Longitudinal Proteomic Analysis of Plasma across Healthy Pregnancies Reveals Indicators of Gestational Age.

Longitudinal changes in the blood proteome during gestation relate to fetal development and maternal homeostasis. Charting the maternal blood proteome in normal pregnancies is critical for establishing a baseline reference when assessing complications and disease. Using mass spectrometry-based shotgun proteomics, we surveyed the maternal plasma proteome across uncomplicated pregnancies. Results indicate a significant rise in proteins that govern placentation and are vital to the development and health of the fetus. Importantly, we uncovered proteome signatures that strongly correlated with gestational age. Fold increases and correlations between the plasma concentrations of ADAM12 (ρ = 0.973), PSG1 (ρ = 0.936), and/or CSH1/2 (ρ = 0.928) with gestational age were validated with ELISA. Proteomic and validation analyses demonstrate that the maternal plasma concentration of ADAM12, either independently or in combination with either PSG1 or CSH1/2, correlates with gestational age within ±8 days throughout pregnancy. These findings suggest that the gestational age in healthy pregnancies may be determined by referencing the concentration of select plasma proteins.

Alterations in tissue microRNA after heat stress in the conscious rat: potential biomarkers of organ-specific injury.

BACKGROUND: Heat illness remains a significant cause of morbidity in susceptible populations. Recent research elucidating the cellular mechanism of heat stress leading to heat illness may provide information to develop better therapeutic interventions, risk assessment strategies, and early biomarkers of organ damage. microRNA (miRNA) are promising candidates for therapeutic targets and biomarkers for a variety of clinical conditions since there is the potential for high specificity for individual tissues and unique cellular functions. The objective of this study was to identify differentially expressed microRNAs and their putative mRNA targets in the heart, liver, kidney, and lung in rats at three time points: during heat stress (i.e., when core temperature reached 41.8 °C), or following a 24 or 48 h recovery period. RESULTS: Rats did not show histological evidence of tissue pathology until 48 h after heat stress, with 3 out of 6 rats showing cardiac inflammation and renal proteinosis at 48 h. The three rats with cardiac and renal pathology had 86, 7, 159, and 37 differentially expressed miRNA in the heart, liver, kidney, or lung, respectively compared to non-heat stressed control animals. During heat stress one differentially expressed miRNA was found in the liver and five in the lung, with no other modulated miRNA after 24 h or 48 h in animals with no evidence of organ injury. Pathway enrichment analysis revealed enrichment in functional pathways associated with heat stress, with the greatest effects observed in animals with histological evidence of cardiac and renal damage at 48 h. Inhibiting miR-21 in cultured cardiomyocytes increased the percent apoptotic cells five hours after heat stress from 70.9 ± 0.8 to 84.8 ± 2.2%. CONCLUSIONS: Global microRNA and transcriptomics analysis suggested that perturbed miRNA due to heat stress are involved in biological pathways related to organ injury, energy metabolism, the unfolded protein response, and cellular signaling. These miRNA may serve as biomarkers of organ injury and potential pharmacological targets for preventing heat illness or organ injury.

Comparative Proteomic Analysis of Liver Steatosis and Fibrosis after Oral Hepatotoxicant Administration in Sprague-Dawley Rats.

The past decade has seen an increase in the development and clinical use of biomarkers associated with histological features of liver disease. Here, we conduct a comparative histological and global proteomics analysis to identify coregulated modules of proteins in the progression of hepatic steatosis or fibrosis. We orally administered the reference chemicals bromobenzene (BB) or 4,4'-methylenedianiline (4,4'-MDA) to male Sprague-Dawley rats for either 1 single administration or 5 consecutive daily doses. Livers were preserved for histopathology and global proteomics assessment. Analysis of liver sections confirmed a dose- and time-dependent increase in frequency and severity of histopathological features indicative of lipid accumulation after BB or fibrosis after 4,4'-MDA. BB administration resulted in a dose-dependent increase in the frequency and severity of inflammation and vacuolation. 4,4'-MDA administration resulted in a dose-dependent increase in the frequency and severity of periportal collagen accumulation and inflammation. Pathway analysis identified a time-dependent enrichment of biological processes associated with steatogenic or fibrogenic initiating events, cellular functions, and toxicological states. Differentially expressed protein modules were consistent with the observed histology, placing physiologically linked protein networks into context of the disease process. This study demonstrates the potential for protein modules to provide mechanistic links between initiating events and histopathological outcomes.

