bluuebunny/crossref_metadata_2025_split huggingface dataset:
ΔNp63α suppresses cell invasion through downregulating Rac1 activity
Authors: Aljagthmi Amjad Ahmed , Cooke Mariana , Kazanietz Marcelo , Kadakia Madhavi P. 
Published: April 2020

ΔNp63α, a member of the p53 family of transcription factors, is overexpressed in a number of cancers and plays a role in proliferation, differentiation, migration and invasion. We showed previously that ΔNp63α suppresses cell invasion by positively regulating miR‐320a, resulting in the downregulation of its direct target protein kinase C gamma (PKCg) and a corresponding reduction in phosphorylated Ras‐related C3 botulinum toxin substrate 1 (Rac1). Here, we further show that ΔNp63α inhibits Rac1 GTP activity. Using a Rac1 pulldown assay to measure Rac1‐GTP levels, we showed that silencing ΔNp63α in A431 cells led to a significant upregulation of Rac1‐GTP. Silencing ΔNp63α in A431 cells also led to an increase in p21‐activated kinase (PAK1) phosphorylation, confirming the negative regulation of ΔNp63α on Rac1. Inhibiting Rac1 GTP activity using Ehop‐016 led to a significant reduction in both pRac1 and pPAK1 levels. Treatment with Ehop‐016 reversed the increase in cell invasion observed upon silencing ΔNp63α. RT‐PCR analysis performed on an array of 43 Rac guanine exchange factors (Rac‐GEFs) showed that 6 GEFs are significantly upregulated by silencing ΔNp63α in A431 cells. Taken together, our data suggest that ΔNp63α negatively regulates Rac1 GTP activity via downregulation of a Rac‐GEF, thereby reducing Rac1‐GTP levels and inhibiting cancer cell invasion.Support or Funding InformationThis work was supported by a grant from the National Cancer Institute [1R01CA154715] to M.P.K.
Production of Ethanol from Lactose in Engineered S. cerevisiae
Authors: Vinz Cami Alyssa 
Published: April 2020

Wisconsin is the dairy capitol of the United States. However, all this cheese production leaves behind a lot of unused by product in the form of cheese whey. If cheese whey could be fermented into consumable ethanol there would be less discarded into the trash and could also be a new revenue stream for Wisconsin dairies. Our plan is to modify yeast that can already produce ethanol from sucrose to also be able to make the ethanol from cheese whey that contains lactose.By engineering two strains of Saccharomyces cerevisiae to metabolize lactose we predict they will be able to digest and ferment cheese whey. This is done by allowing the yeast to express lactose permease and beta galactosidase, LAC12 and LAC4 respectively. These genes can be found in Kluyveromyces marxianus and Kluyveromyces lactis.With this production of ethanol from cheese whey we plan to patent this newly found technique which in return will lead to lots of vodka and money.Support or Funding InformationThis project is funded by UW ‐ Stout's Office of Research and Sponsored Programs.
Evaluation of Virus‐based Vectors for Cystic Fibrosis Gene Therapy
Authors: Rodríguez-Villafañe Andrea , Kuntz Leah , Limberis Maria P. 
Published: April 2020

