bluuebunny/crossref_metadata_2025 huggingface dataset:
Dynamic effects of diet diversity and breastfeeding on weight in months 6‐24 of life (1024.9)
Authors: Wright Melecia , Adair Linda
Published: June 2021

Nutrition is a primary contributor to infant growth. We estimated the association of diet diversity and weight in 6‐24 month old infants. Breastfeeding, dietary and weight data collected bimonthly in the Cebu Longitudinal Health and Nutrition Survey (n=3080) were analyzed using a random‐effects longitudinal regression model. Diet diversity (based on 7 food groups) was classified as low (<3 food groups) or high (3+ groups) for each survey. As time‐varying exposures, diet diversity and any breastfeeding (yes/no) captured the dynamic nature of the diet. Interactions between breastfeeding, age and diet diversity were also included in the model. Weight was measured in kg. We adjusted for fixed (sex, maternal SES, height & education) and time‐varying (diarrheal/respiratory illness) variables.

Diet diversity (β=0.23, p<0.05) and breastfeeding (β=0.48, p<0.05) positively contributed to weight. When interactions between breastfeeding status, age and diet diversity were predicted, exclusively‐breastfed babies weighed the most at 6 months but were just as heavy as mixed fed babies by 10 months. Non‐breastfed babies fed a diverse diet weighed the least from 6‐16 months. Weights in the subgroups were indistinguishable after 16 months. The results illustrate the importance of exclusive breastfeeding in the earliest months of life and the need for time‐sensitive introduction of nutritious complementary foods.

Grant Funding Source: Supported by an educational grant from Wyeth Nutrition
Plasma fetuin‐A and phosphofetuin‐A (Ser312) responses to a single or short‐term repeated bout of exercise in obese and normal‐weight individuals (1028.2)
Authors: Mathews Suresh , Ren Guang , He Xiaoming , Bowers Robert , Araya‐Ramirez Felipe , Littlefield Laurel , Grandjean Peter
Published: June 2021

Fetuin‐A, an inhibitor of insulin‐stimulated insulin receptor tyrosine kinase (IR‐TK) and glucose uptake, has been shown to be associated with obesity, insulin resistance, and incident diabetes. In circulation, fetuin‐A exists in both phosphorylated and dephosphorylated forms. Phosphorylation status of fetuin‐A has been shown to be critical for its IR‐TK inhibitory activity. The objective of this study was to characterize the effects of single and repeated bouts of exercise on serum total fetuin‐A and phosphofetuin‐A (Ser312) in obese individuals. 31 obese and 11 normal‐weight men underwent a single bout of treadmill walking expending 500 kcals. Insulin, glucose, HOMA, QUICKI, and glucose/insulin ratio were not altered immediately after or 24 hours following a single bout of exercise. Total fetuin‐A concentrations were significantly elevated immediately after exercise, which decreased to pre‐exercise levels in the recovery period (24 hours after the single bout of exercise). Area under the curve for serum total fetuin‐A, phosphofetuin‐A, glucose, and insulin during an oral glucose tolerance test was significantly decreased 24 hours after a single bout of exercise. Next, we investigated the influence of repeated bouts of exercise, expending 350 kcal each session, on four consecutive days. Compared to baseline, insulin and HOMA‐IR were decreased significantly on days 2, 3, and 24 hours following the last bout of exercise. Unlike total fetuin‐A, serum phosphofetuin‐A levels were decreased significantly 24 hours after the last bout of exercise. Taken together, these results suggest that the observed lowering of serum phosphofetuin‐A (Ser312) levels, 24 hours after a single or short‐term repeated bout of exercise, in obese individuals, may contribute to the improvement of insulin sensitivity.

Grant Funding Source: Supported by Alabama Agricultural Experiment Station Grants Program
Trend and features of top 100 grossing health and fitness iPhone apps (1028.5)
Authors: Meng Yu , Wong Siew Sun
Published: June 2021

There is a fast‐growing trend in using smartphone apps and wearable devices to track health and fitness. Consumer health apps quadrupled from 3,000 in 2010 to 13,600 in 2012. As of Oct 2013, about half of the top 100 grossing health and fitness apps in the Apple Store were designed to track physical activities (PA), especially cardio fitness. Forty percent of them specifically track running, and 12% cycling. The remaining 48% health and fitness apps track other activities, such as strength training, including yoga. Aside from using wireless wearable device, many of these apps automatically tracks PA using the built‐in Global Positioning System (GPS), accelerometers, and environment sensor in the smartphone. Otherwise users self‐report their PA using these apps to track their progress. Popular features of these PA apps include real‐time data storage, goal setting, feedback, social networking, and synchronization with wearable devices. There was no evidence that these top 100 grossing apps have been validated for scientific research use. To align technology advancement with vigorous research, academic researchers should highly consider partnering more closely with the industry to conduct large‐scale, multilevel approach, longer‐term interventions.

Grant Funding Source: PacificSource Healthcare
Relationship of triglyceride levels with A1C and adiponectin in Haitian Americans with type 2 diabetes (1029.4)
Authors: Antwi Janet , Cheema Amanpreet , Ajabshir Sahar , Exebio Joel , Zarini Gustavo , Vaccaro Joan , Huffman Fatma
Published: June 2021

Background: Increased serum levels of triglycerides may dysregulate metabolic processes. We investigated the associations of triglyceride levels with Hemoglobin A1C (A1C) and adiponectin in Haitian Americans with type 2 diabetes.

Methods: This is a cross‐sectional study of 140 Haitian Americans with type 2 diabetes. Following informed consent, information on sociodemographics, dietary and lifestyle factors were obtained. Serum triglyceride levels were assayed by enzymatic methods. Percentages for A1C were measured from whole blood samples using Roche Tina Quant method. Serum adiponectin was determined by enzyme linked immunosorbent assay.

Results: Serum triglyceride levels were inversely associated with adiponectin levels (β = ‐0.238, p = 0.007), positively associated with A1C (β = 0.172, p = 0.046), and milk consumption (β = 0.215, p = 0.013). Serum triglyceride levels were significantly associated with TV watching (β = 0.273, p = 0.001).

