bluuebunny/crossref_metadata_2025 huggingface dataset:
Active transcription factors in the development of the small intestine during early life
Authors: Zhang Man , Liao Yalin , Lönnerdal Bo
Published: June 2021

Transcription factors (TFs) regulate downstream gene expression individually or cooperatively in different cell types and tissues. Human milk contains growth factors that maintain gastrointestinal mucosal homeostasis. We chose two major growth factors in human milk, TGF‐â (10 ng/ml) and IGF‐1 (30 ng/ml) to study global activation of TFs during early intestinal development in IEC‐6 (crypt cell) and FHs74 cells (fetal intestine). Active transcription factors were detected by Protein‐DNA Array. Among all the active TFs, EGR‐1 expression showed the most substantial up‐(IEC‐6 cell differentiation) and down‐regulation (FHs74 cell proliferation) after stimulation of the two cell lines. For IEC‐6 cells, EGR‐1 is predicted to have binding sites in the promoters of the Axin1 (‐258bp/260bp) and LRP5 (‐84bp/113bp) genes, which are mediators involved in the Wnt signaling pathway. The binding sites were verified by chromatin immunoprecipitation (CHIP) and EMSA. Changes in the Wnt/â‐catenin downstream targets Cyclin D and E‐cadherin further indicate that EGR1 interferes with the Wnt pathway through Axin1 and LRP5. In conclusion, we showed transcription factors regulated by growth factors during small intestinal development. EGR1 was further found to regulate the Wnt pathway through the multiprotein complex, which shows that human milk growth factors are likely to impact small intestinal development considerably.

Grant Funding Source: research grant from Mead Johnson Nutrition
Bacterial flora in human milk in health and disease
Authors: Kvist Linda , Schalén Claes
Published: June 2021

Background

the presence of bacteria in breast milk of lactating women was described more than 30 years ago and it was assumed that the cause was contamination of specimens from the mothers’ skin and infants’ mouths (Gavin & Ostovr 1977). A recent Swedish study showed that many healthy women harbour potential pathogens in their breast milk at similar concentrations to those with mastitis (Kvist et al, 2008). New methods of DNA sequencing of bacteria will revolutionize our understanding of the bacterial content of human milk. New knowledge must either be synthesized with the old or replace it.

Aim

to provide a synthesis and a comparison of findings from recent empirical research on microbial content of human breast milk in healthy women and women with mastitis in order to clarify present day knowledge.

Method

a synthesis of the literature regarding bacterial content of breast milk from healthy women and women with mastitis, published between 1990 and 2010. This resulted in an analysis of milk from 844 healthy women and 624 women with mastitis.

Findings

seven articles reported bacterial content in breast milk from healthy women, four reported findings from women with symptoms of mastitis and one study reported on bacterial content of milk from both groups. The isolate most commonly found in women with mastitis was S. epidermis (45.5%) and in healthy women the most common bacterial isolate was CNS (77.4%). S.aureus was reported in 5.1% of women with mastitis.

Conclusions

a bacterial flora appears to exist in all breast milk. It is unclear why reports of sterile breast milk occur in the literature from time to time. The results of this synthesis challenge the conventional claim, that S. aureus is the most common cause of lactational mastitis.
Multiplexing of Human Preterm and Term Milk Cytokines
Authors: Groer Maureen Edith , Williams S. Nicole , Kane Bradley Patrick
Published: June 2021

The first objective was to use an 18‐plex cytokine panel to compare preterm and term colostrum collected at day 1 postpartum. After observing matrix effects, we worked to develop a milk assay matrix for Luminex and MagPix technologies. After matrix development proved satisfactory, we have progressed to a longitudinal study of immunobiology of milk from 100 mothers of Very Low Birth Weight (VLBW) infants.

We multiplexed milk whey using bead based methods and performed linear spiking experiments. We used a variety of matrix solutions and these matrices did not give reliable results. When we used serum matrix for controls and standards, and added serum matrix to milk samples, the spiking experiments were successful and showed linearity.

Based on our preliminary experiments on preterm and term colostrum and the matrix solution, we designed a plex of 8 cytokines that differed in preterm compared to term colostrum (IL‐4, IL‐6, IL‐8, IL‐10, IP‐10, MIP‐1α, IP‐10, MCP‐1, and TNFα). We are now measuring milk cytokines from 100 mothers of VLBW infants across 6 weeks of the NICU stay. From these data we will describe the trajectory of human milk immunobiology over the first six weeks postpartum.

Multiplexing provides excellent screening for important immune molecules. However there are no plexes specifically for milk. We have developed an assay for multiplexing using serum matrix that is sensitive and reliable.

Funded by 5R21NR013094–02
Anti‐obesity Effect of Bamboo Salt In Vivo
Authors: Park Kun‐Young , Ju Jae‐hyun , Song Jiale
Published: June 2021

This study was aimed to determine the anti‐obesity effect of bamboo salt in vivo. For this purpose, we divided C57BL/6 mice with various groups on the basis of the treatment (normal diet, high‐fat diet (HFD), HFD + purified salt treatment, HFD + solar salt treatment, HFD + bamboo salt treatment (1×, 3× and 9× baked), feeding mice of each salt‐fed group with the dosage corresponding with the concentration twice as high as recommended salt intake of human (5 g). After 8 weeks, we measured serum lipid profile (triglyceride and total cholesterol) and epididymal adipose tissue. Concomitant feeding of HFD and bamboo salt reduced the overall body weight, weight of epididymal adipose tissue and food efficacy ratio, compared with high‐fat diet group and/or other high‐fat diet + salt samples. Feeding of bamboo salt also lowered serum lipid profiles and ALT/AST in comparison with the high‐fat diet group. We also observed changes in expression of obesity‐related factors (C/EBPα, SREBP‐1c and FAS) and inflammation‐related factors (PPAR‐γ and IL‐6). Taken together, this study suggests that treatment of bamboo salt in mice might have a preventive effect on obesity.
Tea extracts modulate fat accumulation and distribution and demonstrate anti‐inflammatory effects in C57BL/6 mice fed a high fat/Western diet
Authors: Zhang Yanjun , Yang Jieping , Henning Susanne , Heber David
Published: June 2021