Utility of Serum miR-122, Liver Enzymes, and Hepatic Histopathology in Response to Hepatotoxicants in Sprague-Dawley Rats.

More than 80,000 chemicals are in commercial use worldwide. Hepatic metabolism to toxic intermediates is often a key mechanism leading to tissue damage and organ dysfunction. Effective treatment requires prompt detection of hepatotoxicity, ideally with rapid, minimally invasive diagnostic assays. In this study, archetypal histologic features of chemically induced hepatic injury were compared with clinical chemistries (including liver enzymes) and serum concentrations of microRNA-122 (miR-122, the processed form miR-122-5p), a biomarker of liver injury. The hepatotoxicants 4,4'-methylenedianiline (4,4'-MDA), allyl alcohol (AA), or carbon tetrachloride (CCl4) were orally administered to male Sprague-Dawley rats for 1, 5, 14, or 28 days to induce liver damage. Formalin-fixed, paraffin-embedded liver sections were evaluated histologically for inflammation, fibrosis, necrosis, and lipid accumulation. Liver enzymes were measured in serum, and serum miR-122 concentrations were assessed by quantitative polymerase chain reaction (qPCR). Histologic features of hepatic injury dose-dependently increased in both severity and frequency. Increases in liver enzymes and bilirubin were more pronounced in response to AA or 4,4'-MDA than to CCl4 at early time points. Elevated serum miR-122 levels in animals administered CCl4, AA, or 4,4'-MDA were more strongly associated with degree of hepatic histopathology than with dosage. Given this sensitive expression pattern postexposure, liver-specific miR-122 may improve the diagnostic accuracy of early hepatic injury.

Mining kidney toxicogenomic data by using gene co-expression modules.

BACKGROUND: Acute kidney injury (AKI) caused by drug and toxicant ingestion is a serious clinical condition associated with high mortality rates. We currently lack detailed knowledge of the underlying molecular mechanisms and biological networks associated with AKI. In this study, we carried out gene co-expression analyses using DrugMatrix-a large toxicogenomics database with gene expression data from rats exposed to diverse chemicals-and identified gene modules associated with kidney injury to probe the molecular-level details of this disease. RESULTS: We generated a comprehensive set of gene co-expression modules by using the Iterative Signature Algorithm and found distinct clusters of modules that shared genes and were associated with similar chemical exposure conditions. We identified two module clusters that showed specificity for kidney injury in that they 1) were activated by chemical exposures causing kidney injury, 2) were not activated by other chemical exposures, and 3) contained known AKI-relevant genes such as Havcr1, Clu, and Tff3. We used the genes in these AKI-relevant module clusters to develop a signature of 30 genes that could assess the potential of a chemical to cause kidney injury well before injury actually occurs. We integrated AKI-relevant module cluster genes with protein-protein interaction networks and identified the involvement of immunoproteasomes in AKI. To identify biological networks and processes linked to Havcr1, we determined genes within the modules that frequently co-express with Havcr1, including Cd44, Plk2, Mdm2, Hnmt, Macrod1, and Gtpbp4. We verified this procedure by showing that randomized data did not identify Havcr1 co-expression genes and that excluding up to 10 % of the data caused only minimal degradation of the gene set. Finally, by using an external dataset from a rat kidney ischemic study, we showed that the frequently co-expressed genes of Havcr1 behaved similarly in a model of non-chemically induced kidney injury. CONCLUSIONS: Our study demonstrated that co-expression modules and co-expressed genes contain rich information for generating novel biomarker hypotheses and constructing mechanism-based molecular networks associated with kidney injury.

Predicting Rat and Human Pregnane X Receptor Activators Using Bayesian Classification Models.