Conclusion: Elevated serum triglyceride levels need to be identified early in the management of type 2 diabetes in Haitian Americans to prevent diabetes complications. Dietary and lifestyle factors may independently modulate triglyceride levels.

Grant Funding Source: Supported by NIH/NIDDK sponsored grant.
Resistant starch improves insulin resistance and reduces adipose tissue weight and CD11c expression in the adipose tissues of OLETF rats (1030.2)
Authors: Goda Toshinao , Harazaki Tomomi , Inoue Seiya , Imai Chihiro , Mochizuki Kazuki
Published: June 2021

CD11s/CD18 dimers induce monocyte/macrophage infiltration into many tissues, including adipose tissues. In particular, it was reported that β2‐integrin CD11c‐positive macrophages in adipose tissues are closely associated with the development of insulin resistance. In this study, we examined whether resistant starch (RS) intake reduces macrophage accumulation in adipose tissues and inhibits the development of insulin resistance at an early stage in Otsuka Long‐Evans Tokushima Fatty (OLETF) rats. 22‐week‐old male OLETF rats were fed a control diet (55% α‐corn starch) or an RS diet (55% RS) for 5 weeks. An oral glucose tolerance test was performed after 4 weeks of feeding, while tissues (mesenteric and epididymal adipose tissues, and liver) and tail vein blood were collected after 5 weeks of feeding the test diets. Feeding the RS diet to OLETF rats for 5 weeks improved insulin resistance, reduced the mesenteric adipose tissue weight and enhanced the number of small adipocytes. CD68 expression, a macrophage infiltration marker, was not changed by the RS diet, while the gene expression levels of integrins such as CD11c, CD11d, and CD18, but not CD11a and CD11b, were significantly reduced. CD11c protein expression was reduced by the RS diet. These findings suggest that part of the mechanism for the improved insulin resistance by the RS diet involves a reduction of CD11c expression in adipose tissues.
Effects of pregnancy vitamin D status on adipose tissue development and inflammation in lean, male adult mice offspring (1037.4)
Authors: Belenchia Anthony , Jones Karen , Beversdorf David , Will Matthew , Vieira‐Potter Victoria , Scroggins Rebecca , Peterson Catherine
Published: June 2021

The bioactive form of vitamin D, 1,25‐dihydroxyvitamin D, and its receptor have been identified for a role in preadipocyte differentiation to adipocytes through the modulation of adipogenic transcript factor genes. We sought to determine if maternal vitamin D status influences adipose tissue development or inflammation in adult offspring. Female C57BL/6J mice were randomized to either a vitamin D deficient diet or a control diet (AIN‐93G) for 21 days prior to mating and maintained on this diet throughout pregnancy until postnatal (PN) day 7, at which point both dietary groups were switched to the control diet. At PN day 75, perigonadal (PGAT) and subcutaneous (SQAT) adipose tissue were collected from adult male offspring for general histology and mRNA expression of adipogenic genes. Serum was also collected. There were no differences between groups in whole body weight, fat pad weights, or adipocyte size or number. Mice born to vitamin D deficient dams had elevated serum concentrations of the pro‐inflammatory markers interleukin‐2 and resistin. Maternal vitamin D status during pregnancy does not appear to influence the development of adipose in lean male offspring maintained on a standard diet; however, increases in circulating resistin, an adipose‐derived protein, suggests an adipocyte response.

Grant Funding Source: Supported by: UM Research Investment Fund and SIRC Research Grant
Effects of an acute fructose or fructose and glucose load in a test meal on fructose disposal (1039.3)
Authors: Theytaz Fanny , De Giorgi Sara , Stefanoni Nathalie , Rey Valentine , Hodson Leanne , Fielding Barbara , Lecoultre Virgile , Schneiter Philippe , Tappy Luc
Published: June 2021

Gluconeogenesis, indirect oxidation of fructose‐derived glucose, lactate and glycogen synthesis are major pathways for the disposal of a pure oral fructose load, while de novo lipogenesis (DNL) constitutes a minor pathway. The metabolic fate of fructose co‐ingested with glucose and other nutrients has however not been evaluated. To address this issue, we studied 6 healthy subjects over 6 hours after ingestion of test meals composed of 20±0g lipids and 19±0g protein either without sugars (Ctrl), or with 32±1g 13C‐labelled fructose (F), or with 32±1g 13C‐labelled fructose + 32±1g unlabelled glucose (F+G). Subjects consumed a weight‐maintenance diet containing 20% sugar during 3 days preceding the measurements. Fructose oxidation was assessed from 13CO2 production, gluconeogenesis from 13C‐glucose production, and DNL from post‐prandial 13C‐VLDL‐palmitate concentrations.

Ctrl
F
F+G

Net Carbohydrate oxidation (g/6h)
21.0 ± 3.3
29.9 ± 2.8
34.5 ± 3.9

Fructose oxidation (g/6h)
‐
12.5 ± 0.7
11.0 ± 0.5#

Gluconeogenesis (g/6h)
‐
9.1 ± 0.2
6.4 ± 0.3#

Net glycogen synthesis (g/6h)
‐
4.4 ± 2.1
34.9 ± 5.2#

Fructose storage (g/6h)
‐
21.8 ± 1.2
23.1 ± 1.2

iAUC lactate (mM*6h)
‐51.1 ± 19.6
108.3 ± 11.7*
124.9 ± 26.3*

iAUC13C‐VLDL‐palmitate (Î¼mol*6h)
‐
3.9 ± 1.2
3.7 ± 0.6

All P <0.05, * vs Ctrl, # vs F. iAUC: incremental area under the curve Compared with fructose alone, co‐ingestion of glucose and fructose incorporated in a meal reduced fructose oxidation by 12 ± 5%, and gluconeogenesis by 28 ± 12% by increasing net glycogen storage; but did not enhance 13C‐VLDL palmitate responses. Chronic consumption may lead to increase DNL due to limited glycogen storage.
Dried plum snacking influences nutrient intake in individuals with metabolic syndrome, prediabetes, and overweight (1040.1)
Authors: Reiter Benjamin , Clayton Zachary , Fusco Elizabeth , Kass Lisa , Carpenter Jennifer , Hooshmand Shirin , Hong Mee Young , Kern Mark
Published: June 2021

