bluuebunny/crossref_metadata_2025 huggingface dataset:
Platelet bioenergetics and its implications in disease
Authors: Prakhya Kanakanagavalli Shravani , Wang Qingjun , Whiteheart Sidney W
Published: June 2021

Bioenergetics drives energy production for mechanical processes that are critical to platelet functions, e.g., adhesion, spreading, aggregation, clot formation and contraction. Storage and pathological conditions e.g., stroke, sepsis, diabetes etc., alters platelet energetics and function; however, the specific mechanisms governing these alterations are not clear. It is not well understood which metabolic fuels can be used and which are required. We are investigating the roles of different metabolic pathways in the energy consuming process of clot retraction. Using a spectrum of techniques from in vitro platelet functional assays to biochemical methods, whole animal models, and measurement of bioenergetic functions of mitochondria we are elucidating platelet bioenergetics and its implications in disease. We developed a simple method for measuring clot contraction kinetics using time lapse imaging. Our system is efficient, high throughput and is favorable to study platelet bioenergetics of clot formation and retraction. We hypothesize that, upon activation with thrombin, fuel use can be highly plastic, depending on the fuels available. We found that glycolysis and oxidative phosphorylation can both contribute to provide energy for platelet function. Inhibitors of glycolysis and oxidative phosphorylation using 2‐Deoxyglucose and oligomycin inhibited clot retraction showing that both these processes are important.

We have also begun to examine bioenergetics in platelets from diabetics. Platelets from type2 diabetes patients have been shown to have abnormal mitochondrial membrane potential, membrane fluidity, ATP content, and oxidative stress. However, the impact of diabetes on platelet bioenergetics is less clear. We have developed a mouse model to probe the role of mitochondria and oxidative phosphorylation in platelet function. TFAM (Mitochondrial Transcription factor A) regulates mitochondria DNA copy number and transcription of mitochondrial DNA encoded proteins. The regulation of mitochondrial DNA is necessary for mitochondrial biogenesis. We have selectively deleted TFAM in the megakaryocyte lineage using the Platelet factor 4 promoter driven Cre‐Lox system. This model will be very useful, and it supports our in vitro studies with metabolic inhibitors in understanding the role of mitochondria and oxidative phosphorylation in platelet function in platelet bioenergetics contributing to the thrombosis

Support or Funding Information

Supported by HL56652, HL138179 and VA Merit Award

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Akt1 and Akt2 Isoforms Play Distinct Roles in the Development of Inflammation and Fibrosis during Alcoholic Liver Disease
Authors: Gordillo Karina Reyes , Shah Ruchi , Tuma Pamela L , Robledo Jaime Arellanes , Cheng Ying , Lakshman Raj M
Published: June 2021

Akt isoforms (Akt1, Akt2, and Akt3), the key phosphorylating kinases, play vital and unique roles in all aspects of alcoholic liver disease (ALD). In vitro, cell specific silencing of Akt isoforms in acetaldehyde+LPS and/or ethanol+LPS treated human hepatic stellate cells (HSC), Kupffer cells and/or VL17A hepatocyte cultures; and in vivo, Akt1 and Akt2 silencing in a chronic ethanol/binge/lipopolysaccharide (EBL) induced mouse liver injury model using pharmacological inhibitors, were used to gain a better understanding of how each Aktisoform differentially regulates cellular functions during ALD. This study involves hepatocyte and Kupffer cells inflammation, and hepatic stellate cells proliferation, migration and fibrogenesis, which cumulatively lead to the pathogenesis of ALD. Herein, our in vitro results showed that siRNA‐directed silencing of Akt2, but not Akt1, significantly suppressed cell inflammatory markers in HSC and Kupffer cells. While Akt1 and Akt2 inhibited cell proliferation in HSC; Akt2 alone inhibited cell migration in HSC; and Akt1 and Akt2, but not Akt3 inhibited the fibrogenic markers in hepatocytes and HSC. Moreover, although Akt isoforms show extensive sequence similarity, they are localized in different subcellular compartments in HSC. On the other hand, our in vivo results showed that EBL treatment in wild‐type C57BL/6 mice stimulated all three isoforms of Akt with concomitant increases in phosphorylated species of phosphoinositide dependent kinase‐1, mammalian target‐o‐rapamycin complex 2, and phosphatidylinositol‐3‐kinase, resulting in increases in inflammatory, proliferative, and fibrogenic genes. In addition, pharmacological blocking of Akt2, but not Akt1, inhibited EBL‐induced inflammation; and of Akt1 and Akt2 inhibited the pro‐fibrogenic markers as well as fibrosis. Our finding suggest that Akt isoforms play unique roles against inflammation, cell proliferation, migration, and fibrogenesis during EBL‐induced liver injury. These in vitro and in vivo results also suggest that close attention must be paid when targeting all Akt isoforms as a therapeutic intervention.

Support or Funding Information

NIAAA/NIH K01 award ‐ number: K01AA025140‐02

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
In Vivo Knockdown of the IP3R1 Calcium Channel Disrupts ER and Actin Morphology, as well as TBC Bulb Maturation in the Apical Processes of Sertoli Cells in Rat Testes
Authors: Adams Arlo , Vogl Wayne
Published: June 2021

