bluuebunny/crossref_metadata_2025 huggingface dataset:
Overexpression of apelin in the paraventricular nucleus increases blood pressure and sympathetic nerve activity in normotensive rats
Authors: Zhang Qi , Yao Fanrong , Pingili Ajeeth , O'Rourke Stephen T. , Sun Chengwen
Published: June 2021

Studies of the peripheral apelin/APJ receptor system indicate that it plays a significant role in cardiovascular regulation. Here, we determined the in vivo actions of apelin in the paraventricular nucleus (PVN) on the central regulation of blood pressure (BP) and sympathetic outflow. Microinjection of exogenous apelin‐13 into the PVN caused dose‐dependent increases in mean arterial pressure (MAP), heart rate (HR) and renal sympathetic nerve activity (RSNA). To further ascertain the role of apelin in PVN, we overexpressed apelin within PVN by AAV2 vector‐mediated gene transfer. Western blot analysis showed that microinjection of AAV2‐apelin into the PVN resulted in a significant increase in apelin expression and that overexpression of intrinsic apelin caused chronic BP elevation and a significant increase in plasma norepinephrine levels. In addition, APJ receptors were localized in RVLM projecting PVN neurons using an immunofluorescent retrograde labeling approach and superfusion of these neurons with exogenous apelin‐13 significantly increased neuronal firing activity, which was measured with whole‐cell patch clamp recording in current‐clamp mode. In summary, the data demonstrate that the apelin/APJ receptor system in the PVN may play an important role in the regulation of sympathetic outflow and in the central control of BP.
Angiotensin (1‐7) Antagonizes the Enhanced Chronotropic Effects of Angiotensin II in Spontaneously Hypertensive Rat (SHR) Brain Neurons
Authors: Modgil Amit , Zhang Qi , Yao Fanrong , O'Rourke Stephen T. , Sun Chengwen
Published: June 2021

Ang (1‐7) plays a major role in counter‐regulating the actions of Ang II in the periphery, but its actions in the CNS are not fully understood. This study was designed to determine the interaction between Ang II and Ang (1‐7) in regulating neuronal activity in neurons cultured from hypothalamus of SHR vs. WKY rats using current clamp recording. Superfusion of neurons with Ang (1‐7) (10 nM‐1 μM) did not alter the neuronal firing rate in either SHR or WKY neurons. Pretreatment of neurons with Ang (1‐7) (100 nM) decreased the Ang II‐induced enhancement in neuronal firing rate in SHR neurons, but not in WKY neurons. This counter‐regulatory action of Ang (1‐7) was blocked by A‐779 (10 μM), a Mas‐receptor antagonist. Previously we demonstrated that the enhanced chronotropic effect of Ang II in SHR neurons is mediated by PI3K. Thus, the role of PTEN (phosphatase and tensin homologue), which dephosphorylates phosphoinositide substrate, was studied in the action of Ang (1‐7). The PTEN inhibitor, BPV (10 μM), abolished the effect of Ang (1‐7) on Ang II action. Incubation of SHR neurons with Ang (1‐7) increased PTEN activity by 116%. The data demonstrate that Ang (1‐7) stimulates PTEN activity via Mas‐R, dephosphorylates phosphatidylinositol‐3,4,5‐trisphosphate, and depresses PI3K signaling pathway, which leads to a counter‐regulatory effect of Ang (1‐7) on Ang II action in neuronal activity of SHR neurons.
Evidence that a Singing Impediment is Caused by Impaired Interhemispheric Coordination of Brain Activity
Authors: Behbahaninia Milad , Vu Eric T
Published: June 2021

The neural network that mediates learned vocalizations in oscine songbirds provides a useful animal model for understanding the neural mechanisms mediating certain aspects of human speech. We studied the song impairment in zebra finches caused by unilateral lesion of nucleus Uvaeformis (Uva), a thalamic nucleus important for song production, as a model for the human speech impediments that can arise after brain injury. We observed an immediate degradation of singing premotor neural activity in the HVC (a brain nucleus encoding higher‐order features of song) after unilateral Uva lesion, as well as a loss of coordination of singing premotor activity across the hemispheres. To establish a more direct link between interhemispheric coordination of brain activity and the quality of song syllables produced, we compared HVC neural activity in lesioned birds during production of the most consistently formed song syllables (“Good”) vs the most poorly formed syllables (“Evil”). We discovered a significant difference in time‐specific synchronizing signals between Good and Evil syllables. This provides some of the most conclusive evidence that impairment of interhemispheric coordination is the physiological basis for some vocal impediments. Supported by the Howard Hughes Medical Institute through the Undergraduate Science Education Program, the ASU School of Life Sciences and the Barrow Neurological Foundation.
Evaluation of PI3K expression and its interaction with angiotensin II on central autonomic nuclei of hypertensive animals
Authors: Antunes Vagner Roberto , Buttler Leila
Published: June 2021

There is evidence that PI3K protein is directly linked to the intracellular signaling of the angiotensin II (ANG II) pathways (via AT1 receptors) that act as a neuromodulator of autonomic outflow in spontaneous hypertensive rats (SHR). In the present study we evaluate the difference on the PI3K expression as well as its interaction with ANGII in the PVN, NTS and RVLM of SHR compared to WKY and wistar rats. Immunoblotting was performed to quantify the regulatory subunit p85 of the PI3K and phosphorilated PKB (active form) in the PVN, NTS and RVLM of SHR and normotensive rats. Moreover, we also evaluated the role of ANG II and its interaction with PI3K at the PVN level in regulating the lumbar sympathetic nerve activity (LSNA) by using an in situ arterially perfused preparation. The expression of p85 subunit did not differ among all nuclei studied at any experimental group, but the phosphorilated PKB was significantly more expressed in the PVN of SHR (1.5x) when compared to wistar and WKY. ANG II (10 μM) microinjected into the PVN elicited an increase in LSNA (18%), which was attenuated at 5 min after LY294002 (50 μM) microinjected into the same site (2%), with recovering 20 min later (17%). If there is any interaction between ANG II and PI3K pathway related to hypertensive stage it seems that PVN is the pivotal autonomic nucleus involved in the sympathoexcitation of this pathology.

