bluuebunny/crossref_metadata_2025 huggingface dataset:
Extracellular Matrix Degradation by Cultured Mesangial Cells: Mediators and Modulators
Authors: Baricos William H. , Reed J. Christopher , Cortez Shirley L.
Published: July 2017

Decreased degradation of the glomerular extracellular matrix (ECM) is thought to contribute to the accumulation of glomerular ECM that occurs in diabetic nephropathy and other chronic renal diseases. Several lines of evidence indicate a key role for the plasminogen activator/plasminogen/plasmin system in glomerular ECM degradation. However, which of the two plasminogen activators (PAs) present in renal tissue, tissue plasminogen activator (tPA) or urokinase-type plasminogen activator (uPA), is responsible for plasmin generation and those factors that modulate the activity of this system remain unclear. This study utilized mesangial cells isolated from mice with gene deletions for tPA, uPA, and plasminogen activator inhibitor 1 (PAI-1) to further delineate the role of the PA/plasminogen/plasmin system in ECM accumulation. ECM degradation by uPA-null mesangial cells was not significantly different from controls (92% ± 1%, n = 12). In contrast, ECM degradation by tPA-null mesangial cells was markedly reduced (–78 ± 1%, n = 12, P < 0.05) compared with controls, whereas tPA/uPA double-null mesangial cells degraded virtually no ECM. Previous studies from this laboratory have established that transforming growth factor-β1 (TGFβ1) inhibits ECM degradation by cultured mesangial cells by increasing the production of PAI-1, the major physiological PA inhibitor. In keeping with this observation, TGFβ1 (1 ng/ml) had no effect on ECM degradation by PAI-1-null MC. High glucose levels (30 m M) in the presence or absence of insulin (0.1 m M) caused a moderate increase in ECM degradation by normal human mesangial cells. In contrast, glycated albumin, whose concentration is known to increase in diabetes, produced a dose-dependent (0.2–0.5 mg/ml) inhibition of ECM degradation by normal human mesangial cells. Taken together, these results document the importance of tPA versus uPA in renal plasmin production and indicate that in contrast to elevated glucose, glycated albumin may contribute to ECM accumulation in diabetic nephropathy.
Erythropoietin-Induced Metallothionein Gene Expression: Role in Proliferation of K562 Cells
Authors: Abdel-Mageed Asim B. , Zhao Fusheng , Rider Barbara J. , Agrawal Krishna C.
Published: July 2017

Recent evidence has demonstrated an appreciable expression of metallothionein (MT) in erythrocytes. However, the induction of the MT protein by hematopoietic growth factors and its subsequent functional significance on clonal expansion or differentiation of erythroid progenitor cells remain elusive. We therefore examined the effects of growth factors erythropoietin (EPO), granulocyte-monocyte colony-stimulating factor (GM-CSF), and interleukin-3 (IL-3) on MT gene expression in erythroid progenitor K562 cell line. EPO, but not IL-3 or GM-CSF, induced a 3-fold increase in MT transcripts in K562 cells. MT induction was associated with EPO-induced cellular proliferation, suggesting a specific role for MT induction by EPO in erythroid progenitor cells. However, EPO- or sodium butyrate-induced differentiation as monitored by hemoglobin formation was inhibited in K562 cells stably transfected with an expression vector containing human MT-IIA gene. This inhibition of differentiation was partially reversed in these cells by an MT antisense phosphorothioate oligonucleotide. Furthermore, the MT-induced inhibition of differentiation was associated with downregulation of EPO receptor transcripts in K562 cells. These data suggest that, among growth factors required for erythropoiesis, EPO is a potent inducer of MT, and that MT may plays a significant role in EPO-induced proliferation, but not in the erythroid-specific differentiation of the progenitor cells.
Nitric Oxide as a Local Mediator of Prostaglandin F2α-Induced Regression in Bovine Corpus Luteum: An In Vivo Study
Authors: Jaroszewski Jerzy J. , Skarzynski Dariusz J. , Hansel William
Published: July 2017

To test whether nitric oxide (NO) is involved in prostaglandin (PG) F2α-induced regression of the bovine corpus luteum (CL) in vivo, heifers were treated as follows: Group 1, saline (3 ml/h); Group 2, dinoprost, an analogue of prostaglandin F2α (aPGF2α; 5 mg/0.5 h); Group III, Nω-nitro-L-arginine methyl ester (L-NAME; 200 mg/4 h), an inhibitor of nitric oxide synthase; and Group IV, L-NAME (400 mg/4 h) and aPGF2α (5 mg/0.5 h). All treatments were administered by an intraluteal microdialysis system (MDS) on day 15 of the cycle. Perfusate and jugular plasma samples were collected at half-hour intervals; additionally, jugular plasma samples were collected once daily from day 16 to day 21 of the cycle. In the perfusate samples, aPGF2α increased P4 ( P < 0.05), PGE2 ( P < 0.001), and LTC4 ( P < 0.05) concentrations; L-NAME increased P4 ( P < 0.05) but did not change PGE2 and LTC4 ( P > 0.05) concentrations as compared with the period before treatment. Simultaneous perfusion of CL with L-NAME and aPGF2α caused a further increase of P4 concentration ( P < 0.05) induced by L-NAME or aPGF2α treatment and increased PGE2 and LTC4 ( P < 0.001) concentrations to the level observed after aPGF2α treatment. Perfusion of CL with aPGF2α caused luteal regression within 24 h, while perfusion with L-NAME prolonged the life span of CL to day 21 ( P < 0.05). Concomitant L-NAME and aPGF2α treatment partially counteracted ( P < 0.05) the luteal regression caused by aPGF2α administration. These results show that NO is involved in the process of luteolysis in the bovine CL and suggest that the luteolytic effect of aPGF2α may be mediated by NO as an important component of an autocrine/paracrine luteolytic cascade.
Dissecting Autocrine Effects on Pulsatile Release of Gonadotropin-Releasing Hormone in Cultured Rat Hypothalamic Tissue
Authors: Woller Michael J. , Meyer Scott , Ada-Nguema Aude , Waechter-Brulla Daryle
Published: July 2017