Spermatogenesis is the process by which sperm cells are produced, but at one phase of this process, late stage spermatids are temporarily attached to the apex of Sertoli cells in the seminiferous epithelium. Late spermatids are anchored by large adhesion like junction complexes termed ‘ectoplasmic specializations’ (ESs) that are proposed to be disassembled by novel endocytic structures only found in mammalian Sertoli cells, called ‘tubulobulbar complexes’ (TBCs). Both ESs and TBCs are heavily actin related, with parallel actin arrays and dendritic actin networks respectively being hallmarks of their identities. The endoplasmic reticulum (ER) in the Sertoli cell apical processes surrounding the late spermatids forms an extensive and continuous network. Cisternae of this network form part of the structure of the ESs, participate in a membrane contact site with the bulbs of TBCs, and have a parallel alignment with the surrounding Sertoli cell plasma membrane. Previously, we've localized the IP3R calcium channel to the TBC bulb‐ER contacts as well as other calcium exchange machinery to various parts of the ER in the apical process. Calcium release and exchange have been implicated in processes related to both actin dynamics as well as in fusion and acidification of endocytic components such as endosomes. We've proposed that calcium exchange may be a crucial mechanism in regulating both the actin networks of ESs and TBCs, as well as in facilitating the maturation and fusion of TBC bulbs as they develop into putative endosomes. If calcium exchange from the ER is responsible for regulating the local actin networks and maturation of endocytic components in the apical process, then knocking down the primary calcium release channel of the ER should alter local actin and TBC bulb morphology. To test this, we injected the testes of Sprague Dawley rats with siRNAs against IP3R1 (ITPR1) and collected tissues at 1 day, 2 day, and 3 days post injection and processed the tissues for either western blot, immunofluorescence, or electron microscopy. ITPR1 siRNA treated tissues showed significant morphological alterations to the ER, actin, and TBC bulbs in the apical processes of late spermatids compared to controls. This data supports the hypothesis that ER calcium exchange is an integral component to actin dynamics and TBC bulb maturation in the apical processes of Sertoli cells.

Support or Funding Information

Natural Sciences and Engineering Research Council of Canada (NSERC) Discovery Grant awarded to AWV; Alexander Graham Bell Scholarship (NSERC) awarded to AA

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
α‐Adducin is crucial for endothelial Tight Junction Integrity
Authors: Ponce Alexander García , Radeva Mariya Yosifova , Waschke Jens
Published: June 2021

Adducin (Add) is a membrane and cytoskeleton‐associated protein participating in the assembly of spectrin‐actin networks, which in turn regulate adherens junction (AJ) and tight junction (TJ) integrity. This process is important for endothelial barrier homeostasis. However, the mechanisms by which Add participates in endothelial barrier regulation are not well understood. Therefore, we generated immortalized Add knock‐out (KO) endothelial cell lines and investigated barrier dynamics. In absence of Add, transendothelial electrical resistance (TER) measurements showed impaired barrier function under basal conditions. Moreover, cAMP‐mediated barrier stabilization induced by treatment with Forskolin (F) and Rolipram (R) was compromised. Additionally, cells lacking Add showed higher intracellular cAMP concentration although the cAMP‐degrading enzyme phosphodiesterase 4D was overexpressed. Strikingly, ablation of Add led to a complete absence of TJ protein claudin‐5. Furthermore, in contrast to WT cells, Add KO cells displayed reduced membrane staining of other TJ molecule such as ZO‐1, both under resting conditions and after F/R treatment. However, the cellular distribution of this protein remained similar. In addition, localization of AJ protein VE‐cadherin at cell‐cell contacts was altered in Add KO cells, which was paralleled by significantly reduced protein levels. Taken together, in this study we show that Add is involved in endothelial barrier dynamics by regulating AJ and TJ integrity as well as cAMP‐mediated‐signalling

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Comparing mitochondrial dysfunction in acute and chronic mouse models of Parkinson's disease
Authors: Patki Gaurav , Lau Yuen‐Sum
Published: June 2021

Parkinson's disease (PD) is a neurodegenerative disorder associated with a progressive loss of nigrostriatal dopaminergic neurons. The etiology of PD is unsolved, although many hypotheses based on genetic, environmental and cytotoxic factors have been examined. According to in vitro and acute in vivo studies, oxidative stress and mitochondrial dysfunction leading to neuronal death have been implicated. It is not known whether mitochondrial dysfunction is an immediate response to cytotoxic‐inducing agents or is a long‐term result of neuronal death. We investigated the striatal mitochondrial function in a chronic mouse model of PD in 8‐month old, male, C57/BL retired breeders treated with 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP, 15 mg/kg) and probenecid (250 mg/kg) twice a week for 5 weeks. We compared the chronic results with that of an acute model treated with 2 doses of MPTP (15 mg/kg), 2 hours apart. Data were collected 6 weeks after the chronic model and 2 hours after the acute model treatment. In the acute model, we detected a significant inhibition of the State 3, but not the State 4 respiration in the striatal mitochondria. No changes in mitochondrial cytochrome c (cytc) and Cu/Zn superoxide dismutase (SOD) protein levels were observed. In contrast, we did not detect mitochondrial respiratory deficiency in the chronic model but the levels of cytc and SOD were significantly decreased. Our results indicate that striatal mitochondrial toxicity is different between acute and chronic models of PD. In the chronic model, there is a long‐term loss of key mitochondrial enzymes in association with neuronal loss. The remaining intact mitochondria appear to respire in a normal rate. Supported by NINDS R01‐47920.
Coq2p function in the Saccharomyces cerevisiae coenzyme Q biosynthetic pathway
Authors: Marbois Beth Noelle , Xie Letian , Shi Yuchen , Hyman Kyle , Clarke Catherine F.
Published: June 2021

Respiratory electron transport in the plasma membrane of prokaryotes and in the inner mitochondrial membrane of eukaryotes is supported by the crucial lipid component, ubiquinone (coenzyme Q or Q). Here we further characterize the Coq2 polypeptide, a key resident polypeptide of a complex of Coq polypeptides that is necessary to manufacture Q. One mutant allele of Coq2p and a hemaglutanin tagged Coq2p are described and compared to the wild‐type polypeptide in S. cerevisiae via Western blotting, growth curves and lipid analyses. Although stable levels of Coq2 polypeptide are evident in mitochondria isolated from the coq2‐1 mutant, the strain is incapable of aerobic respiration. We also present biochemical evidence that Coq2p is expressed in two forms, one at 30 kDa and another at 41kDa. Growth on non‐fermentable carbon sources invokes the 41 kDa isoform. We speculate that the 41kDa form is necessary for Q synthesis since it is lacking in the coq2‐1 mutant, and not highly expressed in cell extracts of cells grown in media containing fermentable sugars. We describe the coenzyme Q intermediates present in lipid extracts of the coq2‐1mutant. Our data demonstrate that Coq2p is essential for Q synthesis.

