Characterization and ecological risk assessment of microplastics accumulated in sea water, sand, sediment, shell water and selected tissues of hermit crab of Sundarban Biosphere Reserve.

Sundarban, a Ramsar site of India, has been encountering an ecological threat due to the presence of microplastic (MP) wastes generated from different anthropogenic sources. Clibanarius longitarsus, an intertidal hermit crab of Sundarban Biosphere Reserve, resides within the abandoned shell of a gastropod mollusc, Telescopium telescopium. We characterized and estimated the MP in the gills and gut of hermit crab, as well as in the water present in its occupied gastropod shell. The average microplastic abundance in sea water, sand and sediment were 0.175 ± 0.145 MP L-1, 42 ± 15.03 MP kg-1 and 67.63 ± 24.13 MP kg-1 respectively. The average microplastic load in hermit crab was 1.94 ± 0.59 MP crab-1, with 33.89 % and 66.11 % in gills and gut respectively. Gastropod shell water exhibited accumulation of 1.69 ± 1.43 MP L-1. Transparent and fibrous microplastics were documented as the dominant polymers of water, sand and sediment. Shell water exhibited the prevalence of green microplastics followed by transparent ones. Microscopic examination revealed microplastics with 100-300 µm size categories were dominant across all abiotic compartments. ATR-FTIR and Raman spectroscopy confirmed polyethylene and polypropylene as the prevalent polymers among the five identified polymers of biotic and abiotic components. The target group index indicated green and black as the preferable microplastics of crab. The ecological risk analysis indicated a considerable level of environmental pollution risk in Sundarban and its inhabiting organisms. This important information base may facilitate in developing a strategy of mitigation to limit the MP induced ecological risk at Sundarban Biosphere Reserve.

Microplastics contamination in two species of gobies and their estuarine habitat of Indian Sundarbans.

Sundarbans, a Ramsar site of India is contaminated with heterogeneous microplastic wastes. Boddart's goggle eye mudskipper and Rubicundus eelgoby, were common gobies of Sundarbans estuary which accumulated microplastics during their normal biological activities. We estimated the abundance of microplastics in water, sediment; skin, gills, bucco-opercular cavity and gastrointestinal tract of these two goby fishes. Microplastic load estimated in gobies were 0.84 and 2.62 particles per fish species with a dominance of transparent, fibrous microplastics with 100-300 µm in length. ATR-FTIR and Raman spectroscopy revealed polyethylene as prevalent polymer. Surface degradations and adsorption of contaminants on microplastic surface were investigated by SEM-EDX analysis. Presence of hazardous polymers influenced high polymer hazard index and potential ecological risk index which indicated acute environmental threat to Sundarbans estuary and its resident organisms. Current study will provide a new information base on the abundance of microplastics and its ecological hazard in this biosphere reserve.

Body Mass Index, Adverse Pregnancy Outcomes, and Cardiovascular Disease Risk.