Research Support: FAPESP, CAPES and CNPQ.
Double Inferior Vena Cava in a patient with lymphoma
Authors: Schoneboom Bruce A , Rengasamy Padmanabhan
Published: June 2021

The inferior vena cava (IVC) develops by a complex process from diverse embryonic sources and drains blood from a large number of structures inferior to the diaphragm. Congenital malformations of the IVC are reported to be rare and most of these malformations are accidental findings through abdominal imaging studies. Although compatible with life, these anomalies are of major clinical importance and can be a contributing factor in thrombus formation. This case report describes a double IVC in an adult male cadaver who died of lymphoma and cardiac events. The common iliac veins were found to continue superiorly and fuse together slightly cranial to the level of the superior mesenteric artery origin from the aorta at the L1 vertebral level. The left IVC received the left gonadal vein. The left suprarenal vein opened into the corresponding renal vein which extended superiorly and fused side to side with the left IVC as well as with the common IVC. Multiple factors may contribute to embryonic IVC malformations including genetic, and environmental including the expression and regulation of growth factors that control epithelial–mesenchymal transformation (e.g. TGF‐â), formation of endothelium, vascular endothelial growth factors (VEGF) and various VEGF receptors.
The selection and early discovery of an anti‐HGF aptamer
Authors: Spielberger Kevin , Genga Ryan
Published: June 2021

Hepatocyte Growth Factor (HGF) specifically binds the c‐Met receptor by forming a homodimer from two heterodimeric molecules. HGF binding to c‐Met results in a multifunctional signaling cascade that activates the upregulation of cellular proliferation under normal conditions. HGF is overexpressed in many types of cancers and increased activity of the HGF:c‐Met pathway causes tumorigenesis and metastasis. We sought to generate an aptamer to HGF that would inhibit the functional interaction between HGF and c‐Met.

Aptamers are oligonucleotides that fold into three‐dimensional structures and specifically bind to ligands with high affinity. Aptamers bound to proteins can modify and/or inhibit protein‐protein interactions. Through Systematic Evolution of Ligands by EXponential enrichment (SELEX), an anti‐HGF aptamer that inhibits the HGF:c‐Met pathway was generated from a modified RNA pool. A systematic analysis was carried out to identify the aptamer's smallest functional core and to further optimize the aptamer, resulting in a high affinity, nuclease‐resistant aptamer. Addition of a polyethylene glycol moiety allows for use of aptamer in vivo. The resulting aptamer inhibits cell mobility in a cell scatter assay and reduces tumor growth in the tumor model. These results demonstrate that the anti‐HGF aptamer inhibits tumor proliferation and has the potential to be used as a cancer therapeutic.
Activation of renal NMDA by Hcy causes ECM remodeling by modulating MMP/TIMP axis
Authors: Gargoum Riyad Rachid , Rosenberger Dorothea , Abe Oluwasegun , Lawal Rasheed , Sen Utpal , Tyagi Suresh
Published: June 2021

Elevated levels of homocysteine (Hcy) also known as hyperhomocysteinemia (HHcy) is one of the major causes of end‐stage renal disease. Disruptions in glomerular extracellular matrix (ECM) resulting from remodeling by activated matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinase (TIMPs) is the primary cause of impaired renal filtration. Although Hcy is a ligand to the ionotropic N‐methyl D‐aspartate (NMDA) receptor and the NMDA‐R1 subunit has been shown to be involved in renovascular calcium‐mediated toxicity, the mechanism of this toxicity is unclear. To test the hypothesis that HHcy causes glomerulosclerosis in part by causing imbalances in the MMP/TIMP axis, we created HHcy in mice and treated them with or without 0.5mg/kg MK801 (NMDA receptor blocker) by intraperitoneal injection for the last 10 days. Mice were grouped into: 1. Wild type (WT), 2. HHcy, 3. WT+MK801, 4. HHcy+MK801. The kidneys were isolated, weighed and homogenized. To measure NMDA‐R1, MMP‐2 and‐9, TIMP‐1, ‐3, and ‐4, western blot analysis was performed. The results suggest that homocysteine is involved in modulating the MMP/TIMP axis by changes in TIMP‐1, and ‐4 in a manner that is reversed by MK801.
EGF cooperates with TGFβ to regulate PAI‐1 expression by a synergistic increase in transcription and stabilization of mRNA
Authors: Song Xiaoling , Nilsen‐Hamilton Marit
Published: June 2021

PAI‐1, the primary inhibitor of plasminogen‐activators, plays an important role in extracellular matrix remodeling and fibrinolysis. Over‐expression of PAI‐1 is found in many diseases including cancer, fibrosis and metabolic syndrome. EGF and TGFβ synergistically elevate PAI‐1 expression up to 80‐fold in mink lung epithelial cell (Mv1Lu) and human hepatoma (HepG2) cells. The mechanism of synergism was investigated in Mv1Lu cells in which it was found that, while EGF and TGFβ synergistically increase PAI‐1 mRNA transcription, EGF alone increases PAI‐1 mRNA stability. TGFβ also increases the sensitivity of the cellular response to EGF. The results of experiments using specific chemical inhibitors of protein kinases, including MEK1/2, p38 MAPK, PI3K, SRC and JNK, showed that synergism between EGF and TGFβ was lost only when ERK1/2 phosphorylation was blocked. Growth factors other than EGF that activate the MAPK pathway, including IGF‐1, FGF‐2 and TNFα, also synergize with TGFβ to increase PAI‐1 expression. Investigation of the responses to EGF and TGFβ of several recombinant promoters containing different combinations of transcription elements identified an interaction between AP‐1 and a SMAD as the probable mechanism of synergism. A model to explain the synergism is proposed in which Fos is uniquely activated in response to the combination of EGF and TGFβ but not in response to either growth factor alone.
Thanks to Frances K: A case study for developmental biology
Authors: Holzman Jennifer
Published: June 2021

