bluuebunny/crossref_metadata_2025 huggingface dataset:
A‐type proanthocyanidins from cranberry inhibit the ability of extraintestinal pathogenic E. coli to invade gut epithelial cells and resist killing by macrophages
Authors: Shanmuganayagam Dhanansayan , Johnson Rachael Erin , Meudt Jennifer J , Feliciano Rodrigo Pedro , Kohlmann Kristin L , Nechyporenko Anatoliy V , Heintz Joseph A , Krueger Christian G , Reed Jess D
Published: June 2021

Extraintestinal pathogenic E. coli (ExPEC) are the major cause of most varieties of extra‐intestinal infection, including urinary tract infection, septicemia, and neonatal meningitis. Persistence of ExPEC in the gut increases the risk of subsequent extraintestinal infection. The mechanisms of persistence and effects of dietary Atype proanthocyanidins (PAC) on the mechanisms are poorly understood. Using a strain of ExPEC expressing both P‐ and type‐1 fimbriae, we studied the ability of ExPEC to invade gut epithelial cells and resist killing by macrophages. We also determined the effects of PAC on these mechanisms. ExPEC attached to and invaded enterocytes by hijacking the host cytoskeletal system, and survived within intracellular vacuoles without causing overt pathology. The ExPEC also resisted post phagocytic killing by macrophages. Exposure of ExPEC to PAC significantly inhibited invasion in a PAC dose dependent manner; higher molecular weight PAC were more effective in inhibiting invasion. Scanning electron microscopy revealed that PAC exposure disrupted surface structures on ExPEC. PAC also induced agglutination of ExPEC and increased killing by macrophages. This study provides novel mechanisms by which ExPEC may persist in the gut and suggests potential use of PAC in preventing gut colonization by ExPEC. Research was funded by Wisconsin Cranberry Board and Cranberry Institute.
Evaluation of in vitro anti‐hyperglycemic effect of Cinnamon cassia derived phenolic phytochemicals
Authors: Kang Bou‐Hee , Racicot Kenneth , Kwon Young‐In , Apostolidis Emmanouil
Published: June 2021

Cinnamon cassia (Cinnamon) proanthocyanidins (PACs) are shown to have anti‐hyperglycemia potential. The present study investigates the carbohydrate hydrolysis enzyme inhibition of cinnamon PACs. Five grams of cinnamon bark powder were extracted in 100 mL acetone solution (CAE) (acetone: water: hydrochloric acid, 70:29.9:0.01) for 2 hours at room temperature and in 100 mL deionized water for 30 min at 90 □ (CWE). PACs were purified from CAE using LH‐20 (CAE‐PAC) to be further evaluated. PAC contents were evaluated by 4‐ Dimethylaminocinnamaldehyde (DMAC) assay and yielded 795, 177 and 123 mg/g, for CAE‐PAC, CAE and CWE respectively. The total phenolic contents of CAE and CWE were determined to be 152 and 134 mg/g respectively. All extracts were adjusted to the same PAC content (180, 90, 45 and 20 μg) and the inhibitory activity against rat α‐glucosidase was determined. The CAE‐PAC fraction had very low rat α‐glucosidase inhibitory activity, CAE had the highest (IC50 3.13 mg/mL DW) followed by CWE (IC50 5.21 mg/mL DW). The specific maltase and sucrase inhibitory activities were determined and CAE (IC50 2.47 and 0.63 mg/mL DW) had higher inhibition than CWE (IC50 5.56 and 2.79 mg/mL DW). Results suggest that the observed bioactivity is not PAC dependent and that CAE has higher anti‐hyperglycemic potential than CWE via inhibition of carbohydrate hydrolysis enzymes.

This research was supported by US Army Natick Soldier Research Engineering Center
Aberrant beta‐catenin signaling as a molecular target for bioactive phytochemicals
Authors: DEY MOUL , LI FENG , WISE MITCHELL , LIU YI , WANG DAN
Published: June 2021

Phytochemicals are rich source of chemoprevention agents but their effects on modulating the Wnt/β‐catenin signaling pathway and attenuating cancer stem cell proliferation have remained largely uninvestigated. Aberrantly activated Wnt signaling can result in the abnormal stabilization of β‐catenin, a key causative step in a broad spectrum of cancers. Here we report the modulation of lithium chloride‐activated canonical Wnt/β‐catenin signaling by phytochemicals that have antioxidant, antiinflammatory or chemopreventive properties. The compounds were first screened with a cervical cancer‐derived stable Wnt signaling reporter HeLa cell line. Positive hits were subsequently evaluated for β‐catenin degradation, suppression of β‐catenin nuclear localization and down‐regulation of downstream oncogenic targets of Wnt/β‐catenin pathway. Our study shows a novel degradation path of β‐catenin protein in HeLa cells by Avenanthramide 2p, a polyphenol present in wholegrain oats. To further investigate the underlying mechanism of the action, we isolated cervical cancer stem cells and confirmed multiple stem cell surface biomarkers using fluorescence‐activated cell sorting analysis. Our continuing and future goals are to evaluate effects of Avenanthramides and other phytochemicals on growth of isolated cancer stem cell like populations as well as in animal models of carcinogenesis and metastasis. Research support came from National Institutes of Health and SD Agriculture Experiment Station.
The unfolded protein response is not involved in trans‐10, cis‐ 12 CLA induced hepatic steatosis in the mouse
Authors: Hussein Mahmoud , Giesy Sarah , Krumm Christopher , Long Oiaoming , Bauman Dale , Boisclair Yves
Published: June 2021