BACKGROUND: Obesity is a well-established risk factor for both adverse pregnancy outcomes (APOs) and cardiovascular disease (CVD). However, it is not known whether APOs are mediators or markers of the obesity-CVD relationship. This study examined the association between body mass index, APOs, and postpartum CVD risk factors. METHODS: The sample included adults from the nuMoM2b (Nulliparous Pregnancy Outcomes Study: Monitoring Mothers-To-Be) Heart Health Study who were enrolled in their first trimester (6 weeks-13 weeks 6 days gestation) from 8 United States sites. Participants had a follow-up visit at 3.7 years postpartum. APOs, which included hypertensive disorders of pregnancy, preterm birth, small-for-gestational-age birth, and gestational diabetes, were centrally adjudicated. Mediation analyses estimated the association between early pregnancy body mass index and postpartum CVD risk factors (hypertension, hyperlipidemia, and diabetes) and the proportion mediated by each APO adjusted for demographics and baseline health behaviors, psychosocial stressors, and CVD risk factor levels. RESULTS: Among 4216 participants enrolled, mean±SD maternal age was 27±6 years. Early pregnancy prevalence of overweight was 25%, and obesity was 22%. Hypertensive disorders of pregnancy occurred in 15%, preterm birth in 8%, small-for-gestational-age birth in 11%, and gestational diabetes in 4%. Early pregnancy obesity, compared with normal body mass index, was associated with significantly higher incidence of postpartum hypertension (adjusted odds ratio, 1.14 [95% CI, 1.10-1.18]), hyperlipidemia (1.11 [95% CI, 1.08-1.14]), and diabetes (1.03 [95% CI, 1.01-1.04]) even after adjustment for baseline CVD risk factor levels. APOs were associated with higher incidence of postpartum hypertension (1.97 [95% CI, 1.61-2.40]) and hyperlipidemia (1.31 [95% CI, 1.03-1.67]). Hypertensive disorders of pregnancy mediated a small proportion of the association between obesity and incident hypertension (13% [11%-15%]) and did not mediate associations with incident hyperlipidemia or diabetes. There was no significant mediation by preterm birth or small-for-gestational-age birth. CONCLUSIONS: There was heterogeneity across APO subtypes in their association with postpartum CVD risk factors and mediation of the association between early pregnancy obesity and postpartum CVD risk factors. However, only a small or nonsignificant proportion of the association between obesity and CVD risk factors was mediated by any of the APOs, suggesting APOs are a marker of prepregnancy CVD risk and not a predominant cause of postpartum CVD risk.

A longitudinal epigenome-wide association study of preeclamptic and normotensive pregnancy.

Background: While preeclampsia (PE) is a leading cause of pregnancy-related morbidity/mortality, its underlying mechanisms are not fully understood. DNA methylation (DNAm) is a dynamic regulator of gene expression that may offer insight into PE pathophysiology and/or serve as a biomarker (e.g., risk, subtype, a therapeutic response). This study's purpose was to evaluate for differences in blood-based DNAm across all trimesters between individuals eventually diagnosed with PE (cases) and individuals who remained normotensive throughout pregnancy, did not develop proteinuria, and birthed a normally grown infant (controls). Results: In the discovery phase, longitudinal, genome-wide DNAm data were generated across three trimesters of pregnancy in 56 participants (n=28 cases, n=28 controls) individually matched on self-identified race, pre-pregnancy body mass index, smoking, and gestational age at sample collection. An epigenome-wide association study (EWAS) was conducted, using surrogate variable analysis to account for unwanted sources of variation. No CpGs met the genome-wide significance p value threshold of 9×10-8, but 16 CpGs (trimester 1: 5; trimester 2: 1; trimester 3: 10) met the suggestive significance threshold of 1×10-5. DNAm data were also evaluated for differentially methylated regions (DMRs) by PE status. Three DMRs in each trimester were significant after Bonferonni-adjustment. Since only third-trimester samples were available from an independent replication sample (n=64 cases, n=50 controls), the top suggestive hits from trimester 3 (cg16155413 and cg21882990 associated with TRAF3IP2-AS1/TRAF3IP2 genes, which also made up the top DMR) were carried forward for replication. During replication, DNAm data were also generated for validation purposes from discovery phase third trimester samples. While significant associations between DNAm and PE status were observed at both sites in the validation sample, no associations between DNAm and PE status were observed in the independent replication sample. Conclusions: The discovery phase findings for cg16155413/cg21882990 (TRAF3IP2-AS1/TRAF3IP2) were validated with a new platform but were not replicated in an independent sample. Given the differences in participant characteristics between the discovery and replication samples, we cannot rule out important signals for these CpGs. Additional research is warranted for cg16155413/cg21882990, as well as top hits in trimesters 1-2 and significant DMRs that were not examined in the replication phase.

Can allostatic load in pregnancy explain the association between race and subsequent cardiovascular disease risk: A cohort study.

