bluuebunny/crossref_metadata_2025 huggingface dataset:
The effect of nitrogen treatment on the anthacyanine and polyphenols content and ORAC factor of Aronia melanocarpa grown in Maryland (1042.5)
Authors: Ndam Tina , Aroh Blessing , Volkis Baruch , Ristvey Andrew , Mathew Sadip , Volkis Victoria
Published: June 2021

Black chokeberry or Aronia melanocarpa is a small fruit‐bearing shrub in the rose family. Although it is native to Maryland, its range nowadays is from Newfoundland, west to Ontario, south into Alabama, and east to Georgia, and hardy to Zone 3. Aronia is a landscape quality plant, susceptible to few pests and diseases that persist in soils and temperate climatic conditions. It is an ideal candidate for organic fruit production. The Aronia fruit has nutraceutical qualities, heightening its marketability and sales potential as a value‐added product. There is currently great interest in fruits and vegetables that contain high concentrations of flavonoids, considered potent antioxidants. Some recent studies have implicated the relationship between in‐field plant nutrient fertility and antioxidant production in aronia. Here we present the data for the antioxidant content of Aronia melanocarpa as a function of the difference of age, amount of time spent in the sun or shade, and nitrogen treatment levels of crops. We have shown that the level of nitrogen treatment in the soil influences the antioxidant capacity significantly. Detailed measurements and analysis of anthocyanin and polyphenols as well as ORAC factor will be presented and discussed. The aim of the project is to determine the treatment that produces the highest capacity of antioxidants in aronia.

Grant Funding Source: Supported by Grant T34GM008411, from the National Institute of General Medical Sciences
Marginal zinc intake impairs mammary gland involution, increases oxidative stress and disrupts ductal integrity abrogating the protective effect of lactation on breast tumorigenesis in a mouse model (1043.3)
Authors: Mack Ronald , Bostanci Zeynep , Alam Samina , Lee Sooyeon , Soybel David , Kelleher Shannon
Published: June 2021

Breast cancer is the most common cancer among women. While breastfeeding can reduce breast cancer risk, several studies found no protective effect. We showed that marginal zinc (Zn) intake compromises mammary gland (MG) expansion and function during lactation and is associated with hallmarks of pre‐neoplastic lesions such as collagen deposition, macrophage infiltration, and ductal hyperplasia in the MG of nulliparous mice. Herein, we hypothesized that marginal Zn deficiency abrogates the protective effect of breastfeeding on breast cancer risk. C57BL/6 mice were fed a Zn adequate (ZA; 30mg Zn/kg) or Zn deficient (ZD; 15mg Zn/ kg) diet. Mice were bred, and 2 wk post‐weaning were gavaged with 7,12‐dimethylbenzanthracene (DMBA; 1mg/0.1mL corn oil) or corn oil (control) weekly for 4 wk. Tumor progression was monitored for 24 wk. ZD mice had lower matrix metalloproteinase 2 activity (p<0.01) and insufficient involution, shorter tumor latency (p=0.05), and tended to have greater palpable (35% vs 12% p=0.137) and non‐palpable tumors (p=0.08). ZD mice had greater oxidative stress, e‐cadherin expression and collagen deposition in the MG. Moreover, mammary tumors in ZD mice accumulated Zn and had characteristics of a more invasive phenotype. Our results suggest that consuming inadequate dietary Zn may impair post‐ lactational regression and abrogate the protective effect of breastfeeding on breast cancer.

Grant Funding Source: Intramural support, Departments of Surgery and Nutritional Sciences, Penn State University
The relationship between toenail Zn and glycemic control in healthy adults (1043.6)
Authors: Lee Okhee , Chung Yongsam , Moon Jongwha
Published: June 2021

Zinc plays important roles for the maintenance of immunity and anti‐oxidative capacity. It participate in many enzymatic reaction and known to protect from inflammation. Therefore its adequacy is important in decreasing the risks of various degenerative diseases such as diabetes or cardiovascular disease. Toenail Zn content is recognized as indicator for their long‐term exposure. Furthermore, the toenail is non‐invasive human sample, many countries reported various toenail zinc in their populations. In this study we measured the toenail zinc content in healthy young adults living around Keunggido in Korea by Instrumental Neutron Activation analysis(INAA)‐method using research reactor. Also we assessed the relationship with indicators for glycemic control such as insulin, glucose. The toenail Zn content showed a significant difference by gender. The toenail Zn content correlated positively with serum insulin, but negatively with serum glucose level. However, any relationship between toenail Zn and HOMA‐IR was observed. The toenail Zn content in young adults showed any association with neither hs‐CRP nor neutrophil proportion in blood. In conclusion, this study suggested that the long term Zn status in healthy young adults is associated with status of glycemic control, but not with inflammatory marker.

Grant Funding Source: Korea National Research Foundation
Changes of gene expressions in zinc deficiency and diet restriction in rats (1043.8)
Authors: Konomi Aki , Yokoi Katsuhiko
Published: June 2021

We previously reported that both zinc deficiency and diet restriction affect total RNA concentration and expression of Gapdh and Actb in kidney. In this study, we have examined whether zinc deficiency and diet restriction affect Crebbp and Taf5l in kidney. Thirty 3‐week‐old male Sprague‐Dawley rats were assigned into 3 dietary treatment groups of 10 for the four‐week study: Control group (AIN‐93G), Zn‐deficient group (4.5 mg Zn/kg) and pair‐fed group (AIN‐93G). Kidney was separated into cortex and medulla for RNA isolation. Quantitative real time RT‐PCR using TaqMan proves was performed for mRNAs of Crebbp and Taf5l. Difference between groups was statistically analyzed by Fisher's PLSD. P value less than 0.05 was considered significant. Kidney cortex Crebbp per total RNA was not changed by the treatments. Kidney medulla Crebbp per total RNA was significantly increased by zinc deficiency and diet restriction, respectively. Kidney cortex Taf5l per total RNA was significantly increased by zinc deficiency and diet restriction, respectively. Kidney medulla Taf5l per gram tissue was significantly decreased by zinc deficiency and diet restriction, respectively. These results suggest that zinc deficiency and diet restriction differently affect expression of Crebbp and Taf5l in kidney cortex and medulla. This work was supported by KAKENHI grant‐in‐aid for Young Scientists (B) (23700928).
Cytokine release by Jurkat T‐lymphocytes is modulated following treatment with polyphenols or Lactobacillus rhamnosus probiotic extracts (1044.10)
Authors: Richardson Sian , Ford Christopher , Crozier Alan , McArdle Francis , McArdle Anne , Jackson Malcolm
Published: June 2021