The pregnane X receptor (PXR) is a ligand-activated transcription factor that acts as a master regulator of metabolizing enzymes and transporters. To avoid adverse drug-drug interactions and diseases such as steatosis and cancers associated with PXR activation, identifying drugs and chemicals that activate PXR is of crucial importance. In this work, we developed ligand-based predictive computational models for both rat and human PXR activation, which allowed us to identify potentially harmful chemicals and evaluate species-specific effects of a given compound. We utilized a large publicly available data set of nearly 2000 compounds screened in cell-based reporter gene assays to develop Bayesian quantitative structure-activity relationship models using physicochemical properties and structural descriptors. Our analysis showed that PXR activators tend to be hydrophobic and significantly different from nonactivators in terms of their physicochemical properties such as molecular weight, logP, number of rings, and solubility. Our Bayesian models, evaluated by using 5-fold cross-validation, displayed a sensitivity of 75% (76%), specificity of 76% (75%), and accuracy of 89% (89%) for human (rat) PXR activation. We identified structural features shared by rat and human PXR activators as well as those unique to each species. We compared rat in vitro PXR activation data to in vivo data by using DrugMatrix, a large toxicogenomics database with gene expression data obtained from rats after exposure to diverse chemicals. Although in vivo gene expression data pointed to cross-talk between nuclear receptor activators that is captured only by in vivo assays, overall we found broad agreement between in vitro and in vivo PXR activation. Thus, the models developed here serve primarily as efficient initial high-throughput in silico screens of in vitro activity.

Longitudinal analysis of maternal plasma apolipoproteins in pregnancy: a targeted proteomics approach.

Minimally invasive diagnostic tests are needed in obstetrics to identify women at risk for complications during delivery. The apolipoproteins fluctuate in complexity and abundance in maternal plasma during pregnancy and could be incorporated into a blood test to evaluate this risk. The objective of this study was to examine the relative plasma concentrations of apolipoproteins and their biochemically modified subtypes (i.e. proteolytically processed, sialylated, cysteinylated, dimerized) over gestational time using a targeted mass spectrometry approach. Relative abundance of modified and unmodified apolipoproteins A-I, A-II, C-I, C-II, and C-III was determined by surface-enhanced laser desorption/ionization-time of flight-mass spectrometry in plasma prospectively collected from 11 gravidas with uncomplicated pregnancies at 4-5 gestational time points per patient. Apolipoproteins were readily identifiable by spectral pattern. Apo C-III(2) and Apo C-III(1) (doubly and singly sialylated Apo C-III subtypes) increased with gestational age (r(2)>0.8). Unmodified Apo A-II, Apo C-I, and Apo C-III(0) showed no correlation (r(2) = 0.01-0.1). Pro-Apo C-II did not increase significantly until third trimester (140 ± 13% of first trimester), but proteolytically cleaved, mature Apo C-II increased in late pregnancy (702 ± 130% of first trimester). Mature Apo C-II represented 6.7 ± 0.9% of total Apo C-II in early gestation and increased to 33 ± 4.5% in third trimester. A label-free, semiquantitative targeted proteomics approach was developed using LTQ-Orbitrap mass spectrometry to confirm the relative quantitative differences observed by surface-enhanced laser desorption/ionization-time of flight-mass spectrometry in Apo C-III and Apo C-II isoforms between first and third trimesters. Targeted apolipoprotein screening was applied to a cohort of term and preterm patients. Modified Apo A-II isoforms were significantly elevated in plasma from mothers who delivered prematurely relative to term controls (p = 0.02). These results support a role for targeted proteomics profiling approaches in monitoring healthy pregnancies and assessing risk of adverse obstetric outcomes.

Patterns of gene expression associated with recovery and injury in heat-stressed rats.

BACKGROUND: The in vivo gene response associated with hyperthermia is poorly understood. Here, we perform a global, multiorgan characterization of the gene response to heat stress using an in vivo conscious rat model. RESULTS: We heated rats until implanted thermal probes indicated a maximal core temperature of 41.8°C (Tc,Max). We then compared transcriptomic profiles of liver, lung, kidney, and heart tissues harvested from groups of experimental animals at Tc,Max, 24 hours, and 48 hours after heat stress to time-matched controls kept at an ambient temperature. Cardiac histopathology at 48 hours supported persistent cardiac injury in three out of six animals. Microarray analysis identified 78 differentially expressed genes common to all four organs at Tc,Max. Self-organizing maps identified gene-specific signatures corresponding to protein-folding disorders in heat-stressed rats with histopathological evidence of cardiac injury at 48 hours. Quantitative proteomics analysis by iTRAQ (isobaric tag for relative and absolute quantitation) demonstrated that differential protein expression most closely matched the transcriptomic profile in heat-injured animals at 48 hours. Calculation of protein supersaturation scores supported an increased propensity of proteins to aggregate for proteins that were found to be changing in abundance at 24 hours and in animals with cardiac injury at 48 hours, suggesting a mechanistic association between protein misfolding and the heat-stress response. CONCLUSIONS: Pathway analyses at both the transcript and protein levels supported catastrophic deficits in energetics and cellular metabolism and activation of the unfolded protein response in heat-stressed rats with histopathological evidence of persistent heat injury, providing the basis for a systems-level physiological model of heat illness and recovery.