OBJECTIVE: To assess the relationship between allostatic load, a measure of cumulative chronic stress in early pregnancy and cardiovascular disease risk, 2-7 years postpartum, and pathways contributing to racial disparities in cardiovascular disease risk. DESIGN: Secondary analysis of a prospective cohort study. SETTING MULTICENTER POPULATION: Pregnant women. METHODS: Our primary exposure was high allostatic load in the first trimester, defined as at least 4 of 12 biomarkers (systolic blood pressure, diastolic blood pressure, body mass index, cholesterol, low-density lipoprotein, high-density lipoprotein, high-sensitivity C-reactive protein, triglycerides, insulin, glucose, creatinine and albumin) in the unfavourable quartile. Logistic regression was used to test the association between high allostatic load and main outcome adjusted for confounders: time from index pregnancy and follow up, age, education, smoking, gravidity, bleeding in the first trimester, index adverse pregnancy outcomes, and health insurance. Each main outcome component and allostatic load were analysed secondarily. Mediation and moderation analyses assessed the role of high allostatic load in racial disparities of cardiovascular disease risk. MAIN OUTCOME MEASURE: Incident cardiovascular disease risk: hypertension, or metabolic disorders. RESULTS: Cardiovascular disease risk was identified in 1462/4022 individuals (hypertension: 36.6%, metabolic disorder: 15.4%). After adjustment, allostatic load was associated with cardiovascular disease risk (adjusted odds ratio [aOR] 2.0, 95% CI 1.8-2.3), hypertension (aOR 2.1, 95% CI 1.8-2.4) and metabolic disorder (aOR 1.7, 95% CI 1.5-2.1). Allostatic load was a partial mediator between race and cardiovascular disease risk. Race did not significantly moderate this relationship. CONCLUSIONS: High allostatic load during pregnancy is associated with cardiovascular disease risk. The relationships between stress, subsequent cardiovascular risk and race warrant further study.

Epigenomic Links Between Social Determinants of Health and Symptoms: A Scoping Review.

Social determinants of health (SDoH) impact health and wellness. The link between SDoH and adverse health outcomes, including symptom occurrence and severity, may be explained by an individual's physiologic response to one or more SDoH. One potential mechanism underlying this physiologic response linking SDoH and symptoms is the dynamic epigenome. The purpose of this scoping review of the literature was to examine differential susceptibility for symptoms by identifying and summarizing research linking SDoH and symptoms through epigenomic mechanisms. PubMed was searched to identify empirical research where at least one SDoH was an independent or dependent variable, at least one symptom was investigated, and the investigation included an epigenomic measure. Of the 484 articles initially retrieved, after thorough vetting, 41 articles met eligibility. The most studied symptom was depressive symptoms followed by anxiety, cognitive function, sleep dysfunction, and pain. The most frequently studied SDoH were: 1) stress, particularly early life stress and acculturative stress; and 2) trauma, predominantly childhood trauma. DNA methylation and telomere length were the most studied epigenomic measures. Four genes (SLC6A4, BDNF, NR3C1, OXTR) had evidence from multiple studies and across methodological approaches linking SDoH to symptoms. This review supports the inclusion of epigenomic approaches to better understand the link between SDoH and symptoms and provides evidence that SDoH impact telomere length and the methylation of genes involved in neurotransmitter signaling, neuronal survival, behavior, inflammation and stress response.

Advancing Nursing Research Through Interactive Data Visualization With R Shiny.

Scientific data visualization is a critical aspect of fully understanding data patterns and trends. To date, the majority of data visualizations in nursing research - as with other biomedical fields - have been static. The availability of electronic scientific journal articles (which are quickly becoming the norm) has created new opportunities for dynamic and interactive data visualization which carry added cognitive benefits and support the ability to understand data more fully. Therefore, here we highlight the benefits of R, an open-source programming language, for scientific data visualization, with a specific focus on creating dynamic, interactive figures using the R shiny package. For R users, we have included a tutorial with example code to create three increasingly complex shiny applications. For individuals more interested in understanding the potential of R shiny as an innovative tool to interact with research data, we have included links to online versions of the examples that do not require any programming or R experience. We believe that widespread adoption of dynamic and interactive scientific data visualization will further support nurse scientists' higher-level mission of advancing our understanding of health and wellness of individuals and communities.

