bluuebunny/crossref_metadata_2025_split huggingface dataset:
Energy production in patients with chronic obstructive pulmonary disease: Evidence of mitochondrial and not O2 supply limitation
Authors: Layec Gwenael , Hart Corey , Trinity Joel , Haseler Luke , Le Fur Yann , Jeong Eun‐Kee , Richardson Russell 
Published: November 2023

It is well documented that chronic obstructive pulmonary disease (COPD) is associated with both muscular and vascular dysfunctions. Interestingly, although both peripheral oxygen (O2) transport and muscle O2 utilization capacity appear to be impaired in patients with COPD, it is still unclear which peripheral component of the O2 cascade actually limits overall muscle aerobic capacity. Therefore, purpose of the present study was to determine whether mitochondrial function in vivo is limited by convective O2 delivery or the intrinsic capacity of mitochondria to synthesize ATP in patients with COPD. To this aim, we combined phosphorus magnetic resonance spectroscopy (31P‐MRS), near‐infrared spectroscopy (NIRS), and Doppler ultrasound, to examine the effect of superimposing reactive hyperaemia (RH), induced by a period of brief ischemia during the last min of exercise, on O2 delivery and mitochondrial function in the calf muscle of 16 patients with moderate to severe COPD in comparison to free‐flow conditions (FF). Limb blood flow [area under the curve (AUC), FF: 2.5 ± 1.2 L; RH: 3.3 ± 1.0 L, P<0.05] and convective O2 delivery (AUC, FF: 0.46 ± 0.23 L; RH: 0.61 ± 0.21 L, P=0.08) were increased in RH in comparison to FF. RH was also associated with significantly higher capillary blood flow (AUC, FF: 26 ± 14 L; RH: 37 ± 25 L, P<0.05). However, despite an increased muscle perfusion and O2 delivery, the peak rate of mitochondrial ATP synthesis did not differ significantly between conditions (FF: 28 ± 14 mM.min−1; RH: 27 ± 9 mM.min−1, P=0.83). Taken together, these results indicate that an increase in convective O2 delivery does not improve mitochondrial function in the plantar flexor muscles of the patients with COPD, thereby revealing that the peak rate of mitochondrial ATP synthesis of these patients appears to be limited by the intrinsic mitochondrial capacity and not O2 availability.Support or Funding InformationThis work was funded in part by grants from the Flight Attendant Medical Research Institute (FAMRI), NIH National Heart, Lung, and Blood Institute (K99HL125756, PO1 HL 091830) and VA Merit grant E6910R.
Adipose‐Derived Exosomes From Severely Obese Individuals Regulate Skeletal Muscle Metabolism
Authors: Park Sanghee , Zheng Donghai , Barberio Matt , Hubal Monica , Houmard Joseph 
Published: November 2023

Accumulation of subcutaneous and visceral fat mass with severe obesity (BMI > 40 kg/m2) is closely associated with various metabolic derangements, including insulin resistance, type 2 diabetes and cardiovascular disease. The mechanism in which expanded visceral tissue promotes insulin resistance in peripheral tissues (i.e. skeletal muscle) is unclear. Exosomes, extracellular nano‐vesicles secreted from almost all types of organs in the body, are known to regulate cellular signaling through endocrine mechanism suggesting that exosomes released from visceral fat can be one of important mediators to communicate with skeletal muscle. In addition, a previous study has shown that certain mature miRNAs from visceral exosomes, which target many confirmed or putative mRNAs, are differentially expressed in adipose‐derived exosomes from obese vs. lean individuals, indicating that adipose‐derived exosomal miRNAs in obesity may play an important role in developing metabolic dysfunction in human skeletal muscle. We, therefore, hypothesize that adipose‐derived exosomes may induce insulin resistance in skeletal muscle. Exosomes were isolated from visceral adipose tissues from severely obese individuals (BMI > 40kg/m2) using ExoQuick‐TC (System Biosciences). Skeletal muscle cells were obtained from lean and several obese individuals (N=8: BMI < 25kg/m2 and BMI > 40kg/m2, respectively). Following proliferation and differentiation, fully mature myotubes were incubated with adipose‐derived exosomes (0.3ug/ml) for 24 hours and stimulated by insulin (100nM) for the last10 minutes. Then, cells were harvested to measure protein expressions in glucose metabolism using immunoblotting techniques. To measure the functional importance of exosome treatment, we examined insulin‐stimulated glycogen synthesis and glucose oxidation. While adipose‐derived exosomes significantly suppressed phosphorylation of AktSer 473 in skeletal muscle cells from both lean and obese individuals, we could not observe any changes in insulin‐stimulated glycogen synthesis and glucose oxidation. The main finding from this study is that adipose‐derived exosomes significantly suppressed Akt phosphorylation; however, there were no changes in glucose metabolism. This would suggest that, though it impairs intracellular signaling, exosomes may not be responsible for impaired glucose metabolism with severe obesity.Support or Funding InformationThis study was supported by the National Institute of Health (DK 56112).