Dietary intake is an important modifiable risk factor for the metabolic syndrome, prediabetes, and overweight. Dried plums are nutrient‐dense and promote modest acute glycemic and insulinemic responses, which are considered favorable for these populations. Twenty‐one adults meeting the criteria for the metabolic syndrome, prediabetes, or clinical overweight consumed a whole food (dried plums;n=11) or refined (low‐fat muffins;n=10) snack, twice daily for eight weeks. Snacks provided 100 kcal/feeding and were similar in macronutrients but differed in fiber and other constituents. Intake was assessed via multiple 24‐hour food recalls obtained prior to and during the test food intervention. Dried plum consumers ingested greater (p<0.05) daily amounts of potassium, copper, and magnesium during the intervention period than pre‐intervention, whereas muffin eaters exhibited no significant differences in those nutrients but increased thiamin, riboflavin and iron intake. The muffin group consumed more energy during the intervention period than pre‐intervention (p<0.05, 95% CI∆ 61 to 762). Results suggest that proper snack selection is a viable avenue for improving dietary intake in the populations studied.

Grant Funding Source: Supported by the California Dried Plum Board
A pre‐meal glucose drink, but not video game playing, suppresses food intake in overweight and obese boys (1040.8)
Authors: Gheller Brandon , Akhavan Tina , Pollard Damion , Gladanac Branka , Constantino Monika , Luhovyy Bohdan , Bellissimo Nick
Published: June 2021

We previously reported decreased food intake (FI) after a glucose drink and 30 min of video game playing (VGP) in normal weight boys, but the effect on FI in boys with increased adiposity is unknown. Therefore, we examined the effect of 30 min of pre‐meal VGP on subjective appetite, emotions and FI in overweight/obese (>85th BMI percentile) boys after a glucose drink. On four mornings, in random order and one‐week apart, boys (n = 22; age = 11.9 ± 0.3 y) consumed equally sweetened drinks (250 mL) of sucralose (0 kcal) or 50 g (200 kcal) glucose, with or without 30 min of subsequent VGP, 2 h after a standardized breakfast. Immediately after all test conditions FI (mean ± SEM kcal) from an ad libitum pizza meal was measured. Subjective appetite was measured at baseline (0 min), 20, 35, and 65 min (post‐meal). While glucose (p < 0.01) decreased FI (∆ = ‐103 ± 48 kcal) compared with the sucralose control, cumulative FI (drink kcal + meal kcal) was higher (p < 0.01). Fullness (p < 0.05) was lower, and subjective aggressiveness (p < 0.01), excitement (p < 0.01), upset (p = 0.05), and frustration (p < 0.05) higher after VGP; however, subjective emotions did not correlate with FI. In conclusion, the stronger effect of glucose compared with VGP on FI is indicative of the primary role of physiologic factors in the short‐term regulation of FI in overweight/obese boys.

Grant Funding Source: Supported by The Danone Institute of Canada, Grant‐in‐aid program
d‐δ‐Tocotrienol promotes the differentiation of murine MC3T3‐E1 preosteoblasts (1045.37)
Authors: Shah Anureet , Yeganehjoo Hoda , Juma Shanil , DiMarco Nancy , Mo Huanbiao
Published: June 2021

Osteoblasts and osteoclasts play essential and opposite roles in maintaining bone homeostasis. Osteoblasts fill cavities excavated by osteoclasts. The mevalonate pathway provides essential prenyl pyrophosphates for the activities of GTPases that promote differentiation of osteoclasts but suppress that of osteoblasts. Preclinical and clinical studies suggest that mevalonate suppressors such as statins increase bone mineral density and reduce risk for bone fracture. Tocotrienols down‐regulate 3‐hydroxy‐3‐methylglutaryl coenzyme A (HMG CoA) reductase, the rate‐limiting enzyme in the mevalonate pathway. In vivo studies have shown the bone‐protective activity of tocotrienols. We hypothesize that d‐δ‐tocotrienol, a mevalonate suppressor, induces differentiation of murine MC3T3‐E1 pre‐osteoblasts. Alizarin staining showed that d‐δ‐tocotrienol (0‐15 μmol/L) induced mineralized nodule formation in a concentration‐dependent manner in MC3T3‐E1 preosteoblasts. d‐δ‐Tocotrienol (0‐25 μmol/L), but not d‐α‐tocopherol (25 μmol/L), significantly induced alkaline phosphatase activity, an indicator of preosteoblast differentiation. Concomitantly, western‐blot analysis showed that d‐δ‐tocotrienol down‐regulated HMG CoA reductase. d‐δ‐Tocotrienol (0‐25 μmol/L) had no impact on the viability of MC3T3‐E1 preosteoblasts following 48‐h incubations, suggesting lack of cytotoxicity at these doses. Tocotrienols and other mevalonate suppressors have potential in maintaining bone health.

