bluuebunny/crossref_metadata_2025_split huggingface dataset:
Depletion of Programmed Cell Death Protein 4 (PDCD4), an mTORC1/S6K1 Substrate, Attenuates Proteolysis in L6 Myotubes
Authors: Adegoke Olasunkanmi John , Sawan Sidney Abou , Cho Hankyul 
Published: November 2023

The mammalian (mechanistic) target of rapamycin complex 1/ribosomal S6 Protein kinase 1 (mTORC1/S6K1) signaling pathway is a critical regulator of skeletal muscle protein synthesis and mass. Because mTORC1 signaling is also implicated in autophagy control, regulation of muscle protein mass by mTORC1 may be mediated by regulation of protein synthesis and/or degradation. We recently characterized the expression of programmed cell death protein 4 (PDCD4), a substrate of mTORC1/S6K1, in skeletal muscle. This pro apoptotic protein normally inhibits mRNA translation by binding to eukaryotic translation initiation factor 4A (eIF4A), but upon phosphorylation by S6K1, it is targeted for proteolysis by the ubiquitin dependent proteolytic system. Depletion of PDCD4 in myoblasts attenuated the suppression of protein synthesis that is normally observed in response to amino acid starvation. Surprisingly in myotubes, PDCD4 depletion led to a reduction in protein synthesis. This led us to hypothesize that PDCD4 likely regulates both protein synthesis and degradation, and that its depletion would lead to a greater suppression of proteolysis (compared to suppression of protein synthesis), thus leading to positive protein balance. L6 myotubes were depleted of PDCD4 by RNA interference. Thirty‐six hours later, cells were pulsed with S‐35 methionine/cysteine for 12 hours. To activate proteolysis, cells were then incubated either in amino acid‐free medium (starvation), or in 1 μM dexamethasone for varying lengths of time. Samples were then harvested and subjected to autoradiography to measure residual radioactivity. At 24–36 hours, starvation increased the disappearance of radiolabeled protein by 50–80% compared with non‐starved cells (P<0.05), consistent with increased proteolysis. In PDCD4 depleted cells, starvation‐induced proteolysis was suppressed by 40–60% (P<0.05). Ubiquitinated proteins accumulated to a greater extent (40–100%) in starved PDCD4‐depleted cells compared to cells treated with control siRNA (P<0.05), consistent with inhibition of degradation of ubiquitinated proteins. In dexamethasone treated cells, proteolysis was inhibited by ~20% in PDCD4 depleted cells compared to cells treated with control siRNA (P<0.001). These data are consistent with a role for PDCD4 in regulating not only protein synthesis but also proteolysis. Therefore, interventions that target PDCD4 may help in limiting muscle proteolysis seen in catabolic conditions like cancer and insulin resistance states.Support or Funding InformationNSERC; York University
A Plasma Proteome is Associated with Anthropometric Status in School‐aged Children in Nepal
Authors: Lee Sun Eun , Christian Parul , Schulze Kerry , Cole Robert N. , Wu Lee S. F. , Yager James D. , Groopman John , Stewart Christine P. , West Keith P. 
