2005, Wang and Zheng, 2012). In addition, it is well established that defective placental angiogenesis is linked to severe pregnancy complications such as preeclampsia and intrauterine growth restriction, leading to negative pregnancy outcomes (Gilbert et al., 2008, Khankin et al., 2010, Mayhew et al., 2004). Thus, any factor that impairs fetoplacental angiogenesis will jeopardize pregnancy and fetal development.
2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a ubiquitous environmental pollutant and toxicant, which belongs to the halogenated aromatic hydrocarbons family (Denison et al., 2011, Pocar et al., 2005). Exposure of humans to TCDD could cause severe intoxication including chloracne, hepatotoxicity, neurotoxicity, and cardiotoxicity (Denison et al., 2011, Pelclová et al., 2006, Pocar et al., 2005). Humans are primarily exposed to TCDD by ingesting contaminated food and during industrial accidents (Pelclová et al., 2006, Pocar et al., 2005). After ingested, TCDD is fairly stably stored in the fatty acid and resistant to metabolic degradation, potentially posing chronic adverse effects on humans (Denison et al., 2011, Pelclová et al., 2006, Pocar et al., 2005). In the general population, average TCDD equivalents (TEQ) concentrations are estimated at 58ng/kg serum lipid, corresponding to 13ng TEQ/kg body weight (DeVito et al., 1995). TCDD concentrations in human infant tissues (0.16–0.38ng/kg lipid of adipose tissue and 0.29–0.49ng/kg lipid of liver) (Kreuzer et al., 1997), which are believed to be lower than or comparable to those in adults (Kreuzer et al., 1997, Leung et al., 1990, Neubert, 1988). However, when humans are accidentally exposed to moderate to high levels of TCDD, the serum lipid TCDD concentrations can reach much high levels, ranging from 130 to 144,000ppt (Aylward et al., 2005).
During pregnancy, TCDD can easily cross the placental barrier and enter the fetus (Bell et al., 2007, Nau and Bass, 1981). Thus, repeated prenatal exposure to TCDD can induce fetotoxicity, leading to increased prenatal mortality in many species including mouse, rat, monkey, and perhaps human too (Hernández-Ochoa et al., 2009, Peterson et al., 1993). Importantly, the TCDD-induced fetotoxicity may be partially attributed to abnormal vascular formation, growth, and function. For example, prenatal exposure of chicks to TCDD impairs coronary vasculogenesis in chicken embryos by attenuating responsiveness of embryos to angiogenic factors including vascular endothelial growth factor-A (VEGFA; Ivnitski-Steele et al., 2005). In addition, TCDD exposure suppresses vascular remodeling in the labyrinth zone of rat placentas (Ishimura et al., 2006, Wu et al., 2014) and also induces vascular inflammatory response, contributing to the development of atherosclerosis in adult mice (Wu et al., 2011).
The majority of TCDD’s activities is mediated via activation of aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor (Denison et al., 2011). While most of unliganded AhR resides in the cytoplasm of cells (Denison et al., 2011), upon binding to its ligands, AhR will transfer to the nucleus, where it binds to aryl hydrocarbon nuclear receptor translocator to form a heterodimer (Denison et al., 2011). This heterodimer subsequently binds to the dioxin response element, activating expression of its downstream genes such as CYP1A1 (cytochrome P450, family 1, subfamily A, polypeptide 1) and CYP1B1 (cytochrome P450, family 1, subfamily B, polypeptide 1) (Denison et al., 2011). The AhR-induced downstream signaling also includes the MEK/ERK1/2 and PI3K/AKT1 pathways (Tan et al., 2002, Tan et al., 2004, Wu et al., 2007). After activation, AhR is translocated back to the cytoplasm and rapidly degraded by the proteasome (Denison et al., 2011). In addition, AhR activation can also be suppressed by the AhR repressor (Denison et al., 2011). Intriguingly, TCDD may exert its actions independent of AhR (Ahmed et al., 2005, Kondraganti et al., 2003, Park et al., 2003, Park et al., 2005, Sanders et al., 2005). For example, TCDD induces an antiproliferative response in human breast cancer cells and reduces expression of p16ink4a (a cell cycle regulator and tumor suppressor) in human endothelial cells via an AhR independent manner (Akintobi et al., 2007, Yoshioka et al., 2012).
