bluuebunny/crossref_metadata_2025 huggingface dataset:
Stress‐induced premature senescence as a mechanism of nucleoside reverse transcriptase inhibitor‐induced endothelial dysfunction (547.18)
Authors: Glover Mitzi , Hebert Valeria , Xue Stephen , Robinson Corie , Crooks Samuel , Anderson Joi , Slaybaugh Kate , Dugas Tammy
Published: June 2021

Nucleoside reverse transcriptase inhibitors (NRTIs) are a major component of the current combination therapy for HIV, known as highly active antiretroviral therapy (HAART). Studies have demonstrated that HAART has significantly decreased mortality and reduced viral load in HIV‐infected patients, but has also been associated with cardiovascular complications including endothelial dysfunction, an early marker of atherosclerosis. Our studies have shown that mice treated chronically with some NRTIs exhibit endothelial dysfunction, which is prevented by overexpression of the mitochondrial antioxidant manganese superoxide dismutase (MnSOD). Moreover, our in vitro data indicates that mitochondrial toxicity via oxidative stress may result in stress‐induced premature senescence (SIPS), a possible mechanism for the development of this dysfunction. In this study, human aortic endothelial cells (HAECs) were treated with NRTIs zidovudine (AZT), lamivudine (3TC), or tenofovir (TEN), through multiple passages to determine whether the drugs induce premature senescence. Cells were also cotreated with coenzyme Q10, a critical cofactor for mitochondrial electron transport, to determine whether a reduction in oxidative stress could prevent premature senescence. Senescence was assessed using population doubling, senescence‐associated β galactosidase staining, and measurement of cell cycle associated proteins, p16 and p21. Our findings indicate that oxidative stress induced by NRTIs enhances premature senescence in endothelial cells, and that the use of the antioxidant coenzyme Q10 attenuates this response.
Risk factors and characteristics of lesions of the cardiovascular system in young aged patients with psoriasis (547.2)
Authors: Batpenova Gulnar , Kazieva Aliya , Pshembaeva Saule , Sadykova Gulnaz
Published: June 2021

To study lipid metabolism,as well as the identification of risk factors,the characteristics of the cardiovascular system lesions in young aged patients with psoriasis.

It was found in clinical and epidemiological study that among patients with psoriasis most men were ‐ 61.1%, women ‐ 38.9%. Mean age ‐ 46,4±2,3years. In the study of the spectrum the concomitant somatic pathology in adult patients 61.3% was identified with underlying diseases. In patients with severe psoriasis concomitant diseases were observed in 78.4% of patients, that is 1.3 times higher than in patients with psoriasis vulgaris and exudative. Lipid profile were as follows: hypercholesterolemia was diagnosed in 39.5% patients,74.4% patients, LDL cholesterol was above normal. Values of triglycerides (TG) averaged 1,82±0,15 mmol/l. In carrying out the cluster analysis between lipid profile and the main clinical and laboratory parameters in patients with psoriasis, revealed strong correlation rates (p<0.001) HDL with disease duration (r=‐0,95),with the affected area (r=‐0,83).

In a study of the cardiovascular system in patients with psoriasis most often ‐ in 70% of cases met various cardiac arrhythmias. Signs of connective tissue dysplasia, namely, small malformations of the heart were detected in 25.6% patients. Hypertension was diagnosed in 9.3%, signs of peripheral atherosclerosis‐6.9% of the patients, ischemic heart disease‐4.7%.
Insulin suppresses ischemia/reperfusion‐induced myocardial injury via inhibiting mitochondrial JNK translocation (547.20)
Authors: Xu Jie , Qin Xinghua , Zhang Xing , Yang Lu , Xing Yuan , Gao Feng
Published: June 2021

Objective: Insulin exerts cardioprotective effect against ischemia/reperfusion (I/R) injury, but the underlying mechanism is unclear. This study aimed to examine the role of mitochondria in insulin‐induced cardioprotection against myocardial I/R injury.

Methods: Adult male rats were subjected to 30 min of myocardial ischemia and 2 h of reperfusion. Rats randomly received vehicle, insulin, and Tat‐SabKIM1 administered 5 min before reperfusion. Incidence of myocardial apoptosis was determined by TUNEL. JNK from the extractions of mitochondria, cytosol/nucleus, and whole cell was detected by Western blot.

Results: Myocardial I/R produced significant myocardial apoptosis, both mitochondrial and cytosolic/nuclear JNK activation and myocardial infarction (n=6, P<0.05). Importantly, we found JNK translocation from mitochondria to cytosol/nucleus together with cytochrome c release following myocardial I/R, which were blunted by insulin treatment. Inhibiting JNK translocation to mitochondria with Tat‐SabKIM1, a retro‐inverso peptide which disrupting the JNK interaction with mitochondria, not only decreased mitochondrial JNK activation but also exerted cardioprotective effect as evidenced by reduced myocardial apoptosis (10.2±1.3% vs. 16.0±1.2%, P<0.05) and myocardial infarct size (15.9±2.1% vs. 23.7±1.5%, P<0.05) after myocardial I/R.

Conclusion: These data show that insulin exerts mitochondrial protective action via disrupting JNK translocation to mitochondria and thus inhibiting cytochrome c release, which contributes to insulin‐elicited cardioprotection against I/R injury.