The control of reproductive function is manifested centrally through the control of hypothalamic release of gonadotropin-releasing hormone (GnRH) in episodic events or pulses. For GnRH release to occur in pulses, GnRH neurons must coordinate release events periodically to elicit a bolus of GnRH. We used a perifusion culture system to examine the release of GnRH from both intact hypothalami and enzymatically dispersed hypothalamic cells after challenge with GnRH analogs to evaluate the role of anatomical neuronal connections on autocrine/paracrine signals by GnRH on GnRH neurons. The potent GnRH agonist des-Gly10-D-Ala6-GnRH N-ethylamide, potent GnRH antagonists D-Phe2-D-Ala6-GnRH and D-Phe2,6-Pro3-GnRH or vehicle were infused, whereas GnRH release from hypothalamic tissue and cells were measured. PULSAR analysis of GnRH release profiles was conducted to evaluate parameters of pulsatile GnRH release. Infusion of the GnRH agonist resulted in a decrease in mean GnRH (P < 0.001), pulse nadir (P < 0.01), and pulse frequency (P < 0.05) but no effect on pulse amplitude. Infusion of GnRH antagonists resulted in an increase in mean GnRH (P < 0.001), pulse nadir (P < 0.05), and pulse frequency (P < 0.05) and in GnRH pulse amplitude only in dispersed cells (P < 0.05). These results are consistent with the hypothesis that GnRH inhibits endogenous GnRH release by an ultrashort-loop feedback mechanism and that treatment of hypothalamic tissue or cells with GnRH agonist inhibits ultrashort-loop feedback, whereas treatment with antagonists disrupts normal feedback to GnRH neurons and elicits an increased GnRH signal.
Different Responses of Atrial Natriuretic Peptide Secretion and Its Receptor Density to Salt Intake in Rats
Authors: Lee Kyung Sun , Kim So Young , Han Jeong Hee , Kim Yun Ah , Cao Chunhua , Kim Sung Zoo , Cho Kyung Woo , Kim Suhn Hee
Published: July 2017

This study investigated whether high-salt intake influences atrial natriuretic peptide (ANP) system, atrial content, and release rate of ANP as well as receptor density in the kidney were measured in salt intake rats. Male Sprague-Dawley rats received either 0.9% or 2% salt in their drinking water for 10 days. The stretch-induced ANP secretion from isolated perfused nonbeating left atria was accentuated, and the production of cGMP by ANP in renal cortical tissue membranes were pronounced in rats exposed to 0.9% salt for 10 days but not in rats exposed to 2% salt. The levels of ANP receptor density and expression in renal cortex were decreased in 2% salt intake rats but not in 0.9% salt intake rats. No significant differences in atrial and plasma concentrations of ANP and water balance were observed in both salt intakes. Therefore, these results suggest that atrial ANP secretion and its binding sites in the kidney may respond differently to ingested salt concentrations in rats.
Inhibition of Stimulated Ascorbic Acid and Luteinizing Hormone-Releasing Hormone Release by Nitric Oxide Synthase or Guanyl Cyclase Inhibitors
Authors: Karanth Sharada , Yu Wen H. , Mastronardi Claudio A , McCann Samuel M.
Published: July 2017

Ascorbic acid (AA), an antioxidant, is present in high concentrations in the hypothalamus. Previously, we have shown that AA inhibited stimulated release of luteinizing hormone-releasing hormone (LHRH) from medial basal hypothalami in vitro. We have also demonstrated that cell membrane depolarization by high [K+] media-induced AA release that is blocked by NG-monomethyl-l-arginine, a competitive inhibitor of nitric oxide synthase (NOS), indicating that the release process is mediated by NO. The release of LHRH is also mediated by NO. We hypothesized that AA is a co-transmitter released with classical transmitters from synaptic vesicles that acts to reduce chemically the NO formed, thereby providing feed-forward inhibitory control over LHRH release. Because NO acts by activating guanylyl cyclase (GC) resulting in production of cGMP, in the present investigation we studied the effects of an NOS inhibitor LY 83583 and GC inhibitor, O.D.Q. to further characterize the role of NO in high [K+]-induced AA and LHRH release. Medial basal hypothalami were incubated in 0.5 ml of Krebs-Ringer Bicarbonate buffer or medium containing increased potassium [K+ = 56 mM] for 1 hr or combinations of high [K+] + LY 83583 or O.D.Q. for 1 hr. AA and LHRH released into the incubation medium were measured by high-pressure liquid chromatography and radioimmunoassay, respectively. Cell membrane depolarization with high [K+] produced a significant increase in both AA and LHRH release. A combination of high [K+] + LY 83583 or high [K+] + O.D.Q. decreased basal AA and completely blocked high [K+]-induced AA and LHRH release. As in the case of high [K+], LHRH release induced by the excitatory amino acid N-methyl-d-aspartic acid (NMDA) was blocked by both the inhibitors. NMDA alone failed to alter AA release, but the combined presence of NMDA and the inhibitors totally blocked AA release. Because LY 83583 and O.D.Q. were shown to inhibit NOS and soluble GC, respectively, the data demonstrate that basal and high [K+]-induced AA and high [K+] and NMDA-stimulated LHRH release were mediated by NO by its activation of GC and consequent generation of cGMP.
Surfactant Releases Internal Calcium Stores in Neutrophils by G Protein–Activated Pathway
Authors: Boston Mark E. , Frech G. Christopher , Chacon-Cruz Enrique , Buescher E. Stephen , Oelberg David G.
Published: July 2017

