bluuebunny/crossref_metadata_2025_split huggingface dataset:
Anion and fluid secretory response of the murine jejunum to the heat‐stable Eschericia coli enterotoxin (STa) analogue linaclotide: Involvement of NHE3, Slc26a6, CFTR, proteinkinase GII (cGKII) and NHERF1‐3.
Authors: Seidler Ursula Elisabeth , Liu Yongjian , Qian Jiajie , Tan Qinghai , Riederer Brigitte , Hofmann Franz , de Jonge Hugo 
Published: June 2021

The different PDZ‐adaptors of the NHERF family differentially associate with intestinal electrolyte transporters and modulate their activity in an agonist‐specific fashion. STa induces secretory diarrhea via binding to the guanylate cyclase C, eliciting cGMP production, resulting both in a stimulation of CFTR‐mediated anion secretion and an inhibition of NHE3‐mediated fluid absorption, and these effects are modulated by different NHERF adaptor proteins in cell lines.AimWe studied the STa‐mediated short circuit current response in isolated jejunal mucosa as well as the STa action on in vivo fluid transport in anethstized mice, deficiet for NHE3, Slc26a6, CFTR and NHERF1‐3, to study how well in vitro and in vivo modulation of fluid transport by the above named transporters and NHERF proteins may correlate.Results10−7M of the STa analogue linacolide was applied to the luminal bath of chambered isolated jejunal mucosa of CFTR‐, NHE3‐, Slc26a6‐, cGKII, and NHERF1–3 deficient and WT mice, and short circuit current (Isc) response was measured. The same concentration of linaclotide was added to the jejunal perfusate during single pass perfusion of a jejunal segment in isoflurane‐anesthetized, acid/base‐ and blood pressure controlled KO and WT mice, and net fluid balance before and after linaclotide application assessed gravimetrically. The peak Isc response to linaclotide in chambered jejunal mucosa was absent in CFTR KO, reduced by 88% in cGKII KO, by 40% in NHERF1 KO, and not significantly in NHERF2, NHERF (PDZK1) and SLC26a6 KO isolated jejunal mucosa. In contrast, the magnitude of change from jejunal fluid absorption to secretion in vivo was reduced by 51% in cGKII KO, 49% in CFTR KO, 45% in NHERF1 KO, by 34% in NHERF2 KO, and not significantly in NHERF3 KO and Slc26a6 KO anesthetized mice. NHE3 deletion (48%), pharmacological NHE3 inhibition (50%) or Slc26a6 deletion (44%) also reduced the STa‐elicited change in fluid movement.ConclusionsThe results demonstrate that Isc measurements in isolated small intestinal mucosa are excellent models to selectively study agonist‐mediated CFTR activation, but do not accurately predict in vivo “secretory responses”, i.e. a relative decrease in the basal fluid absorptive flux. Activating the GCC receptor elicited a decrease in fluid absorption in CFTR KO mice, making this substance an interesting drug to improve gut fluidity in CF patients. Apart from direct interference with guanylate cyclase C activation, potential pharmacological strategies to prevent STa‐mediated fluid loss may be targeting both NHERF1 and NHERF2.Support or Funding InformationFunded by SFB621/C1, DFG grants SE460/13‐4, 19‐1 and 21‐1.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Chlorogenic Acid Enriched in Coffee Pulp Extract Attenuates Hepatic Steatosis by Modulating Gene‐regulated Lipid Metabolism and Lipid Transporters
Authors: Ontawong Atcharaporn , Pasachan Tipthida , Soodvilai Sunhapas , Duangjai Acharaporn , Pongchaidecha Anchalee , Amornlerdpison Doungporn , Srimareong Chutima 
Published: June 2021

