bluuebunny/crossref_metadata_2025_split huggingface dataset:
Benzoylecgonine, a Major Cocaine Metabolite, Enhances HIV‐1 Integration and Replication in Immune Cells
Authors: Addai Amma , Pandhare Jui , Dash Chandravanu 
Published: June 2021

Several studies have indicated an association between drug abuse and an increased risk for HIV/AIDS. The presence of benzoylecgonine (BE), a major cocaine by‐product, is almost a certain indication that a person has consumed the drug. Therefore, BE testing in urine is commoner than testing of cocaine itself in Drug Screening Tests. It has been demonstrated that cocaine influences HIV‐1 replication in peripheral blood mononuclear cells (PBMCs), CD4+T cells, and humanized mice. However, the effects of cocaine and its metabolite on HIV‐1 replication have not been elucidated. The goal of this study is to examine the effects of BE and cocaine on HIV‐1 integration in immune cells that serve as the primary targets HIV in vivo. PBMCs and CD4+ T cells were isolated from human blood by Ficoll‐based reagents. HIV‐1 integration and replication were measured by quantitative nested PCR and flow cytometry. We demonstrate that BE and cocaine increase HIV‐1 integration in immune cells in a dose dependent manner. A direct role of cocaine on HIV‐1 integration was further confirmed by in vitro integration assay using HIV‐1 pre‐integration complexes (PICs) isolated from infected cells treated with cocaine. We observed a maximum increase in integration at 50uM. We also demonstrate that cocaine can localize to the nucleus of CD4+ T cells by utilizing ESI based mass spectrometry. Given that cocaine abuse serves as a powerful cofactor for HIV‐1 infection, replication, and pathogenesis in vivo, our data may have implications in the worsening of disease among cocaine abusing HIV patients. *Amma Addai is supported by T32 grant #5T32HL007735‐19 from NIH/NHLBI to Dr. S. E. Adunyah. The research is in part supported by grants DA024558, DA30896, DA033892 and DA021471 from NIDA/NIH to CD.
Free Actin Impairs Macrophage Bacterial Defense: Reversal by Plasma Gelsolin
Authors: Ordija Christine , Chiou Terry , de Oliveira Valdo Melina , Yang Zhiping , DeLoid Glen , Kobzik Lester 
Published: June 2021

Alveolar macrophage (AM) bacterial clearance is critical to host defense against lung infections. We sought to assess: 1) the effects of free‐actin released during lung injury on AM function and 2) the therapeutic potential of the actin‐binding protein plasma gelsolin (pGSN). AM binding and uptake of bacteria in the presence or absence of free actin and pGSN was measured in vitro using assays based on CFU quantitation or scanning cytometry and fluorescence microscopy of bacteria. We demonstrate impaired binding of Streptococcus pneumoniae by murine B6 AMs in the presence of free actin and reversal of impaired binding by pGSN. In contrast, albumin, heat‐denatured actin or vehicle control do not affect AM bacterial binding, thus establishing specificity. Similarly, primary human AM‐like macrophages also show impaired binding and uptake of Staphylococcus aureus in the presence of free actin or cytochalasin D and improved binding and uptake by pGSN. In conclusion, free actin was found to significantly impair AM binding and phagocytosis of microbial pathogens in vitro, an effect that was reversed in the presence of pGSN. The scanning cytometry method used in these studies will be useful to investigate the mechanism(s) by which free actin interferes with AM bacterial binding. Free actin release after lung injury may contribute to the risk of secondary bacterial pneumonia, and pGSN may offer a therapeutic strategy to reduce this risk.Funding provided by NIH HL007118
A non‐human primate model for Yersinia pestis infection: metabolomic analysis pilot study
Authors: Gautam Aarti , Miller Stacy , Hoke Allison , Hammamieh Rasha , Jett Marti 
Published: June 2021

