bluuebunny/crossref_metadata_2025_split huggingface dataset:
Insight into flavonoid structure for mammalian α‐amylase and α‐glucosidase inhibition to control starch digestion rate
Authors: Lim Jongbin , Zhang Xiaowei , Ferruzzi Mario , Hamaker Bruce R. 
Published: June 2021

In this study we investigated the structural specificity of flavonoids for starch digestive enzyme inhibition to modulate glucose release from glycemic carbohydrate digestion. Using three different flavonoids [eriodictyol, luteolin, and quercetin], chosen based on the degree of hydroxylation and planarity of the C‐ring, structural specificities were found for selectively inhibiting α‐amylase and α‐glucosidases, resulting in different inhibition constants (Ki) and mechanisms. The double bond between C2 and C3 on the C‐ring of flavonoids was particularly important for α‐amylase inhibition, which leads to a π‐stacking interaction between flavonoids and α‐amylase, while the hydroxyl group at C3 of the C‐ring played a key role in inhibiting α‐glucosidases. These structural specificities of flavonoids toward starch degrading enzymes are likely the result of different protein structures of α‐amylase and α‐glucosidases, as they belong to different families, GH13 and GH31. Our findings provide insights into structure‐function aspects of flavonoids in controlling starch digestion rate.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
O‐GlcNAc signaling orchestrates metabolic adaptation to prolonged fasting
Authors: Li Mindian , Liu Jiayu , Sehgal Raghav , Wu Jing , Zhang Pei , Han Weiping , Jha Abhishek , Yang Xiaoyong 
Published: June 2021

Prolonged fasting reprograms metabolic pathways to foster stress resistance. O‐linked b‐D‐N‐acetylglucosamine (O‐GlcNAc) glycosylation is essential in metabolic regulation and stress resistance. It is unknown whether and how O‐GlcNAc signaling catalyzed by O‐GlcNAc transferase (OGT) reprograms metabolic pathways during prolonged fasting. Here we report that OGT coordinates purine, lipid, amino acid and carbohydrate metabolism during prolonged fasting. Unbiased metabolomic profiling of mouse livers after 48‐hour fasting reveals that hepatic ablation of OGT impedes fasting‐associated changes in purine metabolism, phospholipid degradation and fatty acid oxidation. Loss of OGT also impedes amino acid catabolism through reduced flux through the urea cycle and reduced aspartate‐argininosuccinate shunt. Notably, the citric acid cycle is also inhibited in part through inhibition of succinate dehydrogenase activity, which is consistent with decreased oxygen consumption and energy production. These catabolic defects are correlated with reduced gluconeogenesis. Together, these results define OGT as a master regulator of metabolic adaptation during prolonged fasting in liver.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Diacylglycerol kinase δ interacts with sphingomyelin synthase‐related protein via their sterile α motif domains
Authors: Murakami Chiaki , Sakane Fumio 
Published: June 2021