Tea may be effective in the prevention of obesity‐related chronic disease. We compared the effect of 3 teas including green (GT), black (BT) and oolong tea (OT) on body weight, fat distribution and inflammation. 120 C57BL/6 male mice were randomly assigned to 5 treatment groups: control diet (10% kcal from fat), high fat (HF)/Western diet (32% kcal from butter fat and corn oil), or a HF diet with GT, OT or BT extract (0.25% GT polyphenols) for 20 weeks. 3–6 mice per group were sacrificed every 4 weeks. Fat distribution was measured by magnetic resonance imaging (MRI) and fat pads were weighed. Gene expression of interleukin (IL)‐6, tumor necrosis factor (TNF) α and MCP‐1 was determined in liver, epididymal (E‐fat) and mesenteric fat (M‐fat) using qRT‐pCR. The addition of all 3 tea extracts significantly reduced the HFdiet‐induced increase in body weight, liver fat, M‐fat, and subcutaneous fat. GT and BT significantly decreased the amount of E‐fat and GT and OT significantly decreased perirenal fat. GT and BT significantly inhibited HFdiet‐induced MCP‐1 gene expression in the liver, and BT in both E‐fat and M‐fat. All 3 teas significantly reduced the HFdiet‐induced TNF α expression in E‐fat, but not in M‐fat. The addition of BT to the HFdiet led to significant inhibition of IL‐6 concentration in E‐fat. In summary the addition of tea extracts to an HFdiet differentially alleviated HFdiet‐induced fat accumulation and inflammation
Caffeic acid and ferulic acid improves diet induced metabolic syndrome on mice but induces inflammatory process in white adipose tissue
Authors: Bocco Barbara Miranda Leite da Costa , Lorena Fernada Beraldo , Fernandes Gustavo Werpel , Lancelloti Carmem Lucia Penteado , Cysneiros Roberta Monterazzo , Ribeiro Miriam Oliveira
Published: June 2021

Several studies have been shown that Caffeic Acid (CA) and Ferulic Acid (FA) present antioxidant and anti‐inflammatory properties and improve several abnormalities related to metabolic syndrome. The aim of this study was to analyze the effect of CA and FA combined on diet induced obesity in mice. For that, C57 mice fed with high fat diet (HFD) (40%) received CA plus FA concomitantly for 60 days. Our results showed that the combination of CA (0.9 or 1.8 mg/kg/day) with FA (50 mg/kg/day) reduced body weight gain induced by HFD (8.6±1.8 vs. −4.2±2.0 (CA0.9+FA50) and −3,7±1,9 (CA1.8+FA50); p<0.0001), fasting hyperglycemia (223±40 vs. 181±44 (CA0.9+FA50) and 168±41 (CA1.8+FA50); p<0.05), hypercholesterolemia (134±20 vs. 110±7.3 (CA0.9+FA50) and 104±2.7 (CA1.8+FA50); p<0.05), hypertriglyceridemia (45±4 vs. 40±9 (CA0.9+FA50) and 21.1±9 (CA1.8+FA50); p<0.001). Liver histology showed that CA and FA combined improved the hepatic steatosis induced by high fat diet. On the other hand, significant inflammation was observed in WAT of the mice treated with both acids, with neutrophil and macrophage infiltration. In conclusion, the combination of CA and FA improves the abnormalities observed in metabolic syndrome induced by high fat diet but leads to inflammatory process in white adipose tissue.

Supported: CAPES.
Dietary Cod Protein Improves the IGF1‐Akt/PKB Signaling Pathway in Rat Skeletal Muscle during Recovery from Injury
Authors: Dort Junio , Leblanc Nadine , Côté Claude H , Jacques Hélène
Published: June 2021

This study was designed to assess the mechanisms by which dietary cod protein may facilitate skeletal muscle recovery from injury through analyses of the IGF1‐Akt/PKB signaling pathway. Male Wistar rats were fed isoenergetic diets containing either casein (C), cod protein (CP), or casein supplemented with a mixture of arginine, glycine, taurine and lysine (C+), matching their respective levels as in CP. Muscle injury was induced by bupivacaine injection and downstream IGF1‐Akt/PKB effectors were measured post‐injury. At day 2 post‐injury, C+ upregulated (p=0.0008) whereas CP tended to upregulate phospho‐Akt Thr308 compared with C (p=0.075), suggesting that the myogenic effect of CP is due in part to its high levels of arginine, glycine, taurine and lysine. At day 2 post‐injury, CP reduced the expression of MuRF‐1 by 38% compared with C (p=0.03), supporting a decrease in the ubiquitylation of muscle proteins and their degradation. At day 5 post‐injury, CP tended to increase the muscle IGF‐1 level compared with C (p=0.098); phospho‐Akt Ser473, a phosphorylation required for maximum activation of Akt in addition to phosphorylation at Thr308, was also increased (p<0.0001). These findings suggest that at early time‐points of recovery, CP can reduce muscle protein degradation through modulation of the ubiquitination process and upregulate protein synthesis through enhanced Akt activation.