Quercetin and (‐)‐epigallocatechingallate (EGCG) are dietary polyphenols claimed to exert anti‐inflammatory effects. Jurkat T‐lymphocytes were incubated with 5μM quercetin or ECGC for 48h, with or without 25ng/ml phorbol myristoyl acetate and 5μg/ml phytohaemagglutinin (PMA/PHA) at 24h to induce cytokine release. Inflammatory cytokines interleukin 2 (IL2), 8 (IL8) and tumour necrosis factor α (TNFα) were measured. Unexpectedly, polyphenols had no significant anti‐inflammatory effects and EGCG increased IL8 release from stimulated cells by 174±18%. Polyphenols are digested to phenolic acids by the colonic microflora, thus we hypothesised anti‐inflammatory effects may be mediated by these metabolites. Quercetin and EGCG were fermented with normal gut bacteria (E. coli) or probiotic strain (L. rhamnosus) for 6h and phenolic acids were then isolated by solid phase extraction. Jurkat cells were incubated with the extracts for 48h with or without PMA/PHA stimulation at 24h.

E. coli extract increased TNFα release from stimulated cells by 268±51% while L. rhamnosus extract reduced IL8 release from unstimulated cells by 34±2%. EGCG fermented with L. rhamnosus increased IL8 release from stimulated cells by 128±16%, consistent with effects of unfermented EGCG. The findings showed anaerobic bacterial fermentation did not modify polyphenol activity, but supported anti‐inflammatory effects of L. rhamnosus.

Grant Funding Source: Supported by BBSRC DRINC
Strain‐specific profiles of hepatic xanthones in mice chronically fed α‐mangostin, the most abundant xanthone in mangosteen fruit (1044.7)
Authors: Chitchumroonchokchai Chureeporn , Mah Eunich , Gutierrez‐Orozco Fabiola , Bruno Richard , Failla Mark
Published: June 2021

Little is known about the tissue distribution and metabolism of ingested mangosteen xanthones . We investigated hepatic content and profiles of xanthone metabolites in four murine strains (C57BL/6, Athymic nude, C3H and BALB/c mice; n = 5 group) fed AIN‐93G diet containing 0.1% α‐MG‐rich mixture of xanthones for 4 weeks. α‐MG accounted for 88.5% of dietary xanthones with γ‐MG, garcinone D and others representing 3.7%, 3.0% and 4.8%, respectively. Estimated xanthone intake during the feeding trial was approximately 300 nmoL/g BW. Mice were fasted for 4h before necropsy. Hepatic xanthones were extracted without and with glucuronidase/sulfatase to assess free and conjugated compounds. The presence and concentrations of xanthones in diet and liver was determined by LC‐MS. Hepatic concentration of total xanthones was greatest in C57BL/6 mice (15 ± 6 nmol/g) followed by C3H (9.4 ± 5 nmol/g), Athymic nude (8.8 ± 4 nmol/g), and BALB/c mice (3.1 ± 0.8 nmol/g). α‐MG accounted for 77‐82% of hepatic xanthones in C57BL/6, C3H and Athymic nude mice, but only 55% in BALB/c mice. Relative concentrations of garcinone D and 9‐hydroxy‐calabaxanthone in liver accounted for 9‐15% and 6‐14% of total xanthones, respectively, which exceeded their percentage in diet. Approximately 80% of xanthones in liver were phase II conjugates. Results suggest that dietary xanthones and their metabolites are present in murine liver, that various dietary xanthones are biotransformed, preferentially accumulated in liver and/or eliminated, and that absorption and/or whole body metabolism of these compounds appears is/are influenced by genotype.

Grant Funding Source: CAFFRE and OSU CCC MCC
Anti‐inflammatory bioactives in Alaskan seaweed: implications in metabolic syndrome (1045.14)
Authors: Kellogg Joshua , Esposito Debora , Komarnytsky Slavko , Lila Mary Ann
Published: June 2021

A dominant factor underlying the development of metabolic syndrome is abdominal obesity, which is associated with chronic inflammation in adipose tissue. Alaska Native populations, who suffer disproportionately high rates of obesity, have shifted away from traditional dietary foods, like seaweeds, that are a rich source of polyphenols with potential to counteract metabolic syndrome. In this work, Alaskan seaweeds Fucus distichus (FD), Saccharina latissima (SL), Saccharina groenlandica (SG), Alaria marginata (AM), Pyropia fallax (PF), and Ulva lactuca (UL) were evaluated for their potential to ameliorate symptoms of obesity‐linked inflammatory pathways. High levels of phenolics were discovered in ethyl acetate and butanol fractions of AM, FD, and SG (326.8 ‐ 557.2 μg phloroglucinol equivalents (PGE)/mg extract). Fractions of AM, FD, SG, and SL reduced nitric oxide levels in LPS‐induced RAW 264.7 macrophages to basal levels, and assays with 3T3‐L1 adipocytes demonstrated that organic fractions of AM, SL, and UL decreased triglyceride accumulation 13.3 ‐ 24.7%. Subsequent analysis of AM fractions revealed reductions in mRNA expression levels for the inflammatory adipokines IL‐10, MCP‐1, iCAM, and TNF‐α by 92.6%, 50.7%, 67.3%, and 65.4%, respectively. These results support the potential of Alaskan algae to reduce inflammation and obesity in adipose tissue.