We have recently reported that AhR is present in human placentas, primarily in endothelial cells within villous cords, and in endothelial cells of umbilical cord veins, and arteries (Jiang et al., 2010). We have also shown that one dose of TCDD at 1.6μg/kg body weight interferes with vascular remodeling of rat placentas (Wu et al., 2014). However, the role of TCDD in human fetoplacental angiogenesis is not known, although 3-methylcholanthrene, another exogenous AhR ligand, has been shown to inhibit cell proliferation of human umbilical vascular endothelial cells in vitro (Juan et al., 2006). Therefore, in this study, we hypothesize that TCDD inhibits the angiogenic responses of fetoplacental endothelial cells via AhR, which may ultimately lead to adverse pregnancy outcomes. Specifically, we examined the effect of TCDD on the fetoplacental endothelial proliferation and migration, two key steps of angiogenesis, in response to a complete growth medium containing 10% FBS and growth supplement using human umbilical vein (HUVECs) and artery (HUAECs) endothelial cells as cell models since endothelial cells in vein and artery are different tremendously in their global gene expression profiles (Chi et al., 2003, Jiang et al., 2013a, Jiang et al., 2013b), possibly leading to different responses to TCDD. We have also attempted to explore the mechanisms underlying TCDD’s effects on these cells.
Section snippets
Cell isolation and culture
HUVECs and HUAECs, two widely used endothelial cell models for fetoplacental endothelial cells were isolated from umbilical cords of patients with normal term pregnancies using a standard collagenase enzyme digestion as described (Jiang et al., 2013a,b). After isolation, cells were cultured in basal RPMI 1640 media (BM; Life Technology, Grand Island, NY) supplemented with 10% FBS (Thermo Scientific, Waltham, MA), 1% penicillin/streptomycin, 100mg/L heparin (EMD Chemicals, San Diego, CA), and
TCDD inhibits proliferation of HUVECs and HUAECs, and migration of HUAECs
As compared to the vehicle control (DMSO), TCDD dose- and time-dependently significantly inhibited (p<0.05) proliferation of HUVECs and HUAECs induced by CGM (Fig. 1A and B). In both HUVECs and HUAECs, TCDD at 10 and 100nM caused a maximal inhibition on Days 4 and 6, respectively, whereas TCDD at 0.1nM had no effects on Days 2, 4, and 6. On Day 6, TCDD at 100nM decreased (p<0.05) proliferation of HUVECs and HUAECs by 40% and 35%, respectively. The relative IC50 values for HUVECs and HUAECs on
Discussion
In this study, we have demonstrated that TCDD inhibits proliferation of HUVECs and HUAECs and migration of HUAECs grown in a complete growth medium without significantly altering cell progression and apoptosis. These TCDD-induced inhibitory actions are associated with a decrease in cell viability. Intriguingly, these TCDD-inhibited cellular responses are mediated independent and dependent of AhR in HUVECs and HUAECs, respectively. These data suggest that TCDD may attenuate proliferation and/or
Conclusion
In conclusion, our current data indicate that TCDD differentially suppresses angiogenic responses of HUVECs and HUAECs, although the underlying signaling mechanisms remain elusive. Moreover, since that placental angiogenesis is vital for the placental growth (Magness and Zheng, 1996, Reynolds et al., 2005, Wang and Zheng, 2012), and TCDD can easily cross the placenta barrier (Bell et al., 2007, Nau and Bass, 1981), maternal exposure of exogenous AhR ligands such as TCDD may impair the
Conflict of interest
The authors declare that there is no conflict of interest.
Acknowedgements
This work was supported, in part, by the US National Institutes of Health Grants HD38843 (JZ & RRM), an R & D grant from Departmental of Ob/Gyn., University of Wisconsin-Madison (JZ), and the National Natural Science Foundation of China No. 81100429 (KW).