Metformin Limits Loss of Mitochondrial Respiration Seen With Doxorubicin Treatment Without Affecting Muscle Function
Authors: Murphy Tyler Steadman , Crandall Amy Dee , Louw Makensie Joann , Bernhisel Drew , Reynolds Merrick , Hancock Chad R 
Published: November 2023

Doxorubicin (DOX) is a commonly‐used chemotherapeutic drug with a well characterized side effect of skeletal muscle wasting and weakness. It has been shown that DOX contributes to skeletal muscle damage through increasing oxidative stress. It has been suggested that this is due to DOX reacting with free cellular iron which then catalyzes the production of reactive oxygen species (ROS). ROS contribute to cell damage and dysfunction, and thus may play a role in the skeletal muscle wasting seen with DOX treatment. Recent work has shown that Metformin (MET) may protect against this oxidative environment by increasing iron storage capacity, thus preventing skeletal muscle wasting. We hypothesized that mice treated with DOX would see decreased muscle function and decreased respiratory capacity, and that pretreatment with MET would ameliorate these negative side effects. Male C57BL/6 mice were randomly placed into three groups: control, DOX or MET+DOX. Control mice received five doses of saline orally for five days before sacrifice, and one dose of saline intraperitoneally three days before sacrifice. DOX mice received one dose of DOX at 15 mg/kg three days before sacrifice. MET+DOX mice were given MET at 500 mg/kg two days before DOX treatment at 15 mg/kg. They were given this same dosage of MET every day until they were sacrificed three days after DOX treatment. Muscle function was measured in the gastrocnemius, plantaris and soleus muscle complex in situ, with mice under anesthesia (100 msec tetanic contractions at 0.5 Hz for 6 min). Mitochondrial respiratory function was measured in permeabilized fiber bundles from red gastrocnemius muscle using high‐resolution respirometry (OROBOROS Oxygraph‐2k). Treatment with DOX resulted in a 16.0–20.2% reduction in overall body weight in both the DOX and MET+DOX groups as compared to control (p‐value < 0.001). The weight difference between DOX and MET+DOX groups was not significant. Compared to control, DOX treatment caused a reduction in complex I stimulated, complex II stimulated, and uncoupled respiration (20–60% with p‐value < 0.001). MET significantly prevented these reductions (i.e. max uncoupled respiration was 81% of control with p‐value 0.08). Surprisingly we saw no significant differences in absolute force, twitch force, half relaxation time, or muscle fatigue between any of the three groups. These findings show that pretreatment with MET prevents decreases in mitochondrial respiration caused by DOX, but may not prevent muscle wasting. More research is needed in this area to find treatment options for DOX‐induced skeletal muscle dysfunction.Support or Funding InformationBrigham Young University
Overexpression of PGC‐1α Increases Peroxisomal Biogenesis and Mitochondrial Lipid Oxidation in Human Primary Myotubes
Authors: Huang Tai‐Yu , Zheng Donghai , Houmard Joseph A , Brault Jeffrey J , Hickner Robert C , Cortright Ronald N 
Published: November 2023