Grant Funding Source: Supported by the UNC General Administration and NCSU's Plants for Human Health Institute
Antioxidant and anti‐proliferative capacity of extractable phytochemicals from tropical fruits (pineapple, mango and papaya) (1045.18)
Authors: Olivas‐Aguirre Francisco , Velderrain‐Rodriguez Gustavo , Wall‐Medrano Abraham , González‐Aguilar Gustavo , Robles‐Zepeda Ramón , Astiazarán‐García Humberto
Published: June 2021

Fruits and vegetables (F&V) are universally promoted as healthy and prospective cohort studies support the evidence that their consumption is associated with a lower risk of all‐cause mortality and death from cardiovascular disease & cancer. In particular, tropical fruits such as Mango (MA, Mangifera indica cv. Ataulfo), papaya (PM, Carica papaya cv. Maradol) and pineapple (PE, Anana comosus cv. Esmeralda) have a vast array of phytochemicals with antioxidant and anti‐carcinogenic potential. The aim of this study was to compare the antioxidant profile and anti‐proliferative capacity of hydro‐methanolic (20:80 v/v) extracts obtained from freeze‐dried pulps of these fruits. MA had 2.3 & 2.7 more phenolic compounds (9.9±0.2 mgGAE/g DM) and 0.6 & 54.6 more ascorbic acid (956±2.2 mg/g DM) than PM or PE, respectively while PE had 0.2 & 2.8 more flavonoids (0.5±0.1 mgQE/g DM) than MA or PM. MA was more effective in scavenging DPPH and ABTS radicals (IC50 5.0 and 5.6 mg/mL) than PM (IC50 18.0 and 17.5 mg/mL) or PE (IC50 29.0 and 48.0 mg/mL). MA was more effective to inhibit the proliferation of RAW264.7 (mouse leukaemic monocyte/macrophage), L929 (mouse fibroblast) and HeLA (Cervix adenocarcinoma) cells at IC50 = 90, 138 and 187 mg/mL, respectively than PE (IC50 187, >200 and >200 mg/mL) or PM (IC50 >200 mg/mL) as assayed by the MTT Proliferation assay. However, all three fruit extracts were no effective (>200 mg/mL) to inhibit A549 Human lung adenocarcinoma cells. The specific phenolic profile and antioxidant capacity is related to the anti‐proliferative properties of these three tropical fruits.
Trigonelline and 3,3‐diindolymethane regulate cell growth in non‐malignant colonocytes via estrogen signaling (1045.21)
Authors: Yoo Gyhye , Allred Clinton
Published: June 2021

Epidemiological and clinical data suggest that estrogen exposure reduces the incidence of colorectal cancer. However, use of traditional hormone replacement therapy containing estradiol is controversial as it is associated with an elevated incidence of breast cancer. As such, there is considerable interest in identifying novel, plant‐derived, phytoestrogens that can mimic the protective actions of estrogen in the colon. Trigonelline (Trig) and 3,3‐diindolylmethane (DIM) are phytoestrogens which have distinct chemical structures compared to other well described phytoestrogens and both compounds have been reported to elicit estrogenic responses without directly interacting with the binding domain of the estrogen receptor (ER). In this study, we evaluated the ability of these compounds to influence non‐malignant colonocytes in vitro which we have demonstrated is predictive of chemopreventative effects in vivo. Both Trig and DIM suppressed cell growth of young adult mouse colonocytes (YAMCs). Likewise, Trig and DIM disrupted cell cycle progression in the G0/G1 phase and Trig induced programmed cell death. Co‐treatment with ICI 182,780, an ER antagonist, inhibited the effects of Trig but not DIM. Interestingly, DIM did induce transcriptional activity of the ER. In addition, both compounds altered gene expression associated with apoptosis and cell proliferation in unique ways. In conclusion, Trig and DIM alter cell physiology and gene expression in YAMCs via unique estrogenic actions and these data suggest that consumption of these dietary compounds may reduce risk of colon cancer development.
(‐)‐Epigallocatechin 3‐gallate decreases markers of inflammation in a mouse model of ulcerative colitis (1045.24)
Authors: Bitzer Zachary , Lambert Joshua
Published: June 2021

Ulcerative colitis (UC) is an inflammatory bowel disease that manifests as constant/cyclic episodes of colonic inflammation, oxidative stress, increased intestinal permeability, and ulceration followed by periods of quiescence. UC increases risk of colon cancer. Green tea polyphenols, such as epigallocatechin‐3‐gallate (EGCG), have been shown to inhibit oxidative stress in animal and cell studies. Here, we examined the anti‐inflammatory effects of EGCG in the dextran sulfate sodium (DSS)‐induced CF‐1 mouse model of UC. Following induction of colitis by treatment with 1.5% DSS for 7 days, mice were treated with EGCG (0.32%) as the sole source of drinking fluid for 3 days. In the colon homogenate, inflammatory cytokine levels of IL‐6 and TNF‐α were decreased by 50% and IL‐1β by 25% as a result of the EGCG administration. Plasma inflammatory markers and endotoxin levels were also decreased. These results suggest that EGCG can decrease both inflammatory response and intestinal permeability in the DSS‐induced UC mouse model. Ongoing mechanistic studies are focused on determining the effect of EGCG on gut barrier function.
Kaempferol inhibits endoplasmic reticulum stress‐associated mucus hypersecretion in airway epithelial cells and ovalbumin‐sensitized mice (1045.32)
Authors: Gong Ju‐Hyun , Kang Young‐Hee
Published: June 2021