Cystic fibrosis (CF) is a life‐threatening genetic disease caused by mutations in the CF transmembrane conductance regulator gene. It is the most common genetic disease among Caucasians. There is an urgent need to create more therapies for CF since current drugs are not effective in treating all the mutations that cause this disease. Furthermore, gene therapy is a potential treatment for CF airway disease. Because of this, various viral‐based gene transfer vectors have been evaluated for their efficiency in correcting the CF airway disease phenotype. This project aimed to compare side‐by‐side the differences in the transduction efficiency and targeting of specific cell types of adenovirus (Ad)‐based and adeno‐associated virus (AAV)‐based vectors in different models of the airway. Different serotypes of AAV, namely 1, 2, 5, 6, and Ad5‐based vectors, expressing green fluorescent protein (GFP), firefly luciferase (ffLuc) or ß‐galactosidase (LacZ) were assessed in in vitro and in vivo gene transfer studies. Madin‐Darby Canine Kidney (MDCK) cells were grown submerged on plastic, and human airway epithelial (HAE) cells were grown on transwells and fed only from the basolateral side to attempt to model the human conducting airways. The viral vectors expressing GFP were tested at doses of 102–103 genome copies (GC) for AAV or plaque‐forming unit (pfu) for Ad, and the level of transduction was assessed at different time points. The data demonstrates that in MDCK and HAE cells, the highest transduction was achieved by the Ad5 vector followed by the AAV2 vector, while AAV6 had the lowest transduction. Additionally, as examined in HAE cells, the age of the culture impacts transduction efficiency. In parallel, AAV vectors 1, 2, 5, 6 (at a dose of 8×1010 – 3×1011 GC) and Ad5 vector (at a dose of 1×1011 pfu) expressing ffLuc or LacZ were intranasally delivered to mice to assess the transduction efficiency in the epithelium of both the nose and lungs. AAV6 was the most efficient at transducing the cells of the nasal epithelium of mice and AAV5, at transducing lung cells. AAV2 did not transduce airway cells in mice. In conclusion, we confirmed that the viral vectors have different transduction profiles in different models of airway epithelium. More airway models should be studied to better understand which are more useful for preclinical assessments of CF gene therapeutics.Support or Funding InformationThis project was supported by the National Heart, Lung, and Blood Institute (R25‐HL084665‐14).
Mentoring from Both Sides: How to Find, Be and Utilize a Great Mentor
Authors: Kamens Joanne 
Published: April 2020

Mentoring should not be a scary or imposing concept ‐ it's really just about getting and giving advice, support and encouragement for ongoing learning. This workshop is intended for everyone: trainees and faculty ready to learn practical tactics in identifying mentors, making the "ask" for mentoring support and how to take the best advantage of mentoring relationships‐‐from both sides.To learn more about how a strong mentoring culture benefits organizations of any size, Attend the "Re‐Imagining STEM: Who We Are and What We Do" workshop on Sunday, April 5 at 6 PM.
In Utero Exposure to Second‐Hand Cigarette Smoke Promotes Epigenetically‐Mediated Cardiac Dysfunction in High‐Fat Fed Male Mice
Authors: ronis martin , Xia Huijing , rodrigues samya lima , El Haji Eli , Noel Alexandra , Penn Arthur , Pulliam Casey , Gardner Jason , shankar kartik , Varner Kurt 
Published: April 2020

We investigated whether in utero exposure to second‐hand cigarette smoke (SHS) would adversely affect cardiac structure/function in the exposed offspring fed a high‐fat diet. Pregnant mice (C57B1/6J) were exposed to SHS or air (4 hr/d) from gestational days 6–19. The male offspring (age: 65 days) from the SHS and air exposure groups were fed either normal chow, or high‐fat liquid diet (35% fat; HFD) for 16 weeks (n=12/group). Cardiac function was measured by echocardiography at baseline (30 days) and at the end of pair feeding. Mice were then sacrificed and heart tissue collected for histological and mRNAseq analysis. There was no significant difference in body weight or heart weigh/tibial length between the in utero SHS+HFD and in utero air+HFD treated offspring. However, compared to in utero air+HFD mice, in utero SHS+HFD mice had decreased left ventricular internal diameters and volumes during diastole (3.95±0.09 vs. 4.34±0.24 mm; 67.8±1.5 vs. 85.4±4.4 μl, P<0.05) and systole (2.91±0.07 vs. 3.34±0.15 mm; 32.5±1.9 vs. 46.3±5.1 μl, P<0.05), accompanied by increases in left ventricular wall thickness during diastole (0.72±0.03 vs. 0.66±0.03 mm, P<0.01) and septal thickness during systole (1.2±0.06 vs. 0.99±0.05 mm, P<0.05) and increased fractional shortening. There was also a significant increase in collagen in the hearts of the in utero SHS+HFD mice. These data are consistent with the development of concentric cardiac hypertrophy in in utero SHS+HFD mice. mRNAseq analysis of cardiac tissue revealed 100 up‐regulated and 120 down‐regulated genes in in utero air+HFD vs in utero SHS+HFD. Top Wiki Pathway terms included genes linked to adaptive cardiac hypertrophic responses: adrenergic receptor a1a, MAP2K1, MAP2K3, CDK7, CHUK, PDPK1 and IKBKE, microRNAs in cardiomyocytes, exercise induced changes in circadian rhythm (upregulated BMAL1, downregulated Per2) and signaling via estrogens, TNF alpha and MAP kinases. GO pathways identified included cardiac muscle contraction, protein acetylation and regulation of DNA methylation. We conclude that in utero exposure to SHS produces epigenetically‐regulated differential cardiac responses following challenge with obesogenic high fat diet. (NIH P20 GM103514).Support or Funding InformationNIH P20 GM103514
Polyamine Regulation in Diabetic Breast Cancer Cells
Authors: Capellen Caleb , Ortega Jose , Dunworth Matthew , Casero Robert A. , Chandra Surabhi 
Published: April 2020

