bluuebunny/crossref_metadata_2025 huggingface dataset:
Using family‐based experiential learning to improve nutrition knowledge, dietary intake, physical activity, and food purchasing behaviors among Latina WIC participants and their children: a pilot study (1021.3)
Authors: Gewa Constance , Pawloski Lisa , Kanianthra Anna , Forrester Mena , Long Erika , Legum Margaret
Published: June 2021

Objective: To examine the impact of a family‐based nutrition education program on nutrition knowledge, diet, physical activity, and food purchasing behaviors of Latina mothers and children participating in Northern Virginia Women, Infant, Children (WIC) programs.

Methods: Surveys were administered to mothers (n=15) using a pre‐test/post‐test design. The family‐based nutrition intervention included 1) Discussion and lecture on food labels, food purchasing, portion sizes, and healthy meals, 2) Experiential learning focused on preparation and storage of low‐cost, healthy meals incorporating WIC foods, and 3) A Zumba class and discussion on physical activity.

Results: The data revealed improved diet such that mothers reported increased fruit and vegetable consumption, decreased juice consumption among their children. Mothers reported their children were more physically active. Further, mothers prepared more meals at home using raw ingredients.

Conclusions: The findings are significant in that they support growing literature of the success of family based interventions. Further, these data show the importance of integrating experiential learning activities such as cooking and physical activity with the more traditional didactic methods.

This research was supported the Virginia Department of Health and the HRSA funded Virginia Commonwealth Public Health Training Center.

Grant Funding Source: Supported by the Virginia Department of Health and HRSA funded Virginia PHTC
Food frequency questionnaire as an indicator of serum essential n‐3 and n‐6 polyunsaturated fatty acids concentrations in early pregnancy, according to body mass index (1022.3)
Authors: Lepsch Jaqueline , Vaz Juliana , Pinto Thatiana , Rebelo Fernanda , Farias Dayana , Franco‐Sena Ana Beatriz , Kac Gilberto
Published: June 2021

Objective: To investigate whether food frequency questionnaire (FFQ) is indicative of serum essential n‐3 and n‐6 polyunsaturated fatty acids (PUFA) concentrations in early pregnancy and if correlations differ between Body Mass Index (BMI) categories. Methods: Cross‐sectional study of 248 pregnant women between 6th and the 13th weeks of gestation. Analyses comprised Spearman’s correlation (rs) for total sample and also stratified according to BMI (kg/m2) [underweight/normal weight (<25; n=139) and overweight (蠅25; n=109)]. Results: In the total sample, dietary report was significantly correlated with serum PUFA (rs=0.243, P<0.001), linoleic acid (LA, 18:2n‐6; rs=0.278, P<0.001), eicosapentaenoic acid (EPA, 20:5n‐3; rs=0.277, P<0.001) and docosahexaenoic acid (DHA, 22:6n‐3; rs=0.207, P<0.001) concentrations. We observed that underweight/normal weight women presented significant correlations between FFQ and serum concentrations of total PUFA (rs=0.339, P<0.001), LA (rs=0.332, P<0.001), EPA (rs=0.355, P<0.001) and DHA (rs=0.179, P=0.034) and overweight women only with alpha linolenic acid (ALA, 18:3n‐3; rs=0.195, P=0.042) and DHA (rs=0.241, P=0.011). Conclusions: FFQ may be indicative of the serum composition of essential n‐3 and n‐6 PUFAs in early pregnancy. Higher correlations were observed in the underweight/normal weight women.

Grant Funding Source: Supported by FAPERJ, Cnpq and CAPES
Butyrate increased lifespan in C. elegans (1025.13)
Authors: Gao Chenfei , Cao Tuan , Martin Roy , Keenan Michael , Greenway Frank , Finley John , Burton Jeff , Johnson William , Enright Frederick , Zheng Jolene
Published: June 2021

Obesity is a growing public health dilemma around the world. Resistant starch (RS), fermented resistant starch, and short‐chain fatty acids (SCFAs) reduce intestinal fat deposition (IFD) in wild type Caenorhabditis elegans (N2, C. elegans), and increase the healthspan marker, pharyngeal pumping rate (PPR). We hypothesized that the fermentation products, SCFAs, would improve lifespan in C. elegans model. N2 was maintained in liquid culture media supplied with 5‐fluoro‐2’‐deoxyuridine (FUdR, 0.24mM). Control group was fed with E. coli OP50 only. Experimental group received additional sodium butyrate (0.3mM, 1mM), sodium acetate trihydrate (0.3mM), sodium propionate (100mM), and tributyrin (3mM). Butyrate (0.3mM) increased mean survival time during 35 day study (P=0.03) in the C. elegans. Other treatments mildly sustained the lifespan of C. elegans. These data proved that the SCFA butyrate extended lifespan in addition to creating a negative energy balance in C. elegans model.
Association between serum vitamin D and insulin resistance in African Americans with type 2 diabetes (1030.3)
Authors: Exebio Joel , Zarini Gustavo , Vaccaro Joan , Ajabshir Sahar , Cheema Amanpreet , Antwi Janet , Antwi Janet , Huffman Fatma
Published: June 2021

The purpose of the study was to determine the association between serum vitamin D and insulin resistance measured by HOMA2‐IR index in a sample of African Americans with type 2 diabetes residing in South Florida. Subjects were recruited by community outreach. One hundred and eleven subjects (n=111) were included in the study. Fasting plasma insulin and glucose were used to calculate HOMA2‐IR. Serum vitamin D was measured with an enzyme‐immunoassay kit by absorbance. HOMA2‐IR values were log transformed to achieve normality. Unadjusted and adjusted multilinear regressions were used to determine the association between log HOMA2‐IR and serum vitamin D. Log HOMA2‐IR was the dependent variable. The mean age, BMI, A1C, energy intake, serum vitamin D, and log HOMA2‐IR values were 54.73 ± 10.21 years, 35.90 ± 8.22 kg/m2, 7.61 ± 1.92 (%), 2088.47 ± 1116.58 kcal, 19.12 ± 8.86 (ng/mL), and 0.22 ± 0.29 respectively. In the unadjusted model, serum vitamin D was associated with log HOMA2‐IR (β=‐0.203, P=0.030). However, the association did not hold after adjusting for covariates (P=0.109). Age (P=0.020), BMI (P=0.001), and A1C (P=0.002) were strongly associated with HOMA2‐IR. The association among serum vitamin D and HOMA2‐IR was confounded by age, BMI and A1C.