Yersinia pestis, the etiological agent of the plague, is a potential biothreat agent. Upon inhalation, the Gram‐negative bacterium causes pneumonic plague, a rapidly progressing, severe lung infection that is usually fatal. Only limited work using metabolite profiles to study the progression of the disease has been done, and exact mechanisms that facilitate disease are not well‐characterized. In this study, African Green monkeys were exposed to aerosolized highly virulent Y. pestis CO92 and metabolomic profiling of plasma samples was conducted. Comparison of control counterparts to the disease states at earlier time points (6 hours to 18 hours) versus at later time points (24 hours onwards) showed mainly two‐fold higher increases in the levels of certain polyunsaturated fatty acids, long chain fatty acids and metabolites from fatty acid metabolism pathway. Significant alterations in metabolites related to early inflammation and oxidative stress, increased energy demands to the host, and potential renal dysfunction were revealed. The metabolomics of plasma have the potential to provide a more thorough understanding of the Y. pestis pathogenesis, which may assist future treatment of this disease.The grant # TMTI0029_09_WR_T from Department of Defense Chemical and Biological Defense program through the Defense Threat Reduction Agency is acknowledged.DISCLAIMER“Research was conducted in compliance with the Animal Welfare Act and all other Federal requirements. The views expressed are those of the authors and do not constitute endorsement by the U.S. Army.”
Trimeric G Protein Activation by the Non‐Receptor Protein GIV/ Girdin is Required for Integrin Signaling
Authors: Leyme Anthony , Perez‐Gutierrez Lorena , Nguyen Lien , Garcia‐Marcos Mikel 
Published: June 2021

Signal transduction via both integrins and G protein‐coupled receptors (GPCRs) is critical to control cell behavior in health and disease. These two receptor classes have been traditionally believed to trigger distinct and independent signaling cascades. Here we report a novel mechanism of integrin signal transduction that requires activation of the trimeric G protein Gαi by the non‐receptor Guanine nucleotide Exchange Factor (GEF) GIV (aka Girdin), a pro‐metastatic protein. We demonstrate that GIV is necessary and sufficient to enhance responses to extracellular matrix (ECM) stimulation and make tumor cells more invasive. These responses include remodeling of the actin cytoskeleton and PI3K‐dependent signaling, resulting in enhanced haptotaxis and haptoinvasion. Tumor cells engineered to express GEF‐deficient GIV fail to transduce integrin signals into pro‐invasive cell responses. Mechanistically, we show that GIV and Gαi3 are recruited to integrin complexes upon ECM stimulation and activate a Gβγ‐PI3K‐Akt signaling cascade. In summary, our discoveries delineate a novel mechanism by which integrin signaling is rewired during metastasis to result in increased tumor invasiveness.This work is supported by the American Cancer society (RGS‐13‐362‐01‐TBE), the Elsa U. Pardee Foundation and NIH (R01GM108733).
Identification and Characterization of a Group of Polymorphic, Single Domain Peptidoglycan Hydrolases of the NlpC/P60 Superfamily in Dust Mites
Authors: Tang Vivian , Stewart Geoffrey , Chang Barbara 
Published: June 2021

A 14 kDa bacteriolytic enzyme, LytFM, previously purified from the growth medium of dust mite Dermatophagoides pteronyssinus, shows high sequence homology with cysteine peptidases belonging to the NlpC/P60 Superfamily. This study aimed to identify genes encoding similar proteins in dust mite species and to characterize the putative encoded proteins. PCR and bioinformatic analysis revealed homologous genes encoding this single domain protein in Psoroptes ovis, Blomia tropicalis, Euroglyphus maynei and D. farinae. A variant, LytFM2, was detected in D. pteronyssinus indicating that the D. pteronyssinus proteins are polymorphic. The lytFM sequence was amplified from a D. pteronyssinus gDNA library as an intronless gene and its 3′ end flanking sequence contained a prokaryotic promoter sequence suggesting a prokaryotic origin. Homology studies showed that the putative mite proteins possessed a cysteine peptidase catalytic triad characteristic of members of the NlpC/P60 Superfamily involved in peptidoglycan processing. However, they all lacked a conserved Asp residue characteristic of NlpC/P60 proteins recognizing a free L‐Ala residue of the peptidoglycan stem peptide suggesting that the mite proteins cleave GlcNAc‐MurNAc‐L‐Ala‐γ‐D‐Glu‐mDAP/Lys peptidoglycan substrates. These enzymes thus represent a conserved group of proteins in several mite species that may have an important role in protection of the gut from pathogens or utilization of bacteria as a food source, or both. As at least two of these mite proteins are secreted, they may represent a novel group of allergenic proteins.Work supported by Asthma Foundation WA and NH&MRC Australia.
Deciphering Differential Distortion of Purine Substrates by Human and Plasmodium falciparum HGPRT
Authors: Karnawat Vishakha , Gogia Spriha , Balaram Hemalatha , Puranik Mrinalini 
Published: June 2021