Polyamines (spermine, spermidine, and putrescine) are involved in basic cellular processes such as cell growth, replication, and transcription. They are highly regulated cations that are elevated in cancer. Upregulation of polyamines can thus develop tumors due to hyper‐proliferation of cells as observed in cancers of the colon, skin, prostate and breast. Inhibitors of polyamine pathway could prevent tumor growth as shown in animal models of colon and skin cancer. Intracellular polyamine is regulated by biosynthesis, intestinal uptake and microbial production. Thus inhibitors of synthesis alone have not been very effective as cytotoxic agents but exhibit cytostatic properties. Diabetic conditions further accelerate the growth and metastasis of cancers such as pancreas, breast, liver, and colorectal, however the role of polyamines in such states has not been investigated. We hypothesized that polyamine regulation affects proliferation of breast cancer cells and normal breast epithelial cells in diabetic conditions. Breast cancer cells (MDA‐MB‐231) and normal breast epithelial cells (MCF‐10A) were treated with normal glucose (NG, 5mM) or high glucose (HG, 25 mM) in the presence/absence of polyamine pathway inhibitors. Cells were then treated with polyamine supplements, following depletion with polyamine pathway inhibitors. There was a time dependent increase in cell proliferation with HG from 48–72 hr compared to NG. This was correlated with an increase in polyamines (putrescine, spermidine) though spermine levels were not changed considerably. Simultaneous treatment with an ornithine decarboxylase (polyamine synthesis enzyme) inhibitor, difluoromethylornithine (DFMO) could prevent cell proliferation, restore polyamines, and normalize polyamine catabolic enzymes with high glucose exposure. In conclusion, polyamine regulation is responsible for proliferation of breast cancer cells and normal breast epithelial cells in diabetic states, which can be prevented using inhibitors affecting polyamine levels in cells.Support or Funding Information
Funding: Grants from the UNK Undergraduate Research Fellows Program and UNK Research Services Council Seed Grant
Equipment: Microplate Reader (Dr. Nuxoll’s lab), Grant from the National Institute for General Medical Science (NIGMS) (5P20GM103427), a component of the National Institutes of Health (NIH)
Characterization of Novel Intron Splice Mutations in Patients with Propionic Acidemia
Authors: Smith Lorenzo Jason , Stoeger Braden Frederick , King Kaitlyn , Caporale Diane 
Published: April 2020