Grant Funding Source: Supported by NIH/NIDDK sponsored grant. JE was supported by NIH/NIGMS R25 GM061347.
Analysis of the differentiation capacity of muscle satellite cells derived from FOXO1‐ genetically modified mice (1033.5)
Authors: Yamashita Atsushi , Hatazawa Yukino , Yoshimura Ryoji , Ogawa Sachika , Ono Yusuke , Kamei Yasutomi
Published: June 2021

Muscle satellite cells are specific stem cells of skeletal muscle. They play important roles in muscle growth, hypertrophy, and regeneration. The transcriptional factor FOXO1, known to induce muscle atrophy, has been reported to be involved in muscle differentiation; however, the experiments have been performed using the immortalized cell line C2C12. In this study, we evaluated the differentiation capacity of satellite cells isolated from muscle‐specific FOXO1‐transgenic (Tg) mice because these cells are believed to mimic in vivo skeletal muscles. Extensor digitorum longus muscles were isolated from mice and digested in collagenase‐type 1 solution until they separated into single fibers. The single viable muscle fibers were then purified using Pasteur pipettes with fire‐polished tips and cultured in growth medium in Matrigel‐coated wells. After cultivation for 5 days in growth medium, followed by 3 days in differentiation medium, we evaluated the differentiation capacity of the satellite cells. Thus, satellite cells were successfully isolated from FOXO1‐Tg mice. These cells tended to exhibit suppressed myotube formation compared with those from the wild‐type mice. These findings suggested that FOXO1 plays a role in not only mature skeletal muscles but also muscle satellite cells.
Organ inflammation and oxidative damages in fructose‐fed adult offspring born of fructose‐fed dams: modulation by maternal bitter melon supplementation (1033.6)
Authors: Li Edmund , Ching Rachel
Published: June 2021

Hyperinsulinemia and dyslipidemia in offspring born of high fructose (F)‐fed dams could be reversed by supplementing bitter melon juice extract to the F dams (FBM). However, when offspring were also fed F, only serum free fatty acid but not insulin level was normalized in those born of FBM dams. This study further examined the impacts of cross‐generation F intake on organ inflammation in the offspring and the effects of maternal BM supplementation. Virgin female rats received control (C), F (60%), or BM‐supplemented fructose (FBM; 1%) diet before conception until d 21 of lactation. Weaned offspring were fed to the C or F diet for 20 wk, forming C/C, C/F, F/F, FBM/F groups. Compared to the C/F and FBM/F rats, F/F rats had higher PPARγ gene expression in adipose tissue but lower gene methylation at the promoter region. Expression of TNFα, IL‐6 and leptin followed that of PPARγ responses to maternal F and FBM. Maternal BM dampened F‐induced hepatic inflammation, lowering SOCS3 mRNA level and JNK1 phosphorylation. There were higher activities of antioxidant enzymes and lower malondialdehyde level and JNK1 phosphorylation in the brain of FBM/F compared with the F/F group. The results show plasticity of fetal programming in that maternal BM supplementation mitigated F‐induced peripheral organ dysfunction and inflammation, and exerted neuroprotective effects against oxidative damages in the brain of adult offspring.

Grant Funding Source: Supported by a HKU grant
Effect of a dietary portfolio (nopal, chia, soy, oat and inulin) on serum lipoprotein profile in subjects with metabolic syndrome (1035.7)
Authors: Aguilar Miriam , Medina Isabel , Guevara Martha , Radyx Miriam , Orozco Ximena , Jimenez Edith , Flores Adriana , Castaño‐Jameson Marisol , Ayala Brenda , Vazquez Natalia , Tovar Armando , Torres Nimbe
Published: June 2021

Approximately one of each two Mexicans adults present metabolic syndrome. The initial treatment includes changes in lifestyle, such as the consumption of specific dietary portfolio (DP). The aim of this study was to assess the effect of this DP on subjects with metabolic syndrome on serum lipoprotein profile, anthropometric measurements, blood pressure and HbA1c. Thirty‐two patients after two and a half months of dietary treatment had a significant decrease in body weight (‐3.5%), BMI (‐6.2%), percentage of fat mass (‐2.7%), systolic and diastolic blood pressure (‐7%, ‐9.8%) and HBA1c (‐1.7%). There was a significant decrease in serum total triglycerides (‐20.9%), VLDL and chylomicron triglyceride (‐6.3%), IDL particles (‐20.7), LDL particles (‐6.3%) particularly the small LDL particles (‐13.6), whereas the large LDL particles increased (29.9%). In summary, the addition of functional food to the DP may reduce the cardiovascular risk by producing a less atherogenic lipoprotein profile.