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See AlsoLncRNA EPIC1 protects human osteoblasts from dexamethasone-induced cell deathA synergistic negative effect of gestational smoke-exposure and small litter size on rat placental efficiency, vascularisation and angiogenic factors mRNA expression - DocumentToxicol. Lett.
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Polychlorinated biphenyls, polybrominated biphenyls, polychlorinated dibenzo-p-dioxins, and polychlorinated dibenzofurans
2022, Reproductive and Developmental Toxicology
Polychlorinated biphenyls (PCBs), polybrominated biphenyls, polychlorinated dibenzo-p-dioxins, and polychlorinated dibenzofurans belong to a group of compounds that are structurally related and are environmentally and biologically persistent. These chemicals have a tendency to bioaccumulate and biomagnify in the food chain. Residues of these chemicals have been detected in remote areas of the world and in a variety of animal species, including humans. Exposure to these chemicals has been linked to a broad spectrum of effects. Fetal and early developmental exposures are particularly sensitive and can have more profound and irreversible outcomes when compared to adult exposure. Latent effects of early exposures include, but are not limited to, depressed circulating thyroid hormone levels and abnormal thyroid cytology; developmental effects of the heart, palate, and kidney; delayed cognitive development; altered sensory and motor abilities; and reproductive impairment and compromised neuronal function. Although AhR activation has been attributed to several dioxin-like coplanar compounds, some PCBs that are noncoplanar in nature seem to exert their toxic effects through different mechanisms including calcium signaling, oxidative stress, thyroid hormone perturbations, and neurotransmitter imbalance. While certain congeners and isomers can pose a very serious threat to the health of animals and humans, environmental exposure situations are generally such that risks of health effects are generally low. The most significant problem by these compounds involved in accidental poisoning via food supply or consumption of contaminated food from contaminated areas. Additionally, there are areas of the environment that are heavily contaminated by these chemicals because of past industrial activities. Animals and humans residing in or near contaminated locations certainly are at risk of serious health effects. Efforts must continue to reduce exposure to protect wildlife and humans. The best way to accomplish is to modernize technological processes to prevent the release of these chemicals into the environment.
2,7-Dibromocarbazole interferes with tube formation in HUVECs by altering Ang2 promoter DNA methylation status
2019, Science of the Total Environment
Citation Excerpt :
Thus, Ang2 could work as an angiogenic agonist (Fiedler and Augustin, 2006). TCDD-induced abnormal vascular formation, growth, and function could result in fetotoxicity in many species including mice, rats, monkeys, and perhaps humans (Li et al., 2015; Hernández-Ochoa et al., 2009; Peterson et al., 1993). 2,7-DBCZ has also been reported to cause dioxin-like toxicity and induce cardiovascular anormogenesis in zebrafish (Ji et al., 2019; Fang et al., 2016).
2,7-Dibromocarbazole (2,7-DBCZ) is one of the most frequently detected polyhalogenated carbazoles (PHCZs) in the environmental media. 2,7-DBCZ has attracted public attention for its potential for dioxin-like toxicity and cardiovascular toxicity. However, researches on the potential mechanism of angiogenesis inhibition by 2,7-DBCZ is still insufficient. Herein, human umbilical vein endothelial cells (HUVECs) were applied to explore the angiogenic effect of 2,7-DBCZ and the potential underlying mechanisms. 2,7-DBCZ significantly inhibited tube formation in HUVECs in the non-toxic concentration range. PCR array showed that 2,7-DBCZ reduced the expression proportion between VEGFs and Ang2, thereby inhibiting tube formation in HUVECs. Then, small RNA interference and DNA methylation assays were adopted to explore the potential mechanisms. It has been found that angiopoietin2 (Ang2)-silencing recovered the tube formation inhibited by 2,7-DBCZ. The DNA methylation status of Ang2 promoter also showed a demethylation tendency after exposure. In conclusion, 2,7-DBCZ could demethylate the Ang2 promoter to potentiate Ang2 expression, thus altering angiogenic phenotype of HUVECs by reducing the proportion between Ang2 and VEGFs. The data presented here can help to guide safety measures on the use of dioxin-like PHCZs for their potential adverse effects and provide a method for identifying the relevant biomarkers to assess their cardiovascular toxicity.