Supported by NSERC
The effects of arginine and nitric oxide synthase inhibitors on intestinal damage in guanylyl cyclase‐G gene knockout mice with intestinal ischemia and reperfusion
Authors: Lo Hui‐Chen , Lee Chien‐Hsing , Chin Shao‐Hua
Published: June 2021

Previous study demonstrated that membrane‐bound guanylyl cyclase (mGC)‐G gene knockout (KO) mice with intestinal ischemia and reperfusion (II/R) had significantly alleviated jejunal damage and apoptosis compared to wild type (WT) mice. It is known that apoptosis is controlled by guanylyl cyclase‐generated cGMP and arginine‐derived nitric oxide (NO). Therefore, we investigated the roles of NO and mGC‐G gene in II/R‐induced intestinal damage. mGC‐G KO mice suffered with 45 min intestinal ischemia were intra‐peritoneally administered with arginine (300 mg/kg) or inhibitors of nitric oxide synthase (NOS), i.e., aminoguanidine (100 mg/kg) or L‐NAME (10 mg/kg), and were executed after 3 or 24 hours of reperfusion. KO mice had significantly decreased jejunal mass and villus height and increased plasma IL‐6 and mucosal nitrate/nitrite (NOx) and cGMP compared to WT mice. However, II/R‐induced jejuanl atrophy and systemic and local inflammatory responses were attenuated in KO mice as shown in the less increases in plasma NOx and mucosal TNF‐alpha (p<0.05). At 3 hr reperfusion, arginine significantly increased plasma IL‐6 and mucosa TNF‐alpha; AMG increased mucosal TNF‐alpha; and L‐NAME increased plasma TNF‐alpha and mucosal cGMP in KO mice. At 24 hr reperfusion, arginine, AMG, and L‐NAME decreased mucosal TNF‐alpha, plasma IL‐6, and muscularis width, respectively. In addition, AMG and L‐NAME further decreased plasma NOx at 3 and 24 hr reperfusion. However, mGC‐G KO‐and II/R‐induced jejunal atrophy was not significantly altered by arginine or NOS inhibitors. These results suggest that mGC‐G may play certain roles in affecting the effects of NO on jejunal histology and inflammatory response (NSC 96–2320‐B‐309–003‐MY2).
Flood‐primed continuous tracer infusion complies with the precursor‐labeling assumptions for measuring the muscle fractional synthesis rate using a single biopsy approach
Authors: Reitelseder Søren , Dideriksen Kasper , Nielsen Rie Harboe , Kjaer Michael , Holm Lars
Published: June 2021

In certain clinical settings it is allowed only to obtain one single tissue biopsy. Therefore, we developed a tracer administration approach that complies with assumptions for determining the fractional synthesis rate (FSR) by obtaining only one biopsy at the end of the infusion.12 males were recruited: 7 were given a flood‐primed (8.96 mmol, enriched 12.5% tracer:tracee ratio (TTR)) continuous (8 μmol/kg lean body mass (LBM)/h) infusion of ring‐13C6‐phe and muscle biopsies were obtained after 10, 30, 60, 120, and 360 min. 5 men were given a primed (9 μmol/kg LBM) continuous (9 μmol/kg LBM/h) infusion of 1,2‐13C2‐leu. After 120 and 300 min muscle biopsies were obtained to determine FSR. Hereafter, a flood‐primed continuous infusion of phe was given to mimic the protocol and a third muscle biopsy was obtained at 480 min to determine FSR for 3 h following the flood‐prime. Muscle free phe enrichment were up at steady state level from 10‐min after the flood‐prime: 6.98±0.70%, 6.87±0.71%, 6.42±0.58%, 6.58±0.70%, and 6.23±0.81% TTR at 10, 30, 60, 120, and 360 min, respectively. Basal myofibrillar FSR was 0.08±0.01 %/h and was unaffected by the flood‐primed continuous infusion of phe (0.07±0.01 %/h, p>;0.05). The flood‐prime immediate introduced the phe tracer into the intramuscular pools and the continuous infusion kept the enrichment steady without affecting the muscle FSR. Funding: Danish Council of Independent Research.
Iron Metabolism in a Rodent Model of Menkes Disease (Brindled Mice)
Authors: Gulec Sukru , Collins James F.
Published: June 2021

The Menkes copper‐transporting ATPase (Atp7a) is induced in the rodent duodenum during iron deficiency. To understand its role in intestinal iron metabolism, we thus initiated an investigation in Atp7a mutant mice (Brindled). As mutant mice die perinatally of severe copper deficiency, IP copper injections were administered at 7 and 9 days‐of‐age. Mice developed normally thereafter and were similar in size and activity level to wt littermates. Subsequently, mice were deprived of dietary iron for 3 wks and iron and copper homeostasis was studied. Several important observations arose from these analyses, including: 1) mutant mice on control diet were copper deficient and anemic with no changes in iron status; 2) mutant mice on iron‐deficient diet were iron and copper deficient and more significantly anemic; 3) the transcriptional response to iron deficiency in enterocytes was seen only in mutant mice on low Fe diet, suggesting that the HIF2α‐mediated induction of iron and copper homeostasis‐related genes is triggered by iron levels and not hypoxia per se; and 4) Brindled mice had increased hepatic copper levels during iron deprivation, leading to a concomitant increase in serum ferroxidase activity, while no such changes were noted in control or low Fe wt mice. This study has thus revealed novel aspects of the iron‐copper connection and is the first to investigate iron homeostasis in a model of genetic copper deficiency.