This work was supported by NIH grants GM 45952 and NIA AG 19777.
Detection of the Five Transmembrane Vasoactive Intestinal Peptide Receptor‐1 Isoform in Mouse Tissue
Authors: Hermann Rebecca , Palmer Megan , Dorsam Glenn
Published: June 2021

Vasoactive intestinal peptide (VIP) is a pleiotropic secretagogue involved in crucial biological processes necessary for life. Its biological effects are mediated by two structurally similar integral membrane receptors called vasoactive intestinal peptide receptor‐1 and 2 (VPACR‐1/2). VPACR‐1 is a member of the group IIA G protein coupled receptors (GPCR) which are characterized by 7 transmembrane domains (7TM). VIP binding to VPACR‐1 elicits adenylate cyclase/cAMP/PKA and phospholipase C/D signaling cascades. VPACR‐1 has a wide expression profile in the mouse, including the nervous and immune systems. Recently, a novel VPACR‐1 isoform was identified that lacks exons 10 and 11 to form a 5 transmembrane receptor (5TM), and in contrast, elicits significant tyrosine phosphorylation through a G protein independent mechanism. The biological effects mediated by this recently discovered non‐GPCR linked isoform, 5TM, are not known. To determine the expression profile of this isoform, mRNA will be collected from various mouse tissues, cDNA generated with anchored primers and the expression profile of the 7TM and 5TM quantitated using qPCR. Identifying which mouse tissues express the 5 TM VPACR‐1 isoform is a major first step in understanding its contribution to cellular function and physiology. Supported by NIH‐KO1 1K01DK064828 and COBRE 2P20RR05566.
IF‐S, a phosphorylated alpha glucomannan polysaccharide, induces functional human dendritic cell maturation
Authors: Majano Pedro L , Martin‐Vilchez Samuel , Molina‐Jimenez Francisca , Alonso‐Lebrero Jose L , Sanz‐Cameno Paloma , Rodriguez Yolanda , Roda‐Navarro Pedro , Trapero Maria , Aragoneses‐Fenoll Laura , Gonzalez Salvador , Corbi Angel L , Lopez‐Cabrera Manuel , Moreno‐Otero Ricardo
Published: June 2021

Dendritic cells (DC) maturation is a critical step for the induction of antigen‐specific T cell responses. We investigated the effects of IF‐S, a commercial drug whose active principle is the active component of GLPH‐1, on human monocyte‐derived DC (hMDDC). iDC was generated by culturing hPBM in medium supplemented with GM‐CSF and IL‐4. hMDDC derived from healthy was stimulated with IF‐S. Cell surface protein expression (HLA‐DR, CD54/ICAM‐1, ICAM‐3, CD86 and CD83), and endocytosis were analyzed by flow cytometry. IL‐12p70 and IL‐10 were assayed by ELISA. IF‐S induces or enhances the expression of CD54, CD83, CD86, HLA‐DR. Consistent with the effect on MDDC phenotype changes, IF‐S‐stimulated MDDC also exhibite a lower capacity of particle uptake when compared to unstimulated MDDC. IF‐S significantly induces the expression of IL‐12p70, although to a lesser extent than lipopolysaccharide (LPS). Both IF‐S and LPS strongly induce the expression of IL‐10. Finally, we observed that IF‐S increases the expression of the chemokines (MIP‐1α, MIP‐1β IL‐8 and MCP‐1) and chemokine receptors (CXCR4 and CCR7) mRNA levels. IF‐S also reduces CCR1 mRNA levels. These data underline the maturation‐promoting effect of IF‐S on MDDC from healthy donors suggesting that IF‐S might be a useful reagent for regulating immune responses in patho‐physiologycal situations requiring functional DC maturation.
Immunological synapse formation modulates affinity discrimination in Bcells
Authors: Raychaudhuri Subhadip , Tsourkas Phiippos , Tolar Pavel
Published: June 2021

B cells are known to use their signaling mechanism to recognize antigens over a wide affinity range (106 – 1010 M−1) and generate a graded response that depends on the affinity of antigen binding, a phenomenon known as affinity discrimination. Recent experiments have revealed that B cell signaling is crucially modulated through the immunological synapse ‐ an ordered protein segregation structure consisting of BCR/Antigen and LFA‐1/ICAM‐1 molecules that forms at the cell‐cell contact area. We have developed a Monte Carlo computer model to investigate whether synapse formation leads to differential B cell signaling upon ligation with antigens of varying affinity. For a given antigenic affinity, our numerical experiments give rise to stochastically varying levels of activated signaling molecules, such as phosphorylated BCR Igα/Igβ and Syk molecules, as observed in single cell experiments. Hence our model's stochastic nature can capture cell‐to‐cell stochastic fluctuations in B cell signaling. Based on our stochastic simulations we show that synapse formation modulates the B cell affinity discrimination. We also try to corroborate our computer simulation results with a simple mathematical model.
P53‐dependent and ‐independent components of IL‐2 mediated radioprotection in IL‐2 dependent T cell lines
Authors: Kibe Ryoko , Zvonic Sanjin , Iwakuma Tomoo , Durum Scott K , Cui Yan
Published: June 2021