Propionic Acidemia (PA) is a rare genetic metabolic disorder characterized by a deficiency in propionyl CoA carboxylase (PCC), an enzyme responsible for the proper breakdown of certain proteins and lipids. As a result, patients exhibit an abnormal accumulation of toxic organic acids, which can cause detrimental health problems. PA derives from mutations in PCCA and PCCB genes, which are responsible for expressing the vital subunits of PCC. Mutations in intronic sequences may cause errors in RNA processing, specifically with RNA splicing. If intronic segments are improperly included in the final mRNA form, the resultant products become nonfunctional. The goal of this project was to determine whether novel intron splice site mutations induce improper mRNA processing and, thus, nonfunctional proteins. To achieve this, we collaborated with genetics testing company Prevention Genetics to construct minigene plasmids containing intron mutations from patients. The minigene system was transfected into mouse fibroblast cells to allow mRNA processing to occur. RNA was isolated, reverse‐transcribed into cDNA, and sequenced to determine whether the mutated introns are splicing differently from the wildtype. Initial results indicate several identified intronic mutations contribute to an altered splicing pattern in patients with PA. Knowledge of the effect of these mutations on gene expression will strengthen the genetic database to help doctors diagnose PA in their patients, as well as help develop preventive and personalized treatments.Support or Funding InformationUWSP OSCAR Travel Grant (Up to $3,000 for three students)
CD4+ T Cells from RUPP rat model activate NK cells and cause mitochondrial oxidative stress and hypertension in normal pregnant rats
Authors: Deer Evangeline , Reeve Kristin E. , Amaral Lorena M. , Vaka Venkata Ramana , Franks Michael , Fitzgerald Sara , Herrock Owen , Ibrahim Tarek , LaMarca Babbette 
Published: April 2020

Preeclampsia (PE) is new onset hypertension during pregnancy and is associated with elevated inflammatory response such as CD4+ T cells, NK cells, and cytokines. We have previously shown that women with PE exhibit increases in placental mitochondrial (mt) dysfunction and ROS compared to normal pregnant (NP) women. The Reduced Uterine Perfusion Pressure (RUPP) rat model produces many characteristics of PE such as hypertension, increases in CD4+ cells, renal and placental NK cells, and mt dysfunction/ROS. Moreover, we have shown an important role for mt ROS in hypertension in RUPPs. We have previously demonstrated that RUPP CD4+T cells cause hypertension in NP rats, however the role of RUPP CD4+ T cells in stimulating NK cells to cause mitochondrial dysfunction/ROS are unknown. Therefore, we examined the effect of adoptive transfer of RUPP CD4+ T cells to activate NK cells in NP rats and cause mt dysfunction and ROS as mechanisms of hypertension. Splenic CD4+ T cells were isolated from RUPP rats, cultured, and injected into NP rats on gestation day (GD) 13. On GD19, MAP blood and tissue samples were collected. Mt respiration and mtROS were measured in isolated mitochondria using the Oxygraph 2K and fluorescent microplate reader, respectively. A student’s t‐test was used for statistical analysis. On GD19, MAP increased to 110±2 mmHg (n=12) in RUPP CD4+ T cell recipients compared to control NP rats 101±2 mmHg (n=7, p<0.05). Circulating cytolytic NK cells increased to 3±0.6% in RUPP CD4+ T cell recipients (n=8) compared to NP controls 0.3±0.2% (n=7, p<0.05). Renal state 3 (194.3±62.33 vs 1004±251.9 pmol/sec/mg, p<0.05) and maximal (151.2±75.19 vs 531.5±99.97 pmol/sec/mg, p<0.05) respiration rates, indicative of ATP production and electron transport chain efficacy respectively, were reduced with RUPP CD4+ T cells (n=6) compared to NP (n=4). Renal mtROS increased from 251.5±81.75% in NP controls (n=4) to 416.9±47.68% in RUPP CD4+ T cell recipients (n=9, p<0.05). Collectively, the data indicate that the adoptive transfer of RUPP CD4+ T cells stimulates cytolytic NK cells and renal mitochondrial dysfunction/ROS during pregnancy as important mechanisms of hypertension in the pathophysiology of preeclampsia.Support or Funding InformationNIH Grants: RO1HD067541‐06 and P20GM121334; T32HL105324
Oxygen Deficit Is a Sensitive Measure of Mild Gas Exchange Impairment at Inspired O2 between 12.5%–21%
Authors: Liu Matthew A. , Stark Phoebe C. , Prisk G. Kim , West John B. 
Published: April 2020