Traditionally, developmental biology is taught as a stand‐alone, upper division course at most colleges and universities. General topics in development are briefly touched upon in introductory level survey biology courses. At Emory University, we currently teach a 6 week long developmental biology unit during the second semester of our introductory biology sequence, Foundations of Modern Biology. During the laboratory portion of the course, we strive to include new and innovative laboratory activities in addition to problem‐based learning (PBL) exercises. The use of case studies in the laboratory has helped students to relate the material they learn in class – which they often view as irrelevant – to a real‐life situation. Our case studies primarily center around medical topics, as the majority of our students declare themselves as intending to attend medical school in the future. For the developmental biology unit, the case study involves a topic with which students may have little previous knowledge – the use of thalidomide and its side effects. This poster describes the case study we have used and its assessments to show that difficult concepts in developmental biology can be taught in creative ways to introductory biology students.
Expression and Purification of the human Lymphocyte Activation Gene‐3 for Structural Studies
Authors: Rivera Amilcar , Zavala‐Ruiz Zarixia
Published: June 2021

The lymphocyte activation gene 3 (LAG‐3) protein was first identified as a cell surface molecule selectively expressed in activated human T lymphocytes, Natural Killer cells and T‐cell‐dependent activated B cells. The LAG‐3 is a transmembrane protein homolog of CD4 protein. LAG‐3 binds Major Histocompatibility Complex (MHC) class II molecules with much higher affinity than CD4 and acts as an inhibitor of antigen‐driven T cell expansion. This protein can also be detected in a soluble form (sLAG‐3), and in fact, it is now known that can be cleaved from the cell surface. In the soluble form, sLag‐3 induces MHC class II mediated signaling in dendritic cells, leading to their activation and maturation as well as production of cytokines. Also high levels of sLAG‐3 protein in the sera of breast cancer patients tend to correlate with good prognosis of the disease. This correlation suggests that sLAG‐3 may well be an important effectors molecule regulating the immune response to tumor antigens. We plan to study the structure of the extracellular part of the protein and the interaction with MHCII. Here we reported the insertion of the different variants of the LAG‐3 gene into the pENTR vector, and the transfection in mammalian cells. In addition, we demonstrated the expression of the LAG‐3 protein. This information will give us a better understanding of the function of sLAG‐3 and why this protein is a good adjuvant for vaccines.
The Function and Evolution of the Mastoid Process and Clavicle in Humans: A Natural Experiment
Authors: Osborn Michelle L , Homberger Dominique G
Published: June 2021

We hypothesize that, in humans, the evolution of a uniquely enlarged mastoid process and clavicle was connected to the acquisition of an upright posture and of freed hands. Our biomechanical model shows that when the arms are moved or loaded, the contracting upper trapezius muscles counterbalance the added weight on the shoulders. The synchronously contracting sternocleidomastoid muscles prevent the head from being retroflexed. To test our hypotheses, we analyzed the asymmetry of biomechanically relevant character pairs in 54 right‐handed male skeletons, based on the premise that bones subjected to greater forces are larger. The humerus diameter, mastoid process width, and rise of the superior nuchal line were more often larger on the right side, while the clavicle length and scapular breadth were so on the left side. These biomechanically relevant characteristics were found in varying aggregates of 2–5 for each individual. This variability may be due to differing occupations. The forces that mold the mastoid process and clavicle in extant humans probably also selected for their robust size within the Homo lineage.

Funding: LSU Robert C. West Fund to MLO; LSU Foundation to DGH
Structural and functional studies of two esterases (Est25and Est‐Y29) from metagenome library
Authors: Kim Seungbum , Yoon Sangyoung , Jo Do Hyun , Kim T. Doohun , Ryu Yeonwoo
Published: June 2021

Enzymes with high enantioselectivity are highly useful for making several industrial processes such as preparation of pharmaceutical intermediates. Two esterases (Est 25 and Est‐Y29), which are identified from a metagenomic library, catalyze the kinetic resolution of (R/S)‐ketoprofen ethylester for the production of the anti‐inflammatory drug, (S)‐ketoprofen. Substrate binging site of these esterases were mutated to increase their stability and enantioselectiviy of enzyme. W68K and L255W of Est 25 and F137W and R237A of Est Y29, present much higher enantioselectivity.
Regulation of eIF4F complex assembly by Mnk1/2 kinase
Authors: Korneeva Nadejda , Rhoads Robert E
Published: June 2021

The rate of translation initiation in most eukaryotic cells depends on both the features of the mRNA and the level and activity of the eIF4F complex, which consists of eIF4G, eIF4E and eIF4A. mRNAs with highly structured 5′UTR encode a disproportionate share of proteins involved in cell growth and cell cycle progression, growth factors, growth factor receptors, and cyclins. Translation of these mRNAs requires high levels of eIF4E that presumably acts through the eIF4F complex. One current model suggests that the eIF4F integrity is regulated by the PI3K/Akt/mTor cascade, while the MAPK pathway targets the eIF4F cap‐binding activity. The activation of mTor kinase leads to increase of eIF4E·eIF4G complex formation via phosphorylation of eIF4E‐binding protein 4E‐BP that releases eIF4E. MAPK signaling targets the Mnk kinase via Erk1/2 and p38 pathways. Mnk is the only known physiological kinase that phosphorylates eIF4E at single site (S209, human eIF4E). Mnk binds eIF4G and phosphorylates eIF4E primarily in a complex with eIF4G. Earlier we showed that inhibition of Mnk activity in cell‐free translation system affects translation of mRNA that contains both a cap‐ and a structured 5′UTR and also affects binding of eIF4E with eIF4G. We suggest the involvement of Mnk kinase in the dynamics of eIF4F assembly. Supported by grant R01GM20818 from the NIGMS.
Studies on the Role of Acid Sphingomyelinase and Ceramide in the Regulation of TACE Activity and TNFalpha Secretion in Macrophages
Authors: Nikolova‐Karakashian Mariana Nikolova , Rozenova Rozenova Krasimira A. , Deevska Gergana , Karakashian Alexander
Published: June 2021