Alteration in circulating metabolites during and after heat stress in the conscious rat: potential biomarkers of exposure and organ-specific injury.

BACKGROUND: Heat illness is a debilitating and potentially life-threatening condition. Limited data are available to identify individuals with heat illness at greatest risk for organ damage. We recently described the transcriptomic and proteomic responses to heat injury and recovery in multiple organs in an in vivo model of conscious rats heated to a maximum core temperature of 41.8°C (Tc,Max). In this study, we examined changes in plasma metabolic networks at Tc,Max, 24, or 48 hours after the heat stress stimulus. RESULTS: Circulating metabolites were identified by gas chromatography/mass spectrometry and liquid chromatography/tandem mass spectrometry. Bioinformatics analysis of the metabolomic data corroborated proteomics and transcriptomics data in the tissue at the pathway level, supporting modulations in metabolic networks including cell death or catabolism (pyrimidine and purine degradation, acetylation, sulfation, redox alterations and glutathione metabolism, and the urea cycle/creatinine metabolism), energetics (stasis in glycolysis and tricarboxylic acid cycle, ß-oxidation), cholesterol and nitric oxide metabolism, and bile acids. Hierarchical clustering identified 15 biochemicals that differentiated animals with histopathological evidence of cardiac injury at 48 hours from uninjured animals. The metabolic networks perturbed in the plasma corroborated the tissue proteomics and transcriptomics pathway data, supporting a model of irreversible cell death and decrements in energetics as key indicators of cardiac damage in response to heat stress. CONCLUSIONS: Integrating plasma metabolomics with tissue proteomics and transcriptomics supports a diagnostic approach to assessing individual susceptibility to organ injury and predicting recovery after heat stress.

A systematic review of amniotic fluid assessments in twin pregnancies.

The objectives of this systematic review were to examine the reproducibility of sonographic estimates of amniotic fluid volume (AFV) in twin pregnancies, compare the association of sonographic estimates of AFV with dye-determined AFV, and correlate AFV with antepartum, intrapartum, and perinatal outcomes in twin pregnancies. Studies were included if they were adequately powered and investigated antepartum, intrapartum, and/or perinatal adverse outcome parameters in twin gestations. Studies with comparable populations and exclusion criteria were merged into forest plots. Data comparing the accuracy of AFV assessment, correlation of AFV with gestational age, and adverse outcomes were tabulated. Five of the 6 studies investigating AFV by the amniotic fluid index as a function of gestational age reported data fitting a quadratic equation, with fluid volumes peaking at mid gestation and then declining. This trend was less pronounced when AFV was assessed by the single deepest pocket (2 of 4 studies reporting a quadratic fit). Polyhydramnios was associated with prematurity in 2 of 4 studies (1 amniotic fluid index and 1 single deepest pocket), and oligohydramnios was associated with prematurity in 1 single deepest pocket study. Stillbirth was the only intrapartum outcome reported in more than 1 study. Perinatal outcomes associated with polyhydramnios included neonatal death (P < .05 in 1 of 2 studies), low Apgar scores (1 of 2 studies), neonatal intensive care unit admission (1 of 2 studies), and low birth weight (2 of 3 studies).

Transcriptional analysis of novel hormone receptors PGRMC1 and PGRMC2 as potential biomarkers of breast adenocarcinoma staging.