IntroductionDiacyglycerol kinase δ isozyme (DGKδ) plays critical roles in lipid signaling by phosphorylating diacylglycerol (DG) into phosphatidic acid (PA). DGKδ regulates a wide variety of physiological and pathological events, such as type II diabetes and obsessive compulsive disorder. Because DGK is one of the components of phosphatidylinositol (PI) turnover, it is thought that DGKδ also utilizes mainly 18:0/20:4‐DG (X:Y; the total number of carbon atoms: the total number of double bonds) derived from PI turnover. Interestingly, we recently demonstrated that DGKδ preferably metabolized palmitic acid (16:0)‐containing DG molecular species, but not arachidonic acid (20:4)‐containing DG species, in response to high glucose stimulation. However, it is still unclear what kind of DG‐generating enzyme provides palmitic acid‐containing DG species.Sphingomyelin synthase (SMS) 1, SMS2 and SMS‐related protein (SMSr) are DG‐generating enzymes utilizing phosphatidylcholine/phosphatidylethanolamine and ceramide. SMS1 and SMSr contain a sterile α motif domain (SAM), which is a protein‐protein interaction module, at their N‐termini. DGKδ also possesses SAM at its C‐terminus, and DGKδ‐SAM is highly homologous to SMSr‐SAM. Therefore, we hypothesised that DGKδ interacts with SMSr through their SAMs.In the present study, we investigated the interaction between DGKδ‐SAM and SMSr‐SAM.ResultsWe first examined whether DGKδ‐SAM interacts with SMSr‐SAM by immunoprecipitation analysis. We found that SMSr‐SAM, but not SMS1‐SAM, was co‐immunoprecipitated with DGKδ‐SAM. Full‐length DGKδ (DGKδ‐FL) was also co‐immunoprecipitated with SMSr‐FL more strongly than with SMS1‐FL and with SMS2‐FL. To examine whether SMSr‐SAM contribute to the interaction, we performed immunoprecipitation analysis using SMSr‐FL and a SAM‐deletion mutant (SMSr‐DSAM). DGKδ‐FL was strongly co‐immunoprecipitated with SMSr, whereas SMSr‐DSAM only weakly interacted with DGKδ‐FL. Immunostaining analysis demonstrated that DGKδ‐FL co‐localized partly with SMSr‐FL in COS‐7 cells overexpressing these proteins. These results strongly suggest that DGKδ interacts with SMSr through their SAMs.ConclusionIn summary, the present study for the first time showed that DGKδ interacted with SMSr mainly through their SAMs. It is possible that SMSr is one of the candidates of up stream DG‐providing enzymes of DGKδ, which composes a new pathway independent of PI turnover.Support or Funding InformationThis work was supported by JSPS KAKENHI (Grant Number JP18J20003) and supported in part by Venture Business Laboratory in Chiba University (Nanohana Competition 2018 Award).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Characterization of the Perilipin 5 C‐Terminus
Authors: Clinton Tim D , Wei Derek T , Tansey John T 
Published: June 2021

Many countries around the globe have been plagued with a recent epidemic in lipid‐associated diseases. These diseases include obesity, diabetes, and non‐alcoholic fatty liver disease among many others. The United States itself has fallen heavily victim to these ailments, placing first in worldwide rankings for obese persons and within the top fiftieth in rankings for the percentage of persons afflicted with diabetes. These diseases pose clear threats to modern society and having a greater understanding of the mechanistic background of them may lead to better treatments options for those afflicted. A common thread through all these devastating illnesses is their connection to lipids and more specifically lipid storage. Cellular lipids are stored in lipid storage droplets, and these droplets are coated by the perilipins, five regulatory proteins that have an integral role in regulating the breakdown of fats. One such of these family members is perilipin 3, a protein that's structure has been partially determined. In Perilipin 5 this structure has not been determined and is the current focus of this study. Previously a crystal structure of the C‐terminal region of perilipin 3 was determined for residues 206 to 431. These regions were aligned with perilipin 5 using BLAST and their analogous location within the Perilipin 5 DNA sequence was determined. This region of perilipin 5 is being expressed as a 6‐his fusion protein for subsequent characterization and structure determination. Using the predicative programs RaptorX and Phyre2 theoretical protein structures were compiled. These were labeled for regions of interest including a putative hydrophobic cleft found in perilipin 3 that was previously found to recruit that protein to lipid droplets. Collectively these findings indicate that it is highly likely that perilipin 5 has a similar structure to perilipin 3 in the carboxy terminus.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Characterization of a Shortened Form of the Perilipin 5 Protein
Authors: Dalton Rebekah C , Tansey John T 
Published: June 2021