Acid Sphingomyelinase (ASMase) activity mediates LPS signaling in various cell types. This study shows that in macrophages, ASMase is a negative regulator of LPS‐induced TNFα secretion. ASMase‐deficient (asm−/−) mice and peritoneal macrophages produce several fold more TNFα than their wild‐type (asm+/+) counterparts when stimulated with LPS, while the addition of exogenous ceramides or SMase reduces the differences. The underlying mechanism for these effects is not transcriptional but post‐translational.

The TNFα converting enzyme (TACE) catalyzes the maturation of the 26kD precursor (proTNFα) to the active 17kD form (sTNFα). Asm−/− macrophages exhibited 2 to 3 fold higher TACE activity than asm+/+ macrophages and adding ceramide or SMase suppressed this difference. Our studies further showed that the activity of TACE was rate‐limiting factor for TNFα production and proTNFα that was not processed was internalized and degraded in the lysosomes. Indirect immunofluorescence experiments involving inhibitors of TACE, endocytosis, and lysosomal proteolysis showed that in asm−/− cells significant portion of proTNFα is sequestered within the early endosomes and instead of undergoing lysosomal proteolysis, it is recycled to the plasma membrane and processed to sTNFα. In conclusion, these experiments define a novel role of ASMase in regulation of TNFα secretion by macrophages.
Alcohol potentiates HIV protease inhibitor‐induced ER stress and hepatic lipotoxicity (1001.3)
Authors: Hinton Michael , Liu Runping , Zhang Xiaoxuan , Wang Xiang , Pandak William , Hylemon Phillip , Zhou Huiping
Published: June 2021

Alcohol consumption, which is highly prevalent in HIV‐infected individuals, has been associated with the amplification of liver toxicities induced by HIV protease inhibitors (PIs). We have demonstrated that HIV PI‐induced endoplasmic reticulum (ER) stress is linked to the dysregulation of hepatic lipid metabolism. However, little is known about the mechanistic pathways by which alcohol promotes HIV PI‐induced hepatic lipotoxicity. The aim of this study was to examine the cellular mechanisms by which alcohol promotes HIV PI‐induced hepatic lipotoxicity. Methods: Primary hepatocytes isolated from rat, wild type mouse or CHOP‐/‐ mouse were treated with individual HIV PIs with or without alcohol. Activation of the ER stress was determined by real time RT‐PCR and Western Blot. Apoptosis was detected by Annexin V‐FITC/propidium iodide staining. Intracellular lipid accumulation was determined by nile red staining. Results: Alcohol and HIV PIs synergistically induced ER stress, lipid accumulation and apoptosis in primary hepatocytes. Both alcohol and HIV PI‐induced lipid accumulation and apoptosis were significantly reduced in CHOP‐/‐ hepatocytes. Conclusions: Activation of the ER stress response is a key cellular mechanism underlying alcohol and HIV PI‐induced hepatotoxicity. Inhibition of ER stress activation may represent a new strategy to prevent alcohol and HIV PI‐associated hepatic lipotoxicity.

Grant Funding Source: VA Merit Award
ABO modulation of endothelial phenotypes and glycome profile in human induced pluripotent stem cell‐derived endothelial cells (1003.2)
Authors: Zhang Hanrui , Mooney Ciarán , Ishihara Mayumi , Kumagai Tadahiro , Stalnaker Stephanie , Park Seongha , Veilion Lucas , Hinkle Christine , Stout Matthew , Yang Wenli , VanDorn Daniel , Chou Stella , Bhang Dong Ha , Ryeom Sandra , Tiemeyer Michael , Wells Lance , Azadi Parastoo , Reilly Muredach
Published: June 2021

Genome‐wide association studies have suggested that, compared to non‐O blood types, the ABO O blood type phenotype and related genotypes are associated with protection from CVD. To test the hypothesis that ABO glycosyltransferase modulates endothelial functional pathways and glycan profile, we differentiated adipose stem cell (ASC)‐derived induced pluripotent stem cell (iPSC) from individuals of different ABO blood types to endothelial cells (ECs). iPSC‐ECs exhibited cobble‐stone morphology, demonstrated capillary tube formation on matrigel, and incorporated acetylated low‐density lipoprotein. FACS analysis indicates expression of both CD31 and vascular endothelial growth factor receptor‐2 (VEGFR2). Immunocytochemical analysis of iPSC‐ECs revealed positive staining for EC markers e.g., CD31, vascular endothelial cadherin (CD144), and von Willebrand factor (vWF). The phenotypic characteristics of iPSC‐ECs were similar to that of human coronary arterial endothelial cells (hCAECs). The adhesion pattern of iPSC‐EC to a variety of extracellular matrix (ECM) was similar across O and non‐O iPSC‐EC, but ABO blood group O showed lesser adhesion to vitronectin and laminin (mean±SD; 2.37±0.19 folds in non‐O vs. O for vitronectin [p=0.02] and 2.30±0.05 folds in non‐O vs. O for laminin [p=0.004]). Further, gap closure assay showed that O type iPSC‐EC had greater migration capacity. We examined glycan profiles of pooled iPSC‐ECs and pooled hCAECs from subjects with matched blood types. N‐linked glycan profiling showed that series of high‐mannose structures were observed as main components of iPSC‐EC and hCAECs. Complex structures were observed as minor components. iPSC‐ECs which gave similar profile to that of hCAECs. Main O‐glycans in iPSC‐ECs and hCAECs was a monosialylated core 1 structure. In summary, we established the feasibility of differentiating EC from subject‐specific ASC‐iPSC as tools to define disease‐relevant ABO modification of EC functions.