Grant Funding Source: NIH
Analysis of plant G flower essential oil composition
Authors: Kim Gun‐Hee , Chang Kyung Mi
Published: June 2021

The volatile aroma constituents of plant G essential oil (PEO) were separated by the steam distillation extraction (SDE) method using a Clevenger‐type apparatus, and analyzed by gas chromatography‐mass spectrometry (GC/MS). Fifty‐six volatile chemical components, which make up 73.38% of the total aroma composition, were tentatively characterized. This PEO contained 27 hydrocarbons (10.05%), 12 alcohols (36.55%), 8 ketones (19.05%), 4 esters (2.59%), 2 aldehydes (1.79%), and 3 miscellaneous components (3.35%). Borneol (12.96%) and camphor (10.54%) were the predominant volatile components in plant G, an aromatic medicinal herbaceous plant.
Regulation of TCF/LEF promoter activity by soy phytoestrogens in 3T3‐L1 fibroblasts
Authors: Davis Jeremy E , Hastings Darcie , Banz William
Published: June 2021

The canonical Wnt pathway inhibits adipogenesis through activation of the transcription factor, TCF/LEF1. Soy phytoestogens, such as genistein (GEN) and daidzein (DAID), are paradoxically shown to both induce and repress adipogenesis. To determine if the concentration of soy phytoestrogen effects adipogenic regulation, 3T3‐L1 fibroblasts were transfected with either a estrogen receptor (ER) or TCF/LEF lentiviral reporter construct. Transfected cells were then treated with GEN or DAID at 100 pM, nM, or uM concentration. Our data showed that both ER and TCF/LEF promoter activity was 2–3 fold greater in cells treated with 100 uM of GEN or DAID (vs. vehicle). In contrast, fibroblasts treated with the pM or nM concentrations exhibited limited promoter activity. Collectively, these data indicate that the anti‐adipogenic properties of soy phytoestrogen may be attributed to high pharmacological doses.
Roles of dietary vitamin A in the regulation of hepatic gene expression in response to fasting‐refeeding cycle
Authors: Li Yang , Chen Guoxun
Published: June 2021

We have shown that vitamin A (VA) status affects the plasma glucose and lipid levels, and hepatic gene expression in rats. Therefore, we hypothesize that dietary VA intake and its hepatic metabolism regulate the expression of genes involved in glucose and lipid metabolism in response to fasting‐refeeding cycle. Male Zucker lean rats were fed with a VA deficient (VAD) or a VA sufficient (VAS) diet for 8 weeks. The VAD or VAS rats were fed ad libitum, fasted for 48 hours (H) or refed a VAD or VAS diet for 6H. The plasma parameters were measured using commercial kits. The hepatic protein and mRNA levels were respectively examined by Western blot and real‐time PCR. The responses of plasma insulin and leptin levels to the feeding cycle were altered in VAD rats compared to VAS ones. The hepatic mRNA levels of Gck, Srebp‐1c, and Fas were reduced after 48H fasting regardless of VA status. The 6H refeeding of a VAS diet induced their mRNA levels in VAD and VAS rats. However, the 6H refeeding of a VAD diet only induced their expression levels in VAS, but not VAD rats. We conclude that dietary VA and its hepatic metabolism contribute to the regulation of plasma hormone levels and hepatic gene expression in rats.

Grant Funding Source: American Heart Association
Vitamin A equivalence of spirulina β‐carotene in US men
Authors: Tang Guangwen , Muzhingi Tawanda , Grusak Michael
Published: June 2021

To evaluate the vitamin A value of spirulina β‐carotene and to assess dose‐related responses, spirulina was grown in a 23 atom% 2H2O cultural solution, harvested, made as capsules, and fed to volunteers. Spirulina β‐carotene showed the highest abundance of enrichment as [2H10] trans β‐carotene. Eight healthy US male subjects (51±6 y) with a mean serum retinol concentration of 1.7 ±0.3 μmole/L and a BMI of 23 ± 3 kg/m2 consumed 5.8 μmole (2.0 mg) of [13C10] retinyl acetate in oil as a reference dose with a standard liquid breakfast containing 13 g of fat (day 1). At day 8, each subject was randomly assigned to consume 4 g or 2 g of spirulina containing 5.1 (2.8 mg) or 2.6 (1.4 mg) μmoles of trans β‐carotene with the same standard breakfast as day 1. Serum samples were collected at various time points over 50 days. Concentrations and enrichments of retinol and β‐carotene in serum samples were determined by using HPLC and GC/MS0.

A comparison of the mean area‐under‐the‐curve (AUC) for the total labeled serum retinol response after the 4 g (n=4) or 2 g (n=4) of spirulina, with the AUC of the reference dose of 2.0 mg [13C10]retinyl acetate, did not show differences related to the amount of dose taken. The mean vitamin A equivalence of spirulina β‐carotene to retinol was 2.7 and 2.8 to 1 by weight, for the 4 g or 2 g spirulina doses, respectively. (Funded by the Nestle Foundation).
Fish oil differentially regulates B cell and dendritic cell activation in response to a T‐independent antigen
Authors: Teague Heather , Rockett Benjamin Drew , Harris Mitchel , Shaikh Saame Raza
Published: June 2021

Fish oil (FO) has clinical utility for regulating immunity. The functional consequences of FO on B cells and dendritic cells (DC) are poorly studied, especially at the animal level. Here we tested the effects of FO, modeling human pharmacological intake, on murine ex vivo B cell and DC activation in response to the T‐independent antigen lipopolysaccharide (LPS). LPS stimulation of B cells increased CD69 surface expression and enhanced secretion of IL‐6, TNFα and IFNγ with FO. In comparison, LPS stimulation of DCs resulted in reduced CD80 surface expression and TNFα secretion with FO, which was associated with decreased phagocytosis of E. coli bioparticles. To reconcile differences between cell types, we measured the formation of plasma membrane lipid microdomains. Although FO promoted an increase in EPA and DHA levels, we discovered differential effects on the ability of B cells and DCs to cluster lipid microdomains. Overall, this study shows FO differentially regulates the activation of B cells and DCs in response to LPS, which may be due to differences in underlying membrane organization. Our data highlight the possibility that FO can be used to exert both immune enhancing and immunosuppressive effects, depending on the cellular target.