Grant Funding Source: Supported by Medix (to NT) and CONACYT grant 181685 (to MG)
Sorghum starch is well digested in developmental pancreatic α‐amylase insufficient children in Mali (1039.11)
Authors: Cisse Fatimata , Diall Hawa , Rahmanifar Atossa , Sylla Mariam , Opekun Antone , Grusak Michael , Lin Amy , Nichols Buford , Hamaker Bruce
Published: June 2021

We hypothesized that moderately malnourished weaned stunted children have impaired ability to digest starch due to developmental pancreatic α‐amylase insufficiency that impedes their normal growth with typical foods used at home. An innovative non‐invasive modified 13C‐breath test was used to assess the ability to digest starch in healthy (16) and moderately stunted infants (32) from 18 ‐ 30 months in Bamako, Mali. Common or modified sorghum porridges and three different 13C substrates (UL‐algal starch, UL‐algal limit dextrins, and greenhouse‐enriched sorghum) were fed in five separate days. Serial breath samples (every 15 min for 3 hours) were collected and analyzed using a 13CO2 infrared spectrophotometer. The obtained results showed a pancreatic α‐amylase insufficiency in both healthy and moderately stunted groups. 90% of all subjects digested well the common sorghum porridge whereas the modified porridge with mechanical treatment presented a reduced digestion rate which was not corrected with further amylase pretreatment. We conclude that the digestion of sorghum porridge starches is poorly related to amylase insufficiency, and these starches are well digested via the mucosal α‐glucosidases in Malian moderately malnourished stunted weaned children.
Mentoring young scientists (104.1)
Authors: Maxfield Fred
Published: June 2021

Mentoring young scientists has taken on an increasingly important role in the past several years. Important factors related to this increased role include the alterations in financial support for biomedical sciences, changes in the job market, changing attitudes about work‐life balance and others. Senior faculty members, who would traditionally play major mentorship roles, are often not particularly well prepared for the non‐scientific aspects of mentoring. The majority of senior faculty members have followed a certain career trajectory (with, of course, some individual variation). However, only a small fraction of the scientists who are being trained will be following a similar trajectory. Thus, it is important for educational and research institutions to develop ways to provide career advice to students, postdocs, and junior faculty members. In a recent ASBMB survey of biochemists and molecular biologists in early stages of their careers, nearly 40% reported that they were unsatisfied or very unsatisfied with the career mentoring they had received. In contrast, a strong majority were satisfied or very satisfied with their scientific mentoring. The career mentoring support provided by institutions varies greatly, and I will mention some successful examples. I will discuss other resources available to young scientists that can provide career planning advice.
Iron bioavailability of low and high phytic aid maize produced via recurrent selection (1042.4)
Authors: Beavers Alyssa , Scott Paul , Moran Lauter Adrienne , Goggi Susana , Reddy Manju
Published: June 2021

Phytic acid is a potent inhibitor of iron absorption and is abundant in staple food crops. Low phytic acid (lpa) maize mutants have been produced, but have poor germination rates. The first objective of this study was to determine if conventional breeding can be used to select for phytic acid content in maize without reductions in germination rate, and the second objective was to determine the iron bioavailability of maize produced through this method. For the first objective, the BS11 maize population was used in a recurrent selection breeding program to selectively breed for both low phytic acid (BS11LPA) and high phytic acid (BS11HPA) content, and germination tests were performed for both groups and compared with the lpa mutant. For the second objective, maize samples were subjected to in vitro digestion and then applied to Caco‐2 cells to assess iron bioavailability. Our results show that phytic acid content is significantly different among all three maize varieties, with BS11HPA>BS11LPA>lpa mutant (p<0.05). Germination rates for both BS11LPA and BS11HPA are significantly (p<0.05) greater than for the lpa mutant. Iron bioavailability was ≍35% higher in the BS11LPA maize than the BS11HPA maize (p<0.05, n=8), but there was no significant difference in bioavailability between BS11LPA and lpa mutant maize (p>0.05, n=8). This study concludes that selective breeding can produce low phytic acid maize with better germination rates than lpa mutant maize, and that this maize has similar iron bioavailability compared with lpa mutants.

Grant Funding Source: Supported by USDA‐ARS
The study on the impact of lysine‐sugar Maillard reaction products on human intestinal microbiome (1042.9)
Authors: Hong Chung‐Oui , Koo Yun‐Chang , Nam Mi‐Hyun , Yang Sung‐Yong , Lee Kwang‐Won
Published: June 2021

Certain ingredients in food caramelized to enhance food flavor during Maillard reaction. However, Maillard reaction products or high‐molecular‐weight materials with different physical and chemical properties were also formed through this reaction. MRPs in food are able to modify the bacterial intestinal ecosystem and metabolite, which is of great importance for the optimal usage of nutrients and maintenance of both intestinal homeostasis and balanced health status of the consumer. The objective of this study was to create lysine‐sugar (glucose, fructose and galactose) conjugates by the Maillard reaction (MRPs) and establish the impact of MRPs on the human intestinal microbiome. We used microbiome models of the human gastrointestinal tract, which have strictly anaerobic conditions and estimated by fluorescence in situ hybridization (FISH). Also we measured fluorescence intensity and analyzed the production of MRPs markers, furosine, HMF and CML and short chain fatty acid (SCFA) metabolite by HPLC. MRPs showed significantly increased level of fluorescence intensity and furosine, HMF and CML formation. The MRPs affect the growth of gut commensal bacteria, particularly lactobacilli and bifidobacteria, whose levels increased significantly.There was a corresponding shift in the bacterial metabolites with increased levels of the SCFAs lactate, propionate and acetate and decreased butyrate. These changes in microbial composition and SCFA may have beneficially impact on the intestinal environment. Finally we have known that what kind of lysine‐sugar conjugates is good for human gastrointestinal health.
Mapping the response of hepatic human fibroblast growth factor 21 promoter to nutrient availability (1045.10)
Authors: Moreau Regis , Yi Xiaohua , Pashaj Anjeza , Xia Mengna
Published: June 2021