Published: November 2023

BackgroundChild undernutrition, often interpreted as protein‐energy malnutrition (PEM), is typically assessed by anthropometry. Rarely are plasma markers of PEM considered in population surveys, masking opportunities to study biological mechanisms of poor growth.ObjectiveThis study aimed to identify plasma protein biomarkers associated with childhood body size measures and indicators of stunting, wasting, underweight and body composition using proteomics.MethodsWe correlated the relative abundance of plasma proteins quantified by mass spectrometry with measures of height, weight, mid‐upper arm circumference (MUAC), triceps skinfold thickness (TSF), and subscapular skinfold thickness (SSF), and estimated differences in relative abundance of proteins by stunting, underweight, and wasting status in 500 children 6–8 yr of age in rural Nepal.ResultsAmong 982 proteins observed in >10% of all child plasma samples, we identified 18, 58, and 28 proteins associated with height‐for‐age Z‐scores (HAZ), weight‐for‐age Z‐scores (WAZ), and MUAC, respectively, passing a false discovery threshold of 10% (q<0.10). Fifteen proteins mainly involved in cell proliferation/differentiation, ossification, and host defenses were commonly associated with both HAZ and WAZ, whereas WAZ was also associated with extracellular matrix proteins. Three proteins involved in muscle contraction and maintaining the integrity of connective tissues were associated with BMI‐for‐age Z‐scores (BMIZ) and one protein, collagen (type 6) alpha1, was positively associated with arm fat area. Four proteins, including breast cancer type 2 susceptibility protein and sex hormone binding protein, were associated with SSF and no proteins were associated with TSF (q<0.10). Twelve proteins including insulin‐like growth factor‐1 (IGF‐1), IGF‐binding proteins (IGFALS and IGFBP3), and tetranectin were 3–15% less abundant in the plasma of stunted children (HAZ<−2) relative to non‐stunted children (q<0.10). Fifteen proteins, including IGF‐1, ‐2, ‐ALS, and ‐BP3 and others involved in angiogenesis, nutrient transport, and nervous system development were 3–24% less abundant while 3 proteins related to regulation of adipogenesis and neuroendocrine secretory function were 7–9% more abundant in underweight children (WAZ<−2) than non‐underweight children (q<0.10). No protein was differentially abundant by low BMI (BMIZ<−2) status.ConclusionsSubsets of the plasma proteome correlated with body length, weight, lean mass and fat mass in school‐aged children. The utility of identified proteins as biomarkers of growth processes leading to underweight, stunting and altered body composition merits further investigation.Support or Funding InformationThis study is funded by the Bill and Melinda Gates Foundation (OPP5241 and GH614).
Maternal Docosahexaenoic Acid (DHA) Supplementation Promotes Lean Body Mass in the Preterm Infant
Authors: Valentine Christina J. , Kleiman Jeanne , Dingess Kelly , Morrow Adrythe , Rogers Lynette K 
Published: November 2023

BackgroundMaternal supplementation with 1000 mg/day docosahexaenoic acid (DHA) increases breast milk concentration to target fetal third trimester DHA accretion levels. Recent studies have observed correlations between n‐3 LCPUFA levels and lean body mass. Given that extremely preterm infants experience rapid “catch‐up” growth after birth, sufficient quantities of DHA in breast milk may decrease fat accumulation and increase lean mass in this population.MethodsMother/infant dyads were enrolled at the time of birth and randomized to receive 200 or 1000 mg/day of single source (algeal) DHA for 8 weeks. Inclusion criteria was that the infants be born <28 weeks gestation and have no congenital anomalies. A total of 18 mother infant dyads completed the study, 8 on 200 mg/day and 10 on 1000 mg/day DHA supplementation. Body composition was assessed by PEA POD®, utilizing air displacement plethysmography and whole body densitometry, at term corrected gestational age or hospital discharge. Data were analyzed by Multivariate General Linear Regression using DHA supplementation as the primary variable and age at analysis as a co‐variant.ResultsComparing infants fed by mothers receiving 200 mg vs 1000 mg DHA per day, data indicated lower percent fat (12.4 vs 11.0, p=0.030); fat mass (0.38 vs 0.33, p=0.014); body mass (3.00 vs 2.90 kg, p=0.042); and body volume (2.91 vs 2.81 L, p=0.039); and higher percent fat free mass (87.6 vs 89.0, p=0.030). No differences were observed in overall length.ConclusionsThe total number of infants in this study is small but we observed decreases in parameters of body fat and body mass and increases in percent fat free mass in the infants nursed by mothers supplemented with 1000 g/day DHA. Larger studies are needed to determine if higher levels of dietary DHA regulates excess fat accumulation in this population of extremely preterm infants.Support or Funding InformationThis work was supported by the National Institutes of Health, NCCIH/ODS (LKR R01AT006880).