A comparative account of coelomocyte of earthworm ecotypes with reference to its morphology, morphometry, density, phagocytosis, autofluorescence, and oxidative status.

Earthworms inhabit different strata of moist soil. Epigeic and endogeic earthworms prefer superficial and inner stratum of soil respectively, whereas, semiaquatic species are distributed around hydrated soil near ponds and lakes. Coelomocytes, the chief immunoeffector cells of coelomic origin, perform diverse physiological functions like phagocytosis, maintenance of cellular homeostasis, and acid-base balance of coelomic fluid, graft rejection, elicitation of cytotoxic, and oxidative responses under the challenges of pathogens and toxins. The present study aims to analyze selected morphological and functional parameters in three differentially adapted Indian earthworms of nonsimilar habitats. Coelomocytes of Glyphidrilus tuberosus (Stephenson, 1916) (semiaquatic), Perionyx excavatus (Perrier, 1872) (epigeic), and Eutyphoeus orientalis (Beddard, 1883) (endogeic) were isolated for morphological and morphometric analyses and subjected to determination of phagocytic, oxidative, and cytotoxic responses. Activities of phenoloxidase, pro, and antioxidant enzymes, and autofluorescence were determined in the extruded coelomocytes of earthworms of three contrasting habitats. The differential result may be correlated with species-specific responses and variation in habitat preference and related adaptation.

Metal accumulation and morphofunctional damage in coelomocytes of earthworm collected from industrially contaminated soil of Kolkata, India.

The current study is aimed to assess the ecotoxicological effects of toxic metals and seasonal shift of the physicochemical characteristics of soil in an endogeic earthworm Metaphire posthuma of industrially contaminated soil of Calcutta leather complex. The accumulation of cadmium, chromium, lead and mercury was quantitated in whole earthworms and coelomocytes. The accumulation of metals was derived to be high in the coelomocytes than whole earthworms. Morphofunctional shift in coelomocytes indicated a high level of metal toxicity in soil inhabitants. The shift in differential coelomocyte count and cellular damage including intense cytoplasmic vacuolation and membrane blebbing of coelomocytes of M. posthuma of contaminated soil were suggestive to a state of immunocompromisation in the same species. Shift in the generation of nitric oxide and activity of inducible nitric oxide synthase indicated a possible immunosuppression in earthworm. Depletion in the acetylcholinesterase activity of coelomocytes indicated neurotoxicity of metals leached from the dumped wastes in Calcutta leather complex. Integrated biomarker response based analysis was carried out to assess the biomarker potential of experimental endpoints of M. posthuma to monitor metal toxicity in soil.

An exploratory study of white blood cell proportions across preeclamptic and normotensive pregnancy by self-identified race in individuals with overweight or obesity.

Objective: Examine white blood cell (WBC) proportions across preeclamptic (n = 28 cases) and normotensive (n = 28 controls) pregnancy in individuals with overweight/obesity.Methods: WBC proportions were inferred from genome-wide DNA methylation data and compared by case/control status and self-identified race.Results: In Trimester 1, ean B cell proportions were suggestively lower in cases in the overall sample and significantly lower in White participants but not in Black participants. More significant WBC proportion changes were observed across normotensive than preeclamptic pregnancy.Conclusions: These findings in a small sample demonstrate need for additional studies investigating the relationship between self-identified race and WBCs in pregnancy.

Bivalve haemocyte adhesion, aggregation and phagocytosis: A tool to reckon arsenic induced threats to freshwater ecosystem.