Cytokines are crucial growth factors for the survival, proliferation and differentiation of hematopoictic cells. Previous studies have shown that many cytokines are radioprotective, i.e. able to rescue hematopoietic cells from radiation‐induced apoptosis. The radioprotective role of IL‐2 in IL‐2 dependent T cells and its molecular mechanism has not been well defined. We hypothesize that IL‐2 protects T cells from radiation‐induced apoptosis via up‐regulating antiapoptotic molecules and enhancing DNA repairs. It is well known that upon radiation induced double strand DNA break, cells immediately activate DNA repair machinery and initiate cell cycle arrest. Using IL‐2 dependent p53‐sufficient HT‐2 T cell line and p53‐deficient D1 cell line, we show that IL‐2 protects them from radiation‐induced apoptosis through up‐regulating antiapoptotic molecules and inducing G2 cell cycle arrest via Akt pathway. Although Akt mediated cell cycle arrest may occur independent of p53, it appears to be crucial for p53‐dependent radioprotection as inhibition of Akt activity delays successful repair of damaged DNA, which results in radiation‐induced apoptosis. Therefore, both p53‐dependent and ‐independent pathways are important for IL‐2 mediated radioprotection. This study is supported by NIH grants to YC (CA112065 and P20RR021970)
Dendritic cell differentiation in the presence of PGE2 primes dendritic cells for a proinflammatory response, induces IL‐23, and promotes Th17 differentiation
Authors: Khayrullina Tanzilya R , Yen Jui‐Hung , Ganea Doina
Published: June 2021

Dendritic cells (DC) play an important role in the induction of the immune responses. The DC phenotype and function is modified by different factors. PGE2 plays an important role in inflammation and autoimmunity through changes in the proinflammatory cytokine profile. The proinflammatory cytokine IL‐23 is associated with autoimmune disorders. In the periphery, macrophages and DC are major producers of IL‐23, whereas microglia play a similar role in the CNS. IL12 and IL‐23 share the p40 subunit, but have specific p35 (IL‐12) and p19 (IL‐23) subunits. The main goal of this study was to define the role of PGE2 in DC differentiation and production of bioactive IL‐23. DC were generated in GM‐CSF (DC) or GM‐CSF+PGE2 (DCp) and characterized following stimulation with LPS. PGE2 functions as an enhancer of DC differentiation and influence their Th‐cells lineage driving ability, by providing factors required for Th17 differentiation and survival. Activated DCp have higher levels of proinflammatory cytokines, such as IL‐6, IL‐1β and TNF, as a result of higher levels of TLR expression and of NFkBp65 phosphorylation and nuclear translocation. In addition, PGE2 shifts the IL‐23/IL‐12 balance in favor of IL‐23 in activated DC. These observations may help to understand the distinct role of PGE2 in the induction of pathogenic Th17‐cells.

This work is support by NIH/NIAID R01AI052306
Lymphoma cell surface expression of CD200 modulates anti‐tumor immunity
Authors: Wong Karrie Ka Wai , Gorczynski Reg , Spaner David
Published: June 2021

CD200 is a transmembrane protein broadly expressed on a variety of cell types and delivers immunoregulatory signals through binding to receptor (CD200Rs) expressed on monocyte/myeloid cells and T lymphocytes.

We showed in earlier reports that infusion of a soluble form of CD200, CD200Fc, into EL4 thymoma‐bearing C57/B6 mice enhanced tumor progression (Clin. Exp. Immunol. 2001). More recently, independent groups have reported CD200 overexpression associated with multiple myeloma, AML, and CLL.

In this study we investigated the in vitro effect of blockade of CD200 expression, using both anti‐CD200 mAbs and CD200‐specific siRNAs, on CTL induction in human PBL using a CD200+ lymphoma cell line (ly5). We found that blockade of CD200 on ly5 cells significantly enhanced anti‐tumor CTL responses in vitro. Production of the inflammatory cytokines TNFa and IFNg were similarly enhanced. Cell depletion studies supported an important role for antigen presenting cells in CD200 regulation of anti‐tumor responses. We also developed a CD200 ELISA assay to measure serum CD200 levels in healthy donor controls and cancer patients, and in preliminary analyses found elevated levels of sCD200 in the plasma of patients with CLL.

Our data provide novel insights into the mechanisms of CD200 mediated suppression of tumor immunity, the understanding of which may open new avenues to development of different cancer therapies.
Food group intake patterns differ between low‐income Puerto Ricans and Non‐Puerto Rican pregnant Latinas
Authors: Bermúdez‐Millán Angela , Hromi‐Fiedler Amber , Damio Grace , Segura‐Pérez Sofia , Pérez‐Escamilla Rafael
Published: June 2021

Latinos are the largest ethnic minority group in the United States. They are a heterogeneous group, represented by people from Mexico, Central and South America, and the Caribbean Islands. A longitudinal study (n=234) was conducted to examine differences in food group intake between low income Puerto Ricans and Non‐Puerto Rican pregnant Latinas. A single 24‐hour dietary recall was collected to assess food group intake. At baseline 94% of the participants were in their second to third pregnancy trimester. Mean age was 24 ± 5y. Puerto Ricans comprised 69% of the sample. Mean length stay in the US was 15 ± 9y for Puerto Ricans and 5 ± 5y for Non‐Puerto Ricans. Puerto Ricans were more likely to be bilingual, have a larger household composition, participate in WIC and the food stamp program and less likely to have a partner. Non‐Puerto Ricans consumed significantly (p<0.05) more servings of dark green‐leafy vegetables, tomatoes, other vegetables, fruits, beans, tortillas and candy/sugars. Puerto Ricans consumed more servings of breakfast cereals, crackers, processed meat, cheese, dairy desserts, soft drinks and sweetened fruit drinks. Acculturation may be playing an important role as Non‐Puerto Ricans had been for a shorter period of time in the US than their Puerto Rican counterparts.