Mucus hypersecretion is an important pathological feature of chronic airway diseases, such as asthma and pulmonary diseases. MUC5AC is a major component of the mucus matrix forming family of mucins in the airways. The initiation of endoplasmic reticulum (ER)‐mediated stress responses contributes to the pathogenesis of airway diseases. The present study investigated that ER stress was responsible for airway mucus production and this effect was blocked by kaempferol. Nontoxic kaempferol at 蠄20 μM inhibited tunicamycin‐promoted induction of XBP‐1‐splicing and ER chaperones such as GRP78bip and HSP70 in human bronchial airway epithelial BEAS‐2B cells. Kaempferol disturbed the tunicamycin activation of the ER transmembrane sensor IRE1 and JNK resulting in mucus production with MUC5AC induction. The in vivo study further explored the demoting effects of kaempferol on airway mucus secretion in BALB/c mice sensitized with OVA. Oral administration of 10 mg/kg kaempferol suppressed mucus secretion observed in the lung of OVA‐challenged mice. These results demonstrate that dietary kaempferol alleviated asthmatic mucus hypersecretion through blocking bronchial epithelial ER stress via inhibition of IRE1‐JNK activation. Therefore, kaempferol may be a potential therapeutic agent targeting mucus hypersecretion‐related pulmonary diseases.
Aplysin administration reduces alcoholic liver injury by inhibiting nitric oxide production (1045.42)
Authors: Liang Hui , Liang Hui
Published: June 2021

Aplysin administration reduces alcoholic liver injury by inhibiting nitric oxide production Liang Hui1, PhD, Ge Na1, Liu Ying1, Ai‐guo Ma1, PhD, Dayong Wu2, MD, PhD, Wen‐cheng Wang1: 1The Institute of Human Nutrition, Medical College, Qingdao University; 2Nutritional Immunology Laboratory, Jean Mayer USDA Human Nutrition Research Center on Aging, Tufts University Alcoholic liver disease is a major cause of morbidity and mortality worldwide. Oxidative stress is known to contribute to alcohol‐induced liver injury. We previously showed that Aplysin isolated from the red alga had hepatoprotective effect in an alcoholic liver injury animal model. To help understand the mechanism underlying this effect, in current study we determined how aplysin affected synthesis of nitric oxide (NO), and inducible NO synthase (iNOS) activity and enzyme protein levels in rats with alcoholic liver injury. Rats were orally administered different doses of Aplysin (0, 50, 100, and 150 mg/kg BW) and ethanol (8‐12 ml/kg BW) or saline (control) by daily gavage. After 6 wk, rats were sacrificed and blood and livers were collected for analysis. Results showed that Aplysin treatment reduced alcohol‐induced hepatic ultrastructural lesions and elevation in serum levels of aminopherase, NO, and NOS activity in a dose‐dependent manner. In addition, Aplysin at medium and high doses inhibited iNOS expression in liver. These results suggest that increased NO and iNOS are associated with chronic liver injury, and Aplysin’s hepatoprotective effect may be mediated through its inhibitory effect on iNOS expression and activity leading to decreased NO output.
Purple‐fleshed potato, even after processing, prevents and reverses high‐fat diet elevated colonic‐mesenteric fatősystemic inflammation cascade in pig model (1045.47)
Authors: Radhakrishnan Sridhar , Kim Sung , Reddivari Lavanya , Vanamala Jairam
Published: June 2021

High‐fat diet (HFD) elevated colonic/adipose tissue inflammation strongly correlates with lipopolysaccharide (LPS) induced TLR‐4 activation. We have previously shown that purple‐fleshed potatoes (PP) suppress colonic/systemic oxidative stress markers in HFD consuming pigs. We hypothesized that PP, even after processing; suppress HFD induced colonic/mesenteric fat inflammation via suppression of LPS/TLR‐4 signalling compared to white‐fleshed potatoes (WP). We performed 2 studies ‐ a prevention study where 64 pigs consumed one of the 8 diets: Standard diet (SD), HFD and HFD supplemented with raw/baked/chips PP or WP (10% w/w) for 13 wk; and a reversal study where pigs (12 wk on HFD) consumed HFD containing 10/20% PP or WP chips for 5 wk (n=8). In the prevention study, all potato diets suppressed (P < 0.05) HFD elevated colonic/mesenteric fat expression of TLR‐4 and downstream NF‐κB and TNF‐α (qPCR), and serum IL‐1β (Immunoassay). In the reversal study, colonic/mesenteric fat expression of TLR‐4, NF‐κB and TNF‐α were consistently suppressed only in PP group (not WP) compared to HFD control (P < 0.05), suggesting that time of intervention is important. In addition, PP extracts suppressed LPS or TNF‐α induced NF‐κB activation in Caco‐2 cells compared to WP. Collectively, these results suggest that PP were more potent in reversing the HFD elevated colonic‐mesenteric fat‐systemic inflammatory cascade.

Grant Funding Source: Supported by USDA‐NIFA NRI Grant 2009‐55200‐05197
Mango and pomegranate polyphenolics in the modification of microbiota and short chain fatty acids in DSS‐induced colitis (1045.6)
Authors: Kim Hyemee , Minamoto Yasushi , Markel Melissa , Suchodolski Jan , Talcott Stephen , Mertens‐Talcott Susanne
Published: June 2021