The pleiotropic hormone‐like polypeptide, fibroblast growth factor 21 (FGF21), is a major modulator of lipid and glucose metabolism and an exploratory treatment strategy for obesity related metabolic disorders. Recombinant FGF21 (rFGF21) is currently being investigated as therapeutic against high blood lipids and type‐2 diabetes. However, the high costs of producing rFGF21 and the mode of delivery by injection are important limitations to its wide therapeutic use. A cost effective and practical alternative is to stimulate endogenous FGF21 production through diet. Our research shows that naturally occurring dietary compound R‐α‐lipoic acid (LA) stimulates liver Fgf21 expression (+175%) and blood FGF21 levels (+330%) and replicates the adaptive metabolic response to fasting supported by FGF21 in vivo. In the present study, we interrogate FGF21 promoter regulation from HepG2 human hepatocellular carcinoma cells under fed and starved states and in the presence of LA. We employed Formaldehyde‐Assisted Isolation of Regulatory Elements (FAIRE)‐coupled qPCR to quantify nucleosome‐depleted regions in the promoter. Our results provide new insights into how nutrient‐responsive transcription factor binding sites regulate FGF21. The potential impact of this work is that LA may prove to be a safe and affordable means to stimulate FGF21 production.

Grant Funding Source: Supported by UNL Faculty Seed Grant, and Layman Seed Award.
Ocular endoplasmic reticulum stress and inflammation is attenuated by supplementation with muscadine grape polyphenols in vitro and in vivo (1045.2)
Authors: Ha Jung‐Heun , Gu Liwei , Li Qiuhong , Chung Soonkyu
Published: June 2021

Chronic ER stress and inflammation in retinal epithelium are critical risk factors for the pathogenesis of vision‐threatening diseases such as diabetic retinopathy. The aim of this study is to investigate the protective effects of muscadine grape polyphenols (MGP) against ocular ER stress and inflammation. In human retinal cells (ARPE‐19), 50‐100 μg/mL of MGP treatment significantly reduced thapsigragin‐mediated 1) vascular endothelial growth factor (VEGF) secretion, 2) ER stress‐markers of p‐eIF2α, ATF4, p‐JNK, p‐p38 and CHOP, and unfolded protein responses, and 3) early apoptotic cell death (~40 %). It was primarily due to a decrease of intracellular calcium release (p<0.001) and subsequent signal transduction downstream by MGP. In addition, MGP treatment was also effective in attenuating TNFα‐induced proinflammatory gene expressions, such as IL‐1β, IL‐6 and MCP‐1, (p<0.001) in ARPE‐19 cells by inhibiting MAP Kinase‐NFκB axis activation. Moreover, supplementation with MGP for 1 week in C57BL/6 mice led to 1) a remarkable decrease of LPS‐inducible proinflammatory cytokine expression and leukocyte infiltration into eyes, and 2) protection of retinal epithelial leakage against inflammation. Our study suggests that MGP may provide a novel dietary strategy to prevent vision‐threatening retinal diseases.
Peptides derived from extruded amaranth (Amaranthus hypochondriacus) improved the anti‐inflammatory effect in LPS‐induced human THP‐1 and mouse RAW 264.7 macrophages by preventing the activation of NF‐κB pathway (1045.3)
Authors: Montoya‐Rodríguez Alvaro , De Mejia Elvira , Dia Vermont , Reyes‐Moreno Cuauhtémoc , Milán‐Carrillo Jorge
Published: June 2021

The objective was to compare the anti‐inflammatory potential of unprocessed and extruded amaranth pepsin/pancreatin hydrolysates in lipopolysaccharide (LPS)‐induced human THP‐1 and mouse RAW 264.7 macrophages. THP‐1 and RAW 264.7 cells were activated with LPS and treated with unprocessed and extruded amaranth hydrolysates (0.25 to 1.00 mg/mL). Peptides in extruded amaranth hydrolysates displayed antioxidant capacity, angiotensin converting enzyme‐inhibitor activity, antithrombotic and dipeptidyl peptidase‐IV inhibitor activity. Extruded amaranth (1 mg/mL) significantly reduced TNF‐α secretion in THP‐1 and RAW 264.7 cells by 36.5% and 33.5%, respectively; with concomitant reduction in PGE2 (15.4% and 31.4%, respectively), and reduction in COX‐2 (38.1% and 67.6%, respectively). Phosphorylation of IKK‐α was significantly reduced by 52.5% and 88.2% leading to reduced phosphorylation of IκB‐α (86.1% and 66.2%, respectively); resulting in reduction in the expression of p‐65 NF‐κB subunits in the nucleus by 64.2% for THP‐1 cells and 70.7% for RAW 264.7 cells. In conclusion, amaranth hydrolysates inhibited LPS‐induced inflammation in human and mouse macrophages by preventing activation of NF‐κB signaling. Extrusion improved the anti‐inflammatory effect of amaranth hydrolysates in both cell lines, which might be attributed to the production of bioactive peptides during processing.

Grant Funding Source: University of Sinaloa Research Grant
Resveratrol compounds are potent inhibitors of human holocarboxylase synthetase and cause a lean phenotype in Drosophila melanogaster brummer mutants (1045.39)
Authors: Cordonier Elizabeth , Ajam Riem , Camara Teixeira Daniel , Onur Simone , Zbasnik Richard , Döring Frank , Schlegel Vicki , Zempleni Janos
Published: June 2021

Acetyl‐CoA carboxylases (ACC) 1 and 2 are key enzymes in fatty acid (FA) synthesis and repression of FA degradation, respectively. Holocarboxylase synthetase (HLCS) catalyzes the covalent binding of the essential coenzyme biotin to ACCs. We hypothesized that dietary inhibitors of HLCS cause a loss of biotinylated ACCs, leading to a lean phenotype. The PEKISH library of natural compounds was screened for HLCS inhibitor activity in vitro; the screen suggested that resveratrol and its metabolites piceatannol and piceid inhibited HLCS by >90% at nutritionally relevant low micromolar levels (P<0.05 vs. vehicle, by ANOVA & posthoc, n=3). When NIH3T3 fibroblasts were treated with 25 µM piceatannol the levels of biotinylated ACC 1 and 2 decreased by 22% compared to controls. When Drosophila melanogaster brummer mutants, which are genetically predisposed to accumulate body fat, were fed diets supplemented with resveratrol compounds, body fat mass decreased by 50% (P<0.05, n=4) in male and female flies. We conclude that resveratrol compounds are potent inhibitors of HLCS‐dependent biotinylation of ACC 1 and 2, thereby contributing toward body fat loss in Drosophila melanogaster brummer mutants. Future studies will include mammalian models.