Key words: insulin, ischemia/reperfusion injury, JNK signaling, mitochondria, cardioprotection

Grant Funding Source: supported by grants from the National Basic Research Program of China (No. 2013CB531204)
Heart rate variability and cardiac function in heart failure rats treated with allogeneic mesenchymal stem cells (547.21)
Authors: Morais Sharon , Lataro Renata , Silva Carlos Alberto , Oliveira Luciano , Carvalho Eduardo , Simões Marcus Vinicius , Meirelles Lindolfo , da Silva Luiz Eduardo , Salgado Hélio , Fazan Rubens
Published: June 2021

The effect of Mesenchymal Stem Cells (MSC) on cardiac function and non‐linear dynamics of heart rate variability was evaluated after myocardial infarction (MI) in rats. MSC (N=7) or saline (N=11) were given iv to male Wistar rats 1 week after sham (N=9) or left coronary artery ligation. Infarct size, assessed by SPECT, was found similar among rats (51±1%). Cardiac function was assessed by isotopic ventriculography before and 4 weeks after MSC therapy. 5 weeks after MI the rats underwent ECG and arterial pressure recording. The infarct size was reduced by MSC (36±2% vs 48±2% saline). Left ventricular ejection fraction (LVEF) was normal in sham but lower after MI (54±3 vs 20±2%). No effect of MSC was seen in LVEF (20±4%). Interstitial collagen was increased in rats with MI (15±2% vs 10±1%), but not in MSC treated ones (11±1%). Arterial pressure and heart rate were similar among groups in conscious rats. Baroreflex sensitivity was reduced after MI (1.80±2 vs 1.10±0.1 bmp/mmHg), while MSC prevented this outcome (1.74±0.2 bpm/mmHg). QT and QTc intervals were lengthened by MI and MSC did not affect this parameter. A reduction in sample entropy (SampEn) and increase in detrended fluctuation analysis (DFA) of RR interval were observed after MI. MSC therapy did not affect SampEn but protected the DFA. In conclusion, MSC reduced MI and interstitial fibrosis in left ventricle, combined with improvement of baroreflex sensitivity and DFA.

Grant Funding Source: Financial support: FAPESP, CAPES, CNPq, NIC‐USP
A viral peptide deformylase‐ribosome complex reveals mechanism of host gene expression control (558.2)
Authors: Giglione Carmela , Grzela Renata , Bron Patrick , Lai Kee Him Joséphine , Fieulaine Sonia , Nusbaum Julien , Pozza Alexandre , Bienvenut Willy , Meinnel Thierry
Published: June 2021

All prokaryotic nascent chains must be co‐translationally deformylated to allow the further essential removal of the first methionine. This early nascent chain processing is ensured by peptide deformylases (PDFs), an enzyme shown to interact with ribosome at the exit tunnel. Recently, studies demonstrating in one hand the acquisition by different marine phages of modified version of bacterial PDF and in the other hand that also viral proteins undergo deformylation led to propose that phage PDFs could be important for viral fitness. Aside from this intriguing hypothesis, not much is known about these phage PDFs. Here, we show that most viral versions of the PDFs do not contain a C‐terminal extension previously proposed to be responsible of direct PDF binding to ribosome. Nonetheless, we show that the smallest identified phage PDF, a fully C‐terminally‐truncated PDF, is still able to bind directly to ribosome via a new dedicated ribosome‐binding domain, playing a master regulator function. Crosslinking experiments and combination of structural data of this phage PDF alone and the complex between the phage PDF and the ribosome show that the phage PDF interacts with ribosome in several distinct regions. Taking together, our findings provide basis for understanding of a previously unrecognized mechanism based on the pathogenic virus control of host viability at the level of the tunnel exit.

Grant Funding Source: CNRS, ANR
Mechanism of HigB‐mediated ribosome‐dependent mRNA degradation (568.1)
Authors: Schureck Marc , Maehigashi Tatsuya , Marquez Jhomar , Dunkle Jack , Miles Stacey , Dunham Christine
Published: June 2021

Proteus vulgaris (P. vulgaris) is a major cause of bacterial urinary tract infections. P. vulgaris grows on medical equipment, such as catheters, in an antibiotic‐resistant state known as biofilms. In biofilms, bacteria are highly adhesive and are protected from antibiotics by a self‐produced extracellular matrix of DNA, protein and polysaccharides. My goal is to elucidate the molecular mechanism of the ribosome‐dependent RNase family of toxins, which play an important, but unknown, role in biofilm formation. Host inhibition of growth B (HigB) is a member of this family and was first identified in P. vulgaris. HigB cleaves ribosome‐bound A‐site mRNA and in contrast to most toxins, HigB recognizes mRNA in a codon‐independent manner. Using microbial, structural and biochemical techniques, we are beginning to understand how HigB can recognize such a large variety of A‐site codons. Our X‐ray crystal structure of HigB in complex with the Thermus thermophilus 70S ribosome and an A‐site codon chemically modified to prevent cleavage reveals that HigB binds the A site and dramatically reorients mRNA for catalysis by two essential HigB amino acids. Biochemically, we have shown that the minimal substrate specificity for HigB is an adenosine in the A‐site wobble position. Taken together, our results indicate that the HigB toxin acts as a novel translation factor and site specifically recognizes the wobble position.

Grant Funding Source: Supported by a NIH NRSA (GM108351) and BCMB graduate training grant (5T32GM8367)
Postnatal cardiac development in transgenic mice with altered expression of the DNA glycosylase Ogg1 (573.4)
Authors: Ruchko Mykhaylo , Gorodnya Olena , Rachek Lyudmila , Pastukh Viktor , Gillespie Mark
Published: June 2021

A switch from cardiomyocyte hyperplasia to hypertrophy underlies postnatal cardiac development in rodents. Interestingly, this transition is associated with robust mitochondrial biogenesis, which, paradoxically, is accompanied by oxidative mitochondrial (mt) DNA damage and repair (Pohjoismaki et al, NAR, 2012). Whether mtDNA damage and repair are functionally linked to mitochondrial biogenesis is unknown, but previous experiments showed that controlled DNA damage and repair are important for transcription of nuclear genes (Al‐Mehdi et al, Sci Signaling, 2012). To determine if a similar mechanism could be operative in the mitochondrial genome during postnatal cardiac development, we measured oxidative damage in the mtDNA D‐loop transcriptional regulatory sequence and the coding region as a function of time in hearts derived from wild type (WT) mice, knock‐out mice deficient in the DNA glycosylase Ogg1 (KO), and in KO mice transgenically over‐expressing human OGG1 targeted to mitochondria. In comparison to WT animals, KO mice displayed higher mtDNA damage which preceded an exaggerated postnatal increase in heart‐to‐body mass ratio. Conversely, KO mice over‐expressing mitochondria‐targeted OGG1 displayed decreased mtDNA oxidation and a blunted elevation in cardiac mass relative to KO mice. These results suggest mtDNA damage and repair may be functionally linked to postnatal cardiac development.