Chronic intake of trans‐10, cis‐12 conjugated linoleic acid (t10, c12 CLA) reduces adiposity in mice. However, it also causes hyperinsulinemia and increases lipogenic gene expression and lipid accumulation in liver. Recently, components of the unfolded protein response (UPR) have been shown to mediate a portion of the effects of hyperinsulinemia on hepatic lipogenesis. To determine whether UPR contributes to the effect of CLA on hepatic steatosis, C57BL/6J female mice were offered diets supplemented with either 1% oleic acid or t10, c12 CLA for 4 weeks. As expected, t10, c12 CLA reduced adipose tissue mass and simultaneously increased plasma insulin and hepatic lipid deposition. t10, c12 CLA increased the hepatic expression of key enzymes involved in fatty acid synthesis, esterification and uptake. t10, c12 CLA, however, did not alter the mRNA expression or protein level of the key UPR components, IRE1α, spliced XBP1, CHOP, BiP and SEL1L. We conclude that the UPR is not involved in mediating the effects of t10, c12 CLA on hepatic steatosis.
Alpha‐lipoic acid protects against weight gain and hyperlipidemia in response to high fat feeding in obese Zucker rats
Authors: Carrier Bradley , Harding Scott V , Rideout Todd C
Published: June 2021

Alpha‐lipoic acid (ALA) supplementation has recently been shown to have anti‐obesity and lipid‐lowering properties, however, the underlying molecular mechanisms are unknown. Our objectives were to examine the effects of ALA supplementation on body weight, blood lipids, and hepatic lipogenic response in obese Zucker rats. Rats (n=8) were randomly assigned to either a Western Diet (WD) or a WD supplemented with 0.25% ALA. Feed intake was matched between the two groups. ALA protected (p=0.01) against diet‐induced obesity (102.5±3.1 vs. 121.5± 3.6, % change BW) and reduced (p<0.05) plasma total cholesterol (11%), non‐HDL cholesterol (25%), and total LDL particle number (46%) compared with the WD group. Plasma triglycerides (TAG) were increased (p=0.09) in response to ALA supplementation, largely due to an increase in VLDL/chylomicron particle number compared with the WD group. Hepatic TAG (mmol/g tissue) was reduced (p<0.05) by 46% in the ALA group compared with the WD animals. ALA supplementation reduced the mRNA expression of hepatic acetyl‐CoA carboxylase (2.4 fold, p=0.03) and fatty acid synthase (5.4 fold, p=0.02) compared with the WD group. FAS protein abundance was also decreased (43%) in the ALA group compared with the WD group. These data suggest that ALA supplementation protects against diet‐induced weight gain and dyslipidemia by modulating hepatic lipogenesis.

Grant Funding Source: UB start up funds
DIFFERENT POLYPHENOLS HAVE DIFFERENT AFFINITIES FOR C‐TERMINAL SUBUNITS OF MALTASE‐GLUCOAMYLASE AND SUCRASEISOMALTASE FOR THE MODULATION OF GLUCOSE RELEASE
Authors: Simsek Meric , Quezada‐Calvillo Roberto , Nichols Buford L. , Hamaker Bruce R.
Published: June 2021

Some polyphenolic compounds inhibit the intestinal alpha‐glucosidases. However, it is not clear how those compounds affect the enzyme activity of the individual C or N terminals subunits of maltase‐glucoamylase (Mgam) and sucrase‐isomaltase (Si).

Objective

To modulate the enzymatic activity of the most active C terminal‐Mgam (Ct‐Mgam) and C terminal Si (Ct‐Si) by selected polyphenols in order to control the glucose release from the starch digestion.

Methods

Maltase activities of recombinant Ct‐Mgam and Ct‐Si subunits were measured by Tris Glucose‐ Oxidase (TGO) method. The reactions were stopped by TGO and the optical density was measured at 450 nm.

Results

Maltase activity of both enzymes was inhibited. IC50 values were 0.58 and 0.571 mM for caffeic acid, 0.039 and 0.014 mM chlorogenic acid, 0.139 and 0.213 mM for (+)‐catechin hydrate, 0.383 and 0.177 mM for gallic acid, 0.041 and 0.014 mM for (−) epigallocatechin gallate for Ct‐Si and Ct‐Mgam, respectively.

Conclusions

The results indicate that the potencies of inhibition of each polyphenol differ against each Ct‐Mgam and Ct‐Si subunits. This opens the possibility for the use of polyphenols as subunit‐specific inhibitors.
Carotenoids and Health
Authors: Jamieson Herbert Morehouse
Published: June 2021

Carotenoids are naturally occurring pigments found in plants, fungi, algae and bacteria. Carotenoids protect plants from sun damage. Hundreds of plant carotenoids have been identified, of which around 50 are present in edible vegetables and fruits. Astaxanthin,Beta‐carotene,Crocetin, Lycopene,Lutein, Zeaxanthin are the most important carotenoids that have beneficial effects on human health.

The consumption of Carotenoids has been associated with reduced risks of cancers, cataract and age related macular degeneration. The primary benefit of carotenoids lies in their antioxidant potential. Antioxidants are compounds that protect the body's cells from damage by unstable oxygen molecules called free radicals. Carotenoids guard against certain types of cancer, apparently by limiting the abnormal growth of cells. For instance, lycopene appears to inhibit prostate cancer formation. Researchers at Harvard University found that men who ate 10 or more servings a week of tomato‐based foods – tomatoes being the richest dietary source of lycopene – cut their risk of prostate cancer by nearly 45 per cent. In addition, carotenoids fight heart disease by blocking the formation of LDL (‘bad’ cholesterol) which leads to heart attacks. The carotenoids lutein and zeaxanthin both reduce the risk of macular degeneration (MD). This article analyzes the health benefits of Carotenoids on human health.
Bioavailability of iron (Fe) and zinc (Zn) from Fe and Zn biofortified pearl millet
Authors: Hambidge K Michael , Bellad Roopa M. , Mahantshetti Niranjana S. , Kodkany Bhalchandra S. , Boy Erick , Westcott Jamie E. , Rai Kedar N. , Cherian Binu , Miller Leland V. , Krebs Nancy F.
Published: June 2021

Millet is a major food staple in Northwestern India and the Sahel in Africa. This grain's drought resistance is resulting in progressively greater use. The purpose of this study was to determine the absorption of iron and zinc from pearl millet biofortified with two micronutrients that are typically deficient in non‐fortified plantbased diets globally.