BACKGROUND: The expression of progesterone receptor membrane component 1 (PGRMC1) in breast cancer has generated interest in this recently discovered protein because of its role in tumorigenesis. However, correlations between patient age, PGRMC1 gene expression, breast cancer morphology, and breast cancer stage have not been adequately studied. Furthermore, very little is known about possible roles for other PGRMC isoforms in breast cancer, like PGRMC2. Thus, we examined the expression of PGRMC1 and PGRMC2 mRNA by relative quantitative PCR (RelqPCR) and determined whether transcript levels correlate with age, breast cancer staging, estrogen receptor alpha (ERα) status, and other morphometric features routinely used during the pathological examination of breast ductal adenocarcinomas. METHODS: Twenty-eight frozen or paraffin embedded breast cancer samples (ductal carcinoma in situ and stages I thru IV invasive ductal adenocarcinoma) and 10 control benign breast tissue samples were randomly selected and interrogated by RelqPCR to determine PGRMC1, 2, and ERα mRNA transcript levels. To control for slight variations in sample preparation, receptor transcript was normalized to the housekeeping gene phosphoglycerate kinase 1 (PGK1). Descriptive statistics and ANOVA of multiparametric datasets were used to correlate transcript levels with pathological staging parameters. RESULTS: PGRMC1 mRNA levels decreased significantly with patient age (Pearson's correlation -0.369; P=0.035), whereas PGRMC2 levels did not. Although the mean relative expression of PGRMC1 significantly decreased in stage II breast cancer compared with controls (P=0.050), it was no longer significant when age was considered a covariance (P=0.371). On the other hand, PGRMC2 mRNA transcript was significantly decreased in stage II breast cancer when compared to stage III cancer (P=0.028) in a manner independent of age (corrected model Bonferroni pair wise comparison, P=0.036). Furthermore, PGRMC2 levels positively correlated with ERα mRNA transcripts in patients with ER positive tumors (Pearson's correlation 0.503, P=0.096). CONCLUSIONS: Decreases in PGRMC1 mRNA are partially explained by increasing patient age. On the other hand, compared to stage III, PCRMC2 mRNA was significantly decreased in stage II adenocarcinoma of the breast in an age-independent manner. Additionally, PGRMC2 mRNA levels displayed a positive correlation with ERα transcripts. Thus, in addition to morphometric pathologic staging criteria, measurements of PGRMC2 mRNA may be useful for distinguishing low stage tumors from higher stages that require more aggressive clinical management, and may be a useful test when tumor ER IHC results are equivocal.

Rapid and sustained nuclear-cytoplasmic ERK oscillations induced by epidermal growth factor.

Although the ERK pathway has a central role in the response of cells to growth factors, its regulatory structure and dynamics are incompletely understood. To investigate ERK activation in real time, we expressed an ERK-GFP fusion protein in human mammary epithelial cells. On EGF stimulation, we observed sustained oscillations of the ERK-GFP fusion protein between the nucleus and cytoplasm with a periodicity of approximately 15 min. The oscillations were persistent (>45 cycles), independent of cell cycle phase, and were highly dependent on cell density, essentially disappearing at confluency. Oscillations occurred even at ligand doses that elicited very low levels of ERK phosphorylation, and could be detected biochemically in both transfected and nontransfected cells. Mathematical modeling revealed that negative feedback from phosphorylated ERK to the cascade input was necessary to match the robustness of the oscillation characteristics observed over a broad range of ligand concentrations. Our characterization of single-cell ERK dynamics provides a quantitative foundation for understanding the regulatory structure of this signaling cascade.

Intramuscular 17α-hydroxyprogesterone caproate administration attenuates immunoresponsiveness of maternal peripheral blood mononuclear cells.

OBJECTIVE: 17α-hydroxyprogesterone caproate (17P) may decrease risk of prematurity by suppressing maternal immunity. We hypothesized that in vivo 17P treatment attenuates immunoresponsiveness of peripheral blood mononuclear cells (PBMCs). STUDY DESIGN: Study subjects were gravidas receiving weekly prophylactic intramuscular 17P injections. Peripheral blood samples were obtained at 21-27 weeks' gestation. Gestational age-matched, drug-naïve gravidas served as controls. To simulate infection, isolated PBMCs were stimulated with lipoteichoic acid (LTA) or lipopolysaccharide (LPS). Extracellular interleukin-6 (IL-6) concentrations were quantified by an enzyme-linked immunosorbent assay. RESULTS: Unstimulated IL-6 levels were comparable in PBMCs derived from drug-naïve and 17P-treated subjects. LPS and LTA induced a dose-dependent elevation of IL-6 in control PBMCs. In patients who received exogenous 17P, LPS, and LTA stimulated induction of IL-6 was significantly decreased compared with controls (P = .005 and P = .02). CONCLUSION: In vivo 17P attenuated immunoreactivity of PBMCs in our in vitro model of Gram-positive and Gram-negative bacterial infection.