Grant Funding Source: Supported by the NIH R21HL102789, R01HL113614
Diverse topological requirements for mitochondrial transcription (573.5)
Authors: Sondheimer Neal , Zollo Ornella
Published: June 2021

That transcription of the three human mitochondrial promoters can be recapitulated in vitro using a limited set of proteins including the mitochondrial RNA polymerase (POLRMT) and two accessory factors, mitochondrial transcription factor A (TFAM) and transcription factor B2 (TFB2M). The complete set of mechanisms that regulate promoter selection are unknown. The objective of this study was to determine whether the topological state of template DNA plays a role in promoter selection. We used a plasmid‐based system for the in vitro analysis of transcription that recapitulates features of the mtDNA to illustrate the relationship between topology and the three mitochondrial promoters and the involvement of TFAM which can bend template DNA. Both heavy‐strand promoters (HSP1 and HSP2) can be simultaneously transcribed from a circular template but increasing TFAM concentrations suppress HSP2 transcription at concentrations that result in HSP1 activation. We identified a requirement for the use of supercoiled DNA at HSP2 for transcription from circular templates which is not present for HSP1 and LSP. We conclude that the mitochondrial promoters have distinct requirements for DNA topology and TFAM dosage in vitro providing a means for the differential regulation of mitochondrial gene expression.

Grant Funding Source: NIH/NICHD
Energy balance induced by fluoxetine: possible involvement of body heat expenditure (575.1)
Authors: da Silva Aline Isabel , Pedroza Anderson Apolonio , Freitas Cristiane , Braz Glauber Ruda , Ferreira Diorginis , Lagranha Claudia , Manhaes‐de‐Castro Raul
Published: June 2021

Innumerous studies have shown that modulation of serotonin system can contributes to modulation of body weight. It is well known that the control of body weight is dependent of calories intake and energy expenditure and the major tissues related to energy expenditure are brown fat and skeletal muscle. During the metabolic process related to energy balance occurs formation of reactive oxygen species‐ROS. With these our aim was evaluate the effect fluoxetine‐FX on body weight, food intake, voluntary activity, body temperature, malondialdehyde production and superoxide dismutase‐SOD, catalase‐CAT and glutathione S‐transferase‐GST activity in brown adipocytes‐BA and extensor digitorum longus‐EDL. The treatment was conducted during the lactation time using 10mg/Kg‐B.W in male Wistar rats. We observe that FX treatment doesn’t induce alteration in food intake and voluntary activity, although induce decrease in body weight and lower heat loss after challenge with cold environment. Related to ROS, in EDL we observe increase in MDA production, decrease in GST activity and no differences in SOD and CAT; in BA we observe also an increase in MDA production and no difference in antioxidant enzymes. Our data suggest that modulation of serotonin levels by fluoxetine promotes decrease in body weight likely due to alteration in body heat expenditure and modulation of cellular ROS detoxification.
Adipose‐tissue specific deletion of TGF‐β receptor 1 kinase protects mice from diet‐induced obesity and diabetes (577.4)
Authors: Wankhade Umesh , Wiese Mohmmad , Kulkarni Ashok , Rane Sushil
Published: June 2021

The adipose tissue depot is one of main organs targeted during pathogenesis of diabetes and obesity. We previously showed that the TGF‐β/Smad3 signaling pathway regulates the appearance of brown adipocyte features in classical white adipose tissue (WAT). Global Smad3 knockout mice exhibit a phenotypic transition and functional transformation of WAT to BAT‐like features that provide protection to the mice from obesity and diabetes (Yadav et al 2011, Cell Metabolism). We also observed increased circulating levels of TGF‐β ligand in obese mice and human obese subjects suggesting an involvement of TGF‐ β receptors in the WAT‐BAT transition and regulation of glucose metabolism. To examine this possibility, we have generated an adipose tissue specific TGF‐β receptor 1 kinase knockout mouse model (TβRI‐KO). Epididymal WAT (EWAT) from TβRI‐KO mice showed increased BAT mRNA transcripts. Further, stromal vascular fractions (SVF) and pure adipocyte fractions from T[[Unsupported Character ‐ Symbol Font □]]RI‐KO mice showed increased levels of BAT progenitor markers. High fat diet (HFD) fed TβR1‐KO mice gain less weight, exhibit leaner body composition and improved glucose tolerance. EWAT and anterior subcutaneous WAT (AWAT) depots show smaller sized adipocytes with UCP‐1 stained multi‐locular cells interspersed in the WAT that resemble brown adipocytes. Consistent with this, EWAT from TβR1‐KO shows elevated BAT gene transcripts and significantly increased expression of BAT progenitor markers. Together, these results suggest that the TGF‐β receptor 1 kinase plays a vital role in adipose tissue biology by promoting its morphological and functional transformation to a metabolically beneficial brown fat like tissue with protective functions against obesity and diabetes.