LONG NON‐CODING RNA HOTAIR ACTIVATION BY TGF‐BETA1 INDUCES EPITHELIAL MESENCHYMAL TRANSITION (EMT) IN T24 BLADDER CARCINOMA CELLS
Authors: de Oliveira Brito Rodrigo Barbosa , Malta Camila Soares , Matos Yves Silva Teles , Souza Diego Mota , Matheus Luiz Henrique Gomes , Dalboni Maria Aparecida , Dellê Humberto 
Published: November 2023

HOTAIR is a long non‐coding RNA (lncRNA) which is implicated in cell programming for initiation of epithelial mesenchymal transition (EMT) in several types of cancer. In bladder carcinoma T24 cells, the TGF‐beta1 promotes invasion and metastasis through induction of EMT, however, there are evidences to believe that HOTAIR is a possible link between TGF‐beta1 and EMT in bladder cancer.The aim of this study was to analyze the expression of IncRNA HOTAIR in human bladder carcinoma T24 cells incubated with TGF‐beta1, in order to correlate it with markers for EMT.Bladder carcinoma cells (T24) were incubated with TGF‐beta1 (5 ng/ml) during 48h and then total RNA was extract to analyze the expression of Hotair, N‐cadherin (nCAD, a mesenchymal cell marker), E‐cadherin (eCAD, an epithelial cell marker), and Twist (an EMT transcription factor) by RT‐qPCR. The results are presented in mean±SEM, considering relative expression (RE).T24 cells stimulated with TGF‐beta1 presented an increased Hotair expression (RE 2.80 ± 0.14 vs. 1.00 ± 0.17 in control, p<0.001). TGF‐beta1 upregulated the expression of nCAD gene (RE 2.22 ± 0.55 vs 1.00 ± 0.32 in control, p <0.0001), while decreased eCAD expression (RE 0.23 ± 0.15 vs 1.00 ± 0.07 in control, p <0.05). The Twist expression was significantly increased by treatment with TGF‐beta1 (RE 3.05 ± 0.54 vs 1.00 ± 0.32 in control, p <0.001).We conclude that TGF‐beta1 upregulates Hotair expression in T24 cells, correlating with the EMT. It is possible that HOTAIR is involved in cell invasion and metastasis in bladder cancer.Support or Funding InformationFAPESP (2012/04423‐0)
The Sensitivity and Specificity of Thyroglobulin as an Index of Iodine Status in Mildly Iodine Deficient Adults
Authors: Skeaff Sheila Anne , Ma Zheng Feei 
Published: November 2023

Iodine deficiency is still one of the most common micronutrient deficiencies in the world. Iodine status is typically assessed by the determination of urinary iodine concentration (UIC) from a single spot urine sample obtained from a group of subjects. The median UIC of the group is then compared against population cut‐offs set by WHO and the Iodine Global Network. Typically only one spot urine sample is collected from each subject, but the large inter‐and intra‐individual variation means that the UIC in a single urine sample cannot be used as an index of iodine status in an individual. The collection of multiple spot urine samples from one subject has been shown to improve the precision of UIC, but this is associated with a increase in respondent burden. Serum thyroglobulin (Tg), a blood protein specific to the thyroid gland, holds promise as a biomarker of individual iodine status; higher levels of Tg are associated with lower UIC. The aim of this study was to assess the sensitivity and specificity of Tg as a index of iodine status.The study was conducted in Dunedin, New Zealand in a group of adults (n=165) aged 18–40 years. From each participant, five spot urine samples were obtained over a one week period. The average UIC of the five samples was used as the “gold standard” to categorize each participant as iodine deficient (UIC<100 μg/L) or sufficient (UIC ≥100 μg/L). A blood sample was collected for the determination of Tg. Three Tg cut‐offs were used including >11 μg/L, >13 μg/L, and >15 μg/L; a Tg concentration >13 μg/L has been recommended for use in children. The sensitivity, specificity, positive predictive value, and negative predictive value of each cut‐off of Tg was subsequently determined.As a group, the median UIC was 74 μg/L (range: 16–350 ) and median Tg was 16 μg/L (range: 0.2–179 ). The sensitivity was 73%, 65%, and 55% for a Tg >11 μg/L, >13 μg/L, and >15 μg/L, respectively. The specificity was 72%, 59%, and 53% for a Tg >11 μg/L, >13 μg/L, and >15 μg/L, respectively. The positive predictive value was 87%, 80%, and 74% for a Tg >11 μg/L, >13 μg/L, and >15 μg/L, respectively. The negative predictive value was 52%, 40%, and 60% for a Tg >11 μg/L, >13 μg/L, and >15 μg/L, respectively.The results of this study indicate that Tg had relatively poor sensitivity and specificity, and a high negative predictive value. This suggests that Tg is not a good index of iodine status in individual adults, however, its usefulness for groups of adults has yet to be elucidated.Support or Funding InformationLaurenson Award from the Otago Medical Research Foundation.