Grant Funding Source: Supported by USDA NIFA Agriculture and Food Research Initiative Grant 2009‐02941, TDA & TWU REP.
Molecular mechanisms of conjugated linoleic acid (CLA) on muscle metabolism in adult onset inactivity‐induced obese mice (1045.46)
Authors: Kim Yoo , Good Deborah , Park Cheon‐Seok , Park Yeonhwa
Published: June 2021

Conjugated linoleic acid (CLA) has been reported to increase voluntary activities in mice. Current study was conducted to investigate how CLA modulates activity levels by determining the molecular targets on skeletal muscle in nescient basic helix‐loop‐helix 2 knock‐out (N2KO) mice, a unique adult‐onset inactivity induced obesity model. 4‐week‐old female N2KO and wild‐type mice were fed either control or CLA containing diet (0.5%) for 10‐weeks. Non‐exercise physical activity (NEPA) was determined biweekly and markers for muscle metabolisms were determined from the gastrocnemius. CLA fed N2KO animals showed significant increase of NEPA (~40%) from 6 weeks compared to controls. The gastrocnemius weights were lower in N2KO animals compared to wild types, while CLA supplementation resulted in heavier the gastrocnemius over controls in N2KO animals. Protein expressions for peroxisome proliferator‐activated receptor‐δ (PPARδ) and peroxisome proliferator‐activated receptor gamma co‐activator 1α (PGC‐1α) in the gastrocnemius were increased in CLA fed N2KO animals compared to controls, 65% and 38% increase, respectively. CLA feeding increased pAMPK/AMPK ratio, 36% increase, compared to controls. These results suggest that CLA act as a potential exercise‐mimetic, resulting in increased voluntary activity and improving muscle metabolisms in both normal and obese animals.
Broccoli bioactives inhibit human prostate cancer cell invasions in vitro (1045.5)
Authors: Huang Ching‐Yu , Jeffery Elizabeth , Erdman John
Published: June 2021

Previous studies have found that various bioactives in broccoli may have anti‐cancer activities, including anti‐invasion ability. We hypothesized that regular consumption of broccoli may inhibit prostate cancer via inhibition of invasion. Thus, the invasive human prostate cancer cell line, DU145, was treated with two broccoli bioactives—sulforaphane and indole‐3‐carbinol. Cells were seeded in matrigel inserts in 24‐well plates containing complete medium as the chemoattractant. Treatments were renewed at the 24th hour. After a total of 48 hours treatment, the matrigel layer was removed, and cells on the bottom membrane were stained and counted. The invasive ability of DU145 was significantly suppressed in a dose‐dependent manner by treatment with sulforaphane at concentrations as low as 3µM. Indole‐3‐carbinol also significantly reduced invasion, but at concentration as low as 0.2µM. In summary, two major bioactives in broccoli show anti‐invasion activity in vitro, at physiologically‐achievable concentrations.
Expressional profiling of carcinoembryonic antigen‐related microRNAs in human gastric cancer (1047.14)
Authors: Zhang Guo , Ye Hui‐Lan , Yao Ping , Nong Bing , Li Jing
Published: June 2021

MicroRNAs (miRNAs) are a group of non‐coding small RNAs (ncRNAs) that negatively regulate gene expression at the post‐transcriptional level in cellular differentiation and carcinogenesis. Given that carcinoembryonic antigen (CEA) is commonly used for diagnosis of GI cancer for decades, the present study aimed to screen and identify the CEA related miRNAs in gastric cancer tissue and determine their potential pathophysiologic relevance as well. In this study, a total of fivety gastric tissue samples inculing 17 pairs of cancer and adjacent tissure and 16 healthy volunteers control were collected and confirmed by endoscopic biopsy. The microRNA expression was then deteted and profiled in these samples by using QuantoBio three steps based q‐PCR. QuantoIC‐1 and QuantoIC‐2 were simultaneously conducted as the internal controls. As results, miR‐10a,miR‐18a* as well as miR‐27a* are significantly up‐regulated and miR‐148a* down‐regulated in the carcinoma tissue and additionally, miR‐10a, miR‐122*, miR‐129‐5p and miR‐378 are dramatically up‐regulated in pericarcinous tissue versus healthy control. Compared with pericarcinous control, miR‐10a, miR‐148a*, miR‐193b and miR‐378 are apparently down‐regulated in cancer. Interestingly, further bioinformatics analysis has shown that CEA‐mRNA molecule was found to be a common target for all these miRNAs. Altogether, our date indicated that multiple miRNAs were substantially involved in the gastric cancer development. Such miRNAs as miR‐18a* and miR‐10a with increase expressional pattern in stomach carcinoma may potentially servers as novel oncogenes during the process of this malignant desease.

Grant Funding Source: Supported by 2010GXNSFC013020 to G.Z.
Is cigarette smoking an independent risk factor of metastatic disease in prostate cancer? (1047.15)
Authors: McHowat Jane , Kispert Shannon , Marentette John , Powell Jordan
Published: June 2021

Cigarette smoking is a major risk factor in many health conditions, including cancer. Platelet‐activating factor (PAF) may contribute to cancer metastasis by increasing adherence of tumor cells to activated endothelium via the PAF‐PAF receptor interaction. We exposed PC‐3 prostate cancer cells to cigarette smoke extract (CSE) and measured PAH acetylhydrolase activity (PAF‐AH, inactivates PAF), PAF accumulation, PAF receptor expression and adherence to human lung microvascular endothelial cells (HMVEC‐L). CSE incubation (20 μg/ml, 4‐24 hours) resulted in a significant inhibition of PAF‐AH activity, and increased PAF accumulation in PC‐3 cells. Incubation of PC‐3 cells with CSE resulted in a 5‐fold increase in PAF receptor expression on the cell surface and increased adherence to HMVEC‐L. Pretreatment of PC‐3 cells with Ginkgolide B blocked adherence to the endothelium. Our data demonstrate that prostate cancer patients who smoke may be at increased risk for metastasis and that the nutraceutical Ginkgo biloba may be an adjunctive therapy to manage this disease.
Antiangiogenic therapies and aged‐associated changes in the prostate: a comparison between the prostate of aging mice (FVB) and TRAMP mice (1048.7)
Authors: Cagnon Valeria , Montico Fabio , Kido Larissa , Hetzl Amanda
Published: June 2021

Senescence and angiogenesis are related to prostate alterations. The aim was to characterize TGF‐β, FGF‐2, IGFR‐1, MMP‐9, VEGF and endostatin responses in the prostate of the elderly mice submitted to antiangiogenic and finasteride therapies in relation to the prostate of TRAMP mice. Experimental groups: Young group; Prostatic Intraepithelial Neoplasia and Prostate Cancer groups; Senile group; SU5416 group; TNP‐470 group; SU5416+TNP‐470 group; Finasteride group; SU5416+TNP‐470 and Finasteride group. After 21 days, prostates were collected for light microscopy, immunohistochemistry and western blotting. Senescence led to inflammation and proliferative processes. Increased MMP‐9, IGFR‐I, VEGF, FGF‐2 and TGF‐β levels were verified in senile mice. TRAMP mice presented MMP‐9, IGFR‐1, VEGF, FGF‐2 increase. TGF‐β level did not increase in the cancer group. Antiangiogenic therapies showed VEGF, TGF‐β, MMP‐9 and IGFR‐1decrease. The indirect angiogenesis inhibitor acted mainly on MMP‐9 and the direct inhibitor in IGFR‐1 levels. Senescence predisposes prostatic malignant changes. The balance between anti and pro‐angiogenic factors was evident in the combination of the three drugs. The activation of distinct pathways by each drug differentially influenced factors involved in tumor‐promoting processes. TGF‐β could be involved in the preneoplastic lesions.