Pulmonary surfactant with surfactant-associated proteins (PS+SAP) decreases pulmonary inflammation by suppressing neutrophil activation. We have observed that PS+SAP inserts channels into artificial membranes, depolarizes neutrophils, and depresses calcium influx and function in stimulated neutrophils. We hypothesize that PS+SAP suppresses neutrophil activation by depletion of internal Ca++stores and that PS+SAP induces depletion through release of Ca++stores and through inhibition of Ca++influx. Our model predicts that PS+SAP releases Ca++stores through insertion of channels, depolarization of neutrophils, and activation of a G protein–dependent pathway. If the model of channel insertion and membrane depolarization is accurate, then gramicidin—a channel protein with properties similar to those of PS+SAP—is expected to mimic these effects. Human neutrophils were monitored for [Ca++] responses after exposure to one of two different PS+SAP preparations, a PS-SAP preparation, gramicidin alone, and gramicidin reconstituted with phospholipid (PLG). [Ca++] responses were reexamined following preexposure to inhibitors of internal Ca++release or the G protein pathway. We observed that (i) 1% PS+SAP—but not PS-SAP—causes transient increase of neutrophil [Ca++] within seconds of exposure; (ii) 1% PLG—but not gramicidin alone—closely mimics the effect of PS+SAP on Ca++response; (iii) PS+SAP and PLG equally depolarize neutrophils; (iv) direct inhibition of internal Ca++stores releases or of G protein activation suppresses Ca++responses to PS+SAP and PLG; and (v) preexposure to either PS+SAP or PLG inhibits Ca++influx following fMLP stimulation. We conclude that PS+SAP independently depolarizes neutrophils, releases Ca++from internal stores by a G protein-mediated pathway, and alters subsequent neutrophil response to physiologic stimulants by depleting internal Ca++stores and by inhibiting Ca++influx during subsequent fMLP activation. The mimicking of these results by PLG supports the hypothesis that PS+SAP initiates depolarization via channel insertion into neutrophil plasma membrane.
The Influence of Estradiol and Diet on Small Intestinal Glucose Transport in Ovariectomized Rats
Authors: Kimura Yasuhiro , Buddington Karyl K. , Buddington Randal K.
Published: July 2017

Although gender differences exist for intestinal absorption of nutrients and drugs, the possible role estradiol may play in modulating nutrient transport has not been established. Therefore, small intestine glucose transport was measured 1 week after administering estradiol to ovariectomized rats fed diets high in carbohydrate (C) or protein (P). Rats treated with estradiol ate 21% less (P < 0.05) and lost body mass (7%; P < 0.05) but did not have smaller intestines. Administration of estradiol increased rates of glucose transport, but only when the rats were fed the C diet. These findings indicate that estradiol causes a disconnect between food intake and the dimensions and nutrient transport capacities of the small intestine. Furthermore, the responses to estradiol are influenced by diet composition, are not of the same magnitude for rats and dogs, and can be predicted to affect systemic availability of nutrients and drugs.
Extract of Perilla frutescens Enriched for Rosmarinic Acid, a Polyphenolic Phytochemical, Inhibits Seasonal Allergic Rhinoconjunctivitis in Humans
Authors: Takano Hirohisa , Osakabe Naomi , Sanbongi Chiaki , Yanagisawa Rie , Inoue Ken-ichiro , Yasuda Akiko , Natsume Midori , Baba Seigo , Ichiishi Ei-ichiro , Yoshikawa Toshikazu
Published: July 2017

Extract of Perilla frutescens enriched for rosmarinic acid, a polyphenolic phytochemical, suppresses allergic immunoglobulin responses and inflammation caused by polymorphonuclear leukocytes (PMNL) in mice. However, few placebo-controlled clinical trials have examined the efficacy and safety of polyphenolic phytochemicals for treatment of allergic inflammatory diseases in humans. The present study determined whether oral supplementation with rosmarinic acid is an effective intervention for patients with seasonal allergic rhinoconjunctivitis (SAR). In this 21-day, randomized, double-blind, age-matched, placebo-controlled parallel group study, patients with mild SAR were treated daily with extract of Perilla frutescens enriched for rosmarinic acid (200 mg [n = 10] or 50 mg [n = 9]) or placebo (n = 10). Patients recorded symptoms daily in a diary. Profiles of infiltrating cells and concentrations of eotaxin, IL-1β, IL-8, and histamine were measured in nasal lavage fluid. Serum IgE concentrations and routine blood tests were also examined. As compared with placebo supplementation, supplementation with extract of Perilla frutescens enriched for rosmarinic acid resulted in a significant increase in responder rates for itchy nose, watery eyes, itchy eyes, and total symptoms (P < 0.05). Active treatment significantly decreased the numbers of neutrophils and eosinophils in nasal lavage fluid (P < 0.05 vs. placebo). Patients reported no adverse events, and no significant abnormalities were detected in routine blood tests. In conclusion, extract of Perilla frutescens enriched for rosmarinic acid can be an effective intervention for mild SAR at least partly through inhibition of PMNL infiltration into the nostrils. Use of this alternative treatment for SAR might reduce treatment costs for allergic diseases.
Combined Treatment with Vessel Dilator and Kaliuretic Hormone in Persons with Congestive Heart Failure
Authors: Patel Hamang , Dietz John R. , Owen Ashley , Miguel Gloria San , Mccormick Michael T. , Schocken Douglas D. , Vesely David L.
Published: July 2017