The effect of magnesium sulfate on the activity of matrix metalloproteinase-9 in fetal cord plasma and human umbilical vein endothelial cells.

OBJECTIVE: Clinical evidence suggests that magnesium sulfate may reduce the risk of fetal neurologic injury in preterm delivery. Matrix metalloproteinase-9 (MMP-9) levels are elevated in preterm labor patients. There is evidence that MMP-9 may break down the blood-brain barrier in humans, causing cytokine mediated cell injury. Our objective was to determine whether the addition of magnesium sulfate attenuates activity of MMP-9, a complex zinc-dependent enzyme, in fetal cord plasma. STUDY DESIGN: We collected cord plasma in 6 term, unlabored patients. Using enzyme-linked immunosorbent assay, we measured the activity of MMP-9 with varying concentrations of magnesium sulfate added in vitro. Results were verified using a human umbilical cord vein endothelial cell (HUVEC) line. RESULTS: Addition of physiologic doses of magnesium sulfate (0.07 mg/mL) resulted in a 25% decrease in active MMP-9 (P = .03). In a HUVEC line, magnesium sulfate resulted in a 32% decrease in MMP-9 activity (P = .00012). CONCLUSION: The addition of magnesium sulfate attenuated MMP-9 activity in cord plasma and in a HUVEC line.

Group B streptococcus serotype prevalence in reproductive-age women at a tertiary care military medical center relative to global serotype distribution.

BACKGROUND: Group B Streptococcus (GBS) serotype (Ia, Ib, II-IX) correlates with pathogen virulence and clinical prognosis. Epidemiological studies of seroprevalence are an important metric for determining the proportion of serotypes in a given population. The purpose of this study was to evaluate the prevalence of individual GBS serotypes at Madigan Healthcare System (Madigan), the largest military tertiary healthcare facility in the Pacific Northwestern United States, and to compare seroprevalences with international locations. METHODS: To determine serotype distribution at Madigan, we obtained GBS isolates from standard-of-care anogenital swabs from 207 women of indeterminate gravidity between ages 18-40 during a five month interval. Serotype was determined using a recently described molecular method of polymerase chain reaction by capsular polysaccharide synthesis (cps) genes associated with pathogen virulence. RESULTS: Serotypes Ia, III, and V were the most prevalent (28%, 27%, and 17%, respectively). A systematic review of global GBS seroprevalence, meta-analysis, and statistical comparison revealed strikingly similar serodistibution at Madigan relative to civilian-sector populations in Canada and the United States. Serotype Ia was the only serotype consistently higher in North American populations relative to other geographic regions (p < 0.005). The number of non-typeable isolates was significantly lower in the study (p < 0.005). CONCLUSION: This study establishes PCR-based serotyping as a viable strategy for GBS epidemiological surveillance. Our results suggest that GBS seroprevalence remains stable in North America over the past two decades.

The orphan nuclear receptor Nr4a1 mediates perinatal neuroinflammation in a murine model of preterm labor.

Prematurity is associated with perinatal neuroinflammation and injury. Screening for genetic modulators in an LPS murine model of preterm birth revealed the upregulation of Nr4a1, an orphan nuclear transcription factor that is normally absent or limited in embryonic brains. Concurrently, Nr4a1 was downregulated with magnesium sulfate (MgSO4) and betamethasone (BMTZ) treatments administered to LPS exposed dams. To understand the role of Nr4a1 in perinatal brain injury, we compared the preterm neuroinflammatory response in Nr4a1 knockout (KO) versus wild type (wt) mice. Key inflammatory factors Il1b, Il6 and Tnf, and Iba1+ microglia were significantly lower in Nr4a1 KO versus wt brains exposed to LPS in utero. Treatment with MgSO4/BMTZ mitigated the neuroinflammatory process in wt but not Nr4a1 KO brains. These results correspond with a reduction in cerebral hemorrhage in wt but not mutant embryos from dams given MgSO4/BMTZ. Further analysis with Nr4a1-GFP-Cre × tdTomato loxP reporter mice revealed that the upregulation of Nr4a1 with perinatal neuroinflammation occurs in the cerebral vasculature. Altogether, this study implicates Nr4a1 in the developing vasculature as a potent mediator of neuroinflammatory brain injury that occurs with preterm birth. It is also possible that MgSO4/BMTZ mitigates this process by direct or indirect inhibition of Nr4a1.