The study was undertaken in 40 children aged 2 y in Karnataka, India (n = 21 test/19 controls). Three test meals providing approximately 84 ± 17 g dry pearl millet flour were fed on a single day for zinc and 2 days for iron between 9 am and 4 pm. Quantities of zinc and iron absorbed were measured with established stable isotope extrinsic labeling techniques and analyses of duplicate diets.

The mean (±SD) quantities of iron absorbed from test and control groups were 0.54 ± 0.39 mg/d and 0.18 ± 0.13 mg/d respectively (P = 0.001). The quantities of zinc absorbed were 0.95 ± 0.47 mg/d and 0.67 ± 0.24 mg/d respectively (P = 0.03). These data did not include absorption of the modest quantities of iron and zinc contained in snacks eaten before and after the three test meals.

In conclusion, quantities of both iron and zinc absorbed when iron and zinc biofortified pearl millet is fed to children aged two y as the major food staple is more than adequate to meet the physiological requirements for these micronutrients.

Supported by HarvestPlus 8057 and IAEA.
Malaria is associated with decreased nutrient intake in Zambian children fed provitamin A biofortified maize
Authors: Bresnahan Kara , Chileshe Justin , Tanumiharjo Sherry
Published: June 2021

Vitamin A (VA) supplementation improves VA status and protects against malarial infection. Provitamin A biofortified staple crops may provide a more sustainable approach to alleviate VA deficiency, but the impact of febrile illness on food intake must be considered in malaria endemic regions. Morbidity data and food logs from a 3‐mo efficacy trial on provitamin A biofortified (orange) maize in Zambian children (n = 189) were systematically analyzed overtime to determine the impact of malaria on food intake. Nutrients examined included macronutrients, iron, zinc, and VA. t‐Test comparisons based on individual intakes in healthy and malarial states over 3‐d intervals were made within and between orange (n = 30) and white maize (n = 30) groups. Malaria prevalence did not differ overall, or between treatment groups over time (all P >; 0.05). No differences were observed between nutrient decreases in orange and white maize groups (P >; 0.05). Lower nutrient intakes were observed for all variables during malaria outbreaks (food 301 ± 441 g; calories 257 ± 367; carbohydrate 45 ± 67 g; protein 8 ± 13 g; fat 6 ± 8 g; iron 1 ±2 mg; zinc 1 ±1 mg; VA 65 ± 104 RAE; all P < 0.05). Considering the impact of malaria on food and nutrient intake and increased VA utilization and excretion due to the acute phase response, biofortification targets may need to be elevated in endemic regions. Funded by Global Health and HarvestPlus.
Ultrasound imaging to monitor prostate tumor progression and metastases in TRAMP mice
Authors: Conlon Lauren E , Zuniga Krystle E. , Hartman Emily , O'Brien William D. , Erdman John W.
Published: June 2021

The Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) model develops and progresses through all stages of carcinogenesis including metastases. The development of high‐resolution small animal ultrasound (US) technology allows us to obtain longitudinal 3‐D volume calculations from 2‐D images of the mouse prostate. The objective of this research is to utilize longitudinal US imaging to monitor prostate carcinogenesis and development of metastases in the TRAMP model. Beginning at 10 weeks of age, male C57BL/6 X FVB TRAMP mice (n=15) underwent US imaging every two weeks until 20 weeks of age. Prostate tumors were detected as early as 10 weeks of age. Average prostate tumor volume at 20 weeks of age was in excess of 2000 mm3. In mice without tumors, average prostate volume at 10 weeks of age was 50 mm3 and increased to 90 mm3 at 20 weeks. US imaging allowed for the detection and growth of metastases to the regional pelvic lymph nodes. Small animal US technology has not yet been utilized to investigate the efficacy of anti‐cancer therapies, including dietary interventions. At 8 weeks of age, TRAMP mice were randomly assigned to consume AIN‐93G control (n=15) or 10% tomato powder diet until 20 weeks of age, and mice were imaged every two weeks. Through the use of non‐invasive 3D US imaging of prostate growth and development of metastases, we are able to monitor and quantify treatment responses in vivo over time.
Tea bioactive compounds inhibit prostate cancer stem cells via downregulation of Bmi1
Authors: Zhou Jin‐Rong , Gong Yi
Published: June 2021

Cancer stem cells (CSCs) are suggested to be responsible for tumor initiation, progression, drug resistance, recurrence and metastasis. Strategies that target CSCs may significantly prevent and/or delay cancer progression. The objective of this study was to evaluate the effect of tea bioactive components on targeting prostate CSCs. The effect of tea components on the self‐renewal ability of CSCs was evaluated by sphere‐forming efficiency (SFE) assay, gene expression was quantified by real time PCR, and Bmi1 gene silencing was manipulated by siRNA. The SFEs of LNCaP, DUCaP, PC3 and DU145 human prostate cancer cell lines were 2.8%, 15.7%, 1%, and 5.6%, respectively. The expression of several CSC related biomarkers, such as c‐met, Bmi1 and CD44 were increased, whereas the expression of androgen receptor (AR) was decreased in prostate spheres. Tea compounds, especially epigallocatechin gallate and theaflavines inhibited dose‐dependently the SFEs of prostate cancer cells and downregulated the expression of Bmi1 gene. Gene function assay showed that silencing of Bmi1 dramatically inhibited the growth and decreased the sphere‐forming ability of prostate cancer cells. The results derived from our preliminary studies suggest that Bmi1 may play an important role in self‐renewal ability of prostate CSCs, and that tea bioactive components may have inhibitory effect on prostate CSC self‐renewal in part via downregulation of Bmi1 gene expression and function. Supported in part by R21 CA153355 from NCI/NIH
Annexin A1 as a putative target of anti‐mutagenic action of dietary flavonoids
Authors: Hirata Fusao , Harada Takasuke , Corcoran George B , Hirata Aiko
Published: June 2021