Many modern health issues arise from aberrant lipid metabolism, among these are type II diabetes and non‐alcoholic fatty liver disease. One commonality these diseases share is neutral lipid metabolism and storage. Regulation of neutral lipid metabolism is of vital importance in the body, and among the proteins responsible for such regulation are the perilipins. Perilipins are a family of conserved proteins that are found on the surface of lipid storage droplets and play a central role in the regulation of cellular neutral lipid metabolism. Perilipin 5, specifically, is expressed in tissues with a high capacity for fatty acid oxidation such as cardiac muscle, oxidative muscle, and fasting liver. Using western blotting and reverse transcriptase PCR, we have identified a splice variant of perilipin 5 which has been termed perilipin 5b. This shortened form of the protein retains the amino terminal 35 kDa of the protein but lacks the 4‐helix bundle found in the C‐terminus. Based on the reports in the literature, this protein would lack the carboxy terminal domains reported to be necessary for interactions with lipases and other proteins but would retain serine 155 that is requisite for PKA phosphorylation and translocation to the nucleus. We have constructed a carboxy terminal truncation of perilipin 5 and are expressing it in CHO cells to further define the function of this protein. When compared with CHO cellular expression of perilipin 5, 5b appears to have a higher number of smaller lipid droplets per cell. Collectively these data indicate that perilipin 5b plays a role in lipid storage, but its specific function of this shortened form is still unknown.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cost‐effective synthesis of polyethylene glycol diacrylate‐acrylic acid (PEGDA‐AA), a tissue engineering hydrogel ideal for C2C12 mouse muscle cells
Authors: Lee Hunter M. , Fischer Kristin M. 
Published: June 2021

Current research in tissue engineering focuses on growing and functionalizing cells in vitro to assemble viable constructs that restore, maintain, or improve damaged tissues or whole organs. Specific to the field of tissue engineering, hydrogels have proven successful in the development of multiple cell types with the goal of one day serving as grafts that will replace large areas of lost or damaged tissue. This assembly depends on the presence of a scaffold, or three‐dimensional structure that guides the growth of new tissue, and hydrogel polymers in particular have been known to be an excellent building block for scaffold formation. This research focused on the synthesis and characterization of the hydrogel polyethylene glycol diacrylate‐acrylic acid (PEGDA‐AA), a hydrophilic material that provides a viable environment for cell maturation because of its favorable characteristics for the development and maturation of mouse C2C12 muscle cells. Using modified procedures adapted from Browe et al. (2016) to create a cost‐effective way of synthesizing this precursor, PEGDA was successfully synthesized and characterized through 13C and 1H‐NMR spectroscopy. Current research focuses on analyzing the effects of the hydrogel on the growth and development of C2C12 cells with the long‐term goal of establishing an efficient model for human tissue engineering.Support or Funding InformationHampden‐Sydney College Office of Undergraduate ResearchThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Investigating Pericyte Dynamics during Angiogenesis in the Mouse Mesometrium Culture Model
Authors: Suarez‐Martinez Ariana Diandra , Wang Jack , Murfee Walter Lee 
Published: June 2021