Grant Funding Source: NIH U01, K24
Intracellular hyaluronan protects murine fibroblasts against apoptosis (1003.3)
Authors: Wang Yan , Lauer Mark , Mack Judith , Maytin Edward
Published: June 2021

Skin repair typically involves apoptotic cell death. We are interested in the role of hyaluronan (HA), an extracellular matrix molecule, during fibroblast‐mediated repair of injured skin. Has1/3 null mice (Has1/3), which lack hyaluronan synthases 1 and 3 (yet retain Has2) show accelerated wound closure, more fibroblasts, and fewer cleaved caspase‐3–positive dermal cells in healing wounds, suggestive of reduced apoptosis. In cultured Has1/3 fibroblasts, HA levels and Has2 mRNA expression were higher than in WT cells. Apoptosis, induced by serum starvation (SS) or UVB exposure, was reduced in the Has1/3 cells as shown by DNA fragmentation and by cleavage of caspases and PARP. Hyaluronidase, added to remove extracellular HA, surprisingly showed no effect on apoptotic susceptibility to SS or UVB. However, cells treated with 4‐MU to inhibit HA synthesis were sensitized to apoptosis induced by SS or UVB. To further establish a role for HA in protection from apoptosis, Has2 RNA interference (RNAi) was done using Has2‐specific siRNA transfection. Has2 RNAi in WT and Has1/3 fibroblasts inhibited HA synthesis by 90% and significantly increased their sensitivity to apoptosis. External addition of high MW HA failed to reverse this effect. We conclude that Has1/3 skin fibroblasts (which make relatively more HA) are more resistant to apoptosis due to the presence of intracellular HA synthesized by Has2.

Grant Funding Source: supported by NHLBI
Glycoproteomics of the Zabrotes subfasciatus larval midgut associated with the insecticidal mechanism of the Olneya tesota PF2 lectin (1006.1)
Authors: Lagarda‐Díaz Irlanda , Guzmán‐Partida Ana , Geiser Dawn , Winzerling Joy , Vazquez‐Moreno Luz
Published: June 2021

Insect midguts, as well as other animals, are rich in glycoconjugates that have important biological functions required for growth and development. Dietary lectins can bind to gut glycoconjugates and induce various insecticidal effects such as blocking of receptors or transporters and inactivation or activation of enzymes. PF2 is a lectin isolated from the seeds of Olneya tesota (Palo Fierro), and reported to cause 100% mortality of Zabrotes subfasciatus larvae (Mexican bean weevil). We evaluated recognition of the PF2 lectin by α‐amylases from Z. subfasciatus larval midgut and the effect of PF2 on α‐amylase activity. Results demonstrated PF2 decreased total midgut amylase activity. In‐gel amylase activity assay showed three enzyme isoforms, but only one isoform was recognized by PF2. The identity of this Z. subfasciatus α‐amylase was confirmed by LC‐MS/MS. We also evaluated the binding of PF2 to midgut glycoproteins at different larval stages using techniques such as chromatography, 2D‐gel electrophoresis and lectin blotting. Lectin interaction revealed differences in glycoprotein patterns at different larval stages. Analysis by LC‐MS/MS and searches of insect databases revealed several glycoproteins that may serve as potential targets for PF2 recognition. Each of the identified proteins is physiologically significant to weevil larvae and may be associated with the insecticidal mechanism of PF2.

Grant Funding Source: Supported by funds from the CONACYT, the Arizona Ag Experiment Station and CALS
Dysregulated affinity of lamin A to SUN1 induces nuclear and endoplasmic reticulum aberrancies in progeric laminopathies (1009.1)
Authors: Chi Ya‐Hui
Published: June 2021

Hutchinson‐Gilford progeria syndrome (HGPS) is a human progeroid disease caused by a point mutation on the LMNA gene. We have previously reported that the accumulation of the nuclear envelope (NE) protein, SUN1, contributes to HGPS nuclear aberrancies. However, how the interaction between the mutant lamin A (also known as progerin or LAΔ50) and SUN1 produces HGPS cellular phenotypes requires further elucidation. Using light and electron microscopy, this study demonstrated that SUN1 contributes to progerin‐elicited structural changes in the NE and the endoplasmic reticulum (ER) network. This study identified the full‐length lamin A associated with SUN1 in two domains, with the farnesylated cysteine within the CaaX motif (C661farnesyl) showing a stronger affinity to SUN1 than to the amino acids within 607 to 656 (LMNA‐SID). Farnesylation of progerin enhances its interaction with SUN1 and reduces SUN1 mobility, thereby promoting the aberrant recruitment of the ER membrane during postmitotic assembly of the nuclear envelope, and resulting in the accumulation of SUN1 over consecutive cellular divisions. These results indicate that the dysregulated interaction of SUN1 and progerin in the ER during NE reformation determines the progression of HGPS.