Grant Funding Source: Supported by NIDDK/NIH
In vitro analysis of novel inhibitors for Escherichia coli MurA (582.4)
Authors: Haile January , Frost Andrea , Boone Benjamin , Song Vanessa , Muzyka Jennifer
Published: June 2021

The widespread of use of antibiotics to treat bacterial infection has selected for antibiotic resistant strains of bacteria. Fosfomycin is an example of a broad spectrum antibiotic that has become less effective because of increased bacterial resistance. Fosfomycin inhibits synthesis of peptidoglycan, a structural component of the bacterial cell wall. Fosfomycin covalently binds to the phosphoenolpyruvate binding site of the enzyme MurA, an essential enzyme in the peptidoglycan synthesis pathway. MurA catalyzes the addition of the enolpyruvate to UDP‐N‐acetylglucosamine. The additional product, an inorganic phosphate, is detected by the indicator, malachite green. Muzyka, et al. has identified putative inhibitors through DOCK and AMBER studies. IC50 values have been determined for BCB33b and VS5 of these molecules. Additional molecules with similar functional groups are being synthesized and evaluated as putative inhibitors. The site of inhibitor binding will be determined with in silico studies and site‐directed mutagenesis.

Grant Funding Source: Centre College Faculty Development Fund and the Alcock Interdisciplinary Fund
The effects of oral administration of bile salt hydrolase on the partition of fatty acid and glucose oxidation in Zucker fatty rats (585.8)
Authors: Spratt Elisabeth , Mo Yiming , Li Yang , Lin Jun , Chen Guoxun
Published: June 2021

Conjugated bile salts take part in micellar formation, a critical step for dietary lipid digestion and absorption. Bile salt hydrolases (BSHs) catalyze the conversion of conjugated bile salts to bile acids and amino acids. Gut bacteria produce this family of enzymes. BSHs could potentially reduce the efficiency of lipid digestion and absorption if applied to the upper portion of the small intestine. Thus, we hypothesize that BSH is a novel lead for therapeutic reduction of lipid absorption needed to control obesity in humans. To test this, recombinant BSH (rBSH) was expressed and purified from an E. coli expression system by Ni‐NTA affinity chromatography. The rBSH or bovine serum albumin (BSA) control (2.7mg/kg of body weight) was gavaged daily into Zucker fatty (ZF) rats (9‐12 weeks old, 4 per group) just before the start of the dark photoperiod (before active feeding starts) for 7 days. After the last treatment, rats were housed individually in Comprehensive Laboratory Animal Monitoring Systems for 24 hours to collect the indirect calorimetry data. The rats treated with rBSH had significantly higher respiratory exchange ratios (RER) during the dark (p=0.028), but not the light photoperiod. The heat production and change in body weight were not different between groups. The higher RER data of ZF rats treated with rBSH during the dark photoperiod (feeding time) indicate a reduction of energy derived from fat oxidation, suggesting that lipid digestion or absorption may be reduced, which leads to higher utilization of glucose. Together, the findings shown in this pilot study suggest BSH is a lead for novel medication for modulating lipid digestion and absorption.
A genomic approach to microbial source tracking of fecal indicator bacteria in recreational surface waters (589.2)
Authors: Pikaart Michael , Pratt Cameron , Skilling Stephen , Best Aaron
Published: June 2021

We present results of a multi‐year study to identify sources of chronic high fecal indicator bacterial levels in the Lake Macatawa (Ottawa County, Michigan) watershed. In this work, we have analyzed community bacterial DNA from environmental samples to characterize total microbial community populations using traditional qPCR and next generation DNA sequencing and analysis techniques. Standard microbiological and 16S rRNA qPCR source tracking techniques have revealed widespread and possibly endogenous nonpoint sources, as well as occasional point source hotspots. These results are compared to microbial community source tracking is based on DNA information in the same marker molecule, 16S rRNA, with the community source tracking approach allowing for the monitoring all microbial groups in the same samples. The baseline information obtained on microbial communities in the watershed has facilitated development of unique microbial profiles for possible point sources and perform qPCR based microbial source tracking that is robust to false positives and false negatives. Results of suites of newly described indicator bacteria that reflect various environmental events and their associate health and environmental risks will be discussed.
Isolation and genomic annotation of two novel bacteriophage known to infect Mycobacterium smegmatis (589.5)
Authors: Paschalis Valerie , Patel Shivani , Aziz Froogh , Chen Alston , Monsen Kirsten , Adams Sandra , Vega Quinn
Published: June 2021

Tuberculosis (TB), a disease caused by Mycobacterium tuberculosis, kills over 1 million people annually worldwide. With the development of multiple drug resistant strains of TB, interest has increased in the development of alternative methods for controlling this disease. As part of the HHMI‐SEA PHAGES project, bacteriophages capable of infecting Mycobacterium smegmatis, an organism in the same genus as M. tuberculosis, were isolated from soil samples in New Jersey. The phages were characterized by studying plaque morphology and by the use of Transmission Electron Microscopy. In addition, novel mycobacteriophages ShiVal and Alsfro were isolated and sequenced. The phage genomes were then annotated using DNA Master, BLAST, Glimmer, and GeneMark. Lastly, the information obtained was published in PhagesDB and GenBank. ShiVal is a B cluster, B1 subcluster phage with a genome size of 68K, 101 putative genes and 66.5% GC content. Alsfro is an A cluster, A1 subcluster phage with a genome size of 52K, 101 putative genes and a 63.3% GC content. Discovering and cataloging these novel phages is significant because a database of novel phages will be available to be tested for activity against M. tuberculosis.

