bluuebunny/crossref_metadata_2025 huggingface dataset:
Cytotoxicity and efficacy evaluation of polymeric nanoparticles containing resveratrol and quercetin for use on drug eluting stents
Authors: Kleinedler Jim , Foley John , Dugas Tammy
Published: June 2021

Current generation coronary drug‐eluting stents (DES) used to treat restenosis are associated with unintended complications including potentially fatal late‐term thrombosis and hypersensitivity reactions. This provides a platform for novel engineered device therapies that exhibit increased biocompatibility and promote vascular healing after angioplasty. Resveratrol and quercetin have antimitogenic actions on vascular smooth muscle cells (VSMC) and inhibit both platelet and inflammatory cell activation. Our aim was to develop a stent coating that releases these drugs within a therapeutic range and provides an extended release profile to inhibit cascades responsible for restenosis and thrombosis. Stainless steel flats were coated with arborescent polyisobutylene‐polystyrene polymer (arbIBS) applied using the novel NanoElectrospray(TM) process. To determine DES‐mediated effects, assays were conducted on flats incubated in transwells in the presence of vascular cells. These included measures of cell viability, platelet activation, and cell proliferation. Results from this study indicate that arbIBS exhibits no cytotoxicity, and that the nanoparticles release resveratrol and quercetin at the therapeutic level, reducing platelet activation and VSMC proliferation. Ongoing experiments are aimed at introducing these DES into an animal model of angioplasty for further evaluation.

Grant Funding Source

R01‐HL082472
Nimesulide induces differentiation in crypt‐villus epithelial cell axis
Authors: Ettarh Raj , Calamai Alvise , Cullen Anthony , Schumacher Udo
Published: June 2021

The role of peroxisome proliferator‐activated receptor beta/delta (PPARβ/d) is not known but is believed to involve modulation of inflammation and mediation of differentiation in high‐turnover cell populations such as the intestinal epithelium. Cyclooxygenase (COX) inhibitors are anti‐inflammatory agents that affect proliferation in intestinal epithelial cells. This study examined the effect of COX‐inhibitor treatment on PPARβ/d expression and differentiation by immunohistochemical methods following treatment with the selective COX inhibitor nimesulide and found that PPARβ/δ expression is induced in the cytoplasm in crypt cells after 24 hours of treatment, the typical pattern of expression that was observed in untreated differentiated villous cells but not in untreated crypt cells. In addition, the differentiation marker Dolichos biflorus agglutinin (DBA) was expressed in greater parts of nimesulide‐treated crypt compared to the basal expression typically seen in untreated crypts indicating that nimesulide induces differentiation in crypt cells in the small intestine. Supported by the UCD School of Medicine/SSRA Research Scheme.
Regulation of the human mitochondrial chaperone mtHsp70 by the Hsp70 escort protein Hep
Authors: Zhai Peng , Silberg Jonathan Joff
Published: June 2021

The Hsp70 escort protein Hep is required for the solubility of yeast mitochondrial hsp70 (mtHsp70) chaperones, which play central roles in protein translocation, Fe/S‐cluster biogenesis, and the stress response. To establish whether escort proteins play a similar role in metazoans, we examined the effect of human Hep on human mtHsp70 solubility and activity. We demonstrated that human Hep is localized to mitochondria like yeast Hep and showed that mtHsp70 exhibits limited solubility unless it is coexpressed with Hep. This low solubility appears to arise from aggregation mediated by the mtHsp70 ATPase domain, as this domain is only soluble when coexpressed with Hep. In vitro studies further indicate that Hep forms a tight complex with mtHsp70 (and its isolated ATPase domain) and stimulates chaperone ATPase activity. These findings provide evidence that metazoan escort proteins regulate both the catalytic activity and solubility of their cognate chaperones by interaction with their ATPase domain. Ongoing research is examining the effects of point mutations on the ability of Hep to bind mtHsp70, promote chaperone solubility, and stimulate chaperone ATPase activity.
Identification and characterization of phostensin, a protein phosphatase 1 F‐actin cytoskeleton targeting subunit
Authors: Wang Tzu‐Fan , Kao Shu‐Chen , Wang Shiu‐Lan , Chen Chun‐Yu , Lai Ning‐Sheng , Huang Hsien‐Bin
Published: June 2021

We have conducted the search for the new protein phosphatase 1 (PP1)‐binding proteins of heart cells by using a yeast two‐hybrid system and identified one novel protein, protein phosphatase 1 F‐actin cytoskeleton targeting subunit (phostensin). Phostensin contains 165 amino acids and was encoded by KIAA1949. We demonstrated that phostensin binds to actin filament pointed ends, but not to actin monomers, sides of filaments, or barbed ends of filaments. Phostensin retarded the elongation and depolymerization rates of gelsolin‐actin seeds, directly associated with pointed ends, and reduced the rate of G‐actin addition at the pointed ends.
Formalin Fixation and Antigen Retrieval: Translating Insights To Clinical Innovation
Authors: Bogen Steven , Vani Kodela , Sompuram Seshi R
Published: June 2021

Tissue fixation and antigen retrieval (AR) are important variables in evaluating protein expression in biopsies. Because of the complexity in modeling fixation and AR with whole proteins, we investigated antibody immunoreactivity to isolated peptide epitopes. We identified epitopes using phage‐displayed peptide combinatorial libraries. The epitopes were all linear peptide sequences from the native protein. Formalin fixation abrogated immunoreactivity if another (antigenically irrelevant) protein was present. AR restored immunoreactivity. Our data suggest a model of steric interference caused by formaldehyde‐induced cross‐linking. We recently translated these findings to practical use, as a QC tool for routine clinical laboratory use. Spots of formalin‐fixed or unfixed peptides, covalently attached to glass microscope slides, can serve as immunohistochemistry (IHC) controls. The peptides can be printed to specified concentrations, are reproducible (CV <8%), and amenable to mass production. In an initial study of 109 clinical IHC laboratories, approximately 18% demonstrated inadequate staining. Errors were more commonly associated with AR than the IHC stain. This technology was recently transferred for commercial production to ThermoFisher Corp. and will offer laboratories a new method for evaluating AR and IHC staining performance from day‐to‐day as well as relative to other laboratories.
Regulation of BRCA‐1 by Xenobiotic and Dietary Ligands of the Aryl Hydrocarbon Receptor
Authors: Papoutsis Andreas J , Romagnolo Donato F
Published: June 2021