Grant Funding Source: Supported by the National Science Concil, Taiwan
Genome‐wide microRNA and mRNA profiling in skeletal muscle aging (1009.5)
Authors: Kwon Ki‐Sun , Kim Ji Young , Lee Seung‐Min , Lee Kwang Pyo , Kim Seon‐Young
Published: June 2021

Skeletal muscle degenerates progressively, loses mass (sarcopenia) along in years, and leads to reduced physical ability, often causing secondary diseases such as diabetes and obesity. MicroRNAs (miRNAs) play critical roles in a variety of biological processes and it is also known that regulation of gene expression by miRNAs is a key event in muscle development and disease. Although there are several report to profile miRNA expression in skeletal muscle with age, these studies are typically medium scale profiling using microarray technique. Here, we performed next‐generation sequencing which allowed us to assess genome‐wide expression profiling of miRNAs during aging in combination with RNA‐Seq using mouse gastrocnemius muscle at two different ages (6‐mo‐old versus 24‐mo‐old). We found that 39 miRNAs were differentially expressed with age, 20 up‐regulated and 19 down‐regulated. We identified miRNAs, such as miR‐34, ‐206, or ‐434, which are differentially expressed in aged muscle, demonstrating that our data covers, at least in part, previous findings. Further detailed comparison of miRNA expression with its target gene expression from RNA‐Seq data revealed that miRNAs contribute to muscle aging possibly through the positive regulation of transcription, metabolic process, kinase activity. Finally, we believe genome‐wide miRNA profiling helps to expand our knowledge of miRNA function in the muscle aging process.

Grant Funding Source: Supported by Grant 20110030134 from NRF of Korea
Cell survival proteins in fermented milk (1010.16)
Authors: Coria Evelyn , Payne Monique
Published: June 2021

Cell Survival Proteins in Fermented Milk Probiotics are living microorganisms that when administered in adequate amounts can have beneficial effects to the host. Fermented milk, which contains probiotics, has shown the potential to protect against excessive oxidative/ nitrosative stress. Overproduction of nitric oxide (NO·) within the cell ultimately leads to cell death. Past studies have alluded to the idea that components produced by the microbiota may serve as growth factors for the cells. The aim of this study was to determine if fermented milk protects cells from nitrosative stress as well as identifying the active component. The results showed that proteins isolated from fermented milk aided in the recovery/survival of cells treated with NO·. Further investigation of the fermentation process revealed new proteins were produced. Identification and cloning of these protein(s) will not only help elucidate the “pro‐survival” mechanism, but may act as potential therapies for any oxidative/nitrosative based phenomenon or disease.

Grant Funding Source: Supported by Title V STEM
N‐(1‐pyrenyl) maleimide induces Bak oligomerization and mitochondrial dysfunction in Jurkat cells (1010.4)
Authors: Wang Tzu‐Chien , Huang Pei‐Rong , Pao Chia‐Chu
Published: June 2021

N‐(1‐pyrenyl) maleimide (NPM) has been recently identified as a telomerase inhibitor in a cell‐free system and shown to display a great differential cytotoxicity against hematopoietic cancer cells. However, the primary cytotoxic effect of NPM against hematopoietic cancer cells is induction of apoptosis, rather than inhibition of telomerase. In this study, the molecular mechanism by which NPM induces apoptosis was examined. Here, we show that treatment of Jurkat cells with NPM leads to Bak oligomerization, loss of mitochondrial membrane potential (Δψm), and release of cytochrome C from mitochondria to cytosol. Induction of Bak oligomerization appears to play a critical role in NPM‐induced apoptosis, as down‐regulation of Bak by shRNA significantly prevented NPM‐induced apoptosis in Jurkat cells. Inhibition of caspase‐8 by Z‐IETD‐FMK and/or depletion of Bid did not affect NPM‐induced oligomerization of Bak. Taken together, these results suggest that NPM‐induced apoptosis is mediated through induction of Bak oligomerization and mitochondrial dysfunction by a pathway that is independent of caspase‐8 activation.
Static high‐gradient magnetic fields affect the functionality of monocytic cells (1010.5)
Authors: Syrovets Tatiana , Schmidt Zoe , Büchele Berthold , Zablotskii Vitalii , Dejneka Alexandr , Dempsey Nora , Simmet Thomas
Published: June 2021

Magnetic fields, different to electric fields, can intrude into deeper layers of tissue, where they could modulate biochemical processes. In this context, we explored the cellular effects of specifically designed high‐performance micro‐magnet arrays which, in contrast to conventional magnets, generate high‐gradient magnetic fields not encountered in the environment. Exposure of monocytic cells to high‐gradient magnetic fields of various strength generated by micro‐magnet arrays induced cell swelling followed by accumulation of reactive oxygen species (ROS), and inhibition of cell proliferation. High levels of ROS may damage cellular structures and induce apoptosis. Consistently, prolonged exposure of cells to high‐gradient magnetic fields induced time‐dependent translocation of phosphatidylserine to the outer leaflet of the membrane. The extent of cell swelling, ROS production, and apoptosis correlated with the degree of the magnetic field gradient. On the contrary, when cells were exposed for prolonged time to a conventional high‐performance bulk magnet, which produces a strong, yet low‐gradient magnetic field, neither ROS nor apoptosis induction became detectable. These data clearly show that the static high‐gradient magnetic fields generated by the micro‐magnet arrays do affect monocytic cell functionality and may be used to manipulate intracellular biochemical processes.