Funded by the Connecticut NIH EXPORT Center of Excellence for Eliminating Health Disparities among Latinos (NIH‐NCMHD Grant # P20MD001765).
Maternal anthropometric measures of risk for gestational diabetes mellitus in Mexican pregnant woman
Authors: Perez Marcela , Casanueva Esther , Haene Jessica de , Parra Adalberto , King Janet C
Published: June 2021

The prevalence of overweight and obesity is rising in pregnant as well as non‐pregnant Mexican women. A pre‐pregnancy BMI ≥ 25 increases the risk of adverse pregnancy outcomes. Simple anthropometric measurements may help identify pregnant women at risk of metabolic diseases, such as impaired glucose tolerance (IGT) and gestational diabetes mellitus (GDM). The aim of this study was to determine if anthropometric measurements are associated with the diagnosis of GDM among pregnant Mexican women. We collected anthropometric data (weight, height, skin folds, hip and waist perimeters) at 20, 24 and 28 wks in 34 women without any metabolic disease at booking. Nine of the 34 women were diagnosed with GDM at 28 wks. At all time points, body fat percentage (BFP), estimated by the Durnin equation with the sum of 4 skin folds, was significantly higher among women with GDM (Normal (n=25) 36.2% ± 0.8, 36.2% ± 0.7, 36.6% ± 0.7 vs GDM (n=9) 37.9% ± 1.3, 38.4% ± 1.3, 38.5% ± 1.2; p<0.001). Waist and hip perimeters at 20 wks correlated with BFP at 28 wks (r=0.547, p≤0.001; r=0.655, p≤0.001). The BFP at 28 wks was also associated with a GDM diagnosis (r=0.400, p=0.026). These data show that simple anthropometric measurements are useful tools for identifying Mexican pregnant women at risk for developing GDM. Partially support by UCMEXUS and Bristol Myers Squibb.
Effects Of Rapid Or Slow Body Mass Reduction On Body Composition In Young Rats
Authors: Tai Shinji , Kondo Emi , Okamura Koji
Published: June 2021

We showed that skeletal muscles did not decrease regardless of rapid (R) or slow (S) body mass (BM) reduction, whereas splanchnic tissues decreased more in R than S in adult rats (EB2005). To investigate if body fat stores affect the results observed in the above study, young rats whose body fat was less than adult rats were subjected to R or S BM reduction. Fifteen rats (6‐wk‐old) were assigned to a group fed ad‐lib (C, n = 5), moderately restricted energy intake for 14 days to reduce BM gain (S, n = 5) or fasted for the last 3 days of the 14‐day study to reach comparable BM in S rapidly (R, n = 5). All rats were trained with a climbing cage 3 times a week (5 min × 6 sets). Drinking water was withdrawn for 19 h before sacrifice in R. Final BM was 316 g (SD 12), 238 (14), 236 (5) in C, R and S, respectively. The mass, water, protein and total lipid of carcass and excised skeletal muscles were less in both R and S than C. These measurements did not differ between R and S except for the gastrocnemius which was lower in S than R. The adipose tissue mass was significantly lower in both R and S than C and the decrease tended to be greater in S than R. The mass of the liver, stomach and small intestine was lower in both R and S than C and the mass of these tissues was lower in R than S due to lower content of water and protein in R than S, but total lipid of these tissues were similar in R and S. These results demonstrate that both skeletal muscle and splanchnic tissues decrease in both R and S in rats with less body fat stores and a decrease in skeletal muscle would not be influenced by the rapidity of BM reduction whereas a decrease in splanchnic tissues is greater in rapid BM reduction. Thus, the less body fat stores, the more skeletal muscle decreases, while body fat stores does not appear to influence a decrease in splanchnic tissues which is greater in rapid BM reduction.
Comparison of BCFA types in vernix and meconium of healthy neonates
Authors: Ran‐Ressler Rinat , Devapatla Srisatish , Lawrence Peter , Brenna J. Thomas
Published: June 2021

Background. Branched Chain Fatty Acids (BCFA) are rare in mammalian lipids but are known to be synthesized in the sebaceous gland. They are richest in vernix caseosa, the substance that covers the fetus in late gestation. During pregnancy, the fetus swallows amniotic fluid and with it sloughed vernix particles. We recently reported the presence of BCFA in meconium, the first pass of the neonate. Since vernix and meconium are the input and output, respectively, of the GI tract, we hypothesized that the BCFA profile of the two would be related in some systematic way.

Methods. Vernix was skimmed off newborns in the delivery room and meconium was collected into diapers and saved for analysis. Samples were processed by a modified Bligh & Dyer extraction, and FAME were analyzed by GC and GC/MS. The BCFA profiles for vernix and meconium was compared for individual neonates and on average.

Results. Vernix BCFA contain iso‐BCFA, anteiso‐BCFA, dimethyl BCFA and middle‐chain monomethyl BCFA and range from 12 to 26 carbons. In contrast, meconium BCFA are mainly iso‐BCFA and range from 16 to 26 carbons. Major differences in profiles were found from infant to infant.

Conclusions. BCFA are a normal component of the term newborn gut throughout its entire length. BCFA with <16 carbons and with branching other than iso disappear in the sterile fetal gut in late gestation. The mechanism is unknown. Funding: NIH GM071534.
Bone Mass and osteoporosis risk determination in a group of students aged 20 to 30 years old attending a University Campus
Authors: Pérez‐Lizaur Ana , Tamayo‐Orozco Juan A , Clark‐Peralta Patricia , Cruz‐Santiago Victoria
Published: June 2021