Grant Funding Source: NIH R15AT006122 (to S.R.S.)
Effect of a new compounds of substituted 5R1, 6H2–1,3,4‐thiadiazine‐2‐amines on the histological pictures in ligation models of experimental acute myocardial infarction and experimental acute pancreatitis in rats.
Authors: Sarapultsev Alexey Petrovich , Chupakhin Oleg Nickolaevich , Sarapultsev Petr Alekseevich , Rantsev Maxim Anatolevich , Medvedeva Svetlana Ur'evna , Danilova Irina Georgievna
Published: June 2021

Models of experimental acute myocardial infarction (EAMI) and experimental acute pancreatitis (EAP) have been developed to solve this problem. The animals got injection of the compound from the group of substituted 5R1, 6H2–1,3,4‐thiadiazine‐2‐amines, dozed 40 mg/kg. Histological study has been carried out at the 1st, 5st and 7th days. All animal experimental procedures were performed in accordance with the APS's Guiding Principles and in accordance with the Laboratory Practice Regulations of RF.

Already during the 1st day of the EAMI no leucopenia was observed and the signs of proliferative inflammation were detected. Later, at the 5th day, the necrosis area got surrounded by demarcation bank, and further on (7th day) had been entirely replaced by granulation tissue. Withal, the application of compound in EAP results in occurrence of proliferative inflammation, prevents lympho‐ and monocytopenia development and cuts down the lethality rate two‐fold.

Conduction research enabled to state repeatability of the mechanism of the compounds impact on ischemia‐induced inflammation process, both in case of EAMI and EAP. The compounds modulated the transition of destructive inflammation into proliferative one, thereby preventing spreading of the inflammation process to tissues surrounding the necrotic zone and promoting reparative processes as well.

Research Grant 12‐Ì‐34–2064 of Ural Branch of RAS.
Characterization of the tissue response to functional multielectrode arrays in the feline spinal nerve
Authors: Kolarcik Christi L , Castro Carlos A , Cui Xinyan Tracy , Weber Douglas J
Published: June 2021

Peripheral nervous system sites are target implantation regions for neural prosthetics that restore function or provide therapy. Despite advances in electromechanical prostheses, current peripheral‐interface technology cannot convey the users’ intent to the prosthesis. Anatomical information and an understanding of the tissue response are needed. In this study, functional microelectrode arrays (FMAs) were implanted into feline dorsal (sensory) or ventral (motor) roots. At predetermined time points or when <10% of electrodes had recordable signals, animals were perfused and the implanted tissue formalin‐fixed and paraffin‐embedded. Hematoxylin and eosin and Nissl/Luxol fast blue staining along with antibody‐based stains [NF200 (axons), S100 (Schwann cells), vimentin (fibroblasts, endothelial cells, astrocytes), NeuN (neuronal nuclei), caspase‐3 (apoptotic cell death), MAC387 (monocytes/macrophages) and occludin (tight junctions/blood‐nerve barrier integrity)] were used. Axonal density, scar formation, neural degeneration (ventral root and spinal cord ventral horn) and inflammation were quantified and compared in implanted and non‐implanted roots. Histological data was correlated with functional metrics to provide insight toward the overall goal of a long‐term, reliable interface with prosthetic limbs. Funding provided by the DARPA RPI grant.
Secondhand smoke exposure augments and accelerates the progression of balloon angioplasty‐induced neointimal formation morphological changes in rat
Authors: LI Pei Chuan , Pan Chun‐Hsu , Wu Chieh‐Hsi
Published: June 2021

Smoking is one of the most important risk factor for cardiovascular diseases. The purpose of the present study is to evaluate the potential detrimental effects of second‐hand smoke on the progression of neointimal formation in a rat model of balloon angioplasty. SD rats were divided randomly into three groups. Group 1, sham control; Group 2, balloon angioplasty alone; Group 3, passive smoke exposure (inhalation) for 3 weeks before and for 2~14 days after balloon angioplasty. The rat carotid arteries of all groups were harvested at 2, 7 and 14 day after balloon injury. The results of histopathological assay showed that passive smoke augments markedly neointimal formation at 14 day post‐angioplasty as compared with balloon angioplasty alone group (ratio of neointima‐to‐media area, 0.44+0.23 vs. 0.32+0.05). Moreover, treatment of passive smoke has been found to accelerate obviously the progression of neointimal formation as compared to that of balloon angioplasty alone group. Besides, our present study suggested that there was a strongly positive correlation between serum level of nicotine, one of typical compound within tobacco smoke, and the severity of balloon injury‐induced neointimal formation. In summary, our study demonstrated that passive smoke is a detrimental factor to augment and accelerate the progression of balloon injury‐induced neointimal formation.
Region‐, dose‐, and state‐dependency of isoflurane‐induced changes in blood‐brain barrier permeability
Authors: Kuo Eva Yuhua , Wei Vivi Chiali , Yen Lola Fenghuei , Liao Kate Hsiurong , Lai Ted Weita
Published: June 2021

Isoflurane opens the blood‐brain barrier (BBB) to inert tracers like Evans blue dye (EBD). However, the dose‐ and the region‐specificity are not known. To avoid confounding effect of anesthesia per se, the conventional protocol has been to subject the laboratory animal to another pre‐existing anesthesia. Here, 1% isoflurane increased EBD extravasation into the prefrontal cortex (PFC), the motor cortex (MC), the striatum (ST), the cerebellum (CE), and the liver of urethane‐anesthetized rats, but had no effect on blood EBD concentration. In marked contrast, 2% isoflurane had no effect on EBD extravasation to central and peripheral tissues, but increased blood EBD concentration. Because pre‐existing anesthesia can compromise the cardiovascular system, we repeated the study in otherwise‐conscious rats. Surprisingly, not only did isoflurane had no effect on EBD extravasation into the MC, the ST, and the CE, it decreased EBD extravasation into the PFC and the liver. The effect of isoflurane in these otherwise‐conscious rats could not be due to alleviation of stress, as there was little noticeable difference between EBD extravasation in rats placed in home cage, rats placed in the anesthetic cage, and rats placed in the anesthetic cage filled with the gaseous vehicle. In summary, isoflurane‐induced changes in BBB was region‐specific, dose‐dependent, and reversed by pre‐existing anesthesia. (Supported by Taiwan NSC)
Synaptic and extrasynaptic GABAA receptors in benzodiazepine withdrawal: effects of pregnanolone and gaboxadol in benzodiazepine‐dependent monkeys
Authors: Zanettini Claudio , France Charles P , Gerak Lisa R
Published: June 2021