The BRCA‐1 protein is involved in repair of DNA damage and mutations of the BRCA‐1 gene confer a high risk of developing breast cancer. Interestingly, mutations of BRCA‐1 account for only 5‐10% of breast cancer cases, whereas sporadic breast tumors represent the remaining 90‐95%. In sporadic breast cancers there is silencing of BRCA‐1 in the absence of mutations. Factors contributing to this etiology include the environmental ligands of the aryl hydrocarbon receptor (AhR) 2,3,7,8 tetrachlorodibenzene(p)dioxin (TCDD) and polycyclic aryl hydrocarbons (PAHs). Dietary antagonists of the AhR with potential protective effects include the phytoalexin resveratrol and 3,3′‐diindolylmethane (DIM), the condensation product of indole‐3‐carbinol. The objective of this study was to investigate the mechanisms responsible for repression of BRCA‐1 expression by the activated AhR. Results of DNA binding studies in MCF‐7 breast cancer cells documented that TCDD‐induced the association of the AhR to xenobiotic responsive elements (XRE) harbored in the BRCA‐1 promoter region. The cotreatment with resveratrol (5, 10, 20µM) reduced AhR recruitment in a dose‐dependent fashion. We are investigating the chromatin modifications induced by the AhR at the BRCA‐1 gene and the preventative effects of resveratrol. These studies may have implications for the use of dietary AhR ligands in prevention of BRCA‐1 silencing induced by AhR agonists.

Grant Funding Source

Susan G. Komen Breast Cancer Foundation and Arizona Biomedical Research Commission
Omega‐3 fatty acids regulate gene expression levels differently in macrophages of men carrying the PPARα‐L162V polymorphism
Authors: Rudkowska Iwona , Garenc Christophe , Couture Patrick , Deshaies Yves , Vohl Marie‐Claude
Published: June 2021

Omega‐3 fatty acids (FAs) are natural ligands of the peroxisome proliferator‐activated receptor‐α (PPARα), a nuclear receptor that modulates expression levels of genes involved in lipid metabolism. The L162V polymorphism located in the ligand binding domain of the PPARα gene is associated with a deteriorated plasma lipid profile. Therefore, we postulate that subjects carrying the V162 allele exhibit differences in the activation of PPARα and its target genes after incubation with omega‐3 FAs compared with L162 homozygotes. Peripheral blood monocytes from 6 men carrying the V162 allele paired for age and for body mass index with 6 L162 homozygotes were differentiated into macrophages and incubated with eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), or mixtures of EPA: DHA. Results demonstrate that gene expression levels of PPARα and apolipoprotein AI were significantly lower for carriers of the V162 allele compared to L162 homozygotes after the addition of DHA and a mixture of EPA: DHA. Additionally, lipoprotein lipase expression displayed a tendency to be lower in carriers of the V162 allele after the addition of a mixture of EPA: DHA. Consequently, subjects bearing the PPARα V162 allele may demonstrate inferior improvements in the plasma lipid profile due to lower PPARα and its target genes expression rates in response to omega‐3 FA supplementation. Funding provided by a CIHR Operating Grant.
Dietary copper deficiency up‐regulates selected cardiac copper chaperone proteins
Authors: Getz Jean , Lin Dingbo , Medeiros Denis M
Published: June 2021

Oxidized copper can damage cell components as a pro‐oxidant and must be sequestered in order to be non‐reactive. Partly due to this, cell levels of copper are bound to proteins and are under the control of import, export, and copper chaperone proteins. Copper is absorbed by the import protein Ctr‐1 and then is ferried to the site of cupro‐enzyme synthesis by chaperone proteins. In this study we evaluated the impact that copper deficiency had upon the levels of CCS, Sco1, Cox 17, and Ctr‐1 using Western blotting. Rats were fed diets either adequate or deficient in copper from weanling for 5 weeks thereafter. Rats fed the copper deficient diets had significantly lower liver copper levels, hematocrit, and body weights, and demonstrated cardiac hypertrophy. Copper deficiency did not result in any changes in Ctr‐1 or Cox 17. However, there were significant up‐regulations in Sco1 and CCS as a result of copper deficiency. Sco1 delivers the copper to cytochrome C oxidase and CCS to the site of Cu, Zn superoxide dismutase synthesis. Cytochrome C oxidase subunits I and IV were studied with monoclonal antibodies. We report that the mitochondria encoded subunit I (Cox I) and the nuclear encoded subunit IV (Cox IV) were significantly reduced. We conclude dietary copper deficiency increases copper chaperone proteins in the heart. This could be an adaptation to deliver copper to selected enzymes when cellular copper levels are limiting.
Induction of Cellular Senescence and DNA Damage Response by Selenium Compounds
Authors: Wu Min , Lartey Fred , Schoene Norberta W , Cheng Wen‐Hsing
Published: June 2021

Although selenium chemoprevention by apoptosis has been well studied, it is not clear whether and how other tumorigenesis barriers participate in selenium chemoprevention. To address this issue, we treated MRC‐5 normal lung fibroblasts with 0‐10 µM sodium selenite, 0‐10 µM methylseleninic acid (MSeA) and 0‐500 µM methylselenocysteine (MSeC) for 48 hours, followed by a recovery of 1‐7 days. There is a selenium dose‐dependent decline in cell survival during the time course. To determine whether senescence response can be induced by sub‐lethal doses of the selenium compounds, we measured senescence‐associated expression of â‐galactosidase (SA‐â ‐gal) and bromodeoxyuridine (5‐bromo‐2‐deoxyuridine, BrdU) incorporation. We observed increased SA‐â ‐gal staining and decreased incorporation of BrdU into the replicative DNA after cellular exposure to the selenium compounds. In response to clastogens, ATM is rapidly activated, which in turn initiates a cascade of DNA damage response including histone H2AX phosphorylation at Ser‐139 (also known as ãH2AX). Results from Western analysis demonstrated increased expression of phosphorylated‐ATM and ãH2AX after the selenium treatment. Taken together, the results suggest a novel mechanism of selenium chemoprevention by which selenium induces DNA damage response and senescence to inhibit tumorigenesis.