There is scarce information regarding bone mass accretion and its determinants in the skeleton of people aged 20 to 30 years. In order to define risk factors that may interfere with PBM acquisition, we did apply simultaneously a validated questionnaire and measured bone mass in the non dominant Os Calcis with a LUNAR DXA (PIXI) in a convenience sample of 347 students (75.5 % women) attending the Universidad Ibero Americana, Mexico City. According to the number of risk factors present 51% had less than 2 risk factors, the low risk group. 45.8% reported less than 4 risk factors (moderate risk group) only 3.1 % of this sample reported more than 5 risk factors, representing a high risk group. Normal BMD (between minus 0.99 and over 0.99 standard deviations), low (between minus 1.0 and minus 1.8s.d.) and very low (values below minus 1.8s.d.). Mean age was 21.6 years (range 20 to 30), 75.5% had normal BMI, 15.6% is overweight, and 6.1% has low body weight. We found a significant correlation between BMD and female sex (ƒÓ2 = 11.82; p< 0.02) BMI (<18.5) and fracture risk factors (ƒÓ2 = 31.15; p< 0.002). We conclude that this screening strategy is useful to define the risk profiles present in a convenience sample of students normally attending to their school activities at the University Campus, enabling us to design intervention strategies to improve PBM as a prevention strategy for future Osteoporosis.
Natural forms of vitamin E inhibited leukotriene B4 generation and 5‐lipoxygenase translocation in Ca2+ ionophore‐activated human HL‐60 cells
Authors: Jiang Qing , Yin xinmin
Published: June 2021

Leukotriene B4 (LTB4) generated by 5‐lipoxygenase (5‐LOX)‐catalyzed reaction plays an important role in regulation of inflammation and contributes to the development of cancer. Vitamin E has eight natural forms, i.e., alpha‐, beta‐, gamma‐ and delta‐tocopherol and four corresponding tocotrienols. Previous studies of the effect of alpha‐tocopherol on 5‐LOX‐catalyzed reaction revealed inconsistent results. In the present study, we examined the effect of various forms of vitamin E on leukotriene B4 generation in neutrophil‐like cells that were differentiated from HL‐60 cells by 1.25% DMSO for 5–6 days. Activation of HL‐60 cells by calcium ionophore A23187 led to a markedly increase of LTB4. Gamma‐tocotrienol, delta‐tocophoerol and gamma‐tocopherol dose‐dependently inhibited LTB4 with apparent IC50s at 25–50uM. In contrast, alpha‐tocopherol did not show any significant inhibition at concentrations of 10–50uM. Gamma‐ and delta‐tocopherol significantly inhibited A23187‐induced 5‐LOX translocation from cytoplasm to nuclear compartment, a critical step for leukotriene formation. These observations support the notion that certain vitamin E forms may have anti‐inflammatory activity which leads to disease prevention and therapy. Studies are currently undertaken to further investigate the mechanism underlying observed effects. (supported by NIH R01AT001821)
Metabolic syndrome, physical activity and diet in early postpartum women
Authors: Lu Hongxing , Cahill Jodi M , Sanghani Bijal V , Freeland‐Graves Jeanne
Published: June 2021

Metabolic syndrome (MS) includes a cluster of risk factors for heart disease and type 2 diabetes. The purpose of this research was to study the relationships of physical activity and diet with MS in early postpartum. At 1–4 months after childbirth, anthropometrics, blood pressure, blood lipids glucose and insulin were measured in 141 low income women. Physical activity level was assessed by pedometers, and diet, by a 195‐item food frequency questionnaire. Metabolic syndrome was defined according to the National Cholesterol Education Program Adult Treatment Panel III guidelines. The incidence of MS was 34.3%, 35.6% and 16.1% for subjects 18–25, 26–30 and 31+ yrs, respectively. Low serum HDL‐cholesterol was the best predictor of metabolic syndrome (R2= 0.315, p<0.01). Physical activity was inversely related to MS and insulin resistance via HOMA (p<0.05). Women with MS had fewer daily servings of fruits (2.7 vs 3.1) and vegetables (4 vs 5.6) than those without MS (p<0.05). Vegetable intake was negatively associated with systolic blood pressure (r=−0.23, p<0.05). Metabolic syndrome was not related to intakes of other food groups. In summary, MS was quite prevalent in these relatively young low‐income women in early postpartum and associated with less physical activity and reduced consumption of fruits and vegetables. Supported by TX Coordinating Board, # UTA00‐377.
Malnutrition in a probabilistic sample of Mexican elderly population
Authors: Gómez Luz María , Villalpando Salvador , Shamah Teresa , Cuevas Lucia
Published: June 2021

Malnutrition in elderly population is a growing problem in Mexico. The objective of this investigation is to describe the prevalence and distribution of malnutrition and its association with some potential determinants in a probabilistic sample of 5430 adults 60 years of age and older from the Mexican National Nutrition Survey (ENSANUT‐2006). Specific questionnaires for demographic information, depression and physical disabilities were applied and height and weight were measured. Malnutrition was defined as a BMI less than or equal to 18.5. The prevalence progressed with age reaching 5% in adults older than 80 years. In a logistic regression higher risks for malnutrition were associated with age (OR 3.39, CI95% 1.86, 6.20) and low socioeconomic status (OR 4.06 CI95% 1.91, 8.63). Depression and physical disabilities were not associated with the risk for malnutrition (OR 0.63 CI95% 0.20, 1.94) and (OR 0.61, CI95% 0.25, 1.48).

Our results alert about a public health problem in a rapid growing elderly population demanding prompt interventions to control it.

Supported by Ministery of Health, Mexico.
Effectiveness of folic acid (FA) supplementation in Mexican women with genetic methylenetetrahydrofolate reductase (MTHFR) polymorphisms
Authors: De Regil Luz Maria , Casanueva Esther , Ramírez Arlete , Meza Carlos , Hernández‐Guerrero César , Viteri Fernando E
Published: June 2021

In Mexico 1 of 250 newborns has a neural tube defect (NTD) and 70% of population has a mutation in the gene C677T that synthesizes MTHFR, a key enzyme in FA metabolism.

Objective. To evaluate the effectiveness of FA supplementation in ensuring an adequate folate nutritional status in women of childbearing age with different MTHFR gene mutations.