Effects of simvastatin on nuclear receptors, drug metabolizing enzymes and transporters expression in Human Umbilical Vein Endothelial Cells
2018, Pharmacological Reports
Citation Excerpt :
In our study HUVEC treatment with TCDD resulted in a significant decrease of AhR protein level compared with the control cells. These observations are in good agreement with other studies [27]. Furthermore, to confirm functional AhR activation, the enzymatic activity of CYP1A1/1B1 was analyzed.
Vascular endothelial cells (EC) are constantly exposed to endo- and exogenous compounds, which may disturb EC function. One of the protecting mechanisms against chemicals consists of drug metabolizing enzymes and transporter proteins regulated by nuclear receptors and transcription factors. Therefore, the aim of the current study was to assess the regulation of nuclear receptors and their coordinated genes in Human Umbilical Vein Endothelial Cells (HUVEC).
HUVEC were exposed to TCDD (10nM), oltipraz (100μM) and simvastatin (1μM) for 24h. Gene expressions were evaluated using quantitative real-time PCR. The protein expression levels were determined by Western blotting. Enzymatic activity of CYP1A1/CYP1B1 was assessed by luciferin-labelled CYPs substrate.
Our study confirmed that nuclear receptor AhR and nuclear factor Nrf2 are highly expressed in HUVECs. Treatment of HUVECs with TCDD (AhR inducer) resulted in a significant induction of AHR target genes CYP1A1, CYP1B1 and NQO1. Oltipraz (Nrf2 inducer) also markedly increased expression of NQO1 but did not affect Nrf2 mRNA nor protein levels. Under simvastatin stimulation PXR and NRF2 target transcripts were not altered, however AHR-regulated genes: CYP1A1, CYP1B1 and MDR1 were significantly induced. Western blot analysis confirmed CYP1B1 induction in TCDD-treated HUVECs, but not in the simvastatin group. Moreover, HUVEC exposure to TCDD resulted in induction of CYP1A1/CYP1B1 enzymatic activity.
This study revealed functional expression of AhR and Nrf2 in HUVECs. Moreover, it was defined that simvastatin induced AhR and its related genes.
Polychlorinated Biphenyls, Polybrominated Biphenyls, Polychlorinated Dibenzo-p-Dioxins, and Polychlorinated Dibenzofurans
2018, Veterinary Toxicology: Basic and Clinical Principles: Third Edition
Polychlorinated biphenyls (PCBs), polybrominated biphenyls (PBBs), polychlorinated dibenzo-p-dioxins (PCDDs), and polychlorinated dibenzofurans (PCDFs) belong to a group of compounds that are structurally related and are biologically and environmentally persistent. These chemicals can bioaccumulate and biomagnify in the food chain. Exposure to these chemicals has been linked to a broad spectrum of effects. Fetal and early developmental exposures are particularly sensitive, and can have different outcomes from adult exposure. Latent effects of early exposures include, but are not limited to, depressed circulating thyroid hormone (TH) levels and abnormal thyroid cytology; developmental effects of the heart, palate, and kidney; delayed cognitive development; altered sensory and motor abilities; and reproductive impairment and compromised neuronal function. Although Aryl Hydrocarbon Receptor (AhR) activation has been attributed to the adverse effects of several dioxin-like coplanar compounds, some PCBs that are noncoplanar in nature seem to exert their toxic effects through different mechanisms, including calcium/protein kinase C signaling, oxidative stress, TH perturbations, and neurotransmitter imbalance. The most significant problem with these compounds involved accidental poisoning via food supply and consumption of contaminated food indicating oral ingestion was a major route of exposure. Additionally, there are areas of the environment that are heavily contaminated by these chemicals because of past industrial activities. Animals and humans residing in or near contaminated locations certainly are at risk of serious health effects. Efforts must continue to reduce exposure to protect wildlife, veterinary animals, and humans. The best way to accomplish this is to modernize technological processes to prevent the release of these chemicals into the environment.