Grant Funding Source

N/A
Estimated intakes of proanthocyanidin in the US population
Authors: Chun Ock K. , Chung Sang‐Jin , Song Won O.
Published: June 2021

Proanthocyanidins (PAs), the polymers of flavan‐3‐ols, have been documented for free radical scavenging and cardioprotective properties in experimental/ clinical studies; yet PA intake in free‐living population has not yet been estimated. We estimated the sum of all forms of PAs in the US diets and identified major food sources of PAs. We matched the recently released USDA PA database with the dietary intake data of the NHANES 1999‐2002. The database was then expanded by adding food codes of the same food which had similar processing or preparation to overcome the limitation of the food data availability. The intake was computed for monomers, dimmers, trimers, 4‐6mers, 7‐10mers and polymers (10+ mers). US adults over 19+y had a total PA intake of 95 mg/d consisted of polymers (30%), monomers (22%), dimers (16%), 4‐6mers (15%), 7‐10mers (11%), and trimers (5%) in the order. When energy intake was adjusted, PA density increased with age (p<0.001) and decreased with non‐leisure time physical activity level (p<0.001); higher in females (p<0.001) and alcohol consumers (p<0.05); and lower in non‐Hispanic Black (p<0.001) than their counterparts. The major food sources, i.e., tea, legumes, and wines, contributed 45 mg (48%) of daily PA intake. The sociodemographic differences in PA intake as in other antioxidant intake pose important question in relation to chronic disease prevalence in the US.
Potential roles of β‐apo‐carotenoids on activation of retinoic acid receptors
Authors: Marsh Rebekah S , Yan Yan , Reed Vanessa M , Hruzkewycz Damian , Curley Robert W , Harrison Earl H
Published: June 2021

β‐Carotene oxygenase 2 catalyzes the cleavage of β‐carotene at non‐central double bonds of the polyene chain, yielding apo‐carotenals. The biological roles and the potential regulation of retinoic acid receptors (RARs) by apo‐carotenoids are not clear. The hypothesis we seek to test is that apo‐carotenoids are able to stimulate transcription by activation of RARs. We will test the effects of all of the long and short chain apo‐carotenals arising from eccentric cleavage of β‐carotene (and the corresponding apo‐carotenoic acids) on the activation of RARα or RARβ transfected into monkey kidney cells along with RARE‐ luciferase reporter constructs. So far, we have obtained or synthesized apo‐8′‐carotenoic acid, apo‐14′‐carotenoic acid, β‐cyclocitral, β‐cyclogernaic acid, β‐ionylideneacetaldehyde, β‐ionylideneacetic acid, β‐ionone, and a C13 ketone. None of these apo‐carotenoids showed significant transactivation activity for RARs when compared to all‐trans retinoic acid (RA). The results suggest that the biological effects of these apo‐carotenoids are through mechanisms other than activation of RARα and β. Supported by NIH grants DK044498 and HL049879.
Parameters of vitamin A (VA) status in children with sickle cell disease (SCD)
Authors: Park Hyunjin , Schall Joan , Zemel Babette , Kawchak Deborah , Ohene‐Frempong Kwaku , Green Michael H. , Stallings Virginia
Published: June 2021

VA status is compromised and there is chronic inflammation in SCD. We examined effects of VA (300‐600 µg/day; n=22) or VA + zinc (AZ; 300‐600 µg VA + 10 mg Zn/d; n=16) vs placebo (PL; n=21) on serum VA and its transporters in SCD children (7.7 ± 2.8 y). Serum samples were analyzed for retinol concentrations (ROH) at baseline and after 3, 6 or 12 mo of supplementation and for retinol‐binding protein (RBP) and transthyretin (TTR) at baseline and 3 or 12 mo. ROH was low at baseline (0.64 ± 0.14 µmol/L, n=59) compared to the cut‐off for VA deficiency (< 0.70 µmol/L). In children with low baseline ROH (< 0.70 µmol/L), ROH was significantly higher in AZ than in PL but VA alone did not significantly increase ROH (0.626 ± 0.117 [AZ], 0.606 ± 0.125 [VA], 0.574 ± 0.143 µmol/L [PL]). Serum ROH was significantly higher at 3 (5 % increase) and 12 mo (6.5 %) vs baseline in children with low baseline ROH. There InInThwere no differences among groups in the molar ratio of ROH:RBP, which was low in all subjects (0.422 ± 0.102). Higher molar ratios of ROH:RBP were associated with lower molar ratios of RBP:TTR (p=0.000). Our findings indicate that, in spite of VA or VA + zinc supplementation, ROH and ROH:RBP were not normalized in SCD children. Funding provided by: Comprehensive Sickle Cell Center (NIH: 5 U54 HL070596), CTRC (UL1‐RR‐024134), and the Nutrition Center of the Children's Hospital of Philadelphia.
Evaluation of Laser Imaging for Assessment of Circumferences
Authors: Freeland‐Graves Jeanne , Pepper Reese , Xu Bugao , Yu Wurong
Published: June 2021