Enzymes of the salvage pathway are essential to understanding the biochemistry of parasitic protozoa. These parasites lack de novo pathway for nucleotide synthesis and depend solely on purines salvage from their host. Hypoxanthine guanine phosphoribosyltransferase (HGPRT) from Plasmodium falciparum has long been regarded as a potential therapeutic target. Although structurally homologous to human enzyme, PfHGPRT differs from human (h) HGPRT in its expanded substrate specificity and activity towards common substrates. We used substrate analogues, 9‐deazapurines, in combination with UV resonance Raman spectroscopy to capture non‐covalent interactions in enzyme‐substrate complexes. Raman spectral band positions are sensitive to sub‐angstrom changes in bond length. We observed perturbations in Raman spectra that provide quantitative information about hydrogen bonding and stacking‐interactions with the enzyme active‐site. The bound ligands are extensively distorted towards the transition‐state of phosphoribosyltransferase reaction. Differential loss in Raman crosssection of the ligands bound to two enzymes suggests that active‐site is more compact in hHGPRT than in PfHGPRT. The rigidity of active‐site in hHGPRT is attributed to tighter stacking interaction between phenylalanine residue of the active‐site and the nucleobase.Research was supported by IISER, India and Council for Scientific and Industrial Research, India.
Vimentin Phosphorylation Underlies Corneal Myofibroblast Sensitivity to Withaferin A
Authors: Bargagna‐Mohan Paola , Lei Ling , Thompson Alexis , Shaw Camille , Mohan Royce 
Published: June 2021

ObjectiveTo compare vimentin phosphorylation in corneal fibroblasts with myofibroblasts and determine their sensitivity to withaferin A.Methods: Primary rabbit corneal fibroblasts and myofibroblasts were subjected to cell spreading assays. Wild type and vimentin‐deficient mice were subjected to corneal alkali injury and treated with withaferin A (WFA). Corneas were subjected to immunohistochemistry and western blot analyses.Results: We show that myofibroblasts produce increased serine‐38 phosphorylated vimentin (pSer38Vim) and display impaired cell spreading and cell polarization compared to fibroblasts. The pSer38Vim isoform is inefficiently incorporated into growing vimentin IFs of myofibroblasts during cell spreading as myofibroblasts maintain higher levels of soluble pSer38Vim compared to fibroblasts. WFA treatment causes a potent blockade of cell spreading selectively in myofibroblasts, targeting pSer38Vim for hyperphosphorylation, which is not induced in fibroblasts. Hyperphosphorylated pSer38Vim in myofibroblasts complexes with adaptor protein filamin A (FlnA), and appears as short squiggles displaced from focal adhesions upon WFA treatment. The extracellular‐signal regulated kinase (ERK) is also phosphorylated (pERK) in response to WFA, but pERK does not enter the nucleus because it remains bound to pSer38Vim in cytoplasmic complexes. Fibrotic corneas of injured wild type mice possess high levels of pERK compared to injured vimentin‐deficient mice. WFA treatment causes a decrease in pERK and downregulates pSer38Vim expressing myofibroblasts in corneas of wild type mice.ConclusionsWe have identified pSer38Vim as a major determinant of myofibroblast sensitivity to WFA. Support: R01 EY016782
Kinetic characterization of the α‐2,9 polysialyltransferase of Neisseria meningitidis
Authors: Ming Shonoi , Peterson Dwight , Vann Willie 
Published: June 2021

Neisseria meningitidis Serogroup C is responsible for the majority of the meningococcal meningitis outbreaks in the United States in the past 20 years. The serogroup polysaccharide, an α‐2, 9 polysialic acid, of the pathogen is essential for virulence. This highlights the importance understanding of the enzymes involved in its biosynthesis as it may lead to the development of new drugs targeting this disease.We are interested in obtaining a detailed understanding of the kinetics of the α‐2, 9 polysialyltransferase involved in the formation of the polysaccharide capsule of N. meningitidis. Studies of the α‐2, 8 polysialyltransferase conducted by the Wakarchuk group obtained an apparent Km in the low mM range for CMP‐sialic acid. Using the α‐2, 9 polysialyltransferase we obtained an apparent Kmvalue in the low µM range for CMP‐sialic acid. We believe that this large discrepancy in Km is due primarily to the differences in assay methods and conditions.Using CMP‐sialic acid as the donor substrate, we compared acceptor substrate specificity of GD3, a ganglioside, with a hexamer of sialic acid. We observed a lower apparent Km value for GD3 compared with the hexamer suggesting that the attachment of a lipid to the oligosaccharide improves substrate binding. However, the enzyme does not appear to follow Michaelis‐Menten kinetics. The complexity of the reaction kinetics may be due to fact that each product formed via the transfer of sialic acid from CMP‐sialic acid in turn produces a new substrate. To address this issue we used CMP‐sialic acid analogs modified at the N‐9 position to observe a single turnover of product, which allowed us to perform detailed kinetic characterization of the enzyme.
Hepatoma‐Derived Growth Factor (HDGF) Acts in Ovarian Cancer via Distinct Intracellular and Extracellular Mechanisms
Authors: Giri Karuna , Pabelick Christina , Mukherjee Priyabrata , Prakash YS 
Published: June 2021