Annexin A1 responds to DNA damaging agents via SUMOylation‐Ubiquitination pathways and is translocated to nuclei. Mono‐ubiquitinated annexin A1 has higher affinity for damaged DNA and promotes in vitro translesion DNA synthesis. In order to explore roles of annexin A1 in chemical mutagenesis, ultimately cancer initiation, we investigated roles of annexin A1 in mutation of the tk gene in L5178Ytk(+/−) lymphoma cells. An anti‐annexin A1 oligonucleotide, but not a scrambled oligonucleotide suppressed the MMS and As3+ induced mutation of the tk gene as judged by number of large colonies resistant to TFT. Silibinin, quercetin and genistein reduced such mutation, while flavone and flavonol were ineffective. The anti‐mutagenic flavonoids inhibited annexin A1‐dependent translesion DNA synthesis by nuclear extracts of L5178Ytk(+/−) lymphoma cells. Since silibinin, quercetin and genistein directly bound to purified mono‐ubiquitinated annexin A1 and inhibited its DNA helicase activity, we propose that nuclear annexin A1 is a putative target of dietary cancer chemopreventive flavonoids to inhibit gene mutation, ultimately cancer initiation. Supported in part by a grant from the Suzan G. Komen Foundation.
Aqueous extract of Rhus coriaria decreases proliferation and promotes apoptosis in human colon cancer cells in vitro
Authors: Munusamy Shankar , Eid Ali Hussein
Published: June 2021

Colon cancer is the third most commonly diagnosed cancer in men and women, and it is the fourth leading cause of cancer‐related deaths in the world. Epidemiological studies strongly suggest that the intake of a diet rich in vegetables and fruits is associated with a reduced risk of colon cancer. Thus, the objective of this study is to investigate the effects of an aqueous extract of Rhus coriaria (RC), a popular middle eastern spice, on the cell proliferation, migration and apoptosis of HCT116 cell line, an in vitro model of colon cancer. HCT116 cells treated with RC extract showed a significant decrease in cell proliferation (42.33% in 100 μg/ml RC treated vs. 100% in vehicle‐treated group) and a marked increase in apoptosis, accompanied by 2.3 fold elevation in caspase‐3/7 activity compared to vehicle‐treated control. In addition, RC treatment was associated with a 2.2‐fold induction in E‐cadherin mRNA expression (an inhibitor of cell migration) in HCT116 cells. Together, our data indicate that RC treatment inhibits proliferation and migration, and promotes apoptosis in colon cancer cells. Further investigations are required to establish the potential therapeutic role of Rhus coriaria in colon cancer.
Differential gene regulation by nutrient restriction and hypoxia in cultured human placental extravillous trophoblasts (HTR‐8/SVneo)
Authors: Jones Sara , Brannon Patsy M
Published: June 2021

Both maternal undernutrition and hypoxia impair placental and fetal growth, but the mechanisms are unknown. We cultured trophoblasts in nutrient‐replete media (C; 20% O2), hypoxia (Hx; 1% O2 in nutrient‐replete media), 75% nutrient‐restricted media (NR; 20% O2), or NR‐Hx. Hx or NR alone reduced growth 40–45% and together by 70% (p<0.001). We determined ≥2 fold changes in gene expression by microarray analysis (Agilent Whole Human Genome Microarray 4X44K V2) and GeneSpring V12 compared to C. NR altered expression of the most genes [335 upregulated (Up) and 417 downregulated (Dn)] in contrast to the fewest in Hx (33 Up and 16 Dn genes) and intermediate numbers in NR‐Hx (171 Up and 75 Dn genes). NR, Hx and NR‐Hx only affected 13 genes in common with 4 Up including FBXO32 and ARC involved in protein catabolism and repressing apoptosis, respectively, and 9 Dn including GLI1, OTUD6A and SUCNR1 involved in, respectively, regulation of proliferation through Hedgehog signaling, deubiquitination and mediation of local stress. Two gene pathways were altered (p<0.001; DAVID/KEGG): Up MAPK signaling pathway in NR and Up aminoacyl‐tRNA biosynthesis pathway in NR‐Hx. Although many mechanisms may contribute to impaired placental growth in NR or NR‐Hx, these results suggest only a few possible shared mechanisms through which both NR and Hx impair placental growth. Supported by Cornell University
Effect of early life Ca and P restriction on renal transporters gene expression in post weaning piglets
Authors: Graugnard Daniel E , Spry Malinda L , Samuel Ryan S , Jolliff James S , Mahan Donald C , Brennan Kristen M
Published: June 2021

The objective of this study was to evaluate the effect of Ca and P restriction during the post‐weaning period in pigs. Piglets (n=5/treatment) were assigned to one of 4 treatments: Control (100% NRC requirements), or 7‐d, 14‐d or 21‐d postweaning Ca and P restriction (33% of NRC requirements). After restriction periods, piglets were fed 100% of NRC requirements. Performance was measured until d 35, at which time kidney gene expression of renal transporters was measured using qPCR relative to the control group. Performance did not differ between treatments. However, during d 21–35 the gain‐to‐feed ratio was greater (P < 0.05) in piglets restricted for 21 d (0.692 G:F) or 14 d (0.672 G:F) compared with 7 d (0.583 G:F). Relative mRNA level of the solute carrier family 34 member 1(SLC34A1) was up‐regulated (P < 0.05) in piglets restricted for 21 d (1.16‐fold) or 14 d (1.18‐fold), consistent with improved gain‐to‐feed ratio and indicating improved active phosphate transport into cells. The solute carrier family 8 member 1 (SLC8A1) was down‐regulated (P < 0.05) in piglets restricted for 7 d (−1.78‐fold), demonstrating a long‐term effect of early life nutrition on Ca metabolism. No differences were observed in ATP2B1, SLC22A and SLC34A2 mRNA levels. These results suggest performance and renal capacity benefits in response to 14 to 21 d post‐weaning Ca and P restriction.