The most common technique for evaluation of waist and hip circumferences, tape measure, is limited by low inter‐rater reliability and may be uncomfortable for the participant. This research compared circumferences measured by a small‐scale, laser imaging device to those collected by inelastic tape measure in order to assess validity. Seventy women (18‐64 yrs) were tested after fasting 4 hours on one occasion. Subjects had a mean Body Mass Index of 25.6±0.6 kg/m2, ranging from 18.9 to 36.8 kg/m2. Measures from laser body imaging were significantly related to tape circumference for both waist and hip girths (r=0.95 and 0.91, respectively, p<0.01). Additionally, estimation of body girth at the waist did not differ between laser and tape methods (103.1±1.2 cm and 103.6±1.2cm, respectively, p>0.05). Comparison of hip circumference measurements showed similar results (90.0±1.5 cm and 88.5±1.9 cm, p>0.05). In a subset of 32 participants, the reproducibility of repeated laser scans was tested ten times, with the body relaxed between scans. The intraclass correlation coefficient was 0.99 for both waist and hip circumferences (p<0.01). These results suggest that laser body imaging is a reproducible and valid measure of waist and hip circumferences. Supported by NIH 1 R21 DK081206‐01.
Use of the World Health Organization (WHO) growth standards to evaluate prevalence and predictors of overweight in preschool children enrolled in the WIC program
Authors: Maalouf Zeina , Metallinos‐Katsaras Elizabeth , Dewey Kathryn G
Published: June 2021

With the current childhood obesity epidemic, tools for accurate diagnosis of childhood obesity are needed. The World Health Organization (WHO) issued new growth standards for children under five that differ considerably from the 2000 Centers for Disease Control and Prevention (CDC) growth charts. We used records of 143,792 children aged 2‐5 years who participated in the Massachusetts Special Supplemental Nutrition Program for Women, Infants and Children from 2001‐06 to compare the prevalence and predictors of childhood overweight using the WHO vs. the CDC cut‐offs. The prevalence of overweight (>95th percentile) was 17.0% using the CDC cut‐offs vs. 23.6% using the WHO cut‐offs; the difference was largest in children 24‐27 mo of age (12.7% vs. 25.4%, respectively). In a logistic regression model controlling for birth weight and child age, factors significantly related to child overweight, assessed using the WHO cut‐offs, were maternal pre‐pregnancy weight (obese vs. normal OR=1.81 (1.72‐1.91)), Hispanic ethnicity (OR=1.50 (1.43‐1.56, ref=non‐Hispanic), maternal education (some college vs. less than high school graduate (HS), OR=0.80, 0.75‐0.85, HS vs less than HS OR=0.90, 0.86‐0.94) and child sex (OR=0.89, 0.85‐0.93, ref=male). Using the WHO charts will result in a higher estimated prevalence of childhood overweight and allow for earlier diagnosis, thus making earlier intervention possible.

Supported by a grant from the United States Department of Agriculture Economic Research Service Research Innovation and Development Grants in Economics

Grant Funding Source

United States Department of Agriculture Economic Research Service Research Innovation and Development Grants in Economics
Diet quality, anthropometric status and self‐reported chronic diseases among Mexican households
Authors: Xochitl Ponce Martínez , Lucia Hernánez Barrera , Simón Barquera Cervera
Published: June 2021

Objective

To evaluate association between diet quality and cardiovascular diseases among Mexican households.

Methods

A secondary analysis from the MxFLS was conducted in 8840 Mexican families. Dietary quality was evaluated considering adherence to WHO dietary recommendations. Households were classified in 4 groups according to their anthropometric status. Self‐reported health information was used to identify nutrition‐related chronic diseases. The association between type of HH and NRCD was analyzed using logistic regression models.

Results

Underweight HH reported a significantly higher consumption of fiber. Overweight HH reported significantly higher intakes of total and saturated fat and cholesterol. HH who reported a NRCD showed the lowest adherence to the recommendation of fruit and vegetables intake. Overweight HH were 2.8 times more likely to report DM and 2.27 likely to report hypertension. Double burden families reported 2.27 times more probability of presenting hypertension.

Conclusions

Lack of adherence to WHO recommendations with regards to saturated fatty acids and cholesterol, as well as fruits and vegetables, could be considered risk factors for development of chronic disease in overweight and double burden HH in Mexico.
Food insecurity is associated with iron deficiency anemia in U.S. adolescents
Authors: Eicher‐Miller Heather A , Mason April C , Weaver Connie M , McCabe George P , Boushey Carol J
Published: June 2021

Food insecurity is a problem faced by many Americans. This condition is associated with decreased intake of nutrients and poor health. Food insecure children are especially vulnerable. Inadequate iron in the diets of food insecure children could eventually lead to iron deficiency and iron deficiency anemia. The purpose of this study was to formally investigate the current relationship of iron status and food insecurity in children. Child participants ages 1 to 19 years (n=12,329) of the recently released NHANES 1999‐2004 were classified as food secure or food insecure using the U.S. Children's Food Security Scale and the U.S. Household Food Security Module. Two or more abnormal values for transferrin saturation, serum ferritin, and erythrocyte protoporphyrin were used to classify iron deficiency with the addition of low hemoglobin to classify iron deficiency anemia. Results indicated that the odds of iron deficiency anemia were 2.61 times more likely (P=0.029) among food insecure children 12 to 15 years compared with food secure children. The results of this study indicate a continuing need for successful interventions to improve iron deficiency and iron deficiency anemia among food insecure children.