The freshwater aquifers of the Indo-Gangetic plains support rich biodiversity which is under the threat of arsenic contamination. The filter feeding bivalve mollusc Lamellidens marginalis is a sessile and sentinel resident of these freshwater habitats. In the present study, the classical cell behaviours of adhesion and aggregation were monitored in the circulating haemocytes of the freshwater bivalve under the exposure of sodium arsenite (NaAsO2) at sublethal concentrations in controlled laboratory conditions for a maximum time-span of sixteen days. The toxic metalloid significantly inhibited non-self adhesion, inter-haemocyte interactions and haemocyte aggregation in a dose and time dependent manner. The natural occurrence of the filopods on the haemocytes was significantly diminished in the bivalves exposed to the inorganic arsenite. Moreover, a significant fall in the kinetics of phagocytosis index and haemocyte adhesion was observed under the in vitro exposure to NaAsO2. Compromised non-self adhesion, cell-cell aggregation and phagocytosis of non-self particles by the bivalve haemocytes probably indicate susceptible immunological status of the bivalve. Such vulnerable immunity of the bivalve probably signifies the nature of imminent threat to the freshwater ecosystem as a whole under inorganic arsenite exposure. The findings would be helpful to design bivalve haemocyte based inexpensive biomonitoring tool to assess the health of freshwater ecosystem under potential arsenic threat.

An Exploratory Study of Epigenetic Age in Preeclamptic and Normotensive Pregnancy Reveals Differences by Self-Reported Race but Not Pregnancy Outcome.

Preeclampsia is a leading cause of maternal and neonatal morbidity and mortality. Chronological age and race are associated with preeclampsia, but the role of these factors is not entirely understood. We hypothesized that DNA methylation age, a measure of biological age, would be higher in individuals with preeclampsia than in individuals with normotensive pregnancy and that DNA methylation age would differ by race across pregnancy. This was a longitudinal, exploratory study of 56 pregnant individuals (n = 28 preeclampsia cases and n = 28 normotensive controls). Genome-wide DNA methylation data were generated from trimester-specific peripheral blood samples. DNA methylation age was estimated using the "Improved Precision" clock, and ∆age, the difference between DNA methylation age and chronological age, was computed. DNA methylation age was compared with chronological age using Pearson correlations. The relationships between ∆age and preeclampsia status, self-reported race, and covariates were tested using multiple linear regression and performed both with and without consideration of cell-type heterogeneity. We observed strong correlation between chronological age and DNA methylation age across pregnancy, with significantly stronger correlation observed in White participants than in Black participants. We observed no association between ∆age and preeclampsia status. However, ∆age was higher in participants with higher pre-pregnancy body mass index in trimester 1 and lower in Black participants than in White participants in trimesters 2 and 3. Observations were largely consistent when controlling for cell-type heterogeneity. Our findings in a small sample support the need for additional studies to investigate the relationship between race and biological age, which could provide further insight into racial disparities across pregnancy. However, this study does not support an association between ∆age and preeclampsia status.

Effects of copper oxide nanoparticle on gill filtration rate, respiration rate, hemocyte associated immune parameters and oxidative status of an Indian freshwater mussel.

Waterbodies of India support a wide range of molluscs including Lamellidens marginalis, a pearl forming edible mussel of ecological significance. Report of copper oxide nanoparticle toxicity in Indian molluscs is limited in scientific literature. L. marginalis is a gill respiring filter feeder, which is toxicologically vulnerable to exposure of copper oxide nanoparticles liberated from electrical, textile and polymer industries. Experimental exposure of copper oxide nanoparticles for 7 days yielded a decrease in gill filtration rate, respiration rate, total count and phagocytic response of hemocytes, the chief immunoeffector cells of L. marginalis. Nanoparticle exposure resulted in decrease of phagocytic response of mussel hemocytes. Decrease in nitric oxide generation and phenoloxidase activity were recorded in L. marginalis exposed to 0.5, 1 and 5 mg copper oxide nanoparticles per litre of water for 7 and 14 days. Superoxide anion generation in hemocytes was increased under the exposure of copper oxide nanoparticles. Increase in superoxide anion and decrease in the activities of superoxide dismutase and catalase were indicative to oxidative stress in mussels. Copper oxide nanoparticle induced shift in filtration and respiration rate along with the hemocyte associated immune parameters were suggestive to an acute immunophysiological stress in L. marginalis. We estimated the functional performance of gill and physiological status of aquatic respiration in L. marginalis exposed to copper oxide nanoparticles. A parallel set of estimation of each parameter was carried out in L. marginalis exposed to identical copper sulphate concentrations to record and compare the ionic toxicity of copper in the same specimen.