Grant Funding Source: Alltech, Inc.
A Bioinformatic Rational Gene Targeting Approach to Treating Pathologies with Healthy Diet
Authors: Saxena Abhinav , Gao Linda , Crawford Dana
Published: June 2021

We hypothesize that select healthy dietary supplements can modulate the expression of disease‐causing genes back toward the normal, slowing the disease process while lowering treatment cost, toxicity, long‐term risks, and improving compliance as compared with current expensive drugs. This novel strategy includes bioinformatic analysis to identify existing diet modulated risk genes. To demonstrate proof of principle, we tested two diverse models: transplantation rejection and Down Syndrome. These pathologies are caused by excessive IL‐2 production and (in part) RCAN1, respectively, and thus, have defined genetic bases. For clinical immunosuppression, the dietary botanicals curcumin, sulforaphane, quercetin and green tea all significantly inhibited T‐cell proliferation and IL‐2 production in human and mouse T‐cells, suggesting their potential in treating transplant and auto‐immunity patients. We also observe that fish oil reduces mouse hippocampal and spleen RCAN1 mRNA and protein expression. These results have been extended by bioinformatic analyses to assess risk genes for multiple pathologies. Combined, these results support our strategy of using healthy dietary supplements to treat genetically‐defined pathologies, an approach that we believe is simple, healthy, and cost‐effective.
Consumption of Lentinula edodes modulates human immune function by altering cytokine secretion of PBMC ex vivo
Authors: Dai Xiaoshuang , Stanilka Joy M , Rowe Cheryl A , Creasy Rebecca A , Percival Susan S
Published: June 2021

Lentinus edodes (Shiitake) is a medicinal mushroom with a long tradition of use in Asia. The literature suggests that mushrooms modulate immune function, but there have been no controlled studies of Shiitake mushroom consumption on the human immune response. Therefore, we recruited 51 healthy adults, who were randomly allocated to consume either 50g/d or 100g/d of dried, whole Shiitake mushrooms for 28d. Peripheral blood mononuclear cells (PBMC) were isolated and cultured for 24hr with PHA‐L. Culture media was harvested and analyzed for cytokines by multiplex technology. There was no significant difference between the two serving sizes, so the data was pooled and analyzed by paired t‐test. After 28d of Shiitake mushroom consumption, secretion levels of IL4 (51.4%, p<0.001), IL10 (33.4%, p<0.001), IL1α (16.6%, p=0.015) and TNFα (16.6%, p=0.009) by PBMC were significantly increased, while IL6 (−41.0%, p<0.001) and MIP1α (−67.4%, p<0.001) levels were significantly reduced, compared to those seen before consumption. Secretion levels of other cytokines, IL1β, IL17, MIP1β and IFNγ, were the same before and after consumption. These results show that supplementing the diet with whole Shiitake mushrooms is immuno‐modulatory, as seen by the altered pattern of cytokine secretion by stimulated PBMC ex vivo. Funded by US Mushroom Council; Australian Mushroom Growers Assn.
Circulating pro‐inflammatory cytokines are elevated and peak power output correlates with 25‐hydroxyvitamin D in vitamin D insufficient adults
Authors: Barker Tyler , Dixon Brian M. , Schneider Erik D. , Weaver Lindell K.
Published: June 2021

The purpose of this study was to measure circulating cytokines, skeletal muscle strength, and peak power output in young adults with contrasting serum 25‐hydroxyvitamin D (25(OH)D) concentrations. Serum 25(OH)D, inflammatory cytokines, muscle strength, and peak power output were, therefore, measured in young adults (25 to 42 y). Data was collected during the winter to avoid the seasonal influence on serum 25(OH)D. After serum 25(OH)D concentration measurements, subjects were separated into one of two groups: (1) vitamin D insufficient (serum 25(OH)D ≤ 32 ng/mL, n = 14), or (2) vitamin D sufficient (serum 25(OH)D >; 32 ng/mL, n = 14). Following group allocation, serum 25(OH)D concentrations were significantly (p < 0.05) lower and pro‐inflammatory cytokines (interleukin (IL)‐2, IL‐1β, tumor necrosis factor (TNF)‐α, and interferon (IFN)‐γ) were significantly (all p < 0.05) greater in vitamin D insufficient adults. An anti‐inflammatory cytokine (IL‐10; p >; 0.05), peak isometric forces (p >; 0.05), and peak power outputs (p >; 0.05) were not significantly different between vitamin D groups. However, peak power outputs correlated with serum 25(OH)D concentrations in vitamin D insufficient (r = 0.55, p < 0.05) but not in vitamin D sufficient adults (r = −0.27, p = 0.36). We conclude that vitamin D insufficiency could result in pro‐inflammatory stress without altering muscular strength or function in young adults.
Immune‐enhancing Effects of Polysaccharides Isolated from Cheonggukjang in Immunosuppressed Mice
Authors: HONG HEE‐DO , HAN CHUN‐JI , CHO CHANG‐WON , RHEE YOUNG KYOUNG , NAM SO‐HYUN , SUNG SU‐KYUNG , KIM YOUNG‐CHAN , KIM KYUNGTACK
Published: June 2021

In this study, the crude polysaccharide (CHW‐0) was isolated from Cheonggukjang (Korean traditional fermented soybean paste) by hot water extraction, ethanol precipitation and dialysis. The major constituents in CHW‐0 were neutral sugar (69.5%), acidic polysaccharide (24.0%) and protein (6.5%). The neutral sugar was composed mainly of galactose (39.9 mole%), arabinose (31.9 mole%), and glucose (8.8 mole%). We also investigated the immune‐enhancing effects of CHW‐0 in 30 male Chinese Kun Ming mice immunosuppressed by treatment with cyclophophamide. The mice were divided into three groups, and treated by oral administration with CHW‐0 into two dosages (100 mg/kg and 200 mg/kg body weight) for 4 weeks. The results showed that CHW‐0 stimulated proliferation of splenic lymphocytes (T and B cells) in cyclophophamide‐induced immunosuppressed mice. CHW‐0 also markedly increased the natural killer‐cell activity and peripheral white blood cell number. This result demonstrated that CHW‐0 could restore deviated immune parameters in immunosupressed mice and can be utilized as an effective immune‐enhancer.
Urinary ATP and bacteria in shed urothelial cells as a superior diagnostic marker for urinary tract infection in renal transplant recipients
Authors: Kelley Stephen Patrick , Birch Rebecca , Scott‐Ward Toby S , Peppiatt‐Wildman Claire. M , Farmer Chris , Delaney Michael , Wildman Scott S.P
Published: June 2021