Vitamin B12 Status in Pregnant Women and their Infants in Southern India
Authors: Finkelstein Julia L. , Dwarkanath Pratibha , Mehta Saurabh , Thomas Tinku , Kurpad Anura V. 
Published: November 2023

ObjectiveTo determine the prevalence of vitamin B12 deficiency in a prospective cohort study of pregnant women, and its associations with infant vitamin B12 status and adverse birth outcomes in Southern India.MethodsPregnant women (n=419) participating in a perinatal cohort study in India were enrolled before 12 weeks gestation and monitored to assess vitamin B12 and folate status during pregnancy. Plasma vitamin B12 concentrations, methylmalonic acid, and homocysteine concentrations and erythrocyte folate concentrations were measured in maternal samples at enrollment and in infant cord blood at delivery. Binomial and linear regression models were used to evaluate the associations between maternal and infant vitamin B12 status.ResultsThe median vitamin B12 concentrations in maternal and infant samples were 26.9 (IQR: 89.6, 172.2) and 173.7 (IQR: 108.6, 264.1) pmol/L. 64% of mothers were vitamin B12 deficient (<150 pmol/L) at their first trimester visit, and 41% of infants were vitamin B12 deficient at birth. Maternal vitamin B12 concentrations at baseline were associated with significantly higher infant vitamin B12 concentrations (p<0.0001) and lower risk of infant vitamin B12 deficiency (RR: 0.58, 95% CI: 0.49–0.69, p<0.0001). Maternal vitamin B12 deficiency was associated with significantly lower infant vitamin B12 concentrations (p<0.0001) and two‐fold higher risk of infant vitamin B12 deficiency (RR: 1.99, 95% CI: 1.44–2.75, p<0.0001) at birth. Maternal vitamin B12 deficiency was also associated with significantly higher risk of infant small‐for‐gestational age (RR: 1.48, 95% CI: 1.04–2.13, p=0.03).ConclusionsInadequate maternal vitamin B12 status during pregnancy predicted increased risk of infant vitamin B12 deficiency. Future interventions are needed to improve vitamin B12 status in women of reproductive age to reduce the risk of vitamin B12 deficiency in mothers and their infants.Support or Funding InformationDivision of Nutritional Sciences, Cornell University
Targeting the Enterohepatic Bile Acid Signaling to Modulate Hepatic Autophagic Activity in Non‐alcoholic steatohepatitis
Authors: Wang Yifeng , Ding Yifeng , Li Jibiao , Chavan Hemantkumar , Matye David , Ni Hongmin , Krishnamurthy Partha , Ding Wen‐xing , Li Tiangang 
Published: November 2023

Non‐alcoholic steatohepatitis is associated with hepatic free cholesterol accumulation and defective autophagy activity, which sensitize hepatocytes to injury and inflammation. Bile acid synthesis is the only quantitatively significant mechanism for hepatic cholesterol catabolism. Stimulating bile acid synthesis prevented hypercholesterolemia and hepatic steatosis in mice. Autophagy delivers intracellularly stored cholesterol to the lysosome where cholesterol ester is hydrolyzed for subsequent cellular distribution, efflux and bile acid synthesis. The objective of this study is to investigate the molecular link between cholesterol and bile acid metabolism and autophagic activity in hepatocytes and in mouse models of fatty liver disease. Confocal microscopy, immunoblot and cathespin B activity assay showed that excessive intracellular free cholesterol accumulation caused by free cholesterol loading and inhibition of cholesterol esterification in the endoplasmic reticulum (ER) with an ACAT inhibitor blocked autophagosome/lysosome