Vessel dilator and kaliuretic hormone, two cardiovascular peptide hormones, enhance urine flow 2- to 13-fold and 4-fold, respectively, in persons with class III New York Heart Association congestive heart failure (CHF). The natriuresis and diuresis secondary to vessel dilator and kaliuretic hormone are not blunted as are atrial natriuretic peptide and brain natriuretic peptide effects in persons with CHF compared with healthy individuals. The present investigation determined if the two peptide hormones that do not have blunted effects in persons with CHF may have added beneficial effects when given simultaneously to individuals with class III CHF. Together with each at 100 ng/kg of body weight per minute, vessel dilator and kaliuretic hormone increased urine flow rate 3.5-fold (P < 0.05) compared with their 60-min baseline and control CHF subjects' urine flow rates. Combined, they enhanced the excretion rate of sodium a maximum of 3.6-fold (P < 0.05) with 2.5- and 2-fold enhancement 2 and 3 hrs after infusion. These data indicate that vessel dilator and kaliuretic hormone have diuretic and natriuretic effects when used in combination, but these effects are not additive over their individual effects in persons with CHF.
Inflammation and Ischemia: Macrophages Activated by Fibronectin Fragments Enhance the Survival of Injured Cardiac Myocytes
Authors: Trial JoAnn , Rossen Roger D. , Rubio Jose , Knowlton Anne A.
Published: July 2017

Proteolytic enzymes, released early in the course of an inflammatory response, hydrolyze fibronectin, producing fragments of the parent molecule that alter monocyte phenotype and migratory behavior. Here we test the hypothesis that macrophages, stimulated by the dominant 110–120 kd fibronectin fragments (FNf), as are found in lymphatic fluid draining sites of cardiac ischemia-reperfusion injury, produce factors that promote the survival of injured parenchymal cells. Rat splenic macrophages stimulated in vitro with purified FNf produced soluble factors that protected hypoxic rat cardiac myocytes from death by apoptosis. Addition of blocking antibodies specific for tumor necrosis factor-α (TNF-α), fibroblast growth factor-1 (FGF-1), insulin-like growth factor I (IGF-I), and leukemia inhibitory factor (LIF) partly reduced the protection against apoptosis provided to hypoxic cardiac myocytes by cell-free culture supernatants from FNf-stimulated macrophages. Complete blockade of this protection was achieved by a combination of antibodies specific for FGF-1, IGF-I, and LIF. Stimulation of human monocyte-derived macrophages in vitro with FNf significantly increased their output of TNF-α, FGF-1, IGF-I, and LIF. These results suggest that tissue degradation products, released in the early hours of an inflammatory response, stimulate tissue-infiltrating macrophages to protect injured but still viable parenchymal cells from death by apoptosis.
Commensal Bacteria, Redox Stress, and Colorectal Cancer: Mechanisms and Models
Authors: Huycke Mark M. , Gaskins H. Rex
Published: July 2017

The potential role for commensal bacteria in colorectal carcinogenesis is explored in this review. Most colorectal cancers (CRCs) occur sporadically and arise from the gradual accumulation of mutations in genes regulating cell growth and DNA repair. Genetic mutations followed by clonal selection result in the transformation of normal cells into malignant derivatives. Numerous toxicological effects of colonic bacteria have been reported. However, those recognized as damaging epithelial cell DNA are most easily reconciled with the currently understood genetic basis for sporadic CRC. Thus, we focus on mechanisms by which particular commensal bacteria may convert dietary procarcinogens into DNA damaging agents (e.g., ethanol and heterocyclic amines) or directly generate carcinogens (e.g., fecapentaenes). Although these and other metabolic activities have yet to be linked directly to sporadic CRC, several lines of investigation are reviewed to highlight difficulties and progress in the area. Particular focus is given to commensal bacteria that alter the epithelial redox environment, such as production of oxygen radicals by Enterococcus faecalis or production of hydrogen sulfide by sulfate-reducing bacteria (SRB). Super-oxide-produclng E. faecalis has conclusively been shown to cause colonic epithelial cell DNA damage. Though SRB-derived hydrogen sulfide (H2S) has not been reported thus far to induce DNA damage or function as a carcinogen, recent data demonstrate that this reductant activates molecular pathways implicated in CRC. These observations combined with evidence that SRB carriage may be genetically encoded evoke a working model that Incorporates multifactorial gene-environment interactions that appear to underlie the development of sporadic CRC.
Characterization of the 5′-Flanking Transcriptional Regulatory Region of Chicken Growth Hormone Gene
Authors: Ip Stephen C. Y. , Lau Joanna S. , Au W. L. , Leung Frederick C.
Published: July 2017

A 1727-bp fragment of 5-flanking region of chicken growth hormone (cGH) gene has been cloned and sequenced. Various lengths of the 5-flanking region (122 to 1775 bp) was linked to a luciferase reporter gene, and its transcriptional regulation was examined by an in vitro transient transfection coupled with luciferase assay. Our results demonstrated that pituitary-specific transcription factor, Pit-1, is necessary and sufficient to confer a strong tissue-specific expression. Co-transfection with goldfish or chicken Pit-1 expression vectors significantly restored the luciferase expression in HeLa cells. Site-directed mutagenesis and mobility gel-shift assays further confirmed the position of the Pit-1 binding site at –113/–104. Moreover, a repressive thyroid hormone response element (TRE) was identified at –137/–74, and we propose that interactions between the TRE and Pit-1 sites may be required for its repressive effect.
Effect of Male Sex and Obesity on Platelet Arachidonic Acid in Spontaneous Hypertensive Heart Failure Rats
Authors: Park Sonhee C. , Liu-Stratton Yiwen , Medeiros Lydia C. , Mccune Sylvia A. , Radin M. Judith
Published: July 2017