Background & AimsCoffee pulp (CP) is the first by‐product of coffee processing. Major constituents in CP include chlorogenic acid (CGA), epicatechin (EC), caffeine exhibited antioxidant, lipid lowering action, and several biological activities. However, it remains unknown whether CP rich CGA improved hepatic steatosis. This study aimed to investigate the effect of CP aqueous extract (CPE) on hepatic steatosis, and identified the mechanisms involved in hepatocellular carcinoma (HepG2) cells.MethodsHepG2 cells at the cell density of 5×105 cells/well were pre‐incubated with CPE (10–200 μg/ml), CGA, and caffeine for 24 hr. Intracellular lipid accumulation was quantified using Nile red staining followed by flow cytometry. Cell viability of HepG2 cells was determined using MTT assay. The mRNA expression of genes involved in lipid metabolism and lipid transporters were also determined using quantitative real‐time PCR.ResultsCPE at the dose of 50–200 μg/ml, CGA, and caffeine significantly decreased lipid accumulation in HepG2 cells. Correspondingly, CPE decreased lipogenesis‐associated genes: PPARγ, and up‐regulated lipolysis‐related genes: PPARα. In addition, CPE reduced the lipid uptake transporter, FAT/CD36, while increased the lipid efflux transporters, ABCG5/8.ConclusionCPE has hepatic lipid‐lowering effects by modulating genes related lipolysis, lipogenesis, and lipid transporters. These findings indicate that anti‐hepatic steatosis by CPE is attributed to hepatic lipid‐lowering action for reversing such pathology. Thus, CPE could potentially be developed into nutraceutical product for prevention of non‐alcoholic fatty liver disease.Support or Funding InformationThis work was supported by the Faculty of Medicine Endowment Fund (066/2560 to CS), Chiang Mai University, the Research and Researcher for Industry (RRI) by Thailand Research Fund (TRF) (PHDI0010 to CS), and the NSTDA Chair Professor grant (the Fourth Grant) of the Crown Property Bureau Foundation and the National Science and Technology Development Agency to Professor Dr. Vatcharin Rukachaisirikul.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Oligomerization of ferroportin and the mechanism of autosomal dominance in ferroportin disease
Authors: Bonamer John P. , Ruwe T Alex , Qiao Bo , Vieth Kyle R. , Ganz Tomas , Nemeth Elizabeta , Mackenzie Bryan 
Published: June 2021

Ferroportin (Fpn)—the only known cellular iron‐export protein—plays a critical role in iron homeostasis by mediating iron export from macrophages and enterocytes. Rare, loss‐of‐function mutations in human Fpn (e.g. V160del) cause classical ferroportin disease, characterized by macrophage iron loading and iron‐restricted anemia. Ferroportin disease displays an autosomal–dominant inheritance pattern. Because all of the disease‐associated mutations are missense, the dominant–negative effect is most readily explained as resulting from oligomerization of Fpn protein. As a first step in testing this hypothesis, we have examined the functional activity of wildtype (wt) and mutant human Fpn co‐expressed in RNA‐injected Xenopus oocytes. We measured 55Fe efflux from control oocytes, or oocytes expressing wtFpn or V160del GFP proteins with or without co‐expression of a wtFpn–mCherry fusion protein. The RNA dose eliciting half‐maximal iron‐export activity was 12.1 ± (SEM) 1.6 ng for wtFpn–GFP (P = 0.005) and 11.7 ± 2.2 ng for wtFpn–mCherry (P = 0.013), so we injected 12 ng RNA of each construct in subsequent co‐expression experiments. 55Fe efflux activity resulting from co‐expression of wtFpn–GFP and wtFpn–mCherry constructs was additive (Fig 1). Whereas expression of V160del–GFP alone did not stimulate 55Fe efflux, co‐expression of the mutant with wtFpn–mCherry stimulated 55Fe efflux to roughly the same degree as either of the wildtype constructs alone. Our results therefore provide no evidence of a dominant–negative effect of V160del. Our next step will be to test co‐expression of wtFpn with other loss‐of‐function mutants. We will examine the oligomeric structure of wildtype and mutant Fpn by using freeze‐fracture electron microscopy to examine the interior (p) face of the inner lamella of the plasma membrane, as previously described for several membrane‐transport proteins [Eskandari S et al (1998) Proc Natl Acad Sci USA 95, 11235–11240]. We anticipate that our study will further elucidate the structure–function of Fpn and help to explain the autosomal dominance of ferroportin disease.Support or Funding InformationNIH–NIDDK grant R01 DK107309This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Regulation of Intestinal Phosphate Transport in Humans and in a Rat Model of Chronic Kidney Disease
Authors: Myakala Komuraiah , Wang Xiaoxin , Levi Moshe 
Published: June 2021