Ovarian cancer is the eighth most common cancer and the fifth leading cause of cancer death among US women. Hepatoma‐derived growth factor (HDGF) is a heparin binding, secreted mitogen with prognostic value in multiple cancers. We previously reported that HDGF is overexpressed in ovarian cancer. Surprisingly, the mechanisms by which HDGF acts in any cell type remains to be fully elucidated. We hypothesized that overexpression of HDGF in ovarian cancer cells is functionally relevant and that HDGF confers pro‐survival and proliferative properties, promoting cancer growth. To elucidate intracellular and extracellular signaling mechanisms, we examined the effect of HDGF knockdown via siRNA, overexpression of HDGF and stimulation by full‐length recombinant protein. Immunofluorescence image analysis showed nuclear localization of HDGF in OV202 cell line. Additionally, HDGF was minimally secreted but passively released in greater amounts by necrotic cells. Extracellular HDGF stimulated phosphorylation of ERK and p38. In contrast, modulating intracellular HDGF did not affect MAPK pathways. In fact, intracellular HDGF functioned in a distinct manner by regulating cellular metabolism. This suggests that HDGF acts in ovarian cancer cells via two distinct mechanisms, both of which could be exploited for ovarian cancer therapy.Supported by the Mayo Graduate School (Giri) and the Department of Anesthesiology, Mayo Clinic (Prakash).
Vitamin D supplementation as adjunct to anti‐tuberculosis drugs in Mongolian TB patients
Authors: Davaasambuu Ganmaa , Bromage Sabri , Erkhembayar Baasansuren , Jaisandavga Erdenetseg , Baatar Munkhzul 
Published: June 2021

Objectiveto test whether vitamin D supplementation in vivo may be a useful adjunct to anti‐TB drugs and could lead to the development of shorter drug regimens.Methods: Double‐blind, placebo‐controlled randomized clinical trial among TB patients in Mongolia. The baseline clinical assessment consisted of chest radiography, measurement of height, weight, mid‐upper arm circumference, and triceps, biceps and subscapular skinfolds and collection of a sputum, a urine, and a blood samples.Results: The results were available for the subsample of the patients and were analyzed by cavity status at baseline. Fewer of the participants were female (31%) compared to the males (69%). At baseline, 46 patients had a cavity (Cav +) and 94 did not have a cavity (Cav ‐) on chest radiograph. The participants in the Cav (‐) and Cav (+) groups were similar in age (mean 34.9 + 11.9 sd cm vs. 34.9 + 15.4 sd cm). Sputum samples are collected and cultured every two weeks. For each subject, the date of conversion from positive to negative was determined. Since un‐blinding has not occurred, we are presenting here pooled time to sputum culture conversion.ConclusionsMean serum concentration of albumin at baseline was 34.7 + 60.5 g/L in the Cav (+) compared to 37.2+4.9 g/L in the Cav (‐) group (p<0.05). Mean concentrations of C Reactive Protein (CRP) and neutrophils were significantly higher in the Cav (+) group compared to the Cav (‐) group.
Effects of zinc and multivitamin supplementation on hematologic status during infancy
Authors: Carter R Colin , Kupka Roland , Manji Karim , McDonald Christine , Aboud Said , Gosselin Kerri , Kisenge Rodrick , Liu Enju , Fawzi Wafaie , Duggan Christopher 
Published: June 2021