Benzodiazepines and neuroactive steroids positively modulate GABAA receptors through different binding sites. These drugs share many behavioral effects including the ability to reverse benzodiazepine withdrawal; however during chronic benzodiazepine treatment, tolerance develops to benzodiazepines and not to neuroactive steroids. This difference might be related to actions of neuroactive steroids at benzodiazepine‐insensitive extrasynaptic GABAA receptors. This study examined the ability of gaboxadol, an orthosteric agonist at extrasynaptic GABAA receptors, to attenuate flumazenil‐lever responding, a sign of benzodiazepine withdrawal. Four rhesus monkeys received 5.6 mg/kg/day diazepam and discriminated 0.1 mg/kg flumazenil. Gaboxadol (1.78–5.6 mg/kg) did not produce flumazenil‐lever responding or shift flumazenil dose‐response curves whereas pregnanolone, a neuroactive steroid, shifted the curves rightward. Because pregnanolone attenuated withdrawal whereas agonist action only at extrasynaptic GABAA receptors (gaboxadol) did not, it is unlikely that those receptors account for the difference between benzodiazepines and neuroactive steroids in benzodiazepine‐treated monkeys. This difference is more likely due to changes in synaptic GABAA receptors, such as subunit composition or coupling of binding sites. Supported by USPHS grants R01DA09157 and K05DA017918 (CPF).
Effects of chronic treatment with the D3 receptor‐selective compound PG619 on cocaine (COC) self‐administration and FDG brain activity in rhesus monkeys
Authors: Nader Susan , Porrino Linda , Miller Mack , Calhoun Tonya , Martelle Susan , Banala Ashwini , Newman Amy , Nader Michael
Published: June 2021

The selective brain distribution of dopamine D3 receptors (D3R) continues to make it a target for drug discovery. Studies suggest that the ability of D3R antagonists to decrease drug self‐administration (SA) depend on the reinforcement schedule ‐ schedules of high response demands and conditioned stimuli appear sensitive to D3R drugs. In the present study, male rhesus monkeys SA COC under a second‐order (SO) schedule. An advantage of SO schedules is that responding can be maintained for extended periods prior to drug infusion, measuring cue‐related behavior. We examined the effects of 30 day treatment (Tx) with the D3R partial agonist PG619 on COC‐maintained responding using a SO schedule, as well as brain glucose utilization using [18F]FDG and PET. Effects of PG619 on rates of responding (RR) were dependent on baseline RR prior to the 30 day drug Tx. Monkeys with low RR showed either no effect or increases while monkeys with high RR showed decreases during Tx. [18F]FDG PET imaging showed that activation of the amygdala, ventral striatum and precuneus during baseline COC cue presentations may predict behavioral response to PG619. Tx with PG619 blunted the cue‐induced elevations in glucose utilization. These studies support the continued examination of D3R compounds for addiction Tx and suggest the need for individualized treatment strategies. DA006634, DA012460, NIDA‐IRP
EXAMINING THE RELATIONSHIP BETWEEN COCAINE AND SLEEP IN A MONKEY MODEL OF DRUG ABUSE
Authors: Brutcher Robert , Nader Michael
Published: June 2021

Clinical literature suggests that sleep deprivation can induce patients to relapse to drug taking. This animal study examined the effects of cocaine (COC) self‐administration on several sleep measures and how sleep disruption affects COC preference. Subjects were 8 adult male rhesus monkeys (Macaca mulatta), each wearing Actical activity monitors on their collar and surgically implanted with an indwelling intravenous catheter for COC self‐administration. Baseline activity was monitored for 1 week in the absence of any behavioral testing. Each morning (0730), monkeys were permitted to respond under choice procedure in which food reinforcement (three, 1‐g banana‐flavored pellets) was contingent upon completing the response requirement on one lever, while COC (0.003–0.1 mg/kg/injection) or saline presentation was contingent on responding on the other lever. After baseline cocaine‐choice curves were determined, the monkeys were sleep disrupted. Our results indicate that COC self‐administration causes significant changes on several sleep measures, providing evidence for possible treatment strategies. We also found that sleep disruption caused significant changes on sleep measures but did not alter the reinforcing strength of COC the next morning. Future studies will examine a medication that may enhance sleep (quetiapine) and may also decrease COC self‐administration. DA025120
Direct CRFR1 antagonism within the VTA prevents the induction and expression of neural cross‐sensitization to cocaine caused by social defeat stress
Authors: Holly Elizabeth N , Boyson Christopher O , Shimamoto Akiko , DeBold Joseph F , Miczek Klaus A
Published: June 2021