Urinary tract infections (UTIs) cause complications for renal transplant recipients (RTRs) and may result in graft loss. Routine urinalysis to detect UTIs is insensitive/inaccurate in RTRs. UTI detection methods with greater accuracy are needed. A ≥50 nmol/l urinary ATP concentration indicates UTI and ATP measurement may be a superior diagnostic test (1). We investigated various urinalysis tests in 53 RTRs. ATP concentrations and intracellular bacteria (IB) in shed urothelial cells were compared to bacterial culture results. 22% of RTRs tested UTI positive by bacterial culture. IB were identified in 44% or RTRs, but only 2% IB positive were also UTI positive by bacterial culture. There was a significant association of the conventional bacterial culture with both urinary ATP concentration and evidence of IB (Fisher's exact test, p<0.05). 6% of RTRs had ATP concentrations ≥50 nmol/l. Those positive were also bacteria positive. 77% patients with ATP concentrations <50 nmol/l were negative by bacteria culture. Diagnostic odds ratios (DORs) indicated that ATP was a much stronger UTI indicator (DOR = 30.580) compared to IB (0.079). However combined urinary ATP concentration and evidence of IB was an even stronger diagnostic indicator of UTI (DOR = 105). Low urinary ATP concentration combined with absence of IB in shed urothelial may provide as superior diagnostic test for absence of UTI in RTRs.

Funding: St Peter's Trust (UK).
Role of Innate Immunity in Immune Enhancing Effects of Candida Skin Test Reagent
Authors: Nakagawa Mayumi , Coleman Hannah , Sikes Jim , Daniels Jaclyn , Nagarajan Uma
Published: June 2021

OBJECTIVES

Serial intra‐lesional injections of common warts with skin testing reagents such as Candida are effective in regressing them. The goal of this study was to test our hypothesis that the Candida skin testing reagent enhances anti‐HPV T‐cell responses through activation of pattern recognition receptors (PRRs) on Langerhans cells and subsequent cytokine(s) secretion.

METHODS

Langerhans cells were treated with Candin® (Allermed, San Diego, CA). LPS and/or zymosan were used as positive controls. Cells were harvested at 8 and 24 hours for RNA extraction, and qRT‐PCR analysis was performed for IL‐1beta, IL‐6, IL‐8, IL‐10, IL‐12p40, IL‐23Ap19, IL‐27p28, IFN‐gamma, and TNF‐alpha. The threshold cycles were normalized to GAPDH expression. In order to remove a small amount of organic solvent (0.4% phenol), Candin was dialyzed using PBS in some experiments. In addition, expressions of PRRs known to associate with Candida albicans (DC‐SIGN, Dectin‐1, Dectin‐2, Galectin‐3, Mincle, mannose receptor, TLRs 1, 2, 4, 6, and 9) were examined. The role of Dectin‐1 was examined by antibody blocking. Supernatants were analyzed for the presence of IL‐12p40 and IL‐12p70 proteins.

RESULTS

Expression of IL‐12p40 mRNA (≥ 5 fold increase over untreated at 8 hour) was detected most frequently with Candin (15 of 16 experiments using Candin and 5 of 6 experiments using dialyzed Candin). All other cytokines examined were detected in multiple experiments in the following order of decreasing frequency: IFN‐gamma, IL23Ap19, IL‐1beta, IL‐27p28, IL‐6, IL‐8, TNFalpha, and IL‐10. IL‐12p40 and IL‐12p70 proteins were detected. All PRRs examined were detected. Dectin‐1 appeared to have a role in IL‐12 production in some subjects.

CONCLUSIONS

Candin appears to promote T‐cell responses through inducing IL‐12 production by Langerhans cells, and may be an attractive new adjuvant for HPV therapeutic vaccines.
Interleukin‐33 drives a proinflammatory endothelial activation that selectively targets non‐quiescent cells
Authors: Haraldsen Guttorm
Published: June 2021

Interleukin‐33 is an alarmin released from stressed or damaged cells. We investigated the effects of interleukin (IL)‐33 on human umbilical vein endothelial cells comparing it to IL‐1β and observing highly similar phosphorylation of signaling molecules and transcription profiles. On the other hand, intradermally injected IL‐33 elicited significantly less endothelial activation when compared to IL‐1β and led us to observe that quiescent endothelial cells (ppRblowp27high) were strikingly resistant to IL‐33. Accordingly, the IL‐33 receptor was preferentially expressed in non‐quiescent cells, corresponding to selective induction of adhesion molecules and chemokines. Multiparameter phosphoflow cytometry confirmed that signaling driven by IL‐33 was stronger in non‐quiescent cells. Manipulation of nuclear IL‐33 expression by siren or adenoviral transduction revealed no functional link between nuclear, endogenous IL‐33 and exogenous IL‐33‐responsiveness.