Environmental Pollutants on Angiogenesis and Vascular Development
2018, Comprehensive Toxicology: Third Edition
Angiogenesis is one of the major mechanisms of neovascularization to form newly sprouting capillaries from the originally existing microvessels, which covers a broad spectrum of physiological and pathophysiological processes from tissue hypertrophy and wound healing to inflammation and even cancer. A recent report estimates that a quarter of the global burden of disease was due to environmental risk factors, and environmental exposures are increasingly recognized to be strongly associated with major human systemic diseases, such as cardiovascular disorders and cancers, especially in susceptible populations. Therefore, angiogenesis and its regulatory factors, along with major environmental exposure factors, such as cigarette smoke, ambient air pollutants, and some major carcinogens, are reviewed.
Toxicity to the Insulin-Secreting β-Cell
2018, Comprehensive Toxicology: Third Edition
The incidence of diabetes in humans is increasing in many countries around the world. In order to minimize the risk of diabetes in our population, it is important to identify and understand the mechanisms that trigger the disease. The Introduction to the article provides a brief overview of the common types of diabetes and how β-cell demise contributes to this. This is followed by an introduction to the normal and abnormal morphology of the endocrine pancreas, the processes of insulin synthesis and secretion, along with commonly used research tools to study diabetes. The bulk of this article will largely focus on the toxicology of the pancreatic β-cells and the mechanism(s) contributing to β-cell demise. While the cause of diabetes has historically been largely attributed to a sedentary culture and poor diet, there appears to be a set of “nontraditional” contributions from medication use, smoking, and environmental contaminants which we have also extensively described.
Research article
Social interactions modulate the virulence of avian malaria infection
International Journal for Parasitology, Volume 43, Issue 10, 2013, pp. 861-867
There is an increasing understanding of the context-dependent nature of parasite virulence. Variation in parasite virulence can occur when infected individuals compete with conspecifics that vary in infection status; virulence may be higher when competing with uninfected competitors. In vertebrates with social hierarchies, we propose that these competition-mediated costs of infection may also vary with social status. Dominant individuals have greater competitive ability than competing subordinates, and consequently may pay a lower prevalence-mediated cost of infection. In this study we investigated whether costs of malarial infection were affected by the occurrence of the parasite in competitors and social status in domestic canaries (Serinus canaria). We predicted that infected subordinates competing with non-infected dominants would pay higher costs than infected subordinates competing with infected dominants. We also predicted that these occurrence-mediated costs of infection would be ameliorated in infected dominant birds. We found that social status and the occurrence of parasites in competitors significantly interacted to change haematocrit in infected birds. Namely, subordinate and dominant infected birds differed in haematocrit depending on the infection status of their competitors. However, in contrast to our prediction, dominants fared better with infected subordinates, whereas subordinates fared better with uninfected dominants. Moreover, we found additional effects of parasite occurrence on mortality in canaries. Ultimately, we provide evidence for costs of parasitism mediated by social rank and the occurrence of parasites in competitors in a vertebrate species. This has important implications for our understanding of the evolutionary processes that shape parasite virulence and group living.