Grant Funding Source: Supported by Research Foundation of the State of Sao Paulo (Fapesp 2012/03010‐4 / 2011/01968‐30
Goniothalamin and Celocoxib therapies on the preneoplastic and neoplastic prostatic lesions in the transgenic adenocarcinoma of the mouse prostate model (TRAMP) (1048.8)
Authors: Cagnon Valeria , Kido Larissa , Montico Fabio , Dall Pozzo Caroline , Carvalho João , Costa Debora , Pilli Ronaldo
Published: June 2021

Inflammation plays an important role on prostate diseases. The aim of this study was to evaluate and compare the structural and molecular features of the ventral lobe from TRAMP mice undergoing Celecoxib treatment, a nonsteroidal anti‐inflammatory, and Goniothalamin, a synthetic substance, exhibiting anti‐inflammatory and antiproliferative properties. The mice were divided into: Control groups; Tramp: 8 week‐old mice; Tramp:12‐week‐old mice. Treated groups: Celecoxib‐10mg/kg; Goniothalamin‐150mg/kg. The ventral lobe was collected after 4 weeks for light microscopy, immunohistochemistry and western blotting. Structural analysis showed that TRAMP mice developed progressive forms of preneoplastic and neoplastic lesions during their lifetime. Both treatments led to a decrease for cell proliferation and also inflammatory cells. The Celocoxib treatment showed decreased COX‐2, IGFR‐1, STAT‐3 and IL‐17 immunoreactivities. Goniothalamin treatment displayed a stronger IGFR‐1, STAT‐3 and IL‐17 reduction, in contrast to COX‐2 with no change. Thus, both treatments were effective in reducing the progression of prostatic lesions and inflammation. However, the Goniothalamin treatment suggested the most significant effect on the pro‐inflammatory cytokine and on a transcription factor signal transducer, acting in many pro‐oncogenic signals, thus delaying prostate cancer progression.

Grant Funding Source: Research Foundation of the State of Sao Paulo (2012/03010‐4/ 2013/01294‐8)
Direct detection of green fluorescent protein in histological preparations of recombinant vaccinia virus in mice (1050.9)
Authors: Lyons Cassandra
Published: June 2021

Green fluorescent protein (GFP)‐expressing recombinant viruses have proven to be invaluable tools for the in‐vivo monitoring of viral kinetics and disease progression in infectious disease models. Histopathological evaluation of the ex‐vivo tissues by way of immunohistochemical (IHC) staining for viral antigens further assists in the characterization of disease pathology, but can be insensitive, nonspecific, and time consuming. In this study, we determine if the direct detection of GFP signal provides a suitable alternative to IHC staining in GFP‐expressing tissues. Fixation and histological processing protocols were developed in a GFP‐expressing 293T cells and then optimized and validated in a GFP‐recombinant rat model. The optimized protocol was then applied to GFP‐recombinant cowpox virus (VACV‐TK‐GFP) infected mouse lungs in addition to lungs infected with a wild‐type virus strain (VACV‐WR). While animals infected with both virus strains manifested lesions reactive to anti‐Vaccinia IHC, GFP fluorescence was only detected in lesions from animals infected with the recombinant virus strain. The method was further validated through IHC staining of both virus strains with anti‐GFP antibodies, which were only reactive in tissues infected with VACV‐TK‐GFP. Digital alignment of sections showed a strong correlation between GFP fluorescence and IHC staining for GFP and viral proteins suggesting direct‐detection of fluorescent proteins is a promising alternative to traditional methods.
Hyperhomocysteinemia aggravates endothelial dysfunction via EDHF impairment in resistant arteries of db/db mice (1051.13)
Authors: Cheng Zhongjian , Jiang Xiaohua , Fang Pu , Yang Xiaofeng , Wang Hong
Published: June 2021

Aims: The objective of this study was to examine the effect of hyperhomocysteinemia (HHcy)─elevated plasma Hcy level on endothelial function in resistant arteries of type 2 diabetic db/db mice.

Methods and Results: Moderate HHcy (plasma tHcy=54.31±5.4 µmol/L) was developed in diabetic db/db mice fed with a high methionine diet (HM, 2% methionine) for 8 weeks starting from age of 8 weeks, whereas non‐diabetic db/+ mice developed mild HHcy (plasma tHcy=34.21±4.15 µmol/L). HHcy increased blood glucose levels from 474±29 mg/dl to 630±83 mg/dl in diabetic mice (p<0.05), whereas HHcy did not affect blood glucose levels in non‐diabetic mice. Endothelium‐dependent vascular relaxation to acetylcholine (Ach) was significantly impaired in the small mesenteric artery (SMA) of diabetic mice. Mild HHcy did not impair endothelium‐dependent vascular relaxation response to Ach in non‐diabetic mice. Moderate HHcy aggravated the impairment of vascular relaxation response to Ach in diabetic mice both in the presence or absence of eNOS inhibitor L‐NG‐nitroarginine methyl ester (L‐NAME). HHcy‐induced impairment of relaxation response to Ach in diabetic mice was diminished in the presence of L‐NAME plus intermediate‐conductance calcium‐activated potassium channel (IKCa) inhibitor tram‐34 and small conductance KCa (SKCa) blocker apamin. Our results suggest that impairment of endothelium‐derived hyperpolarizing factor (EDHF) via inactivation of IKCa and SKCa plays a critical role in HHcy‐aggravated endothelial dysfunction in the resistant artery of diabetic mice. Moreover, 40Gap27 and 43Gap26 significantly improved vascular relaxation response to Ach in mice with moderate HHcy and diabetes. Finally, HHcy significantly impaired endothelium‐independent vascular relaxation to sodium nitroprusside in diabetic mice.