The conventional view of peroxisomal biogenesis has assumed that it is under the transcriptional regulation by PPAR. However, recent studies in hepatic cells have shown that the transcriptional co‐activator, PGC‐1α, can induce peroxisomal gene expression and abundance in a PPAR‐independent fashion. Also, metabolic linkage between mitochondria and peroxisomes may converge in terms of lipid metabolism. It is unknown whether the regulatory mechanism of peroxisomal biogenesis is through the same node as mitochondrial biogenesis in human skeletal muscle and whether fatty acid metabolism is affected. Therefore, we hypothesized that PGC‐1α acts as a biological “driver” for both peroxisomal and mitochondrial biogenesis in human skeletal muscle, leading to metabolic interactions in terms of fatty acid oxidation. Human skeletal muscle cells from vastus lateralis biopsies of lean donors (BMI= 23.6 ± 1.5 kg/m2, age=26.7 ± 3.5 years, n=3) were exposed to adenovirus encoding PGC‐1α or GFP at day 4 post differentiation and harvested at day 7. Peroxisomal biogenesis proteins (peroxins) and genes (PEXs) responsible for proliferation and function were assessed by western blot and quantitative real time‐PCR. 1‐14C lignoceric acid, necessarily oxidized first by peroxisomes, was used to assess mitochondrial oxidation of peroxisomal derived fatty acid metabolites.ResultsFollowing overexpression of PGC‐1α (protein increased 9.7 fold) 1) peroxisomal exclusive PMP70 and mitochondrial citrate synthase protein content were elevated 3 and 2.8 fold respectively (P<0.05) and PEX19 protein increased by 43% compared to the ad‐GFP controls 2) PGC‐1α, PMP70, PEX19 and a peroxisomal β‐oxidation enzyme, peroxisomal acyl‐coenzyme A oxidase 1, mRNA content were increased by 34.7, 4, 1.3, and 1.5 fold respectively (P<0.05) and 3) A concomitant 2.5‐fold increase in peroxisomal derived lignoceric acid metabolite oxidation by mitochondria was observed.ConclusionIn addition to the proliferative power on mitochondria, PGC‐1α is responsible (at least in part) for regulating peroxisomal biogenesis accompanied by elevations in the functional interactions of peroxisomes with mitochondrial lipid disposal in human skeletal muscle.Support or Funding InformationSupported in part by a seed grant from the Diabetes Obesity Institute, East Carolina University (RNC), NIH DK 056112(JAH), NIH 1R15HL113854‐01A1 (RCH)
Estrogen Treatment Restores Muscle Mitochondrial Function and Redox Homeostasis, reversing the Pro‐Diabetogenic State induced by Ovariectomy
Authors: Torres Maria Jose , Reese Lauren , Smith Cheryl , Pennington Edward , Ryan Terence , Shaikh Raza , Neufer Peter Darrell 
Published: November 2023

Menopause comes with a significant decline in 17β‐estradiol (E2) levels, increased adiposity and a higher risk for type‐2 diabetes, but hormone therapy can help revert these effects. However, we still lack a comprehensive understanding of the mechanism(s) by which E2 modulates insulin sensitivity. Herein, 10 week‐old C57BL‐NJ mice were studied after 2 weeks of ovariectomy to avoid the development of obesity (OVX‐2w), and following 2 weeks of E2 replacement (OVX+E2, 1μg/day) administered via a subcutaneous miniosmotic pump. Control groups included normally cycling females (NC) in the Proestrus stage (high physiological E2 levels) and an OVX group implanted with a saline control pump (OVX‐ctl). E2 treatment effectively reversed fasting hyperglycemia developed in the OVX‐2w and OVX‐ctl groups (+25%, p<0.05), decreased fat mass by 48% relative to OVX‐ctl (p<0.05), and restored ex‐vivo insulin‐stimulated muscle glucose uptake. Accompanying these effects, we demonstrate that soon after ovarian E2 withdrawal a decrease in mitochondrial function and an oxidative shift in the cellular redox environment occur; both conditions associated with the etiology of insulin resistance. Furthermore, spectrophotometric assessment of specific activities of the electron transport