fusion and impaired lysosome function in cholesterol‐laden HepG2 cells. In contrast, cholesterol ester accumulation induced by purified human low‐density lipoprotein treatment increased lysosome function without inhibiting autophagy activity. In addition, bile acid treatment blocked autophagosome/lysosome fusion without affecting key autophagy gene expression in HepG2 cells. Interestingly, over‐expression of CYP7A1 by adenoviral gene delivery, which induced ER cholesterol catabolism and caused relative ER cholesterol depletion, strongly induced autophagy flux in both human hepatocytes and HepG2 cells. Based on this novel regulatory relationship between ER cholesterol sensing and autophagy activity, confocal microscopy, electron microscopy and immunoblot further revealed that cholestyramine administration strongly induced hepatic autophagy in lean mice, and restored hepatic autophagy activity and improved hepatic steatosis in obese mice. The cholestyramine‐mediated hepatic autophagy induction can be attributed to CYP7A1‐mediated ER cholesterol catabolism and decreased hepatic bile acid signaling. In conclusion, these results suggest that hepatic autophagy is regulated by an intracellular cholesterol sensing mechanism, and is an integral part of the complex regulatory network that maintains cholesterol homeostasis in hepatocytes. This study supports a new concept of targeting the enterohepatic bile acid‐CYP7A1‐cholesterol‐autophagy axis to selectively modulate hepatic autophagy activity in fatty liver disease.Support or Funding InformationNIDDK and NIAAA
High MUC2 Biosynthesis Induce ROS Production which Increases Goblet Cell Susceptibility to ER stress and Apoptosis
Authors: Tawiah Adelaide , Moreau France , Chadee Kris 
Published: November 2023

MUC2 is a large glycoprotein, produced by goblet cells, which forms the protective mucus blanket overlying the intestinal epithelium. Apart from its role as a physical barrier, other functions of MUC2 have not been explored. During pathological conditions such as inflammatory bowel disease (IBD), MUC2 biosynthesis and secretion are accelerated. There is however little known on how MUC2 production is regulated and whether goblet cells undergo increased endoplasmic reticulum (ER) stress and apoptosis following high MUC2 biosynthesis and secretion. We studied the role of MUC2 in goblet cell ER stress and apoptosis using a high MUC2‐producing goblet cell line (HT29‐H) and a clone of HT29‐H (HT29‐L) in which MUC2 has been silenced using lentivirus shRNA technology. Cells were treated with ER stress‐inducing agents, and markers for ER stress (GRP78, ATF4, sXBP1 and CHOP) and apoptosis (caspase 3 cleavage, DNA fragmentation and PARP cleavage) were quantified by western blotting, confocal microscopy and transmission electron microscopy. Compared to HT29‐L, HT29‐H cells showed significant increase in ER stress and apoptosis in response to ER stress and apoptosis‐inducing agents. HT29‐H cells also exhibited higher mitochondrial activity and reactive oxygen species (ROS) production than HT29‐L although the cells had similar mitochondrial mass. Inhibition of ROS abrogated basal ER stress levels, tunicamycin and thapsigargin‐induced ER stress, and apoptotic events including cytoplasmic calcium influx, loss of mitochondrial membrane potential, and caspases 3 and 4 activation. This indicates that high MUC2 production specifically increases ROS production, which drives ER stress and apoptosis. These findings were corroborated in isolated