Source of Financial Support: Purdue University Extension
Branched‐chain α‐ketoacid dehydrogenase complex (BCKDC): purification from rat liver and inhibition by thiamin pyrophosphate (TPP)
Authors: Shimomura Yoshiharu , Fujimura Yukihiro , Akita Keiichi , Bajotto Gustavo
Published: June 2021

BCKDC is a rate‐limiting enzyme in the branched‐chain amino acid catabolic pathway. This enzyme complex is regulated by a phosphorylation/dephosphorylation cycle. The specific kinase (branched‐ chain α‐ketoacid dehydrogenase kinase (BDK)) is responsible for inactivation of the complex by phosphorylation. We have developed the method for BCKDC purification from rat liver using hydrophobic interaction column chromatography (Shimomura et al., Arch. Biochem. Biophy., 283, 293‐299 (1990)). We here report the modification of the purification method to obtain the highly purified BCKDC associated with BDK. Using the purified enzyme, we analyzed the inhibition of BDK by TPP at two potassium ion (K+) concentrations. IC50 values of 4.6 and 8.0 µM and inhibition constant values of 3.2 and 16.4 µM were obtained in the presence of 20 and 100 mM K+, respectively. These results suggest that BDK is less sensitive to TPP inhibition under physiological TPP and K+ concentrations.
Accumulation of chromium and histological findings on Hybro chickens liver treated with Cr(VI) and vitamin C
Authors: Gashi Agim , Halili Fetah , Bajraktari Ismet , Sopa Ramadan , Bislimi Kemajl , Fetahu Shukri , Behluli Avni , Grapci‐Kotori Linda
Published: June 2021

In this study the role of vitamine C in the acumulation of chromium in the liver and liver lesions of Hybro chicken treated with Cr (VI) were investigated. The chickens were divided into 5 groups (I‐V), with 10 individuals which and were treated orally with different concentrations of chrome (VI) and 10 mg/kg b.w vitamin C for 180 days. The preparation of tissues for the pathological microscopy was done according to the standard procedure whereas the staining was done with hemathoxylin‐eosin. The Cr concentration on the liver was determined by atomic absorbtion spectroscopy (AAS) via UNICAM 929‐AA type spectrometer. The outcome of the research indicates that the six‐month ingestion of two different doses of Cr (VI) results in light histopathological changes in the periportal and pericentrilobular zone of the liver. The fatty vacuolization of hepathocites, dilatation of centrilobular vein, the dilated sinusoids with erythrocytes, proliferation of biliary tubules, the lymphoplasmocitary infiltrates etc. are visible. The protective role of Vitamin C on the applied dose was not noticed. Vitamin C combined with Cr (VI) doses amplifies the bioaccumulation of Cr on the liver to a considerable degree, compared not only with control group of chicken but as well as with the chicken treated only with Cr (VI) . The investigation results show that the bioaccumulation of chrome and the histological fidings of the liver depend on the dose, time of action and the combination of Cr with Vitamin C.
Use of the extract oil of the plant Eupatorium aschenbornianum for wound healing
Authors: Castillo‐Henkel Carlos , Castillo‐Hernandez Maria del Carmen , Sanchez‐Mendoza Maria Elena , Rios‐Guerra Hulme , Arrieta Jesus , Guevara‐Balcazar Gustavo
Published: June 2021

The effect of an oil extract of the plant Eupatorium aschenbornianum was evaluated in Wistar rats by using (treated group) or not (control group) a new model of wound healing in rats in which the participation of the muscle panniculos carnosus is avoided by removing it. In brief, we made a circular incision of 1.5 cm , remove the panniculos carnosus and stich the skin with the latisimus dorsis. A imagen‐j method was employed to evaluate the evolution of the healing. The results show that the time needed for complete healing was 28‐35 days in the control group whereas in the group treated with 2 and 5% extracts of the plant the time needed was 15‐20 days (about 30‐40% less). We conclude that the extrac is useful in shortening the period of wound healing.
Characterization of a novel transcript deleted in breast and prostate cancer
Authors: DeMoor Stephanie Ann , Hall Devon C. , Matoba Fumika , Reveles Xavier T. , Leach Robin J. , Naylor Susan L. , Johnson‐Pais Teresa L.
Published: June 2021

Chromosomal regions showing homozygous deletion in tumors are not commonly found and are usually the site of genes involved in tumor suppression. We previously identified the homozygous deletion of a discrete region at 18q22.3 in 50% of breast tumors and 58% of prostate tumors. There are no known genes in this region. Our hypothesis for this study was the following: Encoded within this 18q region is a transcript that when lost plays a role in the development of breast and prostate cancer. The objective of this study was to determine whether sequences within the region of deletion are transcribed in primary breast and prostate epithelial cells. A microarray experiment using RNA isolated from primary mammary epithelial cells and primary prostate epithelial cells enabled us to detect an approximately 500 basepair region that is transcribed in normal breast and prostate epithelial cells. This is a novel transcribed sequence that has not been described previously. A quantitative real‐time TaqMan assay verified the existence of the transcript in breast and prostate epithelial cell RNA. The novel transcript was also detected in high abundance in RNA isolated from brain, cervix, spleen, and thymus. Through our experiments, we hope to identify a new molecular pathway in breast and prostate cancer that could provide new targets for developing therapeutics.
CSR1 Induces Cell Death through Inactivation of CPSF3
Authors: Luo Jianhua , Zhu Zehua , Yu Yan P
Published: June 2021