Grant Funding Source: Supported by ARD Hatch, NIH, and BMBF.
Glucoraphanin and glucoraphenin suppressed lipid accumulation and ROS production induced by bisphenol A during the adipogenesis in 3T3‐L1 cells (1045.9)
Authors: Lim Jeong‐Ho , Kim Jae‐Hwan , Lee Young‐Jun , Kim Dan‐Bi , Kim Young‐Hyun , Lee Jong Seok , Lee Ok‐Hwan
Published: June 2021

Glucoraphanin and glucoraphenin, one of the glucosinolate, have known as the secondary metabolite of Brassicaceae species such as kohlrabi, chinese cabbage and radish sprout. Recent studies have suggested that the Bisphenol A (BPA), an endocrine disrupting compound, promotes adipose tissue and oxidative stress. The aim of this study was to investigate the inhibitory effect of the glucoraphanin and glucoraphenin on adipogenesis and oxidative stress induced by BPA. Our results show that the BPA promotes the lipid accumulation and ROS production in 3T3‐L1 cells compared to the control cells. Nevertheless, the BPA treated with glucoraphanin and glucoraphenin remarkably inhibits lipid accumulation and ROS production compared with negative control cells that treated with the BPA alone. We also identified that glucoraphanin and glucoraphenin showed down regulation of adipogenic transcription factors. These results indicate that glucoraphanin and glucoraphenin may inhibits both ROS production and lipid accumulation, which protect against oxidative stress that is liked to BPA‐induced adipocyte dysfunction.
Matrix metalloproteinase‐19 deficient fibroblasts display a profibrotic phenotype (1046.4)
Authors: Pardo Annie , Jara Paul , Calyeca Jazmin , Romero Yair , Placido Luis , Yu Guoying , Maldonado Vilma , Selman Moises
Published: June 2021

Idiopathic pulmonary fibrosis (IPF) is a progressive and usually lethal interstitial lung disease of unknown etiology characterized by aberrant activation of epitelial cells that induce the migration, proliferation and activation of fibroblast/myofibroblasts. The resulting distinctive fibroblastic foci are responsible of the excessive extracellular matrix production. We have recently found that Mmp19‐/‐ mice develop an exaggerated bleomycin‐induced lung fibrosis but the mechanisms are unclear. In this study we explored the effect of MMP19 deficiency on fibroblast behavior. Compared with wild type mice, Mmp19‐/‐ lung fibroblasts showed a decreased expression of Th1‐1 (p<0.01), and alpha 1 (I) collagen gene and collagen protein production at baseline and after TGF‐β treatment (p< 0.05). Likewise, they showed a significant increase in growth rate (p< 0.01), and in transmigration over Boyden chambers coated with type I collagen or with Matrigel (p< 0.05). Microarray analysis revealed the upregulation of several profibrotic pathways in Mmp19‐/‐ lung fibroblasts. These findings suggest that the absence of MMP19 induces in lung fibroblasts a profibrotic phenotype.

Grant Funding Source: CONACyT 177641
The role of platelet activation in the expression and function of platelet hyaluronidase‐2 (1046.7)
Authors: Albeiroti Sami , Ayasoufi Katayoun , de la Motte Carol
Published: June 2021

Background: The deposition of hyaluronan (HA), which is a major glycosaminoglycan component of the extracellular matrix, increases during the course of inflammation. The inflammatory HA matrix, which can promote leukocyte binding and recruitment, can be fragmented be the hyaluronidase enzymes, specifically HYAL1 and HYAL2. Our lab has recently shown that human platelets express HYAL2 and that they can degrade HA from the surfaces of intestinal cells into smaller fragments capable of promoting inflammatory cytokine production. Objective: Define the mechanism by which platelets degrade HA and assess the role of platelet activation. Methods: Platelet degradation of purified HA in solution, immobilized HA and cell surface HA was assessed by carbohydrate gel electrophoresis, hyaluronidase assay, and HA ELISA‐like assay, respectively. HYAL2 expression was assessed by immunoblotting and flow cytometry. Results: 1) Platelets degrade purified HA only under acidic pH; 2) Platelets degrade cell surface HA in a HYAL2 dependent manner; 3) Activation of platelets results in increased surface HYAL2 expression and activity; 4) Platelet activation causes an increase in surface expression of HA binding proteins. Significance: Platelets circulate in an activated state during inflammatory diseases. Increased HYAL2 on activated platelets may lead to higher levels of HA fragments that perpetuate inflammation.

Grant Funding Source: Supported by the NHLBI
Cyclin A2 as a regulator of DNA strand stability (1047.6)
Authors: Gygli Patrick , Catacutan Fay , Gokozan Hamza , Schmidt Theresa , Stephens Julie , Palanichamy Kamalakannan , Otero Jose
Published: June 2021

Cyclin A2 regulates replication origin firing during S‐phase. It binds to Cdk1 and Cdk2 to direct phosphorylation of proteins for DNA synthesis. The goals of this study were to determine the role of Cyclin A2 in the prevention of DNA double strand breaks (DSB). We deleted Cyclin A2 in mouse neural stem cells (NSC) using a cre/lox approach. Cyclin A2‐null NSC showed increased BrdU incorporation, as well as increased staining for the mitotic marker pH3. However, the growth rate in these Cyclin A2‐null cells was slowed. To explain this discordance, we examined the DSB marker γH2AX. Cyclin A2‐null NSC show increased levels of γH2AX as compared to Cyclin A2‐preserved cells. When a DSB forms, the cell cycle is arrested until the break is repaired. Rad51, a marker of homologous recombination repair, and 53BP1, a marker of non‐homologous end joining, both localized to the site of DSB in Cyclin A2‐null cells. This indicates that Cyclin A2‐null NSC can repair these breaks. Our results suggest that Cyclin A2 does not function in the repair of DSB, but instead plays a role in DNA strand stability. Because NSC display similarities to glioma initiating cells, our results may be applied to development of a treatment of glioblastoma, which is currently treated by a combination of temozolomide and radiation. Novel therapies for this fatal brain tumor are needed, and Cyclin A2 is a potential target for radiosensitizing therapy for glioblastoma.