complexes in isolated skeletal mitochondria revealed a ~40% decrease in Complex I (p<0.005) and Complex III (p<0.05) activities in OVX mice. While Complex II activity was not altered, transfer of electrons between Complex I and III, as well as between II and III was reduced by 50% in OVX‐2w (p<0.05), and 35% in OVX‐ctl (p<0.05), but completely restored in OVX+E2. Concomitant with a decrease in mitochondrial membrane fluidity (~‐50%, p<0.05), OVX‐2w mice also showed reduced electron transport supercomplex formation (p<0.05), but all changes were fully rescued in OVX+E2. Overall, we show that E2 protects mitochondrial function and redox homeostasis via non‐genomic pathways; by increasing Complex I and Complex IIII activities, enhancing electron transfer supercomplex assembly, promoting electron transfer efficiency within the Q‐pool, preventing mitochondrial free radical leak, and increasing fluidity of mitochondrial membranes. Our findings offer new insights into the mechanism(s) by which menopause sets, and E2 therapy reverses a pro‐diabetogenic state. A deeper understanding of E2 mechanisms of action in metabolism is of paramount importance to advance in the development of alternative non‐hormonal therapies; that can optimize the metabolic benefits, without the reproductive, oncogenic, and cardiovascular risks associated with HRT, for natural or surgically‐induced menopausal women.Support or Funding InformationNIH DK096907
Triclosan is Excluded from Fetal Brain When Administered to the Pregnant Ewe
Authors: Quevedo Darla , Bryan Daphne , Zarate Miguel , Wood Charles E. 
Published: November 2023

Triclosan (TCS: 5‐chloro‐2‐(2,4‐dichlorophenoxy)‐phenol) is a synthetic antibacterial compound that is commonly added to personal care products. TCS acts as an antibacterial because it interrupts fatty acid synthesis by inhibiting enoyl reductase in bacteria. Because of its action on a bacterial enzyme that is not expressed in the human being, it is thought to be safe for use in humans. We have previously found that TCS administration to the late‐gestation pregnant ewe stimulates a genomic response in the fetal hypothalamus that is consistent with disturbance in lipid biosynthesis and metabolism. We performed the present experiment to test whether the action of TCS on the fetal brain is direct or indirect. Late‐gestation pregnant ewes and their fetuses were chronically catheterized, allowed to recover from surgery, then infused with TCS (0.1 mg/kg/day, iv) for 2 days. TCS concentrations were measured in plasma and after extraction from tissues using an ELISA (Abraxis Corp). The infusion increased maternal plasma TCS concentration to 3.0±0.5 ng/mL compared to 0.2±0.5 ng/mL in saline‐infused ewes. TCS was detected in relatively high concentrations in placenta (0.73±0.18 ng/g, n=4, compared to 0.07±0.05 ng/g, n=4, in untreated fetuses), but not in fetal cerebral cortex (0.24±0.04 ng/g, n=8, compared to 0.27±0.12 ng/g in untreated fetuses, n=8). Interestingly, TCS was measured in plasma and tissue of some untreated animals, perhaps resulting from environmental TCS exposure. This was confirmed by measurement of low levels of TCS in the drinking water in this facility (0.13±0.01 ng/mL, n=3). We conclude that there is a functional barrier that prevents TCS infused into the pregnant ewe from entering the fetal brain. We speculate, therefore, that fetal brain genomic responses to maternal infusions of TCS are an indirect response.Support or Funding InformationThis work was supported by NIH grants R01 HD033053, R01 HD057561, and R25 HL103181.