Dysregulation of the interactions between gut microbiota and the immune system is speculated to be involved in triggering colitis. Gut microbiota can be modulated by dietary compounds such as fiber and polyphenolics. To evaluate the effects of mango and pomegranate polyphenolics on fecal microbiota and SCFAs production, rats were administered control, mango, or pomegranate juice, and were exposed to three cycles of 3% DSS followed by 2‐week recovery period. Fecal samples were collected to characterize the fecal microbiota composition using selected representative genus by qPCR and to measure SCFAs concentration. Mango and pomegranate juice decreased the level of pro‐inflammatory cytokines, such as IL‐1b and GM‐CSF in serum, and induced the level of IL‐10. Pomegranate juice induced compositional changes in the fecal microbiota. In particular, the anti‐inflammatory Ruminococcaceae significantly was increased by pomegranate juice in the feces but the concentrations of SCFAs were not significantly affected. In response to the mango juice, fecal SCFAs isovalerate and valerate were increased, while there was no significant change in the composition of the intestinal microbiota at the genus level. Polyphenolics decreased the TLR4 expression, but only mango increased the GPR43 expression. In summary, mango juice induced changes in the SCFAs production while pomegranate juice induced changes in the composition of microbiota.
1’‐Acetoxychavicol acetate inhibits adipogenesis in 3T3‐L1 adipocytes and in high fat‐fed rats (1045.7)
Authors: Kojima‐Yuasa Akiko , Ohnishi Rie , Deguchi Yohei , Yaku Keisuke , Tabuchi Masaki , Munakata Hiroshi , Matsui–Yuasa Isao
Published: June 2021

The compound 1’‐acetoxychavicol acetate (ACA), which is naturally obtained from the rhizomes and seeds of Languas galangal and Alpinia galangal, is the pungent component of galangal and has various biological properties, including anti‐inflammatory, anti‐allergic and antitumor properties. In this study, we investigated the anti‐obesity effects of ACA in 3T3‐L1 adipocytes and in a high fat diet (HFD)‐induced rat model of obesity.

ACA caused a significant decrease in the activity of glycerol‐3‐phosphate dehydrogenase (GPDH) in 3T3‐L1 adipocytes without eliciting cell cytotoxicity, and it inhibited cellular lipid accumulation through the down‐regulation of transcription factors such as PPARγ and C/EBPα. ACA also induced a dose‐dependent phosphorylation of AMP‐activated protein kinase (AMPK). In the animal model, rats fed an HFD containing 0.05% ACA gained less weight than rats fed an HFD alone. The visceral fat mass in rats fed an HFD containing 0.05% ACA tended to be lower than that in rats fed an HFD alone. Furthermore, a histological examination of livers from rats fed an HFD showed steatohepatitis. However, rats fed an HFD containing 0.05% ACA showed no histopathological changes in the liver tissue.

Our results show that ACA exerts anti‐obesity activities both in vitro and in vivo and suggests that ACA may have a novel preventive activity against obesity and possibly other metabolic diseases.
Characterization of force sensing apparatus in human periodontal ligament fibroblasts (1046.10)
Authors: Macarak Edward , Braud Martin , Wei Wenjie , Giglou Kamyar , Howard Pamela
Published: June 2021

Characterization of force sensing apparatus in human periodontal ligament fibroblasts A putative force transfer complex (FSC) similar to that found in muscle tissue (dystrophin glycoprotein complex or DGC) has been characterized in human periodontal ligament fibroblasts (HPDLFs). Similar to the DGC, the FSC is a physical bridge that links the cell to the basement membrane (BM). HPDLFs were also subjected to compressive strain in vitro to mimic that used in orthodontic tooth movement and its effects on the gene expression of 14 genes evaluated.

Methods: Using immunohistochemistry and Western blotting, HPDLFs were characterized with respect to either the presence or absence of DGC and BM components. To evaluate the effects of compressive force on HPDLF gene expression, mRNAs were quantified by RT‐ PCR.

Results: DGC and BM proteins localized to HPDLFs by immunohistochemistry and further characterized in HPDLFs extracts by Western blotting; however, dystrophin was not identified. RT‐PCR revealed significant changes in gene expression of α‐sarcoglycan (p = 0.0009) and γ‐sarcoglycan (p < 0.0001), collagen III (p = 0.0191), and laminin (p = 0.0077) in HPDLFs that underwent cyclic compression.

Conclusions: DGC and BM proteins, with the exception of dystrophin, are present in HPDLFs. Cyclic compressive forces caused significant changes in gene expression of some DGC and ECM components.
Regulation of versican expression by bacterial infection is TLR4‐dependent but MyD88‐independent (1046.3)
Authors: Chang Mary , Delaney Martha , Hajjar Adeline , Wight Thomas , Frevert Charles
Published: June 2021

In the acute response to bacterial lung infection, gram‐negative bacteria activation of toll‐like receptors (TLRs) stimulates macrophages to secrete a variety of molecules that lead initially to pulmonary inflammation and, ultimately, to resolution of inflammation. Using immunohistochemistry and qPCR we have shown that versican, a chondroitin sulfate proteoglycan, co‐localizes with macrophages in the lungs of mice with gram‐negative pneumonia, and is produced by alveolar and bone marrow‐derived macrophages (BMDM) exposed to live E. coli or lipopolysaccharide (LPS) in vitro. The goal of the current study is to define the TLR signaling pathway responsible for versican production by macrophages. BMDM were isolated from wild‐type mice (WT) and mice lacking TLR2 (TLR2 KO), TLR4 (TLR4 KO), or MyD88 (MyD88 KO). Versican mRNA was detected at low levels in untreated BMDM from all mice. In WT BMDM, versican mRNA was significantly increased by 4 h after treatment with 10 ng/ml LPS (50.5 ± 10.8‐fold vs PBS control; n = 3, p < 0.0001), and returned to basal levels by 24 h. The versican response to LPS was similarly elevated in TLR2 KO BMDM but was completely abrogated in TLR4 KO BMDM, indicating that LPS induction of versican is mediated by TLR4. Further, the response in MyD88 KO BMDM (48.3 ± 5.0‐fold vs PBS control; n = 3, p < 0.0001) was similar to that in WT BMDM, indicating that LPS induction of versican is MyD88‐independent. Further studies to define downstream signaling components in more detail are in process and will contribute to our understanding of the pro‐ vs anti‐inflammatory role of versican in the acute response to bacterial infection.