Grant Funding Source: Supported by DFG; ASCR, grant M100101219; and ANR, grant ANR‐11‐BSV5‐0101.
Comparative heat shock response of wild yeast isolated from spontaneous mezcal fermentation and controlled laboratory yeast (1011.2)
Authors: Mejía‐Barajas Jorge , Arellano‐Plaza Melchor , Saavedra‐Molina Alfredo
Published: June 2021

The yeast manufacture is one of the oldest and the most widespread technologies in the field of industrial biotechnology, but yeast are subjected to many types of stress. These range from the physical to the chemical, secondary metabolite toxicity and heat. Oxidative stress plays a major role in the lethal effect of heat in yeast. This stress generates reduced forms of molecular oxygen, known as reactive oxygen species (ROS). In this work, we study the capacity to tolerate heat shock, production of ROS and their relation with mitochondrial respiratory electron transport, lipoperoxidation and glutathione production, of different yeast strains, isolated from spontaneous mezcal fermentation, two Kluyveromyces marxianus (OFF1 and SLP1), one Saccharomyces cerevisiae (MC4) and one controlled laboratory yeast S. cerevisiae (W303‐1A). Results observed different growth kinetics and growth rate, where the strains of K. marxianus showed faster growth. The strain SLP1 showed the best growing at 40 °C and the rest of the strains at 30 °C. The yeasts more tolerant to heat (OFF1 and SLP1) showed the biggest mitochondrial membrane potential. The higher resistance heat strain SLP1 exhibited the lesser lipoperoxidation and bigger glutathione production of the strain. The results suggest that the wild strains, mainly SLP1 strain, may differ from the responses to heat shock that have been characterized for W303‐1A. Acknowledgements: The authors appreciate the partial economic support grants from CIC‐UMSNH (2.16 to ASM), CONACYT (169093 to ASM). JMB is CONACYT fellow.
Activation of the farnesoid X‐receptor suppresses cyclin D1 expression and decreases proliferation (1011.3)
Authors: Idelman Gila , Vogel Megan , Zucker Stephen
Published: June 2021

Bile acids have increasingly been recognized to play a modulatory role in cancer formation. We previously have shown that administering oral sodium taurocholate to APCmin/+ mice decreases cellular proliferation in intestinal crypts and markedly reduces adenoma formation . The current study validates that bile acids exhibit an anti‐proliferative effect through FXR‐mediated transcriptional activation of Small heterodimer partner (Shp) which, in turn, inhibits cyclin D1 expression. Human colon (HT‐29), breast (MCF7), hepatocellular (HepG2) carcinoma cells and nontransformed mouse hepatocytes (AML12) were incubated in the presence of a synthetic (GW4064) or native (chenodeoxycholic acid, CDCA) FXR agonist for 48 hours. All cell lines except HepG2 monolayers exhibited a significant decrease in cell proliferation at 24 h and 48 h. As expected, Shp mRNA and protein expression was significantly higher in treated cells (confirming activation of FXR), with a corresponding decrease in the expression of cyclin D1 mRNA and protein. Notably, cyclin D1 expression in HepG2 cells remained unchanged despite a significant increase in Shp expression. These data support the hypothesis that activation of FXR inhibits cell proliferation through down regulation of cyclin D1. The uncoupling of this effect in HepG2 cells might be a result of higher basal levels of FXR activity and Shp expression.
Reactivities of the sulphydryl groups of horse (Equus caballus) haemoglobin (1013.11)
Authors: Omotosho Omolola , Shalom Chinedu , Omotosho Temidayo , Okonjo Kehinde
Published: June 2021

The kinetics of the reaction of Ellman’s reagent (DTNB) with CysF9[93]β sulphydryl group of the horse haemoglobins were studied at neutral and physiological pH (6.8 < pH > 7.6) ranges under pseudo first order conditions. The reaction is of first order with respect to the DTNB concentration. The reactions are pH dependent of the observed rate constant gave a complex trend. The observed rate shows that at neutral pH, the presence of inositol hexakisphosphate (inositol‐P6) increases the pseudo first order rate constant. For the first time, inositol‐P6 increases rate of forward reaction, kF at neutral pH values by increasing the mean value of the transition constant, Krt3. The Krt3 for the haemoglobin without inositol‐P6 gave the value of 0.138 ± 0.1 while the haemoglobin in the presence of inositol‐P6 gave the Krt3 value of 0.325 ± 0.2. The results show that inositol‐P6 increases the relative population of the t tertiary conformation. So, it increases the reactivity of CysF9[93]β by changing the relative distribution of two protein conformations.
Mechanistic insights into nitric oxide activation of H‐NOX sensor proteins (1013.18)
Authors: Herzik Mark , Jonnalagadda Rohan , Marletta Michael
Published: June 2021

Nitric oxide (NO) is a freely diffusible gas that is critical for normal cardiovascular function through the regulation of processes such as blood vessel homeostasis and inhibition of platelet aggregation. An integral component of mammalian NO signaling is soluble guanylate cyclase (sGC), the primary receptor for NO, whereby sensitive and selective binding of NO to its heme‐containing H‐NOX domain results in conversion of guanosine triphosphate (GTP) to the secondary messenger cyclic guanosine monophosphate (cGMP). The heme cofactor of sGC exists in a 5‐coordinate ferrous state (Fe(II)) with histidine as the proximal ligand. NO binding to the distal heme site yields a transient 6‐coordinate complex where, the trans effect exhibited by NO weakens the iron‐histidine bond resulting in histidine dissociation to yield an active, 5‐coordinate NO complex (Fe(II)‐NO). Several biochemical studies have determined that severing of the iron‐histidine bond is required for activation, however, the lack of a 5‐coordinate NO H‐NOX structure has limited molecular level insights into the NO activation mechanism. Toward this end, crystal structures of a bacterial H‐NOX protein free and complexed with NO have been determined and reveal a conformational change resulting from the breakage of the iron‐histidine bond. It is hypothesized that this conformational change is the mechanism by which H‐NOX proteins regulate their signaling effector partners.
Empowering adolescent girls against anemia in West Bengal, India (1014.6)
Authors: Chakraborty Priyanka , Bhattacharyya Indrani
Published: June 2021

Objective: To describe the baseline characteristics (2011) and intervention strategies of the government funded scheme SABALA, an effort to empower adolescent girls (AG), 11‐18 yrs to prevent and manage iron deficiency anemia in six high priority districts of West Bengal, India.