Angiogenesis requires multiple cells and systems working in concert to develop new blood vessels from existing ones. Hence, understanding the exact dynamics between cells and systems proves to be beneficial when developing innovative therapies. In the microcirculation, endothelial cells that line the blood vessels are supported by perivascular cells that are elongated cells called pericytes. Cell membrane proteins that modulate cellular responses of different growth factors play an important role during microvascular remodeling. For pericytes, the neural glial antigen‐2 (NG2) proteoglycan is a cell membrane protein that is exclusively expressed by mural cells in mouse microvasculature. Pericytes' multifaceted roles in angiogenesis and where they originate still requires further research. Our lab has recently introduced the mouse mesometrium culture model that allows for the investigation of cell dynamics within intact microvascular networks. The objective of this study was to demonstrate the usefulness of the mouse mesometrium culture model for investigating NG2 inhibition and cell‐lineage tracing. For the NG2 inhibition studies, female C57BL/6 (WT) and NG2 KO were euthanized to aseptically harvest mesometrium tissues and culture them in MEM + 1% PS and 20% FBS at standard incubating conditions. WT and NG2 KO tissues were cultured for 3 days before labeling with PECAM and NG2 antibodies to quantify angiogenesis. PECAM and NG2 labeling confirmed the lack of NG2‐positive pericytes on capillaries from NG2 KO tissues. The angiogenic response after culture from NG2 KO tissues was significantly decreased when compared to WT tissues (NG2 KO: 0.98 ± 0.29; WT: 2.11 ± 0.32 number of sprouts per total vessel length, p = 0.019). For cell‐lineage tracing studies, female NG2dsRed mice were given a daily IP injection of 0.1 mL of sunflower oil for 5 consecutive days to increase microvascular density. Mesometrium tissues were harvested 21 days post last injection and cultured in MEM + 1% PS and 20% FBS at standard incubating conditions for 5 days. The tissues were imaged for time‐lapse at least every 12 hours to track individual cell interactions. Time‐lapse imaging taken every 2 hours for 12 hours on days 1–3 of NG2dsRed tissues revealed that NG2‐positive pericytes migrated along pre‐existing vessels to new vessel segments. Moreover, a subpopulation of NG2‐positive interstitial cells were observed to migrate throughout the tissue. These results validate that inhibiting NG2 expression has a direct effect on angiogenesis in the mouse mesometrium culture model. Furthermore, it highlights the usage of the mouse mesometrium culture model to track individual cell‐to‐cell dynamics of pericytes in intact microvasculature during angiogenesis.Support or Funding InformationNIH R01AG049821This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Chronic GPER1 Activation Prevents H9c2 Cardiac Myoblast Death after Oxidative Stress by Protecting Mitochondrial Structure and Function Via a Non‐Canonical MST1/YAP Pathway
Authors: Madungwe Ngonidzashe , Feng Yansheng , Bopassa Jean Chrisostome 
Published: June 2021

IntroductionEstrogen (17β‐estradiol; E2) is known to be cardioprotective against the damaging effects of ischemia/reperfusion (I/R) injury. We found that acute pre‐ischemic E2 treatment reduces cell death in cardiac tissue via direct effects on mitochondria by the G protein‐coupled estrogen receptor 1 (GPER1). We recently showed that acute application of E2 at the onset of reperfusion results in similar cardioprotection against I/R injury via GPER1, a non‐genomic receptor that mediates rapid E2 actions. Here, we investigated the impact and mechanism underlying chronic E2 protective actions in H9c2 cardiac myoblasts subjected to treatment with the cytotoxic agent, H2O2.MethodsUsing the H9c2 cardiac myoblasts, we subjected cells to H2O2 oxidative stress with no serum, followed by reoxygenation with E2 for 2 and 4 days. Cells were then collected and analyzed for cellular, mitochondrial and genetic function. We measured cell death, gene upregulation, mitochondrial structure, morphology and function using flow cytometry, cell assays, qRT‐PCR, along with fluorescence and electron microscopy. We also collected cell lysates and analyzed them by Western blot to assess the mechanism involved.ResultsWe found that chronic treatment of cells with E2 after oxidative stress results in decreased cell death but remarkably does not affect cell cycle activity. Surviving cells showed improved mitochondrial dynamics as evidenced by better cristae morphology and increased resistance to opening of the mitochondrial permeability transition pore, but with little change to mitochondrial fusion/fission. Additionally, we found that this chronic E2 action predominantly occurs through mitochondria to reduce cytochrome c release and decrease phosphorylation of mammalian sterile‐20‐like kinase (MST1) and increase yes‐associated protein (YAP) translocation to the nucleus for upregulation of TGF‐β and PGC‐1α. All these E2‐observed effects were prevented by G15, a GPER1 antagonist, suggesting a GPER1‐dependent mechanism.ConclusionWe have found that chronic E2 treatment to H9c2 cardiac myoblasts after stress leads to rescue from cell death by reducing apoptotic signaling and protecting mitochondrial morphology and function. We also found that this process occurs via GPER1 acting directly on mitochondria in a non‐canonical MST1/YAP mechanism that leads to genetic upregulation of cell growth.Support or Funding InformationNIH RO1 Grant/Award number: HL138093;American Heart Association, Grant/Award Numbers:18PRE34030307, 17SDG33100000This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Contribution of Membrane Raft Redox Signaling to Visfatin‐Induced Inflammasome Activation and Podocyte Injury
Authors: Singh Gurinder Bir , Patibandla Sai , Zhang Yang , Li Pin‐Lan , Koka Saisudha , Boini Krishna M 
Published: June 2021