Grant Funding Source: Supported by HL098067 & RR030249
CoCl2, chemical inducer of Hif1‐alpha, and CD44 expression in salivary glands malignant neoplasm cell line (1047.2)
Authors: Soave Danilo , Celes Mara Rubia , Ribeiro‐Silva Alfredo
Published: June 2021

Introduction: The use of neoplastic cell lines to evaluate and understand the biological behavior of neoplastic cell is widely spread. Some studies argue that hypoxia promotes the genetic instability, ability to evade apoptosis, increased aggressiveness, as well as promotes extracellular matrix breakdown, tumor progression and metastasis. CD44 is involved in cell‐cell, cell‐matrix interactions and their expression seems to be unregulated in many tumors.

Objective: the present study evaluated the chemically induced hypoxia and CD44 expression in salivary glands malignant neoplasm cell line.

Material and Methods: The chemically induced hypoxia was achieved by exposing normoxic cultures of HTB‐41 (ATCC) cells to CoCl2. The cells were plated (5x105 cells/well) and incubated with medium alone ( or 50, 150, 300 μM of CoCl2 for 24 h at 37 ̊C, 5% CO2. The chemically induced hypoxia was identified and evaluated through analysis of Hif1‐α, CAIX, Glut1 and then correlated to CD44 expression through Western Blotting assay.

Results: The present study demonstrated that HTB‐41 supports the CoCl2 stimulus in different concentrations (50, 150, 300 μM). We also observed that Hif1‐α, CAIX and Glut1 expression were associated with chemically induced hypoxia and the expression were accentuated at 300 μM CoCl2. However, we observed that CD44 expression was decreased in chemically induced hypoxia.

Conclusion: In summary, we presented evidence that Hif1‐α; CAIX and Glut1 expression were associated with chemically induced hypoxia at different concentrations. Additionally, CD44 expression seems to be down regulated in the hypoxic microenvironment.

Grant Funding Source: Suported by FAPESP (2011/18427‐5)
Upregulation of ADP‐ribosylation factor 1 contributes to the enhanced activation of the MAPK ERK1/2 in prostate cancer cells (1047.4)
Authors: Davis Jason , Teng Yong , Guo Jianhui , Wu Guangyu
Published: June 2021

It is well known that the hyperactivation of the MAPK ERK1/2 pathway correlates with the progression of prostate cancer. However, the molecular mechanism responsible for the enhanced activation of this pathway remains ambiguous. Our recent studies have demonstrated that ADP‐ribosylation factor 1 (ARF1), a Ras‐related small GTPase that has been well‐characterized to coordinate vesicle‐mediated intracellular trafficking, regulates the activation of the Raf1‐MEK‐ERK1/2 pathway (Dong C., et al.: J. Biol. Chem. 286:43361‐43369, 2011). This prompted us to define the function of ARF1 in activating ERK1/2 in prostate cancer cells. We found that both ARF1 expression and ERK1/2 activation were markedly augmented in human metastatic prostate cancer cells compared to normal prostate cells. Conversely, the total expression of ERK1/2 was significantly downregulated in prostate cancer cells. To further confirm whether ARF1 expression level is directly linked to ERK1/2 activation, we determined the effect on ERK1/2 activation of lentiviral‐mediated overexpression of ARF1 in normal prostate cells and shRNA‐mediated depletion of ARF1 in prostate cancer cells. Our results showed that ERK1/2 activation was dramatically increased in cells overexpressing ARF1, whereas ERK1/2 activation was markedly decreased in cells infected with shRNA targeting on ARF1. These data have demonstrated for the first time that the enhanced activation of the MAPK ERK1/2 pathway in prostate cancer is at least partially induced by ARF1 upregulation. These data also suggest that, by virtue of its ability to activate the MAPK ERK1/2, ARF1 may play an important role in the progression of prostate cancer.

Grant Funding Source: Supported by: NIH RO1GM076167
BMI‐1 silencing significantly reduces gene expression of DNA damage repair markers in human breast cancer (1047.5)
Authors: Silveira Giórgia , Celes Mara , Soave Danilo , Oliveira‐Costa João Paulo , Ribeiro‐Silva Alfredo
Published: June 2021

Homologous recombination has a major role in DNA double‐strand break repair. BMI‐1 has been associated with cell immortalization through the supression of INK4a/ARF locus, thereby allowing that DNA double‐strand breaks occur. However, the influence of BMI‐1 in homologous recombination markers is still poorly understood. Our objective was to evaluate the influence of BMI‐1 silencing on expression of homologous recombination genes, cell proliferation and cell damage. BMI‐1 was silenced through lipofectamine‐based siRNA (Invitrogen). We accessed gene expression through Real‐Time PCR reactions in MCF‐7 cell line. Cells were subjected to MTT assay for determination of proliferation rate. Immunofluorescence was also performed to access BMI‐1 and γH2AX (DNA damage marker) protein expression. BMI‐1 silenced cells proliferated significantly less than control (p<0.001). BMI‐1 silencing significantly reduced ATM (p<0.001), ATR (p<0.001), BRCA‐1 (p=0.001), BRCA‐2 (p<0.001), H2AX (p=0.001), TP53 (p<0.001), RAD51 (p<0.001) and TOP3β (p=0.037) expression. Immunofluorescence staining showed that cells positive for BMI‐1 were also positive for γH2AX, while silenced BMI1 cells had reduced expression of γH2AX (nuclear staining). Our results showed that BMI‐1 positive cells may have a more aggressive behavior in breast cancer.