We conclude that in contrast to other inflammatory cytokines IL‐33 selectively targets non‐quiescent endothelial cells. By this novel concept quiescent cells may remain non‐responsive to a proinflammatory stimulus that concomitantly triggers a powerful response in cells that have been released from contact inhibition.
Several PDE4‐family enzymes differentially regulate EPAC1‐mediated actions during VEC tubule formation
Authors: Kaczmarek Milosz , Brzezinska Paulina B , Umana Maria B , Maurice Donald H
Published: June 2021

Studies have established that several angiogenesis‐related actions of vascular endothelial cells (VECs) are differentially affected by cAMP‐elevating agents. This selectivity is coordinated, in part, by actions of cAMP at individual VEC cAMP‐signaling complexes. Indeed, recently we showed that EPAC1‐integration into a phosphodiesterase 3B (PDE3B)‐based VEC cAMP‐signalosome allows PDE3 inhibitors to activate PI3Kã and promote VEC adhesion to matrigel, an early event in tubules formation. Since EPAC1 integrates into a PDE4D‐containing, VE‐cadherin‐based, cAMP signalosomes that allows EPAC1 to control VEC intercellular contacts, but not adhesions, we investigated how PDE4 inhibitors impacted VEC tubule formation. Our findings identify distinct roles for each PDE4B and PDE4D in the control of EPAC1‐mediated effects in tubule formation and identify potential novel therapeutic strategies to control these angiogenesis‐related events.
KS‐370G prevents cardiac hypertrophy via modulation of autophagy activity
Authors: Chiao Chin‐Wei , Huang Wei‐Pang , Kuo Yueh‐Hsiung , Su Ming‐Jai
Published: June 2021

KS‐370G, a synthetic polyphenolic caffeamide derivative, had hypoglycemic and cardiovascular protective effects similar to other polyphenolic compounds, which also improved left ventricular function and inhibited cardiac hypertrophy in pressure‐overload mice heart in our previous study. Here, we performed the study of its effect on cultured neonatal rat ventricular myocytes. Treatment of neonatal ventricular myocytes with phenylephrine for 24 hours produced marked hypertrophy as proved by the increase of ANP, β‐MHC gene expression and cell surface area. KS‐370G had a concentration‐dependent effect with a complete inhibition at 3 μM of phenylephrine‐induced ventricular myocytes hypertrophy. Recent evidence suggests that autophagy has been observed in failing myocardium, such as cardiac hypertrophy and heart failure by pressure overload. Treatment with phenylephrine 24 hours prominently induced increase of lysosome production in ventricular myocytes by acridine orange assay, which indicated the induction of autophagy. This effect could be inhibited by pre‐treatment with KS‐370G. Since both the induction of hypertrophy and autophagy by phenylephrine was suppressed by KS‐370G, we suggest that the anti‐hypertrophic effect of this compound may be mediated via inhibition of the autophagic signaling pathway.
Cardioprotective Mechanism of Resveratrol: Mitochondrial Permeability Transition Pore
Authors: Ardic Gizem , Cebe Gürhan , Turan Nilüfer Nermin
Published: June 2021

Resveratrol has cardioprotective effect against ischemia‐reperfusion injury. Opening of MPTP causes mitochondrial swelling, ATP depletion, oxidative phosphorylation and apoptosis/necrosis. The aim of this study is investigated the effects of Atractyloside(Atr), MPTP opener, on cardioprotective effects of resveratrol. After anesthesia rat heart was mounted on Langendorff apparatus. In the control group‐global ischemia; preconditioning group‐hearts were perfused with resveratrol (10–5 M), before global ischemia; third group‐Atr (3–100 μM) and resveratrol were perfused before global ischemia. All groups were made 120 min. reperfusion after 30 min. ischemia. Left ventricular development pressure (LVDP), left ventricular end‐diastolic pressure (LVEDP), ±dP/dt, rate pressure product (RPP) were evaluated, infarct size was evaluated at end of reperfusion. In control hearts, LVDP and LVEDP were significantly reduced during reperfusion as before ischemia. Resveratrol was reversed LVDP, LVEDP, ±dP/dt value as compared Atr. Co‐administration with Atr (100,30 μM) and resveratrol were significantly changed LVDP, LVEDP, ±dP/dt values of resveratrol group. RPP values were changed resveratrol group compared with control and high doses of Atr. Infarct size was significantly increased in Atr+resveratrol groups as resveratrol group. MPTP may mediate the effect of resveratrol.
Quantification of the blood‐brain barrier solute permeability and brain tissue transport by multiphoton microscopy
Authors: Fu Bingmei M , Shi Lingyan , Zeng Min
Published: June 2021

To quantify the blood‐brain barrier (BBB) solute permeability (P) and effective diffusion coefficients in brain tissue (Deff) in vivo, we used laser scanning multiphoton microscopy with 800~850nm excitation wavelength to determine P and Deff in the rat brain tissue 200–400 μm below the pia mater. Sodium fluorescein, or FITC‐dextrans, or Alexa Fluor 488‐IgG in 1% BSA mammalian Ringer was injected into the rat (SD, 250–300g) cerebral circulation via the ipsilateral carotid artery by a syringe pump at a constant rate of ~3 ml/min. P and Deff were determined from the rate of tissue solute accumulation and the radial concentration gradients around individual microvessels in the brain tissue. The mean P was 19.6, 7.4, 1.9, 1.4, 1.3 and 0.58 × 10−7cm/s, respectively, for sodium fluorescein (MW 376Da), dextran‐4k, ‐20k, ‐40k, ‐70k, and IgG (MW ~160kDa). These P values were not significantly different from those of rat pial microvessels for the same sized solutes except small solute sodium fluorescein, suggesting that pial microvessels at the brain surface can be a good model for studying the BBB transport of relatively large solutes. The mean Deff/Dfree was 0.46, 0.19, 0.12, 0.12, 0.11, 0.12 for sodium fluorescein, dextran‐4k, ‐20k, ‐40k, ‐70k, and IgG, respectively. Quantification of BBB permeability and brain tissue transport is important in developing better drug delivery strategies. Supported by NIH SC1CA153325–01.
Ethanol‐evoked reduction in myocardial contractility and autonomic dysregulation in freely moving female rats
Authors: Ibrahim Badr Mostafa , Fan Ming , Abdel‐Rahman Abdel A
Published: June 2021