Study design and methods: This is a cohort study of AGs, selected as beneficiaries under the SABALA scheme. Target sample size (1080 AGs) was estimated using non‐probability purposive sampling for survey, final sample size was 804. For FGDs and interviewing 108 AGs and stakeholders were selected respectively. Knowledge assessment on general nutrition, cause and symptoms of anemia and awareness on IFA tablets were conducted. Information on demography and family background were also collected. Intervention strategies proposed by the scheme will include peer‐mentoring, whereby 1‐3 AGs in Anganwadi (ICDS) Centers of the districts will be trained on health and nutrition issues to promote anemia prevention strategies and motivate other AGs to join the scheme. Other interventions will include IFA supplementation, health check‐up and referral services and Nutrition and Health Education. SPSS 19.0 was used for all data analysis.

Baseline characteristics (Results): Early marriage (40%) and school dropouts (50%) were among the prominent problems in all the 6 districts. While 79% of AGs were aware of anemia in Cooch Bihar district, only 25% were aware in the remaining 5 districts (p < 0.05). Other than Cooch Bihar, > 70% of AGs had never consumed IFA tablets (p < 0.05). Study results indicate stakeholders have limited knowledge and awareness on AG health issues and IFA sources, which can be potential barriers for implementing the scheme. Logistic regression results showed no significant association with nutrition knowledge and anemia prevalence among the respondents in each district.

Conclusion: Results from this baseline study identify the priority areas and barriers for the successful implementation of the SABALA scheme.
Body mass index influences breastfeeding initiation and duration in a central Texas WIC population (1016.6)
Authors: Thornton Hannah , Crixell Sylvia , Shumake Jason , Reat Amanda , Von Bank Julia , Friedman B.J.
Published: June 2021

US breastfeeding rates fall short of federal goals. Choices to initiate and continue breastfeeding are made in the context of demographic, biological, social, and psychological variables. The objective of this study was to explore determinants of breastfeeding initiation and duration among participants in the Special Supplemental Nutrition Program for Women, Infants, and Children (WIC) in a small city in central Texas. Following IRB protocols, caregivers of children ages 4‐24 months were recruited before and after federally mandated changes to the WIC package, and completed telephone interviews (n=204). Chi‐squared analysis revealed no significant differences in outcome variables before and after the package change. Logistic regression revealed caregiver BMI (OR 0.928, 95% CI [0.874, 0.985], p<0.05) and caregiver insurance (OR 0.379, 95% CI [0.164, 0.874], p<0.05) had a significant negative effect on breastfeeding initiation; Spanish spoken at home (OR 3.42, 95% CI [1.44, 8.10], p<0.01), caregiver having a live‐in partner (OR 2.27, 95% CI [1.16, 4.46], p<0.05), and caregiver age (OR 1.06, 95% CI [1.00, 1.12], p<0.05) had a significant positive effect on breastfeeding duration; and caregiver BMI (OR 0.921, 95% CI [0.868, 0.977], p<0.01) had a significant negative effect on breastfeeding duration.
Anti‐diabetic efficacy of Fenfuroœ, a novel, patented Trigonella foenum‐graecum seed extract (1019.14)
Authors: Bagchi Debasis , Swaroop Anand , Kumar Pawan , Bagchi Manashi
Published: June 2021

Anti‐diabetic efficacy of a novel Trigonella foenum‐graecum seed extract (FenfuroTM) enriched in furostanolic saponins was investigated. Sprague‐Dawley rats were given high‐fat diet for 2 weeks followed by streptozotocin (STZ, 35 mg/kg ip). Animals were checked for plasma glucose level (PGL) 2 days post+STZ injection to confirm diabetes. Rats showed >300 mg/dl of PGL were included. FenfuroTM (150‐ and 450 mg/kg po) and glipizide (5 mg/kg po) were administered once daily for 20 days and then twice daily for 10 days over 30 days. Blood was collected on 0, 10, 20 and 30 days and estimated for fasting plasma triglyceride (PTG), TC and Insulin levels. On day 30, FenfuroTM and glipizide‐treated diabetic animals were treated with/without metformin (100 mg/kg) twice daily for the next 10 days. FenfuroTM didn’t affect the body weight, feed and water intake. FenfuroTM (150 mg/kg po) reduced the PTG levels by 22%, 24.6% and 29% on 10‐, 20‐ and 30‐days of treatment, respectively, while glipizide (5 mg/kg po) reduced the PTG levels by 57.4%, 46.2% and 39.4%. FenfuroTM (450 mg/kg) treatment exhibited significant hypoglycemic activity (31.5%) as compared to insulin (48.2% with 1 U/kg i.p.). Fenugreek (150 mg/kg po) and Metformin (100 mg/kg po) combination produced significant reduction in PGL (20.7%). These studies demonstrated the efficacy of this FenfuroTM extract in type 2 diabetes.

Grant Funding Source: Supported by Cepham Inc, NJ
Healthy kids, healthy Cuba: strategies to increase healthy food access in an ethnically diverse, rural Southwest community through policy and environmental change (1019.3)
Authors: Keane Patricia , Ortega Alejandro
Published: June 2021

Objective: The Healthy Kids, Healthy Cuba project aims to increase access to fresh, healthy food and safe places to walk and play through policy and environmental change in this small, tri‐ethnic rural community in northern New Mexico. Strategies include the development of a farmers’ market, expansion of community garden, vending policies at county fairgrounds, and promotion of backyard gardening in outlying American Indian communities.

Methods: Qualitative Interviews, focus groups, field notes, environmental assessments.

Results: Unique challenges in small, rural communities can be overcome to improve access to fresh healthy foods, but require persistence, identification of key champions, and innovative, culturally‐responsive approaches. Additional results will be reported.

Conclusions: Healthy Kids, Healthy Cuba may serve as a model for other rural communities facing similar challenges and limited resources.

Grant Funding Source: Supported by the Robert Wood Johnson Foundation’s Healthy Kids, Healthy Communities Initiative