Shift in phagocytosis, lysosomal stability, lysozyme activity, apoptosis and cell cycle profile in the coelomocytes of earthworm of polluted soil near a tannery field of India.

Calcutta Leather Complex of the state of West Bengal, India has been designated as an industrially active zone with around 400 active tannery units. This area spanning 4.5 km2 is surrounded by human habitation. The soil of this region is contaminated with metal pollutants and exhibited an alteration in selected physicochemical parameters, namely cation exchange capacity, moisture content, pH, total nitrogen, total organic carbon and water holding capacity. Metaphire posthuma, a common variety of endogeic earthworm inhabiting this region is thus continuously exposed to these toxic metals. Coelomocytes, the chief immune effector cells of earthworm presented a shift in phagocytosis, lysosomal membrane stability, lysozyme and phosphatase activity, physiological apoptosis and cell cycle profile of M. posthuma sampled from the soil of tannery industry. Presence of high concentration of toxic metals and change in the physicochemical characteristics of soil led to a state of cellular stress and immunocompromisation in M. posthuma, a common inhabitant of soil of this region. Experimental endpoints bear ecotoxicological significance as biomarkers of physiological stress in earthworm for monitoring the health of soil around this tannery industrial zone.

Regulation of Stress Granule Formation by Inflammation, Vascular Injury, and Atherosclerosis.

OBJECTIVE: Stress granules (SGs) are dynamic cytoplasmic aggregates containing mRNA, RNA-binding proteins, and translation factors that form in response to cellular stress. SGs have been shown to contribute to the pathogenesis of several human diseases, but their role in vascular diseases is unknown. This study shows that SGs accumulate in vascular smooth muscle cells (VSMCs) and macrophages during atherosclerosis. Approach and Results: Immunohistochemical analysis of atherosclerotic plaques from LDLR-/- mice revealed an increase in the stress granule-specific markers Ras-G3BP1 (GTPase-activating protein SH3 domain-binding protein) and PABP (poly-A-binding protein) in intimal macrophages and smooth muscle cells that correlated with disease progression. In vitro, PABP+ and G3BP1+ SGs were rapidly induced in VSMC and bone marrow-derived macrophages in response to atherosclerotic stimuli, including oxidized low-density lipoprotein and mediators of mitochondrial or oxidative stress. We observed an increase in eIF2α (eukaryotic translation initiation factor 2-alpha) phosphorylation, a requisite for stress granule formation, in cells exposed to these stimuli. Interestingly, SG formation, PABP expression, and eIF2α phosphorylation in VSMCs is reversed by treatment with the anti-inflammatory cytokine interleukin-19. Microtubule inhibitors reduced stress granule accumulation in VSMC, suggesting cytoskeletal regulation of stress granule formation. SG formation in VSMCs was also observed in other vascular disease pathologies, including vascular restenosis. Reduction of SG component G3BP1 by siRNA significantly altered expression profiles of inflammatory, apoptotic, and proliferative genes. CONCLUSIONS: These results indicate that SG formation is a common feature of the vascular response to injury and disease, and that modification of inflammation reduces stress granule formation in VSMC.

Analysis of oxidative stress and cellular aggregation in the coelomocytes of earthworms collected from metal contaminated sites of industrial and agricultural soils of West Bengal, India.