In chronic kidney disease (CKD) hyperphosphatemia is a common occurrence and plays important roles in cardiovascular and bone disease. The mechanisms however still remain unknown and the role of intestinal phosphate (Pi) transport is subject of ongoing debate. We have determined that Na/Pi cotransport activity and NaPi‐2b protein is expressed in the duodenum, jejunum, and ileum of human intestinal samples. We further studied regulation of intestinal phosphate transport in a model of 5/6 nephrectomy (Nx) induced CKD in the rat fed a relatively high Pi diet (1.5% Pi, 0.6% Ca). Male rats with 5/6 Nx had a marked increase in serum BUN (37.7 ± 2.0 vs. 150.5 ± 29.8 mg/dl in sham control, p<0.02), serum creatinine (0.5 ± 0.06 vs. 1.65 ± 0.23 mg/dl in sham control, p<0.008), and serum Pi (7.48 ± 0.85 vs. 18.19 ± 1.84 mg/dl in sham control, p<0.001). We isolated apical brush border membrane vesicles (BBMV) from the duodenum and the jejunum and studied sodium gradient dependent Pi (Na+/Pi) cotransport as a function of pH. Across every single pH studied, including 5.5, 6.0, 6.5, 7.0, and 7.5 Na+/Pi transport activity was increased in BBM from 5/6 Nx rats. This was associated with a 2‐fold increase in NaPi‐2b protein abundance in the duodenum and a1.4‐fold increase in NaPi‐2b protein abundance in the jejunum. To determine if there are sex‐dependent differences in CKD and intestinal Na+/Pi transport, we also studied in parallel female rats with 5/6 Nx. We found that female rats with 5/6 Nx also had a marked increase in serum Pi (4.75 ± 0.39 vs. 10.7 ± 1.24 mg/dl in sham control, p<0.003). In addition, there were marked increases in intestinal Na+/Pi transport activity paralleled by increases in NaPi‐2b protein abundance.Our study therefore indicates that in both male and female rats with CKD, increased intestinal Na+/Pi transport activity and increased NaPi‐2b protein expression mediate the hyperphosphatemiaThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The local environment of cysteine 621 determines the rapid electrophilic adduction and activation of hTRPA1
Authors: Bahia Parmvir K. , Parks Thomas A. , Stevens Stanley M. , Varma Sameer , Taylor‐Clark Thomas E. 
Published: June 2021

Transient Receptor Potential Ankyrin 1 (TRPA1) is a non‐selective cation channel expressed largely on nociceptive sensory nerves. TRPA1 is rapidly activated by electrophiles, such as iodoacetamide, through covalent modification of intracellular cysteine (Cys) residues. We have previously shown that, at physiologically relevant exposures, iodoacetamide rapidly labels four Cys residues at positions 273, 621, 655, 1085. Of these, Cys residue at position 621 (C621) appears to be crucial for binding and activation of the human channel (hTRPA1). Interestingly, when comparing hTRPA1 with the electrophile‐insensitive rattlesnake homolog, while the amino acids lysine K620 and C621 are conserved the proline at P622 is not. We hypothesized that this rapid binding of C621 residue in human TRPA1 is mediated by neighboring amino acids K620 and P622 rendering it more sensitive to electrophilic adduction. We expressed V5‐His tagged hTRPA1 constructs in HEK293 cells including the wild‐type and the point mutants C621A, K620A and P622A. We used pulse‐chase experiments to label TRPA1 protein with iodoacetamide‐alkyne (IA‐alk), then used click chemistry to label with a fluorescent tag. Binding was measured using immunoprecipitated TRPA1 separated by SDS‐polyacrylamide gel electrophoresis and western blotting. TRPA1 channel activation was measured using ratiometric calcium imaging. The K620A mutant had the effect of greatly reducing electrophilic binding and activation of the channel. Modeling of deprotonation energies suggests that K620 contributes to C621 reactivity by reducing its pKa. The K620 mutant therefore likely reduces the probability that C621 will be thiolate and therefore able to rapidly react with electrophiles. The P622A mutant was also largely insensitive to electrophilic adduction and activation. IA‐alk binding was ~10% of that seen with the wild‐type channel. We hypothesize this is because, in the wild‐type hTRPA1, P622 induces a kink in the protein resulting in the positively charged K620 being closer to C621 compared with the nearby negatively charged glutamates, E625 and E681. The work described here highlights the importance of the amino acids K620 and P622 flanking C621. Our data demonstrate how the integrity of the local environment is essential to TRPA1 function as an electrophilic sensor.Support or Funding InformationThis work was supported by the National Heart Lung and Blood Institute in Bethesda, USA(R01‐HL119802‐S1)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Age‐dependent Changes in miRNA Profile in F344 rat and C57BL/6J mice: Role of sodium hydrogen exchanger regulatory factor‐1 (NHERF1)
Authors: Jain Anish , Aubee Joseph I. , Thompson Karl M. , Lee Dexter L. , Khundmiri Syed Jalal 
Published: June 2021