Characterization of Calcium Signaling in TαT1 Thyrotrophs
Authors: Tomic Melanija , Bargi‐Souza Paula , Leiva‐Salcedo Elias , Nunes Maria Teresa , Stojilkovic Stanko S 
Published: November 2023

TαT1 cells are mouse pituitary thyrotroph cell line used in studies on thyroid‐stimulating hormone beta subunit gene (Tshb) expression and other cellular functions. Here we studied activation of calcium mobilization and influx pathways in single isolated TαT1 cells in response to application of thyrotropin‐releasing hormone (TRH), a native hypothalamic agonist for thyrotrophs, and extracellular ATP, an intrapituitary agonist for two classes of receptors: G protein‐coupled P2Y receptors and ATP‐gated P2X channels. We also examined contribution of voltage‐gated and voltage‐independent calcium influx pathway to calcium signaling, and the role of triiodothyronine (T3) in cellular calcium homeostasis. Our investigations indicate that TαT1 cells are excitable and fire action potentials spontaneously. The spontaneous electrical activity is coupled to small amplitude fluctuations in intracellular calcium ion concentration. Calcium influx is to a large extent mediated by voltage‐gated calcium channels, as demonstrated by the effects of Bay K 8644, an L‐type voltage‐gated calcium channel agonist. TRH stimulates both calcium mobilization from intracellular pools and calcium influx. Non‐receptor‐mediated depletion of intracellular pool also leads to a prominent facilitation of calcium influx. Both receptor and non‐receptor stimulated calcium influx is substantially attenuated but not completely abolished by inhibition of voltage‐gated calcium channels, suggesting that depletion of intracellular calcium pool in these cells provides a signal for both voltage‐dependent and ‐independent calcium influx. We also found that TαT1 cells express P2Y1 receptors and their activation by extracellular ATP mimics TRH action on calcium mobilization and influx. The thyroid hormone T3 prolongs duration of TRH‐induced calcium spikes during 30‐min exposure. In conclusion, the expression of functional TRH and P2Y1 receptors in TαT1 cells indicates that embryonic thyrotrophs are equipped to respond to feed‐forward action of hypothalamic TRH and intrapituitary effects of released ATP with a rapid calcium mobilization and sustained calcium influx. Moreover, TRH‐induced calcium mobilization in TαT1 cells is amplified by T3, an action consistent with the positive feedback effect of this hormone on calcium‐secretion coupling, presumably through a non‐transcriptional mechanism, in contrast to the transcriptional and post‐transcriptional negative effects of T3 on Tshb expression.Support or Funding InformationThis work was supported by the Intramural Research Program of the Eunice Kennedy Shiver National Institute of Child Health and Human Development (ZIA DH000195‐22).
mTOR signaling coordinates glucose and lipid metabolisms in liver
Authors: Li Ziru 
Published: November 2023

ObjectiveAlthough aberrant mTOR activity has been linked to the development of obesity and diabetes, its role in hepatic glucose and lipid metabolisms remains largely unknown. We aimed to explore the effects of hepatic mTOR activity on the glucose and lipid homeostatsis.MethodsTSC1flox/flox mice and mTORflox/flox mice were injected with Cre adenovirus to activate or inhibit mTOR signaling respectively. By breeding albumin‐Cre mice with TSC1flox/flox mice (AT) or mTORflox/flox mice (Am), liver‐specific mTOR signaling was activated or suppressed. OGTT and glucose metabolism related genes were evaluated. Oil‐red O staining and lipid metabolism related genes were assessed as well.ResultsTSC1flox/flox mice with tail vein injection of Ad‐Cre demonstrated an improvement in glucose tolerance relative to wildtype littermates. Levels of genes related to gluconeogenesis, such as PEP carboxykinase (pepck) and pyruvate carboxylase (pc) were significantly decreased. Oil‐red staining demonstrated a significant decrease of hepatic lipid accumulation. Hepatic lipogenesis related genes such as pparγ2, srebp1c, acaca, gpat and dgat were markedly decreased. Interestingly, levels of chREBP mRNA were significantly decreased. This alteration was associated with a decrement in L‐type pyruvate kinase (lpk) and fatty acid synthase (fasn) mRNAs. Inversely, mTORflox/flox mice with tail vein injection of Ad‐Cre showed deteriorated glucose tolerance compared with wildtype littermates. Genes related to gluconeogenesis were significantly increased. Lipids deposition and lipogenesis related genes were increased as well. Similar results were observed in animal models with genetic interference of mTOR signaling including hepatic deletion of TSC1 (AT) and mTOR (Am ). Activation of mTOR signaling in AT mice demonstrated a decrease in hepatic lipid deposit and triglyceride contents. On the other hand, suppression of hepatic mTOR activity in Am mice increased lipid contents in liver.ConclusionOur studies indicated that hepatic mTOR signaling plays an important role in glucose and lipid metabolisms. Targeting hepatic mTOR signaling may provide a promising therapeutic approach for hepatic steatosis.Support or Funding InformationThis research was supported by grants from American Diabetes Association grant #1‐13‐BS‐225.