96 healthy women stratified according to their C677T genotype [CC (wild type), CT or TT] entered a double‐blind clinical trial. They were supplemented with 400μg FA/day. Red cell (RBC) folate and homocysteine were measured at baseline, after 30 days of FA supplementation and 30 days after it was stopped. Data were analyzed by Repeated Measures ANOVA.

3% had NTD history. Independent of genotype, baseline RBC folate and plasma homocysteine were similar and FA equally increased RBC folate even after stopping supplementation. A quadratic reduction (p = 0.001) in homocysteine levels occurred independent of genotype. The highest reduction was found in the TT group, but in all cases basal homocysteine levels were reestablished 30 days after ceasing FA.

Conclusion. FA is effective in increasing RBC folate and in reducing homocysteine levels, independently of the C677T genotype. Partially supported by UC_MexUS.
Ghrelin, adrenaline and neostigmine stimulate the in vitro expression of ghrelin in lambs’ abomasal tissue slices?
Authors: González‐Dávalos Laura , Pérez Eleazar , Anaya Ana M. , Mora Ofelia , Romano José‐Luis
Published: June 2021

Ghrelin is a 28 amino acid acylated peptide secreted by several digestive organs; X/A cells of stomach's fundic region seem to be its main producers‐ Ghrelin's positive effect on food intake has a negative connotation in humans; in contrast, the same effect could be a positive factor in farm animals’ production; therefore, our aim was to evaluate the effect of administration of human synthetic ghrelin, adrenaline, and neostigmine, on ghrelin expression in lambs′ abomasal tissue in vitro. Samples from 12 lambs were used. At slaughter, tissue slices were washed with saline solution, stored in tubes with media culture (DMEM‐F12, FBS 20%, penicillin/streptomycin and amphotericin) and transported to the lab. Slices were placed in polypropylene inserts, in 24‐well multidishes with 500 μl of medium, and distributed to five treatments: control; adrenaline 10 μM; neostigmine 0.1 μM; ghrelin (500 and 1000 pg), in four exposure times (0, 1, 2, 3 h). Ghrelin expression was identified by purifying RNA from tissue slices using the RT‐PCR technique. Total RNA was extracted from the cells by QIAGEN™ protocol (RNeasy Mini Kit). RT was made using oligo (dT)12–18 primer and PCR were made using the primers: Forward 5′‐CATCTACAGCCTGCTGCTG and Reverse 5′‐CACATCCGGGTCAAACTG. The expression of ghrelin was detected in agarose gel with a band of 361 bp. The results showed no differences among treatments; pending confirmation with Real Time PCR.
MARKERS IN EXPANSION OF REGENERATING & EARLY POSTNATAL PANCREAS
Authors: Hua Hong , Sarvetnick Nora
Published: June 2021

Type 1 diabetes results from destruction of insulin‐producing pancreatic beta cells. A possible treatment is to replace these cells by genetic engineering or the use of stem cells. This approach requires that we understand the signals that regulate beta cell proliferation. Inhibition of DNA binding proteins (Ids), are implicated in a number of cellular processes, including control of proliferation and differentiation. Ids bind to and inhibit the function of bHLH transcription factors including those that regulate pancreatic development. Moreover, bone morphogenetic proteins (BMPs) are shown to regulate the expression of Ids. We found that neutralization of BMP4 significantly reduced duct epithelial cell expansion in a mouse model of islet regeneration. BMP4 stimulation promotes Id2 binding to the bHLH transcription factor NeuroD, which is required for the differentiation of pancreatic islet cells. Moreover, using BTC cells we demonstrated that Id2 enhanced proliferation through the Rb/E2F pathway. The early postnatal pancreas represents an intriguing model since expansion of beta cells is still evident. We found BMP4 enhanced two surface markers that may be relevant in proliferation in neonatal pancreatic cells and during islet regeneration. Collectively, our data provide novel insights into the signaling pathways required for beta cell proliferation, which may be useful for treating diabetes.
Preprotachykinin‐A mRNA changes in response to cisplatin‐induced emesis in the least shrew
Authors: Dey Dilip C , Ray Andrew P. , Abad Joseph , Darmani Nissar A.
Published: June 2021

Cisplatin (CIS) and related drugs cause vomiting in cancer patients. We used the least shrew (Cryptotis parva) emesis model to study this effect, as our previous work has shown that CIS‐treated shrews mimic the biphasic vomiting pattern seen in humans. The acute phase, due presumably to gut and/or medullary serotonin release, occurs within hours. The delayed phase, due presumably to medullary release of substance P (SP), occurs after an emesis‐free period of one (shrews) to several days (humans). We studied the role of SP in CIS‐induced emesis by injecting least shrews with 10 mg/kg CIS (i.p.) and measuring changes in mRNA levels of preprotachykinin‐A (PPTA), the precursor for SP. At different post‐CIS times, tissues from CIS or control shrews were collected and processed for quantitative PCR (QPCR). Whole brain tissue showed a modest increase in PPTA up to 3.5h, a 3 to 14‐fold increase until 33h, then a return to control levels. QPCR of gut tissue showed a rapid rise to 6h, followed by a decrease to below control levels. At 20h, it increased gradually until 33h, then fell below the basal level. Overall, brain PPTA was constantly up‐regulated for 33h, while gut PPTA was upregulated biphasically, coincident with the emetic phases. Thus, SP production in both brain and gut is coordinated with, and may play a role in the modulation of, both phases of CIS‐induced emesis. This study was supported by NIH grant #R01CA115331 to Dr. Darmani.
Phenylephrine induces conducted constrictions in the uterine vasculature of the pregnant mouse in situ.
Authors: Leonard Sean , Junuzovic Jasna , Croy B. Anne , Murrant Coral L
Published: June 2021