Sexual dimorphism is observed in the progression to congestive heart failure and, ultimately, in longevity in spontaneously hypertensive heart failure (SHHF) rats. As platelet activation may impact development of cardiovascular diseases, we studied the effects of obesity and sex on platelet polyunsaturated fatty acid (PUFA) profile and its relationship to platelet aggregation in 6-month-old SHHF rats. After a 24-hr fast, blood was obtained for measurement of platelet phospholipid omega-6 (n-6) and omega-3 (n-3) PUFA. Collagen-induced platelet aggregation was measured by whole-blood impedance aggregometry. Obese male (OM) SHHF had significantly more platelet arachidonic acid (AA) and total n-6 PUFA than lean males (LMs), lean females (LFs), or obese females (OFs). Platelet aggregation was enhanced in males compared to females, with OMs by 45% compared to OFs and with LMs by 28% compared to LFs. Though no difference was found between OFs and LFs, platelet agregation was increased in OMs by 20% compared to LMs. Though not significantly different, lag time to initiate platelet aggregation tended to be shortest in OMs and then, in Increasing duration, LMs, LFs, and OFs, suggesting that Platelets from male rats were quicker to aggregate than those from females. Platelet aggregation was correlated with platelet AA and total n-6 PUFA content. There was no relationship between n-3 PUFA and platelet aggregation. In SHHF rats, elevated AA and n-6 PUFA levels in platelets are associated with Chanced platelet aggregation. This relationship is potentiated by obesity and male sex.
Resuscitation Affects Microcirculatory Polymorphonuclear Leukocyte Behavior After Hemorrhagic Shock: Role of Hypertonic Saline and Pentoxifylline
Authors: Yada-Langui Margareth M. , Anjos-Valotta Edna A. , Sannomiya Paulina , Silva Mauricio Rocha E , Coimbra Raul
Published: July 2017

We have previously shown that lung injury following fluid resuscitation either with hypertonic saline (HS) or lactated Ringer's (LR) plus pentoxifylline (PTX) attenuated acute lung injury when compared with LR resuscitation. The objective of the present study is to determine whether our previous observations are accompanied by changes in polymorphonuclear leukocyte (PMN) behavior. To study this, PMN-endothelial cell interactions, microcirculatory blood flow, lung histology, lung PMN infiltration (WIPO, Myeloperoxidase), and lung intracellular adhesion molecule-1 (ICAM-1) expression were assessed in a controlled hemorrhagic shock model followed by LR, HS, and LR+PTX resuscitation in rodents. Rats (240-300 g) were bled to a mean arterial pressure (MAP) of 35 mm Hg for 1 hr and then randomized into three groups: HS (7.5% NaCl, 4 ml/kg); LR (3× shed blood); and LR+PTX (25 mg/kg). Additionally, total shed blood was reinfused. A sham group underwent no shock and no treatment. The internal spermatic fascia was exteriorized and the microcirculation was observed by closed-circuit TV coupled to a microscope, 2 and 6 hrs after treatment. The number of leukocytes sticking to the venular endothelium was determined 2 hrs after fluid resuscitation. Microcirculatory blood flow was measured by an optical Doppler velocimeter. Lung histology and lung MPO immunostaining were assessed at 6 hrs, and lung ICAM-1 expression was determined by immunostaining at 2 hrs following fluid resuscitation. Two hours after treatment, HS (1.4 ± 0.4), LR+PTX (1.7 ± 0.3), and sham (0.4 ± 0.2) groups presented significant reductions in leukocyte adherence (cells/100 urn venule length), compared with the LR group (4.0 ± 0.9, P < 0.05). No differences were observed 6 hrs after treatment on leukocyte adherence and microcirculatory blood flow. ICAM-1 expression was significantly higher in LR- treated animals compared with the HS, LR+PTX, and sham groups (P < 0.01). PMN infiltration and overall lung injury were significantly attenuated by HS and LR+PTX. These results support earlier studies that indicated the potential application of HS and PTX in shock therapy and the increase in PMN- endothelial cell interaction and lung injury after LR resuscitation.
Genomic Approaches That Aid in the Identification of Transcription Factor Target Genes
Authors: Kirmizis Antonis , Farnham Peggy J.
Published: July 2017

It is well-established that deregulation of the transcriptional activity of many different genes has been causatively linked to human diseases. In cancer, altered patterns of gene expression are often the result of the inappropriate expression of a specific transcriptional activator or repressor. Functional studies of cancer-specific transcription factors have relied upon the study of candidate target genes. More recently, gene expression profiling using DNA microarrays that contain tens of thousands of cDNAs corresponding to human mRNAs has allowed for a large-scale identification of genes that respond to increased or decreased levels of a particular transcription factor. However, such experiments do not distinguish direct versus indirect target genes. Coupling chromatin immunoprecipitation to microarrays that contain genomic regions (ChIP-chip) has provided investigators with the ability to identify, in a high-throughput manner, promoters directly bound by specific transcription factors. Clearly, knowledge gained from both types of arrays provides complementary information, allowing greater confidence that a transcription factor regulates a particular gene. In this review, we focus on Polycomb group (PcG) complexes as an example of transcriptional regulators that are implicated in various cellular processes but about which very little is known concerning their target gene specificity. We provide examples of how both expression arrays and ChIP-chip microarray-based assays can be used to identify target genes of a particular PcG complex and suggest improvements in the application of array technology for faster and more comprehensive identification of directly regulated target genes.
Differential Increases in Syntheses of Newly Identified Trypsinogen 2 Isoforms by Dietary Protein in Rat Pancreas
Authors: Hara Hiroshi , Shiota Hiromichi
Published: July 2017