Endogeic earthworm Metaphire posthuma (Valliant, 1868) is a common biological component of the tropical soil of India and other countries. The species is reported to influence fertility and porosity of soil and bear a high composting potential. Intensive agricultural, industrial, and mining activities increase the amount of toxic metals in soil causing physiological adversity in earthworm and other biotic components in soil. Coelomocytes, the chief immunoeffector cells of earthworm, perform diverse physiological functions under the challenge of toxins and pathogens. The experimental earthworms collected separately from soils with agricultural and tannery activities were subjected to quantitation of prooxidation and antioxidation parameters for estimation of oxidative stress. Total count, cellular aggregation, generation of reactive oxygen species (ROS), superoxide anion, nitric oxide, activities of phenoloxidase, superoxide dismutase, catalase and glutathione-s-transferase, and amount of total protein were estimated in the coelomocytes of M. posthuma as experimental end points of toxicity screening. Concentrations of cadmium, chromium, lead, and mercury were determined in the soil samples to assess the degree of toxic contamination. The increase in the amount of prooxidants and decrease in the activities of antioxidant enzymes indicated the signs of oxidative stress in the coelomocytes of the organism. Aggregation of circulating coelomocytes is considered as an immune response involved in pathogen encapsulation response as reported in many invertebrates. Decrease in coelomocyte aggregation in earthworm collected from contaminated sites suggested a state of inappropriate shift of the innate immune status. Toxin-induced oxidative stress and reductions in cell aggregation response are the signs of immunocompromisation of M. posthuma. Present findings bear a prospect of this experimental species as an indicator of soil pollution.

Regulation of pro- and anti-atherogenic cytokines.

Despite advances in prevention and treatment, vascular diseases continue to account for significant morbidity and mortality in the developed world. Incidence is expected to worsen as the number of patients with common co-morbidities linked with atherosclerotic vascular disease, such as obesity and diabetes, continues to increase, reaching epidemic proportions. Atherosclerosis is a lipid-driven vascular inflammatory disease involving multiple cell types in various stages of inflammation, activation, apoptosis, and necrosis. One commonality among these cell types is that they are activated and communicate with each other in a paracrine fashion via a complex network of cytokines. Cytokines mediate atherogenesis by stimulating expression of numerous proteins necessary for induction of a host of cellular responses, including inflammation, extravasation, proliferation, apoptosis, and matrix production. Cytokine expression is regulated by a number of transcriptional and post-transcriptional mechanisms. In this context, proteins that control and fine-tune cytokine expression can be considered key players in development of atherosclerosis and also represent targets for rational drug therapy to combat this disease. This review will describe the cellular and molecular mechanisms that drive atherosclerotic plaque progression and present key cytokines that participate in this process. We will also describe RNA binding proteins that mediate cytokine mRNA stability and regulate cytokine abundance. Identification and characterization of the cytokines and proteins that regulate their abundance are essential to our ability to identify therapeutic approaches to ameliorate atherosclerotic vascular disease.

RNA stability protein ILF3 mediates cytokine-induced angiogenesis.

Interleukin enhancer-binding factor 3 (ILF3), an RNA-binding protein, is best known for its role in innate immunity by participation in cellular antiviral responses. A role for ILF3 in angiogenesis is unreported. ILF3 expression in CD31+ capillaries of hypoxic cardiac tissue was detected by immunohistochemistry. Proangiogenic stimuli induce ILF3 mRNA and protein expression in cultured human coronary artery endothelial cells (hCAECs). Angiogenic indices, including proliferation, migration, and tube formation, are all significantly reduced in hCAECs when ILF3 is knocked down using small interfering RNA (siRNA), but are significantly increased when ILF3 is overexpressed using adenovirus. Protein and mRNA abundance of several angiogenic factors including CXCL1, VEGF, and IL-8 are decreased when ILF3 is knocked down by siRNA. These factors are increased when ILF3 is overexpressed by adenovirus. ILF3 is phosphorylated and translocates from the nucleus to the cytoplasm in response to angiogenic stimuli. Proangiogenic transcripts containing adenine and uridine-rich elements were bound to ILF3 through RNA immunoprecipitation. ILF3 stabilizes proangiogenic transcripts including VEGF, CXCL1, and IL-8 in hCAECs. Together these data suggest that in endothelial cells, the RNA stability protein, ILF3, plays a novel and central role in angiogenesis. Our working hypothesis is that ILF3 promotes angiogenesis through cytokine-inducible mRNA stabilization of proangiogenic transcripts.-Vrakas, C. N., Herman, A. B., Ray, M., Kelemen, S. E., Scalia, R., Autieri, M. V. RNA stability protein ILF3 mediates cytokine-induced angiogenesis.