Grant Funding Source: HHMI SEA‐PHAGE
Epigenetic regulation of OPA1 sensitizes hepatocellular carcinoma to sorafenib‐induced apoptosis (59.1)
Authors: Puszyk William , Dong Hui‐Jia , Zhao Frank , Shisgal Preston , Cabrera Roniel , Neslon David , Liu Chen
Published: June 2021

The aim of this study is to identify epigenetic markers of DNA methylation for the Optic Atrophy gene OPA1, which is an indication of Sorafenib sensitivity in HCC. Sorafenib has been shown to act as a multi‐kinase inhibitor in other cancers, and is used in the treatment of un‐resectable hepatocellular carcinoma (HCC). Patient response to this therapy is variable. We have previously shown that treatment with Sorafenib does not inhibit the AKT pathway in hepatocellular carcinoma cells and instead induces rapid mitochondrial fragmentation, which is associated with the deregulation of mitochondria‐fusion‐related protein OPA1, thus inducing apoptosis. Exposure of HCC cells or isolated mitochondria to Sorafenib induces cytochrome c release. Our data indicate that siRNA‐mediated OPA1 knockdown significantly sensitizes HCC cells to Sorafenib‐induced apoptosis. Furthermore, Sorafenib inhibits HCC xenograft tumor growth in vivo, and does not induce apoptosis in healthy primary hepatocytes. Murine xenograft tumor tissue samples revealed mitochondria fusion protein OPA1 expression levels are strongly associated with the sensitivity of HCC to Sorafenib treatment. We have shown that Sorafenib suppresses the tumorigenesis of HCC through the induction of mitochondrial injury via OPA1. We now show that epigenetic analysis of the OPA1 gene promoter indicates a role for DNA methylation in the suppression of OPA1 expression, and a potential biomarker which can be utilized for the identification of patients whom may benefit from treatment with Sorafenib. Our results provide new insights into the pathogenesis of HCC and suggest that OPA1 is a novel therapeutic target in patients with HCC.

Grant Funding Source: NIH
XIAP:p19/p12‐casp7 complex serves as a target for selectively killing malignancies with caspase‐3 downregulation (59.2)
Authors: Lin Yuan‐Feng , Lai Tsung‐Ching , Jan Yi‐Hua , Tian Wei‐Ting , Hsiao Michael , Liang Po‐Huang
Published: June 2021

Caspase‐3 down‐regulation (CASP3/DR) in tumors frequently confers resistance to cancer therapy and is significantly correlated with a poor prognosis in cancer patients. Because CASP3/DR cancer cells rely heavily on the activity of caspase‐7, an executioner caspase that is similar to caspase‐3, to initiate apoptosis, the inhibition of activated caspase‐7 (p19/p12‐casp7) by XIAP is considered a possible mechanism for the prevention of apoptosis in those cancer cells. Here, we identify the pocket surrounding the Cys246 residue of p19/p12‐casp7 as a target for the development of a protein‐protein interaction (PPI) inhibitor of the XIAP:p19/p12‐casp7 complex. We demonstrate that interrupting this PPI directly triggers caspase‐7‐dependent apoptotic signaling that bypasses the activation of the apical caspases and selectively kills CASP3/DR malignancies in vitro and in vivo without adverse side effects in non‐tumor cells. Importantly, CASP3/DR combined with p19/p12‐casp7 accumulation correlates with the aggressive evolution of clinical malignancies and a poor prognosis in cancer patients. Moreover, the targeting of this PPI effectively kills cancer cells with multidrug resistance due to miR‐let‐7a1‐mediated CASP3/DR and re‐sensitizes cancer cells to chemotherapy‐induced apoptosis. These findings not only provide an opportunity to treat CASP3/DR malignancies by targeting the XIAP:p19/p12‐casp7 complex but also elucidate the molecular mechanism underlying CASP3/DR in cancers.
The amyloid precursor protein of Alzheimer’s disease associates with axoplasmic organelles (596.10)
Authors: Stevenson James , Moran Sean , Cyr Jennifer , DeGiorgis Joseph
Published: June 2021

The Amyloid Precursor Protein (APP) is a causative agent in Alzheimer’s disease (AD) as the Aβ fragment of this protein builds up in brain lesions of AD patients and mutations in APP lead to heritable forms of the disorder. APP contains a single transmembrane domain and a highly conserved region at the tip of the C‐terminus. Here, we show that APP associates with isolated axoplasmic organelles purified from the axoplasm of the squid giant axon. We have also identified the coding sequence of squid APP from a cDNA library of the North Atlantic Long‐finned Squid, Loligo pealei. We have cloned the full‐length coding sequence and are currently over‐expressing and purifying recombinant squid APP to establish a standard curve for quantitative Western blot to determine the concentration of APP in whole axoplasm and in isolated organelle fractions. Squid provide the only source of pure axoplasm as this organism contains a large axon up to 1 mm in diameter, allowing for the extrusion of cytoplasm away from the cell body, cell membrane, and other types of neuronal tissues. Quantifying the amount of APP in axoplasm and the number of APP molecules per organelle may provide insight into the wild‐type function of APP.

Grant Funding Source: Supported by the Rhode Island INBRE Program NIMGS grant 5 P20 GM103430‐13
The highly conserved C‐terminus and transmembrane domain of the amyloid precursor protein of Alzheimer’s disease is found in highly divergent species (596.11)
Authors: O'Donnell Erin , Granato Juliana , DeGiorgis Joseph
Published: June 2021

The Amyloid Precursor Protein is a causal agent in the pathogenesis of Alzheimer’s disease. A fragment of APP termed A‐beta builds up in the brain lesions of AD patients and mutations in the APP gene have been show to cause familial forms of the disorder. APP exhibits a single transmembrane domain that spans the lipid bilayer of membrane bound vesicles and a C‐terminal that contains a highly conserved sequence that extends into the neuronal cytoplasm. Here, we carried out a blast analysis of the highly conserved C‐terminal and then assayed hits for the presence of a transmembrane domain. Although, in some cases species have lost the homologous region of APP that causes AD (A‐bata) in humans, the C‐terminal and transmembrane domain are found in a wide array of organisms. Currently, we are using antibodies against the C‐terminal fragment to assay the presence and distribution of APP in distantly related species at the confocal and EM levels.