Social defeat stress results in cross‐sensitization to cocaine, both behaviorally, manifested by augmented locomotor activity and neurally, manifested by augmented dopamine (DA) in the nucleus accumbens (NAc) (Holly et al, 2012). We investigated the role of CRF within the ventral tegmental area (VTA) in the induction and expression of neural cross‐sensitization to cocaine. Male Long‐Evans rats were defeated by a larger aggressive resident male in four episodes, separated by 72 hours (induction phase). Ten days later (expression phase) in vivo microdialysis was used to assess DA in the NAc after an acute cocaine challenge (10 mg/kg, ip). In Experiment 1, rats were implanted with bilateral microinjection cannulae aimed at the VTA, as well as a microdialysis cannula aimed at the NAc one week prior to social defeat. The CRFR1 antagonist CP376395 (CP, 500ng/0.25μL/side) was microinjected 20 minutes prior to each defeat. In Experiment 2, rats were implanted with ipsilateral cannulae in the VTA for microinjection and in the NAc for microdialysis after the last defeat. CP was microinjected 20 minutes prior to the cocaine injection during microdialysis. CRFR1 antagonism in the VTA prevented both the induction and expression of neural cross‐sensitization as a result of social defeat stress, indicating a modulatory role of CRF on DAergic function and neuroadaptations in the VTA as a result of intermittent social stress.
Glycogen Synthase Kinase 3β; Inhibiton Improves Diabetes‐induced Cognitive Deficits in Type‐2 Diabetic Rats
Authors: Datusalia Ashok Kumar , Sharma Shyam Sunder
Published: June 2021

Challenges to neural homeostasis in diabetes may produce different responses and damage in CNS and alteration in many measures of cognition in diabetes. This study was carried out to investigate the effect of GSk3β inhibitor (SB216763; SB) on diabetes‐induced cognitive deficit and its mechanism of neuroprotection in type 2 diabetic (T2D) rats. High fat diet with STZ was used to develop T2D in rats. Rats were tested for place navigation, fear & spatial memory using behavioral paradigms on 9, 12, 15 & 18 week after induction of T2D. Brain regions were tested for biochemical and neurotransmitter level. Treatment with SB was given from 15 week to 18 week at 3 doses 0.3, 0.6 & 1.2 mg/kg. T2D rats showed cognitive deficits after 15 week duration of diabetes and continue on 18th week compared to control. Further, elevated brain GABA level was observed after 15th week in T2D rats. Reduced glutamate level was also observed in T2D rats. Treatment with SB (0.6 mg/kg) for 3 weeks improved the diabetes‐induced cognitive deficits in passive avoidance, Y‐maze and MWM test. Improvement in memory also supported with decrease in GABA level and increased glutamate level. Administration of SB in diabetic rats also showed reduction in neuroinflammation in hippocampus and cortex. Improvement in cognitive deficits in diabetic rats by GSK3β inhibitor was attributed to reversal of neurotransmitters changes and reduction of neuroinflammation.
Post‐ischemic thyroid hormone treatment in a rat model of acute stroke
Authors: Campolo Michela , Genovese Tiziana , Impellizzeri Daniela , Ahmad Akbar , Cornelius Carolin , Cuzzocrea Salvatore , Esposito Emanuela
Published: June 2021

This study was performed to investigate the effects of prohormone thyroxine (T4) and the underlying mechanisms in the post‐ischemic rat brain after transient focal cerebral ischemia‐induced brain injury. Ischemic injury was induced for 2 h by middle cerebral artery occlusion (MCAO) followed by 24‐h reperfusion. T4 was administered by intra‐peritoneally injection twice, at 1 after the onset of ischemia and 6 h after reperfusion. The mechanism of neuroprotective effect of T4 was investigated with a focus on inflammatory cells, neurotrophins, and transcriptional factors. The results obtained showed that T4 significantly reduced cerebral infarction, which were accompanied by decreased expression of proapotptic Bax and increased antiapoaptotic Bcl‐2 protein. T4, also suppressed the activation of astrocytes and microglia, increased the expression of neurotrophic factors (BDNF, GDNF), and suppressed up‐regulation of inflammatory‐related prooxidative enzymes (iNOS and COX‐2) in ischemic brain. Moreover, T4 down‐regulated the phosphorylation of p38 and prevented injury‐induced increase of PKCδ. These results revealed that T4 has a promising therapeutic effect in ischemic stroke treatment protecting the brain from I/R injury, probably by its anti‐apoptotic, and anti‐inflammatory mechanism.
Characterization of Post‐Translational Modifications Related to Cardiovascular Disease
Authors: McComb Mark E , Whelan Stephen A , Spencer Jean L , Heckendorf Christian F , Bachschmid Markus M , Siwik Deborah A , Colucci Wilson , Cohen Richard A , Costello Catherine E
Published: June 2021

Unfavorable metabolic conditions (metabolic disorders) associated with obesity and diabetes are major causes for cardiovascular disease. Early detection of the adverse effects of metabolic disease remain elusive. We believe that nonspecific changes which occur in plasma proteins, indicators of inflammation and oxidants, may act as evidence of systemic metabolic disease. Here we elucidate potential biomarkers of CVD including both protein changes and changes in post‐translational modifications (PTMs). Label‐free mass spectrometry‐based proteomics was used to interrogate changes in differential protein and PTM expression in plasma samples from mouse and human models. Label‐free LCMS/MS typically yielded >;1,000 features ( p<0.05, >;2fold). A number of cardiovascular disease related proteins were observed. Up regulated proteins were: haptoglobin, a known biomarker related to inflammation, low mannose binding protein associated with inflammation and CVD in type 2 diabetes, superoxide dismutase and extracellular matrix protein both implicated in type 2 diabetes. We also observed different PTMs associated with oxidative stress including lipid peroxidation products such as hydroxynonenal and multiple forms of oxidation such as cysteine sulfonic acid. Development of a metabolic disorder/CVD‐specific protein panel will afford the first step in biomarker panel development such that disease diagnosis and progression may be performed directly at the molecular level.