Conclusions: These findings demonstrate that HHcy‐induced impairment of EDHF may be resulted from inactivation of IKCa/SKCa and suppression of connexin 40 and connexin 43.

Grant Funding Source: NIH
Reversal of diabetic stem‐cell mobilopathy by angiotensin‐(1‐7) via rho‐kinase downregulation (1051.5)
Authors: Vasam Goutham , Raizada Mohan , Jarajapu Yagna
Published: June 2021

Mobilization of bone marrow stem/progenitor cells (BMSPCs) that maintain cardiovascular health is impaired in diabetes. We have previously shown that Angiotensin (Ang)‐(1‐7) increases the vasoreparative potential of dysfunctional BMSPCs from individuals with diabetes. In this study we tested the effect of Ang‐(1‐7) on the impaired mobilization of BMSPCs in diabetes. Furthermore, the role of Rho‐kinase in the beneficial effects of Ang‐(1‐7) was evaluated. Lean control or obese db/db (type 2 diabetic) (DB) mice were treated with continuous infusion of saline or Ang‐(1‐7) (1 µg/Kg/min) for four weeks by using subcutaneous osmotic pumps. Subsets of peripheral blood lineage‐negative cells, Lin‐Sca‐1+, Lin‐cKit+ and Lin‐Sca‐1+cKit+ cells, were enumerated after four weeks of treatment. All subsets of BMSPCs were lower in DB (compared to control, P<0.05), and were increased by Ang‐(1‐7) treatment to control levels. The number and proliferation of BM‐resident SPCs, and the migration to stromal‐derived factor‐1α (SDF, 100 nM) ex vivo were reduced in DB compared to control group (P<0.01) and this dysfunction was reversed to normal by Ang‐(1‐7) treatment. Expression of Rho‐kinase isoforms, ROCK‐1 and ROCK‐2, were higher in DB‐BMSPCs (compared to control, P<0.01) that was down‐regulated by Ang‐(1‐7) to normal levels. Furthermore, ex vivo treatment of BMSPCs from DB with a non‐specific ROCK inhibitor, Y27632 (10 μM), restored their migration to SDF. These results suggest that Ang‐(1‐7) increases BMSPC‐mobilization in diabetes, in part, via down‐regulation of ROCK. Together with our previous observations, Ang‐(1‐7) appears to be a promising target for the treatment of diabetic stem‐cell mobilopathy and for enhancing the outcomes of cell‐based therapies in individuals with diabetes.

Grant Funding Source: Supported by American Heart Association‐13SDG16960025
Modulating myofibroblast transition in systemic sclerosis through inhibition of Rho/MRTF regulated transcription (1054.9)
Authors: Haak Andrew , Tsou Pei‐Suen , Fox David , Khanna Dinesh , Larsen Scott , Neubig Rick
Published: June 2021

Systemic sclerosis (SSc) or scleroderma, like many fibrotic disorders is a deadly disease lacking effective therapies. Targeted strategies which are currently in development are focusing on blocking individual receptors known to be involved in the myofibroblast transition. However, in light of recent evidence that Rho GTPase/myocardin‐related transcription factor (MRTF) regulated transcription is an essential and convergent downstream mechanism for myofibroblast transition; we explore the hypothesis that inhibitors of this pathway may represent novel antifibrotics. Consistent with previous findings, fibroblasts from SSc patients expressed markers of activated myofibroblasts but additionally, Rho/MRTF regulated genes were found to be elevated (4‐6 fold) in these samples. At low micromolar concentrations, a novel small‐molecule inhibitor of Rho/MRTF regulated transcription (CCG‐203971) blocked RNA and protein expression of multiple pro‐fibrotic markers including connective tissue growth factor (CTGF), alpha‐smooth muscle actin (α‐SMA), and collagen 1 (COL1A2) in both SSc fibroblasts and transforming growth factor β (TGFβ)‐stimulated normal donor samples. In a mouse model for scleroderma, treatment with CCG‐203971 also prevented bleomycin‐induced skin thickening and collagen deposition. These data suggest targeting the Rho/MRTF gene transcription pathway could provide a novel approach to therapy for SSc and other fibrotic disorders.
TPA‐induced invasion of HT29 cells are mediated through ROS production and AMPK deactivation (1055.5)
Authors: Banskota Suhrid , Kim Jung‐Ae
Published: June 2021

AMP‐activated protein kinase (AMPK) is a member of serine/threonine protein kinase which serves as an energy sensor in all eukaryotic cells. In the present study, the role of AMPK in cancer cell metastasis was investigated. We found that 12‐O‐tetradecanoyl phorbol‐13‐acetate (TPA), a tumor promoter, induced invasive behavior of HT‐29 colorectal cancer cells, which corresponded to increased ROS production and MMP‐7 expression, whereas TPA suppressed phosphorylation of AMPK in the cells. The TPA‐induced reactive oxygen species (ROS) production and invasion of HT29 cells were inhibited by NADPH oxidase inhibitors (apocynin and diphenyleneiodinium), ROS scavengers (Vitamin C and Vitamin E), and AMPK activator (AICAR). Taken together, TPA induces transition of epithelial HT29 colon carcinoma cells to invasive mesenchymal phenotype through NADPH oxidase activation and AMPK inactivation.