Acute ethanol reduces blood pressure (BP) in proestrus female rats via reduction in cardiac output, which reflected reduced cardiac contractility based on indirectly measured dP/dtmax (maximal rise of ventricular pressure over time). In this study we directly measured cardiac contractility indices [left ventricular developed pressure (LVDP), dP/dtmax and the ventricular isovolumic relaxation time constant (Tau)] in conscious freely moving female rats via fluid filled catheter. We validated this technique using isoproterenol, which produced the expected dose‐related increases in dP/dtmax. To test whether ethanol cardiac depressant effect precedes the hypotensive response in freely moving female rats, we directly measured cardiac function along with BP, HR, in two animal groups received either intragastric ethanol (1 g/kg) or water (control). Further, we conducted offline analysis of sympathovagal activity (spectral analysis) to elucidate ethanol effect on autonomic balance. Acute ethanol, compared to water, caused slowly developing reduction in BP that was significant at 40 min, reached its nadir at 70 min, and lasted till the experiment ended at 90 min. However, significant reductions in myocardial contractility indices (LVDP and dP/dtmax) started 10 min after ethanol, which preceded the hypotensive response. The high‐frequency (HFα) component of spectral analysis of RRI (cardiac vagal control) increased by intragastric ethanol starting at 20 min and continued throughout the experiment. However, the spectral measure of sympathetic activity, low‐frequency (LFα) component of RRI, was not affected by ethanol resulting in net decreases in the sympathovagal, LFRRI/HFRRI, ratio. Collectively, our findings directly implicate the reduction in cardiac contractility and enhancement of cardiac vagal control, at least partly, in acute ethanol‐evoked hypotensive effect in proestrus female rats.

Supported by NIAAA Grant 2R01 AA14441–06
Defining the molecular mode of binding for α‐linolenic acid and the synthetic ligand TUG891 at GPR120
Authors: Hudson Brian , Shimpukade Bharat , Ulven Trond , Milligan Graeme
Published: June 2021

GPR120 is a receptor for long chain polyunsaturated fatty acids that has received growing interest in recent years for the treatment of type 2 diabetes and obesity. However, its development and validation as a therapeutic target has been hindered by a lack of selective ligands for GPR120 over another long chain fatty acid receptor, FFA1. While the recent description of 3‐(4‐((4‐fluoro‐4’‐methyl‐[1,1’‐biphenyl]‐2‐yl)methoxy)phenyl) propanoic acid (TUG891) as a potent and selective GPR120 agonist improved this situation, the need remains for novel GPR120 ligands. To facilitate ligand development at GPR120 we employed homology models and extensive mutagenesis to define the mode of binding at this receptor for an endogenous ligand, α‐linolenic acid (aLA), and TUG891. These studies confirmed that an ionic interaction between the carboxylate of the ligand with R99 of GPR120 is critical to both aLA and TUG891. Several additional residues, including W104, F115, F211, and W277, were found to define the hydrophobic binding pocket and their mutation reduced function to both aLA and TUG891. Finally, mutation of a group of residues including T119, I280 and I281 predicted to form a narrow binding pocket for the planar biphenyl moiety of TUG891 altered potency of TUG891 but not aLA. Together, our findings define key aspects of ligand interaction for both aLA and TUG891 and will be valuable in future drug development at GPR120.
A novel fluorescent ligand binding assay for free fatty acid receptor FFA1
Authors: Watson Sarah‐Jane , Brown Alastair , Holliday Nick
Published: June 2021

Obtaining affinity estimates at FFA1 receptors using radioligand binding is made difficult by lipophilic ligands. We characterised the properties of a new fluorescent ligand 40Ag‐Cy5 (CellAura) using 293TR cells expressing FLAG‐FFA1. Whole cell competition binding experiments using 40AG‐Cy5 were performed on 96‐well plates, followed by imaging and granularity analysis on a confocal platereader. Curve fitting used GraphPadPrism.

Both 40Ag‐Cy5 and its non‐fluroescent precursor 40Ag‐cg were agonists at FFA1 (Fluo4 measurements of intracellular Ca2+), with respective pEC50 values of 5.8±0.2 and 7.9±0.3. In binding experiments 40Ag‐Cy5 (100 nM) was displaced by synthetic ligands such as GW1100 (pIC50 6.0 ± 0.2) and GW9508 (pIC50 6.8 ± 0.2). However oleic acid (up to 100 μM) did not displace 40Ag‐Cy5 binding. By solving the Cheng‐Prusoff correction for GW1100 competition curves using 100nM or 1μM 40Ag‐Cy5 (GW1100 pIC50 5.6), we obtained pKi estimates for 40‐Ag Cy5 (6.2), and GW1100 (6.0; equivalent to its functional pKb). Fluorescence correlation spectroscopy experiments also confirmed 40Ag‐Cy5 binding to FFA1. Thus 40Ag‐Cy5 provides a novel fluorescent ligand which enables estimation of synthetic ligand affinities at FFA1.

Supported by EPSRC and AstraZeneca.
Molecular Mechanisms of PLCβ Activation
Authors: Lyon Angeline , Dutta Somnath , Boguth Cassandra , Skiniotis Georgios , Tesmer John
Published: June 2021

Phospholipase C β (PLCβ) converts the membrane lipid PIP2 into the second messengers inositol 1,4,5‐triphosphate and diacylglycerol, and is activated by extracellular signals via direct interactions with the heterotrimeric G proteins Gαq and Gβγ. However, the molecular mechanisms governing this regulation are only beginning to be understood. Using structural and biochemical studies, we have begun to elucidate how heterotrimeric G proteins relieve autoinhibition of PLCβ leading to dramatic increases in activity. We have also clarified the molecular basis for how the characteristic C‐terminal coiled‐coil domain of PLCβ contributes to membrane association and activation by Gαq. A better understanding of PLCβ function is important for developing novel therapeutic approaches to tackle such diverse diseases as cardiac hypertrophy and uveal melanomas. Support for this work was provided by NIH grants HL086865 and HL071818 to J.J.G.T. and an American Heart Association postdoctoral fellowship to A.M.L.