Grant Funding Source: 1R21EB017539‐01A1, 8UL1TR000090‐05
Screening of some natural products and nanoparticles for their anti‐breast cancer activities (1048.13)
Authors: Ahweyevu Charles , Taha Murtada
Published: June 2021

Screening of some natural products and nanoparticles for their anti‐breast cancer activities Charles T. Ahweyevu1, Murtada A. Taha2, 1Department of Natural Science, 2Albany State Univeristy, Albany, Ga Cancer refers to a broad group of diseases that involve unregulated cell growth that form malignant tumors, which ultimately lead to death. Breast cancer is the most common type of cancer in women living in Western nations. It is estimated that approximately 1 in 8 North American females will develop breast cancer throughout their lifetime. The best way to battle cancer or any disease is prevention. One of the most researched aspects of chemoprevention is the use of natural products due to its cost effectiveness and little toxic side effects as opposed to other synthetic methods of chemo‐treatment. Herein, we employed both biochemical and microscopic techniques to investigate the anti‐breast cancer activities of seven different preparations namely; Milk thistle extract (MTE), Ginkgo biloba extract (GBE), Selenium (Se), Se‐MTE, Se‐GBE, chitosan, and Se‐chitosan nanoparticles. These natural products have previously shown to have various forms of anti‐cancer activities. Our results showed that only selenium and chitosan have promising anti‐breast cancer activities according to the MTT and microscopy results. Further studies are required to confirm this result.
Examination of ubiquitination and SUMOlyation of GRK4γ (1055.3)
Authors: Nguyen Jenny , Andresen Bradley , Liu Tammy , Alkaddumi Runa , Adams Craig , Liu Jamie , Chiang Steven
Published: June 2021

G protein‐coupled receptor kinases 4γ (GRK4γ) is associated with hypertension. However, the molecular mechanism of action of GRK4γ remains largely unknown. From a proteomic examination of the binding partners of GRK4γ, we identified Gαs, Heat Shock Protein 90 (Hsp90) and an E3 ubiquitin ligase as potential binding partners. Additionally, previous studies suggested that GRK4γ is ubiquitinated. GRK2 and GRK5 are known to be ubiquitinated while SUMOylation, a small Ub‐like modifier protein that also attaches to lysine, has not been extensively studied in any GRKs. Therefore, we tested the hypothesis that GRK4γ can be ubiquitinated and SUMOylated. To demonstrate Ubiquitination and SUMOylation, we immunoprecipitated (IPed) GRK4γ and blotted for Ub or SUMO. The data clearly showed that GRK4γ is ubiquitinated but not SUMOylated. Interesting, GRK4γ ubiquitination is significantly enhanced after stimulation of MG132, a cell‐permeable proteasome inhibitor, but remains unchanged in the presence of two other proteasome inhibitors (Lactacystin and proteasome inhibitor I) which is counter to the previous studies. Further studies will be conducted to identify the role of GRK4γ activity in relation to GRK4γ ubiquination as well as identify the ubiquitination sites on GRK4γ and the role that ubiquitination plays in GRK4γ activity.
Derivatives of 1,2,3‐triazole lead found to be selective and potent agonists at the α7 nicotinic acetylcholine receptor (1059.8)
Authors: Chen Bill , Arunrungvichian Kuntarat , Taylor Palmer , Vajragupta Opa , Fokin Valery , Yamauchi John , Nemecz Akos
Published: June 2021

The α7 nicotinic acetylcholine receptor (nAChR), which is a member of pentameric ligand‐gated ion channel receptors, is a novel candidate target for treatment of Alzheimer’s disease, schizophrenia, other CNS disorders, peripheral pain and inflammation. Proteins homologous to the extracellular domain of nAChRs from Aplysia californica (Ac) and Lymnaea stagnalis (Ls)acetylcholine binding proteins (AChBPs) are expressed and purified as templates for drug design, x‐ray crystallography, and binding affinity assays. Based on the binding poses of α 7‐nAChR agonists obtained from x‐ray crystallography and previous in vitro screening of 1,2,3‐triazole derivatives, approximately 50 molecules were designed in order to enhance the selectivity and efficacy profiles of lead compounds. Lead optimization of 1,2,3‐triazole products started with an azide building block modified to obtain a suitable cationic center followed by a variation of alkyne building blocks to fit a hydrophobic region in the binding pocket of α 7 nAChRs. All modified compounds were synthesized by copper‐catalyzed azide‐alkyne cycloaddition (CuAAC) or click chemistry. They were characterized by binding affinity and functional properties using the AChBPs and fluorescent cell‐based assays using a genetically encoded Ca2+ sensor. The results showed that both selectivity and efficacy profiles of the synthesized congeners were superior to the lead compound. IND1, a six‐membered monocyclic ring with a 2 methylene linker, is the most potent and selective α7‐nAChR agonist in this series. Its potency is 3‐fold greater than the lead compound with high selectivity for α7‐nAChR over the α4β2‐nAChR and 5HT3A receptors. Three compounds (IND8, QND2, and QND8) from a quinuclidine azide building block are the most potent in this series. Their potencies are increased 20‐fold compared to the lead compound while the tropane selectivity is maintained.
Aquaporin‐1 translocation and degradation mediates the water transportation (1062.2)
Authors: Li Xuejun , Zhang Jianzhao , An Yu
Published: June 2021