CSR1 (cellular stress response 1), a newly characterized tumor suppressor gene, undergoes hypermethylation in over 30% of prostate cancers. Re‐expression of CSR1 inhibits cell growth and induces cell death, but the mechanism by which CSR1 suppresses tumor growth is not clear. In this study, we screened a prostate cDNA library using a yeast two‐hybrid system and found that the cleavage and poly‐adenylation specific factor 3 (CPSF3), an essential component for converting heteronuclear RNA to mRNA, binds with high affinity to the CSR1 C‐terminus. Further analyses determined that the binding motifs for CPSF3 are located between amino acids 440‐543. The interaction between CSR1 and CPSF3 induced CPSF3 translocation from the nucleus to the cytoplasm, resulting in inhibition of polyadenylation both in vitro and in vivo. Down‐regulation of CPSF3 using small interfering RNA (siRNA) induced cell death in a manner similar to CSR1 expression. A CSR1 mutant unable to bind to CPSF3 did not alter CPSF3 subcellular distribution, did not inhibit its polyadenylation activity, and did not induce cell death. In summary, CSR1 appears to induce cell death through a novel mechanism by hijacking a critical RNA processing enzyme.
SYNTHESIS OF STEROID 6‐CARBOXYLATE ESTERS: ANALOGS OF METHYL 11β , 17 α , 21‐TRIHYDROXY‐3, 20‐DIOXO‐PREGNA‐1, 4‐DIENE‐6‐CARBOXYLATE
Authors: Gassett Sandi Rena , Lee Henry J.
Published: June 2021

Systemic side effects of anti‐inflammatory steroids may be minimized by incorporation of a labile group which is metabolized to make inactive steroid acids upon entry into the systemic circulation (antedrug concept). Thus, antedrugs act locally and are devoid of systemic toxicities. Flonase® was the first clinically available steroidal thiocarbonate inhalation antedrug marketed by Glaxo SmithKline. In the continuing efforts to increase local to systemic activity ratios of potent steroidal anti‐inflammatory antedrugs, a series of anti‐inflammatory steroids with a carboxylate ester at the carbon 6 position and derivatives were synthesized. These derivatives were evaluated for anti‐inflammatory activity and the effects to bind to glucocorticoid receptors, and to inhibit nitric oxide (NO) generation in vitro. The new compounds are designed to rapidly metabolized to inactive compounds upon entry into the systemic circulation. The objective of our research is to develop a highly potent, and yet safer anti‐inflammatory steroid. These results provide a solid rational basis for the development of therapeutically useful non‐systemic corticosteroids for prolonged local use.
Expression of phosphorylated CREB in the fasted rat hippocampus
Authors: Estrada Nelly M. , Isokawa Masako
Published: June 2021

Fasting facilitates synaptic plasticity and promotes learning consolidation. However, cellular signaling mechanisms underlying the effect of fasting on memory consolidation are not completely understood. We investigated the effect of fasting on the hippocampal neuron by testing the hypothesis of whether fasting promoted phosphorylation of CREB (c‐AMP response element binding protein), an important nuclear protein to consolidate memories by inducing gene expression. We used in vivo rat model of fasting and applied anti‐phospho‐CREB (Ser133) raised in the rabbit to cardially‐perfused rat brain sections (40 μm thick). The cell layer of CA1 hippocampal subfield in fasted animals exhibited strong expression of phosphorylated CREB (pCREB). As reported previously, the hypothalamus was also strongly immunoreactive and provided a positive control in the present study. Our finding suggests fasting may stimulate cAMP‐dependent signaling cascades in the hippocampal CA1 neuron. A molecule involved in and responsible for the activation of this signaling process is currently under the investigation. Supported by NIH grants R15DA021683 and SC1GM081179 (to MI). NME is an American Physiological Society Undergraduate Summer Research Fellow in 2008.
Losartan affects Tyrosine Hydroxylase Protein Expression in the A2 Area and Hypoglossal Nucleus of Normal and Dystrophic Hamsters
Authors: Ambur Lindsay Christen , Schlenker Evelyn Heymann
Published: June 2021

Angiotensin AT1 receptors affect production of tyrosine hydroxylase (TH), the rate‐limiting enzyme for production of catecholamines. AT1 receptors are colocalized with TH in the A2 region of the nucleus tractus solitaries and hypoglossal nucleus. We hypothesized that blocking AT1 receptors with losartan (L), an AT1 receptor antagonist, would decrease TH expression in the A2 and hypoglossal nucleus in normal (N) and dystrophic (D) hamsters. Two month old male N and D hamsters received tap water or L in drinking water at a dose of 50 mg/kg/day for 2.5 months. At 4.5 months, animals were sacrificed, brains perfused, cryoprotected, and frozen at ‐70° C. Brains were sliced at 35‐40 micrometers. Immunohistochemistry was performed using a mouse monoclonal TH primary antibody (1:10,000) visualized with DAB. ImageJ was used to evaluate A2 and hypoglossal areas and number of TH positive neurons. Losartan had no effect on hypoglossal areas, but decreased the number and density of TH positive neurons and in D hamsters, but not in N hamsters. In N and D hamsters TH positive neurons were predominately located in ventral regions of the hypoglossal nuclei. Losartan treatment eliminated this localization in both groups. In the A2 region no significant differences were observed in areas or number of positive TH neurons, but in D hamsters L decreased TH positive neuron density.

Funded by: APS Undergraduate Summer Fellowship & P20 RR‐015567
Heat acclimation improves mitochondrial function following ischemia reperfusion insult
Authors: Assayag Miri , Saada Ann , Horowitz Michal
Published: June 2021

Heat acclimation (AC) is an evolutionary conserved feature allowing organisms to adjust to prolonged/persistent changes in ambient temperature. An inseparable outcome of acclimation is the protection from novel stressors e.g. ischemia reperfusion (I/R) insult. In this study we evaluated the intrinsic apoptotic pathway and mitochondrial metabolic state of AC versus non‐acclimated rats (C) hearts. We here report that AC attenuates the death signaling pathway invoked in response to acute heat stress or I/R via increased BclXl/Bad ratio, thereby abolishing cytochrome c trafficking to the cytosol. This mechanism led to an almost 3 fold reduction in apoptosis level in the AC heart. The beneficial effect of AC was also reflected in mitochondrial metabolic performance. In contrast to C hearts that demonstrated a drop in ATP production post I/R, AC hearts maintained intact mitochondrial membrane potential, respiratory chain enzyme‐complex activities and unimpaired ATP production. Concomitantly we observed upregulation of the mitochondria biogenesis gene marker PGC‐1α transcript, indicative of the beneficial AC effect on mitochondrial performance. We conclude that AC enhances mitochondrial metabolic function and cytoprotection.
Dissociation of Hsp90 interaction with eNOS impaired resistance to myocardial ischemia in BN/Mcw rat hearts
Authors: An Jianzhong , Du Jianhai , Wei Na , Xu Hao , Pritchard kirkwood A , Shi Yang
Published: June 2021