Correction: The orphan nuclear receptor Nr4a1 mediates perinatal neuroinflammation in a murine model of preterm labor.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Sciatic nerve ligation-induced proliferation of spinal cord astrocytes is mediated by kappa opioid activation of p38 mitogen-activated protein kinase.

Partial sciatic nerve ligation (pSNL) markedly increased glial fibrillary acidic protein immunoreactivity (GFAP-IR) 1 week after lesion in the L4-L5 spinal dorsal horn of wild-type, but not in dynorphin knock-out, mice lacking kappa opioid receptors (KOR-/-) or in wild-type mice pretreated with the KOR antagonist nor-binaltorphimine (norBNI). A direct effect of KOR on glial cell proliferation was suggested by the findings that primary cultures of type II GFAP-immunoreactive astrocytes isolated from mouse spinal cord express KOR. Sustained treatment with the kappa agonist U50,488 (trans-3,4-dichloro-N-methyl-N-[2-(1-pyrolytinil)-cyclohexyl]-benzeneacetamide methane sulfonate) significantly increased the proliferation rate of GFAP-immunoreactive astrocytes isolated from wild-type mice, and this effect was blocked by norBNI pretreatment. Proliferation of cultured type II astrocytes may have been stimulated by mitogen-activated protein kinase (MAPK) activation by KOR because (1) U50,488 treatment increased phospho-p38 MAPK-immunoreactivity 247 +/- 44% over untreated cells, (2) the increase in phospho-p38 induced by U50,488 was blocked by norBNI and not evident in KOR-/- cultures, and (3) GFAP-immunoreactive astrocyte proliferation induced by U50,488 was blocked by the p38 MAPK inhibitor SB 203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole]. Similar mechanisms of astrocyte activation may also be responsible in vivo because intrathecal injection of SB 203580 blocked the increased GFAP-IR in lumbar spinal cord induced by pSNL. Although the relationship between kappa-stimulated astrocyte proliferation and neuropathic pain mechanisms was not directly established in these studies, the results support the hypothesis that KOR activation induces spinal astrocyte proliferation, which may contribute to cellular reorganization after sciatic nerve damage.

Far-Forward Diagnostics in Toxic Industrial Chemical and Material Exposure Scenarios and Biomarker Identification.

: This study describes key technical solutions for detecting environmental toxicants and diagnosing adverse health effects in military operational settings as outlined at a symposium cosponsored by the Department of Defense and the Johns Hopkins University-Applied Physics Laboratory (October 27 to 28, 2015). Such technologies are urgently needed in order to provide critical decision-aid tools and prognostic assessment of potential clinical sequelae. This review summarizes the state-of-the-science on (1) prioritization of adverse health effects, (2) existing technologies and diagnostic tools available for use in theater, (3) challenges to advancing diagnostic tools far-forward, and (4) the potential utility of anchoring diagnostic tools to adverse outcome pathways. Emerging technologies are increasingly available for physiological, environmental, and individual exposure monitoring. Challenges to overcome in austere environments include cold chain requirements and determination of adequate sampling intervals.

Potential and Actual Health Hazards in the Dense Urban Operational Environment: Critical Gaps and Solutions for Military Occupational Health.

: This paper presents environmental health risks which are prevalent in dense urban environments.We review the current literature and recommendations proposed by environmental medicine experts in a 2-day symposium sponsored by the Department of Defense and supported by the Johns Hopkins University Applied Physics Laboratory.Key hazards in the dense urban operational environment include toxic industrial chemicals and materials, water pollution and sewage, and air pollution. Four critical gaps in environmental medicine were identified: prioritizing chemical and environmental concerns, developing mobile decision aids, personalized health assessments, and better real-time health biomonitoring.As populations continue to concentrate in cities, civilian and military leaders will need to meet emerging environmental health concerns by developing and delivering adequate technology and policy solutions.