Acetazolamide (AZA), a prototype of carbonic anhydrase (CA) inhibitor, acts as a diuretic agent with its site of action on renal proximal tubules. These nephron segments also contain aquaporin‐1 (AQP1) water channels, which is required for the formation of concentrated urine. In the present study we examined the role of AQP1 on the diuretic effect of AZA. By using in vivo and in vitro models, we found that the AZA increased the urine volume from 8 hrs to 14 days. However, total CAs activity was decreased within 8 hours and then gradually recovered to the normal level and the CAs protein expression was increased at the beginning of AZA treatment. AQP1 was significantly reduced after AZA administration for 7 days. When combined with NaHCO3, AZA had a more pronounced diuretic effect. But it didn’t induce CA isozymes expression and the CAs activity anymore. However, AQP1 expression was still inhibited from day 7. AZA increased AQP1 ubiquitination in the HK‐2 cell membrane followed by the degradation induced by proteasome. By using the coimmunoprecipitation and the surface plasmon resonance (SPR) methods, we found that AZA facilitated AQP1 translocation onto cell membrane by promoting the interaction of AQP1 and myosin heavy chain (MHC). And this process was mediated by ERK and myosin light chain kinase (MLCK) pathway. These findings indicate that AZA promotes AQP1 translocation and degradation, which may partially mediates the effect of AZA.

Grant Funding Source: the National Natural Science Foundation of China (No. 91129727, 81020108031, 30973558)
Transient knockdown of SULT4A1 induces upregulation of phototransduction proteins (1064.19)
Authors: Crittenden Francis , Ethen Cheryl , Wu Zhengliang , Chen Dongquan , Kraft Timothy , Parant John , Falany Charles
Published: June 2021

Since it’s identification in 2000 by Falany et al, sulfotransferase (SULT) 4A1 has presented an enigma to the field of drug metabolism. The SULTs are a superfamily of enzymes responsible for the sulfation of a wide variety of endogenous and exogenous compounds. They catalyze a Phase II conjugation reaction in which a sulfonate group is transferred onto the substrate primarily for conjugation to hydroxyl groups, leading to increased water solubility and excretion. Key structural features shared between SULT4A1 and other SULTs include the KXXFTVXXXE dimerization domain, active site histidine, and conserved PAPS binding site. SULT4A1 is primarily expressed in neurons of the central nervous system and is highly conserved in vertebrates, suggesting an important, conserved function within the nervous system. A function or enzymatic activity for SULT4A1, however, has yet to be elucidated. The goal in this study is to identify a function for this highly conserved orphan SULT. Human SULT4A1 and zebrafish SULT4A1 exhibit 88.4% sequence identity, making zebrafish an excellent model organism for genetic manipulation and functional characterization of this protein. Splice blocking morpholino oligonucleotides (MOs) were used to suppress SULT4A1 expression in zebrafish larvae over the first several days of development. Knockdown (KD) efficacy was demonstrated by qPCR and immunoblot analysis. RNA‐seq was used to investigate the effects of the inhibition of SULT4A1 expression on the transcriptome of the KD larvae. Analysis of the RNA‐seq data using Ingenuity Pathway Analysis showed the phototransduction pathway to be significantly dis‐regulated by SULT4A1 KD. A total of 13 genes known to be involved in phototransduction were shown to be up‐regulated by SULT4A1 knockdown. qPCR was used to validate up‐regulation of selected genes identified by RNA‐seq. This study is the first to identify a cellular process whose regulation appears to be associated with SULT4A1 expression.

Grant Funding Source: Supported by NIH grant R21MH095946
Piperine and ketoconazole increase plasma domperidone concentrations in the rat (1064.2)
Authors: Alhumayyd Mohammad , Bukhari Ishfaq , Almotrefi Abdulrahman
Published: June 2021

This study was carried out to investigate the effect of piperine, a major pungent constituent of the black and red peppers and ketoconazole on the pharmacokinetics of domperidone. Animals received a single dose of domperidone(20mg/kg, p.o.) alone or together with piperine (60mg/kg, p.o.) or ketoconazole(50 mg/kg, p.o.) daily for 5 days. Plasma samples were collected at 0.25, 0.5, 1.0, 2.0, 3.0, 4.0, 6.0, 8.0, 10.0 and 12h after drug administration. The concentrations of domperidone in the plasma were measured using an HPLC method. The concomitant administration of piperine or ketoconazole with domperidone resulted in a significant (p<0.05) increase in the maximum plasma concentration(Cmax), the mean area under the plasma concentration‐ time curve(AUC), and the elimination half‐life(t1/2) of domperidone as compared to those obtained for domperidone alone. These results suggest that an important pharmacokinetic interaction may occur if piperine or ketoconazole are administered concurrently with domperidone.
Glycosylation of protease‐activated receptor‐1 regulates G12/13 versus Gq signal pathway bias (1066.13)
Authors: Smith Thomas , Soto Antonio , Chen Buxin , Trejo JoAnn
Published: June 2021

The objective of this study was to examine the role of N‐linked glycosylation in protease‐activated receptor‐1 (PAR1) biased signaling. Thrombin cleaves the N‐terminus of PAR1, generating a new N‐terminal domain that functions as a tethered ligand by binding intramolecularly to extracellular loop 2 (ECL2) to elicit signaling through multiple G protein subtypes including Gq, G12/13 and Gi. Activated PAR1 displays biased signaling in response to different activating proteases. We discovered that N‐linked glycosylation of PAR1’s ECL2 dictates differential coupling of PAR1 to Gq versus G12/13 signaling. Thrombin activation of PAR1 WT caused robust G12/13‐dependent RhoA activation and stress fiber formation, whereas these responses were diminished in a PAR1 mutant lacking glycosylation. Accordingly, activated PAR1 WT associated more robustly with G12/13 proteins than did the PAR1 NA ECL2 glycosylation mutant. In contrast, activated PAR1 NA ECL2 mutant exhibited a greater capacity to associate with Gq, elicit Gq signaling, and promote Gq‐dependent cellular proliferation compared to WT receptor. These findings suggest that N‐linked glycosylation at ECL2 is critical for stabilizing different PAR1 active conformations that facilitate coupling to distinct G protein subtypes. Thus, we hypothesize that N‐linked glycosylation at ECL2 is critical for regulating PAR1 biased signaling.