Reduced bioavailability of nitric oxide ( •NO) appears to increase myocardial ischemia reperfusion injury. We previously reported that hearts from BN/Mcw rats are more resistant to global ischemia than hearts from SS/Mcw rats by a •NO dependent mechanism that was promoted by the association of hsp90 with eNOS. Recently, we developed a novel cell permeable decoy peptide TSB2 that specifically disrupts hsp90 association with eNOS. Here, we determine if TSB2 decreases resistance to myocardial ischemia in BN/Mcw rat hearts. Isolated BN/Mcw and SS/Mcw rat hearts were perfused with TSB2 (50 μg/ml) for 60 min prior to 35 min ischemia and 120 min reperfusion. Recovery of left ventricular developed pressure (RLVDP) and infarct size (IS) were measured. Immunoprecipitation and western blot analysis of heart homogenates for eNOS and hsp90 were performed. RLVDP in BN/Mcw rats (59.6±4.1%) was significantly higher than in SS/Mcw rats (32.1±4.5%). IS (%LV) in BN/Mcw rats was ~63% less than in SS/Mcw rats ((27.6±3.1% vs. 44±3.5%). TSB2 decreased RLVDP and increased IS in BN/Mcw hearts (46.4±4.7%, 36.5±4%) but not in SS/Mcw hearts (34.5±5.9%, 44.3±3.8%). Immunoblots showed that TSB2 decreased eNOS hsp90 association by 32% in BN/Mcw hearts. Taken together, TSB2 decreased resistance to myocardial ischemia by inhibiting hsp90 interactions with eNOS in isolated BN/Mcw rat hearts.
Left ventricular function in hemorrhagic shock and after Hextend® resuscitation in anesthetized hamster
Authors: Chatpun Surapong , Cabrales Pedro , Tsai Amy , Intaglietta Marcos
Published: June 2021

Left ventricular function during moderate hemorrhagic shock and after resuscitation was studied using a miniature pressure‐volume conductance catheter in an anesthetized hamster model. Moderate hemorrhagic shock was induced by 40% blood volume withdrawal in 15 minutes. A single volume of Hextend®, a comparatively higher viscosity plasma expander, was infused after 30 min of hemorrhagic shock. Cardiac function‐related parameters were measured and derived. Volume filling rate, a load dependent parameter, and a load independent parameter negative pressure‐time derivative (dP/dtmin) were used to assess the diastolic function. Systolic function was assessed by the peak positive pressure‐time derivative (dP/dtmax) and maximum ventricular power divided by square of end‐diastolic volume. Preliminary results showed that volume filling rate was restored to 95 ± 16% of baseline 30 min after resuscitation, this being 15 ‐ 20% greater than during shock (p<0.05). dP/dtmin recovered from 53 ± 15% of baseline during shock to 67 ± 20% of baseline 30 min after resuscitation. dP/dtmax improved from 75 ± 16% of baseline during shock to 94 ± 24% of baseline after resuscitation. It is concluded that Hextend® improved both systolic and diastolic function after moderate hemorrhagic shock in the early stage of resuscitation.
Regulation of calcium transport by the mammalian retinal pigment epithelium
Authors: Echtenkamp Stephen F. , Kennedy Brian G. , Mangini Nancy J.
Published: June 2021

The retinal pigment epithelium (RPE) is a transporting epithelial monolayer that regulates the composition of the subretinal space bathing the photoreceptor outer segments. Calcium concentration in the subretinal space varies with illumination of the retina. In this study we characterized the transepithelial calcium transport by the mammalian RPE. By immunocytochemistry we localized plasma membrane calcium transport proteins PMCA, TRPV5 and NCX to both apical and basolateral RPE membranes in bovine or human native tissues. Calcium transport activity was assessed as transepithelial 45‐Ca flux in voltage‐clamped bovine RPE/choroid Ussing preparations. Apical‐to‐basal and basal‐to‐apical Ca fluxes were measured in both short and open circuit conditions. We found that bovine RPE/choroid catalyzes a net, active calcium transport in the basal‐to‐apical (choroid‐to‐retina) direction. In normal open circuit condition, the positive apical transepithelial potential drives a paracellular calcium flux that roughly balances this active transport. Application of low sodium Ringer solution to the apical surface decreased net basal‐to‐apical flux by almost 90%, suggesting that NCX activity contributes to the active transepithelial calcium transport in bovine RPE.

Supported by American Health Assistance Foundation, Macular Degeneration Research
The cytoskeleton reduces the diffusional dimensionality of CD36 and promotes its aggregation and signaling
Authors: Jaqaman Khuloud , Kuwata Hirotaka , Touret Nicolas , Trimble William , Danuser Gaudenz , Grinstein Sergio
Published: June 2021

We used single‐molecule imaging and single‐particle tracking to investigate the dynamic behavior of CD36, a receptor that mediates oxidized LDL (oxLDL) uptake by macrophages. Free (unliganded) receptors existed as metastable multimers that split and reformed spontaneously. A subpopulation of receptors ( ≈30%) moved in linear patterns radiating from the nucleus. Unexpectedly, the linear motion was not motor‐driven but reflected instead diffusion within a confined space delimited by trough‐like cytoskeletal structures. These were attributed to microtubule‐induced discontinuities in the cortical actin meshwork. The resulting reduction in diffusional dimensionality increased the probability for receptors to collide and aggregate into multimers. CD36 is activated and undergoes endocytosis when clustered by multivalent ligands like oxLDL. Disruption of the cytoskeletal organization reduced multimer formation and inhibited CD36‐mediated signaling and oxLDL internalization. These observations demonstrate that the cytoskeleton can control signal transduction by dictating the confinement of receptors within regions where their collision frequency is increased.
Regulation of stromelysin‐2 in inflammation
Authors: Chen Ann J , Birkland Timothy P. , Chen Peter , Altemeier William A. , Parks William C. , McGuire John K.
Published: June 2021