Grant Funding Source: Supported by: FAPESP (2011/23845‐2)
Alteration of pro‐apoptotic and anti‐apoptotic gene expression in breast cancer cell lines (1047.7)
Authors: Edson Leanne , Hunter Dylan , Coleman William
Published: June 2021

A subset of human triple‐negative breast cancer cell lines exhibit aberrant DNA hypermethylation that is associated with DNA methyltransferase hyperactivity, overexpression of DNMT3b, and methylation‐dependent silencing of numerous genes. These breast cancer cell lines are resistant to standard chemotherapeutic agents, but can be sensitized to chemotherapy with demethylating drugs (5aza). In this study, we surveyed expression of 328 genes associated with apoptosis and survival in Hs578T cells using BioRad Prime PCR Pathway Plates. This analysis identified several pro‐apoptotic genes that are not expressed (undetected) or expressed at negligible levels in Hs578T cells, including FASLG, IGF1, CD27, and BIK. Epigenetic silencing of these genes (directly or indirectly) would convey a survival advantage to the cancer cells through elimination of pro‐apoptotic signaling. These pro‐apoptotic genes and/or members of their pathways have been shown to be subject to methylation‐dependent silencing in cancer cells. In addition, several anti‐apoptotic genes were found to be abundantly expressed in Hs578T cells, including HSPB1 and NPM1. It is not known if epigenetic mechanisms contribute to overexpression of anti‐apoptotic genes in breast cancer. The contributions of aberrant DNA hypermethylation to the silencing of pro‐apoptotic genes (and/or activation of anti‐apoptotic genes) as a mechanism of chemotherapeutic resistance was explored further using a panel of breast cancer cell lines. The results suggest that treatment of triple‐negative breast cancer cell lines that display aberrant DNA hypermethylation with demethylating agents (like 5aza) sensitizes them to standard chemotherapeutic drugs (Doxorubicin, Paclitaxel, 5‐fluorouracil) by reactivation of pro‐apoptotic genes. This experimental observation forms the basis for designing new treatment approaches for patients with triple‐negative breast cancer that combine epigenetic and cytotoxic drugs.
Loss of tumor necrosis factor superfamily genes in breast cancer cell lines (1047.8)
Authors: Hunter Dylan , Edson Leanne , Coleman William
Published: June 2021

A subset of human triple‐negative breast cancer cell lines exhibit aberrant DNA hypermethylation that is associated with DNA methyltransferase hyperactivity, overexpression of DNMT3b, and methylation‐dependent silencing of numerous genes. These breast cancer cell lines are resistant to standard chemotherapeutic agents, but can be sensitized to chemotherapy with demethylating drugs (5aza). In this study, we surveyed expression of 328 genes associated with apoptosis and survival in Hs578T cells using BioRad Prime PCR Pathway Plates. This analysis identified several members of the tumor necrosis factor (TNF) and TNF receptor superfamilies that are not expressed (undetected) or expressed at negligible levels in Hs578T cells, including TNFSF8, TNFSF10, TNFSF11, TNFSF14, TNFSF15, TNFRSF13B, TNFRSF17, and TNFRSF18. Among these TNF/TNFR superfamily genes, TNFSF10, TNFSF14, TNFSF15, TNFRSF1B, and TNFRSF8 have been directly implicated as activators of apoptosis. Epigenetic silencing of these genes (directly or indirectly) would convey a survival advantage to the cancer cells through elimination of pro‐apoptotic signaling. TNFSF10, TNFRSF1B, and TNFRSF8 have been shown to be subject to methylation‐dependent silencing in cancer cells. The contributions of aberrant DNA hypermethylation to the silencing of pro‐apoptotic genes in the TNF/TNFR superfamilies as a mechanism of chemotherapeutic resistance was explored further using a panel of breast cancer cell lines. The results suggest that treatment of triple‐negative breast cancer cell lines that display aberrant DNA hypermethylation with demethylating agents (like 5aza) sensitizes them to standard chemotherapeutic drugs (Doxorubicin, Paclitaxel, 5‐fluorouracil) by reactivation of pro‐apoptotic genes in the TNF/TNFR superfamilies. This experimental observation forms the basis for designing new treatment approaches for patients with triple‐negative breast cancer that combine epigenetic and cytotoxic drugs.
Metallothioneins are potential prognostic biomarkers in breast cancer patients (1048.11)
Authors: Malepati Dhananjaya , S Lakshminarasaiah
Published: June 2021

Metallothioneins (MTs) are highly conserved low molecular weight proteins. The main functions of MTs are binding, chelating and detoxification of various toxic substances. It also protects tissue damage from reactive oxygen species, radiation and mutagens. MTsexpression levels were evaluated in order to determine its role in different clinical stages of breast tumors during pre‐ and post‐ menopausal women. In the present study the expression levels of metallothioneinisoforms MT‐1F, MT‐1E and MT‐2A were assessed by semi‐quantitative PCR in breast carcinoma tissues of seventy two individuals. Similarly, MT levels were also quantified in non tumor breast tissue samples of respective individual subjects, during different clinical stages of pre‐and postmenopausal breast cancer patients through RT‐PCR technique.

.The percentage of expression levels were quantified by integrative density value (IDV). As the stage progresses the selected MT isoforms expression also increases in both pre‐ and post‐ menopausal breast tumors. This indicates that the MTs expression is closely associated with pre and postmenopausal malignant tumors along with non tumor tissues. So, MT isoforms are promising prognostic biomarkers in the prospective patient studies and may be validated to establish MT assays for use in clinical practice. Therefore the MT assays may be used as the Diagnostic tools in the diagnosis and treatment of women breast cancer.