The oxygen deficit (OD) is the difference between the end‐tidal alveolar PO2 and the estimated PO2 of arterial blood derived from the SpO2 that has the potential to be used clinically as a proxy for the alveolar‐arterial PO2 difference. Previous work has shown that a recently developed technology, the Alveolar Gas Meter, is accurate in measuring the efficiency of pulmonary gas exchange, via the OD, in patients with a history of lung disease as well as in normal subjects breathing a hypoxic mixture of 12.5% O2. This study sought to determine how the OD varied as the percent of inspired O2 changed from 21% to 12.5% in 36 healthy subjects split by age (young 22–31, n=23; old 42–90, n=13) to assess the possibility of use in a clinical setting. The PO2 and PCO2 were continually measured as the subject breathed through a mouthpiece using rapidly responding electrochemical cells, and the end‐tidal PO2 and PCO2 were calculated and averaged over 5 steady‐state breaths, while a fingertip pulse oximeter measured the SpO2. After correcting for any change in P50 resulting from the PCO2 differing from 40 mmHg, the arterial PO2 was calculated from the SpO2 by inverting the Hill‐equation. Subjects breathed 12.5, 15, 17.5 and 21% oxygen mixtures (balanced order) and their OD was measured in triplicate. Across all inspired O2 levels, the OD was higher in the older cohort than the young (p <0.0001, repeated measures ANOVA, Table
1). The older cohort had a larger OD at all four oxygen levels (all p<0.0001), with that difference and its variance being greater at higher inspired O2 values (age × O2 interaction p=0.002). The OD at an inspired O2 of 12.5% was significantly less than at 21% for both age cohorts. This is consistent with the known increase in ventilation‐perfusion mismatching that occurs in healthy aging. The decrease in OD magnitude as the inspired oxygen was reduced in both cohorts is consistent with the increased accuracy of the estimated arterial PO2 when based on saturation values on the steep part of the O2‐Hb dissociation curve. Further, as arterial saturation decreases there is an expected decrease in the alveolar‐arterial PO2 difference. The persisting higher OD seen in older subjects, irrespective of inspired O2, suggests that the measurement of oxygen deficit remains sensitive to mild gas exchange impairment, even when breathing air (21% O2).Support or Funding InformationFinancial support from UCSD

Measured Oxygen Deficit by Age

Young

Old

p‐value

All Inspired O2 (mean±SE)

−0.4±0.6

10.6±1.0

<0.0001

Inspired O2 by Age Interaction

0.002

12.5%

−1.4±0.7

5.7±0.6

‐

15%

−2.7±1.0

5.2±1.8

15% vs 12.5%: NS

17.5%

−2.7±1.0

9.4±1.8

17.5% vs 12.5%: NS

21%

5.1±1.3

21.9±2.1

21% vs 12.5%: <0.0001
Empagliflozin Reduces Arrhythmic Events and Improves Ca2+ Transient in Hypoxia‐induced Injury Rat Cardiomyocytes
Authors: dos Santos Danúbia Silva , Turaça Lauro Thiago , da Silva Coutinho Keyla Cristiny , Brunswick Tais Hanae Kasai , de Carvalho Antonio Carlos Campos , Girardi Adriana C. 
Published: April 2020