Research article
Bisphenol AF stimulates transcription and secretion of C-X-C chemokine ligand 12 to promote proliferation of cultured T47D breast cancer cells
Toxicology, Volume 338, 2015, pp. 30-36
Bisphenol AF (4,4′-hexafluoroisopropylidene-2-diphenol, BPAF), an endocrine disruptor, has been shown to stimulate the proliferation of human breast cancer cells. However, the underlying mechanism has not been fully elucidated. We found that BPAF promoted the in vitro proliferation of estrogen receptor α (ERα)-positive breast cancer cells (T47D and MCF7), but not ERα-negative cells (MDA-MB-231 and MDA-MB-435s). BPAF significantly stimulated the proliferation of cultured T47D cell in a dose-dependent manner, and the half-maximal effective concentration (EC50) was approximately 123nM. We employed lentivirus-mediated short hairpin RNA (shRNA) to knockdown ERα and ER antagonist ICI 182780 to inhibit ER activation, which resulted in the repression of BPAF-induced proliferation of T47D and MCF7 cells. We observed that C-X-C chemokine ligand 12 (CXCL12) was up-regulated in T47D cells under treatment with BPAF. Quantitative real-time PCR results showed that BPAF caused a time and dose dependent increase in mRNA level of CXCL12. Furthermore, treatment of T47D cells with BPAF increased CXCL12 secretion according to ELISA assay. BPAF-induced CXCL12 transcription and secretion was significantly attenuated by small interfering RNA (siRNA) targeting ERα and ICI 182780, indicating BPAF-induced CXCL12 expression is mediated through ERα. Notably, knockdown CXCL12 in T47D cells significantly attenuated BPAF-induced cell proliferation. We also observed that inhibition of CXCL12 binding to its receptors CXCR4 and CXCR7 by chalcone 4 blocked BPAF-induced cell growth. Our results indicated that CXCL12 facilitated BPAF-induced proliferation of T47D cells. Taken together, our data provided support that BPAF stimulated transcription and secretion of CXCL12 depending on ERα, and ERα/CXCL12 signaling positively regulated BPAF-induced proliferation of cultured T47D breast cancer cells.
Research article
Urinary bisphenol A concentrations and cytochrome P450 19 A1 (Cyp19) gene expression in ovarian granulosa cells: An in vivo human study
Reproductive Toxicology, Volume 42, 2013, pp. 18-23
Exposure to bisphenol A (BPA), a chemical widely used in consumer products, has been associated with in vitro Cyp19 gene expression.
To evaluate an in vivo human model of Cyp19 gene expression in granulosa cells.
A subset of an ongoing prospective cohort study of women undergoing in vitro fertilization (IVF) at Massachusetts General Hospital.
Mixed effect models were used to evaluate the association of urinary BPA concentrations with granulosa cell Cyp19 mRNA expression.
In 61 women undergoing 76 IVF cycles, adjusted changes in mean Cyp19 expression (β estimate (95% CI)) for quartiles 2, 3 and 4 as compared to the lowest quartile were: −0.97 (−2.22, 0.28); −0.97 (−2.18, 0.24) and −0.38 (−1.58, 0.82).
An in vivo model for evaluation of Cyp19 gene expression was developed for use in epidemiologic studies. In this pilot study, we found no statistically significant linear association between urinary BPA concentrations and Cyp19 expression.
Research article
Clinical significance of Tim3-positive T cell subsets in patients with multiple sclerosis
Journal of Clinical Neuroscience, Volume 34, 2016, pp. 193-197
The present study evaluated associations between the percentages of T cell immunoglobulin and mucin domain 3 (Tim3)-positive T cells and related cytokines and multiple sclerosis (MS). We collected peripheral blood samples from 30 MS patients and 30 healthy controls. Flow cytometry was used to determine the proportions of CD3+Tim3+, CD4+Tim3+, and CD4+CD25+Tim3+ in peripheral blood mononuclear cells (PBMCs) and related cell subsets. The serum concentrations of galectin-9, IL-17, and IFN-γ also were determined using enzyme-linked immunosorbent assays (ELISA). The percentages of Tim3-positive T cells in CD4+ and CD4+CD25+ T cell subsets were significantly lower among MS patients than among controls. This difference was particularly evident in the CD4+CD25(high) T cell subset. The proportions of CD4+Tim3+ and CD4+CD25+Tim3+ cells in PBMCs were significantly lower in the MS group than in the control group, whereas no significant differences were detected regarding the percentages of CD3+Tim3+ in PBMCs and T cell subsets. The serum concentrations of galectin-9, IL-17, and IFN-γ all were increased in MS patients compared with healthy controls. Our results support that Tim3 and related cytokines may be involved in the onset of MS.