OBJECTIVETo determine the separate effects of zinc and multivitamin supplementation on infant hematologic status.METHODSIn a double‐blind RCT, infants born to HIV‐negative women (n=2400) in Dar es Salaam, Tanzania were randomized to daily oral supplementation with zinc (Zn), multivitamins (MV), Zn and MV, or placebo at age 6 wk. Hemoglobin concentration (Hb) and red cell indices were measured at baseline and ages 6, 12, and 18 mo. A subset of infants (n = 589) was examined for iron deficiency at 6 mo, defined as ferritin < 12 μg/L or, where α‐glycoprotein 蠅 1.0 g/L, serum transferrin receptor > 8.3 mg/L.RESULTSInfants treated with MV had higher mean Hb than those given placebo or Zn alone at 18 mo (9.9 vs. 9.6 g/dL, p = .0009). Infants treated with Zn had RR of 1.71 (95% CI 1.01 – 2.89) for iron deficiency at 6 mo, while those treated with MV had RR 0.52 (95% CI 0.30 – 0.90). In Cox models, MV were associated with a 28% reduction in risk of severe anemia (HR = 0.72 [95%CI 0.56 – 0.94]) and a 26% reduction in risk of severe microcytic anemia (HR = 0.74 [95% CI 0.56 – 0.96]) through age 18 mo. No effects of Zn on risk of anemia were seen.CONCLUSIONSMV were associated with reduced risk of both iron deficiency and severe microcytic anemia, while Zn was associated with increased risk of iron deficiency but no longer term increase in risk of anemia.FundingNICHD (R01 HD048969; K24HD058795); NIAAA (K23 AA020516); Bill and Melinda Gates Foundation (OPP1066203)
Behavioral and Predictive Validation of a Self‐Assessment of Parental Feeding Style in Low‐Income Parents of Young Children
Authors: Ontai Lenna , Sutter Carolyn , Shilts Mical , Townsend Marilyn 
Published: June 2021

To improve the capacity of researchers and practitioners to work toward integrating parenting styles into their work on child obesity, valid assessment tools aimed at measuring these dimensions of parenting are needed. Behavioral and predictive validation of My Child at Mealtime (MCMT), a visually enhanced self‐assessment tool designed to measure parent feeding style in low‐income parents of young children. Parents (n=103) from Head Start and WIC provided data about themselves and their young children. Longitudinal data collection included the parent self‐administered MCMT and child height and weight for BMI percentile, a continuous variable. MCMT is scored for child and parent centered feeding (CCF, PCF). A subsample of 60 parents and children were videotaped during a family mealtime at the start of the study. Coded parent behaviors were grouped based on MCMT scoring to create “parent focused feeding” (PFF) and “child focused feeding (CFF) scores. Significant correlations were found between PFF and MCMT CCF (r = .36, p < .01) and PCF (r = .22, p < .10). Assessment of predictive validity (n=48) found that MCMT PCF was significantly associated with BMI percentile two years later (r = ‐.32, p<.05). MCMT is a valid parent self‐assessment tool to measure parent‐centered feeding behaviors in low‐income parents of young children. Further analyses will assess the validity of MCMT in assessing child‐centered feeding behaviors.Supported by National Research Initiative #2009‐55215‐05019 & Agriculture and food Research Initiative #2010‐85215‐20658 from the USDA National Institute of Food and Agriculture, Human Nutrition and Obesity 93330.
WAVE Pilot Study: Feasibility of Recruiting, Enrolling, Assessing, and Retaining High School Soccer Players Using Soccer Coaches in a 2‐Year Integrative Obesity Prevention Research Study
Authors: Wong Siew Sun , Meng Yu , Manore Melinda , Pilolla Kari , Johnson Tonya , Curiel Cristian , Dursch Gretchen 
Published: June 2021

Objective:To evaluate participant recruitment, enrollment, assessment, and retention in a 2‐year intervention designed to prevent excess weight gain among high school (HS) soccer players ages 14‐19.Method:Five soccer teams in 2 school districts were recruited through soccer coaches. Prior to enrollment, parents, coaches and youth all received a printed information packet. Participants were then enrolled into an intervention/control group; non‐randomized by stratified by HS and soccer teams. At baseline, youth and parent's demographics were collected. Youth also had height, weight, and body composition measured and completed questionnaires related to sport nutrition knowledge, eating behavior, bowel health, physical activity, and sleep.Result:Overall 44% (n=54) of available athletes were enrolled (intervention: 16F, 11M; control: 21F, 6M). We identified the following approaches to help us to be successful in future intervention: 1) Engage coaches early in the process; 2) have coaches sign an ‘agreement' statement regarding their role and deliverables including: attend all sessions, help to collect forms/documents, and etc.; 3) engage school districts; and 4) use computers/tablets for data collection through Qualtrics instead of paper/pencil forms.ConclusionBased on this approach, soccer coaches, athletic directors, and parent coordinators were pivotal to the success of a longer‐term obesity prevention intervention.Sponsor: NIFA AFRI Award No. 2013‐67001‐20418