Grant Funding Source: This work was supported by the National Research Foundation of Korea(NRF) grant funded by the Korea
Symmetry considerations provide a new novel open channel blocker of GABA‐A receptors (1059.2)
Authors: Sigel Erwin , Pangerl Michael , Carta Valentina , Baur Roland , Trauner Dirk
Published: June 2021

The major GABA‐A receptor isoform in the mature central nervous system is α1β2γ2. Upon activation, the GABA‐A receptor conducts Cl‐ through its pore, causing an inhibitory effect on neurotransmission by diminishing the chance of action potential firing. The GABA‐A receptor is inhibited by epileptogenic agents such as picrotoxin, which act as a noncompetitive inhibitor of GABA‐induced currents. We screened molecules with near five‐fold symmetry for inhibition of GABA‐A receptors. α1β2γ2, receptors were expressed in Xenopus oocytes. One of the molecules, MP7, inhibited currents elicited by GABA with comparable potency as picrotoxin. This inhibition showed all characteristics of an open channel block. At lower concentration of GABA an additional potentiation was evident. MP7 also inhibited α1β1γ2 and α1β2δ GABA‐A receptors, homomeric and heterogenic glycine receptors and Drosophila insecticide resistant GABA‐A receptors. At higher concentrations (> 10 μM) MP7 by itself induced small permanent and huge transient currents upon voltage jumps, indicating that MP7 can dissolve in the membrane and permeate to some extent. To identify important residues of the pore‐lining segment M2 involved in the interaction with the blocking molecule MP7, point mutations to cysteine were introduced. The reactivity with water‐soluble cysteine‐reactive reagents and the reduction of this reactivity by MP7 showed that the blocker acts in the transmembrane segment M2 at the level of α1V257 / α1T261. MP7 could potentially have use as insecticide.

Grant Funding Source: Supported by the Swiss National Science Foundation grant 31003A‐132806/1 to E.S.
Assessment of drug‐induced changes in visual processing using the optomotor response in rodents (1060.5)
Authors: Hsu Cathleen , Tse Karen , Cannon Keri
Published: June 2021

Drug‐induced ocular toxicity is one of the major obstacles that would halt clinical development of a drug. One nonclinical method of detecting alterations in visual processing may be to assess the effects of a drug on optomotor responses (OR). The present study examined OR in rats as an indication of visual dysfunction using two compounds, quinine and sodium iodate, that are known to produce visual impairment in humans. Male Wistar Han rats were administered vehicle or quinine (100 or 300 mg/kg, p.o.) and OR was assessed at predose, 30, and 120 minutes postdose using the OptoMotry system (CerebralMechanics Inc.). Quinine elicited a dose‐dependent decrease in OR at both timepoints . Furthermore, locomotor activity was assessed in these rats at 30 minutes postdose. Quinine (100 mg/kg, p.o.) resulted in a trend towards decreased activity whereas 300 mg/kg produced significant decreases in locomotor activity. This suggests that decreases in activity did not interfere with the OR at 100 mg/kg but may have affected the measurement at 300 mg/kg. In another study, OR was assessed in rats up to 4 weeks after administration of sodium iodate (50 mg/kg, i.v.) Sodium iodate produced a significant reduction in OR from 2 hours to 1 week postdose. Reduced locomotor activity was detected at 2 hours postdose in sodium iodate‐treated rats. These decreases subsided by 24 hours postdose, suggesting that decreased OR observed at 24 hours and 1 week postdose were not due to decreased activity. Although factors such as sedation may interfere with the ability to assess the OR, the results were in agreement with published literature. Altogether, assessing OR in rodents may be beneficial in characterizing the adverse effects of test compounds on visual perception and acuity.
In vivo inhibition of hepatitis C virus infection by anti‐Claudin‐1 monoclonal antibodies (1062.9)
Authors: Iida Manami , Nagase Shotaro , Yamashita Mayo , Shirasago Yoshitaka , Fukasawa Masayoshi , Tada Minoru , Ishii Akiko , Watari Akihiro , Yagi Kiyohito , Kondoh Masuo
Published: June 2021

More than 170 million people worldwide are infected with hepatitis C virus (HCV). Despite concerted research efforts, anti‐HCV agents resistant to the high mutability of the HCV RNA genome have not yet been produced. Claudin 1 (CLDN1) is a co‐receptor for HCV entry into hepatocytes and is an attractive potential target for HCV therapy, because such host factors are genetically stable. Here, we investigated the effects of 4 previously generated mouse anti‐CLDN1 antibodies on HCV infection. All 4 clones attenuated the infection of Huh7 cells by HCV (genotype 2a) and HCV pseudoviral particles (HCVpp, genotypes 1b and 2a). Two of 4 clones also prevented HCV (genotype 1b) infection of human‐liver‐chimeric mice. Injection of mouse anti‐CLDN1 antibodies did not adversely affect serum human albumin levels, apparent character, and motility. For future clinical application, we prepared human‐mouse chimeric anti‐CLDN1 antibodies. The chimeric antibodies prevented HCV and HCVpp infection of Huh7 cells as effectively as did the mouse antibodies. Our findings suggest that mouse and mouse‐human chimeric anti‐CLDN1 antibodies prevent HCV infection and may therefore be promising candidate anti‐HCV agents.

Grant Funding Source: supported by the Ministry of Health, Labor, and Welfare of Japan
Contribution of guanine nucleotide exchange factor Vav2 to homocysteine‐induced NLRP3 inflammasome activation in mouse podocytes (1063.6)
Authors: Abais Justine , Xia Min , Boini Krishna , Li Pin‐Lan
Published: June 2021