Identification of Glycoproteins on the Surface of Cow's Milk Exosomes that Mediate the Uptake of Exosomes into Human Colon Carcinoma Caco‐2 Cells
Authors: Sukreet Sonal , Zhang Hanyuan , Adamec Jiri , Cui Juan , Zempleni Janos 
Published: November 2023

BackgroundMicroRNAs (miRNAs) play essential roles in gene regulation. A substantial fraction of miRNAs in tissues and body fluids is encapsulated in exosomes (exos), thereby conferring protection against degradation. Traditionally, miRNAs have been considered endogenous regulators of gene expression. Recently, we have made the paradigm‐shifting discovery that humans absorb dietary miRNAs from cow's milk (encapsulated in exos) by endocytosis, and dietary miRNAs have biological activity in humans. Inhibitor and competitor studies implicated glycoproteins on the surface of exos and cells in intestinal transport.ObjectiveIdentify glycoproteins on cow's milk exos as putative substrate recognition feature in intestinal transport.MethodsExos were isolated from cow's milk using ultra centrifugation; identity, purity, and integrity were confirmed using nanoparticle tracker, western blot, and transmission electron microscopy. Where applicable, exos were treated with the proteases Glu‐C (targeting glu), trypsin (targeting arg and lys), or proteinase K (non‐specific); controls were not treated with proteases. For transport studies, exos were labeled using fluorophore (FM‐4), and uptake rates were analyzed in human intestinal Caco‐2 cells. Surface peptides released by treatment with trypsin were identified using LC/MS‐MS and Mascot, and glycoproteins were identified using SwissProt. One way Anova and Bonferroni's Multiple comparison was used to test for statistical significance.ResultsWe identified 417 exosomal surface proteins, including N‐glycans (4), O‐glycans (2), and C‐glycans (2). When exos were treated with Glu‐C or trypsin, rates (Vmax) decreased compared with controls (arbitrary units): 88±1 for control, 40±0.6 for trypsin, 19±0.3 for Glu‐C (P<0.05 vs. control). For comparison, we identified 504 proteins in breast milk exos (6‐N‐glycans, 4 O‐glycans, and 3 C‐glycans). The identities of surface glycoproteins were distinct in bovine and human milk exos.ConclusionEight glycoproteins were identified on surface of cow's milk exos, which might be essential for intestinal transport. We propose that glycan features, as opposed to protein features, are important for exo recognition by intestinal cells.Future directionsWe will use site‐directed mutagenesis and gene knockout protocols in cell cultures and mice to provide unambiguous evidence for the importance of distinct glycoprotein features on the mucosal surface in exo recognition.Support or Funding InformationSupported by the National Institute of Food and Agriculture, U.S. Department of Agriculture, under award number 2015‐67017‐23181, NIH 1P20GM104320 and P30GM103335, the Gerber Foundation, the Egg Nutrition Center, the University of Nebraska Agricultural Research Division (Hatch Act), and USDA multistate group W3002.