Recently we have shown that adipokine visfatin‐induced NLRP3 inflammasome activation contributes to podocyte injury. However, the molecular mechanisms of how visfatin‐induces the Nlrp3 inflammasome activation and podocyte damage still unknown. The present study tested whether ceramide and membrane raft (MR) redox signalling pathway plays a central role in visfatin‐induced inflammasomes formation and activation in podocytes. Using confocal microscopy, visfatin was found to increase MRs clustering in the membrane of podocytes in a dose dependent manner. Upon visfatin stimulation an aggregation of ASMase product, ceramide and NADPH oxidase subunits, gp91(phox) and p47(phox) was observed in the MR clusters, forming a MR redox signaling platform. The formation of this signalling platform was blocked by prior treatment with MR disruptor MCD, NOX inhibitor DPI, ASMase inhibitor amitriptyline. In addition, electron spin resonance (ESR) spectrometry analysis showed that visfatin treatment significantly increased the superoxide (O2·−) production compared to control cells. However, prior treatment with ASMase inhibitor amitriptyline significantly attenuated the visfatin‐induced superoxide production. Furthermore, immunofluorescence analysis demonstrated that visfatin treatment significantly decreased the podocin and nephrin expression (podocyte damage), and prior treatments with DPI, WEHD (inflammasome inhibitor), MCD and amitriptyline attenuated this visfatin‐induced podocin reduction. In conclusion, our results demonstrate that visfatin‐induced the ceramide production via ASMase and thereby stimulates membrane raft clustering in the membrane of podocytes to form redox signalling platforms by aggregation and activation of NADPH oxidase subunits, enhancing O2·− production and leading to Nlrp3 inflammasome formation and activation in podocytes and ultimate glomerular injury (supported by NIH grant, DK104031).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Carcinogenic arecoline shows a paradoxical effect in modulating nitric oxide production and the subsequent protein S‐nitrosylation in endothelial cells
Authors: Cheng Bi‐Ling , Fu Yaw‐Syan , Su Yuan‐Shun , Huang Bin 
Published: June 2021

Arecoline exhibits a carcinogenic tone and is remarkable for the refreshing effect due to the increase of vasodilation and blood flow. Therefore, if arecoline can promote the production of nitric oxide (NO) and the subsequent protein S‐nitrosylation in endothelial cells (ECs) is interesting. Treating arecoline increased the phosphorylation of eNOSSer1177. NO molecular was observed by a fluorescent probe FA‐OMe followed with the measurement of flowcytometry. Arecoline treatment up‐/down‐regulated 914/647 proteins identified by showing by iTRAQ 114/115 labeling and LC‐MS/MS. IPA analysis indicated that these proteins were implicated in eNOS signaling. By using biotin switch, 224/159 S‐nitrosoproteins were up‐/down‐regulated by arecoline. The increased S‐nitrosylation level of chaperonin and the decreased S‐nitrosylation level of calnexin were verified by western blot. Our results suggest a paradoxical effect of arecoline in producing NO and therefore maintains endothelial homeostasis, even though the toxicological tone in carcinogenesis remains concerned.Support or Funding InformationTB0357 from Ministry of Education, TaiwanThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.