We have found that dietary protein markedly induced pancreatic serine protease activity via a mechanism independent of luminal trypsin activity in pancreatico-biliary–diverted (PBD) rats. The aim of this study was to examine the effects of dietary protein on the synthesis of trypsinogen isoforms by comparing in vivo incorporation of [35S] l-methionine into isoform proteins in PBD and sham-operated rats. A small duodenal segment including the ampulla of Vater was sectioned and transposed to the upper ileum with end-to-side anastomosis (PBD) or duodenal transection was followed by reanastomosis (sham) in male Sprague-Dawley rats. After recovery, PBD and sham rats were fed a 25% or 60% casein-sucrose–based diet (NC or HC) for 14 days. Rats were then intravenously injected with [35S] l-methionine (15 MBq/kg body weight) 30 mins before being sacrificed for analysis of pancreatic enzymes by two-dimensional SDS-polyacrylamide gel electrophoresis. By using electrophoresis with narrow range of isoelectric focusing (pl 4.5–5.5), five trypsinogen 2 (2-x) isoform spots were identified using both [35S] incorporation and Coomassie brilliant blue (CBB) staining in PBD rats, but not in sham rats. N-terminal sequences of these trypsinogen 2-x spots were identical to known rat trypsinogen 2 with the exception that the third valine was changed to isoleucine in one isoform. In PBD rats, feeding of HC specifically increased the [35S] and CBB intensities of these trypsinogen 2-x isoforms and trypsinogen 3. The degree of induction of the five trypsinogen 2-x molecules by HC varied greatly. Trypsinogen 1 and 4, which are the major trypsinogens in normal rats, showed no changes. We conclude that increases in synthesis of a few newly identified trypsinogen 2-x isoforms mainly contribute to the induction of trypsin activity in the pancreas by HC in PBD rats.
Effects of Cardiac Hormones on Arterial Pressure and Sodium Excretion in NPRA Knockout Mice
Authors: Dietz John R. , Landon Carol S. , Nazian Stanley J. , Vesely David L. , Gower William R.
Published: July 2017

These studies were designed to determine if the atria contains natriuretic substances that act through a non–natriuretic peptide type A (NPRA) receptor mechanism. C57BL/6 mice, either wild-type NPRA++ (WT) or NPRA —- knockout (KO), were anesthetized with pentobarbital. Catheters were placed in the trachea, carotid artery, jugular vein, and bladder. Urine was collected for six 30-min periods. Both groups received an iv injection of 100 ng of rat atrial natriuetic peptide (rANP) in 200 μl of saline after the first period (30 mins) and 200 μl of rat atrial extract after the fourth period (120 mins). ANP injection increased urine flow (UF) to 2.7 ± 0.5 μl/min in the WT versus 1.9 ± 0.2 in KO. Extract increased UF to 7.9 ± 1.5 μl/min in WT versus 2.7 ± 0.4 in KO (P < 0.01). ANP increased sodium excretion (ENa) to 0.47 ± 0.10 μmoles/min in WT versus 0.27 ± 0.04 in KO (P < 0.05). Extract increased ENa to 1.44 ± 0.47 μmoles/min in WT versus 0.26 ± 0.06 in KO (P < 0.05). Extract decreased mean arterial pressure (MAP) to 62 ± 3 mm Hg in the WT versus 81 ± 5 in KO (P < 0.01). ENa and MAP responses to extract in KO were not different from responses to 200 μl of saline. A constant 150-min infusion of rat atrial extract increased urine flow by 3-fold and ENa by 5-fold (both P < 0.05) in the WT mice but had no significant effect in the KO mice. Thus, acute renal and MAP responses to atrial extracts require the NPRA receptor.
Dietary Organic Isothiocyanates are Cytotoxic in Human Breast Cancer MCF-7 and Mammary Epithelial MCF-12A Cell Lines
Authors: Tseng Elaine , Ramsay Elizabeth A. Scott , Morris Marilyn E.
Published: July 2017

Organic isothiocyanates (ITCs) are dietary components present in cruciferous vegetables. The purpose of this investigation was to examine the cytotoxicity of 1-naphthyl isothiocyanate (NITC), benzyl isothiocyanate (BITC), β-phenethyl isothiocyanate (PEITC), and sulforaphane in human breast cancer MCF-7 and human mammary epithelium MCF-12A cell lines, as well as in a second human epithelial cell line, human kidney HK-2 cells. The cytotoxicity of NITC, BITC, PEITC, and sulforaphane, as well as the cytotoxicity of the chemotherapeutic agents daunomycin (DNM) and vinblastine (VBL), were examined in MCF-7/sensitive (wt), MCF-7/Adr (which overexpresses P-glycoprotein), MCF-12A, and HK-2 cells. Cell growth was determined by a sulforhod-amine B assay. The IC50 values for DNM and VBL in MCF-7/Adr cells were 7.12 ± 0.42 μM and 0.106 ± 0.004 μM (mean ± SE) following a 48-hr exposure; IC50 values for BITC, PEITC, NITC, and sulforaphane were 5.95 ± 0.10, 7.32 ± 0.25, 77.9 ± 8.03, and 13.7 ± 0.82 μM, respectively, with similar values obtained in MCF-7/wt cells. Corresponding values for BITC, PEITC, NITC, and sulforaphane in MCF-12A cells were 8.07 ± 0.29, 7.71 ± 0.07, 33.6 ± 1.69, and 40.5 ± 1.25 μM, respectively. BITC and PEITC can inhibit the growth of human breast cancer cells as well as human mammary epithelium cells at concentrations similar to those of the chemotherapeutic drug DNM. Sulforaphane and NITC exhibited higher IC50 values. The effect of these ITCs on cell growth may contribute to the cancer chemopreventive properties of ITCs by suppressing the growth of preclinical tumors, and may indicate a potential use of these compounds as chemotherapeutic agents in cancer treatment.
Extracellular Cyclic AMP and Adenosine Appearance in Adipose Tissue of Sus scrofa: Effects of Exercise
Authors: Carey Gale B. , Wotjukiewicz Lindsay J. , Goodman Jessie M. , Reineck Kevin E. , Overman Kristen C.
Published: July 2017