colonic epithelial cells and colonic tissues from Muc2+/+ and Muc2−/− mice that showed significant increase in ER stress in Muc2+/+ mice as compared to Muc2−/− mice. Finally, goblet cells in Muc2+/+ mice exhibited severe ER stress and apoptosis during progressive development of DSS‐induced colitis leading to disruption of the mucus barrier. We conclude that high ROS output, as a consequence of high MUC2 production, induces ER stress and apoptosis in goblet cells that could subsequently impair mucus barrier function in disease pathogenesis.Support or Funding InformationCanadian Institutes of Health Research (CIHR)
Hydroxylase inhibition reduces transforming growth factor‐β1 induced fibrosis associated with colitis
Authors: Manresa Mario , Tambuwala Murtaza , Radhakrishnan Praveen , Harnoss Jonathan , Cavadas Miguel , Keogh Ciara , Cheong Alex , Cummins Eoin , Barrett Kim , Schneider Martin , Taylor Cormac 
Published: November 2023

Intestinal fibrosis is a severe complication of inflammatory bowel disease (IBD) that often leads to surgical interventions, highlighting an urgent need for alternative therapies. The inhibition of key oxygen‐sensing hydroxylases that regulate the response to hypoxia ameliorates inflammation in mouse models of colitis. Here we investigated the impact of hydroxylase inhibition on intestinal fibrosis. Intestinal fibrosis was induced in mice secondary to colitis through exposure to dextran sodium sulfate (DSS) in drinking water. Cultures of human colonic (CCD‐18Co) and mouse embryonic (MEF) fibroblasts were used to investigate the actions of hydroxylase inhibitors on the response to transforming growth factor‐β1 (TGF‐β1). Mice allowed to recover for 8 or 14 days after exposure to DSS developed fibrosis with profound alterations in the pattern of collagen deposition and infiltration of α‐smooth muscle actin (α‐SMA)‐positive fibroblasts. Treatment with the pan‐hydroxylase inhibitor, dimethyloxalylglycine (DMOG) during the recovery period reduced signs of fibrosis. In contrast, exposure of prolyl hydroxylase‐2+/− (PHD‐2+/−) mice to DSS resulted in severe intestinal fibrosis suggesting that the effects of DMOG are independent of the PHD‐2‐HIF axis. In vitro, hydroxylase inhibitors reduced TGF‐β1 mediated production of key fibrosis markers such as α‐SMA, Connective Tissue Growth Factor (CTGF) and collagen‐1‐α, in both CCD‐18Co and MEF, strongly suggesting an effect on TGF‐β1 signalling. Furthermore, these antifibrotic effects were present in PHD‐2+/− fibroblasts even though these cells showed enhanced responsiveness to TGF‐β1. In order to investigate the molecular mechanisms of the anti‐TGF effects of hydroxylase inhibitors, we analyzed their impact on the activation of TGF‐β1 signalling pathways. While no major effects were detected on the ability of TGF‐β1 to activate the Smad pathway, hydroxylase inhibition strongly inactivated TGF‐ERK signalling. Furthermore, areas of the intestinal mucosa of mice exposed to DSS where α‐SMA positive fibroblasts were concentrated, exhibited abundant positive phosphoERK nuclear staining. This observation highlighted the importance of the activation of the ERK pathway in fibrotic colons. Taken together, our studies showed that hydroxylase inhibition ameliorates intestinal fibrosis, likely via effects on TGF‐ERK signalling. We propose hydroxylase inhibition as a potential new therapeutic approach for the treatment of intestinal fibrosisSupport or Funding InformationThis work was funded by Science Foundation Ireland