Grant Funding Source: Supported by R01 GM090689 and F31 HL116187‐01A1
Ubiquitination of PAR1 nucleates a non‐canonical p38 signaling pathway to regulate thrombin‐induced vascular leakage (1066.15)
Authors: Grimsey Neil , Smith Thomas , Le Philip , Trejo JoAnn
Published: June 2021

Thrombin activation of the protease‐activated receptor‐1 (PAR1), a GPCR, triggers inflammation, endothelial barrier permeability and vascular leakage. Multiple signaling pathways contribute to vascular leakage including activation of the p38 MAP kinase. However, the precise mechanism of PAR1‐stimulated p38 activation is not known. In this study, we examined the role of PAR1 ubiquitination in the activation of p38 signaling and endothelial barrier dysfunction. We demonstrate that thrombin activation of PAR1 stimulates p38 signaling via a novel ubiquitin/TAB1/TAB2‐induced p38 autophosphorylation pathway. Thrombin‐induced ubiquitination of PAR1 is mediated by the activation and recruitment of the E3 ubiquitin ligase NEDD4‐2. The adaptor protein TAB2 then binds to ubiquitinated PAR1 and recruits another adaptor TAB1. TAB1 also binds and triggers p38 activation through autophosphorylation and not the MKK3/MKK6 pathway. This pathway is conserved for ADP‐dependent activation of the P2Y1 purinergic receptor. We further demonstrate that thrombin activation of PAR1 requires non‐canonical p38‐signaling pathway to regulate barrier permeability of cultured human endothelial cells. Collectively, these data suggest that a subset of GPCRs utilize ubiquitin to assemble and activate a TAB2 and TAB1 non‐canonical p38 signaling complex that regulates endothelial barrier permeability and vascular leakage.

Grant Funding Source: Supported by NIH R01 HL073328 and Western States Affiliate AHA Postdoctoral Fellowship
Understanding the mechanism of biased agonism at chemokine receptors (1066.17)
Authors: Rajagopal Sudarshan , Bassoni Daniel , Campbell James , Gerard Norma , Gerard Craig , Wehrman Tom
Published: June 2021

Chemokines display considerable promiscuity with multiple ligands and receptors shared in common. Their receptors are G protein‐coupled receptors (GPCRs), which are capable of signaling with different efficacies to their multiple downstream pathways, a phenomenon referred to as biased agonism. Biased agonism has been primarily reported as a phenomenon of synthetic ligands and the biologic importance of such signaling is unclear. To assess the presence of biased agonism that may underlie differential signaling by chemokines targeting the same receptor, we performed a pharmacologic analysis of a set of chemokine receptors using assays for G protein signaling, β‐arrestin (βarr) recruitment and receptor internalization. We found that chemokines targeting the same receptor can display marked differences in their efficacies for G protein‐ or βarr‐mediated signaling or receptor internalization. Moreover, some chemokine receptors display significant bias for all their ligands, acting as “biased receptors”. Ligand bias correlates with changes in leukocyte migration, consistent with different mechanisms underlying the signaling downstream of these receptors induced by their ligands. These findings demonstrate that biased agonism is a common biological mechanism for generating qualitatively distinct patterns of signaling via the same receptor in response to different endogenous ligands.

Grant Funding Source: Supported by HL07101‐34 (SR); R21AI092388, R21AI097468 (JC); R01AI039795‐14, 5R01HL51366‐15 (CG)
The ubiquitin ligase Deltex‐3L regulates endosomal sorting of the G protein‐coupled receptor CXCR4 (1066.18)
Authors: Holleman Justine , Marchese Adriano
Published: June 2021

G protein‐coupled receptor (GPCR) sorting into the degradative pathway is important for attenuating signaling. Upon agonist activation of the chemokine receptor CXCR4, a GPCR, it is rapidly ubiquitinated, internalized to endosomes and sorted to lysosomes via the endosomal sorting complex required for transport (ESCRT) pathway. ESCRT‐0 ubiquitination is linked to the efficiency with which CXCR4 is sorted for lysosomal degradation, however mechanistic insight is lacking. Here, we show for the first time that the RING‐domain E3 ubiquitin ligase Deltex‐3‐like (DTX3L) mediates CXCR4 sorting from early endosomes to lysosomes. Using several biochemical and immunochemical techniques including fixed cell confocal immunofluoresence microscopy, co‐immunoprecipitation and in vitro ubiquitination assays, we show that DTX3L localizes to early endosomes upon CXCR4 activation and interacts directly with and inhibits the activity of the E3 ubiquitin ligase atrophin‐1 interacting protein 4 (AIP4). This serves to limit the extent to which ESCRT‐0 is ubiquitinated and is able to sort CXCR4 for lysosomal degradation. Therefore we have defined a novel role for DTX3L in GPCR endosomal sorting and propose that DTX3L may play a broad role in endosomal sorting. In addition, our data reveal an unprecedented link between two distinct E3 ubiquitin ligases to control the activity of the ESCRT machinery.

Grant Funding Source: GM075159