Cyclic AMP (cAMP) appears extracellularly in a variety of tissues including brain, liver, and kidney; whether it appears in adipose tissue and responds to physiological perturbation is unknown. The purpose of this study was to examine adipose tissue extracellular cAMP appearance and metabolism in situ and in vitro in physiologically challenged animals. Littermate swine were either sedentary or exercise trained on a treadmill for 3 months and subjected to acute exercise on experiment day. In situ, microdialysis probes in subcutaneous back fat were perfused before, during, and after animals performed 20 mins of acute exercise, and dialysate was analyzed for cAMP and adenosine. In vitro, isolated adipocytes were hormonally stimulated to provoke cAMP synthesis and efflux, and plasma membrane phosphodiesterase and 5′-nucleotidase activities were measured. Extracellular cAMP and adenosine levels in adipose tissue of sedentary swine averaged 5.2 ± 1.7 and 863 ± 278 nM, respectively. Exercise training tended to increase extracellular cAMP (11.3 ± 1.7 n/lf) and reduce extracellular adenosine (438 ± 303 n/lf), although neither change was statistically significant. Acute exercise caused a significant 3-fold and 16-fold increase in extracellular cAMP and adenosine, respectively, compared to rest. These changes occurred despite a 2- to 3-fold increase in adipose tissue blood flow during acute exercise. In vitro, cAMP efflux from exercise-trained swine was 42% greater than that from adipocytes of sedentary swine, yet adipocyte plasma membranes from exercise-trained and sedentary swine did not differ in maximal phosphodiesterase and 5′-nucleotidase activities. We conclude that cAMP appears extracellularly in swine adipose tissue and that the levels of extracellular cAMP and adenosine in intact swine adipose tissue are influenced by both acute and chronic exercise. The subsequent impact of the changes in these biochemicals on local cellular metabolism and growth remains to be determined.
Leptin Acts Peripherally to Limit Meal-Induced Increases in Plasma Insulin Concentrations in Mice: A Brief Communication
Authors: Mistry Anahita M. , Swick Andrew G. , Romsos Dale R.
Published: July 2017

Leptin inhibits food intake and lowers plasma insulin concentrations. This study was designed to determine whether leptin acts independent of food-intake regulation to affect meal-induced increases in plasma insulin concentrations. Leptin-deficient, Lepob/Lepob mice were administered 1 μg leptin intracerebroventricularly (ICV) or intraperitoneally. Food intake and plasma insulin concentrations of mice administered leptin ICV before a meal were lower, as expected, than were intakes and plasma insulin concentrations of mice administered vehicle ICV. However when food intake was controlled, meal-induced increases in plasma insulin were unaffected by ICV administration of leptin. Intraperitoneal administration of 1 μg leptin before a meal lowered meal-induced increases in plasma insulin concentrations without influencing the size of the meal. We conclude that plasma leptin concentrations can affect meal-induced insulin secretion independent of the central nervous system actions of leptin associated with food-intake regulation.
Aminoguanidine Prevents Fructose-Induced Arterial Stiffening in Wistar Rats: Aortic Impedance Analysis
Authors: Lin Yi-Tsen , Tseng Yung-Zu , Chang Kuo-Chu
Published: July 2017

Fructose has been reported as a potent agent in forming advanced glycation end products (AGEs) and, thus, may play a significant role in the pathogenesis of diabetic complications. Herein, we determined the effects of aminoguanidine (AG), an inhibitor of AGEs, on the mechanical properties of the arterial system in fructose-fed (FF) rats, using aortic impedance analysis. Rats at 2 months were given 10% fructose in drinking water for 2 weeks and compared with untreated age-matched controls. Meanwhile, FF rats were treated for 2 weeks with AG (daily peritoneal injections of 50 mg kg−1) and compared with the untreated FF group. Neither fructose nor AG affects body weight, blood glucose level, and basal heart rate. In comparison with controls, FF rats showed a decrease in cardiac output in the absence of any significant changes in mean aortic pressure, having increased total peripheral resistance (Rp), at 51.1 ± 2.9 versus 66.2 ± 1.9 mm Hg sec ml−1 (P < 0.05). Fructose also contributed to an increase in aortic characteristic impedance (Zc), from 1.528 ± 0.094 to 1.933 ± 0.084 mm Hg sec ml−1 (P < 0.05) and a decrease in wave transit time (τ), from 22.6 ± 0.6 to 19.2 ± 0.7 msec (P < 0.05). The elevated Zc and the reduced τ suggest that fructose may cause a detriment to the aortic distensibility in animals. After exposure to AG, FF rats exhibited a significant improvement in physical properties of the resistance vessels, as evidenced by the reduction of 21.3% in Rp. Meanwhile, AG retarded the fructose-induced decline in aortic distensibility, as reflected in the decrease of 16.0% in Zc (P < 0.05) and the increase of 18.1% in τ (P < 0.05). By contrast, AG exerted no effects on the mechanical properties of Windkessel vessels, as well as resistance vessels, in normal diet controls. We conclude that AG may prevent the fructose-derived changes in arterial stiffening, possibly through inhibition of the fructose-derived advanced glycation end product formation in Wistar rats.
Low-Density Lipoprotein Inhibits Secretion of Phospholipid Transfer Protein in Human Trophoblastic BeWo Cells
Authors: Tu An-Yue , Cheung Marian C. , Zhu Xiaodong , Knopp Robert H. , Albers John J.
Published: July 2017