Sodium glucose cotransporter 2 inhibitors (SGLT2i) emerged as promising antidiabetic drugs that decreasing glucose reabsorption as well as effectively reduce heart failure hospitalization and cardiovascular death in type 2 diabetes patients. Recently, it has been shown that the SGLT2i also exert beneficial cardiovascular effects in non‐diabetic heart failure patients. However, the cardioprotective mechanisms of these drugs remain unknow. The present study aimed to test the hypothesis that the cardioprotective effects of SGLT2i empagliflozin (EMPA) may be associated with changes of cardiac electrical activity and Ca2+ homeostasis in hypoxia‐induced injury ventricular myocytes. Cardiomyocytes isolated from neonatal rats submitted to chemical hypoxia (200 μM CoCl2) or normoxia were incubated with EMPA (1 μM) or vehicle during 24 hours. Electrophysiological recordings were obtained using multielectrode array (MEA) system and intracellular microelectrode. Cytoplasmic calcium transients were investigated via fura‐2 fluorescence. Compared with vehicle, MEA analysis showed a significant decrease in field potential duration in EMPA‐group in both conditions [hypoxia: 83 ± 4 (N=19) vs. 37 ± 3 (N=44); normoxia: 213 ± 5 (N=53) vs. 62 ± 4 (N=36) ms, p<0.001]. EMPA reduced action potential duration at 30%, 50% and 90% repolarization (APD30, APD50 and APD90, respectively) in hypoxia [APD30: 109 ± 2 (N=41) vs. 98 ± 1 (N=30); APD50: 150 ± 3 (N=41) vs. 134 ± 1 (N=30); APD90: 292 ± 4 (N=41) vs. 271 ± 5 (N=30) ms, p<0.05] and normoxia [APD30: 94 ± 2 (N=30) vs. 81 ± 1 (N=39); APD50: 129 ± 2 (N=30) vs. 111 ± 1 (N=39); APD90: 259 ± 4 (N=30) vs. 244 ± 4 (N=39) ms, p<0.05]. No differences were observed in amplitude and resting membrane potential. Action potential recordings also revealed that the percentage of arrhythmic cells was lower in EMPA‐group in both conditions [hypoxia: 19 (N=41) vs. 10 (N=30) %; normoxia: 7 (N=30) vs. 3 (N=39) %]. In addition, EMPA significantly increased in Ca2+ transient amplitude in hypoxia [F/F0: 0.16 ± 0.01 (N=21) vs. 0.21 ± 0.01 (N=19), p<0.01] and normoxia [F/F0: 0.17 ± 0.01 (N=19) vs. 0.23 ± 0.01 (N=20) μM, p<0.01]. These results suggest that EMPA modulates electrical activity, reduces arrhythmic events and improves Ca2+ transient, which may contribute to cardioprotective effects of SGLT2i.Support or Funding InformationFAPESP and CNPq
17β‐Estradiol Induces Temozolomide Resistance through Redox Optimization in Glioblastoma
Authors: Lin Hong-Yi , Hsu Tsung-I , Ko Chiung-Yuan 
Published: April 2020

Glioblastoma is the most fatal cancer of all brain tumors, and the standard treatment of glioblastoma patients is surgical tumor resection followed by radiotherapy and temozolomide (TMZ)‐mediated chemotherapy. According to the TCGA databases, female patients exhibit poorer prognosis compared with male patients, suggesting that the gender‐specific hormone plays an important role in glioblastoma malignancy. In addition to acting as a female sex hormone, 17β‐estradiol is an important neuro‐steroid for brain health, including learning and memory. Numerous studies indicate that 17β‐estradiol aggressively participates in the pathogenesis of endocrine‐related cancers, such as breast and prostate cancers. However, the role of 17β‐estradiol is studied poorly in glioblastoma which exhibits strong neurosteroidogenesis. In this study, we attempt to elucidate whether 17β‐estradiol is involved in acquired resistance of glioblastoma to TMZ. Herein, we found that the level of 17β‐estradiol is significantly increased in TMZ‐resistant glioblastoma. Moreover, 17β‐estradiol attenuated TMZ‐induced cell death in A172 and U87MG cells. To dissect mechanisms underlying 17β‐estradiol‐induced resistance, we investigated whether 17β‐estradiol affected TMZ‐induced DNA damage and ROS production. Particularly, 17β‐estradiol significantly attenuated TMZ‐induced ROS production and hydrogen peroxide accumulation without affecting DNA damage. Further, 17β‐estradiol obviously prevented TMZ‐induced mitochondria dysfunctions characterized by the elevation of mitochondrial ROS and the reduction of ATP production. In addition, 17β‐estradiol treatment increased expression of multiple anti‐oxidant proteins, including superoxide dismutase (SOD) 1, SOD2, catalase (CAT), glutathione reductase, and nuclear factor erythroid 2 like 2. In particular, 17β‐estradiol promoted degradation of hydrogen peroxide through activating CAT, not through glutathione‐ and redox‐in‐mediated catabolic pathways. Based on these findings, we conclude that 17β‐estradiol counteracts TMZ‐induced oxidative stress through enhancing mitochondrial activity and CAT, leading to TMZ resistance.Support or Funding InformationTsung‐I Hsu