This project was funded by NIH‐NCRR grants P41 RR010888/ GM104603, S10 RR015942, S10 RR020946, S10 RR025082 and NIH‐NHLBI contract N01 HV00239.
TARGETING THE REGULATOR OF G PROTEIN SIGNALING 17 (RGS17) FOR LUNG AND PROSTATE CANCER TREATMENTS
Authors: Mackie Duncan Ian , Roman David L
Published: June 2021

G‐Protein Coupled Receptors (GPCR) are one of the most important target in disease treatment with over 50% of all drugs on the market targeting these receptors. Regulator of G‐Protein Signaling (RGS) proteins are known to regulate the complex signaling pathways activated by GPCRs. Recent studies have implicated RGS‐proteins in the development and progression of pathologies, including some cancers. RGS17, the most‐recently identified family member of the RZ family of RGS proteins has been implicated in the growth, proliferation, metastasis and migration of prostate cancer cells as well as small‐cell and non‐small cell lung cancers. RGS17 is up‐regulated in lung and prostate tumors as much as 13 fold. RGS17 knockdown inhibits colony formation and decreases tumorigenesis in nude mice. Our studies implemented a high‐throughput screen campaign to determine the first Gαo: RGS17 protein: protein interaction (ppi) inhibitors. Immediate goals include determining the effectiveness of these compounds on inhibiting the GTPase acceleration activity of RGS17 in a steady‐state GTPase assay. The goal of this study is to establish lead compounds and develop a pharmacophore model for future avenues in cancer treatment. Research is supported by American Foundation for Pharmaceutical Education (AFPE), University of Iowa′s Center for Biocatalysis and Bioprocessing and The National Institutes of Health (R01CA160470).
Acrolein enhances anti‐HIV‐ HAART medication‐ induced hepatotoxicity
Authors: Ghare Smita , Donde Hridgandh , Joshi‐Barve Swati , McClain Craig , Barve Shirish
Published: June 2021

Highly active antiretroviral therapy (HAART) has significantly increased the life expectancy of patients with HIV infection. However, HAART‐ associated hepatotoxicity is becoming a major clinical problem and can be potentially compounded by exposure to diet/environment ‐ derived reactive aldehydes, particularly acrolein. The present study investigates the combined effects of acrolein along with HAART drug, azidothymidine (AZT) in a human hepatocyte cell line (HepG2). Our results showed that exposure to either acrolein or AZT had minimal to no effect on hepatocyte survival. However acrolein in combination with AZT led to a significant increase in hepatocyte apoptotic death. The increase in apoptosis was accompanied by transcriptionally permissive H3 histone acetylation at FasL‐promoter and consequent FasL gene expression. Interestingly, concomitant with the induction of FasL and apoptotic death there was a significant attenuation of autophagy as indicated by a decrease in Beclin‐1 and an increase in LC3II/LC3I ratio. These data strongly suggest that the mechanisms underpinning acrolein‐AZT mediated hepatotoxicity involves the suppression of autophagy and induction of FasL mediated apoptosis. Overall, the data suggest that presence of diet/environment derived reactive aldehydes can play a pathogenic role in HIV patients by enhancing HAART‐hepatotoxicity. This work was supported by NIH grants.
Cadherin peptide‐induced enhancement of blood brain barrier (BBB) permeability
Authors: On Ngoc Hong , Laksitorini Marlyn , Kiptoo Paul K , Siahaan Teruna J , Miller Donald W
Published: June 2021

Objective

To examine the effects of two cadherin peptides, a short linear peptide containing His‐Ala‐Val (HAV) sequence and a cyclic peptide containing the Ala‐Asp‐Thr (cADT) sequence, on BBB integrity.

Methods

The effects of the HAV and cADT peptides on BBB permeability were examined in Balb/c mice. Blood‐brain barrier permeability was assessed with gadolinium contrast agent (Gad; a small hydrophilic permeability marker), IRdye 800cw PEG (a large hydrophilic permeability marker), and Rhodamine 800 (a P‐gp substrate) under control conditions and following exposure to HAV (1.0–32 mM/kg) or cADT peptides (1mM/kg) using magnetic resonance imaging (MRI) and near infrared fluorescence imaging.

Results

Mice treated with HAV displayed a dose‐dependent increase in BBB permeability as assessed with Gad enhanced MRI with doses of 1.0 mM/kg having no effect and maximal increases (18‐fold) observed with 32 mM/kg. Increases in BBB permeability were also observed with IRdye 800cw PEG (3‐fold) and Rhodamine 800 (60% increase). The increase in permeability was transient (return to normal within 1 hr). Increases in BBB permeability were also observed with cADT, albeit at lower dose (1.0 mM/kg).

Conclusions

Cadherin peptides produced a rapid and reversible increase in BBB permeability and the combination of these peptides with therapeutic agents can enhance drug delivery to the brain.

Funding:

Supported by NSERC and NIH.
Application of a genomics‐guided warfarin dosing nomogram for hospitalized patients
Authors: Gryn Steven E. , Facca Nadia , Kim Richard B.
Published: June 2021

Adverse drug reactions (ADRs) due to warfarin are an important cause of emergency department visits, and also occur frequently among hospitalized patients. We now know common genetic variations in CYP2C9‐mediated metabolism of warfarin, as well as its target VKORC1, account for much of the dosing variation. Accordingly, we hypothesized that application of a pharmacogenomics‐guided warfarin dosing algorithm that our group developed, known as WRAPID, may be clinically useful in guiding warfarin initiation and dosing for hospitalized patients. We initiated a personalized medicine‐based inpatient consult service with a goal of providing WRAPID‐based warfarin dosing recommendations. We now have data on a series of patients who were initiated on warfarin as inpatients who subsequently developed a supratherapeutic international normalized ratio (INR). When assessed by our service, most were shown to possess genotypes associated with marked sensitivity to warfarin, including one patient with the rare CYP2C9 *3/*3 genotype, as well as several with CYP2C9 *2/*2 and/or VKORC1‐1639 A/A genotype. Based on our WRAPID‐guided dosing, warfarin dose was accurately predicted to be as low as 0.5 mg. Accordingly, we believe genomics‐guided warfarin dosing is useful for identifying hospitalized patients at risk for ADRs, thereby reducing length of stay and preventing readmissions. Funding: AMOSO Drug Innovation Fund.