Our laboratory demonstrated decreased NHERF1 expression in aging F344. Loss of NHERF1 expression prevented dopamine‐dependent inhibition of NKA in renal PTCs from aged F344. We hypothesized that loss of NHERF1 expression in aged F344 rats is due to changes in miRNA expression. To address this hypothesis we analyzed the miRNA expression profile in 4mo and 22mo old F344 rats and 2 and 18mo old C57BL/6J wild‐type and NHERF1 knock‐out mice. Total RNA from kidney cortex was isolated using Trizol (miRVana, Ambion) and miRNA profile was analyzed using kits and software from Nanostring. The top 100 miRNAs with 25% counts above the negative miRNA counts were considered for analysis. Counts from young animals (n=4, 2M2F) were averaged together and considered as 1. Counts from old animals (n=4, 2M2F) were calculated as fold change. In F344 rats 10 miRNAs increased with age (miR29a, miR29b, miR29c, and miR21), while miR96 and miR328a decreased. In WT‐ mice 51 miRNAs increased with age (miR‐let‐7g, b, and d; and miR22), while 28 miRNAs decreased (miR21 and miR‐let‐7e). In NHERF1‐KO mice 31 miRNAs increased with age (miRNA21, miR29a, miR29b, and miR29c), while 22 miRNAs decreased (miR‐let‐7a, miR‐let‐7b, miR‐let‐7c, and miR24). Our results suggest that miRNAs that regulate inflammation increased with age in both F344 rats and NHERF1‐KO mice. Interestingly, these miRNAs decreased in WT mice. These data may explain the mechanisms of increased renal inflammation in NHERF1 deficient models of hypertension.Support or Funding Information7R21AG047474‐03 NIHR25AG047843‐NIH16GRNT31030019‐AHAThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effect of Gut Microbiota Modulation on Hepatic Lipid Metabolism
Authors: Pereira Louise Tavares Garcia , Beserra Bruna Teles Soares , Engel Daiane Fátima , de Paula Gabriela Cristina , de Bema Andreza Fabro , Amato Angélica Amorim 
Published: June 2021

Non‐alcoholic fatty liver disease is characterized by fat accumulation in the liver and commonly seen in patients with components of the metabolic syndrome. The mechanisms underlying its development are not completely defined, but there has been an increasing body of evidence linking gut microbiota to the development and progression of fat accumulation in the liver. Our objective was to investigate the effect of gut microbiota modulation on hepatic lipid metabolism.MethodsSix week old C57BL/6 male mice, fed with a control diet (CD, n=18) or high‐fat diet (HFD, n=30). At the age of 7 weeks, they were randomly assigned into 5 groups to receive (i) CD and vehicle, (ii) CD and antibiotics, (iii) HFD and vehicle, (iv) HFD and antibiotics, and (v) and HFD and antibiotics followed by fecal transplantation from CD‐ fed donors. Antibiotics (ciprofloxacin 0.2g l−1 and vancomycin 0.5 g l−1) were administered from the 10th to the 15th week, in drinking water. Fecal transplantation was carried out at the 13th week, following antibiotic withdrawal and maintenance of recipient mice on HFD. Weight was measured weekly and water intake twice a week. An oral glucose tolerance test (GTT) was performed at the 12th weeks. Mice were euthanized by decapitation and liver samples were stored for histological analysis and gene expression by real time quantitative polymerase chain reaction.ResultsHFD‐fed mice showed significantly increased weight gain when compared with CD‐fed mice. Antibiotic treatment did not change weight gain in CD‐fed mice but increased weight gain in HFD‐fed mice. Mice fed a CD and treated with antibiotics showed increased glucose tolerance when compared with control (CD, vehicle), and antibiotic treatment of HFD‐fed mice did not change glucose tolerance. CD‐fed mice treated with antibiotics exhibited increased expression of mRNA levels of PPAR‐α in liver and no change in lipogenesis‐related transcripts (Srepb1 and Acsl1), when compared with CD‐fed mice treated with vehicle. Mice fed a HFD and treated with antibiotics showed no change in PPARa, Srepb1 and Acsl1 levels when compared with control (HFD, vehicle), whereas HFD‐fed mice receiving fecal transplantation from CD‐fed mice showed a trend towards increased PPARa levels in the liver.ConclusionOur preliminary data may suggest that gut microbiota modulation with antibiotics may affect fatty acid metabolism in the liver by changing the balance between expression of lipogenesis and fatty acid oxidation‐related genes. Further studies are needed to clarify the extension and mechanisms underlying these effects.Support or Funding InformationThis study was supported by the Fundação de Amparo à Pesquisa do Distrito Federal (FAP‐DF).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
CART‐GPR160 Regulation of Food Intake
Authors: Haddock Christopher J. , Yosten Gina L.C. , Kolar Grant R. , Samson Willis K. 
Published: June 2021