Grant Funding Source: Supported by the Rhode Island INBRE Program NIMGS grant 5 P20 GM103430‐13
A new function for an old enzyme: Src protein tyrosine kinase directs excitotoxic neuronal death in stroke (596.3)
Authors: Cheng Heung‐Chin , Hossain Mohammed , Roulston Carli , Ng Dominic , Williamson Nicholas , Hoque Ashfaqul , Hill Andrew
Published: June 2021

Excitotoxicity resulting from over‐stimulation of ionotropic glutamate receptors is a major cause of neuronal death in cerebral ischemic stroke. The over‐stimulated receptors exert their neurotoxic effects in part by over‐activation of calpains which induce neuronal death by catalyzing limited proteolysis of specific cellular proteins. Here, we report that in cultured cortical neurons and in vivo in a rat model of focal ischemic stroke, the tyrosine kinase Src is cleaved by calpains at a site in the N‐terminal unique domain. This generates a truncated Src fragment of ~52 kDa localised predominantly in the cytosol. A synthetic cell membrane‐permeable fusion peptide derived from the Src unique domain prevents calpains from cleaving Src in neurons and protects against excitotoxic neuronal death. To explore the role of the truncated Src fragment in neuronal death, we expressed a recombinant truncated Src fragment in cultured neurons, and demonstrated that expression of this fragment, which lacks the unique domain, was sufficient to induce neuronal death. Furthermore, inactivation of the pro‐survival kinase Akt is a key step in its neurotoxic signaling pathway. As Src maintains neuronal survival, our results implicate calpain cleavage as a key step of neuronal death in excitotoxicity, and suggest blockade of calpain cleavage of Src as a potential therapeutic strategy to minimize brain damage in ischemic stroke.

Grant Funding Source: NHMRC Australia
Retrieval trafficking plays an important role in homeostasis of hERG expression at the plasma membrane (604.2)
Authors: Chen Jeffery , Guo Jun , Li Wentao , Yang Tonghua , Zhang Shetuan
Published: June 2021

The human ether‐a‐go‐go‐related gene (hERG) encodes the α‐subunit of the rapidly activating delayed rectifier potassium channel (IKr). A reduction in hERG expression on the plasma membrane causes long QT syndrome, predisposing affected individuals to high risk of cardiac arrhythmias and sudden death. We previously reported that plasmalemmal hERG channels are vigorously internalized under certain conditions such as hypokalemia. Whether internalized channels can recycle back to the membrane remains unknown. In this study, using Western blot and patch clamp techniques, we observed that an enhanced recovery rate of IKr is associated with an increased endocytosis of hERG channels after exposure of cells to hypokalemic conditions. This observation is not due to increased synthesis since hERG mRNA expression was not increased by hypokalemic treatment and the enhanced recovery was not affected by the inhibition of protein synthesis using cycloheximide. Using fluorescent labeling, we directly monitored the retrieval trafficking of internalized hERG channels to the plasma membrane. Co‐immunoprecipitation experiments reveal that hERG channels are associated with the GTPase Rab11, but not Rab4. Interference with Rab11 function but not Rab4 led to decreased hERG expression and function. Our data provide novel insights into the recycling mechanisms that regulate the homeostasis of membrane‐bound hERG channels.

Supported by the Canadian Institutes of Health Research ‐ CIHR
Role of brain stearoyl‐CoA desaturase‐1 in metabolism, obesity, and glucose homeostasis (605.2)
Authors: Harrington Kristin , Burhans Maggie , Ntambi James
Published: June 2021

The high prevalence of obesity, coupled with its potential to develop into serious health issues including type II diabetes, insulin resistance and coronary artery disease, reveals it is a significant health problem. To investigate how excess adipose tissue manifests into these health issues, we have observed the effects of a disruption in stearoyl‐CoA desaturase 1 (SCD1), which is involved in the synthesis of endogenous monounsaturated fatty acids from saturated fatty acids. We have previously demonstrated that whole‐body deficiency of SCD1 in mice protects from high‐fat diet‐induced obesity, hepatic steatosis and insulin resistance. However, the tissue‐specific contribution of SCD1 to these phenotypes remains to be fully elucidated. In the current study, we investigated the role of brain SCD1 in mediating the metabolic consequences of a high‐fat diet (HFD). We fed nestin‐cre+;SCD1 flox/flox brain‐specific SCD1 knockout (BKO) mice a HFD for 18 weeks. We observed that both male and female BKO mice weighed significantly less than Lox controls. In addition, male BKO mice had lower hepatic triglyceride levels than Lox mice, and enhanced glucose tolerance relative to Lox mice. Overall, these results indicate brain SCD1 plays a role in HFD‐induced adiposity and glucose homeostasis.
Inactivation of heparan sulfate 2‐O‐sulfotransferase reduce neutrophil bactericidal activity and endothelial integrity to exacerbate host survival upon Group B Streptococcus infection (607.14)
Authors: Chang Yung‐Chi , Xu Ding , van Wijk Xander , Esko Jeffrey , Nizet Victor
Published: June 2021

Neutrophil recruitment and extravasation at sites of inflammation together with the barrier functions provides by the endothelial cells play a vital role for host defense. Inactivation of uronyl 2‐O‐sulfotransferase (Hs2st) in endothelial cells has been shown to result in enhanced neutrophil recruitment in acute sterile inflammation models. Surprisingly, under real infection situation elicited by Group B Streptococcus (GBS), the leading cause for the neonate pneumonia, sepsis and meningitis, the host survival is not beneficial from the enhanced recruitment of neutrophils in the Hs2st deficient animals. In contrast, they showed increased mortality and uncontrolled bacteria organ disseminations after GBS challenge. Moreover, neutrophils from the Hs2st deficient mice were less capable to eliminate bacteria due to reduced extracellular traps formation which is critical for neutrophil bactericidal activity. In addition, increased adhesion junction disruption and apoptosis on the endothelial cells from Hs2st deficient mice was also observed. Deficiency on both neutrophils and endothelial cells contributes to the susceptibility of Hs2st deficient animals upon GBS challenge.
Intercellular adhesion molecule‐1 deficiency correlates with impaired inflammatory response in congenital disorder of glycosylation (607.2)
Authors: He Ping , Freeze Hudson
Published: June 2021