FXR1 Is an IL-19-Responsive RNA-Binding Protein that Destabilizes Pro-inflammatory Transcripts in Vascular Smooth Muscle Cells.

This work identifies the fragile-X-related protein (FXR1) as a reciprocal regulator of HuR target transcripts in vascular smooth muscle cells (VSMCs). FXR1 was identified as an HuR-interacting protein by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The HuR-FXR1 interaction is abrogated in RNase-treated extracts, indicating that their association is tethered by mRNAs. FXR1 expression is induced in diseased but not normal arteries. siRNA knockdown of FXR1 increases the abundance and stability of inflammatory mRNAs, while overexpression of FXR1 reduces their abundance and stability. Conditioned media from FXR1 siRNA-treated VSMCs enhance activation of naive VSMCs. RNA EMSA and RIP demonstrate that FXR1 interacts with an ARE and an element in the 3' UTR of TNFα. FXR1 expression is increased in VSMCs challenged with the anti-inflammatory cytokine IL-19, and FXR1 is required for IL-19 reduction of HuR. This suggests that FXR1 is an anti-inflammation responsive, HuR counter-regulatory protein that reduces abundance of pro-inflammatory transcripts.

Genetic Deletion of IL-19 (Interleukin-19) Exacerbates Atherogenesis in Il19-/-×Ldlr-/- Double Knockout Mice by Dysregulation of mRNA Stability Protein HuR (Human Antigen R).

OBJECTIVE: To test the hypothesis that loss of IL-19 (interleukin-19) exacerbates atherosclerosis. APPROACH AND RESULTS: Il19-/- mice were crossed into Ldlr-/- (low-density lipoprotein receptor knock out) mice. Double knockout (dKO) mice had increased plaque burden in aortic arch and root compared with Ldlr-/- controls after 14 weeks of high-fat diet (HFD). dKO mice injected with 10 ng/g per day rmIL-19 had significantly less plaque compared with controls. qRT-PCR and Western blot analysis revealed dKO mice had increased systemic and intraplaque polarization of T cells and macrophages to proinflammatory Th1 and M1 phenotypes, and also significantly increased TNF (tumor necrosis factor)-α expression in spleen and aortic arch compared with Ldlr-/- controls. Bone marrow transplantation suggests that immune cells participate in IL-19 protection. Bone marrow-derived macrophages and vascular smooth muscle cells isolated from dKO mice had a significantly greater expression of inflammatory cytokine mRNA and protein compared with controls. Spleen and aortic arch from dKO mice had significantly increased expression of the mRNA stability protein HuR (human antigen R). Bone marrow-derived macrophage and vascular smooth muscle cell isolated from dKO mice also had greater HuR abundance. HuR stabilizes proinflammatory transcripts by binding AU-rich elements in the 3' untranslated region. Cytokine and HuR mRNA stability were increased in dKO bone marrow-derived macrophage and vascular smooth muscle cell, which was rescued by addition of IL-19 to these cells. IL-19-induced expression of miR133a, which targets and reduced HuR abundance; miR133a levels were lower in dKO mice compared with controls. CONCLUSIONS: These data indicate that IL-19 is an atheroprotective cytokine which decreases the abundance of HuR, leading to reduced inflammatory mRNA stability.