Cocaine and amphetamine regulated transcript (CART) mRNA was identified in 1995 as a novel transcript increased in the striatum after acute cocaine or amphetamine administration. This transcript matched the protein sequence of an Ovine hypothalamic peptide partially sequenced by Spiess and Vale in 1981, implying that this transcript was indeed translated into an active peptide product. CART peptide is a multifunctional neuromodulator with roles in pain, reward, addiction, and food intake. Using our deductive reasoning strategy (Yosten et al. AJP303:R941, 2012) we have identified the previously orphaned G‐protein coupled receptor, GPR160, as a candidate CART receptor. Colocalization, proximity ligation, and immunoprecipitation studies have confirmed the association of CART and GPR160. We translated this evidence into in vivo rat food intake studies. The action of CART to inhibit food intake (4V administration) was lost in animals pre‐treated with siRNA targeting the GPR160 transcript. Additionally, passive immunoneutralization of GPR160 achieved by injecting antibody against the second extracellular loop of GPR160 into the fourth ventricle also blocked the anorexic effects of pharmacologic doses of fourth ventricle CART peptide. Taken together, these data identify GPR160 as a necessary component of the CART peptide signaling pathway and further demonstrate the requirement for this interaction to mediate the food intake reducing effects of CART peptide. Research on the function of CART peptide had been severely hindered by the lack of knowledge of a cognate receptor. The discovery of a CART peptide receptor will not only revitalize research in the CART field, but also potentially lead to novel therapeutic agents for the treatment of obesity and pain management.Support or Funding InformationHL121456This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Modeling Effects of a High‐Fat (Almonds) vs. High‐Carbohydrate (Graham Crackers) Snack on Metabolic Outcomes in College Freshmen
Authors: Barron Marily , Dhillon Jaapna , Asghar Syed A. , Kuse Quintin , De La Cruz Natalie , Vu Emily , Sindi Suzanne S. , Ortiz Rudy M. 
Published: June 2021

Biomedical research has provided support for the benefits of consuming almonds, a high‐fat food. However, most of the studies to date have considered only adult populations (age 40+). The objective of the study was to model the impact of consuming a high‐fat almond snack vs. a high‐carbohydrate graham cracker (control) snack for 4–8‐weeks in college freshmen. Seventy‐three UC Merced freshmen (Age: 18–19 years, BMI: 18–45 kg/m2) underwent a dietary intervention for eight weeks. Thirty‐eight participants consumed 2 ounces (325 kcal) of almonds and 35 participants consumed an isoenergetic control snack of graham crackers for 8 weeks. Anthropometric, biochemical and clinical measurements were conducted before the study began and at the fourth and the eighth week of the intervention. Several multivariate models were generated to model the effects of chronic almond and graham cracker snacking on the aforementioned outcomes. The most profound effect was the retention of high‐density lipoprotein (HDL) cholesterol and improvements in glucose tolerance in the almond snack group in comparison to the graham cracker snack group. This suggests that chronic almond snacking has the potential to improve the metabolic profile independent of profound changes in body mass or adiposity. We continue to perform the robust multivariate analyses factoring in BMI and sex effects.Support or Funding InformationAlmond Board of California, MARC U STAR, and UROCThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.