Congenital Disorders of Glycosylation (CDGs) result from mutations in various N‐glycosylation genes. Many patients are prone to infections, but the mechanism is unknown. We found that CDG patient fibroblasts have less intercellular adhesion molecule‐1 (ICAM‐1), and because of its role in innate immune response, we hypothesized that its reduction may explain recurrent infections. In support of this, tunicamycin blocks TNF‐α induced ICAM‐1 production, suggesting that many CDG cells may show poor ICAM‐1 response. We, therefore, investigated a mouse model of MPI‐CDG, deficient in phosphomannose isomerase. We challenged MPI‐deficient mice with an intraperitoneal injection of pro‐inflammatory mediator (Zymosan) and found decreased neutrophil extravasation. Immunohistochemical staining of mesenteries showed attenuated neutrophil egress, presumably due to poor ICAM‐1 response to acute peritonitis. Since MPI‐CDG patients and their cells improve glycosylation when given mannose, we provided MPI‐deficient mice with mannose‐supplemented water for 7 days. This increased ICAM‐1 expression on mesenteric endothelial cells of Zymosan‐challenged mice and enhanced neutrophils exudation compared to untreated controls. Attenuated inflammatory response in glycosylation‐deficient mice may result from ICAM‐1 deficiency on the vascular endothelial surface and may help explain high mortality in patients.

Grant Funding Source: supported by the Rocket Fund & NIH R01 DK55615
Identification of novel relationships between glycosyltransferase genotypes and protein glycoforms in pancreatic cancer (608.6)
Authors: Tang Huiyuan , Partyka Katie , Maupin Kevin A. , Westra Hannah , Zeh Herbert , Huang Ying , Brand Randall , Haab Brian
Published: June 2021

Specific glycans or protein glycoforms are potential biomarkers of cancer, but variability in expression between people limits their value. A factor influencing the variation between people is the germline genetic variants in specific genes encoding glycosyltransferases. Therefore, identifying the relationships between genetic variants and glycan expression in cancer patients could lead to better methods to detect cancers. To pursue this goal, we measured the levels of multiple glycans and protein glycoforms in the blood plasma of 123 pancreatic cancer patients and 104 control subjects, and we determined the germline genotypes of the same patients for five different genes involved in the biosynthesis of the glycans. We first tested if certain glycans or glycoforms are selectively high in patients with defined genotypes. We confirmed the known relationships between the cancer marker CA 19‐9 and mutations in the FUT2 or FUT3 fucosyltransferase genes, and we found a new relationship between polymorphims in ABO and higher or lower CA 19‐9 levels, depending on the nature of the polymorphism. We next found that cancer‐associated elevations of certain glycoforms are accentuated in patients with specific genotypes. For example, only blood type B cancer patients showed elevations in a unique marker defined by proteins containing both the blood group B glycan and the CA 19‐9 antigen. This study provides a first look at several candidate genotype‐glycan relationships and suggests routes for enhanced detection of cancer.
Regulator of G protein Signaling 6 suppresses Ras and DNA‐methyltransferase 1 expression and tumorigenesis in a mouse model of urinary bladder cancer (609.10)
Authors: Yang Jianqi , Platt Lance , Ibeawuchi Stella‐Rita , Stewart Adele , Maity Biswanath , Huang Jie , Fisher Rory
Published: June 2021

Urinary bladder cancer, the fifth most common cancer in the U.S., frequently results from exposure to toxic chemicals including those present in cigarette smoke. A previous epidemiological study demonstrated that a polymorphism in RGS6 leading to increased translation/stability of RGS6 mRNA was associated with a pronounced reduction in the risk of bladder cancer, particularly in smokers. However, no studies have evaluated whether RGS6 functions as a tumor suppressor in bladder. Here, we employed RGS6‐/‐ mice to investigate the role of RGS6 in BBN (N‐butyl‐N‐(4‐hydroxybutyl) nitrosamine)‐induced bladder cancer, a model which matches well to its human counterpart. We found that RGS6 was highly enriched in mouse bladder, particularly in urothelium where transitional cell carcinoma originates. RGS6 loss was associated with increases in Ras and Dnmt1 expression in the bladder urothelium and marked decreases in Ras‐association domain family 1 isoform A (RASSF1A), a tumor suppressor and target of Dnmt1‐mediated silencing. A similar phenomenon was observed in BBN‐treated WT mice, an effect further exacerbated by BBN treatment of RGS6‐/‐ mice. In keeping with these observations, RGS6‐/‐ mice exhibited accelerated bladder tumor formation. Our findings suggest that RGS6 is a previously unrecognized, but critical repressor for bladder carcinogenesis.

Grant Funding Source: Supported by NIH CA161882, AHA 11SDG7580008
A role for the tyrosine kinase Pyk2 in depolarization‐induced contraction of vascular smooth muscle (609.3)
Authors: Mills Ryan , Mita Mitsuo , Sutherland Cindy , Walsh Michael
Published: June 2021

Background: Depolarization of vascular smooth muscle induces rapid (phasic) contraction followed by sustained (tonic) contraction. The latter is due to activation of the RhoA/Rho‐kinase pathway via genistein‐sensitive tyrosine phosphorylation.

Objective: To determine if the Ca2+‐dependent tyrosine kinase Pyk2 is involved in depolarization‐induced sustained contraction.

Methods: Depolarization of endothelium‐free rat caudal arterial smooth muscle was induced by high extracellular [K+] in the absence or presence of Pyk2 inhibitors. Isometric force was measured with a force transducer and Pyk2 activation (autophosphorylation of Y402) was quantified by western blotting with phosphospecific antibodies.