Human plasma phospholipid transfer protein (PLTP) plays an important role in lipoprotein metabolism. In this study, we investigated the effects of lipoproteins on the secretion of PLTP in cultured BeWo choriocarcinoma cells. Low-density lipoproteins (LDLs) decreased PLTP secretion in a dose- and time-dependent manner, whereas very low density lipoproteins and high-density lipoproteins (HDLs) had little effect. LDL suppression of PLTP secretion was not altered by the inhibition of both LDL receptor and LDL receptor–related protein with receptor-associated protein. Mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor, U0126, could abolish the LDL-mediated inhibition of PLTP secretion. Furthermore, LDL, but not HDL, could stimulate the expression of MAPK phosphatase-1 (MKP-1) in BeWo cells that resulted in the inactivation of p44/p42 extracellular signal-regulated kinase (ERK) 1 and 2, the family members of MAPKs. These results support the conclusion that LDL-mediated suppression of PLTP secretion in BeWo cells is through a LDL receptor-independent MAPK signaling pathway.
Role of Estrogens in Adipocyte Development and Function
Authors: Cooke Paul S. , Naaz Afia
Published: July 2017

Estrogen has historically been viewed as a major regulator of adipose tissue in adult females, but recent work has indicated that estrogen's role in adipose biology may be broader than initially appreciated and has also provided important insights into the mechanism of estrogen effects on adipose tissue. Estrogen has direct effects on adipocytes to inhibit lipogenesis and may also have direct effects on other cellular constituents of adipose tissue, as well as metabolic effects on other target organs that can regulate adipose tissue. Estrogen has central effects on food consumption and energy expenditure that contribute to its overall inhibitory effects on adipose deposition. Estrogen also plays an important role in regulating adipose deposition in males and recently has been shown to be an important factor in the determination of adipocyte number, indicating that it regulates key developmental events in adipogenesis. Although critical questions still remain in our understanding of the overall role of estrogen in adipose tissue, it is clear that estrogen plays a more important role in adipose tissue than originally realized and that it is a major regulator of adipose tissue in both sexes during development and adulthood.
Reducing Arginase Activity via Dietary Manganese Deficiency Enhances Endothelium-Dependent Vasorelaxation of Rat Aorta
Authors: Ensunsa Jodi L. , Symons J. David , Lanoue Louise , Schrader Heather R. , Keen Carl L.
Published: July 2017

L-Arginine is a common substrate for the enzymes arginase and nitric oxide synthase (NOS). Acute inhibition of arginase enzyme activity improves endothelium-dependent vasorelaxation, presumably by increasing availability of substrate for NOS. Arginase is activated by manganese (Mn), and the consumption of a Mn-deficient (Mn-) diet can result in low arginase activity. We hypothesize that endothelium-dependent vasorelaxation is greater in rats fed Mn– versus Mn sufficient (Mn+) diets. Newly weaned rats fed Mn– diets (0.5 μg Mn/g; n = 12) versus Mn+ diets (45 μa Mn/g; n = 12) for 44 ± 3 days had (i) lower liver and kidney Mn and arginase activity (P < 0.05), (ii) higher plasma L-arginine (P ≤ 0.05), (iii) similar plasma and urine nitrate + nitrite, and (iv) similar staining for endothelial nitric oxide synthase in thoracic aorta. Vascular reactivity of thoracic aorta (~720 μm i.d.) and small coronary arteries (~110 μm i.d.) was evaluated using wire myographs. Acetylcholine (ACh; 10−8–10−4 M) produced greater (P ≤ 0.05) vasorelaxation in thoracic aorta from Mn– rats (e.g., maximal percent relaxation, 79 ± 7%) versus Mn+ rats (e.g., maximal percent relaxation, 54 ± 9%) at 5 of 7 evaluated doses. Tension produced by NOS inhibition using NG monomethyl-L-arginine (l-NMMA; 10−3 M) and vasorelaxation evoked by (i) arginase inhibition using difluoromethylornithine (DFMO; 10−7 M). (ii) ACh (10−8–10−4 M) in the presence of DFMO, and (iii) sodium nitroprusside (10−9–10−4 M) were unaffected by diet. No differences existed between groups concerning these responses in small coronary arteries. These findings support our hypothesis that endothelium-dependent vasorelaxation is greater in aortic segments from rats that consume Mn– versus Mn+ diets; however, responses from small coronary arteries were unaffected.
Prevention of Spontaneous and Experimentally Induced Diabetes in Mice with Zinc Sulfate-Enriched Drinking Water is Associated with Activation and Reduction of NF-κB and AP-1 in Islets, Respectively
Authors: Schott-Ohly Patricia , Lgssiar Abdelhakim , Partke Hans-Joachim , Hassan Mohamed , Friesen Nadira , Gleichmann Helga
Published: July 2017

Recently, we reported that zinc sulfate-enriched (25 mM) drinking water (Zn2+) protected male C57BL/6 mice from diabetes induced by multiple low doses of streptozotocin (MLD-STZ) and that MLD-STZ activates the transcription factors nuclear factor (NF)-κB and activator protein (AP)-1 in islets of these mice. Therefore, we studied the effect of Zn2+on spontaneous diabetes in female nonobese diabetic (NOD) mice and on the activity of NF-κB and AP-1 in islets of NOD and MLD-STZ–injected male C57BL/6 mice. We hypothesized that Zn2+may affect NF-κB, which may play a key role in immune-mediated diabetogenesis. Here we continuously administered Zn2+to NOD mice, to both parents and their F1offspring, and treated C57BL/6 male mice with MLD-STZ either alone or in addition to Zn2+. We assessed effects of Zn2+on insulitis and peri-insulitis in 8-week-old NOD mice and analyzed NF-κB and AP-1 activities in islets. Zn2+significantly prevented diabetes in female F1offspring and significantly reduced insulitis and peri-insulitis. Zn2+significantly stimulated NF-κB and AP-1 activation in NOD mice, in contrast, in C57BL/6 mice, Zn2+significantly reduced their activation by MLD-STZ. These data demonstrate that NF-κB may play a critical role in immune-mediated diabetes. Depending on the mode of β-cell destruction, Zn2+may prevent apoptosis through activation of NF-κB in NOD mice or prevent inflammatory immune destruction through inhibition of NF-κB in MLD-STZ-treated C57BL/6 mice.