Research article
PECAM-1 phosphorylation and tissue factor expression in HUVECs exposed to uniform and disturbed pulsatile flow and chemical stimuli
Journal of Vascular Surgery, Volume 61, Issue 2, 2015, pp. 481-488
We analyzed the relationship between platelet endothelial cell adhesion molecule-1 (PECAM-1) activation and tissue factor (TF) RNA expression in human umbilical vein endothelial cells (HUVECs) exposed to mechanical and chemical stimulation.
Fifty percent confluent and 100% confluent HUVEC cultures were exposed to pulsatile forward flow, as a model for uniform flow, or pulsatile to-and-fro flow, as a model for disturbed flow, using a parallel-plate flow chamber system for up to 4hours in the presence or absence of 4 U/mL thrombin. Protein lysates were immunoprecipitated for PECAM-1 and then immunoblotted with anti-phospho-tyrosine antibody. TF RNA expression was determined using quantitative reverse transcription polymerase chain reaction.
HUVECs exposed to disturbed flow induced higher TF expression at 4hours than HUVECs exposed to uniform flow in sparse cultures (16.8± 5.8 vs 5.1± 1.2; P< .05). HUVECs exposed to disturbed flow and thrombin induced higher TF RNA expression at 4hours than cultures exposed to uniform flow and thrombin in both confluent (47.0± 6.0 vs 30.2± 4.9; P< .05) and sparse (72.3± 10.7 vs 49.8± 4.7; P< .05) cultures. In confluent HUVEC cultures, PECAM-1 is minimally phosphorylated by disturbed and uniform flow, while in sparse HUVEC cultures, PECAM-1 phosphorylation at 15minutes is greater in both disturbed and uniform flow (2.0± 0.2 and 2.1± 0.4 respectively; P< .05). Thrombin treatment of static HUVECs exhibited greater PECAM-1 phosphorylation at 15minutes in confluent compared with sparse cultures (3.0± 0.5 vs 2.3± 0.1; P< .05). PECAM-1 phosphorylation of HUVECs exposed to both flow and thrombin is significantly higher in sparse cultures compared with either flow or thrombin stimulation alone but was suppressed in confluent cultures.
The significantly higher TF RNA expression induced by disturbed flow and cell confluence indicates that suppression of PECAM-1 phosphorylation may be an important contributory mechanical signal pathway that promotes TF expression when HUVECs are exposed to disturbed flow.
Research article
The transcription factor Nrf2 is decreased after spontaneous term labour in human fetal membranes where it exerts anti-inflammatory properties
Placenta, Volume 36, Issue 1, 2015, pp. 7-17
Inflammation plays a central role in the terminal processes of human labour and delivery, including the rupture of fetal membranes. Recent studies show a role for the transcription factor Nrf2 (NF-E2-related factor 2) in regulating inflammation. The aims of this study were to determine the effect of human spontaneous term and preterm labour on Nrf2 expression in fetal membranes; and Nrf2 siRNA knockdown on pro-inflammatory cytokines.
Fetal membranes, from term and preterm, were obtained from non-labouring and labouring women. Primary amnion cells were used to determine the effect of Nrf2 siRNA knockdown on pro-inflammatory cytokines in the presence of inflammation or infection.
Nrf2 mRNA expression and nuclear protein expression were significantly decreased after spontaneous term labour and delivery. There was, however, no effect of spontaneous preterm labour and delivery on Nrf2 mRNA expression and nuclear protein expression. On the other hand, Nrf2 gene expression was significantly lower in fetal membranes obtained from women at preterm with histologic chorioamnionitis compared to fetal membranes obtained from women at preterm without histologic chorioamnionitis. Nrf2 silencing by siRNA in primary amnion cells was associated with a significant increase in IL-6 and IL-8 mRNA expression and release induced by IL-1β, TNF-α, flagellin and poly(I:C).
Nrf2 has an anti-inflammatory effect in human fetal membranes. It is decreased with term labour and preterm chorioamnionitis; and Nrf2 silencing increases inflammation- and infection-induced pro-inflammatory cytokines. Further studies are required to determine if agents that can increase Nrf2 expression may be a potential therapeutic strategy in the treatment and management of infection-induced preterm labour.
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