Results: Depolarization induced Pyk2‐Y402 phosphorylation in a time‐dependent manner. Pyk2 inhibitors (sodium salicylate, PF431396 and others) prevented Y402 phosphorylation and inhibited K+‐induced contraction. Vanadate (tyrosine phosphatase inhibitor)‐, but not calyculin A (Ser/Thr phosphatase inhibitor)‐induced contraction was also inhibited by Pyk2 inhibition.

Conclusion: Membrane depolarization induces phosphorylation and activation of Pyk2, which is required for the tonic phase of K+‐induced contraction.
Low density lipoprotein‐related protein 1 regulates lung inflammation (609.9)
Authors: Garcia‐Arcos Itsaso , Cummins Neville , Dabo Abdoulaye , Pillai Manju , Geraghty Patrick , Goldberg Ira , Foronjy Robert
Published: June 2021

The low density lipoprotein related protein 1 (LRP1) is a widely expressed transmembrane receptor that binds a large number of ligands‐ApoE being one of them‐and is involved in inflammatory signaling cascades in multiple cell types. Recently, LRP1 was linked with lung function in a genomic study, but the implications of this association are unknown. We aimed at describing the role of LRP1 in lung inflammation. Mice were treated with the LRP1 ligand COG133, an ApoE mimetic peptide, for 1 month and they were compared with vehicle‐treated mice. The treatment with COG133 increased whole‐lung baseline protein phosphatase 2A (PP2A) activity, which is considered a defense mechanism often observed after cigarette smoke exposure, and decreased the mRNA expression of TNFα and IL8 in bronchoalveolar cells. In cultured lung epithelial cells, silencing LRP1 increased mRNA expression of MCP1, IL1β and TNFα; activated the transcription factors STAT1 and STAT3; and prevented the activation of PP2A in response to a challenge with cigarette smoke extract. In conclusion, LRP1 can modulate the expression of inflammatory cytokines and the activation of interferon‐mediated signaling cascades in lung.

Grant Funding Source: Supported by NIH 5R01HL098528‐05
NODs‐dependent OmPGRP‐L1 expression modulates the excessive inflammatory response to bacterial infection in rainbow trout liver cells (611.1)
Authors: Jang Ju Hye , Kim Hyun , Kim Yu Jin , Cho Ju Hyun
Published: June 2021

Peptidoglycan recognition proteins (PGRPs) recognize bacterial peptidoglycan and function in antibacterial innate immunity. OmPGRP‐L1, a long form of PGRP from the rainbow trout, is known to have an anti‐inflammatory role in liver cells. In this study, we examined the regulation of the expression of OmPGRP‐L1 in a rainbow trout hepatoma cell line RTH‐149 upon stimulation with microbial ligands. γ‐D‐glutamyl‐meso‐DAP (iE‐DAP) and muramyl dipeptide (MDP) markedly increased the expression of OmPGRP‐L1, as well as pro‐inflammatory cytokines. Silencing of nucleotide‐binding oligomerization domain‐containing protein (NOD)1 and NOD2 specifically inhibited the upregulation of OmPGRP‐L1 expression induced by their cognate ligands. Suppression of NF‐κB activation prevented the induction of OmPGRP‐L1 expression induced by iE‐DAP and MDP. In silico analysis and electrophoretic mobility shift assay revealed that the promoter of OmPGRP‐L1 had NF‐κB binding sites, suggesting that OmPGRP‐L1 is produced through NODs‐NF‐κB signaling pathway. Loss‐of‐function and gain‐of‐function experiments indicated that OmPGRP‐L1 down‐regulated the induction of pro‐inflammatory cytokine expression induced by iE‐DAP and MDP. Overall, these findings suggest that rainbow trout liver cells may modulate the excessive inflammatory response to bacterial infection by producing OmPGRP‐L1 through NODs.

Grant Funding Source: Supported by the National Research Foundation of Korea (NRF‐2011‐0015180 and NRF‐2013M3A6A8073556)
Use of mid‐upper arm muscle circumference to identify undernutrition in hospitalized adults in Hanoi, Vietnam (620.1)
Authors: Manders Aaron , Pham Thi Thu Huong , Nguyen Thi Lam , Nghiem Nguyet Thu , Tran Chau Quyen , Dinh Thi Kim Lien , Nguyen Quoc Anh , Henry Elizabeth , Young Lorraine , Apovian Caroline , Ziegler Thomas , Lenders Carine
Published: June 2021

Objective: To determine the utility of MUAC to screen underweight hospitalized patients in a major Hanoi teaching hospital.

Methods: Data from a one‐day study of patients 蠅19 years old admitted at Bach Mai Hospital, Hanoi, Vietnam (2010) were utilized. Data included actual weight and height, MUAC, age, sex, presence of edema, reason for admission, ward, and prior subjective weight loss (major, >10%) over six months. BMI was calculated and underweight defined as BMI < 18.5 (kg/m²; WHO standards). Receiver operating characteristic curve (ROC) analyses was performed.

Results: A total of 420 inpatients were studied (51.4 ± 0.8 years old; 177 (42%) female. Patients were admitted for acute (n=174, 41%), chronic (n=186, 44%), elective (n=31, 7%), or other (n=29, 7%) reasons. Mean ± SD for BMI was 20.0 ± 0.2 kg/m² and for MUAC 24.5 ± 0.2 cm. Rates of underweight status, and moderate and severe, or severe thinness were 34%, 17%, and 8%. A total of 22% subjects exhibited edema and 15% reported major weight loss prior to admission. For underweight status, sensitivity of MUAC <23 cm was 71%, specificity 90%, (+) predictive value 79%, and (‐) predictive value 86%. Area Under Curve (AUC) for MUAC<23 cm was 0.81.

Conclusion: MUAC <23 cm appears to be a good method to identify undernutrition in hospitalized adults in Vietnam (AUC 0.80‐0.90 is good). Further evaluation of MUAC in larger samples and association with hospital morbidity and mortality in Vietnam are needed.

Grant Funding Source: Supported by the Abbott Fund