NADPH oxidase‐derived reactive oxygen species (ROS) have been demonstrated to mediate the activation of podocyte NLRP3 inflammasomes in response to elevated levels of homocysteine (Hcys). We have also previously provided evidence that the guanine nucleotide exchange factor Vav2 exhibits high specificity to Rac1‐mediated NADPH oxidase activation by elevated Hcys. The precise role of Vav2 in NLRP3 inflammasome activation remains unknown. The present study was designed to test whether Vav2 contributes to Hcys‐induced NLRP3 inflammasome activation through activation of NADPH oxidase. In our experiments, podocytes were transfected with either a scramble, constitutively active form of Vav2 (oncoVav2) or Vav2 shRNA plasmid directly to the nucleus via nucleofection. Confocal microscopic analysis showed Hcys treatment as well as oncoVav2 transfection alone increased colocalization between inflammasome proteins NLRP3 with ASC and NLRP3 with caspase‐1, suggesting NLRP3 inflammasome formation. Additionally, both Hcys treatment as well as oncoVav2 transfection alone resulted in increased caspase‐1 activation and IL‐1β production, signifying NLRP3 inflammasome activation. These Hcys‐induced measures of inflammasome formation and activation were all inhibited by Vav2 shRNA transfection, observed in decreased colocalization of inflammasome components, caspase‐1 activity, and IL‐1β production in podocytes. Furthermore, Vav2 shRNA transfection also prevented podocyte damage by restoring impaired VEGF secretion as well as decreased podocyte marker podocin expression seen in Hcys injured podocytes. Together, these findings suggest that NADPH oxidase activation by Vav2 overexpression by oncoVav2 was sufficient to activate NLRP3 inflammasomes, independent of Hcys treatment. However, Vav2 inhibition was able to inhibit Hcys‐induced NLRP3 inflammasome activation and podocyte injury, further substantiating the role of Vav2 in this mechanism.

Grant Funding Source: Supported by NIH grants DK54927 and 1F31AG043289
Application of physiologically‐based pharmacokinetic modeling in hepatic impairment populations (1064.9)
Authors: Sager Jennifer , Hsu Matt , Isoherranen Nina , Wienkers Larry , Wahlstrom Jan , Foti Robert
Published: June 2021

Diagnosis of hepatic impairment (HI) is associated with a reduction in hepatocyte expression, a decrease in albumin and α‐1‐acid glycoprotein and a reduction in renal function, the combination of which can result in alteration of the pharmacokinetic (PK) properties of a drug. Therefore, regulatory agencies such as the U.S. Food and Drug Administration often require the assessment of PK parameters in HI populations. These studies can entail single or multiple‐dose approaches and must be of sufficient duration to capture the terminal half‐life of the drug, a parameter which may be extended in HI populations. The use of physiologically‐based pharmacokinetic (PBPK) modeling, a technique combining demographic and genetic factors, anatomical and physiological parameters and drug‐specific properties, can aid in the prediction of PK parameters and the design of clinical trials in HI populations. To that end, the utility of PBPK modeling in predicting unique clinical scenarios in HI populations was evaluated for a number of compounds including sensitive P450 substrates and drugs cleared by multiple P450s. Drug interactions, enantioselective PK changes and effects on hepatic versus intestinal CYP3A4 metabolism due to HI were predicted. Results suggest that PBPK modeling is a useful tool for predicting changes in PK parameters under multiple conditions and for contributing to the design of HI clinical trials.
Early life stress induces bladder dysmotility in adult rats (1065.17)
Authors: Kennedy Dawn , Mohammadi Ehsan , Hurst Robert , Greenwood‐Van Meerveld Beverley
Published: June 2021

Symptoms of painful bladder syndrome (PBS) including urgency and pain are common in female patients who report early life stress (ELS) such as neglect or abuse during childhood. We hypothesized that ELS contributes to PBS by inducing abnormal bladder detrusor muscle contractility in adulthood. This study used an odor‐attachment model in which female rats were conditioned on postnatal days (PND) 8‐12 using predictable or unpredictable odor‐shock conditioning, or an odor only control. In adulthood, bladder muscle strips were harvested and mounted in a muscle bath to measure contractile responses to neurally mediated signalling (electrical field stimulation‐EFS), muscarinic receptor‐dependant signalling with carbachol, and membrane depolarization with KCl. Detruser muscle responses to EFS were significantly decreased (P<0.001) in the unpredictable (12.6±2g/cm2) compared to the predictable (24.9±2g/cm2) or odor only (24.0±1.3g/cm2) groups. Detrusor muscle responses to CCh or KCl were not significantly different between groups. Our data indicate that ELS alters neurally mediated detrusor muscle contractility in the absence of abnormalities in smooth muscle myogenic responses. These findings suggest that therapeutics targeting neural innervation of the bladder musculature could effectively reduce PBS symptoms by normalizing bladder contractility in adulthood.

Grant Funding Source: This work was supported by a department of Veterans Affairs Merit Grant to Dr. Beverley Greenwood‐Va
Essential role of mTORC1 in the hypertension induced by overactivation of the brain renin‐angiotensin system (1067.2)
Authors: Muta Kenjiro , Grobe Justin , Sigmund Curt , Rahmouni Kamal
Published: June 2021

Angiotensin II (Ang II) is a major effector peptide of the renin‐angiotensin system (RAS). Ang II‐induced activation of angiotensin type 1 receptor (AT1R) signaling in cardiovascular‐related brain nuclei such as the subfornical organ (SFO) is thought to be important for the development of hypertension. However, the molecular machinery underlying the central pressor actions of Ang II is poorly defined. We previously demonstrated the involvement of neuronal mammalian target of rapamycin complex 1 (mTORC1) in blood pressure (BP) regulation. Interestingly, using in vitro systems we also found that mTORC1 is a downstream element of neuronal AT1R signaling. To assess the relevance of mTORC1 for Ang II action, we first examined mTORC1 activity in the cardiovascular brain regions in hypertensive transgenic mice with brain‐specific hyperactive RAS (sRA mice). Strikingly, the number of phospho‐S6 positive cells (used as a readout of mTORC1 activity) was significantly higher in the SFO of sRA mice (64±6) relative to controls (36±8, p<0.05, n=5). Moreover, central administration of an adenovirus encoding a dominant negative S6 kinase (an mTORC1 substrate) lowered systolic BP in sRA (from 148±8 to 130±8 mmHg, p<0.05, n=3) to comparable levels as control mice receiving GFP adenovirus (from 125±7 to 126±1 mmHg, n=3). These results indicate that neuronal mTORC1 activation is a critical event in brain RAS‐induced BP elevation.