Grant Funding Source: INDIAN GOVT. FUNDING AGENCIES
Epigenetic regulation of CFTR during cancer development and implications in cancer treatment (1048.9)
Authors: Huang Wenqing , Fok Kin Lam , Jiang Xiao Hua , Chan Hsiao Chang
Published: June 2021

Cystic fibrosis transmembrane conductance regulator (CFTR), a member of the ATP‐binding cassette (ABC) transporter family, is expressed in a wide variety of epithelial tissues. Recently, it has been found that CFTR is downregulated in various human cancers, which functionally suppress cancer malignancy. Previous studies have indicated that there are at least two promising epigenetic mechanisms for regulation of CFTR expression, including promoter methylation and microRNAs targeting CFTR. However, the exact interaction between CFTR and epigenetic modulation in human cancers is poorly understood. In the present study, we found that the promoter of CFTR was methylated and could be reversed by DNA methyltransferase (DNMT) inhibitor 5‐aza‐2’‐deoxycydine (5‐aza) in lung cancer cell lines. Meanwhile, CFTR expression was also upregulated by two microRNA inhibitors including mir‐145 and mir‐494 inhibitors in A549 cell line and could be suppressed by mir‐384 mimic in Calu3 cell line. In addition, we demonstrated that both DNMTs and candidate CFTR regulating microRNAs levels were altered by hypoxia in lung cancer cell lines. These results may shed new light into our understanding of CFTR regulation during cancer development.
Histomorphologic characterization of human tissues stained with selected plant‐based dyes (1050.10)
Authors: Bassey Rosemary , Hu Yan , Tomaszewski John , Nickerson Peter , Holmes Jasmine , Colón Luis , Osinubi Abraham , Oremosu Ademola
Published: June 2021

Background: The use of eco‐friendly natural dyes has become a matter of significance due to increased environmental awareness. However, the scientific studies and systematic reports with natural dyes are insufficient. In this study, we have explored the histomorphological properties of several plant‐based dyes.

Methodology: Paraffin‐embedded human tissue sections were stained with a variety of plant‐based dyes sourced from Nigeria to determine factors that produce the optimal staining efficiency in the dyes. The stained tissues were studied under bright‐field and also using a multi‐spectral imaging system to histomorphologically analyze the staining characteristics on the different tissues.

Results: The staining characteristics and factors influencing staining efficiency of the different dyes on human tissues were determined. L. cyanescens stained the nuclei and cytoplasm with strong nuclear labeling in all the tissues. H. sabdariffa selectively and strongly labeled the nuclei in the tissues stained. The deep pink from H. sabdariffa and the blue from L. cyanescens gave good contrast, making a good combination for staining of tissues.

Conclusion: Our study has shown L. cyanescens and H. sabdariffa as effective, cheap and readily available histological stains derived from our rich biota.
In vivo demonstration of okadaic acid internalization in glutamatergic spinal motor neurons (1050.3)
Authors: Torres‐Vazquez Irma , Torrech‐Santos Christian , Serrano‐Velez Jose , Rein Kathleen , Rosa‐Molinar Eduardo
Published: June 2021

The development of new techniques and reagents has facilitated the detailed study and characterization of endocytic pathways. We used okadaic acid (OA), a naturally occurring toxin produced by marine dinoflagellates of the genus Prorocentrum, that is a potent and selective inhibitor of the serine/theorine protein phosphatase, phosphoprotein phosphatase 1, combined with a minimally invasive method to retrogradely label neurons and their processes in an adult spinal motor circuit. We demonstrate the prevalent expression of connexin 35 gap junction proteins and plaques at dendrodendritic excitatory mixed synapses between spinal motor neurons and interneurons. In addition, we discovered that OA appears to be entering spinal motor neurons through a clathrin‐independent internalization mechanism that we are currently investigating.

Grant Funding Source: Supported by MH‐086994; NSF‐10 62963, and NSF‐0964114
Evidence for mitotic recombination in the sub‐telomeric regions of the Plasmodium falciparum genome, with implications for malaria virulence (1050.8)
Authors: Singhakul Suriya , Manary Micah , Winzeler Elizabeth
Published: June 2021

The var gene family undergoes differential expression in the Plasmodium falciparum parasite, resulting in malaria’s variable presentation to the human immune system. This haploid parasite also undergoes variable recombination between chromosomes. Whole genome sequencing provides evidence that this recombination occurs frequently and in a patterned manner in the subtelomeric regions containing the var genes. PCR amplification of the targeted regions show in‐frame, reproducible recombination in a laboratory setting between the suspected regions, exclusively within the exonic regions of var genes. qPCR data verifies the existence of replacement recombination events, resulting in an amplification of one chromosomal region and corresponding the deletion of another. This data has major implications for understanding malaria virulence patterns and presentation to the immune system, as it appears as if variable recombination is a driven parasite method of inducing variability within var genes.
Simvastatin, atorvastatin, and pravastatin are equally effective in improving the hemodynamic status of type 1 diabetic rats (1051.14)
Authors: Quidgley Jose , Cruz Nildris , Melendez George , Crespo Maria
Published: June 2021

Statins improve cardiovascular (CV) status of diabetic patients. The underlying mechanisms remain unclear. We compared the effects of a four‐week administration of 10 mg/kg/day atorvastatin (AV), simvastatin (SV), and pravastatin (PV) on the CV function of (STZ)‐diabetic rats. Echocardiographic parameters, endothelial function, cholesterol levels, perivascular fibrosis and oxidative stress markers were evaluated. Cholesterol levels (mg/dl) were higher in STZ (249±16) than in CT (156±7) (n=10, P<0.05), and were not modified with this statin dose. AV and SV improved ejection fraction, stroke volume and cardiac output index (n=10, P<0.05), while PV only improved ejection fraction and stroke volume. Vascular relaxation was improved by all statins, and abolished by 1 mM LNAME, suggesting a NO‐dependent mechanism. In addition, all statins significantly reduced perivascular fibrosis (µm/100g BW) from 10.59±0.40 in untreated STZ to 9.0±0.3 (AV), 8.8±0.4 (SV), and 9.0±0.4 (PV), and levels of the oxidative stress markers MDA and 4‐HAE. The cholesterol‐independent hemodynamic benefits observed with all statins may be secondary to reductions of oxidative stress. The latter, by improving endothelial function, will reduce vascular tone and remodeling, decrease cardiac workload and improve systolic performance.

Grant Funding Source: Supported by MBRS‐RISE R25‐GM061838.