Introduction

Specific members of the matrix metalloproteinases (MMPs) family are key mediators of inflammation. Our preliminary data suggest stromelysin‐2 (MMP‐10) also functions to moderate inflammation. Stromelysin‐2 is not found in healthy tissue but is highly induced in injured epithelium. However pathways controlling stromelysin‐2 up‐regulation in inflammatory processes are not known.

Data

We found high levels of stromelysin‐2 gene expression in various models of lung injury, including mice challenged with Pseudomonas aeruginosa, influenza, and bleomcyin. We found mice lacking stromelysin‐2 had increased lung injury and higher mortality in response to P. aeruginosa. Bioinformatic analysis of the five kilobase region upstream of the gene identified several binding regions essential to stromelysin‐2 gene regulation.

Conclusion

We conclude that increased expression of stromelysin‐2 is a generalized protective host response in inflammation. We report several transcription factor binding sites responsible for the regulation of stromelysin‐2 in inflammation.

This research is supported by grants from the American Lung Association and the National Institute of Health (HL029594, HL077555).
A re‐examination of the striated organelle in vestibular endorgans
Authors: Vranceanu Florin , Lysakowski Anna
Published: June 2021

The striated organelle (STO) is a structure located in the subcuticular region of hair cells, consisting of alternating thick and thin bands. Although present in all cochlear and vestibular hair cells, the STO is particularly well‐developed in type I hair cells, where it is shaped like an inverted open ended cone that contacts the cell membrane along its entire circumference and is separated from the cuticular plate by a layer of mitochondria. In other hair cells, it is a much smaller structure and appears to be free‐floating. We studied its structure in EM tomography of the type I hair cell. In three‐dimensional reconstructions, we found that it is connected to at least some actin rootlets. It may also be associated with microtubules, mitochondria and smooth ER. Confocal immunohistochemistry places Yotiao (an AKAP protein) in the same area as the STO and the actin‐binding protein, alpha fodrin (non‐erythroid spectrin), where the STO contacts the cell membrane. The contact with the rootlets suggests that the STO might regulate hair‐bundle stiffness. Its association with the cell membrane suggests that the STO may help in the formation of the constricted neck characteristic of type I hair cells.

Supported by NIH DC‐02521 and the 2008 Tallu Rosen Grant in Auditory Science (from the National Organization for Hearing Research Foundation)

Grant Funding Source

NIH DC‐02521; 2008 Tallu Rosen Grant in Auditory Science (NOHR)
Labeling of inner ear afferents in the transgenic thy1‐YFP‐H mouse
Authors: Chidavaenzi Robstein Lovejoy , Lysakowski Anna
Published: June 2021

We wished to assess the level of inner ear endorgan labeling in a transgenic mouse expressing yellow fluorescent protein (YFP) driven by the thy1 promoter and to examine the possibility of using this thy1‐YFP‐H mouse as a model for studying inner ear afferents. End‐organ YFP fluorescence in a postnatal day 8 (P8) thy1‐YFP‐H mouse was observed using confocal microscopy after anesthetizing the animal, then transcardially perfusing it with PBS and 4% paraformaldehyde. Images from utricular and cochlear whole mounts and crista sections suggest that in the thy1‐YFP‐H P8 mouse, vestibular afferents, including both calyces and boutons and type I and II spiral ganglion afferents are labeled. Afferents in the striolar region were more intensely labeled than extrastriolar afferents, and no supporting cell labeling was observed. Cochlear afferents, including those innervating both inner and outer hair cells, were also labeled. In conclusion, the thy1‐YFP‐H mouse could be a useful model in which to study inner ear endorgan structure and function as it lends itself to in‐vivo imaging and enables easy identification and distinction of afferent types and regions.

Supported by NIH DC‐02521.

Grant Funding Source

NIH DC‐02521
In vivo Induction of Poly(ADP‐ribose)polymerase‐1 following Ultraviolet Radiation
Authors: Hayden Brenee S , Cooper Karen , Hudson Laurie G.
Published: June 2021

Skin is a target tissue for arsenic carcinogenesis. Low arsenic concentrations enhance DNA damage and skin tumors in mice following ultraviolet radiation (UVR) exposure, but, the underlying mechanisms are unclear. Inhibition of DNA repair enzymes such as Poly(ADP‐ribose)polymerase (PARP‐1) by arsenic are under investigation, yet little is known about PARP‐1 activation by UVR in the skin. Our aim is to characterize the in vitro and in vivo kinetics and dose dependence of PARP‐1 activation following UVR exposure. Skh‐1 mice (hairless) were exposed to solar simulated UV irradiation (7 kJ/m2 to 35 kJ/m2). Skin samples were collected from unexposed mice and post exposure, then analyzed for cyclobutane pyrimidine dimers (CPDs) and 8‐hydroxy‐2′‐deoxyguanosine (8‐OHdG), products of DNA damage, and PARP‐1 activity using immunohistochemistry.CPD formation and PARP‐1 activation were evident at 14 kJ/m2. At this UVR dose, CPDs were detected as early as 10 minutes and persisted up to 2 hours post exposure. PARP‐1 activation was maximal at 1 hour and returned to control levels by 4 hours. These experiments reveal conditions for further studies which will focus on the interactions of arsenic and UVR in vivo. This work was supported by NIH award R01 ES015826.