Upon implantation, maternal‐ and fetal‐derived signals adapt the uterine vasculature to ensure adequate blood flow to the developing embryo. Conducted responses are important in matching blood flow to tissue metabolism, therefore they could represent a means by which fetal metabolism may influence maternal blood flow. To determine whether conducted responses exist in the uterine vasculature, we developed an in‐situ mouse model to measure blood vessel reactivity within this vasculature. The blood vessels were visualized by staining the endothelium with Isolectin Alexa‐Fluor 488 (0.067M). 10‐4M phenylephrine (PHE) was applied directly to the uterine artery (max diameter approx. 70 um). Vessel diameter was measured at the site of application and two downstream sites: the first >1000um downstream and the second 1000um further downstream from the site of application. Direct PHE application caused the uterine artery to constrict 31.4 +/− 4.9um (n = 9). Significant conducted constrictions to PHE were observed; 14.5 +/− 3.7um (n = 5) and 17.9 +/– 7.4um (n = 5) for the first and second downstream sites respectively. Direct application of PHE to the first and second downstream sites yielded constrictions of 45.1 +/− 11.7um and 40.3 +/− 12.6um respectively. These data show that conducted constrictions exist within this vasculature and suggest the possibility of fetal control of the maternal vasculature. NSERC and CIHR.
Thyroid hormone induces artery smooth muscle cell proliferation: discovery of a new TRα1‐Nox1 pathway
Authors: Wang Xiuqing , Sun Zhongjie
Published: June 2021

Hypothesis

Thyroid hormone (T3) induces artery smooth muscle cell proliferation by up‐regulating NOX1 dependent on thyroid hormone receptor subtype α1 (TRα1).

Methods and Results

Immunofluoresence confocal microscopy was used to visualize the sub‐cellular localization of NOX1 and TRα1 in rat aorta smooth muscle (RASM) cells. TRα1 and NOX1 co‐localized in the nuclear periphery. T3 induced RASM cell proliferation as determined by the fact that T3 significantly increased cytokinesis cells, proliferating cellular nuclear antigen (PCNA), and smooth muscle α‐actin (SM α‐actin). T3 increased NOX1 expression at both the transcription (mRNA) and translation (protein) levels. T3 also increased the intracellular ROS production. RNAi silence of TRα1 or NOX1 abolished T3‐induced intracellular ROS generation and PCNA and SM α‐actin expression, indicating that TRα1 and NOX1 mediate T3‐induced RASM cell proliferation. Notably, RNAi silence of TRα1 blocked the T3‐induced increase in NOX1 expression whereas silence of NOX1 did not affect TRα1 expression, suggesting a new pathway, i.e., T3‐TRα1‐NOX1‐cell proliferation.

Conclusions

TRα1 expressed in RASM cells. TRα1 and NOX1 colocalized in nuclear periphery. T3 induced RASM cell proliferation by up‐regulating NOX1 in a TRα1‐dependent manner.
Expressions of arterial inflammatory regulatory proteins in experimental diabetes mellitus
Authors: Adeagbo Ayotunde S , Moshal Karni , Fleming John , Joshua Irving G. , Wang Yang , Fernandez‐Botran Rafael G.
Published: June 2021

Vascular inflammation is a hallmark of Type‐1 and to a lesser extent, Type‐2 diabetes mellitus. We investigated the status of arterial cyclooxygenase‐2 (COX‐2), prostaglandin D and E synthases (PGDS and mPGES) and P450 2J2 proteins in healthy and streptozotocin (STZ) diabetic rats. Age‐matched male Sprague Dawley control and STZ‐diabetic rats were used. After 6 or 12 weeks STZ or vehicle, mesenteric arteries were harvested from rats, cleaned and prepared for Western blot analyses of target proteins. Serum levels of 6‐keto‐PGF1a, nitrite/nitrate (NOx), PGE2 and 8‐epi‐isoprostanes were determined with enzyme immunoassay kits. STZ‐diabetic were persistently hyperglycemia; COX‐2, mPGES proteins, 8‐epi‐isoprostanes, PGE2, 6‐keto‐PGF1a and NOx were significantly elevated versus control rats. CYP 2J and PGDS proteins were down‐regulated in diabetic versus control arteries. The significant increases in the expression of COX‐2 and mPGES proteins as well as enhanced serum PGE2 and NOx levels and reduced expressions of CYP 2J and PGDS are compatible with development of arterial inflammation. Support: University of Louisville Intramural Research Incentive Grant.
Chronic inhibition of rho‐kinase augments contractions in diabetes
Authors: Lamping Kathryn Griffin , Nuno Daniel W.
Published: June 2021

Objective: Acute inhibition of rho‐kinase decreases augmented contractions of arteries in diabetic animals. The objective of this study was to determine if chronic inhibition of rho‐kinase with fasudil would prevent enhanced contractions of arteries from diabetic mice.

Methods: Non‐diabetic (ND) and streptozotocin‐induced diabetic (D) male mice (Strep 100–150 mg/kg ip) were treated with fasudil (100mg/kg/day) in their drinking water for 12–14 weeks. Responses of isolated aortic rings to serotonin (5HT, 0.01–10 microM), thromboxane mimetic U46619 (0.01–1 microM), KCl (25–75 mM), and acetylcholine (Ach, 0.1–10 microM) were compared to age‐matched untreated ND and D mice.

Results: Fasudil had no effect on weights or glucose levels of ND (mg/dl minus fasudil 213±10; plus fasudil 200±8) or D mice (mg/dl minus fasudil 428±56; plus fasudil 458±55). 5HT, U46619 and KCl produced concentration‐dependent contractions of aorta from ND. Contractions to 5HT and KCl but not U46619 were increased in D arteries compared to ND. Surprisingly, fasudil increased contractions to 5HT and KCl in ND and D. Relaxation to ACh was not altered in D but was decreased similarly in both ND and D by fasudil.

Conclusion: We conclude that chronic fasudil does not protect arteries of diabetic mice from development of abnormal vasoconstriction.

This study was supported by grants from the Dept of Veterans Affairs and American Diabetes Association.